Vector-Borne Diseases Transmission in Ecuador: Implication of Vertebrate Hosts As

Vector-Borne Diseases Transmission in Ecuador: Implication of Vertebrate Hosts as

Food Source of Triatomines in Chagas Disease, and the Diversity of Anopheles

Mosquites in Malaria

A dissertation presented to

the faculty of

the College of Arts and Sciences of Ohio University

In partial fulfillment

of the requirements for the degree

Doctor of Philosophy

Sofia B. Ocana-Mayorga

May 2020

This dissertation titled

Vector-Borne Diseases Transmission in Ecuador: Implications of Vertebrate Hosts as

Food Source of Triatomines in Chagas Disease and the Diversity of Anopheles

Mosquitoes in Malaria

SOFIA B. OCANA-MAYORGA

has been approved for

the Department of Biological Sciences

and the College of Arts and Sciences by

Mario J. Grijalva

Professor of Biomedical Sciences

Florenz Plassmann

Dean, College of Arts and Sciences 3

ABSTRACT

OCANA-MAYORGA, SOFIA B., Ph.D., May 2020, Biological Sciences

Vector-Borne Diseases Transmission in Ecuador: Implications of Vertebrate Hosts as

Food Source of Triatomines in Chagas Disease and the Diversity of Anopheles

Mosquitoes in Malaria

Director of Dissertation: Mario J. Grijalva

Malaria and Chagas disease are two of the most important vector-borne parasitic infections in Ecuador. A better understanding on the transmission scenarios is needed to propose effective strategies for disease control. In Chagas disease, human-vector dynamics is not well understood; however, the presence of active transmission in endemic areas has been demonstrated. The pattern of blood meals of insects that are vector of diseases is fundamental to unveil transmission dynamics. In this study DNA from intestinal contents of triatomines were used to amplify fragments of cytb mitochondrial genes. The results showed that humans are a main source of food for triatomines, indicating that the vector-human contact is more frequent than previously thought. Although other groups of mammals such as rodents are also an available source of blood, birds (particularly chickens) might have a predominant role in the maintenance of triatomines in these areas. The presence of human blood meal in 42% of the samples, even in peridomiciliar and sylvatic environments, together with high infection rates with

Trypanosoma cruzi (the causal agent of Chagas disease) indicate that humans are not accidental host, but an important source of blood for triatomines in all environments, and with high risk of T. cruzi transmission. Consequently, domestic-centered strategies might 4 not prevent vectorial transmission and data of Chagas disease cases might be underestimated. Strategies for Chagas disease control in Ecuador require to evaluate capacity dispersal of triatomines and outdoor activities as risk factors for parasite transmission in Ecuador. In the case of malaria, the lack of a complete inventory of vector species is a barrier for an understanding of malaria transmission. Even more, implication on transmission of reported species is still poorly understood. Identification of species is mainly carried out by morphological characteristics; however, it fails in discriminating species closely related. Molecular tools are a good alternative to discriminate species. In this study, the internal transcribed spacer 2 (ITS2) and the cytochrome oxidase I (COI), were used for species identification. The Restriction

Fragment Length Polymorphism (RFLP) of the ITS2 is a technique that allows the analysis of a great amount of samples and it is cheaper than sequencing. However, variability within species and populations needs to be further known. The results demonstrated the great complexity of taxonomic identification of species, particularly of individuals from species complex, such as the case of the Oswaldoi group which species are sympathric in the Amazon region. In Ecuador, particularly, in the Amazon region we recommend the sequence of COI for anopheline identification until a better knowledge of the diversity is obtained. This study confirmed Ny. albimanus as the most abundant species in the Pacific coast and it also extends the geographic distribution of An. calderoni. In the Amazon region, Ny. benarrochi B is reported for first time in the

Ecuadorian Amazon. A comprehensive survey of Anopheles and Nyssorhynchus species in Ecuador is required to facilitate and targeted malaria control strategies in Ecuador. 5

DEDICATION

To my parents, to Romina and Martin, and to my brothers and sisters of blood and by

choice.

ACKNOWLEDGMENTS

My gratitude to Dr. Mario J. Grijalva who has been my mentor and inspiration to pursue a carreer in infectious disease. Thank you for teaching me why we do what we do.

To Dr. Simone Frédérique Brèniere for being an incredible support during this journey and for being my female role model in science.

I also want to thank my colleagues, without them I could not have achieve it: Anita

Villacis, César Yumiseva, Ana Lucía Moncayo, Fabían Sáenz, Juan José Bustillos,

Giovan Gómez for sharing your knowledge and time during the field research, collection and analysis of the data. A sincere gratitude to the community members of Loja, Manabi and El Oro provinces, for their selfless support.

An special acknowledgment to the institutions that funded this study: Pontifica

Universidad Católica del Ecuador (N234369, I13054, J13034, K13069)

UNICEF/UNDP/World Bank/WHO Special Programme for Reseach and Training in

Tropical Diseases (TDR) (A20785), Pan American Health Organization (A60655), the

European Commission Framework Programme 7 Project (ChagasEpiNet-contract No.

223034), Fogarty International Center (TW008261).

TABLE OF CONTENTS

Page

Abstract ...... 3 Dedication ...... 5 Acknowledgments...... 6 List of Tables ...... 10 List of Figures ...... 11 Chapter 1: Introduction ...... 12 1.1. Vector-borne Transmission of Chagas Disease and Malaria in Ecuador ...... 12 Chapter 2: Triatomine Feeding Profiles, Trypanosoma cruzi Infection and transmission Implications in Domestic and Sylvatic Habitats ...... 15 2.1. Abstract ...... 15 2.2. Background ...... 17 2.3. Methodology ...... 20 2.3.1. Triatomine Selection ...... 20 2.3.2. DNA Extraction and Natural Infection with Trypanosomes ...... 21 2.3.3. Comparison of Primer Sets for Detection of Blood Meals ...... 22 2.3.4. Amplification and Identification of the Sources of Blood Meals ...... 22 2.4. Results ...... 24 2.4.1. Sensitivity of the Different Primer Sets ...... 24 2.4.2. Vertebrate Blood Meal Source Diversity ...... 24 2.4.3. Blood Meal Diversity and Trypanosoma cruzi Infection in the Different Environments ...... 25 2.4.4. Vertebrate Blood Meal Source per Triatomine Species, Stages and Collection Environment ...... 28 2.5. Discussion ...... 32 2.5.1. Diversity of Mammal Hosts and its Implication in Trypanosoma cruzi Transmission ...... 33 2.5.2. Birds and its Role in Triatomine Population Maintenance ...... 36 2.5.3. Triatomine Species and Blood Meal Preference ...... 37 2.5.4. Triatomine Dispersal and Access to Vertebrate Hosts ...... 38 2.6. Conclusions ...... 40 2.7. References ...... 41 8

Chapter 3: Human Blood Meals in Sylvatic Triatomines Challenges Domestic-Centered Strategies for Prevention of Trypanosoma cruzi Transmission in Ecuador ...... 47 3.1. Abstract ...... 47 3.2. Background ...... 49 3.3. Methodology ...... 52 3.3.1. Study Area and Triatomine collection ...... 52 3.3.2. Amplification and Host Identification of Sources of Blood ...... 53 3.3.3. Natural Infection of Triatomines with Trypanosomes ...... 54 3.3.4. Haplotype Net for Human Sequences ...... 54 3.3.5. Feeding Behavior of R. ecuadoriensis Nymphs in Laboratory Conditions 54 3.4. Results ...... 56 3.4.1. Detection of Human Blood Meal and Trypanosoma cruzi in Triatomines, According to Regions and Environments...... 56 3.4.2. Population Structure, Microenvironment and Trypanosoma cruzi Infection of Triatomines with Human Blood Meals ...... 59 3.4.3. Sylvatic Triatomines with Human Blood Meals and its Distance to Households ...... 60 3.4.4. Haplotype Network of Human Sequences ...... 61 3.4.5. Feeding Behavior of R. ecuadoriensis Nymphs in Laboratory Conditions 62 3.5. Discussion ...... 63 3.5.1. Human as an Important Vertebrate Host for Triatomines and the Presence of Trypanosoma cruzi Infection ...... 64 3.6. Conclusions ...... 71 3.7. References ...... 73 Chapter 4: Discrimination of Anophelinae Species from Coastal and Amazon Ecuador Based on Nuclear and Mitochondrial Markers ...... 79 4.1. Abstract ...... 79 4.2. Background ...... 81 4.3. Methodology ...... 84 4.3.1. Study Area ...... 84 4.3.2. Species Identification ...... 85 4.3.3. ITS2 Amplification and Restriction Fragment Length Polymorphism (RFLP) Patterns ...... 85 9

4.3.4. COI Amplification and Sequencing ...... 88 4.4. Results ...... 89 4.4.1. Morphological Identification ...... 89 4.4.2. RFLP-ITS2 Identification ...... 91 4.4.3. COI Identification ...... 94 4.5. Discussion ...... 97 4.6. References ...... 102 Appendix A: Species, Year of Capture and Localtion of 507 Triatomines Analyzed from Central Coastal and Southern Highlands in Ecuador ...... 108 Appendix B: Identification of Triatomine Source of Blood in Samples from Central Coastal and Southern Highland Regions in Ecuador...... 136 Appendix C: Host Species, Collection Data and Sources of Blood of the 416 Triatomine Intestinal Content Analyzed by Sequence of cytb Fragment...... 155 Appendix D: Distance from Sylvatic Places of Capture to the Closest House...... 171 Appendix E: Identification of Mosquitoes Collected in Coastal Region (El Oro Province) and Amazon Region (Sucumbios and Orellana Province) by Morphology, RFLP-ITS2 Pattern and COI Sequencing...... 176

LIST OF TABLES

Page

Table 2.1 Thermal cycling parameters of PCR primers tested for sensibility and vertebrate source of blood detection from triatomine intestinal content DNA ...... 23 Table 2.2 Diversity of feeding host species identified in triatomines collected in intradomicile, peridomicile and sylvatic environments in Ecuador ...... 27 Table 3.1 Triatomine species and T. cruzi infection of the 168 samples where human blood meal were detected in central coastal (CC) and southern highland (SH) regions .. 57 Table 4.1. Expected RFLP pattern of ITS2 fragment digestion of the Anopheles and Nyssorhynchus species with the restriction enzymes AluI, HaeIII and FspI according to literature. In silico pattern was defined for An. calderoni identification...... 87 Table 4.2. Number or individuals identified by morphological characteristics collected in El Oro province (costal region), Sucumbios and Orellana provinces (Amazon region).. 90 Table 4.3. Identification based on RFLP-ITS2 restriction fragments with enzymes AluI, HaeIII and FspI of mosquitoes collected in El Oro province (costal region), Sucumbios and Orellana provinces (Amazon region)...... 93 Table 4.4. Identification by mithochondrial COI of mosquitoes collected in El Oro province (costal region), Sucumbios and Orellana provinces (Amazon region) ...... 96

LIST OF FIGURES

Page

Figure 2.1. Location of collection points and triatomine species included in the analysis of blood meal sources ...... 21 Figure 2.2. Proportion of blood meal source (species identification) of 416 triatomines collected from central coastal and southern highlands in Ecuador...... 25 Figure 2.3. Infection rates of triatomines captured in intradomiciliar, peridomiciliar and sylvatic environments with the diversity of host as sources of blood...... 28 Figure 2.4. Vertebrate host feeding pattern in triatomines...... 31 Figure 3.1. Geographical origin of the 416 samples included in this study from central costal and southern highlands of Ecuador...... 53 Figure 3.2. Human blood meal rates for each triatomine species ands and collection environment: intradocimiciliary, peridomiciliary and sylvatic...... 58 Figure 3.3 Triatomines with human blood meal, population structure, Trypanosoma cruzi infection, and microenvironments of collection by region...... 60 Figure 3.4 Median-joining network of mithocondrial cyt b haplotypes for the human sequences identified in the intestinal content of triatomines from central coast and southern highlands region...... 61 Figure 4.1. Location of communities where anopheline mosquito collection was carried out and species identified by sequencing of a fragment of the COI gene...... 95

CHAPTER 1: INTRODUCTION

1.1. Vector-borne Transmission of Chagas Disease and Malaria in Ecuador

Malaria and Chagas disease are two of the most important vector-borne parasitic infections in Ecuador. Recently, humanitarian crisis, migration, and underreporting of cases influences public health management and have called the attention of academia and public health personnel to the risk of disease reemergence. Thus, the identification of any gap of information on these diseases, including on contact human-vector interactions is necessary to direct research priorities. The aim of this dissertation was to analyze current transmission escenarios Chagas disease and malaria in Ecuador and how these scenarios might be influenced by human activities and the diversity of vectors. Vectorial transmission of both diseases is mostly associated with domestic environments. For

Chagas disease, there is a good knowledge of the diversity and distribution of triatomine vectors. The analysis of the blood meal of triatomines suggests that certain human-vector dynamics are underappreciated. On the other hand, little information about diversity and behavior of malaria vectors (Anopheles spp. and Nyssorhynchus spp.) is available in

Ecuador. Nyssorhynchus albimanus is the main vector species implicated in malaria transmission in coastal Ecuador, while little is known about the species in the Amazon region.

In chapter 2, the diversity of vertebrates that serve as source of blood for triatomines is addressed. Understanding transmission dynamics is fundamental to develop strategies to decrease vector-human contact. In Ecuador, limited data on human infection are available; however, the presence of active transmission in endemic areas has been 13 demonstrated. The aim of chapter was to determine the diversity of hosts as source of blood for triatomines and the potential reservoirs in two endemic areas of Ecuador

(central coastal and southern highlands regions). Analysis of the cytb mitochondrial gene showed that humans are the main source of food for triatomines. Although rodents are also, chickens might have a predominant role in the maintenance of triatomines in these areas. The diversity of blood source found might indicate a preference driven by triatomine species. Moreover, the results indicated that dispersal of vectors occurs regardless the availability of food. Dispersal capacity of triatomines need to be evaluated to propose and effective strategy that limits the vector-human contact and in consequence to decrease the risk of T. cruzi transmission.

In chapter 3, the transmission scenarios that favor the vector- human contact is discussed. Generally, the risk of transmission of Trypanosoma cruzi has been associated to domestic environments. However, alternative scenarios in which human and triatomines could be in contact have not been elucidated. In this chapter the frequency of human blood meal in the intestinal content of triatomines is evaluated. A high percentage of triatomines (42%) were found with human blood. Moreover, the presence of human blood meal in triatomes collected in chicken nest (29%) indicated that there might be a flow of triatomines between the intradomicile and peridomicile in search of blood sources. In sylvatic triatomines, 48%, had human blood meal and T. cruzi infection rate reaches 30%. These results indicate that humans are not accidental host, but an important blood source for triatomines in all environments, and with high risk of T. cruzi transmission. 14

In chapter 4, diversity of malaria vectors is evaluated. In Ecuador, there is a incomplete inventory of the diversity of malaria vectors of the genera Anopheles and

Nyssorhynchus. Even more, implication on malaria transmission of these vector species is still poorly understood. Morphological identification is the most common and inexpensive tool; however, it fails in discriminating species closely related and with similar morphological characteristics. However, molecular markers such as the internal transcribed spacer 2 (ITS2) and the cytochrome oxidase I (COI) has been used to discriminate species. With ITS2, the restriction length fragment (RFLP) patterns have been defined for some of the main South American species. RFLP is less expensive than sequencing; however, the diversity between and within species is reflected in the variability of the band sizes and the band pattern do not clearly discriminate the species as sequencing. The results of this study demonstrated the complexity of taxonomic identification of species, even with the use of molecular tools, particularly of individuals of the Oswaldoi group which species are sympathric in the Amazon region. The results of this study confirmed that in the coastal region of Ecuador, Ny. albimanus is the most abundant species and it also extends the geographic distribution of An. calderoni. In the

Amazon region, this study confirmed the presence of Ny. rangeli, Ny. oswaldoi B and Ny. aff. konderi. Moreover, molecular analysis increase Ny. benarrochi B in the list of reported species in the Amazon. 15

CHAPTER 2: TRIATOMINE FEEDING PROFILES, TRYPANOSOMA CRUZI

INFECTION AND TRANSMISSION IMPLICATIONS IN DOMESTIC AND

SYLVATIC HABITATS

2.1. Abstract

Understanding blood meal pattern of insects that are vector of diseases is fundamental to unveil transmission dynamics and to develop strategies to impede or decrease vector- human contact. Chagas disease has a complex transmission cycle that implies interactions between vectors, the parasite and vertebrate hosts. In Ecuador, limited data on human infection are available; however, the presence of active transmission in endemic areas has been demonstrated. The aim of this study was to determine the diversity of hosts that serve as source of blood for triatomines in two endemic areas of Ecuador (central coastal and southern highlands regions). Using unspecific primers and DNA intestinal contents of triatomines collected from 2006-20213, we amplified fragments of cytb mitochondrial genes. After sequencing, species were identified achieving ≥ 95% identity with GenBank sequences (NCBI-BLAST tool). The results showed that humans are a main source of food for triatomines, indicating that the vector-human contact is more frequent than previously thought. Although other groups of mammals such as rodents are also an available blood source, birds (particularly chickens) might have a predominant role in the maintenance of triatomines in these areas. However, the diversity of blood source found might indicate a preference driven by triatomine species. Moreover, the presence of more than one blood source from triatomines collected in the same point indicated that dispersal of vectors occurs regardless the availability of food. Dispersal capacity of 16 triatomines need to be evaluated to propose and effective strategy that limits the vector- human contact and in consequence to decrease the risk of T. cruzi transmission.

2.2. Background

Blood meal analysis is an important strategy to understand the implication of a hematophagous vector species in disease transmission and its ecological interactions.

Constant environmental changes, the emergence of outbreaks, and increase of vector distribution pose a challenges for understanding transmission scenarios. Important epidemiological traits of neglected tropical diseases (NTDs) such as transmission of pathogens across multiple taxa, primary vector species, vertebrate species with a role in transmission. Even social and economic conditions that foster transmission risk have been assessed for important vectors such as Culex (Eastwood, Cunningham et al. 2018,

Goodman, Egizi et al. 2018) Phlebotomus (Bennai, Tahir et al. 2018, de Avila, Brilhante et al. 2018); Aedes (Chepkorir, Venter et al. 2018) and triatomines (Dumonteil, Ramirez-

Sierra et al. 2018, Orantes, Monroy et al. 2018).

Chagas disease (CD) is considered as one of the NTDs and it is caused by the infection in humans with Trypanosoma cruzi. This parasite is transmitted, principally, by the feces of infected triatomines. The primitive transmission of T. cruzi was restricted to sylvatic environments (between triatomines and mammals without human participation); however, the gradual process of human settlement near the zoonotic transmission zone opened new niches for triatomines (Guhl 2017).

Understanding both scenarios of transmission, enzootic and human, is fundamental to know the dynamics of T. cruzi in different environments and how its cycle can be targeted to control CD. This protozoan parasite has been reported in more than 100 mammalian species (seven orders) in diverse environments (Noireau, Diosque et al. 2009, 18

Jansen, Xavier et al. 2015, Jansen, Xavier et al. 2017). Transmission of T. cruzi mostly occurs when infected feces of the triatomines are in contact with mucosae and injuries

(scratching). Other less frequent transmission routes are oral with ingestion of contaminated beverages and food (e.g. by scratching, eating the triatomine or infected animals) (Jansen, Xavier et al. 2017), transfusion and transplant of contaminated blood products and organs, congenital route and laboratory accidents (Apt, 2017, Moncayo and

Silveira, 2017). Although mammals can be infected by T. cruzi, not all mammals can be considered reservoirs, because it implies not only the infection but the infective potential and competence of the host (Jansen, Xavier et al. 2017). Moreover, birds, amphibians, reptiles are refractory to T. cruzi, but in particular the birds that represents an important feeding source for triatomines and play an important role in transmission because they can increase the density of vector populations (Jansen, Roque et al. 2017).

Even though transmission cycle of T. cruzi is difficult to delineate, it occurs in three main environments: (i) the enzootic or sylvatic cycle includes wild mammals and triatomines living in nest and borrows. This cycle is a dynamic web with different routes of transmission that may or may not overlap (Miles, Feliciangeli et al. 2003, Guhl 2017,

Jansen, Roque et al. 2017). In the space associated with human dwellings, two cycles can be defined: (ii) the domestic cycle, when the transmission occurs indoors, involving, mainly, humans and domiciliated triatomines (Guhl 2017) and (iii) the peridomestic cycle that occurs between triatomines infesting man-made construction and structures around the house and domestic animals (Walter, Lozano-Kasten et al. 2007). 19

Persistence of these cycles depends on the availability and diversity of the vertebrate hosts (Gottdenker, Chaves et al. 2012). To understand the dynamics of T. cruzi infection and the epidemiology of CD, it is necessary to determine the diversity of vertebrate hosts and its role in the different environments. Moreover, an accurate assessment of the risk of transmission is required in areas where human disturbance affect adjacent forest areas, and thus, vertebrates behavior (Jansen, Roque et al. 2017).

In central coastal and southern highlands regions of Ecuador, five species of triatomines are considered as the main vectors for Trypanosoma cruzi transmission in domestic and peridomestic environments: Rhodnius ecuadoriensis, Panstrongylus chinai,

P. howardi, P. rufotuberculatus and Triatoma carrioni (Grijalva, Villacis, Ocaña-

Mayorga et al. 2015, Grijalva, Villacis, Moncayo et al. 2017). Moreover, R. ecuadoriensis has been found as the main vector of T. cruzi in sylvatic environments and is the species with broader distribution (Grijalva and Villacis 2009, Grijalva, Teran,

Dangles et al. 2014). Regarding vertebrate hosts, T. cruzi has been reported infecting rodents (Rattus rattus, Oryzomys xanthaeolus and Scimosciurus nebouxii), marsupials

(Didelphis marsupialis) and bats (Artibeus fraterculus, Myotis sp. and Glossophaga soricina) (Pinto et al. 2006, 2015, Ocaña-Mayorga et al. 2015). However, how the interactions between vectors and hosts are not fully understood. Thus, the aim of this study is to evaluate the diversity of vertebrate hosts that serves as source of food for triatomines and to assess vector infection rate with T. cruzi in domestic, peridomestic and sylvatic environments in central coastal and southern highlands of Ecuador.

2.3. Methodology

2.3.1. Triatomine Selection Analysis were carried out in 507 selected samples of DNA purified from intestinal content, stored at -20 °C, of triatomines previously collected and from 2006-2013 in central coastal (CC) and southern highlands (SH) regions of Ecuador (Figure 2.1).

Samples were obtained from triatomines from nymphs (n= 228) and adults (n= 188).

Intestinal content were extracted from triatomines that do not exceed 40 days after collection, and that have not been fed before extraction. Triatomines were collected from three environments defined as: intradomicile (indoors) that refers to triatomines collected in any material or structure inside of the house (bed, matrix, indoor wall, animal nest, clothes); peridomicile (space around the house, usually delimited by a fence such as storage rooms, chicken crops, poultry or livestock yard, dog and cat beds, pigeon nests and piles of material); and sylvatic (outside household limits) associated with non- domestic animals (e.g. squirrel, rat and bird nests), but where human activities can take place (e.g. forest patches with trails, harvesting, etc.).

Figure 2.1. Location of collection points and triatomine species included in the analysis of blood meal sources.

2.3.2. DNA Extraction and Natural Infection with Trypanosomes DNA was extracted from the homogenized intestinal content with the DNAzol reagent for isolation of genomic DNA (InvitrogenTM, Waltham, USA) and stored at -20 °C until use. Infection with T. cruzi was assessed by amplification of the variable domain of the minicircle of kinetoplastid DNA (kDNA) by the presence of a 330 bp product (Vallejo et 22 al. 1999, Virreira et al. 2006). A negative sample (without DNA) and a positive control of

DNA for T. cruzi in each PCR assay. Amplification products were detected in a 2% agarose gel stained with 1X SYBR Safe (Thermo Fisher Scientific, Waltham, USA).

2.3.3. Comparison of Primer Sets for Detection of Blood Meals Four different primer sets that amplified mithocondrial gene fragments were assayed with serial dilution of human blood DNA (10 ng, 1ng, 100 pg, 10 pg, 1 pg, 100 fg) to determine the most adequate for vertebrate host identification in DNA from stored intestinal content. Two sets of primers amplified a fragment of cytb gene: FA-CYTB-F

(5´-CCC CTC AGA ATG ATA TTT GTC CTC A-3´) and FA-CYTB-R (5´-CCA TCC

AAC ATC TCA GCA TGA TGA AA-3´) (Buitriago, Depickere et al. 2012); L14841 (5´-

AAA AAG CTT CCA TCC AAC ATC TCA GCA TGA-3´) and H15149 (5´-AAA CTG

CAG CCC CTC AGA ATG ATA TTT GTC-3´) (Kocher, Thomas et al. 1989). Another set of primers amplified a fragment of the 12S-rRNA gene: L1091 (5´-AAA AAG CTT

CAA ACT GGG ATT AGA TAC CCC-3´) and H1478 (5´-TGA CTG CAG AGG GTG

ACG GGC GGT GTG T-3´) (Kocher, Thomar et al. 1989). PCR products were visualized in 2% agarose gels stained with SYBR Safe.

2.3.4. Amplification and Identification of the Sources of Blood Meals Intestinal content DNA samples were amplified with the primers set FACytb, following the amplification conditions described in Table 2.1. The expected 335 pb band was visualized in 2% agarose gels after SYBR Safe staining. Amplification products were sent for sequencing to Macrogen, Inc. (Korea). Forward and reversed sequences were aligned with Mega V6.0 (https://www.megasoftware.net/manual.pdf). Consensus sequences were compared with DNA nucleotide sequences in NCBI BLAST using the 23 megablast query. Matches were based on results achieving ≥ 95% maximum identity and cover.

Table 2.1. Thermal cycling parameters of PCR primers tested for sensibility and vertebrate detection from triatomine intestinal content DNA.

Ta Step and conditions No. of Product Primer set (°C) (°C/sec)ᵃ cycles size (bp)

Cytochrome b

FA-CYTB-F, FA-CYTB-R 52.7 D,95/30; A,55/50; E,72/40 36 355

Cytochrome b

L14841, H15149 56.8 D,93/60; A,50/60; E,72/300 30 307

12S rRNA

L1091, H1478 55.4 D,93/60; A,50/60; E,72/300 30 386

Ta. Optimum annealing temperature calculated by using Thermo Fisher Tm calculator. a. Actual denaturation (D), annealing (A) and extension (E) temperatures and times used in cited publications.

2.4. Results

2.4.1. Sensitivity of the Different Primer Sets The best performance of primer sensitivity using human DNA serial dilutions was achieved for the primers FA-CYTb, where the amplification of a 335 pb fragment was obtained using up to 1 pg of DNA. For the two other primer sets (L14841-H15149 and

L1091-H1478) the expected bands were obtained up to 10 pg of DNA.

2.4.2. Vertebrate Blood Meal Source Diversity Of the 507 samples analyzed (Appendix A), blood meal was identified in 82.05% of the cases; while PCR products were not obtained for 10.45% (n = 53) of the samples. In

3.5% (n = 16) of the samples, the species could not be identified due to sequences with <

95% identity. Within these samples, sequences from birds (Gallus gallus), primates

(Homo sapiens) and rodents (Rhipidomys leucodactylus, Scimosciurus nebouxii, and

Rattus rattus) were found. All these species have been reported in Ecuador. Only the rodent species R. leucodactylus was not reported in the list of species with ˃95% identity although the complete cytb gene is available in GenBank. Non legible chromatograms or the presence of multiple peaks were present in 4.33% (n= 22) of the samples.

The blood of sixteen vertebrate species was identified (Appendix B). Mammals represented 67.8% of the analyzed samples with nine species including five rodents

(Aegialomys xanthaeolus, Mus musculus, Rattus norvegicus, R. rattus, Scimosciurus nebouxii (named before Sciurus stramineus), one carnivore (Canis lupus familiaris), two artiodactyla (Capra hircus, Sus scrofa), one didelphimorpha (Didelphis marsupialis) and primate Homo sapiens (Figure 2.2). Four species of birds were detected (Gallus gallus,

Campylorhynchus fasciatus, Columba livia and Leptotila verreauxi decolor) and 25 represented 31% of the analyzed samples (Figure 2.2). Reptiles (Boa constrictor imperator) and amphibians (Bufo marinus) were found in a small percentage (<1%)

(Figure 2.2). In general, humans (H. sapiens), chicken (Gallus gallus) and rodents (S. nebouxii and R. rattus) were the most abundant vertebrate host (Figure2. 2).

Figure 2.2. Proportion of blood meal source (species identification) of 416 triatomines collected from central coastal and southern highlands in Ecuador.

2.4.3. Blood Meal Diversity and Trypanosoma cruzi Infection in the Different Environments Of the 416 identified blood meal samples, 65.8% presented infection with T. cruzi.

Triatomines that contained mammal and bird blood presented highest infection rates

(64.2% and 69.2%, respectively) (Table 2.2; Appendix A). From the samples with T. cruzi infection, no difference was found between infection of adults (n= 166) and nyphs

(n= 177).

Triatomines from sylvatic environments presented 11 species of hosts (seven species of mammals and five species of birds). Infection rate with T. cruzi reached 61.1%, being triatomines that contained bird blood the ones with highest rate (73.1%) while the ones that have fed from mammals presented 58.8% of infection. In peridomestic triatomines, 26 eleven species were identified as blood meal (seven mammal, two bird, one reptile and one amphibian species); and infection rate with T. cruzi was of 72.7%. Within peridomestic triatomines, the ones were mammals and birds blood was detected, presented similar infection rates with T. cruzi (75.3% and 70.3%, respectively), while triatomines that had fed from reptile and amphibian blood had also high infection rates

(100% and 50%, respectively) although these samples only represented <1% of the analyzed samples. In the domestic triatomines, five species were identified (four mammal and one bird species) with 61% of infection rate with T. cruzi. In this environment, infection rate in triatomines and birds were high (62.3% and 53.8%, respectively) (Table

2.2, Figure 2.3).

Human (Homo sapiens), dog (Canis lupus familiaris), the black rat (Rattus rattus) and chicken (Gallus gallus) blood meals were detected in the intestinal content of triatomines collected in all three environments, all of them presented T. cruzi infection > 50% (Table

2.2, Figure 2.3). The yellowish rice rat (Aegialomys xanthaeolus), the Guayaquil squirrel

(Simociurus nebouxii) and the Fasciated Wren (Campylorhynchus fasciatus) were found mainly associated with sylvatic triatomines, although, they were also detected in peridomiciliary insects. High infection rate with T. cruzi was detected in sylvatic triatomines that contained the Guayaquil squirrel blood (50%) and the Fasciated Wren

(71.4%) (Table 2.2, Figure 2.3). Other species that constitute a source of blood for sylvatic triatomines are the Rock Pigeon (Columba livia), the white-tipped dove

(Leptotila verreauxi decolor), the opossum (Didelphis marsupialis) and mouse (Mus musculus). All of them, but the triatomines were mouse blood was detected, presented 27 infection rates over 66% (Table 2.2, Figure 2.3). Although blood of pig (Sus scrofa), brown rat (Rattus norvegicus), boa (Boa constrictor imperator) and the cane toad (Bufo marinus) was found in few peridomestic triatomines (n = 8), high infection rate with T. cruzi was detected (> 50%). The goat (Capra hircus) was detected in triatomines collected in the intradomicile with 62.5% of T. cruzi infection (Table 2.2, Figure 2.3).

Table 2.2. Diversity of feeding host species identified in triatomines collected in intradomicile, peridomicile and sylvatic environments in Ecuador.

Vertebrate source of blood I P S TOTAL Species Common name n (%) n (%) n (%) n (%) Mammals Capra hircus Goat 8 (62.5) - - 8 (62.5) Sus scrofa Pig - 1 (0.0) - 1 (0) Canis lupus familiaris Dog 2 (100.0) 2 (50) 1 (100.0) 5 (80.0) Didelphis marsupialis Opossum - - 1 (100.0) 1 (100.0) Homo sapiens Human 44 (54.5) 48 (81.3) 77 (63.6) 169 (66.3 ) Aegialomys xanthaeolus Yellowish rice rat - 1 (0) 3 (0) 4 (0) Mus musculus Mouse - - 1 (0) 1 (0 ) Rattus norvegicus Brown rat - 3 (100.0) - 3 (100.0) Rattus rattus Black rat 15 (80.0) 19 (68.4) 7 (85.7) 41 (75.6) Simosciurus nebouxii Guayaquil - 3 (66.7) 46 (50.0) 49 (51.0) squirrel Total mammals 69 (62.3) 77 (75.3) 136 (58.8) 282 (64.2) Birds Columba livia Rock Pigeon - - 4 (100.0) 4 (100.0) Leptotila verreauxi decolor White-tipped ove - - 3 (66.7) 3 (66.77) Gallus gallus Chicken 13 (53.8) 90 (71.1) 12 (66.7) 115 (68.7) Campylorhyncus fasciatus Fasciated Wren - 1 (0) 7 (71.4) 8 (62.5) Total birds 13 (53.8) 91 (70.3) 26 (73.1) 130 (69.2) Reptiles Boa constrictor imperator Boa - 2 (100.0) - 2 (100.0) Total reptiles - 2 (100.0) - 2 (100.0) Amphibians Bufo marinus Cane toad - 2 (50.0) - 2 (50) Total amphibians - 2 (50.0) - 2 (50) TOTAL 82 (61.0) 172 (72.7) 162 (61.1) 416 (65.9) I: intradomicile, P: peridomicile, S: sylvatic.

Figure 2.3. Infection rates of triatomines captured in intradomiciliary (intra), peridomiciliary (peri) and sylvatic (sylv) environments with the diversity of vertebrate host that are sources of blood for triatomines.

2.4.4. Vertebrate Blood Meal Source per Triatomine Species, Stages and Collection Environment Five species of triatomines were included in the analysis: Panstrongylus chinai, P. howardi, P. rufotuberculatus, Rhodnius ecuadoriensis and Triatoma carrioni. Rhodnius ecuadoriensis individuals were collected in all three environments (intradomiciliar, peridomiciliar and sylvatic) in both geographic regions. While P. chinai, P. rufotuberculatus and T. carrioni were only collected in southern highlands, in the intradomiciles, and some P. chinai (n = 4) in the peridomicile. Individuals of P. howardi were collected in central coastal region, mainly in the peridomiciles (Figura 2.4a). A 29 similar proportion of nymphs (45%) and adults (55%) were analyzed. In both groups, human, chicken and rodents (R. rattus and S. nebouxii) were the main source of blood.

However, more diversity of vertebrates were found in nymphs (15 species) compared to adults (10 species) (Figure 2.4b).

In central coastal region, highest diversity species as blood meal was detected in R. ecuadoriensis (11 species). Of them seven species were mammals (S. nebouxii, R. rattus,

R. norvegicus, H. sapiens, D. marsupialis, Canis lupus, A. xanthaeolus) and four birds (L. verreauxi decolor, G. gallus, C. livia, C. fasciatus) (Figure 2.4b). Five species were found to be source of blood for P. howardi, mainly rats (R. rattus) and humans (H. sapiens) and even the pig (S. scrofa), although in lesser extent. However, non-typical sources of blood for triatomines such as toad (B. marinus) and snake (B. constrictor imperator) were also found in this species (Figure 2.4c).

In southern highlands, humans are the main source for triatomines, particularly for the ones collected in the intradomicile (P. rufotuberculatus and T. carrioni) (Figure 2.4b).

However, for species that were found also in peridomestic and sylvatic environments (P. chinai and R. ecuadoriensis), although humans are important source, the diversity of other source of food varies between vector species. In R. ecuadoriesnis, other species that are important source of blood are chicken (G. gallus), squirrel (S. nebouxii) and rats (R. rattus) (Figure 2.4c). For P. chinai, important source of blood are rats (R. rattus), although goats (C. hircus) were detected in triatomines collected in the intradomicile of a single house (Figure 2.4c, Appendix A). 30

Triatomine habitats are diverse; triatomines collected in sylvatic environments were associated with animal shelter (organic matter in palm trees, bird and rodent nest) and presented the greater diversity of sources of blood (10 species) being humans and squirrel the principal species. Other species collected in this environment included rodents (R. rattus, M. musculus, A. xanthaeolus), marsupial (D. marsupialis) and birds (L. verreauxi decolor, G. gallus, C. livia and C. fasciatus) (Figure 2.4d, Appendix A). Most of peridomestic triatomines were associated with animal shelter, particularly chicken nest but also of rodents (rat, mouse, guinea pig). Animal shelter was the place were most triatomines were collected, and seven vertebrate species were identified as sources of blood meal (S. nebouxii, R. rattus, C. lupus, C. fasciatus, H. sapiens, G. gallus), particularly human and chicken. By the other side, triatomines found in pile of material as bricks, tile and wood were fed mainly on rodents (R. rattus, R. norvegicus, A. xanthaeolus) and in a less extent human, chicken, pig, toad and snake (Figure 2.4d,

Appendix A).

Intradomicile triatomines were mostly found in the bedroom; bellow the bed, between clothes and on the wall. The main source of blood was human, although other species were also found such as chicken, rat and dog (Figure 2.4d, Appendix A). Pile of organic material inside of the house was collection places for triatomines fed mainly from rats.

Triatomines collected in animal shelter (chicken nest and guinea pig) were found to have been fed from human and chicken blood. (Figure 2.4d, Appendix A).

Figure 2.4.Vertebrate host feeding pattern in triatomines a) diversity of triatomines analyzed and environment of collection, b) vertebrate diversity found in nymphs and adults, c) diversity of vertebrate blood by species and geographic area, and d) diversity of host sources by structure and environment of collection.

2.5. Discussion

Triatomines feed on a wide diversity of vertebrate species, and the detection of these hosts in a given area is essential to understand the interactions between triatomines and vertebrate hosts. In Ecuador, wide information has been obtained about the diversity and ecological conditions of triatomines and T. cruzi (Ocana-Mayorga, Llewellyn et al. 2010,

Costales, Jara-Palacios et al. 2015, Grijalva, Villacis et al. 2015, Grijalva, Villacis et al.

2017). This information has been crucial to define target species of triatomines for their control but also to evaluate the effectiveness of control strategies. Indeed, limited effectiveness of the insecticide spaying was observed (Grijalva, Villacis et al. 2011) likewise the continuous reinfestation of houses constitute a challenge for effective measures to avoid the contact human-vector. Lately, a first initiativa has been taken with a community-based perspective, which is based in the improvement of house conditions

(Nieto-Sanchez, Bates et al. 2019). To find alternative measures to fight against human contact with triatomines, information about the interactions between the triatomines and vertebrate hosts is needed to define 1) the main source of blood for triatomines, 2) its availability in the area and 3) the circumstances that are promoting the human-vector contact.

The current feeding patterns of triatomines in central coastal and southern highlands regions in Ecuador established the presence of 16 species of vertebrates, being mammals and birds the most important sources of blood. Among them, humans and chicken constitutes the most frequent sources of blood (68.3%). Are those hosts preferred by triatomines? or is their availability greater to that of other mammals in the studied areas? 33

The possible implication of the currently identified host species in T. cruzi transmission cycle and their interaction with humans are discussed in the following sections.

2.5.1. Diversity of Mammal Hosts and its Implication in Trypanosoma cruzi Transmission Mammal species are the most frequent sources of blood found and the most important due to they can be infected and transmit T. cruzi. In this study, mammals contributed with

67.8% of the identified feeding sources, of them, triatomines fed from human, squirrel (S. nebouxii) and black-rat (R. rattus) represented 91.8% of the samples. All three species were found in triatomines collected in human habitat, and also in sylvatic environments.

Human blood has been previously found in the three principal triatomine genus implicated in T. cruzi transmission (Panstrongylus, Rhodnius and Triatoma)

(Christensen and de Vasquez 1981, Carrasco, Torrellas et al. 2005, Cantillo-Barraza,

Garces et al. 2015, Cecere, Leporace et al. 2016); however, the presence of human blood in sylvatic triatomines (especially associated with squirrel nests) was unexpected.

Moreover, high infection rate found in this study (66%) confirm previous reports

(Grijalva, Suarez-Davalos et al. 2012, Grijalva, Teran et al. 2014). Contact of humans with sylvatic triatomines has been proposed to be related to outdoor activities (e.g. camping, harvesting) or to a continuous migration of triatomines from sylvatic to domestic settings and vice versa (Buitrago et al. 2013, Waleckx, Suarez et al. 2014,

Buitrago, Bosseno et al. 2016). In the present context, such human activities and triatomine dispersal need to be further studied.

Endemic and wild species of rodents are important sources of blood for triatomines particularly in sylvatic environments (Bosseno et al. 2009, Almeida, Faucher et al. 2016, 34

Buitrago, Bosseno et al. 2016, Chacon, Bacigalupo et al. 2016). In this study, apart from human, triatomines mostly fed from the squirrel (S. nebouxii) and few other endemic species. In previous studies, triatomines have been searched in burrows, tree holes, under trunks and rocks, but they were mainly collected in nests built in trees, particularly in squirrel nest (Grijalva and Villacis 2009, Suarez-Davalos, Dangles et al. 2010). Thus, these results support the importance of S. nebouxii in the maintenance of triatomine populations in forest patches around rural communities (Ocana-Mayorga, Lobos et al.

2018).

Synanthropic rodents such as rats (R. rattus) have been found to be key sources of blood for triatomines in some areas such as Mexico, Guatemala and Bolivia (Bosseno et al. 2006, Bustamante, De Urioste-Stone et al. 2014, Breniere et al. 2004, Buitrago,

Bosseno et al. 2016). The results of this study showed that rat blood represented 15.6% of the feeding source of triatomine being collected in all three environments. Although rats have the capacity to circulate between sylvatic and domestic environments, rat contact with triatomines might be more related to intra and peridomiciliary environments.

Moreover, the presence of T. cruzi in triatomines that have fed from rat blood indicates and active transmission of the parasite. Although rats are widely distributed, its role in T. cruzi transmission depends on the ecological and social characteristics of a particular area. For example, studies of feeding patterns of triatomines in some areas of Panamá,

Argentina, Venezuela and Colombia, demonstrated that rats are not an important sources of blood for triatomines (Christensen and de Vasquez 1981, Carrasco, Torrellas et al.

2005, Cantillo-Barraza, Garces et al. 2015, Cecere, Leporace et al. 2016). On the other 35 hand, in Mexico, rats play a major role for development and feeding source of peridomicile colonies of T. longipennis and shoud be the main reservoir (Bosseno et al

2006). Similarly, in eastern Guatemala, rodent control was applied as part of a strategy for an integrated vector management, and a major reduction of triatomine infestation was obtained where rodent and triatomine control was applied (De Urioste-Stone, Pennington et al. 2015). In Ecuador, a similar approach could be applied; however, the presence of rats varies according to regions. In southern highlands, rats (Rattus spp.) are the 35% of the small mammals collected in the intra and peridomicile environments; while in the central coastal area, rats represented only 11% of the collected mammals (Ocaña-

Mayorga, unpublished data).

Based on the current results, opossums seem to have limited role on triatomine maintenance and T. cruzi transmission in our studied areas, although this is an abundant species in the area (Ocaña-Mayorga, unpublished data). The opossum is considered a natural host species for the parasite and it is an important reservoir in some areas in

Colombia and Brazil (Travi, Jaramillo et al. 1994, Yeo, Acosta et al. 2005, Bosseno et al.

2006, Cantillo-Barraza, Garces et al. 2015). By the other hand, the role of dogs needs to be further evaluated. In Latin America, particularly in Chagas disease endemic areas, there is a high frequency of dogs infected with T. cruzi and their presence of in households has been associated with higher risk of human infection (Travi, Jaramillo et al. 1994, Xavier et al. 2012). In Colombia, dogs have been found as an important source of blood for T. maculate (Cantillo-Barraza, Garces et al. 2015). In other areas such as

USA (southern of Texas and Louisiana) and Brazil, T. cruzi seroprevalence of dogs 36 ranged from 7 to 76% (Curtis-Robles, Snowden et al. 2017, Freitas, Souza et al. 2018,

Porfirio, Santos et al. 2018, Elmayan, Tu et al. 2019). In northern Argentina, infected dogs constitute a risk factor and have been used as sentinels for T. cruzi domestic transmission (Gurtler, Cecere et al. 1991, Saldana, Calzada et al. 2015). However, in other areas, dogs might not have such an important role, although they have been found to be a source of blood but in a lesser extent ((Mota, Chacon et al. 2007, Arce-Fonseca,

Carrillo-Sanchez et al. 2017). In endemic areas of Chagas disease in Ecuador, dogs are present in the majority of domiciles (Grijalva, Villacis et al. 2015, Grijalva, Villacis et al.

2017). The results of this study indicated that dogs are not a main source of food although its availability.

Other mammals such as the mouse (M. musculus), the brown rat (R. norvegicus), goats

(C. hircus) and pigs (S. scrofa) might be occasional source of food and could be playing a limited role on T. cruzi transmission.

2.5.2. Birds and its Role in Triatomine Population Maintenance Birds do not have a direct role in T. cruzi transmission due to they are refractory to the parasites; they are able to destroy the typomastigote form through the action of their immune system. However, they are a feeding source that plays an important role in survival and maintenance of triatomine populations (Jansen, Roque et al. 2017). Raising chickens in endemic areas for Chagas disease is frequent, thus they are an available source of blood. Experimental studies in T. infestans have demonstrated that this vector species prefers feeding from dog that from chicken (Gurtler, Ceballos et al. 2009); however, in natural conditions a high frequency of triatomines fed from chicken. In

Ecuador having chickens was found to be a main determinant of triatomine infestation 37

(Grijalva, Villacis et al. 2015, Dumonteil, Herrera et al. 2016, Grijalva, Villacis et al.

2017). Most chickens are kept free ranging, except laying hens whose nests are outside against the house walls to be easily monitored by the inhabitants (Patterson, Bates et al.

2018). Large colonies of triatomines have been found in these nests. Although chicken is the expected host for triatomines found in chicken nests, we also found human and rodent blood meals. Moreover, triatomines that contained chicken blood were found infected by

T. cruzi. Thus, dispersal of triatomines among environments might be playing an important role in transmission.

In sylvatic environments, rodents have been described as as important source of blood for triatomines (Buitrago, Bosseno et al. 2016, Chacon, Bacigalupo et al. 2016). Other birds such as doves, pigeons and other birds might contribute with the maintenance of sylvatic populations of triatomines. However, little information is available about the role of birds and other vertebrates in triatomine population maintenance in sylvatic environments.

2.5.3. Triatomine Species and Blood Meal Preference Rhodnius ecuadoriensis is the main species found colonizing domestic and sylvatic environments (Grijalva, Villacis et al. 2015, Grijalva, Villacis et al. 2017). Thus, it is expected that this species presented a higher diversity of vertebrate species as source of blood; however, P. howardi and P. chinai; that are more restricted to domicile and peridomicile, also have a diverse source of vertebrates as source of food. In the central coastal region, this species presented higher blood host diversity than in southern highlands individuals. These differences can be explained by different ecological conditions, as the rate of intradomiciliar infestation and the access or preference to 38 different species of host. For P. chinai, P. rufotuberculatus and T. carrioni that were mainly associated to the intradomiciliary environment, human blood meals are expected.

However, and unlike of what was seen in R. ecuadoriensis, P. chinai presented a more diverse diet (including goats) but limited feeding with birds, especially chickens, that are one of the most abundant species in peridomestic environments. A similar pattern was seen in P. howardi in which no chicken blood was found, however non-common hosts

(snakes, toed and pigs) were detected. These results might indicate a preference for particular sources of blood driven by triatomine species.

2.5.4. Triatomine Dispersal and Access to Vertebrate Hosts The detection of multiple meals in a single triatomine contributes to understand triatomine movements but in our case, the technique used did not allow this level of analysis. In fact, in the case of multiple meals, the sequence obtained after Cytb-PCR is a mixture of sequences that can be resolved only by molecular cloning what was not realized. Another criterion that informs the movements of triatomines is the relationship between their blood meal and place of capture. For example, triatomines collected in chicken nests in the peridomicile and squirrel nests in the sylvatic environment presented a high diversity of sources of blood (10 and 11 species, respectively), showing that these triatomines had fed elsewhere before or after arriving in these habitats. Furthermore, triatomines collected in all environments, including sylvatic, showed the presence of blood samples from human and chicken.

Moreover, it is important to consider than nymphs have a limited capacity for dispersion due to the lack of wings, however, they presented higher diversity of sources of blood (15 species) compared to adults (10 species). This result probably indicates that 39 nymphs might be using other strategies such as cleptohematophagy (stealing blood from a recently fed congener) in searching of sources of blood or even they are passively dispersed by transportation by animals or humans.

Host accessibility is a major factor that shapes triatomine the patterns of sources of blood, and its accessibility is variable according to the area studied (Rabinovich, Kitron et al. 2011). In Panamá, anthropogenic landscape disturbance changed the host community structure and increased R. prolixus infection with T. cruzi (Gottdenker,

Chaves et al. 2012). By the other side, in Argentina, domestic animals and its availability along the year affect host-feeding choices of triatomines and even human-vector contact rates (Gurtler, Cecere et al. 2014). In Ecuador, triatomines have accessibility to domestic animals such as dogs and chickens (Grijalva, Villacis et al. 2017). Although chickens are an important source of blood for triatomines, humans constitutes the most important source of blood. Moreover, it is important to consider than nymphs have a limited capacity for dispersion due to the lack of wings, however, they presented higher diversity of sources of blood (15 species) compared to adults (10 species). This result probably indicates that nymphs might be using other strategies such as cleptohematophagy, coprophagy or even passive dispersion to move in searching for different sources of blood.

Dispersal and dynamics of triatomines, particularly R. ecuadoriensis need to be further evaluated to determine the factors that are influencing the continuous human-vector contact in domestic but also in sylvatic environments in Ecuador.

2.6. Conclusions

Humans are a main source of food for the sampled triatomines species in Ecuador, indicating that the vector-human contact is more frequent than previously thought. This result together with the high infection rates with T. cruzi indicates a high risk of transmission of the parasite in central coastal and southern highlands regions of Ecuador.

Moreover, the high frequency of chicken blood in peridomiciliary triatomines indicates that this species is fundamental in the maintenance and thriving of triatomine populations in household environments. R. ecuadoriensis, which is the most abundant species in both studied regions and widely distributed in domestic and sylvatic environments, appeared to prefer human and chicken blood. Moreover, other species that colonized peridomestic environments such as P. chinai and P. howardi might prefer human and rodent sources of blood but not chicken. Finally, it is important to consider that although nymphs cannot fly, other mechanisms that remain to be elucidated, allowed them to feed from diverse blood meal sources increasing their probability to acquire the infection. The dispersion profile of triatomines is obviously very complex. They are influenced by the availability of hosts, their own trophic preferences, but also environmental disturbances that lead them to move and adapt to new sources of food. In this context, dispersal patterns need to be evaluated to propose and effective strategy that limits the vector-human contact and to decrease the risk of T. cruzi transmission.

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CHAPTER 3: HUMAN BLOOD MEALS IN SYLVATIC TRIATOMINES

CHALLENGES DOMESTIC-CENTERED STRATEGIES FOR PREVENTION OF

TRYPANOSOMA CRUZI TRANSMISSION IN ECUADOR

3.1. Abstract

Generally, the risk of transmission of Trypanosoma cruzi has been associated with the contact human-triatomines in domestic environments, particularly with triatomine populations that have been able to adapt and colonize intradomiciliary environments. In fact, control strategies such as spraying of insecticide have seen to be effective in controlling domiciliary populations of triatomines. However, alternative scenarios in which human and triatomines could be in contact have not been elucidated. To unveil these scenarios, we evaluated the frequency of human blood meal in the intestinal content of triatomines collected in domiciliary, peridomiliciary and sylvatic environments in central coastal and southern highlands of Ecuador. The amplification of a region of the cytochrome oxidase gen demonstrated that human blood meal was present in 42% of the samples. In domestic and peridomestic environments, the contact of triatomines with humans is more likely to occur. However, the presence of human blood meal in triatomes collected in chicken nest (29%) indicated that there might be a flow of triatomines between the intradomicile and peridomicile in search of sources of blood. Moreover, an infection rate with T. cruzi was > 20%. In this scenario, house improvement to avoid the entrance of vectors and insecticide spraying can be effective to avoid transmission.

However, our results indicated that applying control strategies in intradomiciliary and peridomiciliary environments might not avoid T. cruzi transmission in Ecuador. Even if 48 sylvatic triatomines are less likely to be in contact with human, 48%, including adults and nymphs, had human blood meal and T. cruzi infection rate reaches 30%. These results indicate that humans are not accidental host, but an important source of blood for triatomines in all environments, and with high risk of T. cruzi transmission.

Consequently, domestic-centered strategies might not prevent vectorial transmission and data of Chagas disease cases might be underestimated. Further studies need to be conducted to unveil the mechanisms by which humans are in contact with triatomines, including dispersal capacity of triatomines and community-based research to evaluate outdoor activities as risk factors for parasite transmission in Ecuador.

3.2. Background

Hematophagy is one of the main adaptations of some insects. This adaptation has evolved independently at least 21 times in arthropods (Black and Kondratieff, 2005). The study of some of the most important vector-borne diseases depends on the understanding of the interactions among the host, the parasite and the vector. In this regard, studies of blood meal sources have been carried out in insects of medical importance such as ticks, sand flies and triatomines (Allan, Goessling, Storch and Thach, 2010; Baum et al. 2015,

Mota et al. 2007). Triatomines are obligatory hematophagous insects of epidemiological importance since they transmit the parasite Trypanosoma cruzi that causes Chagas disease in humans. For T. cruzi, three overlapping cycles have been defined: domestic, peridomestic and sylvatic. Although some differences exist in their delimitations, generally the domestic cycle refers to transmission of T. cruzi that occurs to hosts that are living inside the dwellings. The peridomestic cycle refers to the transmission to domestic and synanthropic mammals that occupy or circulate in areas and man-made structures that surround houses (delimited by natural or artificial fences) (Walter et al. 2007). The sylvatic cycle refers to transmission to non-domestic mammals occurring outside of the domestic and peridomestic areas where human activities (e.g. harvesting) can take place or not. Until now, vector transmission to humans has been mostly attributed to indoor contact with infected feces of triatomines (Breniere, Villacis and Aznar, 2017). Previous studies on vector feeding sources have reported a high percentage of human blood in triatomines found in domiciliary and peridomiciliary environments (Bosseno et al. 2006,

Cantillo-Barraza et al. 2015, Carrasco, Torrellas, Garcia, Segovia and Feliciangeli, 2005; 50

Cecere et al. 2016). In fact, triatomine species that adapt and colonize human dwellings are more likely to contribute to human infection with T. cruzi (Waleckx, Gourbiere and

Dumonteil, 2015) and its elimination is a main component of the strategies applied to control and prevent Chagas disease transmission (Gorla and Hashimoto, 2017).

Furthermore, some domestic populations of triatomines have been or are close to being eliminated in regions where these species are not endemic (e.g. Triatoma infestans,

Rhodnius prolixus, T. dimidiata) (Gorla and Hashimoto, 2017). However, previous reports have challenged the idea of human transmission being restricted to indoor environments. Human blood have been detected in T. rubida, T. protracta (Stevens et al.

2012), T. sanguisuga (Waleckx, Suarez, Richards and Dorn, 2014) and T. infestans (N. L.

Buitrago et al. 2013) collected in wild environments.

In Ecuador, important advances in understanding the diversity of biological features of triatomines and the ecological conditions in different endemic areas for Chagas disease have been made (Grijalva et al. 2017, Grijalva et al. 2015). Under some conditions, a limited efficacy of standard insecticide-based control and prevention strategies has been reported (Grijalva, Villacis, Ocana-Mayorga, Yumiseva and Baus, 2011). As a consequence, alternative prevention models than include community-based strategies are being studied (Marco-Crespo, Casapulla, Nieto-Sanchez, Urrego and Grijalva, 2018;

Nieto-Sanchez et al. 2019, Nieto-Sanchez, Baus, Guerrero and Grijalva, 2015; Patterson,

Bates, Chadwick, Nieto-Sanchez and Grijalva, 2018). However, there is still a gap of knowledge about biological characteristics of Chagas disease transmission, which poses a threat for the success of transmission control/prevention strategies. In this regard, the aim 51 of this study is to determine the presence of human as source of blood in triatomines collected in domestic, peridomestic and sylvatic environments, and the vectorial transmission scenarios taking place in endemic areas for Chagas disease of southern and coastal Ecuador.

3.3. Methodology

3.3.1. Study Area and Triatomine collection Triatomines were collected by manual search in domicile, peridomicile and sylvatic environments. Analyses were carried out with 416 intestinal content samples from triatomines (228 nymphs and 188 adults) collected from 2006 to 2012 in two regions of

Ecuador: central coastal (n = 146) and southern highlands (n = 270). Triatomine species included Panstrongylus chinai, P. howardi, P. rufotuberculatus, Triatoma carrioni and

Rhodnius ecuadoriensis (Figure 3.1).

Intestinal content of each triatomine was obtained by dissection, following the procedure described in (Grijalva, Suarez-Davalos, Villacis, Ocana-Mayorga and Dangles,

2012) and stored at -20°C in PBS. Triatomine species, developmental stage, environmental data and microenvironment were recorded for each triatomine (Appendix

C).

Figure 3.1. Geographical origin of the 416 samples included in this study from central costal and southern highlands of Ecuador.

3.3.2. Amplification and Host Identification of Sources of Blood DNA was obtained with the DNeasy kit (Qiagen, Germantown, United States). The region of cytb was amplified with cytb FA-CYTB-F (5´-CCC CTC AGA ATG ATA TTT

GTC CTC A-3´) and FA-CYTB-R (5´-CCA TCC AAC ATC TCA GCA TGA TGA AA- 54

3´) according to previous conditions (Buitrago et al. 2012). PCR products were visualized in 2% agarose with SYBR Safe and double strand sequenced (Macrogen, Korea).

Forward and reversed sequences from each sample were edited with Mega V6.0

(https://www.megasoftware.net/manual.pdf). Consensus sequences were compared with

DNA nucleotide sequences in NCBI BLAST using the megablast query. Matches were based on results achieving ≥ 95% of identity.

3.3.3. Natural Infection of Triatomines with Trypanosomes Infection with T. cruzi was assessed by PCR size products of the conserved domain of the minicircle of kinetoplastid DNA (kDNA). A band of 300 bp was expected for T. cruzi following the procedures described in (Ocana-Mayorga et al. 2015).

3.3.4. Haplotype Net for Human Sequences Haplotype diversity of the sequences determined as human blood meal was assessed.

Sequences were analyzed by using median-joining algorithm (Bandelt, Forsterand Rohl,

1999) implemented in Network 4.6 (Fluxus Technology Ltd, United Kingdom). The haplotype network was performed based on polymorphic sites in an alignment of a subset of 93 sequences identified as human cytb gene fragment in the BLAST search. Sequences with a length of 164 bp were used in the alignment for the analysis.

3.3.5. Feeding Behavior of R. ecuadoriensis Nymphs in Laboratory Conditions To test the possibility of cleptohematopaghy, recently feed adults (one or two) were exposed to four to six nymphs NIII and NIV. Previous to the experiment, adults and nymphs were kept apart and were offered pigeon as feeding source. After four weeks of starvation, only the adults were offered a mouse as source of blood. Starving nymphs were placed in a container with one or two adults for 72 hours. After that time, intestinal 55 contents of all individuals (adults and nymphs) were taken and cytb sequence were amplified and analyzed as previously described. This procedure was repeated five times and a negative control only fed once with pigeon was included.

3.4. Results

3.4.1. Detection of Human Blood Meal and Trypanosoma cruzi in Triatomines, According to Regions and Environments. Of 416 blood meals identified, 40.6% were from human (n = 169). Non-human blood included mammals (n = 113, 27%), birds (n = 130, 31%), amphibian (n = 2, 0.5%) and reptiles (n= 2, 0.5%) (Appendix C).

Human blood sequences were detected in all five species of triatomines, of which

16.1% were detected in central coastal region and 24.5% in southern highlands (Table

3.1, Figure 3.2). Rhodnius ecuadoriensis was the most abundant species, representing

83.9% (n = 349) of the total analyzed samples. Although, both P. chinai (n = 51, 12%) and P. howardi (n = 13, 3%) represented only 15.4% of the analyzed samples, human blood was detected in more than 30% of the samples for each species. All the three samples from P. rufotuberculatus (n = 1) and T. carrioni (n = 2), collected in intradomiciles (bed), human blood was detected (Table 3.1, Figure 3.2).

All five triatomine species collected in the intradomicile were tested and 54.5% (n =

44) were positive for human blood (Table 3.1, Figure 3.2). Higher percentage of human blood meal was detected in introdomicile samples from southern highlands (50.6%) compared with central coastal (3.7%). Although the majority of analyzed samples were collected in the peridomicile (n = 173), only 27.7% (n = 48) presented human blood meal

(Table 3.1, Figure 3.2). A similar pattern was found in the sylvatic environment, where only R. ecuadoriensis was collected. Of the 162 analyzed individuals, 47.5% were detected with human blood (Table 3.1, Figure 3.2) with a similar proportion between 57 southern highlands and central coastal (23.5% and 24.1%, respectively) (Table 3.1,

Figure 3.2).

Assessment of T. cruzi infection indicated that 66.3% (n = 112) of the samples with human blood were positive. Rhodnius ecuadoriensis, the species with the majority of human blood samples (n = 138), had 70.3% of T. cruzi infection. Infection rates >50% were found in P. chinai, P. howardi, P. rufotuberculatus and T. carrioni, however, the total number of individuals analyzed from these species was low (n = 31). T. cruzi infection rate was high in all environments (> 50%), being the peridomicile the environment that presented the highest rate (81.3%) (Table 3.1, Figure 3.2). Infection with T. rangeli was found in 13% of the triatomines were human blood were detected

(including mixed infections with T. cruzi).

Table 3.1. Triatomine species and T. cruzi infection of the 168 samples where human blood meal were detected in central coastal (CC) and southern highland (SH) regions.

Human blood meal Non- Triatomine species/ T. cruzi human Total environment CC (%) SH (%) Total (%) infection blood meal (%) P. chinai 24 (47.1) 24 (47.1) 10 (41.7) 27 51 Domicile 21 21 8 25 46 peridomicile 3 3 2 2 5 P. howardi 4 (30.8) 4 (30.8) 3 (75.0) 9 13 Domicile 2 2 2 0 2 peridomicile 2 2 1 9 11 P. rufotuberculatus 1(100.0) 1 (100.0) 1 (100.0) - 1 Domicile 1 1 1 - 1 R. ecuadoriensis 63 (18.1) 75 (21.5) 138 (39.5) 97 (70.3) 211 349 Domicile 1 17 18 12 12 30 peridomicile 23 20 43 36 114 157 Sylvatic 39 38 77 49 85 162 T. carrion 2 (100) 2 (100) 1 (50) - 2 Domicile 2 2 1 - 2 Total 67 (16.1) 102 (24.5) 169 (40.6) 112 (66.3) 247 416 Domicile 3 (3.7) 41(50.6) 44 (54.3) 24 (54.5) 37 81 peridomicile 25 (14.5) 23 (13.3) 48 (27.7) 39 (81.3) 125 173 Sylvatic 39 (24.1) 38 (23.5) 77 (47.5) 49 (63.6) 85 162 58

Figure 3.2. Human blood meal rates for each triatomine species ands and collection environment: intradocimiciliary (intra), peridomiciliary (peri) and sylvatic (sylv).

3.4.2. Population Structure, Microenvironment and Trypanosoma cruzi Infection of Triatomines with Human Blood Meals Of the 169 samples with human blood meal, we found similar proportion of nymphs

(50.8%) and adults (49.1%), as well T. cruzi infection of 32% for nymphs (n = 54) and

35% for adults (n = 58) (Figure 3.3a).

In the domicile, triatomines were collected mainly in bedrooms (in beds or on wall), particularly in southern highlands (Figure 3.3b). Both nymphs and adults of all the triatomine species in this environment presented infection with T. cruzi > 30% (Figure

3.3a). In the peridomicile, triatomines from both regions were mainly associated to animal shelters, in particular to chicken nests (Figure 3.3b). Infection rates of nymphs was > 50% in P. chinai and P. howardi, while in R. ecuadoriensis higher infection rate was recorded in adults (53%) than in nymphs (30%) (Figure 3.3b). In sylvatic environments, R. ecuadoriensis, in both regions, was found associated to animal shelter

(Figure 3.3b), mainly to squirrel nest, and both adult and nymphs presented around 30% of T. cruzi infection (Figure 3.3a).

Figure 3.3. Triatomines with human blood meal, a) population structure and Trypanosoma cruzi infection, and b) microenvironments of collection by region.

3.4.3. Sylvatic Triatomines with Human Blood Meals and its Distance to Households The majority of sylvatic nests where triatomines with human blood meals were identified were located from 200 to a maximum of 500 m far from the households (19 nests); while 13 were located ˂100 m. From the entire sylvatic nest, 49% had triatomines with human blood meal, while in 30% non-human blood was found. In 21% of the nests 61 we found triatomines with evidence of human blood as well as other triatomines with non-human blood. No statistical association was found between the distance and the source of blood (human, non-human) (Chi-square t=3.766, p=0.152), although ˃40% of nests with triatomines with human blood meals in sylvatic environment were near to an infested house. The presence of triatomines with human blood meals in sylvatic environments was not associated with the distance to the nearest house (Fisher's exact test=1.909, p=0.767) (Appendix D).

3.4.4. Haplotype Network of Human Sequences Four different haplotypes were identified using the Median-Joining network. Haplotype

2 (H2) gather most of the sequences (n = 62), which indicates that this is the most common haplotype in these population, followed by the haplotype 1 (H1) (n = 21). Haplotype 3 (H3) and 4 (H4) were represented by eight and two sequences, respectively. Polymorphism positions were detected in two positions along the sequences (position 16 and 138) (Figure

3.4).

Figure 3.4. Median-joining network of mithocondrial cyt b haplotypes for the human sequences identified in the intestinal content of triatomines from central coast and southern highlands region

3.4.5. Feeding Behavior of R. ecuadoriensis Nymphs in Laboratory Conditions Of the seven intestinal contents of adults analyzed by PCR after 72 hours of contact with fasting nymphs, six presented, as expected, sequences corresponding to mouse (Mus musculus) blood. For nymphs, no mouse blood was obtained; from the 21 intestinal contents of nymphs, 67% corresponded to sequences of Rock Pigeon (Columbia livia). In the control assay, 63% of the samples (adults and nymphs) amplified a pigeon sequence.

Low quality products or no amplification was obtained in 31% of the samples. These results showed that in the current experimental conditions, cleptohematophagy was not evidenced.

3.5. Discussion

Control programs of vectors of Chagas disease have been focused on strategies to decrease contacst of the vectors with humans. Indeed, insecticide spraying has been largely applied for control of domiciliated triatomine populations to avoid colonization and human contact indoors (Gorla and Hashimoto, 2017) where transmission was considered to occur.

In this context, in order to develop effective control measures adapted to local conditions in Ecuador, it is important to have a better understanding of the biology of the vector species and of their dynamic of movements considering local characteristics.

Previous studies on relationship of vectors with their vertebrate hosts have been directed to the detection of blood meal sources in various species of triatomines along the Americas such as Mepraia spinolai (Chile), T. longipennis (Mexico), T. brasiliensis

(Brazil), T. infestans and T. eratyrusformis (Argentina), T. protracta and T. rubida

(United States) (Almeida, Faucher, Lavina, Costaand Harry, 2016, Bosseno et al. 2006,

Cecere et al. 2016, Chacon et al. 2016, Curtis-Robles et al. 2018). In the first instance, these studies provide information on the anthropophily of vectors and thus, on the risk of contact between humans and vectors. As expected, humans have been reported as one of the vertebrate host for triatomines collected in domicile, but also in peridomicile environments showing a stream of insects from outside to inside the house and vice versa

(Cantillo-Barraza et al. 2015, Carrasco et al. 2005, Cecere et al. 2016; Ibanez-Cervantes et al. 2013). Surprisingly, several reports of human blood meal in sylvatic populations of triatomines have challenged the idea of human transmission restricted to 64 domicile/peridomicile environments (Breniere et al. 2007, Almeida et al. 2016, Buitrago et al. 2013, Stevens et al. 2012, Waleckx et al. 2014). In this study, the high percentage of triatomines with human blood meal (40.6%) including those collected in sylvatic triatomines (47.5%) indicated that human is an important source of blood for triatomines than previously thought. Moreover, the presence of high infection rate with T. cruzi

(66.3%), including triatomines fed from humans and collected in sylvatic environments, shows that the risk of parasite transmission to humans should be not restricted to domestic/peridomestic environments.

3.5.1. Human as an Important Vertebrate Host for Triatomines and the Presence of Trypanosoma cruzi Infection The presence of human blood in five species of triatomines (P. chinai, P. howardi, P. rufotuberculatus, R. ecuadoriensis and T. carrioni) indicates that human is the principal host for vectors living in human dwellings and their surroundings in both endemic regions under this study, especially for R. ecuadoriensis that is widely distributed and present in all environments. The haplotype analysis demonstrated that different human sources were present, which indicates multiple sources of human blood. Even more, all the species presented infection with T. cruzi, which has important implication in Chagas disease transmission.

In domestic environments, it is expected to find human blood as triatomine meal. This study has reported 54.3% of the triatomines collected in this environment with human blood, and with a 54.5% of T. cruzi infection rate. In this case, humans are frequent hosts and most of the triatomines from this environment were found in bedrooms. However, the presence of triatomines indoors will depend on the local ecological and social 65 characteristics of the area. In southern highlands region, there are established colonies in the intradomicile (mostly associated to bedrooms) of P. chinai, P. rufotuberculatus, R. ecuadoriensis and T. carrioni with an infestation rate of 10% and 11% of T. cruzi infection (Grijalva, Villacis, Ocaña-Mayorga, et al. 2015). In the central coastal region, a low domicile infestation for R. ecuadoriensis and P. howardi (< 0.7%) has been reported, although with high T. cruzi infection (47.5%) (Grijalva, Villacis, Moncayo, et al. 2017).

Thus, intradomicile T. cruzi transmission is active in the southern highlands but it is less likely in the central coastal region.

By the other hand, in the peridomicile, 27.7% of the triatomines were detected with human blood. Most of these triatomines were collected in chicken nests suggesting a frequent move to the house to feed on humans or other available host. Due to birds are refractory to the parasite, the high T. cruzi infection rate found in triatomines collected in chicken nests (81.3%) confirmed that triatomines are acquiring the parasite by feeding from other infected mammal hosts. Thus, the presence of infected triatomines collected in chicken nest demonstrated that the insects, even as nymphs, can shift hosts according to availability. The shifting of host has been previously described for T. infestans in

Argentina where climate conditions, people habits and vertebrate host availability change along the year (Cecere, Gurtler, Canale, Chuitand Cohen, 1997; Gurtler et al. 2009). A similar scenario can be taking place in the study areas. In this case, triatomines in the peridomicile are having easy access to humans. The human-vector contact might be taking place by an active dispersal of the triatomines between the domicile and peridomicile due to the lack of physical barriers, or passive dispersal with domestic 66 animals. However, human activities that require spending long periods of time outdoors

(knitting, cleaning, and embroidering) could also be assed to determine transmission risk of T. cruzi.

In sylvatic environments, the presence of human blood in triatomines has been previously reported (Almeida et al. 2016, Buitrago et al. 2013, Stevens et al. 2012,

Waleckx et al. 2014). In this study, only individuals of R. ecuadoriensis were analyzed from this environment and 47.5% of them were feed with human blood. Moreover, infection rate with T. cruzi was 63.6%. These results agreed with previous reports of high infection rates in sylvatic R. ecuadoriensis (Grijalva et al. 2012, Grijalva, Teranand

Dangles, 2014, Grijalva and Villacis, 2009). The presence of human blood in sylvatic triatomines was as these triatomines were collected in bird and small mammal’s nests in trees and palm trees, particularly in squirrel´s nest.

The human blood meals in triatomines collected in peridomestic and sylvatic environments poses the question on how these triatomines (adults and nymphs) are getting in contact with humans. We suggest three mechanisms: i) dispersion of adults and nymphs from sylvatic and peridomestic environments to indoors, ii) the cleptohematophagy within colonies; and iii) human-triatomine contact outdoors.

 Dispersion of Adult Triatomines

Dispersion of triatomines can be driven by passive transport by vertebrate hosts (e.g. birds) or by their own means walking (nymphs and adults) or flying (adults) (Guhl 2017).

In central coastal region, the majority of houses in rural communities are built with sugar cane walls (79%) and zinc roofs (66%); while in southern highlands, houses are built 67 with adobe walls and tile roofs (Black et al. 2007). Both kind of construction permit easy access of insects from outdoors to indoors due to the presence of open spaces between the walls and roof. In addition, in both areas laying hens are installed against to the outside wall of the house or within less than 50 m distance.

Under these conditions, and considering the capability of triatomines to disperse (e.g. for T. infestans the flying range is from 0.2 to 2 km, Schweigmann et al. 1988), both adults and nymphs can easily walk or fly to indoors and come back to their original habitat if they do not found adequate conditions indoors. Returning to the nest of chickens outside the house shows that the habitat and perhaps the gregarious instinct of triatomes motivate their movements. The active dispersion is usually associated with searching of food and mate for adults (Schofield 1994), however other reason can be suspected. The second senario that can explain human blood meal in triatomines collected outside the house is their passive introduction by domestic animals such as dogs or hens that come and go between the two environments.

In sylvatic environments, active dispersal of adults of R. ecuadoriensis by flying to dwellings could explain the presence of human blood meal. In other triatomine species, there is evidence of light attraction (Minoli and Lazzari, 2006; Pacheco-Tucuch,

Ramirez-Sierra, Gourbiereand Dumonteil, 2012) and also good dispersal capabilities in species such as R. pallescens, T. dimidiata, T. infestans, T. brasiliensis and T. pseudomaculata, T. sanguisuga and T. gerstaeckeri (Carbajal de la Fuente et al. 2007,

Bremond et al. 2014, Castro et al. 2014, Dumonteil et al. 2007, Hamer, Bejcek, Valdez,

Curtis-Roblesand Hamer, 2018). 68

In Ecuador, R. ecuadoriensis is widely distributed and is the only species present in all three environments (Grijalva, Villacis, Moncayo, et al. 2017, Grijalva, Villacis, Ocaña-

Mayorga, et al. 2015). Sylvatic populations of R. ecuadoriesis has been found aggregated at a distance < 100 m from houses and associated to vertebrate host distribution. These data indicates that this vector species seems to have good dispersion capability. Thus, even if the dispersion of R. ecuadoriensis from sylvatic to domestic environments is feasible, the frequency, distance and conditions of the dispersion need to be further assessed. However, no association was found between the distance from the sylvatic place of capture of triatomines and the closest infested house.

 Cleptohematophagy as a Survival Mechanism

Dispersion of sylvatic adults of R. ecuadoriensis might be achieved by flying.

However, our results demonstrated that 51% of the sylvatic triatomines where human blood meals have been detected, were nymphs. These nymphs of R. ecuadoriensis can be found in nests located in a range of 10 m to 900 m far from the closest house. Although, active dispersion of nymphs has been suggested to take place in sylvatic populations of other triatomine species such as T. dimidiata, T. infestans, T. guasayana and T. ryckmani

(Monroy, Marroquin, Rodas, Rosalesand Jaenson 2004, Monroy, Bustamante, Rodas,

Enriquez and Rosales 2003, Vazquez-Prokopec, Ceballos, Kitron and Gurtler 2004,

Vazquez-Prokopec et al. 2006, Bremond et al. 2014), the capability of R. ecuadorienesis nymphs to disperse is not known. An alternative explanation is the cleptohemagophagy.

Adults that have fed from humans can contribute to the maintenance of colonies by

“feeding” nymphs when access to other vertebrate host is scarce. This behavior has been 69 previously reported, under experimental conditions for some hemiptera species such as

Clerada apicicornis (Torres, Cardenas, Perez and Morales, 2000) and the triatomine

Belminus herreri (Sandoval, Joya, Gutierez and Angulo, 2000). Although in our experiment, no cleptohematophagy behavior was observed, further experiments needs to be carried out to assess this behavior and feasibility of T. cruzi transmission in R. ecuadorienesis and other species.

 Outdoor Activities as Possible Risk for T. cruzi Transmission to Humans

Another alternative that needs to be evaluated is the risk of transmission to human by outdoor activities. The high proportion of human blood meals in this current study

(including nymphs) could be due to other factors than triatomine dispersal. For example, the presence of human blood meal in sylvatic triatomines has been associated to an expansion of human settlements areas (Waleckx et al. 2014) or by accidental contact with humans (e.g. camping areas) (Stevens et al. 2012). However, the risk of transmission might be related with outdoor activities in the field (e.g. farming, sheltering cattle, etc.)

(Breniere, Buitrago, et al. 2017, Buitrago et al. 2013). Human population in the study areas are mainly farmers that cultivate small plots of land for family consumption (Nieto-

Sanchez et al. 2015), thus, a major part of its time is dedicated to planting, harvesting or even “mingas” (communitarian activities for building of cleaning common areas). All those activities require a long stay in the field in areas similar to where we collected the sylvatic triatomines. Generally, the population recognizes triatomines but does not know that they are the vectors of a serious disease. Anthropological studies are needed to 70 determine if people have contact with triatomines during outdoors activities and which of those activities could constitute a major risk of transmission.

3.6. Conclusions

This study confirms the complexity of Chagas disease transmission and reinforces the importance of considering local conditions for understanding the dynamics of the transmission disease. It has been reported that human disturbance around their dwellings can result on higher T. cruzi infection risk (Jansen, Roque and Xavier, 2017). However, the impact seems to be different in different geographic areas. For example, in Panamá, landscape disturbance increase vector infection with T. cruzi (Gottdenker, Chaves,

Calzada, Saldana and Carroll, 2012), while in the Atlantic forest in Brazil, domiciliary invasion by sylvatic triatomines seems not to be stimulated by environmental disturbance

(Leite, dos Santos and Falqueto, 2011). Thus, local features, biological as well as anthropological, need to be considered to evaluate and proposed integral control and prevention measures. Although spraying has an effect on decreasing domestic triatomine populations, it might not be the principal strategy for Chagas disease control in several places. This study demonstrated that the contact between humans and vectors is more frequent than previously reported and that might not be restricted to indoors. In addition, triatomines could enter the house, feed and come out. According to this scenario, insecticide spraying indoors would have effect on domestic populations but would not avoid human-vector contact. Moreover, the report of high number of triatomines with human blood may reflect an underestimation of Chagas disease patients in the studied areas. Thus, other studies are required for a better understanding of the dynamics of the parasite transmission in Ecuador. In particular, three areas need to be assessed. 1)

Serological survey to detect chagasic patients and estimate areas of higher risk of 72 transmission. 2) Anthropological studies to estimate transmission risk outdoors associated with some activities such as planting, harvesting, etc.; and, 3) Entomological studies to know the dispersal capability of R. ecuadoriensis and cleptohematophagy behavior.

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Waleckx, E., Suarez, J., Richards, B., and Dorn, P. L. (2014). Triatoma sanguisuga blood meals and potential for Chagas disease, Louisiana, USA. Emerg Infect Dis, 20(12), 2141-2143. doi: 10.3201/eid2012.131576

Walter, A., Lozano-Kasten, F., Bosseno, M. F., Ruvalcaba, E. G., Gutierrez, M. S., Luna, C. E., . . . Breniere, S. F. (2007). Peridomicilary habitat and risk factors for Triatoma infestation in a rural community of the Mexican occident. Am J Trop Med Hyg, 76(3), 508-515.

CHAPTER 4: DISCRIMINATION OF ANOPHELINAE SPECIES FROM

COASTAL AND AMAZON ECUADOR BASED ON NUCLEAR AND

MITOCHONDRIAL MARKERS

4.1. Abstract

In Ecuador, there is a lack of a comprehensive survey of Anopheles and

Nyssorhynchus diversity. Even more, implication on malaria transmission of reported species is still poorly understood. Morphological identification is the most common and available tool; however, it fails in discriminating species closely related and with similar morphological characteristics. Some molecular tools have been proposed for an accurate identification. The most common markers used for species discrimination are nuclear internal transcribed spacer 2 (ITS2) and the mithocondrial cytochrome oxidase I (COI).

The results of this study demonstrated the complexity of taxonomic identification of species, particularly of individuals from species complex, such as the case of the

Oswaldoi group which species are sympathric in the Amazon region. By the analysis of the Restriction Fragment Length Polymorphism (RFLP) of the ITS2 was accurate to identify species that are not closely related, thus this methodology is a good alternative to identify large amounts of individuals from areas where diversity is well known. However, in the Ecuadorian Amazon, this methodology fails to accurate discriminate species from the Oswaldoi group. Species confirmation was carried out by sequencing of COI gene that led to a proper identification of samples. This study confirmed that in the coastal region of Ecuador, Ny. albimanus is the most abundant species and it also extends the geographic distribution of An. calderoni. In the Amazon region, this study confirmed the 80 presence of Ny. rangeli, Ny. oswaldoi B and Ny. aff. konderi. Moreover, molecular analysis increase Ny. benarrochi B in the list of reported species in the Amazon. A comprehensive survey of Anopheles and Nyssorhynchus species in Ecuador is required to have an update and complete inventory of species. Moreover, studies on ecological and behavioral data are needed to facilitate and targeted malaria control strategies in the

Ecuador.

4.2. Background

Mosquitos of the genus Anopheles and Nyssorhynchus are responsible for the transmission of malaria. In Ecuador, malaria is endemic in the coastal and Amazon region. An important reduction of cases has been reported since 2001 (SNEM, 2013) and

Ecuador was one of the countries considered capable of eliminating malaria by 2020

(SNEM, 2013; WHO, 2013, 2018). However, since 2016 the country has experienced an increase in the number of cases in both the coastal and Amazon regions of more than

1,000 cases/year, challenging the elimination goal (WHO, 2018) The administrative transition from the National Service of vector-borne diseases to the Ministry of Public

Health (MSP) is perceived to have weakened the control strategy that was effective in previous years and epidemics are present due to the decrease of surveillance and response capabilities (PAHO, 2016). This increase in cases poses a challenge for the country towards the elimination goal. Some of the major challenges include accurate estimation of malaria cases, influence of asymptomatic cases (Saenz et al. 2017), social factors (e.g. movement of people from active to low transmission areas, increase of agricultural land), and limited information about the diversity and biology of anophelines.

It is the well-known that the complexity of the anopheline hinders the accurate identification by morphology only (Gómez, Cienfuegos, Gutiérrez, Conn and Correa,

2010, Harbach 2004, Lobo et al. 2015). Morever, it requires individuals well preserved that maintained important morphological characteristics for taxonomic identificiation.

Moreover, several Neotropical species are known to comprise isomorphic species (Ruiz-

Lopez et al. 2013) while others are cryptic species (Lehr, Kilpatrick, Wilkerson and 82

Conn, 2005). The genus Anopheles Meigen (1818) is classified in six subgenera:

Anopheles, Stethomya, Cellia, Lophopomyia, Kertesia and Nyssorhynchus (Gonzalez and

Carrejo, 2009), although Nyssorhynchus is now recognized as a monophyletic group and constitutes a separate species (Foster et al. 2017). In Latin America, the subgenera

Nyssorhynchus and Anopheles have a medical importance due to they contain the majority of species incriminated as vectors of malaria. Nine species (including species complexes) are considered as dominant malaria vectors (An. freeboni, Ny. albimanus, Ny. aquasalis, Ny. darlingi, Ny. marajoara and species complex: An. pseudopunctipennis s.l., An. quadrimaculatus s.l., Ny. albitarsis s.l. and Ny. nuneztovari s.l. (Sinka, Bangs, et al. 2010, Sinka et al. 2012).

In Ecuador, historical documents reported 21 species of Anopheles (Levi-Castillo,

1949). A revision based on the available sequences of the mitochondrial DNA

(Cytochrome c Oxydase I) identified 22 species of Anopheles based on literature and available sequences in GenBank including species that have not been previously reported like An. cruzii, An. lepidotus, An. gomezdelatorrei and An. trinkae (Arregui, Enriquez,

Benítez-Ortiz and Navarro, 2015). However, reports with collected mosquitoes reported six species of anophelines in the coastal and highlands areas (An. eiseni, An. calderoni,

An. neivai, An. pseudopunctipennis, An. punctimacula, Ny. albimanus, and Ny. oswaldoi s.l.) (Gonzalez et al. 2010, Pinault and Hunter, 2011, 2012 (Gonzalez et al. 2010, Pinault and Hunter, 2011, 2012, Navarro, Arrivillaga, Morales, Ponce and Cevallos, 2015). Of them, An. calderoni, An. pseudopunctipennis and Ny. albimanus have been found infected with the parasite Plamodium (Collins and Jeffery, 2003, Orjuela et al. 2015, 83

Warren, Collins, Richardson and Skinner 1977). In the Amazon region, eight species were reported (An. apicimacula, An. bombusicolus, An. fluminensis, An. forattinii, An. lepidotus, An. mattogrossensis, An. squamifemur, Ny. sp.nr. konderi, the species complex

Ny. oswaldoi, Ny. rangeli and Ny. trinkae) (Duque et al. 2019, Linton et al. 2013, Ortega et al. 2018). An. darling, the most important vector in the Amazon region, has not been reported in Ecuador, although this species is present in Colombia and Peru in areas that border Ecuador (Montoya-Lerma et al. 2011, Schoeler, Flores-Mendoza, Fernandez,

Davila and Zyzak 2003). A complete list of the Anopheles species in Ecuador is yet to be defined, and no local taxonomic key is available, making species identification a difficult task.

Due to the importance of accurate species identification, molecular tools have been evaluated to confirm morphological identification (Foster et al. 2013). Nuclear (ITS2) and mitochondrial (COI) markers have been used to elucidate phylogenetic relationships among closely related species (Arregui et al. 2015, Cienfuegos et al. 2008, Cienfuegos et al. 2011, Wilkerson, Reinert and Li 2004). Moreover, the use of both markers, increase the resolution because of the high number of copies and the proved resolution to differentiate reproductively isolated populations (Alquezar, Hemmerter, Cooper and

Beebe, 2010, Foster et al. 2013, Loaiza et al. 2013, Matson et al. 2008). The aim of this study was to compare morphological with molecular identification though the restriction fragments length polymprohism (RFLP) analysis of the ITS2 region and the sequencing of the COI in Anopheles mosquitos from the coastal and Amazon region of Ecuador.

4.3. Methodology

4.3.1. Study Area Mosquitoes samples were obtained from different locations in the coastal (El Oro province, n= 608) and the Amazon (Orellana, n= 116 and Sucumbíos, n= 235) regions, during 2011 and 2012 (Figure 4.1). Individuals were coded and stored dried in silica gel.

Mosquitoes from the coastal area were taken from stored samples of previous projects of the Center for Research on Health in Latin America (CISeAL) and correspond to the localities of Barbones (3°10'60"S, 79°55'0"W; 4 meters above sea level, masl), Los

Ángeles (3°17'57"S, 79°57'25.92"W; 6 masl), Huaquillas (3°29'9"S, 80°14'20"W y 9 masl). While the samples from the Amazon were collected by human or animal landing catches by personnel of the Ministry of Public Health (MSP) as an epidemiological study in the area. In Orellana province, mosquitoes were collected in the locality of Kawymeno

(1°1'58.8"S, 75°46'48"W, 182 masl); while in Sucumbíos province, collections were carried out in seven localities: Santa Elena (0°22'16.1"N, 76°11'35.5"W; 231 masl),

Nueva Montepa (0°10'26.4"N, 75°55'18,6"W; 221 masl), Bajo Restrepo (0°20'05.8"N,

76°07'28.0"W; 220 masl), El Porvenir (0°17'50.7"N, 76°0'35.6"W; 219 masl), San José de Wisuya (00°24'07.9"N, 76°14'14.9"W, 229 masl), El Palmar (00°25'28.2"N,

76°17'22.2"W; 240 masl), Bocana de Cuembí (00°22'52.0"N, 76°23'46.7"W; 243 masl).

El Oro, Orellana and Sucumbios are endemic for malaria and are located in areas where risk factors as weather, vegetation, population migration and increase of agricultural land make them susceptible for the presence and proliferation of Anopheles.

Since 2010, El Oro province has shown a sustained decrease in the number of cases

(MSP, 2018). However, in the Amazon region, particularly in Orellana, an increase of 85 cases have been reported with more than 1 000 cases from 2015 to 2019 ((MSP, 2018).

Information about vectors implicated in transmission is scarce. The MSP reported An. albimanus and An. pseudopunctipennis as vectors in the coastal area, while An. nuneztovari and An. trinkae in the Amazon region (SNEM, 2013).

4.3.2. Species Identification Morphological identification to species was made in using taxonomic keys (Gonzalez and Carrejo, 2009; Linthicum, 1988; Suárez, Quiñones and Robayo, 1988).

Molecular identification was conducted by two strategies 1) initial PCR-RFLP for

ITS2 patterns, and 2) COI amplification for comparison with sequences from NCBI and

BOLD databases.

4.3.3. ITS2 Amplification and Restriction Fragment Length Polymorphism (RFLP) Patterns DNA template was obtained by a single leg, added to the PCR as described by (Zapata et al. 2007). For stored individuals, where a leg was not feasible to use, DNA from the abdomen was extracted following the protocol for DNA extraction with protein precipitation with AcK, as described in (Rosero, Gutierrez, Cienfuegos, Jaramillo and

Correa, 2010), with a previous wash of the sample in 100% alcohol. ITS2 fragment was amplified by PCR using primers for the conserved 5.8S and 28S flanking sequences:

ITS2A 5´-TGT GAA CTG CAG GAC ACA T-3´ and ITS2B 5´-TAT GCT TAA CAG

GAC GGG GT-3´ following the amplification conditions, previously reported in (Beebe and Saul, 1995) for a total volume of 30 µl. The presence of PCR product was visualized in a 2% agarose gel with 1X SYBER Safe® (Invitrogen). Restriction digestion analysis of 10 µl of the ITS2 product was performed according to the manufacturer. All samples 86 were analyzed with the enzyme AluI (BioLabs®Inc, RO137S) as recommended by

(Cienfuegos et al. 2008, Zapata et al. 2007). Samples that did not show restriction fragments with AluI were analyzed with HaeIII (BioLabs®Inc, R0108S) and FspI

(BioLabs®Inc, RO135S). Fragments were visualized in 2% agarose gel with 1X SYBER

Safe®. Control DNA from An. albimanus, An. nuneztovari, An. punctimacula and An. darling were used for each digestion set. Resulting gels were photographed and analyzed with the software Doc-It®LSImageAnalysis. Photo analysis was carried out to determine the size of the restriction bands.

Identification was realized according to comparison with the RFLP fragments of control DNA (Ny. albimanus), and patterns reported in literature (Cienfuegos et al. 2008,

Ruiz et al. 2005, Zapata et al. 2007). Identification of An. calderoni was analyzed in silico using sequences of ITS2 available in GenBank with the software NEVcutter V2.0

(Table 4.1). Identification was determined according to the band size of the restriction fragments. An error of 10% was accepted for each fragment size. A consensus identification was carried based on the restriction patterns of the three enzymes (AluI,

HaeIII and FspI) considering the distribution of the species. Table 4.1. Expected RFLP pattern of ITS2 fragment digestion of the Anopheles and Nyssorhynchus species with the restriction enzymes AluI, HaeIII and FspI according to literature. In silico pattern was defined for An. calderoni identification. Alu I Hae III Fsp I Species # RFLP pattern # RFLP pattern # RFLP pattern References/ accession # bands (bp) bands (bp) bands (bp) An. punctimacula 2 334, 81 Zapata et al. 2007 An. pseudopunctipennis 1 566 Zapata et al. 2007 HQ622618.1, HQ622621.1, An. calderoni 2 272, 181 3 282, 97, 68 QF698887.1, HQ622622.1, KF698880.1, KF698879.1 Ny. albimanus 2 377, 127 Zapata et al. 2007 Ny. albitarsis s.l. 3 280, 170, 95 Zapata et al. 2007 Ny. aquasalis 4 282, 135, 60, 5 2 444, 38 2 437, 45 Cienfuegos et al. 2008 Ny. nuneztovari 3 358, 96, 76 Zapata et al. 2007 Ny. nuneztovari 3 341, 83, 77 3 309, 154, 38 3 357, 99, 45 Cienfuegos et al. 2008 Ny. rangeli 2 340, 165 Zapata et al. 2007 Ny. rangeli 2 326, 152 2 440, 38 2 433, 45 Cienfuegos et al. 2008 Ny. triannulatus s.l 3 240, 190, 168 Zapata et al. 2007 Ny. darlingi 3 321, 158, 78 Zapata et al. 2007 Ny. benarrochi 3 341, 83, 76 4 306, 158, 38, 4 2 407, 99 Cienfuegos et al. 2008 Cienfuegos et al. 2008; Ruiz Ny. benarrochi 2 423, 83 3 365, 137, 38 et al. 2005 Ny. oswaldoi 3 284, 125, 83 2 454, 38 3 350, 97, 45 Cienfuegos et al. 2008 Cienfuegos et al. 2008; Ruiz Ny. oswaldoi 2 367, 125 2 493, 38 et al. 2005 Ny. strodei 1 488 4 170, 148, 132, 38 3 356, 87, 45 Cienfuegos et al. 2008

4.3.4. COI Amplification and Sequencing Criteria for selection of samples for COI analysis were the following: i) Random samples that could not been identified by morphology nor PCR-RFLP patterns, ii) a subset of each identified species for confirmation.

PCR was performed using the primers LCO1490 (5'-GGT CAA CAA ATC ATA

AAG ATA TTG G-3') and HCO2198 (5'-TAA ACT TCA GGG TGA CCA AAA AAT

CA-3') and following the amplification conditions described in (Folmer, Black, Hoeh,

Lutz and Vrijenhoek, 1994). PCR products were confirmed in a 2% agarose gel with 1X

SYBER Safe®. Samples that presented more than the expected band were purified from gel with the QIAquick Gel extraction® (Qiagen). PCR products were sequenced in both directions, edited and analyzed using the software Chormas Lite v2.1 and SeaView v4.2.6. Consensus sequences were compared with data on the NCBI BLAST

(https://blast.ncbi.nlm.nih.gov) and BOLD (http://www.boldsystems.org/) database to confirm species identification.

4.4. Results

4.4.1. Morphological Identification Of the 959 individuals, 56% (n= 533) were identified to species, while the rest were identified to genera (Anopheles spp.). In El Oro province (n= 608), four species were detected: Ny. albimanus was the most abundant (72%, n= 436), An. calderoni represented 10% of the samples (n= 59), while only two individuals were identified as

An. pseudopunctipennis and An. puctimacula (0.2%). Eighteen percent of the samples were not identified to species (n= 111). In Orellana province, only 9% of the samples were identified as Ny. dunhami, while 91% (n= 106) were not identified to species. In

Sucumbios, a high percentage of individuals were not able to be identified to species

(89%); however, Nyssorhynchus oswaldoi s.l., Ny. nuneztovari and Ny. dunhami were identified in low proportions (4.3%, 3.4% and 0.9%, respectively), while misidentification of Ny. albimanus and An. calderoni was detected in 3% of the samples

(n= 7) (Table 4.2, Appendix E). Individuals that were identified as Anopheles spp. but not identified to species presented problems with poor specimen preservation, thus some important morphological characteristics were absent. Table 4.2. Number or individuals identified by morphological characteristics collected in El Oro province (costal region), Sucumbios and Orellana provinces (Amazon region).

Province/Locality An. An. An. Ny. Ny. Ny. Ny. Anopheles Total calderoni pseudopuctipennis punctimacula albimanus dunhami nuneztovari oswaldoi spp. (%) (%) (%) (%) (%) (%) (%) (%) El Oro 59 (9.7) 1 (0.2) 1 (0.2) 436 (71.7) - - - 111 (18.3) 608 Barbones 24 (15) - 1 (0.6) 90 (56.3) - - - 45 (28.1) 160 Huaquillas 27 (16.7) - - 114 (70.4) - - - 21 (13.0) 162 Los Angeles 8 (2.8) 1 (0.3) - 232 (81.1) - - - 45 (15.7) 286 Orellana - - - - 10 (8.6) - - 106 (91.4) 116 Kawymeno - - - - 10 (8.6) - - 106 (91.4) 116 Sucumbios 1 (0.4%) - - 6 (2.6) 2 (0.9) 8 (3.4) 10 (4.3) 209 (88.9) 235 Bajo Restrepo ------52 (100) 52 Bocona Cuembí - - - - - 2 (6.3) 2 (6.3) 28 (87.5) 32 Nueva Montepa - - - 1 (3.3) - - - 29 (96.7) 30 Palmar - - - 1 (6.3) - 1 (6.3) 3 (18.8) 11 (68.8) 16 Porvenir 1 (2.2) - - 1 (2.2) 1 (2.2) 2 (4.3) 4 (8.7) 37 (80.4) 46 San José Wisuya - - - - - 3 (13.6) 1 (4.5) 18 (81.8) 22 Santa Elena - - - 2 (5.4) 1 (2.7) - - 34 (91.9) 37 Total 60 (6.3) 1 (0.1) 1 (0.1) 441 (46.0) 12 (1.3) 8 (0.8) 10 (1.0) 426 (44.4) 959

4.4.2. RFLP-ITS2 Identification RFLP pattern analysis were applied to 647 samples and a product for restriction analyses was obtained in 74.3% of the samples (n= 480). In this analysis, we included only 30% of samples that were morphologically identified as Ny. albimanus.

Six species were identified by RFLP patterns (An. calderoni, Ny. albimanus, Ny. benarrochi, Ny. oswaldoi¸ Ny. rangeli and Ny. strodei). Seven samples (1.09%) were not able to define the species due to RFLP patterns range were shared with two species (Ny. rangeli/Ny. oswaldoi and Ny. strodei/Ny. benarrochi) (Table 4.3).

In El Oro province, Alu I RFLP pattern identified two species Ny. albimanus and Ny. strodei, although the latter has not been reported in the Pacific coast. In 52 samples, Ny. albimanus shared band patterns with Ny. oswaldoi, Ny. rangeli and Ny. strodei. With Hae

III and Fsp I, only An. calderoni was identified, based on the expected restriction patterns according to the in silico analysis of six ITS2 sequences available in GeneBank (Appendix

D).

In the Amazon provinces (Orellana and Sucumbios), the restriction pattern with the enzyme Alu I identified five species (An. pseudopunctipennis, Ny. albimanus, Ny. oswaldoi, Ny. rangeli and Ny. strodei), although both An. pseudopunctipennis and Ny. albimanus are not present in the Amazon region. Shared band patterns were found in 13 samples among Ny. oswaldoi, Ny. albimanus and Ny. rangeli. Only two samples produced the expected restriction fragment pattern with the enzyme HaeIII, however, it fails in discriminating between Ny. rangeli and Ny. oswaldoi. The enzyme FspI identified only six samples as Ny. benarrochi (Appendix D). 92

Consensus identification by RFLP-ITS2 considered band and size patterns as well as known distribution of the species. In El Oro province, two species were found (An. calderoni and Ny. albimanus), being Ny. albimanus the most abundant species in the province (71%, n= 210), while An. calderoni represented 18% (n= 52). Not identifiable patterns were found in 11% (n= 32) of the samples. In Orellana and Sucumbios province, four species of the genus Nyssorhynchus were identified (Ny, benarrochi, Ny. oswaldoi,

Ny. rangeli and Ny. strodei). The most abundant species was Ny. strodei (20%, n= 130), followed by Ny. rangeli with 11% (n= 67) (Table 4.3). From 30 to 40% of the samples were not identified due to the presence of non-reported band pattern, while in 1.3% of the samples, shared patterns between Ny. rangeli/Ny. oswaldoi and Ny. strodei/Ny. benarrochi did not allow to identification (Table 4.3).

Agreement between morphological and RFLP-ITS2 identification was lower in individuals from the Oswaldoi complex. Individuals identified as Ny. nuneztovari and Ny. oswaldoi, by morphology, were identified as Ny. rangeli by RFLP-ITS2; while individuals of Ny. dunhami were identified as Ny. strodei. For Ny. albimanus agreement of identification between methods was high, however, five samples from the Amazon region that were morphologically identified as An. albimanus were Ny. rangeli by RFLP-

ITS2. One sample from El Oro province was identified as An. pseudopuctipennis but by

RFLP-ITS2 this individual was Ny. albimanus. Table 4.3. Identification based on RFLP-ITS2 restriction fragments with enzymes AluI, HaeIII and FspI of mosquitoes collected in El Oro province (costal region), Sucumbios and Orellana provinces (Amazon region).

Province/Locality An. Ny. Ny. Ny. Ny. Ny. Nyra/ Nyst/ NI (%) Total calderoni albimanus benarrochi oswaldoi rangeli strodei Nyos Nybe (%) (%) (%) (%) (%) (%) (%) (%) El Oro 52 (17.7) 210 (71.4) ------32 (10.9) 294 Barbones 16 (17.0) 59 (62.8) ------19 (20.2) 94 Huaquillas 25 (30.1) 52 (62.7) ------6 (7.2) 83 Los Angeles 11 (9.4) 99 (84.6) ------7 (6.0) 117 Orellana - - - - 1 (0.9) 77 (66.4) - 3 (2.6) 35 (30.2) 116 Kawymeno - - - - 1 (0.9) 77 (66.4) - 3 (2.6) 35 (30.2) 116 Sucumbios - - 1 (2.2) 13 (5.7) 66 (28.9) 53 (23.2) 2 (0.9) 2 (0.9) 91 (39.9) 228 Bajo Restrepo - - - - - 28 (62.2) 1 (2.2) - 16 (35.6) 45 Bocona Cuembí - - - - 25 (78.1) - - 2 (6.3) 5 (15.6) 32 Nueva Montepa - - - 5 (16.7) 3 (10) 6 (20.0) 1 (3.3) - 15 (50.0) 30 Palmar - - - - 10 (62.5) 2 (12.5) - - 4 (25.0) 16 Porvenir - - - 7 (15.2) 7 (15.2) 8 (17.4) - - 24 (52.2) 46 San José Wisuya - - - - 14 (63.6) 2 (9.1) - - 6 (27.3) 22 Santa Elena - 1 (0.2) 1 (2.7) 7 (18.9) 7 (18.9) - - 21 (56.8) 37 Total 52 (8.2%) 210 (32.9) 1 (0.2 13 (2.0) 67 (10.5) 130 (20.4) 2 (0.3) 5 (0.8) 158 (24.8) 638 An.: Anopheles; Ny.: Nyssorhynchus; Nyra/Nyos: Ny. rangeli / Ny. oswaldoi; Nyst/Nybe: Ny. strodei / Ny. benarrochi; NI: non identified

4.4.3. COI Identification According to availability of samples, a subset (n= 40) was chosen to confirm identification by COI sequencing; however, only 28 samples amplified products which sequence were analyzed. By this analysis, the presence of An. calderoni and Ny. albimanus was confirmed widely distributed in El Oro province (Figure 4.1) and identification was confirmed to species by COI (Table 4.4, Appendix F). In Orellana province (Kawymeno community), only Ny. benarrochi B was identified by COI (Table

4.4, Figure 4.1); while in Sucumbios province, five species were identified. Ny. benarrochi B was reported for Bajo Restrepo and Nueva Montepa. In El Porvenir, three species were reported ( Ny. aff. konderi, Ny. oswaldoi, Ny. rangeli); while Ny. rangeli was present in San José Wisuya and Santa Elena (Table 4.4, Figure 4.1). However, discordance in identification was found in species distributed in the Amazon (Sucumbios and Orellana provinces). One out of the 13 samples of Ny. oswaldoi by RFLP-ITS2 was identified as Ny. aff. konderi (a species of the Oswaldoi complex); while samples of Ny. strodei by RFLP-ITS2 were identified as Ny. benarrochi B or Ny. oswaldoi B.

Concordance in identification between RFLP-ITS2 and COI was found in Ny. rangeli.

COI analysis of samples with shared RFLP-ITS2 patterns between two species (Ny. rangeli / Ny. oswaldoi, Ny. strodei / Ny. benarrochi) were not able to be confirmed due to lack of sample material for sequencing (Appendix F).

Twelve out of the 158 non-identified samples by RFLP-ITS2 were randomly selected for COI analysis. Among these samples five species were identified (An. calderoni, Ny. albimanus, Ny. benarrochi B, Ny. rangeli and Ny. oswaldoi B). Samples identified as An. calderoni, Ny. benarrochi and Ny. oswaldoi by COI were not discriminated by RFLP-

ITS2 due to the 10% error accepted for variability on the band size. In these cases, 95 obtained band size were close to the limits set for the species. However, other individuals were not identified by RFLP-ITS2 because of the presence of non-expected band sizes or patterns (Ny. albimanus, Ny. benarrochi, Ny. oswaldoi) (Appendix F).

Figure 4.1. Location of communities where anopheline mosquito collection was carried out and species identified by sequencing of a fragment of the COI gene.

Table 4.4. Identification by mithochondrial COI of mosquitoes collected in El Oro province (costal region), Sucumbios and Orellana provinces (Amazon region).

Province/Locality An. calderoni Ny. Ny. Ny. aff. Ny. Ny. rangeli NI (%) Total (%) albimanus benarrochi B konderi (%) oswaldoi B (%) (%) (%) (%) El Oro 3 (30) 6 (60) - - - - 1 (10) 10 Barbones 2 (33.3) 4 (66.7) - - - - - 6 Huaquillas 1 (50) - - - - - 1 (50) 2 Los Angeles - 2 (100) - - - - - 2 Orellana - - 9 (69.2) - - - 4 (30.8) 13 Kawymeno - - 9 (69.2) - - - 4 (30.8) 13 Sucumbios - - 1 (5.9) 1 (5.9) 4 (23.5) 4 (23.5) 7 (41.2) 17 Bajo Restrepo - - 1 (33.3) - - - 2 (66.7) 3 Bocona Cuembí ------Nueva Montepa - - - - 1 (33.3) - 2 (66.7) 3 Palmar - - - - - 1 (33.3) 2 (66.7) 3 Porvenir - - - 1 (16.7) 3 (50) 1 (16.7) 1 (16.67) 6 San José Wisuya - - - - - 1 (100) - 1 Santa Elena - - - - - 1 (100) - 1 Total 3 (7.5) 6 (15) 10 (25) 1 (2.5) 4 (10) 4 (10) 12 (30) 40 4.5.Discussion

In Ecuador, there is a lack of a comprehensive survey of Anopheles and

Nyssorhynchus diversity. Even more, implication on malaria transmission of reported species is still poorly understood. The results of this study demonstrated the complexity of taxonomic identification of species, particularly of individuals from species complex, such as the case of the Oswaldoi group. However, molecular analysis increase Ny. benarrochi B in the list of reported species in the Amazon and extend the geographic distribution of An. calderoni in the coastal area.

Morphological identification is a cheap and efficient tool for identification of species with features clearly defined, such as Ny. albimanus. However, it requires trained personnel with a good knowledge on mosquitoes’ anatomy and geographic distribution of the species. Besides, it requires careful handling and storage of the mosquito to keep important taxonomic characteristics such as hind legs, wings and thorax. In this study, misidentification of Ny. rangeli as Ny. albimanus could be caused by these factors. In the case of An. calderoni, the results of this study extend its distribution range. By morphological characteristics, this species was successfully identified, although reports of misidentification with An. punctimacula and An. malefactor (Gonzalez et al. 2010;

Loaiza et al. 2013). Misidentification of An. calderoni could cause an inaccurate report of its distribution. Morphological misidentification was common in individuals from the

Amazon region, particularly in individuals from species complex such as the Oswaldoi group. By morphological characteristics, three species were described (Ny. dunhami, Ny. nuneztovari and Ny. oswaldoi. Of them, Ny. oswaldoi was confirmed by COI sequencing, while the individuals identified as Ny. dunhami, Ny. nuneztovari were 98 identified as Ny. benarrochi B and Ny. rangeli, respectively. Discrepancy of identification in the Oswaldoi group has been previously reported (Calle, Quinones,

Erazo and Jaramillo, 2002; Gonzalez et al. 2010), thus, the use of molecular markers is required, particularly in geographic areas where diversity is little known.

The internal transcribed spacer (ITS2) has been used or reconstructing evolutionary relationships at the level of species in various organisms, including insects (Young and

Coleman, 2004). Restriction fragment length polymorphism (RFLP) of the ITS2 has been used to discriminate species of malaria vectors and RFLP patterns have been reported for species from Colombia (Cienfuegos et al. 2008; Gutierrez et al. 2009; Zapata et al.

2007); which constitutes and advantage to use this molecular tool to evaluate Ecuadorian malaria vectors diversity. Variability of band size was one of the main issues when there is no availability of control DNA for visually comparing band patterns of various species.

Some patterns are shared between species as it was seen with Ny. albimanus and Ny. oswaldoi, Ny. rangeli and Ny. strodei; while other patterns identified species that do not correspond to species reported in the area such as individuals of An. pseudopunctipennis in the Amazon region. In both cases, an analysis of the geographic distribution of the species can resolve the identification. Species identification by COI sequencing confirmed as Ny. albimanus, even in the individuals that presented shared bands with Ny. oswaldoi, Ny. rangeli and Ny. strodei.

In the case of species complex and sympathric species such as Ny. rangeli, Ny. oswaldoi, Ny. strodei and Ny. benarrochi, RFLP-ITS2 did not discriminate to species. In these cases, sequencing is required to resolve identification. In this study, COI 99 sequencing confirmed Ny. rangeli. Although, Ny. strodei were not able to be confirmed by COI, these samples were identified as Ny. benarrochi B, and Ny. oswaldoi B, indicating that RFLP-ITS2 is not the a good mechanism to identify species of the

Oswaldoi group.

Additionally, by RFLP-ITS2 and COI, An. calderoni was reported in southern

Ecuador. This species has few reports in northern and central Ecuador (Gonzalez et al.

2010), thus the results of this study extends southern distribution of An. calderoni.

Moreover, this study report Ny. benarrochi in the Amazon region, a species that haven´t been reported to Ecuador.

Although RFLP-ITS2 is a convenient tool to explore diversity in areas with little information, the influence of population variability within species needs to be further evaluated. In the Amazon region, where complex species are present in sympatry, sequencing is still the tool of choice due to an accurate identification and the amount of available sequences for COI and ITS2.

Epidemiological implication of some of the reported species has been widely described as in Ny. albimanus that is considered the main vector of malaria in the Pacific

Coast (Sinka, Rubio-Palis, et al. 2010). Other species present in the coastal region of

Ecuador is An. calderoni which role in malaria transmission has not been defined. In western Colombia, An. calderoni has been incriminated as a potential vector due to its infection with P. falciparum and P. vivax and its adaptability to different environments

(Galeano-Castañeda, Gomez, Hernandez-Valencia and Correa, 2019). However, in 100 southwest Colombia and western Perú, An. calderoni is a primary vector (Orjuela et al.

2015; Wilkerson, 1991).

In the Amazon region, Ny. darlingi is considered the major vector for malaria.

Although this species has been reported in Colombia and Peru, in areas with border to

Ecuador (Montoya-Lerma et al. 2011; Schoeler et al. 2003), it was not reported in this study. However, the presence of Ny. benarrochi was reported. Implication of this species is not clearly understood. In Peru, this species has been found infected with Plasmodium falciparum and P. vivax in areas were Ny. darlingi did not occur (Flores-Mendoza,

Fernandez, Escobedo-Vargas, Vela-Perez and Schoeler, 2004).

By the other side, Ny. oswaldoi, is a species complex which taxonomy remains unresolved. Although Ny. oswaldoi, from the Putumayo area of Colombia (near the collection area in this study), has been found genetically similar to individuals from

Brazil and the Venezuelan Amazon (Quinones et al. 2006). Further studies have found great diversity within this species with at least four species (Ny. oswaldoi s.s., Ny. oswaldoi A, Ny. oswaldoi B and at least two species of the Ny. konderi complex) (Ruiz-

Lopez et al. 2013). Implications in malaria transmission would depend on the species. In this study we found Ny. oswaldoi B and Ny. aff. konderi. Ny. oswaldoi B is a confirmed vector, however, Ny. konderi has been implicated as secondary vector (Ruiz-Lopez et al.

2013). Also, in the area of Colombian Putumayo, Ny. rangeli is one of the species of interest and has been implicated in transmission of P. vivax due to its anthropophilic behavior and apparent high densities (Quinones et al. 2006). In this study, Ny. rangeli has been reported in most of sampled localities, together with Ny. oswaldoi B. Both species 101 has been reported infected with P. vivax (Quinones et al. 2006), thus, these species could have been responsible for the malaria outbreak registered in 2011 in the Ecuadorian side of the Putumayo area.

A comprehensive survey of Anopheles and Nyssorhynchus species in Ecuador is required to have an update and complete inventory of species. Moreover, studies on ecological and behavioral data are needed to facilitate and targeted malaria control strategies in the Ecuador.

102

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APPENDIX A: SPECIES, YEAR OF CAPTURE AND LOCALTION OF 507 TRIATOMINES ANALYZED FROM CENTRAL

COASTAL AND SOUTHERN HIGHLANDS IN ECUADOR

Collection Sample Place of Microenv. Days* Vector species Stage Locality Area Environment year code capture of capture 1 2006 2 TEX318 R. ecuadoriensis adult La Extensa SH intradomicile animal shelter rodent nest bed/ wall/ 2 2006 2 TEX320 P. chinai nymph La Extensa SH intradomicile room clothes bed/ wall/ 3 2006 3 TBR340 R. ecuadoriensis adult Bramaderos SH intradomicile room clothes bed/ wall/ 4 2006 3 TBR341 R. ecuadoriensis adult Bramaderos SH intradomicile room clothes bed/ wall/ 5 2006 15 TEX342 P. chinai adult La Extensa SH intradomicile room clothes room bed/ wall/ 6 2006 15 TEX349 R. ecuadoriensis adult La Extensa SH intradomicile clothes bed/ wall/ 7 2006 8 TGL356 R. ecuadoriensis nymph Galapagos SH intradomicile room clothes bed/ wall/ 8 2006 8 TGL357 R. ecuadoriensis nymph Galapagos SH intradomicile room clothes 9 2007 9 TTBJ370 P. howardi nymph El Bejuco CC peridomicile animal shelter rodent nest 10 2007 9 TTBJ371 P. howardi nymph El Bejuco CC peridomicile animal shelter rodent nest pile of 11 2007 10 TBJ378 R. ecuadoriensis nymph El Bejuco CC peridomicile material leaves/wood 12 2007 10 TBJ379 R. ecuadoriensis nymph El Bejuco CC peridomicile animal shelter Bird nest 13 2007 10 TBJ384 R. ecuadoriensis adult El Bejuco CC sylvatic animal shelter rodent nest 14 2007 18 TBJ401 R. ecuadoriensis nymph El Bejuco CC peridomicile animal shelter Bird nest pile of 15 2007 18 TBJ402 P. howardi nymph El Bejuco CC peridomicile material leaves/wood 109

pile of 16 2007 18 TBJ403 R. ecuadoriensis adult El Bejuco CC peridomicile material leaves/wood pile of 17 2007 18 TBJ405 P. howardi nymph El Bejuco CC peridomicile material leaves/wood bed/ wall/ 18 2008 4 TCE674 P. chinai adult Coamine SH intradomicile room clothes bed/ wall/ 19 2008 4 TCE675 T. carrioni nymph Coamine SH intradomicile room clothes bed/ wall/ 20 2008 5 TCE681 P. chinai nymph Coamine SH intradomicile room clothes bed/ wall/ 21 2008 8 TCE696 P. chinai adult Coamine SH intradomicile room clothes 22 2008 8 TCE699 R. ecuadoriensis adult Coamine SH sylvatic animal shelter rodent nest 23 2008 8 TCE701 R. ecuadoriensis adult Coamine SH sylvatic animal shelter rodent nest 24 2008 8 TCE702 R. ecuadoriensis adult Coamine SH sylvatic animal shelter rodent nest 25 2008 8 TCE703 R. ecuadoriensis adult Coamine SH sylvatic animal shelter rodent nest 26 2008 8 TCE704 R. ecuadoriensis nymph Coamine SH sylvatic animal shelter rodent nest room bed/ wall/ 27 2008 9 TCE706 P. chinai adult Coamine SH intradomicile clothes 28 2008 9 TCE708 P. chinai adult Coamine SH intradomicile animal shelter rodent nest bed/ wall/ 29 2008 9 TCE709 P. chinai nymph Coamine SH intradomicile room clothes bed/ wall/ 30 2008 22 TCE759 R. ecuadoriensis adult Coamine SH intradomicile room clothes bed/ wall/ 31 2008 22 TCE760 P. chinai nymph Coamine SH intradomicile room clothes 32 2008 22 TCE763 P. chinai nymph Coamine SH intradomicile animal shelter rodent nest 33 2008 22 TCE764 P. chinai nymph Coamine SH intradomicile animal shelter rodent nest 34 2008 22 TCE765 P. chinai nymph Coamine SH intradomicile animal shelter rodent nest 110

bed/ wall/ 35 2008 7 TCQ800 R. ecuadoriensis adult Chaquizhca SH intradomicile room clothes bed/ wall/ 36 2008 7 TCQ801 R. ecuadoriensis adult Chaquizhca SH intradomicile room clothes 37 2008 25 TCQ844 P. chinai adult Chaquizhca SH intradomicile animal shelter Bird nest 38 2008 32 TCQ877 P. chinai nymph Chaquizhca SH peridomicile animal shelter Bird nest bed/ wall/ 39 2008 40 TCQ940 R. ecuadoriensis adult Chaquizhca SH intradomicile room clothes bed/ wall/ 40 2008 40 TCQ942 R. ecuadoriensis adult Chaquizhca SH intradomicile room clothes bed/ wall/ 41 2008 40 TCQ952 P. chinai adult Chaquizhca SH intradomicile room clothes 42 2009 5 TGL1266 R. ecuadoriensis nymph Galapagos SH sylvatic animal shelter rodent nest 43 2009 5 TGL1296 R. ecuadoriensis nymph Galapagos SH sylvatic animal shelter rodent nest 44 2009 5 TGL1299 R. ecuadoriensis adult Galapagos SH sylvatic animal shelter rodent nest 45 2009 5 TGL1300 R. ecuadoriensis adult Galapagos SH sylvatic animal shelter rodent nest 46 2009 10 TGL1301 R. ecuadoriensis nymph Galapagos SH sylvatic animal shelter rodent nest 47 2009 10 TGL1309 R. ecuadoriensis adult Galapagos SH sylvatic animal shelter rodent nest bed/ wall/ 48 2009 5 TBR1367 P. chinai adult Bramaderos SH intradomicile room clothes P. bed/ wall/ 49 2009 5 TBR1368 adult Bramaderos SH intradomicile rufotuberculatus room clothes 50 2009 5 TBR1369 R. ecuadoriensis adult Bramaderos SH peridomicile animal shelter Bird nest 51 2009 5 TBR1370 R. ecuadoriensis nymph Bramaderos SH peridomicile animal shelter Bird nest 52 2009 5 TBR1371 R. ecuadoriensis nymph Bramaderos SH peridomicile animal shelter Bird nest 53 2009 5 TBR1372 R. ecuadoriensis nymph Bramaderos SH peridomicile animal shelter Bird nest 54 2009 5 TBR1373 R. ecuadoriensis adult Bramaderos SH peridomicile animal shelter Bird nest 55 2009 6 TBR1375 R. ecuadoriensis nymph Bramaderos SH sylvatic animal shelter rodent nest bed/ wall/ 56 2009 6 TBR1378 R. ecuadoriensis adult Bramaderos SH intradomicile room clothes 111

57 2009 6 TBR1381 R. ecuadoriensis nymph Bramaderos SH sylvatic animal shelter rodent nest 58 2009 6 TBR1382 R. ecuadoriensis adult Bramaderos SH sylvatic animal shelter rodent nest bed/ wall/ 59 2009 6 TBR1383 R. ecuadoriensis adult Bramaderos SH intradomicile room clothes bed/ wall/ 60 2009 6 TBR1384 R. ecuadoriensis nymph Bramaderos SH intradomicile room clothes 61 2009 6 TBR1386 R. ecuadoriensis adult Bramaderos SH peridomicile animal shelter Bird nest 62 2009 6 TBR1388 R. ecuadoriensis adult Bramaderos SH peridomicile animal shelter Bird nest 63 2009 6 TBR1389 R. ecuadoriensis adult Bramaderos SH peridomicile animal shelter Bird nest 64 2009 6 TBR1390 R. ecuadoriensis adult Bramaderos SH peridomicile animal shelter Bird nest 65 2009 6 TBR1398 R. ecuadoriensis adult Bramaderos SH intradomicile animal shelter Bird nest 66 2009 6 TBR1399 R. ecuadoriensis adult Bramaderos SH intradomicile animal shelter Bird nest 67 2009 6 TBR1400 R. ecuadoriensis adult Bramaderos SH intradomicile animal shelter Bird nest 68 2009 7 TBR1404 R. ecuadoriensis adult Bramaderos SH sylvatic animal shelter rodent nest 69 2009 7 TBR1415 R. ecuadoriensis nymph Bramaderos SH sylvatic animal shelter rodent nest 70 2009 7 TBR1416 R. ecuadoriensis nymph Bramaderos SH sylvatic animal shelter rodent nest 71 2009 7 TBR1417 R. ecuadoriensis nymph Bramaderos SH sylvatic animal shelter rodent nest 72 2009 7 TBR1420 R. ecuadoriensis adult Bramaderos SH sylvatic animal shelter rodent nest 73 2009 7 TBR1421 R. ecuadoriensis nymph Bramaderos SH peridomicile animal shelter Bird nest 74 2009 7 TBR1423 R. ecuadoriensis nymph Bramaderos SH peridomicile animal shelter Bird nest 75 2009 7 TBR1433 R. ecuadoriensis adult Bramaderos SH peridomicile animal shelter Bird nest 76 2009 7 TBR1435 R. ecuadoriensis adult Bramaderos SH peridomicile animal shelter Bird nest 77 2009 8 TBR1445 R. ecuadoriensis nymph Bramaderos SH sylvatic animal shelter rodent nest 78 2009 8 TBR1447 R. ecuadoriensis nymph Bramaderos SH sylvatic animal shelter rodent nest 79 2009 8 TBR1448 R. ecuadoriensis adult Bramaderos SH sylvatic animal shelter rodent nest 80 2009 8 TBR1449 R. ecuadoriensis adult Bramaderos SH sylvatic animal shelter rodent nest 81 2009 8 TBR1455 R. ecuadoriensis nymph Bramaderos SH sylvatic animal shelter rodent nest 82 2009 8 TBR1457 R. ecuadoriensis nymph Bramaderos SH sylvatic animal shelter rodent nest 83 2009 8 TBR1459 R. ecuadoriensis nymph Bramaderos SH sylvatic animal shelter rodent nest 112

84 2009 8 TBR1460 R. ecuadoriensis nymph Bramaderos SH sylvatic animal shelter rodent nest 85 2009 8 TBR1461 R. ecuadoriensis nymph Bramaderos SH sylvatic animal shelter rodent nest 86 2009 8 TBR1462 R. ecuadoriensis nymph Bramaderos SH sylvatic animal shelter rodent nest 87 2009 8 TBR1463 R. ecuadoriensis nymph Bramaderos SH sylvatic animal shelter rodent nest 88 2009 8 TBR1464 R. ecuadoriensis adult Bramaderos SH sylvatic animal shelter rodent nest 89 2009 8 TBR1465 R. ecuadoriensis adult Bramaderos SH sylvatic animal shelter rodent nest 90 2009 8 TBR1466 R. ecuadoriensis adult Bramaderos SH sylvatic animal shelter rodent nest 91 2009 8 TBR1468 R. ecuadoriensis nymph Bramaderos SH sylvatic animal shelter rodent nest 92 2009 8 TBR1469 R. ecuadoriensis adult Bramaderos SH sylvatic animal shelter rodent nest 93 2009 12 TBR1472 R. ecuadoriensis nymph Bramaderos SH sylvatic animal shelter rodent nest 94 2009 12 TBR1474 R. ecuadoriensis nymph Bramaderos SH sylvatic animal shelter rodent nest 95 2009 12 TBR1475 R. ecuadoriensis adult Bramaderos SH sylvatic animal shelter rodent nest 96 2009 12 TBR1476 R. ecuadoriensis nymph Bramaderos SH sylvatic animal shelter rodent nest 97 2009 12 TBR1477 R. ecuadoriensis nymph Bramaderos SH sylvatic animal shelter rodent nest 98 2009 12 TBR1480 R. ecuadoriensis nymph Bramaderos SH sylvatic animal shelter rodent nest 99 2009 12 TBR1481 R. ecuadoriensis adult Bramaderos SH sylvatic animal shelter rodent nest 100 2009 12 TBR1483 R. ecuadoriensis adult Bramaderos SH sylvatic animal shelter rodent nest 101 2009 12 TBR1484 R. ecuadoriensis nymph Bramaderos SH sylvatic animal shelter rodent nest 102 2009 12 TBR1491 R. ecuadoriensis nymph Bramaderos SH sylvatic animal shelter rodent nest 103 2009 12 TBR1494 R. ecuadoriensis adult Bramaderos SH sylvatic animal shelter rodent nest 104 2009 13 TBR1496 R. ecuadoriensis adult Bramaderos SH sylvatic animal shelter rodent nest 105 2009 13 TBR1497 R. ecuadoriensis adult Bramaderos SH sylvatic animal shelter rodent nest 106 2009 13 TBR1506 R. ecuadoriensis nymph Bramaderos SH sylvatic animal shelter rodent nest pile of 107 2009 8 TBJ1519 P. howardi nymph El Bejuco CC peridomicile material leaves/wood pile of 108 2009 7 TBJ1520 P. howardi adult El Bejuco CC peridomicile material bricks/tiles pile of 109 2009 7 TBJ1522 P. howardi nymph El Bejuco CC peridomicile material bricks/tiles 113

110 2009 9 TBJ1524 R. ecuadoriensis adult El Bejuco CC peridomicile animal shelter Bird nest collected by light 111 2009 7 TBJ1530 P. howardi adult El Bejuco CC intradomicile inhabitant attracted 112 2009 9 TBJ1531 R. ecuadoriensis adult El Bejuco CC peridomicile animal shelter Bird nest 113 2009 9 TBJ1532 R. ecuadoriensis adult El Bejuco CC peridomicile animal shelter Bird nest 114 2009 14 TBJ1537 R. ecuadoriensis nymph El Bejuco CC peridomicile animal shelter Bird nest pile of 115 2009 15 TBJ1538 P. howardi nymph El Bejuco CC peridomicile material bricks/tiles pile of 116 2009 13 TBJ1541 P. howardi nymph El Bejuco CC peridomicile material bricks/tiles pile of 117 2009 13 TBJ1542 P. howardi nymph El Bejuco CC peridomicile material bricks/tiles pile of 118 2009 15 TBJ1543 P. howardi nymph El Bejuco CC peridomicile material bricks/tiles pile of 119 2009 13 TBJ1553 P. howardi adult El Bejuco CC peridomicile material leaves/wood 120 2009 14 TBJ1554 R. ecuadoriensis nymph El Bejuco CC peridomicile animal shelter Bird nest 121 2009 14 TBJ1555 R. ecuadoriensis nymph El Bejuco CC peridomicile animal shelter Bird nest 122 2009 14 TBJ1556 R. ecuadoriensis adult El Bejuco CC peridomicile animal shelter Bird nest 123 2009 14 TBJ1557 R. ecuadoriensis adult El Bejuco CC peridomicile animal shelter Bird nest 124 2009 15 TBJ1562 R. ecuadoriensis adult El Bejuco CC peridomicile animal shelter Bird nest 125 2009 15 TBJ1563 R. ecuadoriensis nymph El Bejuco CC peridomicile animal shelter Bird nest bed/ wall/ 126 2009 15 TBJ1565 P. howardi adult El Bejuco CC intradomicile room clothes pile of 127 2009 16 TBJ1568 P. howardi nymph El Bejuco CC peridomicile material bricks/tiles 128 2009 14 TBJ1572 R. ecuadoriensis nymph El Bejuco CC sylvatic animal shelter rodent nest 129 2009 15 TBJ1585 R. ecuadoriensis nymph El Bejuco CC sylvatic animal shelter Bird nest 130 2009 14 TBJ1594 R. ecuadoriensis nymph El Bejuco CC peridomicile animal shelter Bird nest 131 2009 14 TBJ1595 R. ecuadoriensis adult El Bejuco CC peridomicile animal shelter Bird nest 114

132 2009 17 TBJ1598 R. ecuadoriensis nymph El Bejuco CC sylvatic animal shelter rodent nest pile of organic 133 2009 16 TBJ1608 P. howardi adult El Bejuco CC peridomicile material matter 134 2009 21 TBJ1677 R. ecuadoriensis nymph El Bejuco CC peridomicile animal shelter Bird nest 135 2009 20 TBJ1721 R. ecuadoriensis adult El Bejuco CC sylvatic animal shelter rodent nest 136 2009 20 TBJ1722 R. ecuadoriensis adult El Bejuco CC sylvatic animal shelter rodent nest 137 2009 20 TBJ1723 R. ecuadoriensis adult El Bejuco CC sylvatic animal shelter rodent nest TCD174 animal shelter 138 2009 25 R. ecuadoriensis nymph Chade CC sylvatic 7 rodent nest TCD174 animal shelter 139 2009 25 R. ecuadoriensis nymph Chade CC sylvatic 8 rodent nest TCD175 animal shelter 140 2009 27 R. ecuadoriensis nymph Chade CC sylvatic 2 rodent nest TCD175 animal shelter 141 2009 27 R. ecuadoriensis nymph Chade CC sylvatic 6 rodent nest TCD178 animal shelter 142 2009 28 R. ecuadoriensis nymph Chade CC sylvatic 2 rodent nest TCD178 animal shelter 143 2009 28 R. ecuadoriensis adult Chade CC sylvatic 5 rodent nest TCD178 animal shelter 144 2009 28 R. ecuadoriensis adult Chade CC sylvatic 6 rodent nest TCD180 animal shelter 145 2009 29 R. ecuadoriensis adult Chade CC sylvatic 1 rodent nest TCD182 animal shelter 146 2009 30 R. ecuadoriensis nymph Chade CC peridomicile 4 Bird nest TCD182 147 2009 30 R. ecuadoriensis nymph Chade CC peridomicile animal shelter Bird nest 5 TCD182 animal shelter 148 2009 30 R. ecuadoriensis adult Chade CC peridomicile 7 Bird nest 115

TCD182 animal shelter 149 2009 30 R. ecuadoriensis nymph Chade CC peridomicile 8 Bird nest TCD182 animal shelter 150 2009 30 R. ecuadoriensis nymph Chade CC peridomicile 9 Bird nest TCD184 animal shelter 151 2009 28 R. ecuadoriensis nymph Chade CC sylvatic 1 rodent nest TCD186 animal shelter 152 2009 31 R. ecuadoriensis nymph Chade CC peridomicile 0 Bird nest TCD186 animal shelter 153 2009 31 R. ecuadoriensis nymph Chade CC peridomicile 1 Bird nest TCD186 animal shelter 154 2009 31 R. ecuadoriensis nymph Chade CC sylvatic 2 rodent nest TCD186 animal shelter 155 2009 31 R. ecuadoriensis nymph Chade CC sylvatic 3 rodent nest TCD186 animal shelter 156 2009 31 R. ecuadoriensis nymph Chade CC sylvatic 7 rodent nest TCD189 animal shelter 157 2009 34 R. ecuadoriensis nymph Chade CC sylvatic 7 rodent nest TCD189 animal shelter 158 2009 34 R. ecuadoriensis nymph Chade CC sylvatic 8 rodent nest TCD190 159 2009 34 R. ecuadoriensis nymph Chade CC sylvatic animal shelter rodent nest 0 TCD190 animal shelter 160 2009 34 R. ecuadoriensis nymph Chade CC sylvatic 1 rodent nest 161 2009 19 TBJ1938 R. ecuadoriensis adult El Bejuco CC peridomicile animal shelter Bird nest 162 2009 19 TBJ1942 R. ecuadoriensis adult El Bejuco CC peridomicile animal shelter Bird nest 163 2009 19 TBJ1944 R. ecuadoriensis adult El Bejuco CC sylvatic animal shelter rodent nest 164 2009 19 TBJ1945 R. ecuadoriensis nymph El Bejuco CC sylvatic animal shelter rodent nest 165 2009 19 TBJ1948 R. ecuadoriensis adult El Bejuco CC peridomicile animal shelter Bird nest 166 2009 19 TBJ1949 R. ecuadoriensis adult El Bejuco CC peridomicile animal shelter Bird nest 116

167 2009 19 TBJ1950 R. ecuadoriensis adult El Bejuco CC peridomicile animal shelter Bird nest 168 2009 15 TBJ1951 R. ecuadoriensis adult El Bejuco CC sylvatic animal shelter rodent nest 169 2009 19 TBJ1952 R. ecuadoriensis adult El Bejuco CC peridomicile animal shelter Bird nest 170 2009 19 TBJ1953 R. ecuadoriensis adult El Bejuco CC peridomicile animal shelter Bird nest 171 2009 19 TBJ1954 R. ecuadoriensis adult El Bejuco CC peridomicile animal shelter Bird nest 172 2009 15 TBJ1955 R. ecuadoriensis adult El Bejuco CC sylvatic animal shelter rodent nest 173 2009 15 TBJ1956 R. ecuadoriensis adult El Bejuco CC sylvatic animal shelter rodent nest 174 2009 15 TBJ1957 R. ecuadoriensis adult El Bejuco CC sylvatic animal shelter rodent nest 175 2009 15 TBJ1958 R. ecuadoriensis adult El Bejuco CC sylvatic animal shelter rodent nest 176 2009 15 TBJ1960 R. ecuadoriensis nymph El Bejuco CC sylvatic animal shelter rodent nest 177 2009 15 TBJ1961 R. ecuadoriensis nymph El Bejuco CC sylvatic animal shelter rodent nest 178 2009 15 TBJ1962 R. ecuadoriensis nymph El Bejuco CC sylvatic animal shelter rodent nest 179 2009 20 TBJ1966 R. ecuadoriensis adult El Bejuco CC sylvatic animal shelter rodent nest 180 2009 20 TBJ1967 R. ecuadoriensis adult El Bejuco CC sylvatic animal shelter rodent nest 181 2009 20 TBJ1968 R. ecuadoriensis adult El Bejuco CC sylvatic animal shelter rodent nest 182 2009 20 TBJ1970 R. ecuadoriensis adult El Bejuco CC sylvatic animal shelter rodent nest 183 2009 20 TBJ1971 R. ecuadoriensis adult El Bejuco CC sylvatic animal shelter rodent nest 184 2009 20 TBJ1972 R. ecuadoriensis nymph El Bejuco CC sylvatic animal shelter rodent nest Tablada del animal shelter 185 2009 23 TTO1990 R. ecuadoriensis adult CC intradomicile algodón Bird nest Tablada del animal shelter 186 2009 23 TTO1992 R. ecuadoriensis adult CC intradomicile algodón Bird nest Tablada del animal shelter 187 2009 26 TTO2002 R. ecuadoriensis adult CC peridomicile algodón Bird nest 188 2009 18 TDZ2003 R. ecuadoriensis adult Danzarin CC sylvatic animal shelter Bird nest 189 2009 24 TDZ2005 R. ecuadoriensis adult Danzarin CC sylvatic animal shelter Bird nest 190 2009 35 TBJ2008 R. ecuadoriensis adult El Bejuco CC peridomicile animal shelter Bird nest 191 2009 35 TBJ2009 R. ecuadoriensis adult El Bejuco CC peridomicile animal shelter Bird nest 192 2009 35 TBJ2010 R. ecuadoriensis adult El Bejuco CC peridomicile animal shelter Bird nest 117

193 2009 35 TBJ2011 R. ecuadoriensis nymph El Bejuco CC peridomicile animal shelter Bird nest 194 2009 25 TDZ2016 R. ecuadoriensis adult Danzarin CC sylvatic animal shelter rodent nest 195 2009 25 TDZ2022 R. ecuadoriensis adult Danzarin CC peridomicile animal shelter Bird nest 196 2009 27 TDZ2026 R. ecuadoriensis adult Danzarin CC peridomicile animal shelter Bird nest 197 2009 27 TDZ2027 R. ecuadoriensis adult Danzarin CC peridomicile animal shelter Bird nest Tablada del animal shelter 198 2009 29 TTO2051 R. ecuadoriensis nymph CC intradomicile algodón Bird nest 199 2009 31 TDZ2057 R. ecuadoriensis adult Danzarin CC sylvatic animal shelter rodent nest 200 2009 31 TDZ2068 R. ecuadoriensis nymph Danzarin CC sylvatic animal shelter Bird nest 201 2009 31 TDZ2074 R. ecuadoriensis adult Danzarin CC peridomicile animal shelter Bird nest Tablada del bed/ wall/ 202 2009 35 TTO2086 R. ecuadoriensis nymph CC intradomicile algodón room clothes 203 2009 33 TDZ2101 R. ecuadoriensis nymph Danzarin CC peridomicile animal shelter Bird nest 204 TDZ2110 R. ecuadoriensis adult Danzarin CC peridomicile animal shelter Bird nest Tablada del animal shelter 205 2009 35 TTO2114 R. ecuadoriensis adult CC sylvatic algodón rodent nest 206 2009 40 TDZ2198 R. ecuadoriensis nymph Danzarin CC sylvatic animal shelter rodent nest 207 2009 40 TDZ2199 R. ecuadoriensis nymph Danzarin CC peridomicile animal shelter Bird nest 208 2009 40 TDZ2200 R. ecuadoriensis nymph Danzarin CC peridomicile animal shelter Bird nest 209 2009 40 TDZ2201 R. ecuadoriensis nymph Danzarin CC peridomicile animal shelter Bird nest 210 2009 6 TBJ2239 R. ecuadoriensis nymph El Bejuco CC sylvatic animal shelter rodent nest 211 2009 7 TBJ2240 R. ecuadoriensis nymph El Bejuco CC sylvatic animal shelter rodent nest 212 2009 8 TBJ2242 R. ecuadoriensis nymph El Bejuco CC peridomicile animal shelter Bird nest 213 2009 8 TBJ2243 R. ecuadoriensis adult El Bejuco CC peridomicile animal shelter Bird nest 214 2009 8 TBJ2244 R. ecuadoriensis nymph El Bejuco CC peridomicile animal shelter Bird nest 215 2009 8 TBJ2246 R. ecuadoriensis nymph El Bejuco CC peridomicile animal shelter Bird nest 216 2009 8 TBJ2248 R. ecuadoriensis nymph El Bejuco CC peridomicile animal shelter Bird nest 217 2009 8 TBJ2249 R. ecuadoriensis adult El Bejuco CC peridomicile animal shelter Bird nest 218 2009 8 TBJ2250 R. ecuadoriensis nymph El Bejuco CC peridomicile animal shelter Bird nest 118

219 2009 8 TBJ2251 R. ecuadoriensis adult El Bejuco CC peridomicile animal shelter Bird nest 220 2009 8 TBJ2252 R. ecuadoriensis nymph El Bejuco CC peridomicile animal shelter Bird nest 221 2009 8 TBJ2254 R. ecuadoriensis nymph El Bejuco CC peridomicile animal shelter Bird nest 222 2009 6 TBJ2257 R. ecuadoriensis adult El Bejuco CC peridomicile animal shelter Bird nest 223 2009 7 TBJ2258 R. ecuadoriensis nymph El Bejuco CC sylvatic animal shelter rodent nest 224 2009 7 TBJ2259 R. ecuadoriensis nymph El Bejuco CC sylvatic animal shelter rodent nest 225 2009 8 TBJ2260 R. ecuadoriensis adult El Bejuco CC sylvatic animal shelter rodent nest 226 2009 8 TBJ2266 R. ecuadoriensis adult El Bejuco CC peridomicile animal shelter Bird nest 227 2009 8 TBJ2267 R. ecuadoriensis adult El Bejuco CC peridomicile animal shelter Bird nest 228 2009 6 TBJ2268 R. ecuadoriensis nymph El Bejuco CC sylvatic animal shelter rodent nest 229 2009 6 TBJ2270 R. ecuadoriensis nymph El Bejuco CC sylvatic animal shelter rodent nest 230 2009 6 TBJ2271 R. ecuadoriensis nymph El Bejuco CC sylvatic animal shelter rodent nest 231 2009 6 TBJ2272 R. ecuadoriensis nymph El Bejuco CC sylvatic animal shelter rodent nest 232 2009 6 TBJ2273 R. ecuadoriensis nymph El Bejuco CC sylvatic animal shelter rodent nest 233 2009 6 TBJ2274 R. ecuadoriensis adult El Bejuco CC sylvatic animal shelter rodent nest 234 2009 6 TBJ2275 R. ecuadoriensis adult El Bejuco CC sylvatic animal shelter rodent nest 235 2009 6 TBJ2276 R. ecuadoriensis adult El Bejuco CC sylvatic animal shelter rodent nest 236 2009 7 TBJ2277 R. ecuadoriensis nymph El Bejuco CC sylvatic animal shelter rodent nest 237 2009 9 TBJ2278 R. ecuadoriensis nymph El Bejuco CC sylvatic animal shelter rodent nest 238 2009 7 TBJ2279 R. ecuadoriensis nymph El Bejuco CC sylvatic animal shelter rodent nest 239 2009 7 TBJ2280 R. ecuadoriensis nymph El Bejuco CC sylvatic animal shelter rodent nest 240 2009 7 TBJ2281 R. ecuadoriensis nymph El Bejuco CC sylvatic animal shelter rodent nest 241 2009 7 TBJ2282 R. ecuadoriensis nymph El Bejuco CC sylvatic animal shelter rodent nest 242 2009 11 TBJ2291 R. ecuadoriensis nymph El Bejuco CC peridomicile animal shelter Bird nest 243 2009 11 TBJ2292 R. ecuadoriensis nymph El Bejuco CC peridomicile animal shelter Bird nest 244 2009 11 TBJ2295 R. ecuadoriensis adult El Bejuco CC peridomicile animal shelter Bird nest 245 2010 21 TBJ2308 P. howardi nymph El Bejuco CC sylvatic animal shelter rodent nest 246 2010 21 TBJ2310 R. ecuadoriensis adult El Bejuco CC peridomicile animal shelter Bird nest 119

247 2010 5 TBJ2448 R. ecuadoriensis nymph El Bejuco CC sylvatic animal shelter rodent nest 248 2010 9 TBJ2452 R. ecuadoriensis nymph El Bejuco CC sylvatic animal shelter Bird nest 249 2010 7 TBJ2455 R. ecuadoriensis nymph El Bejuco CC sylvatic animal shelter rodent nest 250 2010 7 TBJ2456 R. ecuadoriensis adult El Bejuco CC sylvatic animal shelter rodent nest TCQ249 bed/ wall/ 251 2010 13 P. chinai adult Chaquizhca SH intradomicile room 2 clothes TCQ249 bed/ wall/ 252 2010 12 R. ecuadoriensis adult Chaquizhca SH intradomicile 3 room clothes TCQ249 bed/ wall/ 253 2010 12 R. ecuadoriensis adult Chaquizhca SH intradomicile 4 room clothes TCQ249 bed/ wall/ 254 2010 12 R. ecuadoriensis adult Chaquizhca SH intradomicile room 7 clothes TCQ250 bed/ wall/ 255 2010 12 R. ecuadoriensis adult Chaquizhca SH intradomicile 2 room clothes 256 2010 22 TCE2532 R. ecuadoriensis adult Coamine SH peridomicile animal shelter Bird nest 257 2010 23 TCE2534 R. ecuadoriensis adult Coamine SH peridomicile animal shelter Bird nest 258 2010 33 TCE2538 R. ecuadoriensis adult Coamine SH peridomicile animal shelter Bird nest 259 2010 36 TCE2542 R. ecuadoriensis adult Coamine SH peridomicile animal shelter Bird nest 260 2010 36 TCE2543 R. ecuadoriensis adult Coamine SH peridomicile animal shelter Bird nest 261 2010 23 TCE2557 R. ecuadoriensis adult Coamine SH peridomicile animal shelter Bird nest 262 2010 26 TCE2564 R. ecuadoriensis adult Coamine SH peridomicile animal shelter Bird nest 263 2010 26 TCE2565 R. ecuadoriensis adult Coamine SH peridomicile animal shelter Bird nest 264 2010 26 TCE2566 R. ecuadoriensis adult Coamine SH peridomicile animal shelter Bird nest 265 2010 26 TCE2567 R. ecuadoriensis adult Coamine SH peridomicile animal shelter Bird nest 266 2010 26 TCE2568 R. ecuadoriensis adult Coamine SH peridomicile animal shelter Bird nest 267 2010 26 TCE2569 R. ecuadoriensis adult Coamine SH peridomicile animal shelter Bird nest 268 2010 26 TCE2570 R. ecuadoriensis adult Coamine SH peridomicile animal shelter Bird nest 269 2010 26 TCE2572 R. ecuadoriensis nymph Coamine SH peridomicile animal shelter Bird nest 270 2010 27 TCE2585 R. ecuadoriensis adult Coamine SH peridomicile animal shelter Bird nest 120

271 2010 27 TCE2586 R. ecuadoriensis adult Coamine SH peridomicile animal shelter Bird nest 272 2010 27 TCE2587 R. ecuadoriensis adult Coamine SH peridomicile animal shelter Bird nest 273 2010 27 TCE2588 R. ecuadoriensis adult Coamine SH peridomicile animal shelter Bird nest 274 2010 27 TCE2589 R. ecuadoriensis adult Coamine SH peridomicile animal shelter Bird nest 275 2010 27 TCE2590 R. ecuadoriensis nymph Coamine SH peridomicile animal shelter Bird nest TCQ259 bed/ wall/ 276 2010 19 P. chinai nymph Chaquizhca SH intradomicile 5 room clothes TCQ259 bed/ wall/ 277 2010 19 P. chinai nymph Chaquizhca SH intradomicile room 6 clothes 278 2010 28 TCE2598 R. ecuadoriensis adult Coamine SH peridomicile animal shelter Bird nest bed/ wall/ 279 2010 28 TCE2602 T. carrioni nymph Coamine SH intradomicile room clothes 280 2010 28 TCE2603 R. ecuadoriensis adult Coamine SH peridomicile animal shelter Bird nest TBM277 pile of 281 2011 6 P. chinai nymph Bellamaria SH intradomicile 5 material bricks/tiles TBM277 pile of 282 2011 6 P. chinai nymph Bellamaria SH intradomicile 6 material bricks/tiles TBM277 pile of 283 2011 6 P. chinai nymph Bellamaria SH intradomicile 7 material bricks/tiles TBM277 pile of 284 2011 6 P. chinai nymph Bellamaria SH intradomicile 8 material bricks/tiles TBM277 pile of 285 2011 6 P. chinai nymph Bellamaria SH intradomicile 9 material bricks/tiles TBM278 pile of 286 2011 6 P. chinai nymph Bellamaria SH intradomicile 1 material bricks/tiles TBM278 pile of 287 2011 6 P. chinai nymph Bellamaria SH intradomicile 2 material bricks/tiles TBM278 pile of 288 2011 6 P. chinai nymph Bellamaria SH intradomicile 4 material bricks/tiles 121

TBM278 pile of 289 2011 6 P. chinai nymph Bellamaria SH intradomicile 5 material bricks/tiles TBM278 pile of 290 2011 6 P. chinai nymph Bellamaria SH intradomicile 6 material bricks/tiles TBM278 pile of 291 2011 7 P. chinai nymph Bellamaria SH intradomicile 9 material bricks/tiles TBM279 pile of 292 2011 7 P. chinai nymph Bellamaria SH intradomicile 0 material bricks/tiles TBM279 pile of 293 2011 7 P. chinai nymph Bellamaria SH intradomicile 1 material bricks/tiles TBM279 pile of 294 2011 7 P. chinai nymph Bellamaria SH intradomicile 2 material bricks/tiles TBM279 pile of 295 2011 7 P. chinai nymph Bellamaria SH intradomicile 4 material bricks/tiles TBM279 pile of 296 2011 7 P. chinai nymph Bellamaria SH intradomicile 5 material bricks/tiles TBM279 pile of 297 2011 7 P. chinai nymph Bellamaria SH intradomicile 6 material bricks/tiles TBM279 pile of 298 2011 7 P. chinai nymph Bellamaria SH intradomicile 7 material bricks/tiles TBM279 pile of 299 2011 7 P. chinai nymph Bellamaria SH intradomicile 8 material bricks/tiles TBM280 bed/ wall/ 300 2011 7 P. chinai nymph Bellamaria SH intradomicile 0 room clothes TBM280 bed/ wall/ 301 2011 8 P. chinai nymph Bellamaria SH intradomicile room 3 clothes TBM280 bed/ wall/ 302 2011 8 P. chinai nymph Bellamaria SH intradomicile 4 room clothes TBM280 bed/ wall/ 303 2011 8 P. chinai nymph Bellamaria SH intradomicile 5 room clothes 122

TBM280 bed/ wall/ 304 2011 8 P. chinai nymph Bellamaria SH intradomicile 6 room clothes TBM280 bed/ wall/ 305 2011 8 P. chinai nymph Bellamaria SH intradomicile room 7 clothes TBM280 bed/ wall/ 306 2011 8 P. chinai nymph Bellamaria SH intradomicile 8 room clothes TBM280 bed/ wall/ 307 2011 8 P. chinai nymph Bellamaria SH intradomicile 9 room clothes TBM281 bed/ wall/ 308 2011 8 P. chinai nymph Bellamaria SH intradomicile 1 room clothes TBM281 bed/ wall/ 309 2011 8 P. chinai nymph Bellamaria SH intradomicile 2 room clothes TBM281 bed/ wall/ 310 2011 8 P. chinai nymph Bellamaria SH intradomicile 3 room clothes TCQ281 animal shelter 311 2011 11 R. ecuadoriensis adult Chaquizhca SH peridomicile 4 Bird nest TCQ281 312 2011 11 R. ecuadoriensis adult Chaquizhca SH peridomicile animal shelter Bird nest 5 TCQ281 bed/ wall/ 313 2011 10 P. chinai adult Chaquizhca SH intradomicile room 6 clothes TCQ281 bed/ wall/ 314 2011 4 R. ecuadoriensis adult Chaquizhca SH intradomicile 8 room clothes TCQ281 bed/ wall/ 315 2011 4 R. ecuadoriensis nymph Chaquizhca SH intradomicile 9 room clothes TBM282 bed/ wall/ 316 2011 10 P. chinai nymph Bellamaria SH intradomicile 0 room clothes TBM282 317 2011 10 P. chinai adult Bellamaria SH intradomicile room rodent nest 1 TBM282 bed/ wall/ 318 2011 10 P. chinai nymph Bellamaria SH intradomicile 2 room clothes 123

TBM282 319 2011 10 P. chinai nymph Bellamaria SH intradomicile room rodent nest 3 TBM282 bed/ wall/ 320 2011 10 P. chinai nymph Bellamaria SH intradomicile 4 room clothes TCQ284 animal shelter 321 2011 17 R. ecuadoriensis adult Chaquizhca SH peridomicile 0 Bird nest TCQ284 animal shelter 322 2011 17 R. ecuadoriensis adult Chaquizhca SH peridomicile 2 Bird nest TCQ284 animal shelter 323 2011 17 R. ecuadoriensis adult Chaquizhca SH peridomicile 3 Bird nest TCQ284 animal shelter 324 2011 17 R. ecuadoriensis adult Chaquizhca SH peridomicile 4 Bird nest TBM284 animal shelter 325 2011 13 R. ecuadoriensis adult Bellamaria SH sylvatic 5 rodent nest TBM284 animal shelter 326 2011 13 R. ecuadoriensis adult Bellamaria SH sylvatic 6 rodent nest TBM284 animal shelter 327 2011 19 R. ecuadoriensis nymph Bellamaria SH peridomicile 8 Bird nest TCQ285 pile of 328 2011 18 R. ecuadoriensis adult Chaquizhca SH peridomicile 0 material bricks/tiles TCQ285 pile of 329 2011 18 R. ecuadoriensis adult Chaquizhca SH peridomicile 2 material bricks/tiles TCQ285 animal shelter 330 2011 19 R. ecuadoriensis nymph Chaquizhca SH peridomicile 4 Bird nest TCQ286 bed/ wall/ 331 2011 18 P. chinai adult Chaquizhca SH intradomicile 3 room clothes TCQ286 332 2011 22 R. ecuadoriensis adult Chaquizhca SH peridomicile animal shelter rodent nest 6 TCQ286 333 2011 22 R. ecuadoriensis adult Chaquizhca SH peridomicile animal shelter rodent nest 7 124

TCQ286 334 2011 22 R. ecuadoriensis adult Chaquizhca SH peridomicile animal shelter rodent nest 8 TCQ286 335 2011 22 R. ecuadoriensis nymph Chaquizhca SH peridomicile animal shelter rodent nest 9 TBM287 bed/ wall/ 336 2011 24 R. ecuadoriensis adult Bellamaria SH intradomicile 3 room clothes TCQ287 337 2011 22 R. ecuadoriensis adult Chaquizhca SH peridomicile animal shelter Bird nest 5 TCQ287 animal shelter 338 2011 22 R. ecuadoriensis adult Chaquizhca SH peridomicile 7 Bird nest TCQ287 animal shelter 339 2011 22 R. ecuadoriensis nymph Chaquizhca SH peridomicile 8 Bird nest TCQ288 bed/ wall/ 340 2011 23 R. ecuadoriensis nymph Chaquizhca SH intradomicile 0 room clothes TCQ288 pile of 341 2011 22 R. ecuadoriensis nymph Chaquizhca SH peridomicile 1 material bricks/tiles TCQ288 pile of 342 2011 22 R. ecuadoriensis adult Chaquizhca SH peridomicile 2 material bricks/tiles TCQ288 pile of 343 2011 22 R. ecuadoriensis nymph Chaquizhca SH peridomicile 3 material bricks/tiles TCQ288 pile of 344 2011 22 R. ecuadoriensis nymph Chaquizhca SH peridomicile 5 material bricks/tiles TCQ288 pile of 345 2011 22 R. ecuadoriensis nymph Chaquizhca SH peridomicile 8 material bricks/tiles TCQ289 pile of 346 2011 22 R. ecuadoriensis nymph Chaquizhca SH peridomicile 0 material bricks/tiles TBM289 animal shelter 347 2011 25 P. chinai nymph Bellamaria SH peridomicile 2 dog bed TBM289 animal shelter 348 2011 25 P. chinai nymph Bellamaria SH peridomicile 3 dog bed 125

TBM289 bed/ wall/ 349 2011 24 R. ecuadoriensis adult Bellamaria SH intradomicile 4 room clothes TCQ289 bed/ wall/ 350 2011 22 P. chinai nymph Chaquizhca SH intradomicile 5 room clothes TCQ289 animal shelter 351 2011 21 R. ecuadoriensis adult Chaquizhca SH peridomicile 6 Bird nest TCQ289 animal shelter 352 2011 21 R. ecuadoriensis nymph Chaquizhca SH peridomicile 8 Bird nest TCQ290 bed/ wall/ 353 2011 35 R. ecuadoriensis nymph Chaquizhca SH intradomicile 2 room clothes TBM290 animal shelter 354 2011 26 R. ecuadoriensis nymph Bellamaria SH peridomicile 3 Bird nest TCQ290 animal shelter 355 2011 25 R. ecuadoriensis nymph Chaquizhca SH peridomicile 6 rodent nest TCQ290 animal shelter 356 2011 25 R. ecuadoriensis nymph Chaquizhca SH peridomicile 7 rodent nest TCQ291 357 2011 25 R. ecuadoriensis nymph Chaquizhca SH peridomicile animal shelter rodent nest 0 TCQ291 animal shelter 358 2011 25 R. ecuadoriensis nymph Chaquizhca SH peridomicile 2 rodent nest TCQ292 animal shelter 359 2011 25 R. ecuadoriensis nymph Chaquizhca SH peridomicile 2 rodent nest TCQ292 animal shelter 360 2011 24 R. ecuadoriensis adult Chaquizhca SH peridomicile 8 Bird nest TCQ292 animal shelter 361 2011 25 R. ecuadoriensis adult Chaquizhca SH peridomicile 9 Bird nest TCQ293 animal shelter 362 2011 25 R. ecuadoriensis adult Chaquizhca SH peridomicile 0 Bird nest TCQ293 animal shelter 363 2011 25 R. ecuadoriensis adult Chaquizhca SH peridomicile 4 Bird nest 126

TCQ293 animal shelter 364 2011 25 R. ecuadoriensis nymph Chaquizhca SH peridomicile 6 Bird nest TCQ293 365 2011 17 R. ecuadoriensis adult Chaquizhca SH peridomicile animal shelter Bird nest 8 TCQ293 animal shelter 366 2011 25 R. ecuadoriensis nymph Chaquizhca SH peridomicile 9 Bird nest TCQ294 animal shelter 367 2011 25 R. ecuadoriensis nymph Chaquizhca SH peridomicile 0 Bird nest TCQ294 animal shelter 368 2011 25 R. ecuadoriensis nymph Chaquizhca SH peridomicile 1 Bird nest TCQ294 369 2011 25 R. ecuadoriensis nymph Chaquizhca SH peridomicile animal shelter Bird nest 2 TCQ294 animal shelter 370 2011 25 R. ecuadoriensis nymph Chaquizhca SH peridomicile 3 Bird nest TCQ294 animal shelter 371 2011 25 R. ecuadoriensis nymph Chaquizhca SH peridomicile 4 Bird nest TCQ294 animal shelter 372 2011 25 R. ecuadoriensis nymph Chaquizhca SH peridomicile 6 Bird nest TBM294 373 2011 30 R. ecuadoriensis adult Bellamaria SH peridomicile animal shelter Bird nest 8 TBM294 animal shelter 374 2011 30 R. ecuadoriensis adult Bellamaria SH peridomicile 9 Bird nest TBM295 animal shelter 375 2011 30 R. ecuadoriensis adult Bellamaria SH peridomicile 0 Bird nest TBM295 376 2011 30 R. ecuadoriensis adult Bellamaria SH peridomicile animal shelter Bird nest 2 TBM295 animal shelter 377 2011 30 R. ecuadoriensis adult Bellamaria SH peridomicile 4 Bird nest TBM295 animal shelter 378 2011 30 R. ecuadoriensis adult Bellamaria SH peridomicile 6 Bird nest 127

TBM295 animal shelter 379 2011 30 R. ecuadoriensis adult Bellamaria SH peridomicile 7 Bird nest TBM295 animal shelter 380 2011 30 R. ecuadoriensis adult Bellamaria SH peridomicile 8 Bird nest TBM296 animal shelter 381 2011 30 R. ecuadoriensis adult Bellamaria SH peridomicile 0 Bird nest TBM296 animal shelter 382 2011 32 R. ecuadoriensis adult Bellamaria SH peridomicile 1 Bird nest TBM296 383 2011 32 R. ecuadoriensis adult Bellamaria SH peridomicile animal shelter Bird nest 3 TBM296 384 2011 32 R. ecuadoriensis adult Bellamaria SH peridomicile animal shelter Bird nest 6 TBM296 385 2011 32 R. ecuadoriensis adult Bellamaria SH peridomicile animal shelter Bird nest 7 TBM297 386 2011 35 R. ecuadoriensis nymph Bellamaria SH peridomicile animal shelter Bird nest 7 TBM296 animal shelter 387 2011 32 R. ecuadoriensis nymph Bellamaria SH peridomicile 9 Bird nest TBM297 animal shelter 388 2011 35 R. ecuadoriensis adult Bellamaria SH peridomicile 1 Bird nest TBM297 389 2011 35 R. ecuadoriensis adult Bellamaria SH peridomicile animal shelter Bird nest 8 TBM297 animal shelter 390 2011 36 R. ecuadoriensis adult Bellamaria SH peridomicile 9 Bird nest TBM298 animal shelter 391 2011 36 R. ecuadoriensis adult Bellamaria SH peridomicile 0 Bird nest TBM298 animal shelter 392 2011 36 R. ecuadoriensis nymph Bellamaria SH peridomicile 3 Bird nest TCQ298 animal shelter 393 2011 32 R. ecuadoriensis nymph Chaquizhca SH peridomicile 5 Bird nest 128

TCQ298 animal shelter 394 2011 32 R. ecuadoriensis nymph Chaquizhca SH peridomicile 6 Bird nest TCQ298 animal shelter 395 2011 32 R. ecuadoriensis nymph Chaquizhca SH peridomicile 9 Bird nest TCQ299 animal shelter 396 2011 32 R. ecuadoriensis nymph Chaquizhca SH peridomicile 0 Bird nest TCQ299 animal shelter 397 2011 32 R. ecuadoriensis nymph Chaquizhca SH peridomicile 1 Bird nest TCQ299 398 2011 32 R. ecuadoriensis nymph Chaquizhca SH peridomicile animal shelter Bird nest 2 Estero de animal shelter TNC300 399 2011 30 R. ecuadoriensis adult Noche CC peridomicile 1 Arriba rodent nest Estero de animal shelter TNC300 400 2011 30 R. ecuadoriensis adult Noche CC peridomicile 2 Arriba rodent nest Estero de animal shelter TNC303 401 2011 22 R. ecuadoriensis nymph Noche CC intradomicile 1 Arriba Bird nest Estero de animal shelter TNC303 402 2011 22 R. ecuadoriensis nymph Noche CC peridomicile 7 Arriba rodent nest Estero de TNC303 403 2011 22 R. ecuadoriensis nymph Noche CC peridomicile pile of 9 Arriba material leaves/wood Estero de TNC304 404 2011 22 R. ecuadoriensis adult Noche CC peridomicile pile of 0 Arriba material leaves/wood Estero de animal shelter TNC304 405 2011 22 R. ecuadoriensis nymph Noche CC peridomicile 5 Arriba rodent nest 129

TCQ308 animal shelter 406 2012 12 R. ecuadoriensis adult Chaquizhca SH sylvatic 4 rodent nest TCQ308 407 2012 12 R. ecuadoriensis adult Chaquizhca SH sylvatic animal shelter rodent nest 5 TCQ308 408 2012 11 R. ecuadoriensis adult Chaquizhca SH sylvatic animal shelter rodent nest 6 TCQ308 animal shelter 409 2012 10 R. ecuadoriensis nymph Chaquizhca SH sylvatic 8 rodent nest TCQ308 animal shelter 410 2012 10 R. ecuadoriensis nymph Chaquizhca SH sylvatic 9 rodent nest TCQ309 animal shelter 411 2012 10 R. ecuadoriensis nymph Chaquizhca SH sylvatic 1 rodent nest TCQ309 animal shelter 412 2012 10 R. ecuadoriensis nymph Chaquizhca SH sylvatic 2 rodent nest TCQ309 413 2012 10 R. ecuadoriensis nymph Chaquizhca SH sylvatic animal shelter rodent nest 3 414 2012 5 TBC3098 R. ecuadoriensis nymph Bellamaria SH sylvatic animal shelter rodent nest TBM310 animal shelter 415 2012 6 R. ecuadoriensis adult Bellamaria SH sylvatic 1 rodent nest TBM310 collected by light 416 2012 4 R. ecuadoriensis adult Bellamaria SH intradomicile 2 inhabitant attracted TCQ310 animal shelter 417 2012 11 R. ecuadoriensis nymph Chaquizhca SH sylvatic 3 rodent nest TCQ310 animal shelter 418 2012 11 R. ecuadoriensis nymph Chaquizhca SH sylvatic 4 rodent nest TCQ310 animal shelter 419 2012 14 R. ecuadoriensis nymph Chaquizhca SH sylvatic 7 rodent nest TCQ311 animal shelter 420 2012 24 R. ecuadoriensis adult Chaquizhca SH peridomicile 1 Bird nest TCQ311 animal shelter 421 2012 24 R. ecuadoriensis adult Chaquizhca SH peridomicile 3 Bird nest 130

TCQ311 animal shelter 422 2012 24 R. ecuadoriensis adult Chaquizhca SH peridomicile 4 Bird nest TCQ311 animal shelter 423 2012 24 R. ecuadoriensis adult Chaquizhca SH peridomicile 5 Bird nest TCQ311 424 2012 24 R. ecuadoriensis nymph Chaquizhca SH peridomicile animal shelter Bird nest 6 TBM313 animal shelter 425 2012 27 R. ecuadoriensis nymph Bellamaria SH sylvatic 1 rodent nest TBM313 426 2012 27 R. ecuadoriensis adult Bellamaria SH sylvatic animal shelter rodent nest 3 TBM313 animal shelter 427 2012 27 R. ecuadoriensis adult Bellamaria SH sylvatic 4 rodent nest TCQ313 animal shelter 428 2012 34 P. chinai nymph Chaquizhca SH peridomicile 9 Bird nest TBM314 animal shelter 429 2012 32 R. ecuadoriensis nymph Bellamaria SH sylvatic 3 rodent nest TBM314 animal shelter 430 2012 32 R. ecuadoriensis nymph Bellamaria SH sylvatic 4 rodent nest TBM314 animal shelter 431 2012 30 R. ecuadoriensis nymph Bellamaria SH peridomicile 8 Bird nest TBM315 animal shelter 432 2012 30 R. ecuadoriensis nymph Bellamaria SH peridomicile 1 Bird nest TBM318 animal shelter 433 2012 34 R. ecuadoriensis adult Bellamaria SH peridomicile 5 Bird nest TBM318 animal shelter 434 2012 34 R. ecuadoriensis adult Bellamaria SH peridomicile 6 Bird nest TBM319 animal shelter 435 2012 35 R. ecuadoriensis nymph Bellamaria SH sylvatic 0 rodent nest TCQ319 animal shelter 436 2012 36 R. ecuadoriensis nymph Chaquizhca SH peridomicile 7 Bird nest 131

TCQ321 animal shelter 437 2012 40 R. ecuadoriensis nymph Chaquizhca SH peridomicile 5 Bird nest TCQ321 animal shelter 438 2012 40 R. ecuadoriensis nymph Chaquizhca SH peridomicile 8 Bird nest TCQ321 439 2012 40 R. ecuadoriensis nymph Chaquizhca SH peridomicile animal shelter Bird nest 9 TCQ322 animal shelter 440 2012 40 R. ecuadoriensis nymph Chaquizhca SH peridomicile 2 Bird nest TCQ322 animal shelter 441 2012 40 R. ecuadoriensis nymph Chaquizhca SH peridomicile 3 Bird nest TCQ322 animal shelter 442 2012 40 R. ecuadoriensis nymph Chaquizhca SH peridomicile 4 Bird nest TCQ322 animal shelter 443 2012 40 R. ecuadoriensis nymph Chaquizhca SH peridomicile 5 Bird nest TBM323 animal shelter 444 2012 39 R. ecuadoriensis nymph Bellamaria SH sylvatic 4 rodent nest TBM323 animal shelter 445 2012 39 R. ecuadoriensis nymph Bellamaria SH sylvatic 5 rodent nest TBM323 animal shelter 446 2012 39 R. ecuadoriensis nymph Bellamaria SH sylvatic 6 rodent nest TBM323 animal shelter 447 2012 39 R. ecuadoriensis nymph Bellamaria SH sylvatic 9 rodent nest TBM324 animal shelter 448 2012 39 R. ecuadoriensis nymph Bellamaria SH sylvatic 1 rodent nest TBM324 449 2012 39 R. ecuadoriensis nymph Bellamaria SH sylvatic animal shelter rodent nest 2 TBM328 450 2013 2 R. ecuadoriensis adult Bellamaria SH sylvatic animal shelter rodent nest 3 TBM329 animal shelter 451 2013 2 R. ecuadoriensis adult Bellamaria SH sylvatic 0 rodent nest 132

TBM329 animal shelter 452 2013 2 R. ecuadoriensis adult Bellamaria SH sylvatic 1 rodent nest TBM329 animal shelter 453 2013 2 R. ecuadoriensis adult Bellamaria SH sylvatic 2 rodent nest TBM329 animal shelter 454 2013 2 R. ecuadoriensis adult Bellamaria SH sylvatic 4 rodent nest TBM329 animal shelter 455 2013 2 R. ecuadoriensis adult Bellamaria SH sylvatic 5 rodent nest TBM329 animal shelter 456 2013 2 R. ecuadoriensis adult Bellamaria SH sylvatic 6 rodent nest TBM329 animal shelter 457 2013 2 R. ecuadoriensis adult Bellamaria SH sylvatic 7 rodent nest TBM330 animal shelter 458 2013 2 R. ecuadoriensis nymph Bellamaria SH sylvatic 0 rodent nest TBM330 animal shelter 459 2013 3 R. ecuadoriensis nymph Bellamaria SH sylvatic 2 rodent nest TBM330 animal shelter 460 2013 3 R. ecuadoriensis nymph Bellamaria SH sylvatic 4 rodent nest TBM330 461 2013 3 R. ecuadoriensis nymph Bellamaria SH sylvatic animal shelter rodent nest 5 TBM330 462 2013 3 R. ecuadoriensis nymph Bellamaria SH sylvatic animal shelter rodent nest 7 TBM330 animal shelter 463 2013 3 R. ecuadoriensis nymph Bellamaria SH sylvatic 8 rodent nest TBM330 animal shelter 464 2013 3 R. ecuadoriensis nymph Bellamaria SH sylvatic 9 rodent nest TBM331 animal shelter 465 2013 3 R. ecuadoriensis nymph Bellamaria SH sylvatic 0 rodent nest TBM331 animal shelter 466 2013 3 R. ecuadoriensis adult Bellamaria SH sylvatic 2 rodent nest 133

TBM331 animal shelter 467 2013 3 R. ecuadoriensis adult Bellamaria SH sylvatic 3 rodent nest TBM331 animal shelter 468 2013 3 R. ecuadoriensis adult Bellamaria SH sylvatic 4 rodent nest TBM331 469 2013 3 R. ecuadoriensis adult Bellamaria SH sylvatic animal shelter rodent nest 5 TBM331 animal shelter 470 2013 3 R. ecuadoriensis adult Bellamaria SH sylvatic 6 rodent nest TBM331 animal shelter 471 2013 3 R. ecuadoriensis adult Bellamaria SH sylvatic 7 rodent nest TBM331 animal shelter 472 2013 3 R. ecuadoriensis adult Bellamaria SH sylvatic 8 rodent nest TBM332 animal shelter 473 2013 3 R. ecuadoriensis adult Bellamaria SH sylvatic 0 rodent nest TBM332 474 2013 3 R. ecuadoriensis adult Bellamaria SH sylvatic animal shelter rodent nest 2 TBM332 animal shelter 475 2013 3 R. ecuadoriensis adult Bellamaria SH sylvatic 3 rodent nest TBM332 animal shelter 476 2013 2 R. ecuadoriensis nymph Bellamaria SH sylvatic 6 rodent nest TBM333 animal shelter 477 2013 3 R. ecuadoriensis nymph Bellamaria SH sylvatic 7 rodent nest TBM333 animal shelter 478 2013 3 R. ecuadoriensis nymph Bellamaria SH sylvatic 8 rodent nest TBM334 animal shelter 479 2013 3 R. ecuadoriensis nymph Bellamaria SH sylvatic 0 rodent nest TBM334 animal shelter 480 2013 3 R. ecuadoriensis nymph Bellamaria SH sylvatic 1 rodent nest TBM334 animal shelter 481 2013 3 R. ecuadoriensis nymph Bellamaria SH sylvatic 2 rodent nest 134

TBM334 482 2013 3 R. ecuadoriensis adult Bellamaria SH sylvatic animal shelter rodent nest 3 TBM334 animal shelter 483 2013 3 R. ecuadoriensis adult Bellamaria SH sylvatic 4 rodent nest TBM335 animal shelter 484 2013 4 R. ecuadoriensis adult Bellamaria SH sylvatic 7 rodent nest TBM336 485 2013 4 R. ecuadoriensis nymph Bellamaria SH sylvatic animal shelter rodent nest 1 TBM336 486 2013 4 R. ecuadoriensis nymph Bellamaria SH sylvatic animal shelter rodent nest 2 TBM336 487 2013 4 R. ecuadoriensis nymph Bellamaria SH sylvatic animal shelter rodent nest 3 TBM336 488 2013 4 R. ecuadoriensis nymph Bellamaria SH sylvatic animal shelter rodent nest 4 TBM336 489 2013 4 R. ecuadoriensis nymph Bellamaria SH sylvatic animal shelter rodent nest 5 TBM336 animal shelter 490 2013 5 R. ecuadoriensis adult Bellamaria SH sylvatic 8 rodent nest TBM337 491 2013 5 R. ecuadoriensis nymph Bellamaria SH sylvatic animal shelter rodent nest 1 TBM337 animal shelter 492 2013 5 R. ecuadoriensis nymph Bellamaria SH sylvatic 6 rodent nest TBM337 493 2013 5 R. ecuadoriensis nymph Bellamaria SH sylvatic animal shelter rodent nest 7 TBM338 494 2013 5 R. ecuadoriensis nymph Bellamaria SH sylvatic animal shelter rodent nest 0 TBM328 495 2013 2 R. ecuadoriensis nymph Bellamaria SH sylvatic animal shelter rodent nest 9 TBM340 animal shelter 496 2013 6 R. ecuadoriensis adult Bellamaria SH sylvatic 9 rodent nest 135

Tablada del organic 497 2013 12 TTO3424 R. ecuadoriensis adult CC sylvatic algodón animal shelter matter TCD343 animal shelter 498 2013 7 R. ecuadoriensis adult Chade CC sylvatic 7 rodent nest TCD345 499 2013 13 R. ecuadoriensis adult Chade CC sylvatic animal shelter rodent nest 2 TCD345 animal shelter 500 2013 13 R. ecuadoriensis adult Chade CC sylvatic 3 rodent nest TCD345 animal shelter 501 2013 13 R. ecuadoriensis nymph Chade CC sylvatic 4 rodent nest 502 2013 19 TDZ3455 R. ecuadoriensis nymph Danzarin CC sylvatic animal shelter rodent nest 503 2013 19 TDZ3456 R. ecuadoriensis nymph Danzarin CC sylvatic animal shelter rodent nest 504 2013 19 TDZ3457 R. ecuadoriensis nymph Danzarin CC sylvatic animal shelter rodent nest organic 505 2013 32 TBJ3475 R. ecuadoriensis nymph El Bejuco CC sylvatic animal shelter matter TCD349 animal shelter 506 2013 17 R. ecuadoriensis nymph Chade CC sylvatic 9 rodent nest TCD350 507 2013 17 R. ecuadoriensis nymph Chade CC sylvatic animal shelter rodent nest 0 *Days from collection to analysis

136

APPENDIX B: IDENTIFICATION OF TRIATOMINE SOURCE OF BLOOD IN SAMPLES FROM CENTRAL COASTAL AND

SOUTHERN HIGHLAND REGIONS IN ECUADOR.

Sample Infeccion >=95% Host Blast Order Cover Identity AC number Reference code T. cruzi identity 1 TEX318 no NA NA NA NA NA NA NA Rodentia 100% 100% si 2 TEX320 yes Rattus rattus KT232247.1 Igbokwe et al; 2016 3 TBR340 yes Homo sapiens Primates 100% 100% si KC252523.1 Loo et al; 2014 4 TBR341 yes Homo sapiens Primates 100% 100% si KC252523.1 Loo et al; 2014

5 TEX342 no Homo sapiens Primates 100% 100% si MF565490.1 Direct submission Galliforme 100% 100% si 6 TEX349 no Gallus gallus KX987152.1 Huang et al; 2017 Galliforme 100% 100% si 7 TGL356 no Gallus gallus KX987152.1 Huang et al; 2017 Galliforme 100% 100% si 8 TGL357 no Gallus gallus KX987152.1 Huang et al; 2017 Rodentia 100% 100% si 9 TTBJ370 yes Rattus rattus KT232247.1 Igbokwe et al; 2016 10 TTBJ371 yes Rattus rattus Rodentia 100% 100% si MH186588.1 Viesser et al; 2018

Rodentia 99% 100% si 11 TBJ378 no Aegialomys xanthaeolus EU074632.1 Direct submission

12 TBJ379 yes Rattus rattus Rodentia 100% 100% si MH186588.1 Viesser et al; 2018

Rodentia 99% 99% si 13 TBJ384 no Aegialomys xanthaeolus EU074632.1 Direct submission

14 TBJ401 yes Rattus rattus Rodentia 100% 100% si MH186588.1 Viesser et al; 2018

15 TBJ402 yes Rattus rattus Rodentia 100% 100% si KT232247.1 Igbokwe et al; 2016

16 TBJ403 yes Rattus rattus Rodentia 100% 100% si MH186588.1 Viesser et al; 2018 17 TBJ405 no Sus scrofa Artiodactyla 100% 100% si Direct submission MH880279.1

18 TCE674 no Homo sapiens Primates 100% 100% si MF565490.1 Direct submission

19 TCE675 no Homo sapiens Primates 100% 100% si MF521309.1 Direct submission

20 TCE681 no Homo sapiens Primates 100% 100% si KY410017.1 Olivieri et al; 2017

21 TCE696 no Homo sapiens Primates 100% 100% si MF521309.1 Direct submission

22 TCE699 yes Homo sapiens Primates 100% 99% si MF565490.1 Direct submission 137

23 TCE701 yes Homo sapiens Primates 100% 99% si KY411465.1 Larruga et al; 2017

24 TCE702 yes Homo sapiens Primates 100% 99% si MF565490.1 Direct submission

25 TCE703 yes Homo sapiens Primates 100% 99% si MF521309.1 Direct submission

26 TCE704 yes Homo sapiens Primates 100% 99% si MF565490.1 Direct submission

27 TCE706 no Homo sapiens Primates 100% 99% si MF565490.1 Direct submission 28 TCE708 no NA NA NA NA NA NA NA

29 TCE709 no Homo sapiens Primates 100% 100% si MF565490.1 Direct submission

30 TCE759 no Homo sapiens Primates 100% 100% si MF521309.1 Direct submission

31 TCE760 no Homo sapiens Primates 100% 100% si MF521309.1 Direct submission

32 TCE763 no Homo sapiens Primates 100% 100% si MF521309.1 Direct submission

33 TCE764 no Homo sapiens Primates 100% 100% si MF565490.1 Direct submission

34 TCE765 no Homo sapiens Primates 100% 100% si MF521309.1 Direct submission 35 TCQ800 yes Homo sapiens Primates 100% 100% si KY022422.1 Direct submission 36 TCQ801 no Homo sapiens Primates 100% 100% si KY101631.1 Direct submission 37 TCQ844 no NA NA NA NA NA NA NA 38 TCQ877 no Gallus gallus Galliforme 100% 100% si KX947010.1 Direct submission 39 TCQ940 no Homo sapiens Primates 100% 100% si KY101631.1 Direct submission 40 TCQ942 no Homo sapiens Primates 100% 100% si KY101631.1 Direct submission 41 TCQ952 no Homo sapiens Primates 100% 100% si KY101631.1 Direct submission 42 TGL1266 no NA NA NA NA NA NA NA Leptotila verreauxi Columbiforme 100% 99% si 43 TGL1296 yes Johnson et al; 2011

decolor HQ993503.1 Leptotila verreauxi Columbiforme 100% 99% si 44 TGL1299 yes Johnson et al; 2011

decolor HQ993503.1

45 TGL1300 yes Homo sapiens Primates 100% 98% si MF565490.1 Direct submission 46 TGL1301 no NA NA NA NA NA NA NA 47 TGL1309 yes NA NA NA NA NA NA NA 48 TBR1367 yes Homo sapiens Primates 100% 100% si KY022422.1 Direct submission 49 TBR1368 yes Homo sapiens Primates 100% 100% si KY022422.1 Direct submission 138

50 TBR1369 yes Gallus gallus Galliforme 100% 100% si KX947010.1 Direct submission 51 TBR1370 no Gallus gallus Galliforme 100% 100% si KX947010.1 Direct submission 52 TBR1371 no Gallus gallus Galliforme 100% 100% si KX947010.1 Direct submission 53 TBR1372 yes Gallus gallus Galliforme 100% 100% si KX947010.1 Direct submission 54 TBR1373 yes NA NA NA NA NA NA NA 55 TBR1375 yes NA NA NA NA NA NA NA 56 TBR1378 yes Homo sapiens Primates 100% 100% si KY022422.1 Direct submission 57 TBR1381 yes Simosciurus nebouxii Rodentia 100% 100% si U46183.1 Direct submission 58 TBR1382 no Simosciurus nebouxii Rodentia 100% 100% si u46183.1 Direct submission 59 TBR1383 yes Homo sapiens Primates 100% 100% si KY101631.1 Direct submission 60 TBR1384 yes Homo sapiens Primates 100% 100% si KY101631.1 Direct submission 61 TBR1386 yes Gallus gallus Galliforme 100% 100% si KX947010.1 Direct submission 62 TBR1388 yes Gallus gallus Galliforme 100% 100% si KX947010.1 Direct submission 63 TBR1389 no Gallus gallus Galliforme 100% 100% si KF964328.1 Direct submission 64 TBR1390 yes Gallus gallus Galliforme 100% 100% si KX947010.1 Direct submission 65 TBR1398 no Gallus gallus Galliforme 100% 100% si KX947010.1 Direct submission 66 TBR1399 no Gallus gallus Galliforme 100% 100% si KX947010.1 Direct submission 67 TBR1400 no Homo sapiens Primates 100% 100% si KY101631.1 Direct submission 68 TBR1404 yes Homo sapiens Primates 100% 100% si KY022422.1 Direct submission 69 TBR1415 no Mus musculus Rodentia 100% 99% si AB649478.1 Suzuki et al; 2013 70 TBR1416 no NA NA NA NA NA NA NA 71 TBR1417 no ND ND ND ND ND ND ND 72 TBR1420 no Homo sapiens Primates 100% 100% si KJ154879.1 Duggan et al; 2014 73 TBR1421 yes Gallus gallus Galliforme 100% 100% si KX947010.1 Direct submission 74 TBR1423 no Gallus gallus Galliforme 100% 100% si KX512321.1 Direct submission 75 TBR1433 no Gallus gallus Galliforme 100% 100% si KX947010.1 Direct submission 76 TBR1435 no Gallus gallus Galliforme 100% 100% si KX947010.1 Direct submission

77 TBR1445 yes Simosciurus nebouxii Rodentia 100% 100% si U46183.1 Direct submission 139

Rodentia 100% 100% si 78 TBR1447 yes Rattus rattus KT232247.1 Igbokwe et al; 2016 Rodentia 100% 100% si 79 TBR1448 yes Rattus rattus KT232247.1 Igbokwe et al; 2016 Rodentia 100% 100% si 80 TBR1449 yes Rattus rattus KT232247.1 Igbokwe et al; 2016

81 TBR1455 yes Simosciurus nebouxii Rodentia 100% 100% si U46183.1 Direct submission 82 TBR1457 yes Homo sapiens Primates 100% 100% si KY101631.1 Direct submission 83 TBR1459 yes Homo sapiens Primates 100% 100% si KY101631.1 Direct submission 84 TBR1460 no Homo sapiens Primates 100% 100% si KY022422.1 Direct submission 85 TBR1461 no Homo sapiens Primates 100% 100% si KY022422.1 Direct submission 86 TBR1462 yes Simosciurus nebouxii Rodentia 100% 100% si u46183.1 Direct submission 87 TBR1463 no Simosciurus nebouxii Rodentia 100% 100% si u46183.1 Direct submission 88 TBR1464 yes Simosciurus nebouxii Rodentia 100% 100% si u46183.1 Direct submission 89 TBR1465 yes Simosciurus nebouxii Rodentia 100% 100% si u46183.1 Direct submission 90 TBR1466 no Simosciurus nebouxii Rodentia 100% 100% si u46183.1 Direct submission 91 TBR1468 no Homo sapiens Primates 100% 100% si KY022422.1 Direct submission 92 TBR1469 no Homo sapiens Primates 100% 99% si KY022422.1 Direct submission 93 TBR1472 no Homo sapiens Primates 100% 100% si KY101631.1 Direct submission

94 TBR1474 yes Homo sapiens Primates 100% 100% si MF097961.1 Direct submission

95 TBR1475 yes Homo sapiens Primates 100% 100% si KY411465.1 Larruga et al; 2017 96 TBR1476 no Homo sapiens Primates 100% 100% si KY101631.1 Direct submission 97 TBR1477 no Gallus gallus Galliforme 100% 98% si KX947010.1 Direct submission 98 TBR1480 yes Simosciurus nebouxii Rodentia 100% 100% si u46183.1 Direct submission 99 TBR1481 yes Simosciurus nebouxii Rodentia 100% 100% si u46183.1 Direct submission

100 TBR1483 yes Homo sapiens Primates 100% 99% si KC911568.1 Direct submission 101 TBR1484 yes ND ND ND ND ND ND ND

102 TBR1491 yes Homo sapiens Primates 100% 98% si KP240730.1 Direct submission 103 TBR1494 yes Simosciurus nebouxii Rodentia 100% 100% si u46183.1 Direct submission 104 TBR1496 yes Simosciurus nebouxii Rodentia 100% 100% si u46183.1 Direct submission 105 TBR1497 no NA NA NA NA NA NA NA 140

106 TBR1506 no Simosciurus nebouxii Rodentia 100% 100% si u46183.1 Direct submission

107 TBJ1519 yes Homo sapiens Primates 100% 100% si KX675326.1 Marchi et al; 2017 108 TBJ1520 yes NA NA NA NA NA NA NA

109 TBJ1522 no Bufo marinus Anura 100% 99% si DQ415597.1 Mulcahy et al; 2006

110 TBJ1524 yes Homo sapiens Primates 100% 100% si KY595977.1 Direct submission

111 TBJ1530 yes Homo sapiens Primates 100% 96% si KX457634.1 Kutanan et al; 2016

112 TBJ1531 no Gallus gallus Galliforme 100% 100% si KX534431.1 Johnson et al; 2016 113 TBJ1532 no NA NA NA NA NA NA NA

114 TBJ1537 yes Gallus gallus Galliforme 100% 100% si KX534431.1 Johnson et al; 2016 115 TBJ1538 yes ND ND ND ND ND ND ND

116 TBJ1541 yes Boa constrictor imperator Squamata 100% 100% si KF576747.1 Direct submission

117 TBJ1542 yes Boa constrictor imperator Squamata 100% 100% si KF576747.1 Direct submission 118 TBJ1543 yes ND ND ND ND ND ND ND

119 TBJ1553 yes Rattus rattus Rodentia 100% 95% si KT232247.1 Igbokwe et al; 2016 120 TBJ1554 yes ND ND ND ND ND ND ND

121 TBJ1555 yes Gallus gallus Galliforme 100% 100% si KX534431.1 Johnson et al; 2016

122 TBJ1556 yes Homo sapiens Primates 100% 100% si KY595977.1 Direct submission 123 TBJ1557 yes ND ND ND ND ND ND ND

124 TBJ1562 no Gallus gallus Galliforme 100% 100% si KX534431.1 Johnson et al; 2016

125 TBJ1563 no Gallus gallus Galliforme 100% 100% si KX534431.1 Johnson et al; 2016

126 TBJ1565 yes Homo sapiens Primates 100% 100% si KY595977.1 Direct submission

127 TBJ1568 yes Bufo marinus Anura 100% 95% si DQ415597.1 Mulcahy et al; 2006

128 TBJ1572 no Homo sapiens Primates 100% 99% si KY595977.1 Direct submission 129 TBJ1585 yes NA NA NA NA NA NA NA

130 TBJ1594 yes Homo sapiens Primates 100% 100% si KX675326.1 Marchi et al; 2017 131 TBJ1595 yes NA NA NA NA NA NA NA

132 TBJ1598 no Homo sapiens Primates 100% 97% si KX675326.1 Marchi et al; 2017

133 TBJ1608 no Homo sapiens Primates 100% 100% si KM102144.1 Just et al; 2014 141

134 TBJ1677 no Canis lupus familiaris Carnivora 100% 100% si JX849653.1 Direct submission 135 TBJ1721 yes Homo sapiens Primates 100% 100% si KX675326.1 Marchi et al; 2017

136 TBJ1722 yes Homo sapiens Primates 100% 100% si KY595977.1 Direct submission

137 TBJ1723 yes Homo sapiens Primates 100% 100% si KY595977.1 Direct submission Galliforme 100% 100% si 138 TCD1747 no Gallus gallus KX987152.1 Huang et al; 2017 Galliforme 100% 100% si 139 TCD1748 no Gallus gallus KX987152.1 Huang et al; 2017

140 TCD1752 yes Homo sapiens Primates 100% 100% si MF588865.1 Duggan et al; 2017

141 TCD1756 yes Homo sapiens Primates 100% 99% si MF588865.1 Duggan et al; 2017

142 TCD1782 yes Homo sapiens Primates 100% 99% si MF588865.1 Duggan et al; 2017 Galliforme 100% 99% si 143 TCD1785 yes Gallus gallus KX987152.1 Huang et al; 2017 Galliforme 100% 100% si 144 TCD1786 yes Gallus gallus KX987152.1 Huang et al; 2017

145 TCD1801 yes Homo sapiens Primates 100% 100% si MF588867.1 Duggan et al; 2017

146 TCD1824 yes Gallus gallus Galliforme 100% 100% si KX987152.1 Huang et al; 2017 147 TCD1825 yes ND ND ND ND ND ND ND Campylorhynchus Passeriformes 100% 99% si 148 TCD1827 no

fasciatus DQ004883.1 Direct submission

149 TCD1828 yes Gallus gallus Galliforme 100% 100% si KX987152.1 Huang et al; 2017 Galliforme 100% 100% si 150 TCD1829 yes Gallus gallus KX987152.1 Huang et al; 2017 Galliforme 100% 100% si 151 TCD1841 no Gallus gallus KX987152.1 Huang et al; 2017 152 TCD1860 no Homo sapiens Primates 100% 100% si MF588865.1 Duggan et al; 2017 Galliforme 100% 95% si 153 TCD1861 no Gallus gallus KP211422.1 Direct submission

154 TCD1862 yes Homo sapiens Primates 100% 99% si KT897693.1 Direct submission Galliforme 100% 100% si 155 TCD1863 yes Gallus gallus KX987152.1 Huang et al; 2017

156 TCD1867 yes Gallus gallus Galliforme 100% 100% si KX987152.1 Huang et al; 2017 Galliforme 100% 100% si 157 TCD1897 yes Gallus gallus KX987152.1 Huang et al; 2017 Galliforme 100% 100% si 158 TCD1898 yes Gallus gallus KX987152.1 Huang et al; 2017 159 TCD1900 yes ND ND ND ND ND ND ND

160 TCD1901 yes Homo sapiens Primates 100% 99% si MF588865.1 Duggan et al; 2017 Galliforme 100% 100% si 161 TBJ1938 no Gallus gallus KX534431.1 Johnson et al; 2016 142

162 TBJ1942 no ND ND ND ND ND ND ND

163 TBJ1944 no Homo sapiens Primates 100% 98% si KY573997.1 Direct submission

164 TBJ1945 yes Homo sapiens Primates 100% 100% si KX675326.1 Marchi et al; 2017

165 TBJ1948 yes Homo sapiens Primates 100% 100% si KY595977.1 Direct submission

166 TBJ1949 yes Homo sapiens Primates 100% 99% si GU480002.1 Direct submission

167 TBJ1950 yes Gallus gallus Galliforme 100% 100% si KX534431.1 Johnson et al; 2016 168 TBJ1951 yes ND ND ND ND ND ND ND

169 TBJ1952 yes Homo sapiens Primates 100% 100% si KY595977.1 Direct submission

170 TBJ1953 no Gallus gallus Galliforme 100% 99% si KX534431.1 Johnson et al; 2016

171 TBJ1954 yes Gallus gallus Galliforme 100% 99% si KX534431.1 Johnson et al; 2016

172 TBJ1955 yes Homo sapiens Primates 100% 100% si KY595977.1 Direct submission 173 TBJ1956 yes NA NA NA NA NA NA NA Campylorhynchus Passeriformes 100% 99% si 174 TBJ1957 yes

fasciatus DQ004883.1 Direct submission Campylorhynchus Passeriformes 100% 99% si 175 TBJ1958 yes

fasciatus DQ004883.1 Direct submission Campylorhynchus Passeriformes 100% 99% si 176 TBJ1960 yes

fasciatus DQ004883.1 Direct submission Campylorhynchus Passeriformes 100% 99% si 177 TBJ1961 no

fasciatus DQ004883.1 Direct submission

178 TBJ1962 yes Homo sapiens Primates 100% 100% si KY595977.1 Direct submission 179 TBJ1966 yes NA NA NA NA NA NA NA

180 TBJ1967 yes Homo sapiens Primates 100% 100% si KY573997.1 Direct submission 181 TBJ1968 yes NA NA NA NA NA NA NA 182 TBJ1970 yes NA NA NA NA NA NA NA

183 TBJ1971 yes Homo sapiens Primates 100% 100% si KY595977.1 Direct submission

184 TBJ1972 yes Homo sapiens Primates 100% 96% si KF148520.1 Duggan et al; 2013

185 TTO1990 yes Gallus gallus Galliforme 100% 100% si KX987152.1 Huang et al; 2017 Galliforme 100% 100% si 186 TTO1992 yes Gallus gallus KX987152.1 Huang et al; 2017

187 TTO2002 no Homo sapiens Primates 100% 99% si MF588865.1 Duggan et al; 2017 143

188 TDZ2003 no Homo sapiens Primates 100% 100% si MF362755.1 Margaryan et al; 2017 189 TDZ2005 yes ND ND ND ND ND ND ND

190 TBJ2008 yes Homo sapiens Primates 100% 100% si KY509039.1 Direct submission

191 TBJ2009 yes Homo sapiens Primates 100% 100% si KX675326.1 Marchi et al; 2017

192 TBJ2010 yes Homo sapiens Primates 100% 100% si KY595977.1 Direct submission

193 TBJ2011 yes Homo sapiens Primates 100% 99% si KY595977.1 Direct submission

194 TDZ2016 yes Homo sapiens Primates 100% 100% si MF498725.1 Margaryan et al; 2017

195 TDZ2022 no Homo sapiens Primates 100% 100% si MF362755.1 Margaryan et al; 2017 Galliforme 100% 100% si 196 TDZ2026 yes Gallus gallus KX987152.1 Huang et al; 2017

197 TDZ2027 yes Homo sapiens Primates 100% 100% si MF588865.1 Duggan et al; 2017 Galliforme 100% 100% si 198 TTO2051 yes Gallus gallus KX987152.1 Huang et al; 2017

199 TDZ2057 yes Homo sapiens Primates 100% 100% si MF588865.1 Duggan et al; 2017

200 TDZ2068 yes Homo sapiens Primates 100% 100% si MF588865.1 Duggan et al; 2017 Galliforme 100% 100% si 201 TDZ2074 no Gallus gallus KX987152.1 Huang et al; 2017

202 TTO2086 yes Homo sapiens Primates 100% 100% si MF588865.1 Duggan et al; 2017 Galliforme 100% 100% si 203 TDZ2101 no Gallus gallus KX987152.1 Huang et al; 2017

204 TDZ2110 yes Homo sapiens Primates 100% 100% si MF588865.1 Duggan et al; 2017

205 TTO2114 no Homo sapiens Primates 100% 100% si MF588865.1 Duggan et al; 2017

206 TDZ2198 no Homo sapiens Primates 100% 99% si MF588865.1 Duggan et al; 2017 207 TDZ2199 no ND ND ND ND ND ND ND 208 TDZ2200 no NA NA NA NA NA NA NA

209 TDZ2201 no Homo sapiens Primates 100% 99% si MF588865.1 Duggan et al; 2017

210 TBJ2239 no Homo sapiens Primates 100% 100% si KY595977.1 Direct submission

211 TBJ2240 yes Homo sapiens Primates 100% 100% si KY595977.1 Direct submission

212 TBJ2242 yes Homo sapiens Primates 100% 100% si KY595977.1 Direct submission

213 TBJ2243 yes Gallus gallus Galliforme 100% 100% si KX534431.1 Johnson et al; 2016 Galliforme 100% 100% si 214 TBJ2244 yes Gallus gallus KX534431.1 Johnson et al; 2016

215 TBJ2246 yes Homo sapiens Primates 100% 99% si KY595977.1 Direct submission 144

216 TBJ2248 yes Homo sapiens Primates 100% 100% si KY595977.1 Direct submission

217 TBJ2249 no Homo sapiens Primates 100% 99% si KY595977.1 Direct submission

218 TBJ2250 yes Homo sapiens Primates 100% 100% si KY595977.1 Direct submission Galliforme 100% 98% si 219 TBJ2251 yes Gallus gallus KX534431.1 Johnson et al; 2016 220 TBJ2252 yes ND ND ND ND ND ND ND Galliforme 100% 96% si 221 TBJ2254 yes Gallus gallus AJ971340.1 Oshaghi et al; 2006

222 TBJ2257 yes Homo sapiens Primates 100% 100% si KY595977.1 Direct submission

223 TBJ2258 yes Homo sapiens Primates 100% 100% si KY595977.1 Direct submission

224 TBJ2259 yes Homo sapiens Primates 100% 100% si KY595977.1 Direct submission

225 TBJ2260 yes Homo sapiens Primates 100% 100% si KY595977.1 Direct submission

226 TBJ2266 yes Gallus gallus Galliforme 100% 100% si KX534431.1 Johnson et al; 2016 Galliforme 100% 100% si 227 TBJ2267 yes Gallus gallus KX534431.1 Johnson et al; 2016

228 TBJ2268 no Homo sapiens Primates 100% 98% si KY595977.1 Direct submission Campylorhynchus Passeriformes 100% 99% si 229 TBJ2270 yes

fasciatus DQ004883.1 Direct submission

230 TBJ2271 no Homo sapiens Primates 100% 99% si KP240730.1 Direct submission

231 TBJ2272 yes Homo sapiens Primates 100% 96% si KC993958.1 Delfin et al; 2013 Campylorhynchus Passeriformes 100% 99% si 232 TBJ2273 no

fasciatus DQ004883.1 Direct submission 233 TBJ2274 yes ND ND ND ND ND ND ND 234 TBJ2275 yes Homo sapiens Primates 100% 100% si KY595977.1 Direct submission

235 TBJ2276 yes Homo sapiens Primates 100% 100% si KY595977.1 Direct submission

236 TBJ2277 yes Homo sapiens Primates 100% 100% si KY595977.1 Direct submission

237 TBJ2278 no Homo sapiens Primates 100% 99% si KM277006.1 Direct submission Campylorhynchus Passeriformes 100% 100% si 238 TBJ2279 yes

fasciatus DQ004883.1 Direct submission

239 TBJ2280 yes Homo sapiens Primates 100% 100% si KY595977.1 Direct submission

240 TBJ2281 yes Homo sapiens Primates 100% 100% si KY595977.1 Direct submission Galliforme 100% 96% si 241 TBJ2282 yes Gallus gallus KX534431.1 Johnson et al; 2016 145

Galliforme 100% 100% si 242 TBJ2291 no Gallus gallus KX534431.1 Johnson et al; 2016 Galliforme 100% 100% si 243 TBJ2292 yes Gallus gallus KX534431.1 Johnson et al; 2016

244 TBJ2295 no Gallus gallus Galliforme 100% 100% si KX534431.1 Johnson et al; 2016 245 TBJ2308 yes NA NA NA NA NA NA NA Galliforme 100% 100% si 246 TBJ2310 no Gallus gallus KX534431.1 Johnson et al; 2016

247 TBJ2448 no Aegialomys xanthaeolus Rodentia 100% 100% si EU340015.1 Hanson et al; 2008 Leptotila verreauxi Columbiforme 100% 100% si 248 TBJ2452 no

decolor HQ993507.1 Johnson et al; 2011 249 TBJ2455 no Homo sapiens Primates 100% 100% si KY595977.1 Direct submission

250 TBJ2456 yes Simosciurus nebouxii Rodentia 99% 99% si U46183.1 Direct submission 251 TCQ2492 yes ND ND ND ND ND ND ND 252 TCQ2493 yes Gallus gallus Galliforme 100% 100% si KX947010.1 Direct submission 253 TCQ2494 yes Gallus gallus Galliforme 100% 100% si KX947010.1 Direct submission 254 TCQ2497 yes NA NA NA NA NA NA NA 255 TCQ2502 no Gallus gallus Galliforme 100% 100% si KX947010.1 Direct submission

256 TCE2532 no Gallus gallus Galliforme 100% 100% si KX987152.1 Huang et al; 2017

257 TCE2534 no Gallus gallus Galliforme 100% 98% si KX987152.1 Huang et al; 2017

258 TCE2538 no Gallus gallus Galliforme 100% 100% si KX987152.1 Huang et al; 2017

259 TCE2542 no Gallus gallus Galliforme 100% 100% si KX987152.1 Huang et al; 2017

260 TCE2543 no Gallus gallus Galliforme 100% 100% si KX987152.1 Huang et al; 2017 261 TCE2557 yes NA NA NA NA NA NA NA

262 TCE2564 yes Gallus gallus Galliforme 100% 100% si KX987152.1 Huang et al; 2017

263 TCE2565 yes Gallus gallus Galliforme 100% 100% si KX987152.1 Huang et al; 2017

264 TCE2566 yes Homo sapiens Primates 100% 100% si MF521309.1 Direct submission

265 TCE2567 yes Gallus gallus Galliforme 100% 100% si KX987152.1 Huang et al; 2017

266 TCE2568 yes Gallus gallus Galliforme 100% 100% si KX987152.1 Huang et al; 2017

267 TCE2569 yes Gallus gallus Galliforme 100% 100% si KX987152.1 Huang et al; 2017 268 TCE2570 no Gallus gallus Galliforme 100% 100% si KX987152.1 Huang et al; 2017

269 TCE2572 yes Gallus gallus Galliforme 100% 100% si KX987152.1 Huang et al; 2017 146

270 TCE2585 yes Gallus gallus Galliforme 100% 100% si KX987152.1 Huang et al; 2017 271 TCE2586 yes NA NA NA NA NA NA NA

272 TCE2587 yes Gallus gallus Galliforme 100% 100% si KX987152.1 Huang et al; 2017 273 TCE2588 yes NA NA NA NA NA NA NA

274 TCE2589 yes Homo sapiens Primates 100% 100% si MF565490.1 Direct submission Galliforme 100% 100% si 275 TCE2590 yes Gallus gallus KX987152.1 Huang et al; 2017 276 TCQ2595 yes Homo sapiens Primates 100% 99% si MN05390.4 Direct submission 277 TCQ2596 yes NA NA NA NA NA NA NA

278 TCE2598 yes Gallus gallus Galliforme 100% 99% si KX987152.1 Huang et al; 2017

279 TCE2602 yes Homo sapiens Primates 100% 100% si KJ856729.1 Derenko et al; 2014 280 TCE2603 yes NA NA NA NA NA NA NA Rodentia si 281 TBM2775 no Rattus rattus 100% 100% KT232247.1 Igbokwe et al; 2016 Rodentia si 282 TBM2776 yes Rattus rattus 100% 100% KT232247.1 Igbokwe et al; 2016

283 TBM2777 no Rattus rattus Rodentia 100% 100% si KT232247.1 Igbokwe et al; 2016 Rodentia si 284 TBM2778 yes Rattus rattus 100% 100% KT232247.1 Igbokwe et al; 2016 Rodentia si 285 TBM2779 yes Rattus rattus 100% 100% KT232247.1 Igbokwe et al; 2016

286 TBM2781 no Homo sapiens Primates 100% 100% si KY083725.1 Neparaczki et al; 2017 Artiodactyla si 287 TBM2782 no Capra hircus 100% 100% KY348327.1 Direct submission Artiodactyla si 288 TBM2784 no Capra hircus 100% 100% KY348327.1 Direct submission Rodentia si 289 TBM2785 no Rattus rattus 100% 100% KT232247.1 Igbokwe et al; 2016 Rodentia si 290 TBM2786 yes Rattus rattus 100% 98% KT232247.1 Igbokwe et al; 2016 Rodentia si 291 TBM2789 yes Rattus rattus 100% 100% KT232247.1 Igbokwe et al; 2016 Artiodactyla si 292 TBM2790 no Capra hircus 100% 100% KY348327.1 Direct submission Rodentia si 293 TBM2791 yes Rattus rattus 100% 100% KT232247.1 Igbokwe et al; 2016 Rodentia si 294 TBM2792 yes Rattus rattus 100% 100% KT232247.1 Igbokwe et al; 2016 Artiodactyla si 295 TBM2794 yes Capra hircus 100% 100% KY348327.1 Direct submission Rodentia si 296 TBM2795 yes Rattus rattus 100% 99% KT232247.1 Igbokwe et al; 2016 Carnívora si Direct submission 297 TBM2796 yes Canis lupus familiaris 100% 99% KX379529.1 147

298 TBM2797 yes Capra hircus Artiodactyla 100% 100% si KY348327.1 Direct submission Rodentia si 299 TBM2798 yes Rattus rattus 100% 100% KT232247.1 Igbokwe et al; 2016

300 TBM2800 yes Homo sapiens Primates 100% 100% si KY083725.1 Neparaczki et al; 2017 301 TBM2803 yes NA NA NA NA NA NA NA

302 TBM2804 no Homo sapiens Primates 100% 100% si KY083725.1 Neparaczki et al; 2017

303 TBM2805 no Homo sapiens Primates 100% 100% si KY083725.1 Neparaczki et al; 2017

304 TBM2806 yes Homo sapiens Primates 100% 100% si KY083725.1 Neparaczki et al; 2017 305 TBM2807 yes ND ND ND ND ND ND ND Artiodactyla si 306 TBM2808 yes Capra hircus 100% 100% KY348327.1 Direct submission Gomez-Carballa et al;

307 TBM2809 yes Homo sapiens Primates 100% 98% si HQ384200.1 2011

308 TBM2811 yes Capra hircus Artiodactyla 100% 100% si KY348327.1 Direct submission

309 TBM2812 yes Capra hircus Artiodactyla 100% 100% si KY348327.1 Direct submission

310 TBM2813 yes Homo sapiens Primates 100% 99% si KY083723.1 Neparaczki et al; 2017 311 TCQ2814 yes Rattus rattus Rodentia 100% 100% si KT232247.1 Igbokwe et al; 2016 312 TCQ2815 yes NA NA NA NA NA NA NA 313 TCQ2816 yes ND ND ND ND ND ND ND 314 TCQ2818 no Homo sapiens Primates 100% 100% si KY101631.1 Direct submission 315 TCQ2819 yes Homo sapiens Primates 100% 100% si KY101631.1 Direct submission

316 TBM2820 yes Canis lupus familiaris Carnívora 100% 95% si KX379529.1 Direct submission 317 TBM2821 yes ND ND ND ND ND ND ND

318 TBM2822 yes Rattus rattus Rodentia 100% 100% si KT232247.1 Igbokwe et al; 2016 319 TBM2823 yes ND ND ND ND ND ND ND 320 TBM2824 yes Homo sapiens Primates 100% 100% si KY863512.1 Direct submission 321 TCQ2840 yes Gallus gallus Galliforme 100% 100% si KX947010.1 Direct submission 322 TCQ2842 yes Gallus gallus Galliforme 100% 100% si KX947010.1 Direct submission 323 TCQ2843 yes Gallus gallus Galliforme 100% 100% si KX947010.1 Direct submission 324 TCQ2844 yes Gallus gallus Galliforme 100% 100% si KX947010.1 Direct submission

325 TBM2845 yes Homo sapiens Primates 100% 97% si KX023905.1 Direct submission 148

326 TBM2846 no Homo sapiens Primates 100% 100% si KC911568.1 Direct submission Galliforme si 327 TBM2848 no Gallus gallus 100% 100% KX534431.1 Johnson et al; 2016 328 TCQ2850 yes ND ND ND ND ND ND ND 329 TCQ2852 no Rattus rattus Rodentia 100% 100% si KT232247.1 Igbokwe et al; 2016 330 TCQ2854 yes Gallus gallus Galliforme 100% 100% si KX947010.1 Direct submission 331 TCQ2863 yes Gallus gallus Galliforme 98% 99% si KX947010.1 Direct submission 332 TCQ2866 yes ND ND ND ND ND ND ND 333 TCQ2867 yes ND ND ND ND ND ND ND 334 TCQ2868 yes ND ND ND ND ND ND ND 335 TCQ2869 yes ND ND ND ND ND ND ND

336 TBM2873 yes Homo sapiens Primates 100% 100% si MF002495.1 Direct submission 337 TCQ2875 yes NA NA NA NA NA NA NA 338 TCQ2877 yes Gallus gallus Galliforme 100% 100% si KX947010.1 Direct submission 339 TCQ2878 yes Gallus gallus Galliforme 100% 100% si KX947010.1 Direct submission 340 TCQ2880 yes Homo sapiens Primates 100% 100% si KY101631.1 Direct submission 341 TCQ2881 no Rattus rattus Rodentia 100% 100% si KT232247.1 Igbokwe et al; 2016 342 TCQ2882 no Rattus rattus Rodentia 100% 100% si KT232247.1 Igbokwe et al; 2016 343 TCQ2883 yes ND ND ND ND ND ND ND 344 TCQ2885 yes Rattus rattus Rodentia 100% 97% si KT232247.1 Igbokwe et al; 2016 345 TCQ2888 yes Rattus rattus Rodentia 100% 100% si KT232247.1 Igbokwe et al; 2016 346 TCQ2890 no Rattus rattus Rodentia 100% 100% si KT232247.1 Igbokwe et al; 2016

347 TBM2892 yes Homo sapiens Primates 100% 99% si KY083723.1 Neparaczki et al; 2017 Rodentia si 348 TBM2893 yes Rattus rattus 100% 100% KT232247.1 Igbokwe et al; 2016 Rodentia si 349 TBM2894 yes Rattus rattus 100% 99% KT232247.1 Igbokwe et al; 2016 350 TCQ2895 yes Homo sapiens Primates 99% 99% si KY022422.1 Direct submission 351 TCQ2896 yes Homo sapiens Primates 100% 99% si KY022422.1 Direct submission 352 TCQ2898 yes Homo sapiens Primates 100% 100% si KY101631.1 Direct submission 353 TCQ2902 yes Homo sapiens Primates 97% 100% si KY101631.1 Direct submission 149

Galliforme si 354 TBM2903 yes Gallus gallus 100% 100% KX987152.1 Huang et al; 2017 355 TCQ2906 no Gallus gallus Galliforme 100% 100% si KX947010.1 Direct submission 356 TCQ2907 yes Gallus gallus Galliforme 100% 100% si KX947010.1 Direct submission 357 TCQ2910 yes ND ND ND ND ND ND ND 358 TCQ2912 no Rattus rattus Rodentia 100% 100% si KT232247.1 Igbokwe et al; 2016 359 TCQ2922 no Rattus rattus Rodentia 100% 100% si KT232247.1 Igbokwe et al; 2016 360 TCQ2928 yes Gallus gallus Galliforme 98% 100% si KX947010.1 Direct submission 361 TCQ2929 yes Gallus gallus Galliforme 97% 100% si KX947010.1 Direct submission 362 TCQ2930 yes Gallus gallus Galliforme 100% 100% si fm205718.1 Direct submission 363 TCQ2934 yes Gallus gallus Galliforme 100% 99% si AJ971340.1 Oshaghi et al; 2006

364 TCQ2936 yes Rattus rattus Rodentia 100% 99% si KT232247.1 Igbokwe et al; 2016 365 TCQ2938 yes NA NA NA NA NA NA NA 366 TCQ2939 yes Gallus gallus Galliforme 100% 100% si KX947010.1 Direct submission 367 TCQ2940 yes Gallus gallus Galliforme 100% 100% si KX947010.1 Direct submission 368 TCQ2941 yes Gallus gallus Galliforme 100% 100% si KX947010.1 Direct submission 369 TCQ2942 yes NA NA NA NA NA NA NA 370 TCQ2943 yes Gallus gallus Galliforme 100% 100% si KX947010.1 Direct submission 371 TCQ2944 yes Gallus gallus Galliforme 100% 100% si KX947010.1 Direct submission 372 TCQ2946 yes Gallus gallus Galliforme 100% 100% si KX947010.1 Direct submission 373 TBM2948 yes NA NA NA NA NA NA NA Galliforme si 374 TBM2949 yes Gallus gallus 100% 100% KX534431.1 Johnson et al; 2016

375 TBM2950 yes Homo sapiens Primates 100% 99% si KJ557251.1 Direct submission 376 TBM2952 yes ND ND ND ND ND ND ND Galliforme si 377 TBM2954 yes Gallus gallus 100% 99% AJ971340.1 Oshaghi et al; 2006 Galliforme si 378 TBM2956 yes Gallus gallus 100% 100% KX534431.1 Johnson et al; 2016 Rodentia si 379 TBM2957 yes Rattus rattus 100% 99% KT232247.1 Igbokwe et al; 2016 Galliforme si 380 TBM2958 yes Gallus gallus 100% 100% KX534431.1 Johnson et al; 2016

381 TBM2960 yes Homo sapiens Primates 100% 97% si KY083723.1 Neparaczki et al; 2017 150

Galliforme si 382 TBM2961 yes Gallus gallus 100% 100% KX534431.1 Johnson et al; 2016 383 TBM2963 yes NA NA NA NA NA NA NA 384 TBM2966 yes ND ND ND ND ND ND ND 385 TBM2967 yes NA NA NA NA NA NA NA 386 TBM2977 yes NA NA NA NA NA NA NA

387 TBM2969 yes Homo sapiens Primates 100% 99% si KY863512.1 Direct submission

388 TBM2971 yes Homo sapiens Primates 100% 100% si KY863512.1 Direct submission 389 TBM2978 yes NA NA NA NA NA NA NA 390 TBM2979 yes Simosciurus nebouxii Rodentia 100% 100% Si U46183.1 Direct submission 391 TBM2980 yes Homo sapiens Primates 100% 100% Si KY083723.1 Neparaczki et al; 2017 Galliforme 100% 100% Si 392 TBM2983 yes Gallus gallus KX987152.1 Huang et al; 2017 393 TCQ2985 yes Gallus gallus Galliforme 100% 100% si KX947010.1 Direct submission 394 TCQ2986 yes Gallus gallus Galliforme 100% 100% si KX947010.1 Direct submission 395 TCQ2989 yes Gallus gallus Galliforme 100% 100% si KX947010.1 Direct submission 396 TCQ2990 yes Gallus gallus Galliforme 100% 100% si KX947010.1 Direct submission 397 TCQ2991 yes Gallus gallus Galliforme 100% 100% si KX947010.1 Direct submission 398 TCQ2992 yes ND ND ND ND ND ND ND

399 TNC3001 no Homo sapiens Primates 100% 100% si MF588867.1 Duggan et al; 2017 Rodentia 100% 100% si 400 TNC3002 yes Rattus norvegicus KY356148.1 Liu et al;2017 Galliforme 100% 100% si 401 TNC3031 yes Gallus gallus KX987152.1 Huang et al; 2017 Carnívora 100% 97% si 402 TNC3037 yes Canis lupus familiaris EU789744.1 Direct submission

403 TNC3039 yes Gallus gallus Galliforme 100% 95% si KX987152.1 Huang et al; 2017 Rodentia 100% 100% si 404 TNC3040 yes Rattus norvegicus KY356148.1 Liu et al;2017 Rodentia 100% 100% si 405 TNC3045 yes Rattus norvegicus KY356148.1 Liu et al;2017 406 TCQ3084 yes Gallus gallus Galliforme 100% 100% si KX947010.1 Direct submission 407 TCQ3085 yes NA NA NA NA NA NA NA 408 TCQ3086 no ND ND ND ND ND ND ND

409 TCQ3088 no Simosciurus nebouxii Rodentia 99% 97% si U46183.1 Direct submission 151

410 TCQ3089 yes Simosciurus nebouxii Rodentia 100% 100% si U46183.1 Direct submission

411 TCQ3091 yes Simosciurus nebouxii Rodentia 100% 99% si U46183.1 Direct submission

412 TCQ3092 no Homo sapiens Primates 100% 99% si KY101631.1 Direct submission 413 TCQ3093 yes NA NA NA NA NA NA NA 414 TBC3098 yes ND ND ND ND ND ND ND 415 TBM3101 yes Homo sapiens Primates 100% 99% Si KY083723.1 Neparaczki et al; 2017 416 TBM3102 yes Homo sapiens Primates 100% 100% Si KY083725.1 Neparaczki et al; 2017

417 TCQ3103 no Simosciurus nebouxii Rodentia 100% 99% si U46183.1 Direct submission

418 TCQ3104 no Homo sapiens Primates 100% 100% si KY101631.1 Direct submission 419 TCQ3107 yes Homo sapiens Primates 100% 100% si KY022422.1 Direct submission

420 TCQ3111 yes Homo sapiens Primates 100% 98% si LC088149.1 Bravo-Barriga et al 2016

421 TCQ3113 yes Homo sapiens Primates 100% 99% si KY101631.1 Direct submission

422 TCQ3114 yes Homo sapiens Primates 100% 96% si KM047233.1 Direct submission

423 TCQ3115 no Simosciurus nebouxii Rodentia 100% 95% si U46183.1 Direct submission 424 TCQ3116 yes NA NA NA NA NA NA NA 425 TBM3131 yes Rattus rattus Rodentia 100% 96% Si KT232247.1 Igbokwe et al; 2016 426 TBM3133 yes NA NA NA NA NA NA NA 427 TBM3134 no Homo sapiens Primates 100% 99% Si KX023905.1 Direct submission 428 TCQ3139 yes Homo sapiens Primates 100% 100% si KY101631.1 Direct submission 429 TBM3143 yes Homo sapiens Primates 100% 98% Si KX023905.1 Direct submission 430 TBM3144 yes Homo sapiens Primates 100% 100% Si KY083725.1 Neparaczki et al; 2017 431 TBM3148 no Homo sapiens Primates 100% 99% Si KC911568.1 Direct submission 432 TBM3151 yes Homo sapiens Primates 100% 100% Si KY083723.1 Neparaczki et al; 2017 433 TBM3185 yes Homo sapiens Primates 100% 100% Si KY083723.1 Neparaczki et al; 2017

434 TBM3186 yes Homo sapiens Primates 100% 96% Si LC088151.1 Bravo-Barriga et al 2016

435 TBM3190 no Rattus rattus Rodentia 100% 100% si KT232247.1 Igbokwe et al; 2016 152

436 TCQ3197 yes Gallus gallus Galliforme 99% 99% si KX947010.1 Direct submission 437 TCQ3215 yes Gallus gallus Galliforme 100% 100% si KX947010.1 Direct submission

438 TCQ3218 yes Simosciurus nebouxii Rodentia 97% 99% si U46183.1 Direct submission 439 TCQ3219 yes ND ND ND ND ND ND ND 440 TCQ3222 yes Homo sapiens Primates 100% 100% si KY101631.1 Direct submission

441 TCQ3223 yes Homo sapiens Primates 100% 100% si KY101631.1 Direct submission

442 TCQ3224 yes Homo sapiens Primates 97% 99% si KX697544.1 Auty et al; 2016

443 TCQ3225 yes Homo sapiens Primates 100% 100% si KY101631.1 Direct submission 444 TBM3234 yes Homo sapiens Primates 100% 99% si KY849396.1 Direct submission 445 TBM3235 no Homo sapiens Primates 100% 100% si KY849395.1 Direct submission 446 TBM3236 yes Rattus rattus Rodentia 100% 97% si KT232247.1 Igbokwe et al; 2016 447 TBM3239 no Homo sapiens Primates 100% 99% si KY849396.1 Direct submission 448 TBM3241 no Homo sapiens Primates 100% 100% si KY849395.1 Direct submission 449 TBM3242 no ND ND ND ND ND ND ND 450 TBM3283 no NA NA NA NA NA NA NA 451 TBM3290 yes Simosciurus nebouxii Rodentia 100% 100% si U46183.1 Direct submission 452 TBM3291 yes Simosciurus nebouxii Rodentia 100% 100% si U46183.1 Direct submission 453 TBM3292 yes Simosciurus nebouxii Rodentia 100% 100% si U46183.1 Direct submission 454 TBM3294 yes Simosciurus nebouxii Rodentia 100% 100% si U46183.1 Direct submission 455 TBM3295 no Simosciurus nebouxii Rodentia 100% 100% si U46183.1 Direct submission 456 TBM3296 no Homo sapiens Primates 100% 98% si KP240730.1 Direct submission 457 TBM3297 yes Simosciurus nebouxii Rodentia 100% 100% si U46183.1 Direct submission Rodentia 100% 99% si 458 TBM3300 yes Simosciurus nebouxii U46183.1 Direct submission Rodentia 100% 100% si 459 TBM3302 no Simosciurus nebouxii U46183.1 Direct submission Rodentia 100% 97% si 460 TBM3304 no Simosciurus nebouxii U46183.1 Direct submission 461 TBM3305 no ND ND ND ND ND ND ND 462 TBM3307 no NA NA NA NA NA NA NA Rodentia 100% 99% si 463 TBM3308 no Simosciurus nebouxii U46183.1 Direct submission 153

Rodentia 100% 100% si 464 TBM3309 no Simosciurus nebouxii U46183.1 Direct submission

465 TBM3310 no Simosciurus nebouxii Rodentia 100% 100% si U46183.1 Direct submission Rodentia 100% 99% si 466 TBM3312 no Simosciurus nebouxii U46183.1 Direct submission Rodentia 100% 99% si 467 TBM3313 no Simosciurus nebouxii U46183.1 Direct submission Rodentia 100% 99% si 468 TBM3314 no Simosciurus nebouxii U46183.1 Direct submission 469 TBM3315 yes NA NA NA NA NA NA NA Rodentia 100% 99% si 470 TBM3316 no Simosciurus nebouxii U46183.1 Direct submission Rodentia 100% 99% si 471 TBM3317 no Simosciurus nebouxii U46183.1 Direct submission Rodentia 100% 99% si 472 TBM3318 no Simosciurus nebouxii U46183.1 Direct submission

473 TBM3320 no Homo sapiens Primates 100% 100% si KX023905.1 Direct submission 474 TBM3322 yes ND ND ND ND ND ND ND Rodentia 100% 98% si 475 TBM3323 no Simosciurus nebouxii U46183.1 Direct submission

476 TBM3326 no Homo sapiens Primates 99% 98% si KC911568.1 Direct submission Rodentia 100% 99% si 477 TBM3337 no Simosciurus nebouxii U46183.1 Direct submission Rodentia 100% 100% si 478 TBM3338 no Simosciurus nebouxii U46183.1 Direct submission

479 TBM3340 no Simosciurus nebouxii Rodentia 99% 98% si U46183.1 Direct submission Rodentia 100% 100% si 480 TBM3341 yes Simosciurus nebouxii U46183.1 Direct submission Rodentia 100% 100% si 481 TBM3342 no Simosciurus nebouxii U46183.1 Direct submission 482 TBM3343 yes NA NA NA NA NA NA NA Rodentia 100% 100% si 483 TBM3344 yes Simosciurus nebouxii U46183.1 Direct submission Rodentia 100% 99% si 484 TBM3357 yes Simosciurus nebouxii U46183.1 Direct submission 485 TBM3361 yes NA NA NA NA NA NA NA 486 TBM3362 yes NA NA NA NA NA NA NA 487 TBM3363 yes NA NA NA NA NA NA NA 488 TBM3364 no ND ND ND ND ND ND ND 489 TBM3365 no NA NA NA NA NA NA NA

490 TBM3368 yes Homo sapiens Primates 100% 100% si LC198226.1 Mizuno et al; 2017 491 TBM3371 yes NA NA NA NA NA NA NA 154

Rodentia 100% 98% si 492 TBM3376 yes Simosciurus nebouxii U46183.1 Direct submission 493 TBM3377 yes NA NA NA NA NA NA NA 494 TBM3380 no NA NA NA NA NA NA NA 495 TBM3289 yes ND ND ND ND ND ND ND

496 TBM3409 yes Homo sapiens Primates 100% 100% si LC198226.1 Mizuno et al; 2017

497 TTO3424 yes Didelphis marsupialis Didelphimorphia 100% 99% si KF799996.1 Navia-Gine et al; 2013 Rodentia 99% 100% si 498 TCD3437 no Aegialomys xanthaeolus EU340015.1 Hanson et al; 2008 499 TCD3452 yes NA NA NA NA NA NA NA Carnívora 99% 100% si 500 TCD3453 yes Canis lupus familiaris JX849653.1 Direct submission 501 TCD3454 no ND ND ND ND ND ND ND Columbiforme 100% 99% si 502 TDZ3455 yes Columba livia KY378739.1 Valente et al; 2017 Columbiforme 100% 99% si 503 TDZ3456 yes Columba livia KY378739.1 Valente et al; 2017 Columbiforme 100% 99% si 504 TDZ3457 yes Columba livia KY378739.1 Valente et al; 2017 Columbiforme 100% 100% si 505 TBJ3475 yes Columba livia KP319029.1 Direct submission Rodentia 100% 100% si 506 TCD3499 yes Rattus rattus KT232247.1 Igbokwe et al; 2016 507 TCD3500 no NA NA NA NA NA NA NA

NA: samples that did not amplified a PCR product for analysis ND: samples where blood meal was not identified due to the identity percentage was ˂95% or presented an illegible chromatogram

155

APPENDIX C: HOST SPECIES, COLLECTION DATA AND SOURCES OF BLOOD OF THE 416 TRIATOMINE INTESTINAL

CONTENT ANALYZED BY SEQUENCE OF CYTB FRAGMENT.

Collection Parasite Code Host species Stage Locality Region Blood souce environment infection 1 EX320 P. chinai nymph La Extensa Southern highlands Intradomiciliary non-human T. cruzi 2 TBR340 R. ecuadoriensis adult Bramaderos Southern highlands Intradomiciliary Human T. cruzi 3 TBR341 R. ecuadoriensis adult Bramaderos Southern highlands Intradomiciliary Human T. cruzi 4 TEX342 P. chinai adult La Extensa Southern highlands Intradomiciliary Human T. rangeli 5 TEX349 R. ecuadoriensis adult La Extensa Southern highlands Intradomiciliary non-human no infection 6 TGL356 R. ecuadoriensis nymph Galapagos Southern highlands Intradomiciliary non-human no infection 7 TGL357 R. ecuadoriensis nymph Galapagos Southern highlands Intradomiciliary non-human no infection 8 TTBJ370 P. howardi nymph El Bejuco Central coastal Peridomiciliary non-human T. cruzi 9 TTBJ371 P. howardi nymph El Bejuco Central coastal Peridomiciliary non-human T. cruzi 10 TBJ378 R. ecuadoriensis nymph El Bejuco Central coastal Peridomiciliary non-human T. rangeli 11 TBJ379 R. ecuadoriensis nymph El Bejuco Central coastal Peridomiciliary non-human Mixed 12 TBJ384 R. ecuadoriensis adult El Bejuco Central coastal Sylvatic non-human T. rangeli 13 TBJ401 R. ecuadoriensis nymph El Bejuco Central coastal Peridomiciliary non-human T. cruzi 14 TBJ402 P. howardi nymph El Bejuco Central coastal Peridomiciliary non-human T. cruzi 15 TBJ403 R. ecuadoriensis adult El Bejuco Central coastal Peridomiciliary non-human T. cruzi 16 TBJ405 P. howardi nymph El Bejuco Central coastal Peridomiciliary non-human T. rangeli 17 TCE674 P. chinai adult Coamine Southern highlands Intradomiciliary Human no infection 18 TCE675 T. carrioni nymph Coamine Southern highlands Intradomiciliary Human no infection 19 TCE681 P. chinai nymph Coamine Southern highlands Intradomiciliary Human no infection 20 TCE696 P. chinai adult Coamine Southern highlands Intradomiciliary Human no infection 21 TCE699 R. ecuadoriensis adult Coamine Southern highlands Sylvatic Human T. cruzi 22 TCE701 R. ecuadoriensis adult Coamine Southern highlands Sylvatic Human T. cruzi 23 TCE702 R. ecuadoriensis adult Coamine Southern highlands Sylvatic Human T. cruzi 156

24 TCE703 R. ecuadoriensis adult Coamine Southern highlands Sylvatic Human T. cruzi 25 TCE704 R. ecuadoriensis nymph Coamine Southern highlands Sylvatic Human T. cruzi 26 TCE706 P. chinai adult Coamine Southern highlands Intradomiciliary Human no infection 27 TCE709 P. chinai nymph Coamine Southern highlands Intradomiciliary Human no infection 28 TCE759 R. ecuadoriensis adult Coamine Southern highlands Intradomiciliary Human no infection 29 TCE760 P. chinai nymph Coamine Southern highlands Intradomiciliary Human T. rangeli 30 TCE763 P. chinai nymph Coamine Southern highlands Intradomiciliary Human no infection 31 TCE764 P. chinai nymph Coamine Southern highlands Intradomiciliary Human no infection 32 TCE765 P. chinai nymph Coamine Southern highlands Intradomiciliary Human no infection 33 TCQ800 R. ecuadoriensis adult Chaquizhca Southern highlands Intradomiciliary Human T. cruzi 34 TCQ801 R. ecuadoriensis adult Chaquizhca Southern highlands Intradomiciliary Human no infection 35 CQ877 P. chinai nymph Chaquizhca Southern highlands Peridomiciliary non-human no infection 36 TCQ940 R. ecuadoriensis adult Chaquizhca Southern highlands Intradomiciliary Human no infection 37 TCQ942 R. ecuadoriensis adult Chaquizhca Southern highlands Intradomiciliary Human no infection 38 TCQ952 P. chinai adult Chaquizhca Southern highlands Intradomiciliary Human no infection 39 TGL1296 R. ecuadoriensis nymph Galapagos Southern highlands Sylvatic non-human T. cruzi 40 TGL1299 R. ecuadoriensis adult Galapagos Southern highlands Sylvatic non-human T. cruzi 41 TGL1300 R. ecuadoriensis adult Galapagos Southern highlands Sylvatic Human T. cruzi 42 TBR1367 P. chinai adult Bramaderos Southern highlands Intradomiciliary Human T. cruzi 43 TBR1368 P. rufotuberculatus adult Bramaderos Southern highlands Intradomiciliary Human Mixed 44 TBR1369 R. ecuadoriensis adult Bramaderos Southern highlands Peridomiciliary non-human T. cruzi 45 TBR1370 R. ecuadoriensis nymph Bramaderos Southern highlands Peridomiciliary non-human no infection 46 TBR1371 R. ecuadoriensis nymph Bramaderos Southern highlands Peridomiciliary non-human no infection 47 TBR1372 R. ecuadoriensis nymph Bramaderos Southern highlands Peridomiciliary non-human T. cruzi 48 TBR1378 R. ecuadoriensis adult Bramaderos Southern highlands Intradomiciliary Human T. cruzi 49 TBR1381 R. ecuadoriensis nymph Bramaderos Southern highlands Sylvatic non-human T. cruzi 50 TBR1382 R. ecuadoriensis adult Bramaderos Southern highlands Sylvatic non-human no infection 51 TBR1383 R. ecuadoriensis adult Bramaderos Southern highlands Intradomiciliary Human T. cruzi 157

52 TBR1384 R. ecuadoriensis nymph Bramaderos Southern highlands Intradomiciliary Human T. cruzi 53 TBR1386 R. ecuadoriensis adult Bramaderos Southern highlands Peridomiciliary non-human T. cruzi 54 TBR1388 R. ecuadoriensis adult Bramaderos Southern highlands Peridomiciliary non-human T. cruzi 55 TBR1389 R. ecuadoriensis adult Bramaderos Southern highlands Peridomiciliary non-human no infection 56 TBR1390 R. ecuadoriensis adult Bramaderos Southern highlands Peridomiciliary non-human T. cruzi 57 TBR1398 R. ecuadoriensis adult Bramaderos Southern highlands Intradomiciliary non-human T. rangeli 58 TBR1399 R. ecuadoriensis adult Bramaderos Southern highlands Intradomiciliary non-human no infection 59 TBR1400 R. ecuadoriensis adult Bramaderos Southern highlands Intradomiciliary Human no infection 60 TBR1404 R. ecuadoriensis adult Bramaderos Southern highlands Sylvatic Human T. cruzi 61 TBR1415 R. ecuadoriensis nymph Bramaderos Southern highlands Sylvatic non-human no infection 62 TBR1420 R. ecuadoriensis adult Bramaderos Southern highlands Sylvatic Human T. rangeli 63 TBR1421 R. ecuadoriensis nymph Bramaderos Southern highlands Peridomiciliary non-human T. cruzi 64 TBR1423 R. ecuadoriensis nymph Bramaderos Southern highlands Peridomiciliary non-human no infection 65 TBR1433 R. ecuadoriensis adult Bramaderos Southern highlands Peridomiciliary non-human no infection 66 TBR1435 R. ecuadoriensis adult Bramaderos Southern highlands Peridomiciliary non-human no infection 67 TBR1445 R. ecuadoriensis nymph Bramaderos Southern highlands Sylvatic non-human T. cruzi 68 TBR1447 R. ecuadoriensis nymph Bramaderos Southern highlands Sylvatic non-human Mixed 69 TBR1448 R. ecuadoriensis adult Bramaderos Southern highlands Sylvatic non-human T. cruzi 70 TBR1449 R. ecuadoriensis adult Bramaderos Southern highlands Sylvatic non-human T. cruzi 71 TBR1455 R. ecuadoriensis nymph Bramaderos Southern highlands Sylvatic non-human T. cruzi 72 TBR1457 R. ecuadoriensis nymph Bramaderos Southern highlands Sylvatic Human T. cruzi 73 TBR1459 R. ecuadoriensis nymph Bramaderos Southern highlands Sylvatic Human T. cruzi 74 TBR1460 R. ecuadoriensis nymph Bramaderos Southern highlands Sylvatic Human no infection 75 TBR1461 R. ecuadoriensis nymph Bramaderos Southern highlands Sylvatic Human T. rangeli 76 TBR1462 R. ecuadoriensis nymph Bramaderos Southern highlands Sylvatic non-human Mixed 77 TBR1463 R. ecuadoriensis nymph Bramaderos Southern highlands Sylvatic non-human T. rangeli 78 TBR1464 R. ecuadoriensis adult Bramaderos Southern highlands Sylvatic non-human Mixed 79 TBR1465 R. ecuadoriensis adult Bramaderos Southern highlands Sylvatic non-human Mixed 158

80 TBR1466 R. ecuadoriensis adult Bramaderos Southern highlands Sylvatic non-human T. rangeli 81 TBR1468 R. ecuadoriensis nymph Bramaderos Southern highlands Sylvatic Human no infection 82 TBR1469 R. ecuadoriensis adult Bramaderos Southern highlands Sylvatic Human T. rangeli 83 TBR1472 R. ecuadoriensis nymph Bramaderos Southern highlands Sylvatic Human T. rangeli 84 TBR1474 R. ecuadoriensis nymph Bramaderos Southern highlands Sylvatic Human T. cruzi 85 TBR1475 R. ecuadoriensis adult Bramaderos Southern highlands Sylvatic Human T. cruzi 86 TBR1476 R. ecuadoriensis nymph Bramaderos Southern highlands Sylvatic Human no infection 87 TBR1477 R. ecuadoriensis nymph Bramaderos Southern highlands Sylvatic non-human T. rangeli 88 TBR1480 R. ecuadoriensis nymph Bramaderos Southern highlands Sylvatic non-human T. cruzi 89 TBR1481 R. ecuadoriensis adult Bramaderos Southern highlands Sylvatic non-human Mixed 90 TBR1483 R. ecuadoriensis adult Bramaderos Southern highlands Sylvatic Human T. cruzi 91 TBR1491 R. ecuadoriensis nymph Bramaderos Southern highlands Sylvatic Human T. cruzi 92 TBR1494 R. ecuadoriensis adult Bramaderos Southern highlands Sylvatic non-human T. cruzi 93 TBR1496 R. ecuadoriensis adult Bramaderos Southern highlands Sylvatic non-human Mixed 94 TBR1506 R. ecuadoriensis nymph Bramaderos Southern highlands Sylvatic non-human no infection 95 TBJ1519 P. howardi nymph El Bejuco Central coastal Peridomiciliary Human T. cruzi 96 TBJ1522 P. howardi nymph El Bejuco Central coastal Peridomiciliary non-human no infection 97 TBJ1524 R. ecuadoriensis adult El Bejuco Central coastal Peridomiciliary Human T. cruzi 98 TBJ1530 P. howardi adult El Bejuco Central coastal Intradomiciliary Human T. cruzi 99 TBJ1531 R. ecuadoriensis adult El Bejuco Central coastal Peridomiciliary non-human no infection 100 TBJ1537 R. ecuadoriensis nymph El Bejuco Central coastal Peridomiciliary non-human T. cruzi 101 TBJ1541 P. howardi nymph El Bejuco Central coastal Peridomiciliary non-human T. cruzi 102 TBJ1542 P. howardi nymph El Bejuco Central coastal Peridomiciliary non-human T. cruzi 103 TBJ1553 P. howardi adult El Bejuco Central coastal Peridomiciliary non-human T. cruzi 104 TBJ1555 R. ecuadoriensis nymph El Bejuco Central coastal Peridomiciliary non-human T. cruzi 105 TBJ1556 R. ecuadoriensis adult El Bejuco Central coastal Peridomiciliary Human T. cruzi 106 TBJ1562 R. ecuadoriensis adult El Bejuco Central coastal Peridomiciliary non-human no infection 107 TBJ1563 R. ecuadoriensis nymph El Bejuco Central coastal Peridomiciliary non-human no infection 159

108 TBJ1565 P. howardi adult El Bejuco Central coastal Intradomiciliary Human T. cruzi 109 TBJ1568 P. howardi nymph El Bejuco Central coastal Peridomiciliary non-human T. cruzi 110 TBJ1572 R. ecuadoriensis nymph El Bejuco Central coastal Sylvatic Human no infection 111 TBJ1594 R. ecuadoriensis nymph El Bejuco Central coastal Peridomiciliary Human T. cruzi 112 TBJ1598 R. ecuadoriensis nymph El Bejuco Central coastal Sylvatic Human no infection 113 TBJ1608 P. howardi adult El Bejuco Central coastal Peridomiciliary Human no infection 114 TBJ1677 R. ecuadoriensis nymph El Bejuco Central coastal Peridomiciliary non-human no infection 115 TBJ1721 R. ecuadoriensis adult El Bejuco Central coastal Sylvatic Human T. cruzi 116 TBJ1722 R. ecuadoriensis adult El Bejuco Central coastal Sylvatic Human T. cruzi 117 TBJ1723 R. ecuadoriensis adult El Bejuco Central coastal Sylvatic Human T. cruzi 118 TCD1747 R. ecuadoriensis nymph Chade Central coastal sylvatic non-human no infection 119 TCD1748 R. ecuadoriensis nymph Chade Central coastal sylvatic non-human no infection 120 TCD1752 R. ecuadoriensis nymph Chade Central coastal sylvatic Human T. cruzi 121 TCD1756 R. ecuadoriensis nymph Chade Central coastal sylvatic Human T. cruzi 122 TCD1782 R. ecuadoriensis nymph Chade Central coastal sylvatic Human T. cruzi 123 TCD1785 R. ecuadoriensis adult Chade Central coastal sylvatic non-human T. cruzi 124 TCD1786 R. ecuadoriensis adult Chade Central coastal sylvatic non-human T. cruzi 125 TCD1801 R. ecuadoriensis adult Chade Central coastal sylvatic Human T. cruzi 126 TCD1824 R. ecuadoriensis nymph Chade Central coastal peridomiciliary non-human T. cruzi 127 TCD1827 R. ecuadoriensis adult Chade Central coastal peridomiciliary non-human no infection 128 TCD1828 R. ecuadoriensis nymph Chade Central coastal peridomiciliary non-human T. cruzi 129 TCD1829 R. ecuadoriensis nymph Chade Central coastal peridomiciliary non-human T. cruzi 130 TCD1841 R. ecuadoriensis nymph Chade Central coastal sylvatic non-human no infection 131 TCD1860 R. ecuadoriensis nymph Chade Central coastal peridomiciliary Human no infection 132 TCD1861 R. ecuadoriensis nymph Chade Central coastal peridomiciliary non-human no infection 133 TCD1862 R. ecuadoriensis nymph Chade Central coastal sylvatic Human T. cruzi 134 TCD1863 R. ecuadoriensis nymph Chade Central coastal sylvatic non-human T. cruzi 135 TCD1867 R. ecuadoriensis nymph Chade Central coastal sylvatic non-human T. cruzi 160

136 TCD1897 R. ecuadoriensis nymph Chade Central coastal sylvatic non-human T. cruzi 137 TCD1898 R. ecuadoriensis nymph Chade Central coastal sylvatic non-human T. cruzi 138 TCD1901 R. ecuadoriensis nymph Chade Central coastal sylvatic Human T. cruzi 139 TBJ1938 R. ecuadoriensis adult El Bejuco Central coastal peridomiciliary non-human no infection 140 TBJ1944 R. ecuadoriensis adult El Bejuco Central coastal sylvatic Human T. rangeli 141 TBJ1945 R. ecuadoriensis nymph El Bejuco Central coastal sylvatic Human T. cruzi 142 TBJ1948 R. ecuadoriensis adult El Bejuco Central coastal peridomiciliary Human T. cruzi 143 TBJ1949 R. ecuadoriensis adult El Bejuco Central coastal peridomiciliary Human Mixed 144 TBJ1950 R. ecuadoriensis adult El Bejuco Central coastal peridomiciliary non-human Mixed 145 TBJ1952 R. ecuadoriensis adult El Bejuco Central coastal peridomiciliary Human T. cruzi 146 TBJ1953 R. ecuadoriensis adult El Bejuco Central coastal peridomiciliary non-human no infection 147 TBJ1954 R. ecuadoriensis adult El Bejuco Central coastal peridomiciliary non-human T. cruzi 148 TBJ1955 R. ecuadoriensis adult El Bejuco Central coastal sylvatic Human T. cruzi 149 TBJ1957 R. ecuadoriensis adult El Bejuco Central coastal sylvatic non-human T. cruzi 150 TBJ1958 R. ecuadoriensis adult El Bejuco Central coastal sylvatic non-human T. cruzi 151 TBJ1960 R. ecuadoriensis nymph El Bejuco Central coastal sylvatic non-human T. cruzi 152 TBJ1961 R. ecuadoriensis nymph El Bejuco Central coastal sylvatic non-human no infection 153 TBJ1962 R. ecuadoriensis nymph El Bejuco Central coastal sylvatic Human T. cruzi 154 TBJ1967 R. ecuadoriensis adult El Bejuco Central coastal sylvatic Human T. cruzi 155 TBJ1971 R. ecuadoriensis adult El Bejuco Central coastal sylvatic Human T. cruzi 156 TBJ1972 R. ecuadoriensis nymph El Bejuco Central coastal sylvatic Human T. cruzi Tablada del 157 TTO1990 R. ecuadoriensis adult Central coastal intradomiciliary non-human T. cruzi algodón Tablada del 158 TTO1992 R. ecuadoriensis adult Central coastal intradomiciliary non-human T. cruzi algodón Tablada del 159 TTO2002 R. ecuadoriensis adult Central coastal peridomiciliary Human no infection algodón 160 TDZ2003 R. ecuadoriensis adult Danzarin Central coastal sylvatic Human no infection 161 TBJ2008 R. ecuadoriensis adult El Bejuco Central coastal peridomiciliary Human T. cruzi 161

162 TBJ2009 R. ecuadoriensis adult El Bejuco Central coastal peridomiciliary Human T. cruzi 163 TBJ2010 R. ecuadoriensis adult El Bejuco Central coastal peridomiciliary Human T. cruzi 164 TBJ2011 R. ecuadoriensis nymph El Bejuco Central coastal peridomiciliary Human T. cruzi 165 TDZ2016 R. ecuadoriensis adult Danzarin Central coastal sylvatic Human T. cruzi 166 TDZ2022 R. ecuadoriensis adult Danzarin Central coastal peridomiciliary Human no infection 167 TDZ2026 R. ecuadoriensis adult Danzarin Central coastal peridomiciliary non-human T. cruzi 168 TDZ2027 R. ecuadoriensis adult Danzarin Central coastal peridomiciliary Human T. cruzi Tablada del 169 TTO2051 R. ecuadoriensis nymph Central coastal intradomiciliary non-human T. cruzi algodón 170 TDZ2057 R. ecuadoriensis adult Danzarin Central coastal sylvatic Human T. cruzi 171 TDZ2068 R. ecuadoriensis nymph Danzarin Central coastal sylvatic Human T. cruzi 172 TDZ2074 R. ecuadoriensis adult Danzarin Central coastal peridomiciliary non-human T. rangeli Tablada del 173 TTO2086 R. ecuadoriensis nymph Central coastal intradomiciliary Human T. cruzi algodón 174 TDZ2101 R. ecuadoriensis nymph Danzarin Central coastal peridomiciliary non-human no infection 175 TDZ2110 R. ecuadoriensis adult Danzarin Central coastal peridomiciliary Human T. cruzi Tablada del 176 TTO2114 R. ecuadoriensis adult Central coastal sylvatic Human no infection algodón 177 TDZ2198 R. ecuadoriensis nymph Danzarin Central coastal sylvatic Human no infection 178 TDZ2201 R. ecuadoriensis nymph Danzarin Central coastal peridomiciliary Human no infection 179 TBJ2239 R. ecuadoriensis nymph El Bejuco Central coastal sylvatic Human T. rangeli 180 TBJ2240 R. ecuadoriensis nymph El Bejuco Central coastal sylvatic Human T. cruzi 181 TBJ2242 R. ecuadoriensis nymph El Bejuco Central coastal peridomiciliary Human T. cruzi 182 TBJ2243 R. ecuadoriensis adult El Bejuco Central coastal peridomiciliary non-human T. cruzi 183 TBJ2244 R. ecuadoriensis nymph El Bejuco Central coastal peridomiciliary non-human T. cruzi 184 TBJ2246 R. ecuadoriensis nymph El Bejuco Central coastal peridomiciliary Human T. cruzi 185 TBJ2248 R. ecuadoriensis nymph El Bejuco Central coastal peridomiciliary Human T. cruzi 186 TBJ2249 R. ecuadoriensis adult El Bejuco Central coastal peridomiciliary Human no infection 187 TBJ2250 R. ecuadoriensis nymph El Bejuco Central coastal peridomiciliary Human T. cruzi 162

188 TBJ2251 R. ecuadoriensis adult El Bejuco Central coastal peridomiciliary non-human T. cruzi 189 TBJ2254 R. ecuadoriensis nymph El Bejuco Central coastal peridomiciliary non-human T. cruzi 190 TBJ2257 R. ecuadoriensis adult El Bejuco Central coastal peridomiciliary Human T. cruzi 191 TBJ2258 R. ecuadoriensis nymph El Bejuco Central coastal sylvatic Human T. cruzi 192 TBJ2259 R. ecuadoriensis nymph El Bejuco Central coastal sylvatic Human T. cruzi 193 TBJ2260 R. ecuadoriensis adult El Bejuco Central coastal sylvatic Human T. cruzi 194 TBJ2266 R. ecuadoriensis adult El Bejuco Central coastal peridomiciliary non-human T. cruzi 195 TBJ2267 R. ecuadoriensis adult El Bejuco Central coastal peridomiciliary non-human T. cruzi 196 TBJ2268 R. ecuadoriensis nymph El Bejuco Central coastal sylvatic Human T. rangeli 197 TBJ2270 R. ecuadoriensis nymph El Bejuco Central coastal sylvatic non-human Mixed 198 TBJ2271 R. ecuadoriensis nymph El Bejuco Central coastal sylvatic Human T. rangeli 199 TBJ2272 R. ecuadoriensis nymph El Bejuco Central coastal sylvatic Human Mixed 200 TBJ2273 R. ecuadoriensis nymph El Bejuco Central coastal sylvatic non-human T. rangeli 201 TBJ2275 R. ecuadoriensis adult El Bejuco Central coastal sylvatic Human T. cruzi 202 TBJ2276 R. ecuadoriensis adult El Bejuco Central coastal sylvatic Human T. cruzi 203 TBJ2277 R. ecuadoriensis nymph El Bejuco Central coastal sylvatic Human T. cruzi 204 TBJ2278 R. ecuadoriensis nymph El Bejuco Central coastal sylvatic Human T. rangeli 205 TBJ2279 R. ecuadoriensis nymph El Bejuco Central coastal sylvatic non-human T. cruzi 206 TBJ2280 R. ecuadoriensis nymph El Bejuco Central coastal sylvatic Human T. cruzi 207 TBJ2281 R. ecuadoriensis nymph El Bejuco Central coastal sylvatic Human T. cruzi 208 TBJ2282 R. ecuadoriensis nymph El Bejuco Central coastal sylvatic non-human T. cruzi 209 TBJ2291 R. ecuadoriensis nymph El Bejuco Central coastal peridomiciliary non-human no infection 210 TBJ2292 R. ecuadoriensis nymph El Bejuco Central coastal peridomiciliary non-human T. cruzi 211 TBJ2295 R. ecuadoriensis adult El Bejuco Central coastal peridomiciliary non-human no infection 212 TBJ2310 R. ecuadoriensis adult El Bejuco Central coastal peridomiciliary non-human no infection 213 TBJ2448 R. ecuadoriensis nymph El Bejuco Central coastal sylvatic non-human T. rangeli 214 TBJ2452 R. ecuadoriensis nymph El Bejuco Central coastal sylvatic non-human no infection 215 TBJ2455 R. ecuadoriensis nymph El Bejuco Central coastal sylvatic Human no infection 163

216 TBJ2456 R. ecuadoriensis adult El Bejuco Central coastal sylvatic non-human T. cruzi 217 TCQ2493 R. ecuadoriensis adult Chaquizhca Southern highlands intradomiciliary non-human T. cruzi 218 TCQ2494 R. ecuadoriensis adult Chaquizhca Southern highlands intradomiciliary non-human T. cruzi 219 TCQ2502 R. ecuadoriensis adult Chaquizhca Southern highlands intradomiciliary non-human no infection 220 TCE2532 R. ecuadoriensis adult Coamine Southern highlands peridomiciliary non-human no infection 221 TCE2534 R. ecuadoriensis adult Coamine Southern highlands peridomiciliary non-human no infection 222 TCE2538 R. ecuadoriensis adult Coamine Southern highlands peridomiciliary non-human no infection 223 TCE2542 R. ecuadoriensis adult Coamine Southern highlands peridomiciliary non-human no infection 224 TCE2543 R. ecuadoriensis adult Coamine Southern highlands peridomiciliary non-human no infection 225 TCE2564 R. ecuadoriensis adult Coamine Southern highlands peridomiciliary non-human T. cruzi 226 TCE2565 R. ecuadoriensis adult Coamine Southern highlands peridomiciliary non-human T. cruzi 227 TCE2566 R. ecuadoriensis adult Coamine Southern highlands peridomiciliary Human T. cruzi 228 TCE2567 R. ecuadoriensis adult Coamine Southern highlands peridomiciliary non-human T. cruzi 229 TCE2568 R. ecuadoriensis adult Coamine Southern highlands peridomiciliary non-human T. cruzi 230 TCE2569 R. ecuadoriensis adult Coamine Southern highlands peridomiciliary non-human T. cruzi 231 TCE2570 R. ecuadoriensis adult Coamine Southern highlands peridomiciliary non-human no infection 232 TCE2572 R. ecuadoriensis nymph Coamine Southern highlands peridomiciliary non-human T. cruzi 233 TCE2585 R. ecuadoriensis adult Coamine Southern highlands peridomiciliary non-human T. cruzi 234 TCE2587 R. ecuadoriensis adult Coamine Southern highlands peridomiciliary non-human T. cruzi 235 TCE2589 R. ecuadoriensis adult Coamine Southern highlands peridomiciliary Human T. cruzi 236 TCE2590 R. ecuadoriensis nymph Coamine Southern highlands peridomiciliary non-human T. cruzi 237 TCQ2595 P. chinai nymph Chaquizhca Southern highlands intradomiciliary Human T. cruzi 238 TCE2598 R. ecuadoriensis adult Coamine Southern highlands peridomiciliary non-human T. cruzi 239 TCE2602 T. carrioni nymph Coamine Southern highlands intradomiciliary Human T. cruzi 240 TBM2775 P. chinai nymph Bellamaria Southern highlands intradomiciliary non-human no infection 241 TBM2776 P. chinai nymph Bellamaria Southern highlands intradomiciliary non-human T. cruzi 242 TBM2777 P. chinai nymph Bellamaria Southern highlands intradomiciliary non-human no infection 243 TBM2778 P. chinai nymph Bellamaria Southern highlands intradomiciliary non-human T. cruzi 164

244 TBM2779 P. chinai nymph Bellamaria Southern highlands intradomiciliary non-human T. cruzi 245 TBM2781 P. chinai nymph Bellamaria Southern highlands intradomiciliary Human no infection 246 TBM2782 P. chinai nymph Bellamaria Southern highlands intradomiciliary non-human no infection 247 TBM2784 P. chinai nymph Bellamaria Southern highlands intradomiciliary non-human no infection 248 TBM2785 P. chinai nymph Bellamaria Southern highlands intradomiciliary non-human no infection 249 TBM2786 P. chinai nymph Bellamaria Southern highlands intradomiciliary non-human T. cruzi 250 TBM2789 P. chinai nymph Bellamaria Southern highlands intradomiciliary non-human T. cruzi 251 TBM2790 P. chinai nymph Bellamaria Southern highlands intradomiciliary non-human no infection 252 TBM2791 P. chinai nymph Bellamaria Southern highlands intradomiciliary non-human T. cruzi 253 TBM2792 P. chinai nymph Bellamaria Southern highlands intradomiciliary non-human T. cruzi 254 TBM2794 P. chinai nymph Bellamaria Southern highlands intradomiciliary non-human T. cruzi 255 TBM2795 P. chinai nymph Bellamaria Southern highlands intradomiciliary non-human T. cruzi 256 TBM2796 P. chinai nymph Bellamaria Southern highlands intradomiciliary non-human T. cruzi 257 TBM2797 P. chinai nymph Bellamaria Southern highlands intradomiciliary non-human T. cruzi 258 TBM2798 P. chinai nymph Bellamaria Southern highlands intradomiciliary non-human T. cruzi 259 TBM2800 P. chinai nymph Bellamaria Southern highlands intradomiciliary Human T. cruzi 260 TBM2804 P. chinai nymph Bellamaria Southern highlands intradomiciliary Human no infection 261 TBM2805 P. chinai nymph Bellamaria Southern highlands intradomiciliary Human no infection 262 TBM2806 P. chinai nymph Bellamaria Southern highlands intradomiciliary Human T. cruzi 263 TBM2808 P. chinai nymph Bellamaria Southern highlands intradomiciliary non-human T. cruzi 264 TBM2809 P. chinai nymph Bellamaria Southern highlands intradomiciliary Human T. cruzi 265 TBM2811 P. chinai nymph Bellamaria Southern highlands intradomiciliary non-human T. cruzi 266 TBM2812 P. chinai nymph Bellamaria Southern highlands intradomiciliary non-human T. cruzi 267 TBM2813 P. chinai nymph Bellamaria Southern highlands intradomiciliary Human T. cruzi 268 TCQ2818 R. ecuadoriensis adult Chaquizhca Southern highlands intradomiciliary Human no infection 269 TCQ2819 R. ecuadoriensis nymph Chaquizhca Southern highlands intradomiciliary Human T. cruzi 270 TBM2820 P. chinai nymph Bellamaria Southern highlands intradomiciliary non-human T. cruzi 271 TBM2822 P. chinai nymph Bellamaria Southern highlands intradomiciliary non-human T. cruzi 165

272 TBM2824 P. chinai nymph Bellamaria Southern highlands intradomiciliary Human T. cruzi 273 TCQ2840 R. ecuadoriensis adult Chaquizhca Southern highlands peridomiciliary non-human T. cruzi 274 TCQ2842 R. ecuadoriensis adult Chaquizhca Southern highlands peridomiciliary non-human T. cruzi 275 TCQ2843 R. ecuadoriensis adult Chaquizhca Southern highlands peridomiciliary non-human T. cruzi 276 TCQ2844 R. ecuadoriensis adult Chaquizhca Southern highlands peridomiciliary non-human T. cruzi 277 TBM2845 R. ecuadoriensis adult Bellamaria Southern highlands sylvatic Human T. cruzi 278 TBM2846 R. ecuadoriensis adult Bellamaria Southern highlands sylvatic Human T. rangeli 279 TBM2848 R. ecuadoriensis nymph Bellamaria Southern highlands peridomiciliary non-human no infection 280 TCQ2852 R. ecuadoriensis adult Chaquizhca Southern highlands peridomiciliary non-human T. rangeli 281 TCQ2854 R. ecuadoriensis nymph Chaquizhca Southern highlands peridomiciliary non-human T. cruzi 282 TCQ2863 P. chinai adult Chaquizhca Southern highlands intradomiciliary non-human T. cruzi 283 TBM2873 R. ecuadoriensis adult Bellamaria Southern highlands intradomiciliary Human T. cruzi 284 TCQ2877 R. ecuadoriensis adult Chaquizhca Southern highlands peridomiciliary non-human T. cruzi 285 TCQ2878 R. ecuadoriensis nymph Chaquizhca Southern highlands peridomiciliary non-human T. cruzi 286 TCQ2880 R. ecuadoriensis nymph Chaquizhca Southern highlands intradomiciliary Human T. cruzi 287 TCQ2881 R. ecuadoriensis nymph Chaquizhca Southern highlands peridomiciliary non-human T. rangeli 288 TCQ2882 R. ecuadoriensis adult Chaquizhca Southern highlands peridomiciliary non-human T. rangeli 289 TCQ2885 R. ecuadoriensis nymph Chaquizhca Southern highlands peridomiciliary non-human T. cruzi 290 TCQ2888 R. ecuadoriensis nymph Chaquizhca Southern highlands peridomiciliary non-human Mixed 291 TCQ2890 R. ecuadoriensis nymph Chaquizhca Southern highlands peridomiciliary non-human T. rangeli 292 TBM2892 P. chinai nymph Bellamaria Southern highlands peridomiciliary Human T. cruzi 293 TBM2893 P. chinai nymph Bellamaria Southern highlands peridomiciliary non-human T. cruzi 294 TBM2894 R. ecuadoriensis adult Bellamaria Southern highlands intradomiciliary non-human T. cruzi 295 TCQ2895 P. chinai nymph Chaquizhca Southern highlands intradomiciliary Human T. cruzi 296 TCQ2896 R. ecuadoriensis adult Chaquizhca Southern highlands peridomiciliary Human T. cruzi 297 TCQ2898 R. ecuadoriensis nymph Chaquizhca Southern highlands peridomiciliary Human T. cruzi 298 TCQ2902 R. ecuadoriensis nymph Chaquizhca Southern highlands intradomiciliary Human T. cruzi 299 TBM2903 R. ecuadoriensis nymph Bellamaria Southern highlands peridomiciliary non-human T. cruzi 166

300 TCQ2906 R. ecuadoriensis nymph Chaquizhca Southern highlands peridomiciliary non-human T. rangeli 301 TCQ2907 R. ecuadoriensis nymph Chaquizhca Southern highlands peridomiciliary non-human Mixed 302 TCQ2912 R. ecuadoriensis nymph Chaquizhca Southern highlands peridomiciliary non-human T. rangeli 303 TCQ2922 R. ecuadoriensis nymph Chaquizhca Southern highlands peridomiciliary non-human T. rangeli 304 TCQ2928 R. ecuadoriensis adult Chaquizhca Southern highlands peridomiciliary non-human T. cruzi 305 TCQ2929 R. ecuadoriensis adult Chaquizhca Southern highlands peridomiciliary non-human T. cruzi 306 TCQ2930 R. ecuadoriensis adult Chaquizhca Southern highlands peridomiciliary non-human T. cruzi 307 TCQ2934 R. ecuadoriensis adult Chaquizhca Southern highlands peridomiciliary non-human T. cruzi 308 TCQ2936 R. ecuadoriensis nymph Chaquizhca Southern highlands peridomiciliary non-human T. cruzi 309 TCQ2939 R. ecuadoriensis nymph Chaquizhca Southern highlands peridomiciliary non-human T. cruzi 310 TCQ2940 R. ecuadoriensis nymph Chaquizhca Southern highlands peridomiciliary non-human T. cruzi 311 TCQ2941 R. ecuadoriensis nymph Chaquizhca Southern highlands peridomiciliary non-human T. cruzi 312 TCQ2943 R. ecuadoriensis nymph Chaquizhca Southern highlands peridomiciliary non-human T. cruzi 313 TCQ2944 R. ecuadoriensis nymph Chaquizhca Southern highlands peridomiciliary non-human T. cruzi 314 TCQ2946 R. ecuadoriensis nymph Chaquizhca Southern highlands peridomiciliary non-human T. cruzi 315 TBM2949 R. ecuadoriensis adult Bellamaria Southern highlands peridomiciliary non-human T. cruzi 316 TBM2950 R. ecuadoriensis adult Bellamaria Southern highlands peridomiciliary Human T. cruzi 317 TBM2954 R. ecuadoriensis adult Bellamaria Southern highlands peridomiciliary non-human T. cruzi 318 TBM2956 R. ecuadoriensis adult Bellamaria Southern highlands peridomiciliary non-human T. cruzi 319 TBM2957 R. ecuadoriensis adult Bellamaria Southern highlands peridomiciliary non-human T. cruzi 320 TBM2958 R. ecuadoriensis adult Bellamaria Southern highlands peridomiciliary non-human T. cruzi 321 TBM2960 R. ecuadoriensis adult Bellamaria Southern highlands peridomiciliary Human T. cruzi 322 TBM2961 R. ecuadoriensis adult Bellamaria Southern highlands peridomiciliary non-human T. cruzi 323 TBM2969 R. ecuadoriensis nymph Bellamaria Southern highlands peridomiciliary Human T. cruzi 324 TBM2971 R. ecuadoriensis adult Bellamaria Southern highlands peridomiciliary Human T. cruzi 325 TBM2979 R. ecuadoriensis adult Bellamaria Southern highlands peridomiciliary non-human T. cruzi 326 TBM2980 R. ecuadoriensis adult Bellamaria Southern highlands peridomiciliary Human T. cruzi 327 TBM2983 R. ecuadoriensis nymph Bellamaria Southern highlands peridomiciliary non-human T. cruzi 167

328 TCQ2985 R. ecuadoriensis nymph Chaquizhca Southern highlands peridomiciliary non-human T. cruzi 329 TCQ2986 R. ecuadoriensis nymph Chaquizhca Southern highlands peridomiciliary non-human T. cruzi 330 TCQ2989 R. ecuadoriensis nymph Chaquizhca Southern highlands peridomiciliary non-human T. cruzi 331 TCQ2990 R. ecuadoriensis nymph Chaquizhca Southern highlands peridomiciliary non-human T. cruzi 332 TCQ2991 R. ecuadoriensis nymph Chaquizhca Southern highlands peridomiciliary non-human T. cruzi Estero de Noche 333 TNC3001 R. ecuadoriensis adult Central coastal peridomiciliary Human no infection Arriba Estero de Noche 334 TNC3002 R. ecuadoriensis adult Central coastal peridomiciliary non-human T. cruzi Arriba Estero de Noche 335 TNC3031 R. ecuadoriensis nymph Central coastal intradomiciliary non-human T. cruzi Arriba Estero de Noche 336 TNC3037 R. ecuadoriensis nymph Central coastal peridomiciliary non-human T. cruzi Arriba Estero de Noche 337 TNC3039 R. ecuadoriensis nymph Central coastal peridomiciliary non-human T. cruzi Arriba Estero de Noche 338 TNC3040 R. ecuadoriensis adult Central coastal peridomiciliary non-human T. cruzi Arriba Estero de Noche 339 TNC3045 R. ecuadoriensis nymph Central coastal peridomiciliary non-human T. cruzi Arriba 340 TCQ3084 R. ecuadoriensis adult Chaquizhca Southern highlands sylvatic non-human Mixed 341 TCQ3088 R. ecuadoriensis nymph Chaquizhca Southern highlands sylvatic non-human T. rangeli 342 TCQ3089 R. ecuadoriensis nymph Chaquizhca Southern highlands sylvatic non-human T. cruzi 343 TCQ3091 R. ecuadoriensis nymph Chaquizhca Southern highlands sylvatic non-human T. cruzi 344 TCQ3092 R. ecuadoriensis nymph Chaquizhca Southern highlands sylvatic Human T. rangeli 345 TBM3101 R. ecuadoriensis adult Bellamaria Southern highlands sylvatic Human T. cruzi 346 TBM3102 R. ecuadoriensis adult Bellamaria Southern highlands intradomiciliary Human T. cruzi 347 TCQ3103 R. ecuadoriensis nymph Chaquizhca Southern highlands sylvatic non-human T. rangeli 348 TCQ3104 R. ecuadoriensis nymph Chaquizhca Southern highlands sylvatic Human T. rangeli 349 TCQ3107 R. ecuadoriensis nymph Chaquizhca Southern highlands sylvatic Human T. cruzi 168

350 TCQ3111 R. ecuadoriensis adult Chaquizhca Southern highlands peridomiciliary Human T. cruzi 351 TCQ3113 R. ecuadoriensis adult Chaquizhca Southern highlands peridomiciliary Human T. cruzi 352 TCQ3114 R. ecuadoriensis adult Chaquizhca Southern highlands peridomiciliary Human T. cruzi 353 TCQ3115 R. ecuadoriensis adult Chaquizhca Southern highlands peridomiciliary non-human T. rangeli 354 TBM3131 R. ecuadoriensis nymph Bellamaria Southern highlands sylvatic non-human T. cruzi 355 TBM3134 R. ecuadoriensis adult Bellamaria Southern highlands sylvatic Human T. rangeli 356 TCQ3139 P. chinai nymph Chaquizhca Southern highlands peridomiciliary Human T. cruzi 357 TBM3143 R. ecuadoriensis nymph Bellamaria Southern highlands sylvatic Human T. cruzi 358 TBM3144 R. ecuadoriensis nymph Bellamaria Southern highlands sylvatic Human T. cruzi 359 TBM3148 R. ecuadoriensis nymph Bellamaria Southern highlands peridomiciliary Human no infection 360 TBM3151 R. ecuadoriensis nymph Bellamaria Southern highlands peridomiciliary Human T. cruzi 361 TBM3185 R. ecuadoriensis adult Bellamaria Southern highlands peridomiciliary Human T. cruzi 362 TBM3186 R. ecuadoriensis adult Bellamaria Southern highlands peridomiciliary Human T. cruzi 363 TBM3190 R. ecuadoriensis nymph Bellamaria Southern highlands sylvatic non-human no infection 364 TCQ3197 R. ecuadoriensis nymph Chaquizhca Southern highlands peridomiciliary non-human T. cruzi 365 TCQ3215 R. ecuadoriensis nymph Chaquizhca Southern highlands peridomiciliary non-human T. cruzi 366 TCQ3218 R. ecuadoriensis nymph Chaquizhca Southern highlands peridomiciliary non-human T. cruzi 367 TCQ3222 R. ecuadoriensis nymph Chaquizhca Southern highlands peridomiciliary Human T. cruzi 368 TCQ3223 R. ecuadoriensis nymph Chaquizhca Southern highlands peridomiciliary Human T. cruzi 369 TCQ3224 R. ecuadoriensis nymph Chaquizhca Southern highlands peridomiciliary Human T. cruzi 370 TCQ3225 R. ecuadoriensis nymph Chaquizhca Southern highlands peridomiciliary Human T. cruzi 371 TBM3234 R. ecuadoriensis nymph Bellamaria Southern highlands sylvatic Human T. cruzi 372 TBM3235 R. ecuadoriensis nymph Bellamaria Southern highlands sylvatic Human no infection 373 TBM3236 R. ecuadoriensis nymph Bellamaria Southern highlands sylvatic non-human T. cruzi 374 TBM3239 R. ecuadoriensis nymph Bellamaria Southern highlands sylvatic Human no infection 375 TBM3241 R. ecuadoriensis nymph Bellamaria Southern highlands sylvatic Human no infection 376 TBM3290 R. ecuadoriensis adult Bellamaria Southern highlands sylvatic non-human Mixed 377 TBM3291 R. ecuadoriensis adult Bellamaria Southern highlands sylvatic non-human Mixed 169

378 TBM3292 R. ecuadoriensis adult Bellamaria Southern highlands sylvatic non-human Mixed 379 TBM3294 R. ecuadoriensis adult Bellamaria Southern highlands sylvatic non-human Mixed 380 TBM3295 R. ecuadoriensis adult Bellamaria Southern highlands sylvatic non-human T. rangeli 381 TBM3296 R. ecuadoriensis adult Bellamaria Southern highlands sylvatic Human T. rangeli 382 TBM3297 R. ecuadoriensis adult Bellamaria Southern highlands sylvatic non-human T. cruzi 383 TBM3300 R. ecuadoriensis nymph Bellamaria Southern highlands sylvatic non-human Mixed 384 TBM3302 R. ecuadoriensis nymph Bellamaria Southern highlands sylvatic non-human T. rangeli 385 TBM3304 R. ecuadoriensis nymph Bellamaria Southern highlands sylvatic non-human T. rangeli 386 TBM3308 R. ecuadoriensis nymph Bellamaria Southern highlands sylvatic non-human T. rangeli 387 TBM3309 R. ecuadoriensis nymph Bellamaria Southern highlands sylvatic non-human T. rangeli 388 TBM3310 R. ecuadoriensis nymph Bellamaria Southern highlands sylvatic non-human T. rangeli 389 TBM3312 R. ecuadoriensis adult Bellamaria Southern highlands sylvatic non-human T. rangeli 390 TBM3313 R. ecuadoriensis adult Bellamaria Southern highlands sylvatic non-human T. rangeli 391 TBM3314 R. ecuadoriensis adult Bellamaria Southern highlands sylvatic non-human T. rangeli 392 TBM3316 R. ecuadoriensis adult Bellamaria Southern highlands sylvatic non-human T. rangeli 393 TBM3317 R. ecuadoriensis adult Bellamaria Southern highlands sylvatic non-human T. rangeli 394 TBM3318 R. ecuadoriensis adult Bellamaria Southern highlands sylvatic non-human T. rangeli 395 TBM3320 R. ecuadoriensis adult Bellamaria Southern highlands sylvatic Human T. rangeli 396 TBM3323 R. ecuadoriensis adult Bellamaria Southern highlands sylvatic non-human T. rangeli 397 TBM3326 R. ecuadoriensis nymph Bellamaria Southern highlands sylvatic Human T. rangeli 398 TBM3337 R. ecuadoriensis nymph Bellamaria Southern highlands sylvatic non-human T. rangeli 399 TBM3338 R. ecuadoriensis nymph Bellamaria Southern highlands sylvatic non-human T. rangeli 400 TBM3340 R. ecuadoriensis nymph Bellamaria Southern highlands sylvatic non-human T. rangeli 401 TBM3341 R. ecuadoriensis nymph Bellamaria Southern highlands sylvatic non-human Mixed 402 TBM3342 R. ecuadoriensis nymph Bellamaria Southern highlands sylvatic non-human T. rangeli 403 TBM3344 R. ecuadoriensis adult Bellamaria Southern highlands sylvatic non-human Mixed 404 TBM3357 R. ecuadoriensis adult Bellamaria Southern highlands sylvatic non-human Mixed 405 TBM3368 R. ecuadoriensis adult Bellamaria Southern highlands sylvatic Human T. cruzi 170

406 TBM3376 R. ecuadoriensis nymph Bellamaria Southern highlands sylvatic non-human T. cruzi 407 TBM3409 R. ecuadoriensis adult Bellamaria Southern highlands sylvatic Human Mixed Tablada del 408 TTO3424 R. ecuadoriensis adult Central coastal sylvatic non-human T. cruzi algodón 409 TCD3437 R. ecuadoriensis adult Chade Central coastal sylvatic non-human T. rangeli 410 TCD3453 R. ecuadoriensis adult Chade Central coastal sylvatic non-human T. cruzi 411 TDZ3455 R. ecuadoriensis nymph Danzarin Central coastal sylvatic non-human T. cruzi 412 TDZ3456 R. ecuadoriensis nymph Danzarin Central coastal sylvatic non-human Mixed 413 TDZ3457 R. ecuadoriensis nymph Danzarin Central coastal sylvatic non-human T. cruzi 414 TBJ3475 R. ecuadoriensis nymph El Bejuco Central coastal sylvatic non-human T. cruzi 415 TCD3499 R. ecuadoriensis nymph Chade Central coastal sylvatic non-human T. cruzi 416 TCQ2814 R. ecuadoriensis adult Chaquizhca Southern highlands peridomiciliary non-human T. cruzi

171

APPENDIX D: DISTANCE FROM SYLVATIC PLACES OF CAPTURE TO THE CLOSEST HOUSE.

Sourcess of Triatomine Distance blood, 1:<100m, Closest infestation Collection Place of Microenvironment to the Locality Area 1: human, 2: 101-200, House 1: Infested/ place code capture of capture closest 2: non-human, 3: >201 code 2: non house (m) 3: mixed infested BJP01 El Bejuco CC animal shelter rodent nest (rat) 2 152.57 2 BJ206 2 animal shelter rodent nest 1 285.94 3 CE503 CEN20 Coamine SH (squirrel) 1 animal shelter rodent nest 3 40.99 1 GL103 GLN-44 Galapagos SH (squirrel) 1 BRN-21 Bramaderos animal shelter rodent nest 2 225.39 3 BR533 SH (squirrel) 2 BRN-3 Bramaderos animal shelter rodent nest 1 229.65 3 BR533 SH (squirrel) 2 BRN-20 Bramaderos SH animal shelter rodent nest (rat) 3 125.51 2 BR533 2 BRN-15 Bramaderos animal shelter rodent nest 2 60.45 1 BR108 SH (squirrel) 1 BRN4-1 Bramaderos animal shelter rodent nest 2 251.62 3 BR533 SH (squirrel) 2 BRN-24 Bramaderos animal shelter rodent nest 1 242.97 3 BR107 SH (squirrel) 1 BRN41-1 Bramaderos animal shelter rodent nest 1 251.62 3 BR533 SH (squirrel) 2 BRN-12 Bramaderos animal shelter rodent nest 1 144.73 BR533 SH (squirrel) 2 2 BRN-11 Bramaderos animal shelter rodent nest 2 156.72 BR533 SH (squirrel) 2 2 BRN-6 Bramaderos SH animal shelter rodent nest (rat) 1 246.2 3 BR620 1 172

BRN27-1 Bramaderos animal shelter rodent nest 1 270.17 3 BR620 SH (squirrel) 1 BRN32-3 Bramaderos animal shelter rodent nest 1 300.85 3 BR533 SH (squirrel) 2 BRN32 Bramaderos animal shelter rodent nest 3 260.39 3 BR534 SH (squirrel) 1 BRN-47 Bramaderos animal shelter rodent nest 2 321.85 3 BR603 SH (squirrel) 2 BRN-18 Bramaderos animal shelter rodent nest 1 260.39 3 BR534 SH (squirrel) 1 BRN32-1 Bramaderos SH animal shelter rodent nest (mouse) 1 246.31 3 BR534 1 BRN13 Bramaderos animal shelter rodent nest 2 144.73 BR533 SH (squirrel) 2 2 BRN-2 Bramaderos animal shelter rodent nest 2 175.66 BR533 SH (squirrel) 2 2 BJN046-1 El Bejuco CC animal shelter rodent nest (mouse) 1 140.19 2 BJ205 1 animal shelter rodent nest 1 67.77 1 BJN056-1 El Bejuco CC (squirrel) BJ210 2 animal shelter rodent nest 1 76.86 1 BJ308 BJN064-1 El Bejuco CC (squirrel) 2 animal shelter rodent nest 2 102.14 CD311 CDN223 Chade CC (squirrel) 2 2 animal shelter rodent nest 1 29.70 1 CD121 CDN210 Chade CC (squirrel) 2 animal shelter rodent nest 3 90.02 1 CD414 CDN221 Chade CC (squirrel) 2 animal shelter rodent nest 1 119.37 CD413 CDN213 Chade CC (squirrel) 2 2 animal shelter rodent nest 3 82.66 1 CD402 CDN104 Chade CC (squirrel) 2 173

animal shelter rodent nest 3 74.32 1 CD416 CDN212 Chade CC (squirrel) 2 animal shelter rodent nest 1 156.11 BJ304 BJN078 El Bejuco CC (squirrel) 2 1 animal shelter rodent nest 3 131.59 BJ205 BJN030-1 El Bejuco CC (squirrel) 2 1 animal shelter rodent nest 3 108.94 BJN02-3 El Bejuco CC (squirrel) 2 BJ209 1 BJN027-4 El Bejuco CC animal shelter rodent nest (mouse) 1 132.55 2 BJ304 1 animal shelter rodent nest 1 122.71 BJ208 BJN064-2 El Bejuco CC (squirrel) 2 2 DZN103 Danzarin CC animal shelter Bird nest 1 9.60 1 DZ202 1 animal shelter rodent nest 1 55.90 1 DZ306 DZN203 Danzarin CC (squirrel) 2 animal shelter rodent nest 1 76.26 1 DZ306 DZN202 Danzarin CC (squirrel) 2 DZN106 Danzarin CC animal shelter Bird nest 1 14.58 1 DZ202 1 Tablada del rodent nest TON217 CC animal shelter 1 231.17 3 TO317 2 algodón (squirrel) animal shelter rodent nest 1 17.75 1 DZ306 DZN204 Danzarin CC (squirrel) 2 animal shelter rodent nest 3 121.69 BJ205 BJN049-1 El Bejuco CC (squirrel) 2 1 animal shelter rodent nest 1 103.11 BJ210 BJN013 El Bejuco CC (squirrel) 2 2 animal shelter rodent nest 1 106.42 BJ308 BJN063 El Bejuco CC (squirrel) 2 2 animal shelter rodent nest 1 35.25 1 BJ210 BJN011-1 El Bejuco CC (squirrel) 2 animal shelter rodent nest 1 121.68 BJ205 BJN492 El Bejuco CC (squirrel) 2 1 174

animal shelter rodent nest 1 261.38 3 BJ306 BJN040-1 El Bejuco CC (squirrel) 1 BJN317 El Bejuco CC animal shelter rodent nest (mouse) 2 55.48 1 BJ228 1 BJN019 El Bejuco CC animal shelter Bird nest (pigeon) 2 162.32 2 BJ304 1 animal shelter rodent nest 3 173.29 BJ306 1 BJN107 El Bejuco CC (squirrel) 2 BMN311 Bellamaria SH animal shelter rodent nest 1 224.98 3 BM206 2 CQN04 Chaquizhca animal shelter rodent nest 2 324.99 3 CQ402 1 SH (squirrel) CQN08 Chaquizhca animal shelter rodent nest 3 385.55 3 CQ402 1 SH (squirrel) animal shelter rodent nest 1 286.35 3 BM107 2 BCN01 Bellamaria SH (squirrel) CQN10 Chaquizhca animal shelter rodent nest 1 363.43 3 CQ402 1 SH (squirrel) animal shelter rodent nest 3 172.46 BM107 2 BMN01 Bellamaria SH (squirrel) 2 animal shelter rodent nest 1 138.64 BM110 2 BMN07 Bellamaria SH (squirrel) 2 animal shelter rodent nest 2 127.94 BM110 2 BMN05 Bellamaria SH (squirrel) 2 animal shelter rodent nest 3 172.46 BM107 2 BMNR01 Bellamaria SH (squirrel) 2 animal shelter rodent nest 3 218.11 3 BM111 2 BMN21 Bellamaria SH (squirrel) animal shelter rodent nest 1 529.97 3 BM111 2 BMN27 Bellamaria SH (squirrel) animal shelter rodent nest 2 301.92 3 BM111 2 BMN24 Bellamaria SH (squirrel) animal shelter rodent nest 2 507.22 3 BM111 2 BMN29 Bellamaria SH (squirrel) 175

animal shelter rodent nest 3 240.23 3 BM111 2 BMNR09 Bellamaria SH (squirrel) Tablada del TOP003 CC tree organic matter 2 26.61 1 TO312 2 algodón CDP002- animal shelter 2 94.07 1 CD401 2 N3 Chade CC rodent nest (rat) CDP003- animal shelter 2 46.62 1 CD401 2 N3 Chade CC rodent nest (rat) DZP005 Danzarin CC animal shelter rodent nest (rat) 2 2182.15 3 DZ308 2 BJP001 El Bejuco CC tree organic matter 2 152.57 2 BJ206 2 CDP004- animal shelter 2 25.71 1 CD311 2 N1 Chade CC rodent nest (rat)

176

APPENDIX E: IDENTIFICATION OF MOSQUITOES COLLECTED IN COASTAL REGION (EL ORO PROVINCE) AND

AMAZON REGION (SUCUMBIOS AND ORELLANA PROVINCE) BY MORPHOLOGY, RFLP-ITS2 PATTERN AND COI

SEQUENCING.

RFLP-ITS2

Ident by Ident by COI Code Province Locality Morph. Ident Ident by AluI Consensus ident. HaeIII FspI Ident.

1 AWK0001 Orellana Kawymeno Anopheles spp. Ny. strodei NI NI Ny. strodei NP 2 AWK0002 Orellana Kawymeno Anopheles spp. Ny. strodei NI NI Ny. strodei NP 3 AWK0003 Orellana Kawymeno Anopheles spp. Ny. strodei NI NI Ny. strodei NP 4 AWK0004 Orellana Kawymeno Anopheles spp. Ny. strodei NI NI Ny. strodei NP AWK0005 Orellana Kawymeno NI 5 Anopheles spp. NP NP NI NI Ny. strodei/ An. AWK0006 Orellana Kawymeno NI 6 Anopheles spp. pseudopunctipennis NI Ny. strodei NP Ny. AWK0007 Orellana Kawymeno benarrochi 7 Anopheles spp. NI NP NP NI B 8 AWK0008 Orellana Kawymeno Anopheles spp. Ny. strodei NI NI Ny. strodei NP 9 AWK0009 Orellana Kawymeno Anopheles spp. Ny. strodei NI NI Ny. strodei NP 10 AWK0010 Orellana Kawymeno Anopheles spp. Ny. strodei NI NI Ny. strodei NP Ny. strodei/ An. AWK0011 Orellana Kawymeno NI NI 11 Anopheles spp. pseudopunctipennis Ny. strodei NP 12 AWK0012 Orellana Kawymeno Anopheles spp. NI NI NI NI NP 13 AWK0013 Orellana Kawymeno Anopheles spp. Ny. strodei NI NI Ny. strodei NP 14 AWK0014 Orellana Kawymeno Anopheles spp. Ny. rangeli NI NP Ny. rangeli NA 177

15 AWK0015 Orellana Kawymeno Anopheles spp. Ny. strodei NI NI Ny. strodei NP 16 AWK0016 Orellana Kawymeno Anopheles spp. Ny. strodei NI NI Ny. strodei NP Ny. strodei/ An. AWK0017 Orellana Kawymeno NI 17 Anopheles spp. pseudopunctipennis NI Ny. strodei NP Ny. strodei/ An. AWK0018 Orellana Kawymeno 18 Anopheles spp. pseudopunctipennis NI NI Ny. strodei NP 19 AWK0019 Orellana Kawymeno Anopheles spp. NI NI NI NI NP Ny. strodei/ An. AWK0020 Orellana Kawymeno NI 20 Anopheles spp. pseudopunctipennis NI Ny. strodei NP Ny. Ny. strodei/Ny. AWK0021 Orellana Kawymeno 21 Anopheles spp. Ny. strodei NI benarrochi benarrochi NP Ny. AWK0022 Orellana Kawymeno Ny. strodei/ An. NI NI benarrochi 22 Anopheles spp. pseudopunctipennis Ny. strodei B Ny. strodei/ An. AWK0023 Orellana Kawymeno NI NP 23 Anopheles spp. pseudopunctipennis NI Ny. strodei An. AWK0024 Orellana Kawymeno NI 24 Anopheles spp. pseudopunctipennis NI NI NP An. AWK0025 Orellana Kawymeno NI 25 Anopheles spp. pseudopunctipennis NI NI NP An. AWK0026 Orellana Kawymeno NI 26 Anopheles spp. pseudopunctipennis NI NI NP An. AWK0027 Orellana Kawymeno NI 27 Anopheles spp. pseudopunctipennis NI NI NP Ny. strodei/ An. AWK0028 Orellana Kawymeno NI 28 Anopheles spp. pseudopunctipennis NI Ny. strodei NP Ny. strodei/ An. AWK0029 Orellana Kawymeno 29 Anopheles spp. pseudopunctipennis NI NI Ny. strodei NP 30 AWK0030 Orellana Kawymeno Anopheles spp. NI NI NI NI NP Ny. strodei/ An. AWK0031 Orellana Kawymeno 31 Anopheles spp. pseudopunctipennis NI NI Ny. strodei NP Ny. strodei/ An. AWK0032 Orellana Kawymeno 32 Anopheles spp. pseudopunctipennis NI NI Ny. strodei NP 178

Ny. strodei/ An. Ny. Ny. strodei/Ny. AWK0033 Orellana Kawymeno 33 Anopheles spp. pseudopunctipennis NI benarrochi benarrochi NP Ny. strodei/ An. Ny. Ny. strodei/Ny. AWK0034 Orellana Kawymeno 34 Anopheles spp. pseudopunctipennis NI benarrochi benarrochi NP Ny. strodei/ An. AWK0035 Orellana Kawymeno NI 35 Anopheles spp. pseudopunctipennis NI Ny. strodei NP An. AWK0036 Orellana Kawymeno NI 36 Anopheles spp. pseudopunctipennis NI NI NP An. AWK0037 Orellana Kawymeno NI 37 Anopheles spp. pseudopunctipennis NI NI NP Ny. strodei/ An. AWK0038 Orellana Kawymeno NI NI 38 Anopheles spp. pseudopunctipennis Ny. strodei NP 39 AWK0039 Orellana Kawymeno Anopheles spp. NI NI NI NI NP 40 AWK0040 Orellana Kawymeno Anopheles spp. NI NI NI NI NP Ny. strodei/ An. AWK0041 Orellana Kawymeno NI NI 41 Anopheles spp. pseudopunctipennis Ny. strodei NP Ny. strodei/ An. AWK0042 Orellana Kawymeno NI NI 42 Anopheles spp. pseudopunctipennis Ny. strodei NP Ny. strodei/ An. AWK0043 Orellana Kawymeno NI NI 43 Anopheles spp. pseudopunctipennis Ny. strodei NP Ny. strodei/ An. AWK0044 Orellana Kawymeno NI NI 44 Anopheles spp. pseudopunctipennis Ny. strodei NP Ny. strodei/ An. AWK0045 Orellana Kawymeno NI 45 Anopheles spp. pseudopunctipennis NI Ny. strodei NP 46 AWK0046 Orellana Kawymeno Anopheles spp. Ny. strodei NI NI Ny. strodei NP 47 AWK0047 Orellana Kawymeno Anopheles spp. Ny. strodei NI NI Ny. strodei NP Ny. strodei/ An. AWK0048 Orellana Kawymeno 48 Anopheles spp. pseudopunctipennis NI NI Ny. strodei NP 49 AWK0049 Orellana Kawymeno Anopheles spp. Ny. strodei NI NI Ny. strodei NP 50 AWK0050 Orellana Kawymeno Ny. dunhami Ny. strodei NI NI Ny. strodei NP 51 AWK0051 Orellana Kawymeno Anopheles spp. Ny. strodei NI NI Ny. strodei NP 179

Ny. strodei/ An. AWK0052 Orellana Kawymeno 52 Ny. dunhami pseudopunctipennis NI NI Ny. strodei NP 53 AWK0053 Orellana Kawymeno Ny. dunhami NI NI NI NI NP Ny. strodei/ An. AWK0054 Orellana Kawymeno 54 Ny. dunhami pseudopunctipennis NI NI Ny. strodei NP 55 AWK0055 Orellana Kawymeno Ny. dunhami Ny. strodei NI NI Ny. strodei NP Ny. AWK0056 Orellana Kawymeno Ny. strodei/ An. NI NI benarrochi 56 Ny. dunhami pseudopunctipennis Ny. strodei B Ny. strodei/ An. AWK0057 Orellana Kawymeno NI 57 Ny. dunhami pseudopunctipennis NI Ny. strodei NP 58 AWK0058 Orellana Kawymeno Ny. dunhami NI NI NI NI NI Ny. strodei/ An. AWK0059 Orellana Kawymeno 59 Anopheles spp. pseudopunctipennis NI NI Ny. strodei NP 60 AWK0060 Orellana Kawymeno Anopheles spp. NI NI NI NI NP 61 AWK0061 Orellana Kawymeno Anopheles spp. NI NI NI NI NP Ny. strodei/ An. AWK0062 Orellana Kawymeno NI NI 62 Anopheles spp. pseudopunctipennis Ny. strodei NP 63 AWK0063 Orellana Kawymeno Anopheles spp. Ny. strodei NI NI Ny. strodei NP 64 AWK0064 Orellana Kawymeno Anopheles spp. Ny. strodei NI NI Ny. strodei NI An. AWK0065 Orellana Kawymeno 65 Anopheles spp. pseudopunctipennis NI NI NI NP Ny. strodei/ An. AWK0066 Orellana Kawymeno NI 66 Anopheles spp. pseudopunctipennis NI Ny. strodei NP An. AWK0067 Orellana Kawymeno 67 Anopheles spp. pseudopunctipennis NI NI NI NP Ny. strodei/ An. AWK0068 Orellana Kawymeno NI NI 68 Anopheles spp. pseudopunctipennis Ny. strodei NP 69 AWK0069 Orellana Kawymeno Anopheles spp. NI NI NI NI NP Ny. strodei/ An. AWK0070 Orellana Kawymeno NI 70 Anopheles spp. pseudopunctipennis NI Ny. strodei NP 180

Ny. strodei/ An. AWK0071 Orellana Kawymeno NI 71 Anopheles spp. pseudopunctipennis NI Ny. strodei NP 72 AWK0072 Orellana Kawymeno Anopheles spp. NI NI NI NI NP Ny. strodei/ An. AWK0073 Orellana Kawymeno NI NI 73 Anopheles spp. pseudopunctipennis Ny. strodei NP 74 AWK0074 Orellana Kawymeno Anopheles spp. Ny. strodei NI NI Ny. strodei NP 75 AWK0075 Orellana Kawymeno Anopheles spp. NI NI NI NI NP Ny. strodei/ An. AWK0076 Orellana Kawymeno NI 76 Anopheles spp. pseudopunctipennis NI Ny. strodei NP Ny. strodei/ An. AWK0077 Orellana Kawymeno NI 77 Anopheles spp. pseudopunctipennis NI Ny. strodei NP Ny. AWK0078 Orellana Kawymeno Ny. strodei/ An. NI NI benarrochi 78 Anopheles spp. pseudopunctipennis Ny. strodei B Ny. AWK0079 Orellana Kawymeno benarrochi 79 Anopheles spp. Ny. strodei NI NI Ny. strodei B 80 AWK0080 Orellana Kawymeno Anopheles spp. NI NI NI NI NP 81 AWK0081 Orellana Kawymeno Anopheles spp. Ny. strodei NI NI Ny. strodei NP 82 AWK0082 Orellana Kawymeno Anopheles spp. NI NI NI NI NP Ny. AWK0083 Orellana Kawymeno NI NI benarrochi 83 Anopheles spp. NI NI B Ny. AWK0084 Orellana Kawymeno NI NI benarrochi 84 Anopheles spp. NI NI B 85 AWK0085 Orellana Kawymeno Anopheles spp. Ny. strodei NI NI Ny. strodei NP 86 AWK0086 Orellana Kawymeno Anopheles spp. Ny. strodei NI NI Ny. strodei NP 87 AWK0087 Orellana Kawymeno Anopheles spp. NI NI NI NI NP 88 AWK0088 Orellana Kawymeno Anopheles spp. NI NI NI NI NP 89 AWK0089 Orellana Kawymeno Anopheles spp. NI NI NI NI NP 181

Ny. AWK0090 Orellana Kawymeno NI benarrochi 90 Anopheles spp. Ny. strodei NI Ny. strodei B 91 AWK0091 Orellana Kawymeno Anopheles spp. Ny. strodei NI NI Ny. strodei NP 92 AWK0092 Orellana Kawymeno Anopheles spp. Ny. strodei NI NI Ny. strodei NP 93 AWK0093 Orellana Kawymeno Anopheles spp. Ny. strodei NI NI Ny. strodei NP Ny. AWK0094 Orellana Kawymeno Ny. strodei/ An. benarrochi 94 Anopheles spp. pseudopunctipennis NI ND Ny. strodei B 95 AWK0095 Orellana Kawymeno Anopheles spp. Ny. strodei NI NI Ny. strodei NP 96 AWK0096 Orellana Kawymeno Anopheles spp. NI NI NI NI NP 97 AWK0097 Orellana Kawymeno Anopheles spp. Ny. strodei NI NI Ny. strodei NP Ny. strodei/ An. AWK0098 Orellana Kawymeno 98 Anopheles spp. pseudopunctipennis NI NI Ny. strodei NP 99 AWK0099 Orellana Kawymeno Anopheles spp. Ny. strodei NI NI Ny. strodei NP Ny. strodei/ An. AWK0100 Orellana Kawymeno 100 Anopheles spp. pseudopunctipennis NI NI Ny. strodei NP 101 AWK0101 Orellana Kawymeno Anopheles spp. Ny. strodei NI NI Ny. strodei NP 102 AWK0102 Orellana Kawymeno Anopheles spp. Ny. strodei NI NI Ny. strodei NP 103 AWK0103 Orellana Kawymeno Anopheles spp. Ny. strodei NI NI Ny. strodei NP 104 AWK0104 Orellana Kawymeno Anopheles spp. Ny. strodei NI NI Ny. strodei NP Ny. strodei/ An. AWK0105 Orellana Kawymeno 105 Anopheles spp. pseudopunctipennis NI NI Ny. strodei NP 106 AWK0106 Orellana Kawymeno Anopheles spp. Ny. strodei NI NI Ny. strodei NP 107 AWK0107 Orellana Kawymeno Anopheles spp. Ny. strodei NI NI Ny. strodei NP 108 AWK0108 Orellana Kawymeno Anopheles spp. Ny. strodei NI NI Ny. strodei NP 109 AWK0109 Orellana Kawymeno Anopheles spp. Ny. strodei NI NI Ny. strodei NP AWK0110 Orellana Kawymeno NI 110 Anopheles spp. NP NP NI NP 111 AWK0111 Orellana Kawymeno Anopheles spp. NI NP NP NI NP 182

112 AWK0112 Orellana Kawymeno Anopheles spp. NI NP NP NI NP 113 AWK0113 Orellana Kawymeno Anopheles spp. NI NI NI NI NP 114 AWK0114 Orellana Kawymeno Anopheles spp. NI NP NP NI NP An. ABN0115 El Oro Barbones NI 115 An. calderoni NI calderoni An. calderoni NP An. ABN0116 El Oro Barbones 116 An. calderoni NI calderoni NI An. calderoni NP An. ABN0117 El Oro Barbones 117 An. calderoni NI calderoni NI An. calderoni NP 118 ABN0118 El Oro Barbones Anopheles spp. Ny. strodei NI NI NI NP An. ABN0119 El Oro Barbones 119 An. calderoni Ny. strodei calderoni NI An. calderoni NP An. ABN0120 El Oro Barbones 120 An. calderoni NI calderoni NI An. calderoni NP An. ABN0121 El Oro Barbones 121 An. calderoni NI calderoni NI An. calderoni NP 122 ABN0122 El Oro Barbones An. calderoni NI NI NI NI NP An. ABN0123 El Oro Barbones NI 123 An. calderoni Ny. strodei calderoni An. calderoni NP 124 ABN0124 El Oro Barbones An. calderoni Ny. strodei NI NI NI NP 125 ABN0125 El Oro Barbones An. calderoni NI NI NI NI NP An. ABN0126 El Oro Barbones NI 126 An. calderoni NI calderoni An. calderoni NP 127 ABN0127 El Oro Barbones Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 128 ABN0128 El Oro Barbones Anopheles spp. Ny. strodei NI NI NI NP An. ABN0129 El Oro Barbones 129 Anopheles spp. Ny. strodei calderoni NI An. calderoni NP An. ABN0130 El Oro Barbones 130 Anopheles spp. Ny. strodei calderoni NI An. calderoni NP 131 ABN0131 El Oro Barbones Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 132 ABN0132 El Oro Barbones Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 183

133 ABN0133 El Oro Barbones Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 134 ABN0134 El Oro Barbones Anopheles spp. NA NA NA NA NP 135 ABN0135 El Oro Barbones Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 136 ABN0136 El Oro Barbones Ny. albimanus NP NP NP NP NP 137 ABN0137 El Oro Barbones Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 138 ABN0138 El Oro Barbones Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 139 ABN0139 El Oro Barbones Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 140 ABN0140 El Oro Barbones Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 141 ABN0141 El Oro Barbones Ny. albimanus NP NP NP NP NP 142 ABN0142 El Oro Barbones Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 143 ABN0143 El Oro Barbones Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 144 ABN0144 El Oro Barbones Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 145 ABN0145 El Oro Barbones Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 146 ABN0146 El Oro Barbones Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 147 ABN0147 El Oro Barbones Ny. albimanus NP NP NP NP NP 148 ABN0148 El Oro Barbones Ny. albimanus NP NP NP NP NP 149 ABN0149 El Oro Barbones Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 150 ABN0150 El Oro Barbones Ny. albimanus NP NP NP NP NP 151 ABN0151 El Oro Barbones Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 152 ABN0152 El Oro Barbones Ny. albimanus NP NP NP NP NP 153 ABN0153 El Oro Barbones Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 154 ABN0154 El Oro Barbones Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 155 ABN0155 El Oro Barbones Ny. albimanus NP NP NP NP NP 156 ABN0156 El Oro Barbones Ny. albimanus NP NP NP NP NP 157 ABN0157 El Oro Barbones Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 158 ABN0158 El Oro Barbones Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 159 ABN0159 El Oro Barbones Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 160 ABN0160 El Oro Barbones Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 184

161 ABN0161 El Oro Barbones Ny. albimanus NP NP NP NP NP 162 ABN0162 El Oro Barbones Ny. albimanus NP NP NP NP NP 163 ABN0163 El Oro Barbones Ny. albimanus NP NP NP NP NP 164 ABN0164 El Oro Barbones Ny. albimanus NP NP NP NP NP 165 ABN0165 El Oro Barbones Ny. albimanus NP NP NP NP NP 166 ABN0166 El Oro Barbones Ny. albimanus NP NP NP NP NP 167 ABN0167 El Oro Barbones Ny. albimanus NP NP NP NP NP 168 ABN0168 El Oro Barbones Ny. albimanus NP NP NP NP NP 169 ABN0169 El Oro Barbones Ny. albimanus NP NP NP NP NP 170 ABN0170 El Oro Barbones Ny. albimanus NP NP NP NP NP 171 ABN0171 El Oro Barbones Ny. albimanus NP NP NP NP NP 172 ABN0172 El Oro Barbones Ny. albimanus NP NP NP NP NP Ny. albimanus/Ny. ABN0173 El Oro Barbones Ny. albimanus 173 oswaldoi NP NP Ny. albimanus NP 174 ABN0174 El Oro Barbones Ny. albimanus NP NP NP NP NP 175 ABN0175 El Oro Barbones Ny. albimanus NP NP NP NP NP 176 ABN0176 El Oro Barbones Ny. albimanus NP NP NP NP NP 177 ABN0177 El Oro Barbones Anopheles spp. NI NP NP NI NP Ny. ABN0178 El Oro Barbones NI 178 Anopheles spp. NP NP NP albimanus 179 ASL0179 Sucumbíos Santa Elena Anopheles spp. NI NI NP NI NP 180 ASL0180 Sucumbíos Santa Elena Anopheles spp. NI NP NP NI NP Ny. ASL0181 Sucumbíos Santa Elena 181 Anopheles spp. NI NP NP NI rangeli 182 ASL0182 Sucumbíos Santa Elena Anopheles spp. Ny. rangeli NP NP Ny. rangeli NP 183 ASL0183 Sucumbíos Santa Elena Anopheles spp. Ny. rangeli NP NP Ny. rangeli NP 184 ASL0184 Sucumbíos Santa Elena Anopheles spp. NI NP NP NI NP 185 ASL0185 Sucumbíos Santa Elena Anopheles spp. NI NI NI NI NP 186 ASL0186 Sucumbíos Santa Elena Anopheles spp. NI NI NI NI NP 185

187 ASL0187 Sucumbíos Santa Elena Anopheles spp. NI NP NP NI NP Ny. albimanus/Ny. ASL0188 Sucumbíos Santa Elena 188 Anopheles spp. oswaldoi NP NP Ny. oswaldoi NP 189 ASL0189 Sucumbíos Santa Elena Anopheles spp. Ny. rangeli NI NP Ny. rangeli NP Ny. ASL0190 Sucumbíos Santa Elena NI 190 Anopheles spp. NI benarrochi Ny. benarrochi NP 191 ASL0191 Sucumbíos Santa Elena Anopheles spp. Ny. rangeli NP NP Ny. rangeli NP 192 ASL0192 Sucumbíos Santa Elena Anopheles spp. Ny. strodei NP NP Ny. strodei NP 193 ASL0193 Sucumbíos Santa Elena Anopheles spp. NI NP NP NI NP 194 ASL0194 Sucumbíos Santa Elena Anopheles spp. Ny. strodei NP NP Ny. strodei NP 195 ASL0195 Sucumbíos Santa Elena Anopheles spp. NI NP NP NI NP 196 ASL0196 Sucumbíos Santa Elena Anopheles spp. Ny. strodei NP NP Ny. strodei NP 197 ASL0197 Sucumbíos Santa Elena Anopheles spp. Ny. strodei NI NI Ny. strodei NP An. ASL0198 Sucumbíos Santa Elena NI 198 Anopheles spp. pseudopunctipennis NI NI NP 199 ASL0199 Sucumbíos Santa Elena Anopheles spp. NI NI NI NI NP 200 ASL0200 Sucumbíos Santa Elena Anopheles spp. Ny. rangeli NP NP Ny. rangeli NP 201 ASL0201 Sucumbíos Santa Elena Ny. albimanus Ny. rangeli NP NP Ny. rangeli NP 202 ASL0202 Sucumbíos Santa Elena Anopheles spp. NI NP NP NI NP 203 ASL0203 Sucumbíos Santa Elena Anopheles spp. Ny. strodei NP NP Ny. strodei NP 204 ASL0204 Sucumbíos Santa Elena Anopheles spp. Ny. strodei NP NP Ny. strodei NP 205 ASL0205 Sucumbíos Santa Elena Anopheles spp. Ny. albimanus NP NP NI NP 206 ASL0206 Sucumbíos Santa Elena Anopheles spp. NI NP NP NI NP 207 ASL0207 Sucumbíos Santa Elena Ny. albimanus Ny. rangeli NI NP Ny. rangeli NP 208 ASL0208 Sucumbíos Santa Elena Anopheles spp. NI NP NP NI NP 209 ASL0209 Sucumbíos Santa Elena Anopheles spp. NI NP NP NI NP 210 ASL0210 Sucumbíos Santa Elena Anopheles spp. Ny. strodei NI NI Ny. strodei NP 211 ASL0211 Sucumbíos Santa Elena Anopheles spp. NI NI NI NI NP 212 ASL0212 Sucumbíos Santa Elena Anopheles spp. NI NP NP NI NP 186

213 ASL0213 Sucumbíos Santa Elena Anopheles spp. NI NP NP NI NP 214 ASL0214 Sucumbíos Santa Elena Anopheles spp. NI NP NP NI NP Nueva ANM0215 Sucumbíos 215 Montepa Anopheles spp. Ny. rangeli NP NP Ny. rangeli NP Nueva Ny. albimanus/Ny. ANM0216 Sucumbíos 216 Montepa Anopheles spp. oswaldoi NP NP Ny. oswaldoi NP Nueva ANM0217 Sucumbíos 217 Montepa Anopheles spp. Ny. rangeli NP NP Ny. rangeli NP Nueva Ny. albimanus/Ny. ANM0218 Sucumbíos 218 Montepa Anopheles spp. oswaldoi NP NP Ny. oswaldoi NP Nueva ANM0219 Sucumbíos NI 219 Montepa Anopheles spp. NP NP NI NP Nueva Ny. albimanus/Ny. ANM0220 Sucumbíos 220 Montepa Anopheles spp. oswaldoi NP NP Ny. oswaldoi NP Nueva ANM0221 Sucumbíos NI 221 Montepa Anopheles spp. NP NP NI NP Ny. Nueva ANM0222 Sucumbíos NI oswaldoi Montepa 222 Anopheles spp. NP NP NI B Ny. Nueva ANM0223 Sucumbíos NI rangeli/Ny. Ny. rangeli/Ny. Montepa 223 Anopheles spp. oswaldoi NI oswaldoi NP Nueva Ny. albimanus/Ny. ANM0224 Sucumbíos 224 Montepa Anopheles spp. oswaldoi NI NP Ny. oswaldoi NP Nueva ANM0225 Sucumbíos NI 225 Montepa Anopheles spp. NP NP NI NP Nueva ANM0226 Sucumbíos NI 226 Montepa Anopheles spp. NP NP NI NP Nueva Ny. strodei/ An. ANM0227 Sucumbíos 227 Montepa Anopheles spp. pseudopunctipennis NI NI Ny. strodei NP Nueva ANM0228 Sucumbíos NI 228 Montepa Anopheles spp. NP NP NI NP 187

Nueva ANM0229 Sucumbíos NI 229 Montepa Anopheles spp. NI NI NI NP Nueva ANM0230 Sucumbíos NI 230 Montepa Anopheles spp. NP NP NI NP Nueva ANM0231 Sucumbíos NI NI 231 Montepa Anopheles spp. Ny. strodei Ny. strodei NP Nueva ANM0232 Sucumbíos NI 232 Montepa Anopheles spp. NP NP NI NP Nueva ANM0233 Sucumbíos NI 233 Montepa Anopheles spp. NP NP NI NP Nueva ANM0234 Sucumbíos NI 234 Montepa Anopheles spp. NP NP NI NP Nueva ANM0235 Sucumbíos NI 235 Montepa Anopheles spp. NP NP NI NP Nueva ANM0236 Sucumbíos NI 236 Montepa Anopheles spp. NP NP NI NP Nueva ANM0237 Sucumbíos NI 237 Montepa Anopheles spp. NP NP NI NP Nueva ANM0238 Sucumbíos 238 Montepa Anopheles spp. Ny. strodei NI NI Ny. strodei NP Nueva ANM0239 Sucumbíos 239 Montepa Anopheles spp. Ny. strodei NI NI Ny. strodei NP Nueva ANM0240 Sucumbíos Ny. albimanus 240 Montepa Ny. rangeli NP NP Ny. rangeli NP Nueva ANM0241 Sucumbíos NA 241 Montepa Anopheles spp. Ny. strodei NI NI Ny. strodei Nueva ANM0242 Sucumbíos NI NA 242 Montepa Anopheles spp. NP NP NI Nueva Ny. strodei/ An. ANM0243 Sucumbíos NP 243 Montepa Anopheles spp. pseudopunctipennis NI NI Ny. strodei Nueva Ny. albimanus/Ny. ANM0244 Sucumbíos NP 244 Montepa Anopheles spp. oswaldoi NP NP Ny. oswaldoi 245 ABR0245 Sucumbíos Bajo Restrepo Anopheles spp. NI NI NI NI NP 188

246 ABR0246 Sucumbíos Bajo Restrepo Anopheles spp. Ny. strodei NI NI Ny. strodei NP 247 ABR0247 Sucumbíos Bajo Restrepo Anopheles spp. Ny. strodei NI NI Ny. strodei NP 248 ABR0248 Sucumbíos Bajo Restrepo Anopheles spp. Ny. strodei NI NI Ny. strodei NP 249 ABR0249 Sucumbíos Bajo Restrepo Anopheles spp. NI NI NI NI NA 250 ABR0250 Sucumbíos Bajo Restrepo Anopheles spp. Ny. strodei NI NI Ny. strodei NP 251 ABR0251 Sucumbíos Bajo Restrepo Anopheles spp. NA NA NA NA NP 252 ABR0252 Sucumbíos Bajo Restrepo Anopheles spp. Ny. strodei NI NI Ny. strodei NP 253 ABR0253 Sucumbíos Bajo Restrepo Anopheles spp. NI NI NI NI NP 254 ABR0254 Sucumbíos Bajo Restrepo Anopheles spp. NI NI NI NI NP Ny. strodei/ An. ABR0255 Sucumbíos Bajo Restrepo NI NP 255 Anopheles spp. pseudopunctipennis NI Ny. strodei 256 ABR0256 Sucumbíos Bajo Restrepo Anopheles spp. NI NP NP NI NP 257 ABR0257 Sucumbíos Bajo Restrepo Anopheles spp. NA NA NA NA NP Ny. strodei/ An. ABR0258 Sucumbíos Bajo Restrepo NI NP 258 Anopheles spp. pseudopunctipennis NI Ny. strodei 259 ABR0259 Sucumbíos Bajo Restrepo Anopheles spp. Ny. strodei NI NI Ny. strodei NP 260 ABR0260 Sucumbíos Bajo Restrepo Anopheles spp. Ny. strodei NI NI Ny. strodei NP 261 ABR0261 Sucumbíos Bajo Restrepo Anopheles spp. Ny. strodei NI NI Ny. strodei NP Ny. strodei/ An. ABR0262 Sucumbíos Bajo Restrepo NI NI NP 262 Anopheles spp. pseudopunctipennis Ny. strodei 263 ABR0263 Sucumbíos Bajo Restrepo Anopheles spp. NA NA NA NA NP 264 ABR0264 Sucumbíos Bajo Restrepo Anopheles spp. NA NA NA NA NP An. ABR0265 Sucumbíos Bajo Restrepo NI NI NP 265 Anopheles spp. pseudopunctipennis NI 266 ABR0266 Sucumbíos Bajo Restrepo Anopheles spp. NI NP NP NI NP Ny. strodei/ An. ABR0267 Sucumbíos Bajo Restrepo NI NP 267 Anopheles spp. pseudopunctipennis NI Ny. strodei Ny. strodei/ An. ABR0268 Sucumbíos Bajo Restrepo NI NI NI 268 Anopheles spp. pseudopunctipennis Ny. strodei 189

Ny. strodei/ An. ABR0269 Sucumbíos Bajo Restrepo NI NI NP 269 Anopheles spp. pseudopunctipennis Ny. strodei 270 ABR0270 Sucumbíos Bajo Restrepo Anopheles spp. NI NI NI NI NP 271 ABR0271 Sucumbíos Bajo Restrepo Anopheles spp. NA NA NA NA NP 272 ABR0272 Sucumbíos Bajo Restrepo Anopheles spp. Ny. strodei NI NI Ny. strodei NP Ny. strodei/ An. ABR0273 Sucumbíos Bajo Restrepo NI NI NP 273 Anopheles spp. pseudopunctipennis Ny. strodei Ny. strodei/ An. ABR0274 Sucumbíos Bajo Restrepo NI NI NP 274 Anopheles spp. pseudopunctipennis Ny. strodei 275 ABR0275 Sucumbíos Bajo Restrepo Anopheles spp. NI NP NP NI NP 276 ABR0276 Sucumbíos Bajo Restrepo Anopheles spp. NA NA NA NA NP 277 ABR0277 Sucumbíos Bajo Restrepo Anopheles spp. NI NI NI NI NP An. ABR0278 Sucumbíos Bajo Restrepo NI NP 278 Anopheles spp. pseudopunctipennis NI NI 279 ABR0279 Sucumbíos Bajo Restrepo Anopheles spp. NA NA NA NI NP Ny. strodei/ An. ABR0280 Sucumbíos Bajo Restrepo NI NP 280 Anopheles spp. pseudopunctipennis NI Ny. strodei 281 ABR0281 Sucumbíos Bajo Restrepo Anopheles spp. Ny. strodei NI NI Ny. strodei NP Ny. ABR0282 Sucumbíos Bajo Restrepo Ny. strodei/ An. NI benarrochi 282 Anopheles spp. pseudopunctipennis NI Ny. strodei B 283 ABR0283 Sucumbíos Bajo Restrepo Anopheles spp. Ny. strodei NI NI Ny. strodei NP 284 ABR0284 Sucumbíos Bajo Restrepo Anopheles spp. NA NA NA NA NP Ny. ABR0285 Sucumbíos Bajo Restrepo NI rangeli/Ny. Ny. rangeli/Ny. NP 285 Anopheles spp. oswaldoi NI oswaldoi 286 ABR0286 Sucumbíos Bajo Restrepo Anopheles spp. Ny. strodei NI NI Ny. strodei NP 287 ABR0287 Sucumbíos Bajo Restrepo Anopheles spp. Ny. strodei NI NI Ny. strodei NP 288 ABR0288 Sucumbíos Bajo Restrepo Anopheles spp. NI NI NI NI NP Ny. strodei/ An. ABR0289 Sucumbíos Bajo Restrepo NP 289 Anopheles spp. pseudopunctipennis NI NI Ny. strodei 190

290 ABR0290 Sucumbíos Bajo Restrepo Anopheles spp. Ny. strodei NI NI Ny. strodei NP 291 ABR0291 Sucumbíos Bajo Restrepo Anopheles spp. NI NP NP NI NP 292 ABR0292 Sucumbíos Bajo Restrepo Anopheles spp. Ny. strodei NI NI Ny. strodei NP 293 ABR0293 Sucumbíos Bajo Restrepo Anopheles spp. NI NI NI NI NP 294 ABR0294 Sucumbíos Bajo Restrepo Anopheles spp. Ny. strodei NI NI Ny. strodei NP 295 ABR0295 Sucumbíos Bajo Restrepo Anopheles spp. NI NI NI NI NP 296 ABR0296 Sucumbíos Bajo Restrepo Anopheles spp. Ny. strodei NI NI Ny. strodei NP 297 APR0297 Sucumbíos Porvenir (106) Anopheles spp. Ny. rangeli NI NP Ny. rangeli NP 298 APR0298 Sucumbíos Porvenir (106) Anopheles spp. NI NP NP NI NP 299 APR0299 Sucumbíos Porvenir (106) Anopheles spp. NI NP NP NI NP Ny. albimanus/Ny. APR0300 Sucumbíos Porvenir (106) NP 300 Anopheles spp. oswaldoi NP NP Ny. oswaldoi 301 APR0301 Sucumbíos Porvenir (106) Anopheles spp. Ny. strodei NI NI Ny. strodei NP 302 APR0302 Sucumbíos Porvenir (106) Anopheles spp. NI NP NP NI NP 303 APR0303 Sucumbíos Porvenir (106) Anopheles spp. NI NP NP NI NP Ny. APR0304 Sucumbíos Porvenir (106) oswaldoi 304 Anopheles spp. Ny. strodei NP NP Ny. strodei B 305 APR0305 Sucumbíos Porvenir (106) Anopheles spp. Ny. strodei NP NP Ny. strodei NP 306 APR0306 Sucumbíos Porvenir (106) Anopheles spp. NI NI NP NI NP 307 APR0307 Sucumbíos Porvenir (106) Anopheles spp. Ny. strodei NP NP Ny. strodei NP 308 APR0308 Sucumbíos Porvenir (106) Anopheles spp. NI NP NP NI NP 309 APR0309 Sucumbíos Porvenir (106) Anopheles spp. NI NP NP NI NP 310 APR0310 Sucumbíos Porvenir (106) Anopheles spp. NI NP NP NI NP 311 APR0311 Sucumbíos Porvenir (106) Anopheles spp. NI NP NP NI NP Ny. APR0312 Sucumbíos Porvenir (106) oswaldoi 312 Anopheles spp. NI NP NP NI B 313 APR0313 Sucumbíos Porvenir (106) Anopheles spp. NI NP NP NI NP 191

Ny. APR0314 Sucumbíos Porvenir (106) 314 Ny. nuneztovari Ny. rangeli NP NP Ny. rangeli rangeli Ny. albimanus/Ny. Ny. af.f APR0315 Sucumbíos Porvenir (106) 315 Anopheles spp. oswaldoi NP NP Ny. oswaldoi konderi Ny. APR0316 Sucumbíos Porvenir (106) oswaldoi 316 Ny. oswaldoi NI NI NI NI B 317 APR0317 Sucumbíos Porvenir (106) Anopheles spp. NI NP NP NI NA 318 APR0318 Sucumbíos Porvenir (106) Anopheles spp. NI NP NP NI NP 319 APR0319 Sucumbíos Porvenir (106) Anopheles spp. Ny. rangeli NP NP Ny. rangeli NP 320 APR0320 Sucumbíos Porvenir (106) Anopheles spp. Ny. oswaldoi NP NP Ny. oswaldoi NP 321 APR0321 Sucumbíos Porvenir (106) Anopheles spp. NI NI NP NI NP 322 APR0322 Sucumbíos Porvenir (106) Anopheles spp. Ny. strodei NI NI Ny. strodei NP 323 APR0323 Sucumbíos Porvenir (106) Anopheles spp. NI NP NP NI NP Ny. albimanus/Ny. APR0324 Sucumbíos Porvenir (106) 324 Anopheles spp. oswaldoi NP NP Ny. oswaldoi NP 325 APR0325 Sucumbíos Porvenir (106) Ny. nuneztovari Ny. rangeli NP NP Ny. rangeli NP 326 APR0326 Sucumbíos Porvenir (106) Anopheles spp. NI NP NP NI NP Ny. albimanus/Ny. APR0327 Sucumbíos Porvenir (106) NP 327 Ny. oswaldoi oswaldoi NP NP Ny. oswaldoi 328 APR0328 Sucumbíos Porvenir (106) Anopheles spp. NI NP NP NI NP 329 APR0329 Sucumbíos Porvenir (106) Ny. albimanus Ny. rangeli NP NP Ny. rangeli NP Ny. albimanus/Ny. APR0330 Sucumbíos Porvenir (106) NP 330 Ny. oswaldoi oswaldoi NP NP Ny. oswaldoi 331 APR0331 Sucumbíos Porvenir (106) Anopheles spp. NI NP NP NI NP 332 APR0332 Sucumbíos Porvenir (106) Anopheles spp. NI NP NP NI NP 333 APR0333 Sucumbíos Porvenir (106) Anopheles spp. NI NP NP NI NP Ny. albimanus/Ny. APR0334 Sucumbíos Porvenir (106) 334 Anopheles spp. oswaldoi NP NP Ny. oswaldoi NP 335 APR0335 Sucumbíos Porvenir (106) Ny. oswaldoi Ny. rangeli NP NP Ny. rangeli NP 192

336 APR0336 Sucumbíos Porvenir (106) Anopheles spp. NI NP NP NI NP 337 APR0337 Sucumbíos Porvenir (106) Anopheles spp. Ny. strodei NP NP Ny. strodei NP 338 APR0338 Sucumbíos Porvenir (106) Anopheles spp. Ny. strodei NP NP Ny. strodei NP 339 APR0339 Sucumbíos Porvenir (106) Anopheles spp. Ny. rangeli NP NP Ny. rangeli NP 340 ALA0340 El Oro Los Ángeles Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 341 ALA0341 El Oro Los Ángeles Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 342 ALA0342 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 343 ALA0343 El Oro Los Ángeles Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 344 ALA0344 El Oro Los Ángeles Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP Ny. albimanus/Ny. ALA0345 El Oro Los Ángeles Ny. albimanus 345 oswaldoi NP NP Ny. albimanus NP Ny. albimanus/Ny. ALA0346 El Oro Los Ángeles Ny. albimanus 346 oswaldoi NP NP Ny. albimanus NP Ny. albimanus/Ny. ALA0347 El Oro Los Ángeles Ny. albimanus 347 oswaldoi NP NP Ny. albimanus NP Ny. albimanus/Ny. ALA0348 El Oro Los Ángeles Ny. albimanus 348 oswaldoi NP NP Ny. albimanus NP 349 ALA0349 El Oro Los Ángeles Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP Ny. albimanus/Ny. ALA0350 El Oro Los Ángeles Ny. albimanus 350 rangeli NP NP Ny. albimanus NP Ny. albimanus/Ny. ALA0351 El Oro Los Ángeles Ny. albimanus 351 oswaldoi NP NP Ny. albimanus NP Ny. albimanus/Ny. ALA0352 El Oro Los Ángeles Ny. albimanus 352 rangeli NP NP Ny. albimanus NP Ny. albimanus/Ny. ALA0353 El Oro Los Ángeles Ny. albimanus 353 oswaldoi NP NP Ny. albimanus NP 354 ALA0354 El Oro Los Ángeles Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 355 ALA0355 El Oro Los Ángeles Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP Ny. albimanus/Ny. ALA0356 El Oro Los Ángeles Ny. albimanus 356 strodei NP NP Ny. albimanus NP 193

Ny. albimanus/Ny. Ny. ALA0357 El Oro Los Ángeles Ny. albimanus 357 strodei NP NP Ny. albimanus albimanus Ny. albimanus/Ny. ALA0358 El Oro Los Ángeles Ny. albimanus 358 strodei NP NP Ny. albimanus NP 359 ALA0359 El Oro Los Ángeles Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 360 ALA0360 El Oro Los Ángeles Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 361 ALA0361 El Oro Los Ángeles Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP Ny. albimanus/Ny. ALA0362 El Oro Los Ángeles 362 Anopheles spp. rangeli NP NP Ny. albimanus NP 363 ALA0363 El Oro Los Ángeles Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 364 ALA0364 El Oro Los Ángeles Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP An. ALA0365 El Oro Los Ángeles 365 Anopheles spp. NI calderoni NI An. calderoni NP 366 ALA0366 El Oro Los Ángeles Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP An. ALA0367 El Oro Los Ángeles Anopheles spp. NP 367 Ny. strodei calderoni NI An. calderoni 368 ALA0368 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 369 ALA0369 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP An. ALA0370 El Oro Los Ángeles 370 Anopheles spp. Ny. strodei calderoni NI An. calderoni NP 371 ALA0371 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP An. ALA0372 El Oro Los Ángeles 372 Anopheles spp. NI calderoni NI An. calderoni NP 373 ALA0373 El Oro Los Ángeles Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 374 ALA0374 El Oro Los Ángeles Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 375 ALA0375 El Oro Los Ángeles Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP An. ALA0376 El Oro Los Ángeles 376 Anopheles spp. Ny. strodei calderoni NI An. calderoni NP 377 ALA0377 El Oro Los Ángeles Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 378 ALA0378 El Oro Los Ángeles Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 379 ALA0379 El Oro Los Ángeles Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 194

380 ALA0380 El Oro Los Ángeles Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 381 ALA0381 El Oro Los Ángeles Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP Ny. albimanus/Ny. ALA0382 El Oro Los Ángeles 382 Anopheles spp. oswaldoi NP NP Ny. albimanus NP 383 ALA0383 El Oro Los Ángeles Anopheles spp. Ny. strodei NI NI NI NP An. ALA0384 El Oro Los Ángeles 384 Anopheles spp. Ny. strodei calderoni NI An. calderoni NP An. ALA0385 El Oro Los Ángeles 385 Anopheles spp. NI calderoni NI An. calderoni NP 386 ALA0386 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 387 ALA0387 El Oro Los Ángeles Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 388 ALA0388 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 389 ALA0389 El Oro Los Ángeles Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 390 ALA0390 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 391 ALA0391 El Oro Los Ángeles Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 392 ALA0392 El Oro Los Ángeles Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 393 ALA0393 El Oro Los Ángeles Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 394 ALA0394 El Oro Los Ángeles Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 395 ALA0395 El Oro Los Ángeles Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 396 ALA0396 El Oro Los Ángeles Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 397 ALA0397 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 398 ALA0398 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 399 ALA0399 El Oro Los Ángeles Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 400 ALA0400 El Oro Los Ángeles Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 401 ALA0401 El Oro Los Ángeles Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 402 ALA0402 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 403 ALA0403 El Oro Los Ángeles Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 404 ALA0404 El Oro Los Ángeles Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 195

Ny. albimanus/Ny. ALA0405 El Oro Los Ángeles 405 Anopheles spp. rangeli NP NP Ny. albimanus NP Ny. albimanus/Ny. ALA0406 El Oro Los Ángeles 406 Anopheles spp. rangeli NP NP Ny. albimanus NP 407 ALA0407 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 408 ALA0408 El Oro Los Ángeles Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 409 ALA0409 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 410 ALA0410 El Oro Los Ángeles Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 411 ABN0411 El Oro Barbones Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 412 ABN0412 El Oro Barbones Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 413 ABN0413 El Oro Barbones Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP Ny. albimanus/Ny. ABN0414 El Oro Barbones 414 Anopheles spp. oswaldoi NP NP Ny. albimanus NP 415 ABN0415 El Oro Barbones Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP Ny. albimanus/Ny. Ny. ABN0416 El Oro Barbones 416 Anopheles spp. rangeli NP NP Ny. albimanus albimanus 417 ABN0417 El Oro Barbones Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 418 ABN0418 El Oro Barbones Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 419 ABN0419 El Oro Barbones Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 420 ABN0420 El Oro Barbones An. calderoni NI NI NI NI NP 421 ABN0421 El Oro Barbones An. calderoni NI NI NI NI NP 422 ABN0422 El Oro Barbones An. calderoni NI NI NI NI NP 423 ABN0423 El Oro Barbones An. calderoni NI NI NI NI NP An. ABN0424 El Oro Barbones 424 An. calderoni NI calderoni NI An. calderoni NP An. ABN0425 El Oro Barbones 425 An. calderoni NI calderoni NI An. calderoni NP 426 ABN0426 El Oro Barbones An. calderoni NI NI NI NI NP 427 ABN0427 El Oro Barbones An. punctimacula NI NI NI NI NP 196

Ny. albimanus/Ny. ABN0428 El Oro Barbones 428 Anopheles spp. rangeli NP NP Ny. albimanus NP 429 ABN0429 El Oro Barbones Ny. albimanus NP NP NP NP NP 430 ABN0430 El Oro Barbones Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 431 ABN0431 El Oro Barbones Ny. albimanus NP NP NP NP NP 432 ABN0432 El Oro Barbones Ny. albimanus NP NP NP NP NP 433 ABN0433 El Oro Barbones Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP Ny. albimanus/Ny. ABN0434 El Oro Barbones 434 Anopheles spp. strodei NP NP Ny. albimanus NP 435 ABN0435 El Oro Barbones Ny. albimanus NP NP NP NP NP Ny. albimanus/Ny. Ny. ABN0436 El Oro Barbones 436 Anopheles spp. strodei NP NP Ny. albimanus albimanus 437 ABN0437 El Oro Barbones Ny. albimanus NP NP NP NP NP 438 ABN0438 El Oro Barbones Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 439 ABN0439 El Oro Barbones Ny. albimanus NP NP NP NP NP 440 ABN0440 El Oro Barbones Ny. albimanus NP NP NP NP NP 441 ABN0441 El Oro Barbones Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP Ny. albimanus/Ny. ABN0442 El Oro Barbones 442 Anopheles spp. oswaldoi NP NP Ny. albimanus NP 443 ABN0443 El Oro Barbones Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 444 ABN0444 El Oro Barbones Ny. albimanus NP NP NP NP NP 445 ABN0445 El Oro Barbones Ny. albimanus NP NP NP NP NP 446 ABN0446 El Oro Barbones Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 447 ABN0447 El Oro Barbones Ny. albimanus NP NP NP NP NP 448 ABN0448 El Oro Barbones Ny. albimanus NP NP NP NP NP 449 ABN0449 El Oro Barbones Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 450 ABN0450 El Oro Barbones Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 451 ABN0451 El Oro Barbones Ny. albimanus NP NP NP NP NP 452 ABN0452 El Oro Barbones Ny. albimanus NP NP NP NP NP 197

453 ABN0453 El Oro Barbones Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 454 ABN0454 El Oro Barbones Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 455 ABN0455 El Oro Barbones Ny. albimanus NI NP NP NI NP 456 ABN0456 El Oro Barbones Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP Ny. ABN0457 El Oro Barbones NI 457 Anopheles spp. NI NI NI albimanus 458 ABN0458 El Oro Barbones Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 459 ABN0459 El Oro Barbones Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 460 ABN0460 El Oro Barbones Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 461 ABN0461 El Oro Barbones Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 462 ABN0462 El Oro Barbones Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 463 ABN0463 El Oro Barbones Ny. albimanus NP NP NP NP NP 464 ABN0464 El Oro Barbones Ny. albimanus NP NP NP NP NP 465 ABN0465 El Oro Barbones Ny. albimanus NP NP NP NP NP 466 ABN0466 El Oro Barbones Ny. albimanus NP NP NP NP NP 467 ABN0467 El Oro Barbones Ny. albimanus NP NP NP NP NP 468 ABN0468 El Oro Barbones Ny. albimanus NP NP NP NP NP 469 ABN0469 El Oro Barbones Ny. albimanus NP NP NP NP NP 470 ABN0470 El Oro Barbones Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 471 ABN0471 El Oro Barbones Ny. albimanus NP NP NP NP NP 472 ABN0472 El Oro Barbones Ny. albimanus NP NP NP NP NP 473 ABN0473 El Oro Barbones Ny. albimanus NP NP NP NP NP 474 ABN0474 El Oro Barbones Ny. albimanus NP NP NP NP NP 475 ABN0475 El Oro Barbones Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 476 ABN0476 El Oro Barbones Ny. albimanus NP NP NP NP NP 477 ABN0477 El Oro Barbones Ny. albimanus NP NP NP NP NP 478 ABN0478 El Oro Barbones Ny. albimanus NP NP NP NP NP 479 ABN0479 El Oro Barbones Ny. albimanus NP NP NP NP NP 480 ABN0480 El Oro Barbones Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 198

481 ABN0481 El Oro Barbones Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 482 ABN0482 El Oro Barbones Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 483 ABN0483 El Oro Barbones Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 484 AHU0484 El Oro Huaquillas Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 485 AHU0485 El Oro Huaquillas Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 486 AHU0486 El Oro Huaquillas Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 487 AHU0487 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 488 AHU0488 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 489 AHU0489 El Oro Huaquillas Ny. albimanus NP NP NP NP NP Ny. albimanus/Ny. AHU0490 El Oro Huaquillas 490 Anopheles spp. oswaldoi NP NP Ny. albimanus NP 491 AHU0491 El Oro Huaquillas Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 492 AHU0492 El Oro Huaquillas Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 493 AHU0493 El Oro Huaquillas Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 494 AHU0494 El Oro Huaquillas Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 495 AHU0495 El Oro Huaquillas Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 496 AHU0496 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 497 AHU0497 El Oro Huaquillas Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 498 AHU0498 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 499 AHU0499 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 500 AHU0500 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 501 AHU0501 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 502 AHU0502 El Oro Huaquillas Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 503 AHU0503 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 504 AHU0504 El Oro Huaquillas Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 505 AHU0505 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 506 AHU0506 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 507 AHU0507 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 508 AHU0508 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 199

509 AHU0509 El Oro Huaquillas Ny. albimanus NP NP NP NP NP Ny. albimanus/Ny. AHU0510 El Oro Huaquillas Ny. albimanus 510 oswaldoi NP NP Ny. albimanus NP 511 AHU0511 El Oro Huaquillas Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 512 AHU0512 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 513 AHU0513 El Oro Huaquillas Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 514 AHU0514 El Oro Huaquillas Anopheles spp. NI NP NP NI NP 515 AHU0515 El Oro Huaquillas Anopheles spp. NI NP NP NI NP 516 AHU0516 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 517 AHU0517 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 518 AHU0518 El Oro Huaquillas Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 519 AHU0519 El Oro Huaquillas Ny. albimanus NP NP NP NP NP Ny. albimanus/Ny. AHU0520 El Oro Huaquillas Ny. albimanus 520 oswaldoi NP NP Ny. albimanus NP 521 AHU0521 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 522 AHU0522 El Oro Huaquillas Ny. albimanus NP NP NP NP NP Ny. albimanus/Ny. AHU0523 El Oro Huaquillas Ny. albimanus 523 oswaldoi NP NP Ny. albimanus NP 524 AHU0524 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 525 AHU0525 El Oro Huaquillas Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 526 AHU0526 El Oro Huaquillas Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 527 AHU0527 El Oro Huaquillas Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 528 AHU0528 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 529 AHU0529 El Oro Huaquillas Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 530 AHU0530 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 531 AHU0531 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 532 AHU0532 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 533 AHU0533 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 534 AHU0534 El Oro Huaquillas Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 200

535 AHU0535 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 536 AHU0536 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 537 AHU0537 El Oro Huaquillas Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 538 AHU0538 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 539 AHU0539 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 540 AHU0540 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 541 AHU0541 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 542 AHU0542 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 543 AHU0543 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 544 AHU0544 El Oro Huaquillas Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 545 AHU0545 El Oro Huaquillas Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 546 AHU0546 El Oro Huaquillas Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 547 AHU0547 El Oro Huaquillas Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP San José ASJ0548 Sucumbíos NI 548 Wisuya Anopheles spp. NP NP NI NP San José ASJ0549 Sucumbíos 549 Wisuya Ny. nuneztovari Ny. rangeli NP NP Ny. rangeli NP San José Ny. ASJ0550 Sucumbíos 550 Wisuya Anopheles spp. Ny. rangeli NP NP Ny. rangeli rangeli San José ASJ0551 Sucumbíos NI 551 Wisuya Anopheles spp. NP NP NI NP San José ASJ0552 Sucumbíos 552 Wisuya Anopheles spp. Ny. rangeli NI NP Ny. rangeli NP San José ASJ0553 Sucumbíos 553 Wisuya Anopheles spp. Ny. rangeli NP NP Ny. rangeli NP San José ASJ0554 Sucumbíos 554 Wisuya Anopheles spp. Ny. rangeli NP NP Ny. rangeli NP San José ASJ0555 Sucumbíos 555 Wisuya Anopheles spp. Ny. rangeli NI NP Ny. rangeli NP San José ASJ0556 Sucumbíos 556 Wisuya Ny. nuneztovari Ny. rangeli NP NP Ny. rangeli NP 201

San José Ny. strodei/ An. ASJ0557 Sucumbíos 557 Wisuya Anopheles spp. pseudopunctipennis NI NI Ny. strodei NP San José ASJ0558 Sucumbíos 558 Wisuya Ny. oswaldoi Ny. rangeli NP NP Ny. rangeli NP San José ASJ0559 Sucumbíos 559 Wisuya Anopheles spp. Ny. rangeli NP NP Ny. rangeli NP San José ASJ0560 Sucumbíos 560 Wisuya Anopheles spp. NI NP NP NI NP San José Ny. rangeli/Ny. ASJ0561 Sucumbíos 561 Wisuya Anopheles spp. oswaldoi NP NP Ny. rangeli NP San José ASJ0562 Sucumbíos NI 562 Wisuya Anopheles spp. Ny. strodei NI Ny. strodei NP San José ASJ0563 Sucumbíos 563 Wisuya Anopheles spp. Ny. rangeli NP NP Ny. rangeli NP San José ASJ0564 Sucumbíos NI 564 Wisuya Anopheles spp. NI NI NI NP San José ASJ0565 Sucumbíos 565 Wisuya Anopheles spp. Ny. rangeli NP NP Ny. rangeli NP San José ASJ0566 Sucumbíos 566 Wisuya Anopheles spp. NI NP NP NI NP San José ASJ0567 Sucumbíos 567 Wisuya Anopheles spp. NI NP NP NI NP San José ASJ0568 Sucumbíos 568 Wisuya Ny. nuneztovari Ny. rangeli NP NP Ny. rangeli NP San José ASJ0569 Sucumbíos 569 Wisuya Anopheles spp. Ny. rangeli NP NP Ny. rangeli NP 570 APR0570 Sucumbíos Porvenir (106) Anopheles spp. NI NP NP NI NP 571 ALA0571 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 572 ABN0572 El Oro Barbones Ny. albimanus NP NP NP NP NP 573 AWK0573 Orellana Kawymeno Ny. dunhami Ny. strodei NI NI Ny. strodei NP An. ABN0574 El Oro Barbones 574 Anopheles spp. NI NI NI NI calderoni 202

Ny. strodei/ An. APR0575 Sucumbíos Porvenir (106) 575 Ny. dunhami pseudopunctipennis NI NI Ny. strodei NP 576 ALA0576 El Oro Los Ángeles An. calderoni NI NI NI NI NP 577 ASL0577 Sucumbíos Santa Elena Ny. dunhami NI NP NP NI NP 578 AWK0578 Orellana Kawymeno Ny. dunhami Ny. strodei NI NI Ny. strodei NP 579 APR0579 Sucumbíos Porvenir (106) An. calderoni NI NP NP NI NP 580 ALA0580 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP Ny. albimanus/Ny. ABN0581 El Oro Barbones 581 Anopheles spp. rangeli NP NP Ny. albimanus NP 582 ALA0582 El Oro Los Ángeles Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 583 ALA0583 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 584 ALA0584 El Oro Los Ángeles Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP Ny. ALA0585 El Oro Los Ángeles 585 Anopheles spp. Ny. albimanus NP NP Ny. albimanus albimanus 586 ALA0586 El Oro Los Ángeles Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 587 ALA0587 El Oro Los Ángeles Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 588 ALA0588 El Oro Los Ángeles Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP Ny. albimanus/Ny. ALA0589 El Oro Los Ángeles 589 Anopheles spp. rangeli NP NP Ny. albimanus NP 590 ALA0590 El Oro Los Ángeles Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 591 ALA0591 El Oro Los Ángeles Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 592 APA0592 Sucumbíos Palmar Anopheles spp. Ny. rangeli NP NP Ny. rangeli NP 593 APA0593 Sucumbíos Palmar Ny. nuneztovari Ny. rangeli NI NP Ny. rangeli NI 594 APA0594 Sucumbíos Palmar Anopheles spp. Ny. rangeli NP NP Ny. rangeli NP 595 APA0595 Sucumbíos Palmar Anopheles spp. Ny. rangeli NP NP Ny. rangeli NP 596 APA0596 Sucumbíos Palmar Ny. oswaldoi Ny. rangeli NP NP Ny. rangeli NP 597 APA0597 Sucumbíos Palmar Anopheles spp. NI NI NI NI NP 598 APA0598 Sucumbíos Palmar Ny. oswaldoi Ny. rangeli NI NP Ny. rangeli NI 599 APA0599 Sucumbíos Palmar Ny. albimanus Ny. rangeli NI NP Ny. rangeli NP 203

600 APA0600 Sucumbíos Palmar Anopheles spp. Ny. strodei NI NI Ny. strodei NP 601 APA0601 Sucumbíos Palmar Anopheles spp. NI NP NP NI NP 602 APA0602 Sucumbíos Palmar Anopheles spp. Ny. rangeli NP NP Ny. rangeli NP 603 APA0603 Sucumbíos Palmar Anopheles spp. Ny. rangeli NP NP Ny. rangeli NP 604 APA0604 Sucumbíos Palmar Ny. oswaldoi Ny. rangeli NP NP Ny. rangeli NP Ny. strodei/ An. APA0605 Sucumbíos Palmar 605 Anopheles spp. pseudopunctipennis NI NI Ny. strodei NP 606 APA0606 Sucumbíos Palmar Anopheles spp. NI NP NP NI NP Ny. APA0607 Sucumbíos Palmar 607 Anopheles spp. NI NP NP NI rangeli Bocona ABL0608 Sucumbíos 608 Cuembí Anopheles spp. Ny. rangeli NP NP Ny. rangeli NP Bocona ABL0609 Sucumbíos NI 609 Cuembí Anopheles spp. NP NP NI NP Bocona ABL0610 Sucumbíos 610 Cuembí Anopheles spp. Ny. rangeli NP NP Ny. rangeli NP Bocona ABL0611 Sucumbíos 611 Cuembí Anopheles spp. NI NI NP NI NP Bocona ABL0612 Sucumbíos 612 Cuembí Anopheles spp. NI NP NP NI NP Bocona ABL0613 Sucumbíos 613 Cuembí Anopheles spp. Ny. rangeli NP NP Ny. rangeli NP Bocona ABL0614 Sucumbíos 614 Cuembí Anopheles spp. Ny. rangeli NP NP Ny. rangeli NP Bocona ABL0615 Sucumbíos 615 Cuembí Anopheles spp. NI NI NI NI NP Bocona ABL0616 Sucumbíos 616 Cuembí Anopheles spp. Ny. rangeli NP NP Ny. rangeli NP Bocona ABL0617 Sucumbíos 617 Cuembí Anopheles spp. Ny. rangeli NP NP Ny. rangeli NP Bocona ABL0618 Sucumbíos 618 Cuembí Anopheles spp. NI NP NP NI NP 204

Bocona ABL0619 Sucumbíos 619 Cuembí Anopheles spp. Ny. rangeli NP NP Ny. rangeli NP Bocona ABL0620 Sucumbíos 620 Cuembí Anopheles spp. Ny. rangeli NP NP Ny. rangeli NP Bocona ABL0621 Sucumbíos 621 Cuembí Anopheles spp. Ny. rangeli NP NP Ny. rangeli NP Bocona Ny. Ny. strodei/Ny. ABL0622 Sucumbíos 622 Cuembí Anopheles spp. Ny. strodei NI benarrochi benarrochi NP Bocona ABL0623 Sucumbíos 623 Cuembí Anopheles spp. Ny. rangeli NP NP Ny. rangeli NP Bocona ABL0624 Sucumbíos 624 Cuembí Anopheles spp. Ny. rangeli NP NP Ny. rangeli NP Bocona ABL0625 Sucumbíos 625 Cuembí Anopheles spp. Ny. rangeli NP NP Ny. rangeli NP Bocona ABL0626 Sucumbíos 626 Cuembí Ny. nuneztovari Ny. rangeli NP NP Ny. rangeli NP Bocona ABL0627 Sucumbíos 627 Cuembí Anopheles spp. Ny. rangeli NP NP Ny. rangeli NP Bocona ABL0628 Sucumbíos 628 Cuembí Anopheles spp. Ny. rangeli NP NP Ny. rangeli NP Bocona ABL0629 Sucumbíos 629 Cuembí Ny. oswaldoi Ny. rangeli NP NP Ny. rangeli NP Bocona ABL0630 Sucumbíos 630 Cuembí Anopheles spp. Ny. rangeli NP NP Ny. rangeli NP Bocona ABL0631 Sucumbíos 631 Cuembí Anopheles spp. Ny. rangeli NP NP Ny. rangeli NP Bocona ABL0632 Sucumbíos 632 Cuembí Anopheles spp. Ny. rangeli NP NP Ny. rangeli NP Bocona ABL0633 Sucumbíos 633 Cuembí Ny. nuneztovari Ny. rangeli NP NP Ny. rangeli NP Bocona ABL0634 Sucumbíos 634 Cuembí Anopheles spp. Ny. rangeli NI NP Ny. rangeli NP Bocona ABL0635 Sucumbíos 635 Cuembí Anopheles spp. Ny. rangeli NP NP Ny. rangeli NP 205

Bocona ABL0636 Sucumbíos 636 Cuembí Anopheles spp. Ny. rangeli NP NP Ny. rangeli NP Bocona ABL0637 Sucumbíos 637 Cuembí Ny. oswaldoi Ny. rangeli NP NP Ny. rangeli NP Bocona ABL0638 Sucumbíos 638 Cuembí Anopheles spp. Ny. rangeli NP NP Ny. rangeli NP Bocona Ny. strodei/ An. Ny. Ny. strodei/Ny. ABL0639 Sucumbíos 639 Cuembí Anopheles spp. pseudopunctipennis NI benarrochi benarrochi NP 640 ALA0640 El Oro Los Ángeles Anopheles spp. NI NP NP NI NP 641 ALA0641 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP An. ABN0642 El Oro Barbones NI 642 An. calderoni NI calderoni An. calderoni NP 643 ABN0643 El Oro Barbones Ny. albimanus NP NP NP NP NP 644 AHU0644 El Oro Huaquillas Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 645 AHU0645 El Oro Huaquillas Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 646 ABN0646 El Oro Barbones Ny. albimanus NP NP NP NP NP An. ALA0647 El Oro Los Ángeles NI 647 An. calderoni NI calderoni An. calderoni NP 648 ALA0648 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 649 ABN0649 El Oro Barbones Ny. albimanus NP NP NP NP NP An. ALA0650 El Oro Los Ángeles NI 650 An. calderoni NI calderoni An. calderoni NP An. ABN0651 El Oro Barbones Anopheles spp. 651 Ny. strodei calderoni NI An. calderoni NP 652 ALA0652 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 653 ALA0653 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 654 ALA0654 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP An. ABN0655 El Oro Barbones 655 An. calderoni NI calderoni NI An. calderoni NP 656 ALA0656 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 657 ABN0657 El Oro Barbones Ny. albimanus NP NP NP NP NP 206

658 AHU0658 El Oro Huaquillas Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 659 ABN0659 El Oro Barbones Ny. albimanus NP NP NP NP NP An. ABN0660 El Oro Barbones 660 An. calderoni NI calderoni NI An. calderoni NP 661 ABN0661 El Oro Barbones Anopheles spp. NA NA NA NA NP 662 ALA0662 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 663 ALA0663 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP An. ALA0664 El Oro Los Ángeles 664 An. calderoni NI calderoni NI An. calderoni NP 665 ABN0665 El Oro Barbones Ny. albimanus NP NP NP NP NP An. ALA0666 El Oro Los Ángeles 666 An. calderoni NI calderoni NI An. calderoni NP 667 ALA0667 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 668 ABN0668 El Oro Barbones Ny. albimanus NP NP NP NP NP An. ABN0669 El Oro Barbones 669 An. calderoni NI NI NI NI calderoni 670 ABN0670 El Oro Barbones Ny. albimanus NP NP NP NP NP 671 ABN0671 El Oro Barbones Ny. albimanus NP NP NP NP NP 672 ALA0672 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 673 ALA0673 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 674 ALA0674 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 675 ABN0675 El Oro Barbones An. calderoni NI NI NI NI NP 676 ABN0676 El Oro Barbones Ny. albimanus NP NP NP NP NP 677 ALA0677 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 678 ABN0678 El Oro Barbones An. calderoni NI NI NI NI NP 679 ABN0679 El Oro Barbones Ny. albimanus NP NP NP NP NP 680 AHU0680 El Oro Huaquillas Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 681 ALA0681 El Oro Huaquillas Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 682 AHU0682 El Oro Huaquillas Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 207

An. AHU0683 El Oro Huaquillas 683 An. calderoni NI calderoni NI An. calderoni NP 684 AHU0684 El Oro Huaquillas Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 685 ABN0685 El Oro Barbones Ny. albimanus NP NP NP NP NP 686 AHU0686 El Oro Huaquillas Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 687 AHU0687 El Oro Huaquillas Anopheles spp. Ny. albimanus NP NP Ny. albimanus NP 688 ALA0688 El Oro Los Ángeles Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 689 ALA0689 El Oro Los Ángeles Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP Ny. albimanus/Ny. ALA0690 El Oro Ny. albimanus 690 Los Ángeles oswaldoi NP NP Ny. albimanus NP 691 ALA0691 El Oro Los Ángeles Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 692 ALA0692 El Oro Los Ángeles Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 693 ALA0693 El Oro Los Ángeles Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 694 ALA0694 El Oro Los Ángeles Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 695 ALA0695 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 696 ALA0696 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP Ny. albimanus/Ny. ALA0697 El Oro Ny. albimanus 697 Los Ángeles oswaldoi NP NP Ny. albimanus NP 698 ALA0698 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 699 ALA0699 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 700 ALA0700 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 701 ALA0701 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 702 ALA0702 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 703 ALA0703 El Oro Los Ángeles Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 704 ALA0704 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 705 ALA0705 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 706 ALA0706 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 707 ALA0707 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 708 ALA0708 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 208

709 ALA0709 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 710 ALA0710 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 711 ALA0711 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 712 ALA0712 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 713 ALA0713 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 714 ALA0714 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 715 ALA0715 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 716 ALA0716 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 717 ALA0717 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 718 ALA0718 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 719 ALA0719 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 720 ALA0720 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 721 ALA0721 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 722 ALA0722 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 723 ALA0723 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 724 ALA0724 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 725 ALA0725 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 726 ALA0726 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 727 ALA0727 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 728 ALA0728 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 729 ALA0729 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 730 ALA0730 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 731 ALA0731 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 732 ALA0732 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 733 ALA0733 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 734 ALA0734 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 735 ALA0735 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 736 ALA0736 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 209

737 ALA0737 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 738 ALA0738 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 739 ALA0739 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 740 ALA0740 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 741 ALA0741 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP Ny. albimanus/Ny. ALA0742 El Oro Ny. albimanus 742 Los Ángeles rangeli NP NP Ny. albimanus NP 743 ALA0743 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 744 ALA0744 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 745 ALA0745 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 746 ALA0746 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP Ny. albimanus/Ny. ALA0747 El Oro Ny. albimanus 747 Los Ángeles strodei NP NP Ny. albimanus NP 748 ALA0748 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 749 ALA0749 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 750 ALA0750 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 751 ALA0751 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 752 ALA0752 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 753 ALA0753 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 754 ALA0754 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 755 ALA0755 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 756 ALA0756 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 757 ALA0757 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 758 ALA0758 El Oro Los Ángeles Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 759 ALA0759 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 760 ALA0760 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 761 ALA0761 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 762 ALA0762 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 763 ALA0763 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 210

764 ALA0764 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 765 ALA0765 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 766 ALA0766 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 767 ALA0767 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP Ny. albimanus/Ny. ALA0768 El Oro Ny. albimanus 768 Los Ángeles oswaldoi NP NP Ny. albimanus NP 769 ALA0769 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 770 ALA0770 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 771 ALA0771 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 772 ALA0772 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 773 ALA0773 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 774 ALA0774 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 775 ALA0775 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 776 ALA0776 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 777 ALA0777 El Oro Los Ángeles Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 778 ALA0778 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 779 ALA0779 El Oro Los Ángeles Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 780 ALA0780 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 781 ALA0781 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 782 ALA0782 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 783 ALA0783 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 784 ALA0784 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 785 ALA0785 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 786 ALA0786 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 787 ALA0787 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 788 ALA0788 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 789 ALA0789 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 790 ALA0790 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 791 ALA0791 El Oro Los Ángeles Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 211

Ny. albimanus/Ny. ALA0792 El Oro Ny. albimanus 792 Los Ángeles oswaldoi NP NP Ny. albimanus NP 793 ALA0793 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 794 ALA0794 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 795 ALA0795 El Oro Los Ángeles Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 796 ALA0796 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 797 ALA0797 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 798 ALA0798 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 799 ALA0799 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 800 ALA0800 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 801 ALA0801 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 802 ALA0802 El Oro Los Ángeles Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 803 ALA0803 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 804 ALA0804 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 805 ALA0805 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 806 ALA0806 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 807 ALA0807 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 808 ALA0808 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 809 ALA0809 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 810 ALA0810 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 811 ALA0811 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 812 ALA0812 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 813 ALA0813 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 814 ALA0814 El Oro Los Ángeles Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 815 ALA0815 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 816 ALA0816 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 817 ALA0817 El Oro Los Ángeles Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 818 ALA0818 El Oro Los Ángeles Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 819 ALA0819 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 212

Ny. albimanus/Ny. ALA0820 El Oro Ny. albimanus 820 Los Ángeles rangeli NP NP Ny. albimanus NP Ny. albimanus/Ny. ALA0821 El Oro Ny. albimanus 821 Los Ángeles oswaldoi NP NP Ny. albimanus NP Ny. albimanus/Ny. ALA0822 El Oro Ny. albimanus 822 Los Ángeles oswaldoi NP NP Ny. albimanus NP 823 ALA0823 El Oro Los Ángeles Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 824 ALA0824 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP Ny. albimanus/Ny. ALA0825 El Oro Ny. albimanus 825 Los Ángeles oswaldoi NP NP Ny. albimanus NP 826 ALA0826 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 827 ALA0827 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 828 ALA0828 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 829 ALA0829 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 830 ALA0830 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 831 ALA0831 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP Ny. albimanus/Ny. ALA0832 El Oro Ny. albimanus 832 Los Ángeles oswaldoi NP NP Ny. albimanus NP Ny. albimanus/Ny. ALA0833 El Oro Ny. albimanus 833 Los Ángeles oswaldoi NP NP Ny. albimanus NP Ny. albimanus/Ny. ALA0834 El Oro Ny. albimanus 834 Los Ángeles oswaldoi NP NP Ny. albimanus NP 835 ALA0835 El Oro Los Ángeles Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 836 ALA0836 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 837 ALA0837 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP Ny. albimanus/Ny. ALA0838 El Oro Ny. albimanus 838 Los Ángeles oswaldoi NP NP Ny. albimanus NP 839 ALA0839 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 840 ALA0840 El Oro Los Ángeles Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 841 ALA0841 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 213

842 ALA0842 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 843 ALA0843 El Oro Los Ángeles An. calderoni NI NI NI NI NP 844 ALA0844 El Oro Los Ángeles An. calderoni NI NI NI NI NP An. Ny. albimanus/Ny. ALA0845 El Oro 845 Los Ángeles pseudopunctipennis oswaldoi NP NP Ny. albimanus NP 846 ALA0846 El Oro Los Ángeles An. calderoni NI NI NI NI NP 847 ALA0847 El Oro Los Ángeles Ny. albimanus NI NI NI NI NP 848 ALA0848 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP Ny. albimanus/Ny. ALA0849 El Oro Ny. albimanus 849 Los Ángeles oswaldoi NP NP Ny. albimanus NP 850 ALA0850 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 851 ALA0851 El Oro Los Ángeles Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 852 ALA0852 El Oro Los Ángeles Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 853 ALA0853 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 854 ALA0854 El Oro Los Ángeles Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 855 ALA0855 El Oro Los Ángeles Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 856 ALA0856 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 857 ALA0857 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 858 ALA0858 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 859 ALA0859 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 860 ALA0860 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 861 ALA0861 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 862 ALA0862 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 863 ALA0863 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 864 ALA0864 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 865 ALA0865 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 866 ALA0866 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 867 ALA0867 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 868 ALA0868 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 214

869 ALA0869 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 870 ALA0870 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 871 ALA0871 El Oro Los Ángeles Ny. albimanus NP NP NP NP NP 872 AHU0872 El Oro Huaquillas Ny. albimanus NP NP NP NP NP Ny. albimanus/Ny. AHU0873 El Oro Ny. albimanus 873 Huaquillas oswaldoi NP NP Ny. albimanus NP Ny. albimanus/Ny. AHU0874 El Oro Ny. albimanus 874 Huaquillas oswaldoi NP NP Ny. albimanus NP 875 AHU0875 El Oro Huaquillas Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 876 AHU0876 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 877 AHU0877 El Oro Huaquillas Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 878 AHU0878 El Oro Huaquillas Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP Ny. albimanus/Ny. AHU0879 El Oro Ny. albimanus 879 Huaquillas oswaldoi NP NP Ny. albimanus NP 880 AHU0880 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 881 AHU0881 El Oro Huaquillas Ny. albimanus NP NP NP NP NP Ny. albimanus/Ny. AHU0882 El Oro Ny. albimanus 882 Huaquillas rangeli NP NP Ny. albimanus NP Ny. albimanus/Ny. AHU0883 El Oro Ny. albimanus 883 Huaquillas oswaldoi NP NP Ny. albimanus NP 884 AHU0884 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 885 AHU0885 El Oro Huaquillas Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 886 AHU0886 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 887 AHU0887 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 888 AHU0888 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 889 AHU0889 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 890 AHU0890 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 891 AHU0891 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 892 AHU0892 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 893 AHU0893 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 215

894 AHU0894 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 895 AHU0895 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 896 AHU0896 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 897 AHU0897 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 898 AHU0898 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 899 AHU0899 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 900 AHU0900 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 901 AHU0901 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 902 AHU0902 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 903 AHU0903 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 904 AHU0904 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 905 AHU0905 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 906 AHU0906 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 907 AHU0907 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 908 AHU0908 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 909 AHU0909 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 910 AHU0910 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 911 AHU0911 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 912 AHU0912 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 913 AHU0913 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 914 AHU0914 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 915 AHU0915 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 916 AHU0916 El Oro Huaquillas Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 917 AHU0917 El Oro Huaquillas Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 918 AHU0918 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 919 AHU0919 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 920 AHU0920 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 921 AHU0921 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 216

922 AHU0922 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 923 AHU0923 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 924 AHU0924 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 925 AHU0925 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 926 AHU0926 El Oro Huaquillas Ny. albimanus NP NP NP NP NP 927 AHU0927 El Oro Huaquillas Ny. albimanus Ny. albimanus NP NP Ny. albimanus NP 928 AHU0928 El Oro Huaquillas An. calderoni NI NI NI NI NI Ny. albimanus/Ny. AHU0929 El Oro Ny. albimanus 929 Huaquillas rangeli NP NP Ny. albimanus NP An. AHU0930 El Oro 930 Huaquillas An. calderoni NI calderoni NI An. calderoni NP An. AHU0931 El Oro 931 Huaquillas An. calderoni NI calderoni NI An. calderoni NP An. AHU0932 El Oro 932 Huaquillas An. calderoni NI calderoni NI An. calderoni NP 933 AHU0933 El Oro Huaquillas Ny. albimanus NP NP NP NP NP An. AHU0934 El Oro 934 Huaquillas An. calderoni NI calderoni NI An. calderoni NP An. AHU0935 El Oro 935 Huaquillas An. calderoni NI calderoni NI An. calderoni NP An. An. AHU0936 El Oro 936 Huaquillas An. calderoni NI calderoni NI An. calderoni calderoni An. AHU0937 El Oro 937 Huaquillas An. calderoni NI calderoni NI An. calderoni NP An. AHU0938 El Oro NI 938 Huaquillas An. calderoni NI calderoni An. calderoni NP An. AHU0939 El Oro NI 939 Huaquillas An. calderoni NI calderoni An. calderoni NP An. AHU0940 El Oro NI 940 Huaquillas An. calderoni NI calderoni An. calderoni NP An. AHU0941 El Oro Anopheles spp. 941 Huaquillas NI calderoni NI An. calderoni NP 217

An. AHU0942 El Oro NI 942 Huaquillas An. calderoni NI calderoni An. calderoni NP An. AHU0943 El Oro NI 943 Huaquillas An. calderoni NI calderoni An. calderoni NP 944 AHU0944 El Oro Huaquillas Ny. albimanus NI NI NP NI NP An. An. AHU0945 El Oro 945 Huaquillas An. calderoni NI calderoni calderoni An. calderoni NP An. An. AHU0946 El Oro 946 Huaquillas An. calderoni NI calderoni calderoni An. calderoni NP An. An. AHU0947 El Oro 947 Huaquillas An. calderoni NI calderoni calderoni An. calderoni NP An. An. AHU0948 El Oro 948 Huaquillas An. calderoni NI calderoni calderoni An. calderoni NP An. AHU0949 El Oro 949 Huaquillas An. calderoni NI calderoni NI An. calderoni NP 950 AHU0950 El Oro Huaquillas An. calderoni NI NI NI NI NP 951 AHU0951 El Oro Huaquillas An. calderoni NI NI NI NI NP An. AHU0952 El Oro 952 Huaquillas An. calderoni NI calderoni NI An. calderoni NP Ny. albimanus/Ny. AHU0953 El Oro Ny. albimanus 953 Huaquillas oswaldoi NP NP Ny. albimanus NP An. AHU0954 El Oro 954 Huaquillas An. calderoni NI calderoni NI An. calderoni NP An. AHU0955 El Oro 955 Huaquillas An. calderoni NI calderoni NI An. calderoni NP An. AHU0956 El Oro 956 Huaquillas An. calderoni NI calderoni NI An. calderoni NP Ny. albimanus/Ny. AHU0957 El Oro Ny. albimanus 957 Huaquillas oswaldoi NP NP Ny. albimanus NP An. AHU0958 El Oro 958 Huaquillas An. calderoni NI calderoni NI An. calderoni NP An. AHU0959 El Oro 959 Huaquillas An. calderoni NI calderoni NI An. calderoni NP 218

NI: non-identified NP: non-processed with the restriction enzyme NA: did not amplified a PCR product

219

Appendix F. Identification of mosquitoes by BLAST analysis of the sequences obtained for COI amplification products.

%ID NCBI BLAST ID - Date: 05 Nov Accession Code NCBI Author Country: Locality 2019 Number BLAST

ABN0574 Anopheles calderoni 100 MG701368.1 Alvares et al. Colombia: Tumaco. Nariño ABR0268 - - - - - Ecuador: Guayas. Guayaquil. Yaguachi. Hacienda. AHU0936 Anopheles calderoni 99 HQ642974 González et al. Eulalia

ALA0357 Anopheles albimanus 96 KC354823.1 Herrera et al. Colombia: La Guajira

ALA0585 Anopheles albimanus 96 KC354823.1 Herrera et al. Colombia: La Guajira ALA0593 - - - - - APA0598 - - - - -

APR0315 Anopheles aff. konderi 100 MF381585.1 Foster et al. 2018 Brazil AWK0005 - - - - - AWK0058 - - - - - AWK0064 - - - - -

AWK0083 Anopheles benarrochi B 100% JX205097.1 Orjuela et al. 2012 Colombia: Putumayo

ABR0282 Anopheles benarrochi B 100% JX205097.1 Orjuela et al. 2012 Colombia: Putumayo

AWK0078 Anopheles benarrochi B 97% JX205097.1 Orjuela et al. 2012 Colombia: Putumayo Atiencia-Pineda et

APR0304 Anopheles oswaldoi B 99% MF598468.1 al 2017 Colombia: Cordoba. Montelibano.Puerto Anchica Atiencia-Pineda et

APR0316 Anopheles oswaldoi B 99% MF598468.1 al 2017 Colombia: Cordoba. Montelibano.Puerto Anchica

AWK0056 Anopheles benarrochi B 99% JX205100.1 Orjuela et al. 2012 Colombia: Putumayo

AWK0079 Anopheles benarrochi B 99% JX205097.1 Orjuela et al. 2012 Colombia: Putumayo ABN0178 Anopheles albimanus 96% KC354825 Herrera et al. Colombia: La Guajira ABN0457 Anopheles albimanus 97% KC354825 Herrera et al. Colombia: La Guajira 220

AHU0928 - - - - - APA0607 Anopheles rangeli 99% JX205120 Orjuela et al. 2012 Colombia: Putumayo Ruiz-Lopez et al. Ecuador: Orellana. Coca. Ecuador. Orellana. Coca. APR0312 Anopheles oswaldoi B 99% KF809103 2013 Guamayacu APR0314 Anopheles rangeli 99% HM022394 Ahumada et al. Colombia: Meta

AWK0022 Anopheles benarrochi B 99% JX205097.1 Orjuela et al. 2012 Colombia: Putumayo

AWK0084 Anopheles benarrochi B 99% JX205097.1 Orjuela et al. 2012 Colombia: Putumayo

AWK0090 Anopheles benarrochi B 99% JX205097.1 Orjuela et al. 2012 Colombia: Putumayo Ahumada et al.

ABN0416 Anopheles albimanus 99% KU900789.1 2016 Colombia: Nariño Ahumada et al.

ABN0436 Anopheles albimanus 99% KU900794.1 2016 Colombia: Nariño

AWK0094 Anopheles benarrochi B 99% JX205097.1 Orjuela et al. 2012 Colombia: Putumayo

AWK0007 Anopheles benarrochi B 100% JX205097.1 Orjuela et al. 2012 Colombia: Putumayo Ahumada et al.

ASL0181 Anopheles rangeli 100% HM022394.1 2016 Colombia Ruiz-Lopez et al. Ecuador: Orellana. Coca. Ecuador. Orellana. Coca.

ANM0222 Anopheles oswaldoi B 99% KF809099.1 2013 Guamayacu González et al. Ecuador: Guayas. Guayaquil. Yaguachi. Hacienda. HQ642971.1 ABN0669 Anpheles calderoni 99% 2016 Eulalia Ahumada et al.

ASJ0550 Anopheles rangeli 99% HM022394.1 2016 Colombia

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