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Diversity, Abundance and Prevalence of Medically Important Pathogens of Mosquitoes Caught During the Dry Winter Season in Madagascar

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Diversity, Abundance and Prevalence of Medically Important Pathogens of Mosquitoes Caught During the Dry Winter Season in Madagascar MSc Project Report 2015-2016 Diversity, abundance and prevalence of medically important pathogens of mosquitoes caught during the dry winter season in Madagascar Supervisor: Dr. Thomas Walker Candidate number: 109210 Word count: 9264 words Project length: Standard Submitted in part fulfilment of the requirements for the degree of MSc in Medical Entomology for Disease Control September 2016 iii 1 TABLE OF CONTENTS 2 Abstract ............................................................................................................................... v 3 Acknowledgements ............................................................................................................. vi 3.1 Acknowledgement of Academic Support ...................................................................... vi 3.2 Acknowledgement of Other Support ........................................................................... vii 1 Introduction ......................................................................................................................... 1 1.1 Mosquito-Borne Diseases ............................................................................................ 1 1.2 Malaria ......................................................................................................................... 1 1.3 Arboviruses .................................................................................................................. 5 1.4 Madagascar ................................................................................................................. 6 1.5 Malagasy Mosquitoes .................................................................................................. 8 2 Aims and objectives ...........................................................................................................10 3 Hypothesis .........................................................................................................................10 4 Methodology ......................................................................................................................11 4.1 Madagascar Locations ................................................................................................12 4.1.1 Rationale for Site Selection ..................................................................................13 4.2 Vector sampling ..........................................................................................................13 4.3 Identification, Storage and Transportation ...................................................................17 4.4 Laboratory Screening ..................................................................................................18 4.4.1 Sampling and Pooling Strategy ............................................................................18 4.4.2 RNA Extraction and cDNA Production .................................................................19 iv 4.4.3 PCR Assays ........................................................................................................20 5 Results ...............................................................................................................................23 5.1 Temperature Data and Humidity Data .........................................................................23 5.2 Qualitative Assessment of Sites ..................................................................................26 5.3 Overall Mosquito Abundance ......................................................................................26 5.4 Species Diversity ........................................................................................................30 5.5 Species Screened, Number of Individuals and Pool Sizes ..........................................33 5.6 PCR Analysis ..............................................................................................................33 6 Discussion .........................................................................................................................38 6.1 Mosquito Abundance ..................................................................................................38 6.2 Vector Diversity ...........................................................................................................39 6.3 Pathogen Screening ...................................................................................................40 6.4 Limitations ..................................................................................................................41 6.5 Summary ....................................................................................................................42 7 Recommendations .............................................................................................................42 8 References ........................................................................................................................43 9 Appendix ............................................................................................................................46 v 2 ABSTRACT 2.1 INTRODUCTION Mosquitoes are responsible for significant human and animal disease through transmission of pathogens such as malaria and arboviruses. Outbreaks of malaria and arboviral diseases in Madagascar have typically been confined to the wet season when mosquito abundance is highest. However, there have been limited studies looking at mosquito abundance, activity or role in disease transmission in the dry season in Madagascar. 2.2 METHODS Mosquitoes were sampled using eight CDC light traps and one zebu-baited trap from five sites for two nights each across Madagascar. Traps were selected to sample a variety of night-biting mosquitoes. Caught individuals underwent morphological identification and correct storage for transport and further laboratory analysis. Non-bloodfed females of varying species were selected for RNA extraction and reverse transcription to produce cDNA for PCR analysis according to abundance and previous roles in disease transmission. cDNA underwent real-time PCR assays for malaria parasites and a range of arboviruses. 2.3 RESULTS In total, 2051 mosquitoes were caught with 2050 identified. 29 different species were identified within 6 different mosquito genera. 18 caught species were previously identified as disease vectors. PCR analysis revealed two Culex decens positive for Rift Valley fever virus and seven Anopheles spp. positive for Plasmodium falciparum. 2.4 CONCLUSIONS These results indicate the possibility of disease transmission during the dry season given the abundance of vector species and evidence of pathogen infections within caught females. Further research is needed to determine the importance of different vector species in dry season transmission to Madagascar transmission dynamics. vi 3 ACKNOWLEDGEMENTS 3.1 ACKNOWLEDGEMENT OF ACADEMIC SUPPORT 3.1.1.1 Project Development The project idea originally came from my supervisor’s interest in collaborating on a project with Institut Pasteur de Madagascar (IPM). The project was generally developed by myself, my supervisor and his PhD student while details regarding field work were decided on in collaboration with IPM. My supervisor provided information on which PCRs would be possible to perform in the laboratory while I made the final decision on which to perform. Pooling strategies for PCRs were designed by myself with input by both my supervisor and his PhD student. 3.1.2 Contact, Input and Support I had an initial meeting with my supervisor in order to discuss general proposals for projects. There were two further meetings, one of which was also attended by my supervisor’s PhD student for detailed planning of this project. There were several email exchanges to discuss administrative issues. Alongside this there was email communication with IPM regarding field work. Before travel there was a meeting with my supervisor and his PhD student to review objectives, ensure I had all required equipment and discuss contact details while in the field. 3.1.3 Main Research Work On arrival in Madagascar I met with Dr. Sebastien Boyer of IPM to finalize field sites. I determined trap placement locations at each site with assistance by Dr. Fara Raharimalala and Dr. Luciano Tantely of IPM. Both assisted in all aspects of primary data collection in the field. I maintained sporadic contact with my supervisor. On return to London, while I performed the laboratory work myself I was assisted in high-cost or skill-dependent steps by my supervisor and his PhD student. 3.1.4 Writing-Up My supervisor read one initial draft and provided suggestions focusing on structure and corrections on grammar, spelling, incorrect referencing and minor comments on content. A final draft was submitted to ensure that the report was fit for submission (checking spelling, title pages, table of contents, referencing, word count and anonymity). vii 3.2 ACKNOWLEDGEMENT OF OTHER SUPPORT I am very grateful to both Dr. Luciano Tantely and Dr. Fara Raharimalala for their assistance in the practicalities of field work. Additionally, I would like to thank Dr. Sebastien Boyer and the rest of the management at IPM for their help throughout the project. I would also like to thank the military personnel, local government, and village chiefs who assisted me in the field. I am grateful to the London School of Hygiene and Tropical Medicine and Bayer Pharmaceuticals for providing me with funding without which I would have been unable to undertake this project
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