Ecology and morphology of mouse lemurs ( Microcebus spp.) in a hotspot of microendemism in northeastern Madagascar, with the description of a new species Dominik Schüßler, Marina Blanco, Jordi Salmona, Jelmer Poelstra, Jean Andriambeloson, Alex Miller, Blanchard Randrianambinina, David Rasolofoson, Jasmin Mantilla-contreras, Lounès Chikhi, et al.

To cite this version:

Dominik Schüßler, Marina Blanco, Jordi Salmona, Jelmer Poelstra, Jean Andriambeloson, et al.. Ecology and morphology of mouse lemurs ( Microcebus spp.) in a hotspot of microendemism in northeastern Madagascar, with the description of a new species. American Journal of Primatology, Wiley, 2020, 82 (9), pp.e23180. ￿10.1002/ajp.23180￿. ￿hal-03026869￿

HAL Id: hal-03026869 https://hal.archives-ouvertes.fr/hal-03026869 Submitted on 5 Jan 2021

HAL is a multi-disciplinary open access L’archive ouverte pluridisciplinaire HAL, est archive for the deposit and dissemination of sci- destinée au dépôt et à la diffusion de documents entific research documents, whether they are pub- scientifiques de niveau recherche, publiés ou non, lished or not. The documents may come from émanant des établissements d’enseignement et de teaching and research institutions in France or recherche français ou étrangers, des laboratoires abroad, or from public or private research centers. publics ou privés. American Journal of Primatology

Ecology and morphology of mouse lemurs (Microcebus spp.) in a hotspot of microendemism in northeastern Madagascar, with the description of a new species

Journal: American Journal of Primatology

Manuscript ID AJP-20-0048.R1

Wiley - Manuscript type:ForResearch Peer Article Review

Date Submitted by the n/a Author:

Complete List of Authors: Schüßler, Dominik; University of Hildesheim, Institute of Biology and Chemistry Blanco, Marina; Duke Lemur Center; Duke University, Department of Biology Salmona, Jordi; Laboratoire Evolution & Diversité Biologique, UMR 5174 CNRS – Université Paul Sabatier Poelstra, Jelmer; Duke University, Department of Biology Andriambeloson, Jean-Basile; University of Antananarivo, Department of Zoology and Animal Biodiversity Miller, Alex; Instituto Gulbenkian de Ciencia, Population and Conservation Genetics Group Randrianambinina, Blanchard; Groupe d’Etude et de Recherche sur les Primates de Madagascar (GERP); University of Mahajanga, Faculté des Sciences Rasolofoson, David; Groupe d’Etude et de Recherche sur les Primates de Madagascar (GERP) Mantilla-Contreras, Jasmin; University of Hildesheim, Institute of Biology and Chemistry Chikhi, Lounès; Instituto Gulbenkian de Ciênca, ; Laboratoire Evolution & Diversité Biologique, UMR 5174 CNRS – Université Paul Sabatier, Lewis, Edward; Omaha’s Henry Doorly Zoo and Aquarium, Grewcock Center for Conservation and Research Yoder, Anne; Duke University, Department of Biology Radespiel, Ute; University of Veterinary Medicine, Institute of Zoology

Indicate which taxonomic group was the subject of your Prosimians study (select all that apply or type another option)::

Keywords: evolution, cryptic species, phenotype, habitat use, sympatry

John Wiley & Sons Page 1 of 66 American Journal of Primatology

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 For Peer Review 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60 John Wiley & Sons American Journal of Primatology Page 2 of 66

Schüßler et al. page 1 1 2 3 4 5 1 Ecology and morphology of mouse lemurs (Microcebus spp.) in 6 7 2 a hotspot of microendemism in northeastern Madagascar, with 8 9 10 3 the description of a new species 11 12 13 4 14 15 16 5 Authors: 17 18 19 6 Dominik Schüßler1#, Marina B. Blanco2,3#, Jordi Salmona4, Jelmer Poelstra3, 20 21 7 Jean B. AndriambelosonFor5, Alex Peer Miller6, BlanchardReview Randrianambinina7,8, David 22 23 7 1 4,6 24 8 W. Rasolofoson , Jasmin Mantilla-Contreras , Lounès Chikhi , Edward E. 25 26 9 Louis Jr.9, Anne D. Yoder3, Ute Radespiel10* 27 28 29 10 30 31 32 11 Affiliations: 33 34 35 12 1: Research Group Ecology and Environmental Education, Institute of Biology 36 37 13 and Chemistry, University of Hildesheim, Universitaetsplatz 1, 31141 38 39 14 Hildesheim, Germany 40 41 42 15 2: Duke Lemur Center, Duke University, Durham, NC 27705, USA 43 44 45 16 3: Department of Biology, Duke University, Durham, NC 27708, USA 46 47 48 17 4: CNRS, Université Paul Sabatier, IRD, UMR5174 EDB (Laboratoire Évolution 49 50 51 18 & Diversité 11 Biologique), 118 route de Narbonne, 31062 Toulouse, France 52 53 5 54 19 : Zoology and Animal Biodiversity, University of Antananarivo, Antananarivo 55 56 20 101, Madagascar 57 58 59 60

John Wiley & Sons Page 3 of 66 American Journal of Primatology

Schüßler et al. page 2 1 2 3 4 21 6: Instituto Gulbenkian de Ciência, Rua da Quinta Grande, 6, 2780-156 Oeiras, 5 6 7 22 Portugal 8 9 7 10 23 : Groupe d’Etude et de Recherche sur les Primates de Madagascar (GERP), 11 12 24 BP 779, Antananarivo 101, Madagascar 13 14 15 25 8: Faculté des Sciences, University of Mahajanga, Mahajanga, Madagascar 16 17 18 26 9: Grewcock Center for Conservation and Research, Omaha’s Henry Doorly 19 20 27 Zoo and Aquarium, Omaha, NE 21 For Peer Review 22 23 28 10: Institute of Zoology, University of Veterinary Medicine Hannover, Buenteweg 24 25 29 17, 30559 Hannover, Germany 26 27 28 30 29 30 31 31 32 32 Short title: 33 34 35 33 A new species of mouse lemur 36 37 38 34 39 40 41 35 42 43 44 36 Corresponding author: 45 46 47 37 *Ute Radespiel, Institute of Zoology, University of Veterinary Medicine 48 49 Hannover, Buenteweg 17, 30559 Hannover, Germany; email: 50 38 51 52 39 [email protected] 53 54 55 40 56 57 58 41 #: should be considered as shared first authors 59 60

John Wiley & Sons American Journal of Primatology Page 4 of 66

Schüßler et al. page 3 1 2 3 4 42 Abstract 5 6 7 43 Delimitation of cryptic species is increasingly based on genetic analyses but the 8 9 10 44 integration of distributional, morphological, behavioral and ecological data offers 11 12 45 unique complementary insights into species diversification. We surveyed 13 14 46 communities of nocturnal mouse lemurs (Microcebus spp.) in five different sites 15 16 17 47 of northeastern Madagascar, measuring a variety of morphological parameters 18 19 48 and assessing reproductive states for 123 individuals belonging to five different 20 21 49 lineages. We documentedFor two Peer different Reviewnon-sister lineages occurring in 22 23 50 sympatry in two areas. In both cases, sympatric species pairs consisted of a 24 25 26 51 locally restricted (M. macarthurii or M. sp. #3) and a more widespread lineage 27 28 52 (M. mittermeieri or M. lehilahytsara). Estimated Extents of Occurrence (EOO) of 29 30 53 these lineages differed remarkably with 560 and 1,500 km² versus 9,250 and 31 32 33 54 50,700 km², respectively. Morphometric analyses distinguished unambiguously 34 35 55 between sympatric species and detected more subtle but significant differences 36 37 56 among sister lineages. Tail length and body size were most informative in this 38 39 40 57 regard. Reproductive schedules were highly variable among lineages, most 41 42 58 likely impacted by phylogenetic relatedness and environmental variables. While 43 44 59 sympatric species pairs differed in their reproductive timing (M. sp. #3 / M. 45 46 60 lehilahytsara and M. macarthurii / M. mittermeieri), warmer lowland rainforests 47 48 49 61 were associated with a less seasonal reproductive schedule for M. mittermeieri 50 51 62 and M. lehilahytsara compared to populations occurring in montane forests. 52 53 63 Distributional, morphological and ecological data gathered in this study support 54 55 56 64 the results of genomic species delimitation analyses conducted in a companion 57 58 65 study, which identified one lineage, M. sp. #3, as meriting formal description as 59 60 66 a new species. Consequently, a formal species description is included.

John Wiley & Sons Page 5 of 66 American Journal of Primatology

Schüßler et al. page 4 1 2 3 4 67 Worryingly, our data also show that geographically restricted populations of M. 5 6 7 68 sp. #3 and its sister species (M. macarthurii) are at high risk of local and 8 9 69 perhaps permanent extinction from both deforestation and habitat 10 11 70 fragmentation. 12 13 14 71 15 16 17 72 Research highlights 18 19 20 73  Two pairs of Microcebus species occur in partial sympatry 21 For Peer Review 22 23 74  Morphological distinctiveness supports genomic species delimitation in 24 25 75 cryptic lemurs 26 27 76  High plasticity in reproductive schedules in a lineage of habitat 28 29 30 77 generalists detected 31 32 33 78 34 35 36 79 Keywords: evolution, cryptic species, phenotype, habitat use, sympatry, 37 38 80 conservation 39 40 41 81 42 43 44 82 1. Introduction 45 46 47 83 Madagascar is one of the world’s prime biodiversity hotspots and its endemic 48 49 84 group of primates, the lemurs (Primates; Lemuriformes), are flagships for 50 51 85 species conservation (Myers, Mittermeier, Mittermeier, Da Fonseca & Kent, 52 53 54 86 2000). More than 100 species of lemurs are recognized today making up about 55 56 87 one fifth of all living primate species on earth (Estrada et al., 2017). However, 57 58 88 the full extent of lemur species diversity is not yet fully known as several regions 59 60

John Wiley & Sons American Journal of Primatology Page 6 of 66

Schüßler et al. page 5 1 2 3 4 89 in Madagascar are still poorly studied. Intensified biological inventories during 5 6 7 90 recent years have indeed resulted in a considerable rise in lemur species 8 9 91 numbers. One example of increased taxonomic recognition is the genus of 10 11 92 mouse lemurs (Microcebus). These small-bodied and nocturnal primates can be 12 13 14 93 found in all regions of Madagascar that offer forested habitats, while partially 15 16 94 deforested areas appear to offer at least dispersal opportunities (Knoop, Chikhi 17 18 95 & Salmona, 2017; Miller et al., 2018; Schüßler, Radespiel, Ratsimbazafy & 19 20 96 Mantilla-Contreras, 2018). 21 For Peer Review 22 23 97 Although rather widespread across the island, mouse lemurs suffer from habitat 24 25 26 98 loss due to ongoing deforestation (Vieilledent et al., 2018). According to the 27 28 99 IUCN, eleven species are listed as Endangered, three species are Critically 29 30 100 Endangered, while four are Vulnerable and only two species are categorized as 31 32 33 101 of Least Concern. The remaining four species have not yet been assessed due 34 35 102 to their recent discovery (Andriaholinirina et al., 2014; Hotaling et al., 2016; 36 37 103 Rasoloarison et al., 2013). Integrating ecological and distributional data with 38 39 40 104 molecular analyses in mouse lemurs is often difficult, largely because of their 41 42 105 cryptic morphology and life history and lack of detailed metadata (Zimmermann 43 44 106 & Radespiel, 2014). Analyses of mtDNA datasets have identified divergent 45 46 107 lineages despite similar phenotypes and roughly similar ecological niches, and 47 48 49 108 have led to the description of twelve new species over the past 20 years from 50 51 109 the western part (Louis et al. 2008; Olivieri et al. 2007; Rasoloarison, Goodman 52 53 110 & Ganzhorn, 2000; Yoder et al. 2000; Zimmermann, Cepok, Rakotoarison, 54 55 56 111 Zietemann & Radespiel, 1998) and a further eleven species from the eastern 57 58 112 part of Madagascar (Hotaling et al. 2016; Kappeler, Rasoloarison, 59 60 113 Razafimanantsoa, Walter & Roos, 2005; Louis et al. 2006; Radespiel et al.

John Wiley & Sons Page 7 of 66 American Journal of Primatology

Schüßler et al. page 6 1 2 3 4 114 2008, 2012; Rasoloarison, Weisrock, Yoder, Rakotondravony & Kappeler, 5 6 7 115 2013). 8 9 10 116 Recent studies indicate that some regions appear to be hotspots of 11 12 117 microendemism. One of these is located in northeastern Madagascar where M. 13 14 118 lehilahytsara (Kappeler et al., 2005), M. mittermeieri and M. simmonsi (Louis et 15 16 17 119 al., 2006) are known to occur. Radespiel et al. (2008) surveyed the forests of 18 19 120 the Makira region (Anjiahely, Figure 1) and found evidence for three divergent 20 21 121 lineages occurring inFor sympatry, Peer a phenomenon Review previously undocumented for 22 23 122 mouse lemurs. One of these was identified as M. mittermeieri, while the second 24 25 26 123 was newly described as M. macarthurii. The third lineage, named M. sp. #3, 27 28 124 was hypothesized to be a new species based on mitochondrial sequence data 29 30 125 but could not be formally described given that only a single individual was 31 32 33 126 found. 34 35 36 127 We conducted additional sampling in northeastern Madagascar to fill the gap 37 38 128 between the known distribution of M. simmonsi (Zahamena NP, Betampona 39 40 129 SNR, Tampolo; Louis et al., 2006) and the sympatric species pair M. 41 42 43 130 macarthurii and M. mittermeieri at Anjiahely (Radespiel et al., 2008; Figure 1). 44 45 131 The presence of the M. sp. #3 lineage was indeed confirmed but only for three 46 47 132 study sites south of Anjiahely by genomic data in a companion study (Poelstra 48 49 133 et al., 2020). Based on a comprehensive dataset generated from restriction-site 50 51 52 134 associated DNA sequencing (RADseq) using a total of 63 mouse lemurs from 53 54 135 the entire region (Marojejy NP to Betampona SNR, excluding Ile St. Marie, 55 56 136 Figure 1), these analyses unambiguously supported four different lineages: M. 57 58 59 137 sp. #3, M. macarthurii, M. lehilahytsara/M. mittermeieri and M. simmonsi. While 60

John Wiley & Sons American Journal of Primatology Page 8 of 66

Schüßler et al. page 7 1 2 3 4 138 the two lineages M. sp. #3 and M. macarthurii unambiguously passed all 5 6 7 139 species delimitation tests (i.e., mitochondrial and nuclear monophyly in RAxML 8 9 140 and SVDquartets, clear nuclear clustering in NGSadmix, rejection of a simple 10 11 141 isolation-by-distance pattern, formal species delimitation using SNAPP Bayes 12 13 14 142 factors, BPP and gdi), M. mittermeieri and M. lehilahytsara did not fall into two 15 16 143 separate monophyletic clades (Poelstra et al., 2020; Figure 2). Instead, these 17 18 144 two latter species exhibited a single isolation-by-distance pattern and high 19 20 145 levels of interspecific gene flow, suggesting that separate species status may 21 For Peer Review 22 23 146 not be justified (Figure 2; discussed in detail in Poelstra et al., 2020). For the 24 25 147 purpose of this study, however, we will still treat these two lineages as separate 26 27 148 taxa in order to be able to test their distinctiveness in other domains. 28 29 30 149 Besides using molecular data, species delimitation under an integrative 31 32 150 taxonomic approach (e.g., Padial, Miralles, de la Riva & Vences, 2010) requires 33 34 151 incorporating morphological and ecological information. While phenotypic 35 36 37 152 differences between lineages could, for instance, indicate dietary preferences 38 39 153 (e.g., Viguier, 2004), environmental and reproductive data can help to 40 41 154 understand the role of habitat selection (Dammhahn & Kappeler, 2008; 42 43 Rakotondravony & Radespiel, 2009) and reproductive schedules (Evasoa et al. 44 155 45 46 156 2018) during speciation. 47 48 157 Here, we complement the molecular results presented by Poelstra et al. (2020) 49 50 158 by (1) providing morphological, ecological and distributional data for the M. sp. 51 52 53 159 #3 lineage in comparison to all other species from the same region and (2) by 54 55 160 formally describing this new species. 56 57 58 161 59 60

John Wiley & Sons Page 9 of 66 American Journal of Primatology

Schüßler et al. page 8 1 2 3 4 162 2. Methods 5 6 7 163 Study region 8 9 10 164 Northeastern Madagascar is characterized by a humid climate with abundant 11 12 13 165 precipitation (2,086 ± SD 530 mm SD; Fick & Hijmans, 2017) and tropical 14 15 166 rainforests as primary vegetation (Kottek, Grieser, Beck, Rudolf & Rubel, 2006). 16 17 167 Forest cover has been steadily declining for decades, with lowland rainforests 18 19 20 168 being particularly prone to deforestation (Schüßler, Mantilla-Contreras, 21 For Peer Review 22 169 Stadtmann, Ratsimbazafy & Radespiel, 2020; Vieilledent et al., 2018). By 2018, 23 24 170 about half of the remaining forested areas were under protection by 25 26 171 governmental institutions or non-governmental organizations (Schüßler et al., 27 28 29 172 2020). The study region is subdivided by more than seven large rivers that flow 30 31 173 from the highlands of the central plateau (west of the study region; Figure 1) 32 33 174 eastwards into the Indian Ocean. Large rivers have been considered potential 34 35 36 175 biogeographic boundaries for mouse lemurs (e.g., Olivieri et al., 2007; Martin, 37 38 176 1972). 39 40 41 177 42 43 44 178 Microcebus sampling 45 46 47 179 Mouse lemurs were sampled between 2008 and 2017 at five lowland rainforest 48 49 180 sites ranging in altitude between 42 and 462 m a.s.l. (Figure 1). In particular, 50 51 181 animals were captured around the village of Anjiahely (Makira region; where the 52 53 54 182 holotype of M. macarthurii was obtained; Radespiel et al., 2008), in the 55 56 183 fragmented forests around the village of Ambavala (Schüßler et al., 2018), 57 58 184 within Mananara-Nord NP (Ivontaka-Sud section), around Antanambe village in 59 60

John Wiley & Sons American Journal of Primatology Page 10 of 66

Schüßler et al. page 9 1 2 3 4 185 the vicinity of Mananara-Nord NP, as well as in the Ambodiriana community 5 6 7 186 protected area (Miller et al., 2018). All study sites comprise habitats ranging 8 9 187 from undisturbed near-primary rainforest to heavily degraded secondary shrub-, 10 11 188 grass- and fernlands (Radespiel et al., 2008; Miller et al., 2018; Schüßler et al., 12 13 14 189 2018). 15 16 190 Mouse lemurs were captured using Sherman Live traps (H. B. Sherman Traps 17 18 191 ®) or by hand during nocturnal surveys (e.g., Radespiel et al., 2008). 19 20 192 Morphometric measurements were taken for each individual (see below) and 21 For Peer Review 22 23 193 additional descriptors such as fur coloration were noted and photographed. Ear 24 25 194 biopsies (~2 mm²) were collected to provide DNA samples, and all animals were 26 27 195 released unharmed within 24 hours at their exact location of capture. 28 29 30 196 GPS coordinates and the altitude of capture locations were collected to 31 32 197 estimate the altitudinal range and Extent of Occurrence of mouse lemur species 33 34 198 included in this study. The latter measure follows the definition of the IUCN 35 36 37 199 (2012) in which the “shortest continuous imaginary boundary which can be 38 39 200 drawn to encompass all the known, inferred or projected sites of present 40 41 201 occurrence of a taxon” is used to derive the possible distribution of a certain 42 43 species. 44 202 45 46 203 All procedures adhered to the standards of the International Primatological 47 48 204 Society (Riley, MacKinnon, Fernandez-Duque, Setchell & Garber, 2014) and 49 50 205 the Principles for the Ethical Treatment of Non-Human Primates of the 51 52 53 206 American Society of Primatologists (2001). This research was conducted with 54 55 207 permission from institutional and governmental agencies that regulate animal 56 57 208 research in Madagascar, Germany, France, Portugal, and the United States. 58 59 60 209

John Wiley & Sons Page 11 of 66 American Journal of Primatology

Schüßler et al. page 10 1 2 3 4 210 Morphometric characterization and reproduction 5 6 7 211 Captured mouse lemurs were measured for 13 different morphometric variables 8 9 10 212 (ear length, ear width, head length, head width, snout length, inter- and intra- 11 12 213 orbital distance, lower leg length, hind foot length, third toe length, tail length, 13 14 214 body length and body mass) following Hafen, Neveu, Rumpler, Wilden and 15 16 17 215 Zimmermann (1998) and Zimmermann et al. (1998). Mouse lemurs were 18 19 216 assigned to two age categories based on their body mass and reproductive 20 21 217 state: (1) adult in contrastFor to Peer(2) young mouseReview lemurs (<1 year old) that had a 22 23 218 relatively low body mass (Table 1) and showed no visible nipples (females) and 24 25 26 219 undeveloped, or barely developed testes (males). Young mouse lemurs were 27 28 220 excluded from all morphometric comparisons and only values for adult 29 30 221 Microcebus spp. are used for further analyses. Very light young mouse lemurs 31 32 33 222 of about half of the adult body mass were termed juveniles. 34 35 223 To supplement the comparative dataset, we also included published 36 37 224 morphometric data from 42 M. lehilahytsara individuals from Mantadia NP 38 39 40 225 (holotype locality, Randrianambinina, 2001) and data from 22 M. mittermeieri 41 42 226 individuals that had previously been caught near Anjiahely (Radespiel et al., 43 44 227 2008). The morphometric dataset is provided in the supplementary material 45 46 228 (Table S1). 47 48 49 229 Mouse lemurs are seasonal breeders and can already reproduce during their 50 51 52 230 first year of life (Evasoa et al., 2018; Kraus, Eberle & Kappeler, 2008; 53 54 231 Schmelting, Zimmermann, Berke, Bruford & Radespiel, 2007). Reproductive 55 56 232 states were assessed using several morphological indicators (i.e., vaginal 57 58 59 233 morphology and testis size) frequently used in the literature (e.g., Blanco, 2008; 60

John Wiley & Sons American Journal of Primatology Page 12 of 66

Schüßler et al. page 11 1 2 3 4 234 Randrianambinina, Rakotondravony, Radespiel & Zimmermann, 2003; 5 6 7 235 Wrogemann & Zimmermann, 2001). The reproductive state of females was 8 9 236 defined as anestrous (closed vagina, non-reproductive), pre-estrous (swollen 10 11 237 vagina), estrous (open vagina), pregnant (enlarged belly) or lactating (palpable 12 13 14 238 and enlarged nipples that release milk under soft pressure). Male reproduction 15 16 239 was assessed based on testes state: while being completely regressed in the 17 18 240 non-breeding season, testes increase considerably in size starting about one to 19 20 241 three months prior to female estrus (Evasoa et al., 2018). However, testes size 21 For Peer Review 22 23 242 was measured slightly differently across our dataset (i.e., left and right testes 24 25 243 separately or only total width) and measures may also differ slightly between 26 27 244 different researchers. Therefore, we defined a unified and realistic threshold for 28 29 30 245 classifying total testes width by defining a binary variable for a regressed (< 31 32 246 10.0 mm) or enlarged (> 10.0 mm) width. Total testes width for the regressed 33 34 247 category ranged from 0.0 to 5.3 mm and from 10.2 to 26.2 mm for the enlarged 35 36 37 248 category across all species. 38 39 249 One limitation to the morphometric analyses is that measurements across the 40 41 250 five different lineages were taken by five researchers, thus potentially 42 43 introducing inter-observer error. It is worth reporting, however, that two 44 251 45 46 252 researchers contributed data points to more than one species (DS, DWR), and 47 48 253 that DS was trained by DWR. Furthermore, there was a strict selection of 49 50 254 measurements that fully agreed with collection standards before assembling the 51 52 53 255 dataset. Finally, the dataset was carefully scanned for outliers and 54 55 256 inconsistencies within and across species, and a total of 21 measurements was 56 57 257 excluded for this reason prior to data analyses. Reproductive data can also be 58 59 60 258 found in supplementary Table S1.

John Wiley & Sons Page 13 of 66 American Journal of Primatology

Schüßler et al. page 12 1 2 3 4 259 5 6 7 260 Statistical analyses of morphometrics 8 9 10 261 We performed a principal component analysis (PCA) and linear discriminant 11 12 13 262 analysis (LDA) as well as Analyses of Variance (ANOVA) with Tukey post-hoc 14 15 263 tests for pair-wise comparisons for all 13 morphometric variables. Assumptions 16 17 264 of the respective tests were examined beforehand using Shapiro-Wilk and 18 19 20 265 Levene’s tests in R (R Core Team 2019; RStudio Team, 2016) using the car 21 For Peer Review 22 266 v3.0-2 package (Fox & Weisberg 2011). M. lehilahytsara individuals from 23 24 267 Ambavala were excluded from the ANOVA due to small sample size. For LDA 25 26 268 and ANOVA, mouse lemurs were a priori assigned to their respective taxon 27 28 29 269 based on the results of the parallel phylogenomic study (Figure 2; Poelstra et 30 31 270 al., 2020). For PCA, species assignment was done a posteriori to investigate 32 33 271 clustering under naïve conditions in which the distance between sample points 34 35 36 272 reflects their distance along the major axes of variation in the dataset. 37 38 273 Accordingly, points that cluster closely together are more similar to each other 39 40 274 than points that do not (Abdi & Williams, 2010). In contrast to that, the LDA aims 41 42 43 275 to minimize distances within pre-defined clusters while maximizing distances 44 45 276 among clusters (Balakrishnama & Ganapathiraju, 1998). The PCA was followed 46 47 277 by a permutational multivariate analysis of variances (PERMANOVA) as 48 49 278 implemented in the “vegan” R package (Oksanen et al., 2019), which tests the 50 51 52 279 null hypothesis of no differences in the position of cluster centroids (Anderson, 53 54 280 2017). 55 56 57 281 For both PCA and LDA, we used all measurements except third-toe length, as 58 59 282 this measurement was not available for M. simmonsi. We further only used a 60

John Wiley & Sons American Journal of Primatology Page 14 of 66

Schüßler et al. page 13 1 2 3 4 283 subset of eleven M. sp. #3 individuals for which we had all 13 measurements. 5 6 7 284 The PCA was performed using the “prcomp” command in R (scaled and 8 9 285 centered), while the LDA was calculated using the “MASS” package (v7.3-51.3; 10 11 286 Venables & Ripley, 2002). Model fit of the latter was evaluated by a jackknife 12 13 14 287 cross-validation and calculated as misclassification error. We also computed 15 16 288 Wilks’ Lambda and the P-value to evaluate the ability of the LDA model to 17 18 289 distinguish between the five lineages. 19 20 21 290 For Peer Review 22 23 24 291 3. Results 25 26 27 292 Distribution of Microcebus spp. in northeastern Madagascar 28 29 30 293 First, we confirmed the presence of mouse lemurs at four locations previously 31 32 294 not surveyed in our study region (Figure 1 & 2; species delimitation based on 33 34 35 295 genomic data in Poelstra et al., 2020). At two locations, two different mouse 36 37 296 lemur species were found in sympatry. These are M. macarthurii/M. mittermeieri 38 39 297 in Anjiahely and M. sp. #3/M. lehilahytsara in Ambavala (Figure 1). In 40 41 42 298 Mananara-Nord NP and around Antanambe village (south of the Mananara 43 44 299 River), extensive surveys revealed only the presence of M. sp. #3. At 45 46 300 Ambodiriana (one major river further south from Antanambe; Figure 1), we only 47 48 301 found M. simmonsi. Altitudinal ranges vary among the lineages, with M. 49 50 51 302 macarthurii and its sister species M. sp. #3 being only found in lowland 52 53 303 rainforests. The other three lineages, M. mittermeieri, M. lehilahytsara and M. 54 55 304 simmonsi, were found in lowland as well as montane rainforests (Table 2). The 56 57 58 305 estimated Extent of Occurrence (EOO) is much smaller for the two lowland 59 60 306 species (M. macarthurii [560 km²] and M. sp. #3 [1,500 km²]) compared to M.

John Wiley & Sons Page 15 of 66 American Journal of Primatology

Schüßler et al. page 14 1 2 3 4 307 mittermeieri (9,250 km²) and M. lehilahytsara (50,700 km²; Table 2). Combined 5 6 7 308 EOO for M. lehilahytsara and M. mittermeieri, if considered as a single species, 8 9 309 is estimated with 66,800 km². 10 11 12 310 13 14 15 311 Morphometric distinction between lineages 16 17 18 312 All morphometric parameters differed significantly among lineages (ANOVA; P < 19 20 313 0.001; Table S3) and Tukey post-hoc tests revealed many pairwise differences 21 For Peer Review 22 (Figure 3; Table 3; Figure S1). M. sp. #3 can be statistically differentiated from 23 314 24 25 315 its closest relative, M. macarthurii, by 5 out of 13 parameters. M. macarthurii 26 27 316 has smaller body size and longer tail length, and subtle differences were found 28 29 317 in head-associated parameters (i.e., ear width, head length and width). By 30 31 32 318 comparison, M. lehilahytsara (from Mantadia NP) and M. mittermeieri (from 33 34 319 Anjiahely) differed in 7 out of 13 variables. Major differences were found in 35 36 320 snout and tail length, while other differences were more subtle but statistically 37 38 39 321 significant (Figure 3; Figure S1; Table 3). Both M. lehilahytsara and M. 40 41 322 mittermeieri were significantly smaller than M. sp. #3 and M. macarthurii, which 42 43 323 was mainly reflected in the parameters body mass and length, tail length, lower 44 45 leg, hind foot and third toe length (Figure 3, Figure S1; Table 3). M. simmonsi 46 324 47 48 325 took an intermediate position in most comparisons. M. lehilahytsara from 49 50 326 Ambavala was not compared by ANOVA due to the small number of adults (N = 51 52 327 3). However, the individuals from this population showed remarkably different 53 54 55 328 measures compared to conspecifics from Mantadia NP or M. mittermeieri from 56 57 329 Anjiahely (i.e., body length, tail length; Table 3). 58 59 60

John Wiley & Sons American Journal of Primatology Page 16 of 66

Schüßler et al. page 15 1 2 3 4 330 These patterns are also illustrated in the multivariate analyses: the PCA (a 5 6 7 331 posteriori assignment of species clusters) distinguished two clusters along PC1 8 9 332 and PC2 (Figure 4). The two larger species, M. sp. #3 and M. macarthurii, 10 11 333 clustered with negative values along PC1 and the smaller M. mittermeieri and 12 13 14 334 M. lehilahytsara had positive values along PC1. Both sister-species pairs were 15 16 335 split in two clusters along PC2, while M. lehilahytsara from Ambavala clustered 17 18 336 with M. mittermeieri from Anjiahely. Again, M. simmonsi occupied a position in 19 20 337 between these two major clusters. PC3 mainly separated M. sp. #3 and M. 21 For Peer Review 22 23 338 macarthurii, while all other species clustered together along PC3 and PC4. 24 25 339 These first four principal components (PCs) together explained 84.7% of the 26 27 340 variance in our dataset. These five clusters corresponding to the five lineages 28 29 30 341 were significantly different from each other as indicated by the PERMANOVA (F 31 32 342 = 36.88, df = 77, P < 0.001). 33 34 35 343 The LDA model (a priori assignment of species) could also statistically 36 37 344 distinguish between the five lineages (Wilk’s Lambda = 0.005, F = 10.338, P < 38 39 40 345 0.001). Four distinct clusters that included M. lehilahytsara, M. macarthurii, M. 41 42 346 mittermeieri and M. sp. #3 are illustrated in Figure 5. M. simmonsi fell again 43 44 347 between these major clusters, while M. lehilahytsara and M. mittermeieri 45 46 348 exhibited some minor overlap. M. lehilahytsara from Ambavala again showed 47 48 49 349 more affinity to M. mittermeieri than to conspecifics from Mantadia NP. 50 51 350 Misclassification error after cross-validation was 12.8% and misclassification 52 53 351 occurred mainly with M. simmonsi (Table S4). The first two discriminant 54 55 56 352 functions explained together 86.9% of the variation between the groups. 57 58 59 353 60

John Wiley & Sons Page 17 of 66 American Journal of Primatology

Schüßler et al. page 16 1 2 3 4 354 Reproductive status 5 6 7 355 At Anjiahely, all male M. macarthurii that were captured from late October to 8 9 10 356 December had enlarged testes (N = 8), while the four female M. macarthurii 11 12 357 showed no signs of reproduction from September to November (Figure 6). Only 13 14 358 one out of five females was in pre-estrus (swollen vagina) in early November. 15 16 17 359 No young or juvenile individuals were found. Sympatric M. mittermeieri 18 19 360 individuals (N = 22 adults) were captured across three months: in September, 20 21 361 all males already hadFor well developed Peer testes Review (N = 11), while females showed no 22 23 362 signals of reproduction (N = 5). In November, testes were still well developed (N 24 25 26 363 = 2), and 2 out of 3 females were in estrus. Juveniles weighing 20-24 g were 27 28 364 captured in the population in mid-December (Figure 6). 29 30 31 365 During our surveys at three locations between August 13th and September 17th, 32 33 366 none of the M. sp. #3 females (N = 7) showed indications of estrus, pregnancy 34 35 36 367 or lactation. However, all adult males had well developed testes (N = 15) and no 37 38 368 juveniles were found. In contrast, at Ambavala between September 8th to 19th, 39 40 369 all sympatric M. lehilahytsara females were anestrous (N = 2), while all adult 41 42 43 370 males had regressed testes (N = 6). At the same time, two juvenile males were 44 45 371 captured that were still very light with only 19 and 20 g. In contrast, M. 46 47 372 lehilahytsara females at Mantadia NP were anestrous from May until late 48 49 373 October (N = 5), in estrus in November (N = 1) and pregnant in December (N = 50 51 52 374 3; Figure 6). Males had regressed testes in April and May (N = 8) and enlarged 53 54 375 testes from August to November (N = 13). Although young individuals were 55 56 376 identifiable between March and August, juvenile individuals (20 g body mass, N 57 58 59 377 = 1) were only found in March (no other captures at that time, Figure 6). 60

John Wiley & Sons American Journal of Primatology Page 18 of 66

Schüßler et al. page 17 1 2 3 4 378 Eight M. simmonsi were caught in Ambodiriana in June and July. All males (N = 5 6 7 379 5) had enlarged testes, while females (N = 3) were not reproductively active 8 9 380 (Figure 6). 10 11 12 381 13 14 15 382 4. Discussion 16 17 383 We studied the distribution, morphology and reproductive state of five mouse 18 19 20 384 lemur lineages occurring in a complex spatial pattern across a small region in 21 For Peer Review 22 385 northeastern Madagascar. Comparative phenotypic and ecological data was 23 24 386 previously sparse for four of these taxa, and this study therefore represents an 25 26 387 important step towards deepening our understanding of mouse lemur diversity 27 28 29 388 in this understudied hotspot of microendemism. 30 31 32 389 33 34 35 390 Distribution of Microcebus spp. in northeastern Madagascar 36 37 38 391 Our study, in conjunction with the companion study by Poelstra et al. (2020), 39 40 392 confirmed the presence of five Microcebus spp. lineages, including two pairs of 41 42 393 closely related sister lineages, M. macarthurii / M. sp. #3 and M. mittermeieri / 43 44 45 394 M. lehilahytsara in northeastern Madagascar, with two cases of local sympatry. 46 47 395 Besides the case of M. macarthurii and M. mittermeieri at Anjiahely (Radespiel 48 49 396 et al., 2008), sympatry of two mouse lemur species is so far only known from 50 51 397 five cases from western Madagascar and one case from the northern part of the 52 53 54 398 island (Sgarlata et al., 2019). In the five western cases, geographically 55 56 399 restricted species co-exist with the widely distributed congener M. murinus 57 58 400 (Radespiel, 2016) which probably expanded northwards rather recently 59 60

John Wiley & Sons Page 19 of 66 American Journal of Primatology

Schüßler et al. page 18 1 2 3 4 401 (Schneider, Chikhi, Currat & Radespiel, 2010). Here, we confirm a new case 5 6 7 402 with M. sp. #3 being found in sympatry with M. lehilahytsara in Ambavala 8 9 403 (Figure 1). 10 11 12 404 Ecologically, two sister species, M. macarthurii and M. sp. #3, appear to be 13 14 405 restricted to lowland forests, whereas M. mittermeieri and M. lehilahytsara are 15 16 17 406 present in lowland as well as montane forests (Table 2). Thus, M. macarthurii 18 19 407 and M. sp. #3 both have geographically restricted distributions and possess a 20 21 408 limited Extend of Occurrence,For Peer suggesting Review that they are microendemic, while M. 22 23 409 lehilahytsara and M. mittermeieri are more widely distributed. Consequently, the 24 25 26 410 cases of sympatry between M. sp. #3/M. lehilahytsara and M. macarthurii/M. 27 28 411 mittermeieri, respectively, may to some extent be similar to the cases of 29 30 412 sympatry from western Madagascar, where locally restricted species co-occur 31 32 33 413 with the habitat generalist M. murinus (Kamilar, Blanco & Muldoon, 2016; 34 35 414 Radespiel, 2016). However, the recent expansion of M. murinus seems to have 36 37 415 been a unique event that has no clear equivalent in eastern Madagascar. 38 39 40 416 M. simmonsi was previously reported from Zahamena NP, Betampona SNR 41 42 43 417 and Tampolo (Hotaling et al., 2016; Louis et al., 2006). We can now confirm its 44 45 418 occurrence 75 km and four inter-river systems (IRSs) further north (Figure 1 & 46 47 419 2) which expands its EOO by almost fivefold. The northern range limit for M. 48 49 420 simmonsi appears to be the Anove River, which separates it from M. sp. #3. 50 51 52 421 These two species have (so far) only been found in allopatry (despite intensive 53 54 422 sampling north of the river), and we consider two alternative hypotheses 55 56 423 responsible for this pattern: (1) competitive exclusion at the geographic limits of 57 58 59 424 the respective species ranges (Beaudrot et al., 2013; Hardin, 1960) or (2) an 60

John Wiley & Sons American Journal of Primatology Page 20 of 66

Schüßler et al. page 19 1 2 3 4 425 altitudinal range limit of both species below 640-700 m a.s.l corresponding to 5 6 7 426 the source region of the Anove River (DS, unpubl. data). The latter hypothesis 8 9 427 appears to be unlikely, as both species are distributed over two or more IRSs 10 11 428 that are separated by rivers with much higher headwaters (DS, unpubl. data). 12 13 14 429 Moreover, M. simmonsi has been found at an elevation of around 956 m a.s.l. 15 16 430 (in Zahamena NP, Louis et al., 2006), contradicting the altitudinal limitation 17 18 431 hypothesis. A further expansion of M. simmonsi northwards across the Anove 19 20 432 River, however, may have been precluded by the presence of the larger M. sp. 21 For Peer Review 22 23 433 #3 that may have a higher competitive potential than the smaller M. simmonsi 24 25 434 (Table 3; Thorén, Linnenbrink & Radespiel, 2011). In the case of M. sp. #3, the 26 27 435 subpopulations on both sides of the large Mananara River (Figure 1) were 28 29 30 436 shown to belong to two separate population clusters evolving largely 31 32 437 independently from each other (Figure 2; Poelstra et al., 2020). This suggests 33 34 438 that the Mananara River poses a significant barrier to gene flow within this 35 36 37 439 species (Poelstra et al., 2020). This moderate sensitivity to altitude may have 38 39 440 limited the colonization potential of M. sp. #3 southwards. These complex 40 41 441 patterns demonstrate that the biogeography and phylogeography of mouse 42 43 lemurs in this region of Madagascar are still not completely understood and 44 442 45 46 443 should be re-evaluated. 47 48 49 444 Biogeographic patterns in the region are further complicated by our unexpected 50 51 445 finding that the previously regarded “highland specialist” M. lehilahytsara 52 53 446 (Radespiel et al., 2012) also occurs in the lowland rainforests around the village 54 55 56 447 of Ambavala (233-462 m a.s.l). Prior to our study, this species had never been 57 58 448 observed at altitudes below 800 m a.s.l., although it is known to occur in an 59 60 449 extensive stretch of highland forests in central and northeastern Madagascar

John Wiley & Sons Page 21 of 66 American Journal of Primatology

Schüßler et al. page 20 1 2 3 4 450 (between Riamalandy; Figure 1; (Weisrock et al., 2010) and Tsinjoarivo (430 km 5 6 7 451 further south; (Yoder et al., 2016)). Besides in the IRS of Ambavala, the other 8 9 452 lowland regions are always inhabited by different mouse lemur species (from 10 11 453 north to south: M. simmonsi (Louis et al., 2006), M. gerpi (Radespiel et al., 12 13 14 454 2012), M. marohita (Rasoloarison et al., 2013)). Phylogenomic analyses 15 16 455 indicated, that M. lehilahytsara and M. mittermeieri should rather be considered 17 18 456 a single widely distributed species with extensive population structure, which is 19 20 457 most likely driven by isolation-by-distance between subpopulations (Figure 2; 21 For Peer Review 22 23 458 Poelstra et al., 2020). Under these circumstances, allopatry of southern M. 24 25 459 lehilahytsara populations with lowland taxa, but sympatry of more northern 26 27 460 populations with lowland species (i.e., M. sp. #3 (at Ambavala) and M. 28 29 30 461 macarthurii (at Anjiahely)) requires further investigation concerning the relative 31 32 462 importance of competitive exclusion, different habitat preferences or 33 34 463 distributional barriers, all of which have remained unexplored so far. 35 36 37 464 38 39 40 465 Morphometric differences among mouse lemurs 41 42 43 466 Mouse lemurs are typically regarded as cryptic species exhibiting only subtle 44 45 interspecific morphological differences (Zimmermann & Radespiel 2014). 46 467 47 48 468 Although our measurements of 13 external body parameters generally confirm 49 50 469 their cryptic nature, some differences could be detected that can help to 51 52 470 distinguish different species. 53 54 55 471 All analyses confirmed a noticeable divide between the two larger taxa M. sp. 56 57 58 472 #3/M. macarthurii and the two smaller-bodied lineages M. lehilahytsara and M. 59 60 473 mittermeieri (along PC1, Figure 4, Table 3). Even sister taxa could be

John Wiley & Sons American Journal of Primatology Page 22 of 66

Schüßler et al. page 21 1 2 3 4 474 differentiated by some morphological measurements. Specifically, M. sp. #3 5 6 7 475 could be distinguished from M. macarthurii by body size, tail length and three 8 9 476 head-associated parameters (ear width, head length, and head width). Tail 10 11 477 length has been previously emphasized as a feature distinguishing mouse 12 13 14 478 lemur species (e.g., Radespiel et al. 2012) and can be measured with high 15 16 479 accuracy. The more subtle differences in head-associated parameters must be 17 18 480 interpreted more carefully, particularly because measurements were not always 19 20 481 made by the same person. Nevertheless, it has been suggested that skull 21 For Peer Review 22 23 482 parameters may also vary with feeding habits for lemurs and strepsirrhine 24 25 483 primates in general (e.g., omnivorous, folivorous or frugivorous etc.; Fabre et 26 27 484 al., 2018; Meloro et al., 2015; Viguier, 2004). If validated by future studies, such 28 29 30 485 differences may indicate dietary or even cognitive differentiation between 31 32 486 closely related taxa (Zimmermann & Radespiel 2014). 33 34 35 487 Conversely, M. lehilahytsara and M. mittermeieri differed in tail length, body 36 37 488 length, third toe length and four head-associated parameters (Figure 3; Table 38 39 40 489 3). This comparison was, however, based on two populations that are over 400 41 42 490 km away from each other. Genomic analyses on samples obtained for M. 43 44 491 lehilahytsara at two intermediate locations (Riamalandy and Ambavala, Figure 45 46 492 1) and M. mittermeieri samples from Anjiahely, Anjanaharibe-Sud SR and 47 48 49 493 Marojejy NP revealed only moderate genomic differentiation along a geographic 50 51 494 gradient (Figure 2; Poelstra et al., 2020). It was concluded that these results do 52 53 495 not justify separate species status of these two taxa. Unfortunately, we only 54 55 56 496 caught three adult M. lehilahytsara at Ambavala making it difficult to assess 57 58 497 whether the individuals from this intermediate geographic location also took an 59 60 498 intermediate morphometric position. Measurements from these three

John Wiley & Sons Page 23 of 66 American Journal of Primatology

Schüßler et al. page 22 1 2 3 4 499 individuals, however, suggest that M. lehilahytsara from Ambavala was more 5 6 7 500 similar to M. mittermeieri from Anjiahely than to M. lehilahytsara from Mantadia 8 9 501 NP (Table 3). If considered as one species, these differences could indicate 10 11 502 morphological adaptations to different environmental conditions (highland 12 13 14 503 versus lowland rainforest) or a morphological gradient across its entire range. 15 16 504 These hypotheses, however, will require further testing. 17 18 19 505 M. simmonsi individuals collected at Ambodiriana fell in between the clusters of 20 21 506 larger and smaller-bodiedFor species Peer in all Reviewanalyses. However, when comparing 22 23 507 these individuals to M. simmonsi sampled further south (Zahamena NP and 24 25 26 508 Betampona SNR; Louis et al., 2006), our individuals were smaller (9.2 vs. 8.9 27 28 509 cm body size), had lower body mass (65 vs. 52 g) and had shorter tails (14.2 29 30 510 vs. 13.1 cm). These differences are even more pronounced when comparing 31 32 33 511 our dataset to the holotype specimen for M. simmonsi (9.8 cm, 77 g and 14.9 34 35 512 cm, respectively; Louis et al. 2006). On the other hand, the holotype specimen 36 37 513 of M. boraha, occurring on Ile St. Marie (an island less than 20 km off the coast 38 39 40 514 of Ambodiriana, Figure 1) had about the same average body mass (56.5 g; 41 42 515 Hotaling et al., 2016) as the population in Ambodiriana. While these 43 44 516 comparisons should be interpreted with caution due to small sample sizes and 45 46 517 different collection details, they suggest that M. simmonsi from Ambodiriana and 47 48 49 518 M. boraha from Ile St. Marie may be more closely related than previously 50 51 519 thought. Genomic analyses revealed only slight differentiation between 52 53 520 southern M. simmonsi and the population at Ambodiriana (Poelstra et al., 2020). 54 55 56 521 However, future analyses should also include samples from M. boraha from Ile 57 58 522 St. Marie to clarify relationships among these populations and lineages. 59 60

John Wiley & Sons American Journal of Primatology Page 24 of 66

Schüßler et al. page 23 1 2 3 4 523 5 6 7 524 Reproductive patterns of Microcebus spp. in northeastern Madagascar 8 9 10 525 Differences in reproductive schedules among mouse lemur species depend on 11 12 13 526 phylogenetic relatedness, although environmental parameters (i.e., rainfall and 14 15 527 temperature) likely play a role in fine tuning reproductive function (Evasoa et al., 16 17 528 2018). Although photoperiod is generally considered the main trigger of 18 19 20 529 reproductive physiological function in mouse lemurs and is a relatively good 21 For Peer Review 22 530 predictor of food availability in seasonal habitats, there is substantial variation in 23 24 531 the timing and duration of reproduction within and across mouse lemur species, 25 26 532 as evident in our study populations. 27 28 29 533 Reproductive observations from this study and those from published sources 30 31 32 534 (Evasoa et al., 2018) suggest that M. sp. #3 may start their mating season in 33 34 535 October-November, because males showed developed testes by 35 36 536 August/September, while females were still anestrous and not lactating (Figure 37 38 39 537 6), and juvenile individuals were not found. This timing of reproduction is 40 41 538 comparable to that of M. lehilahytsara from other sites but interestingly not from 42 43 539 Ambavala, where this species is sympatric with M. sp. #3. In September, the 44 45 two adult male M. lehilahytsara from Ambavala did not have enlarged testes 46 540 47 48 541 and we also captured juvenile M. lehilahytsara with less than half of the adult 49 50 542 body mass (19-23 g). A body mass of about 20 g was found around the time of 51 52 543 weaning in juvenile captive M. lehilahytsara of about seven weeks of age 53 54 55 544 (Radespiel, Zimmermann & Wittkowski, unpubl. data). Adding these seven 56 57 545 weeks (49 days) to about 57 days of gestation (for M. lehilahytsara, 58 59 546 Wrogemann & Zimmermann, 2001; for M. rufus, Blanco, 2008), this could point 60

John Wiley & Sons Page 25 of 66 American Journal of Primatology

Schüßler et al. page 24 1 2 3 4 547 towards a mating season of the M. lehilahytsara in Ambavala potentially lasting 5 6 7 548 until May. 8 9 10 549 These findings contrast strikingly with those from M. lehilahytsara inhabiting 11 12 550 montane and high elevation rainforests. For instance, at Mantadia NP (300 km 13 14 551 further south of Ambavala; Figure 1) all adult males had enlarged testes in 15 16 17 552 September, and some adult females were in estrus in November. Juveniles at 18 19 553 this site (20 g body mass) were only found in late March (Randrianambinina et 20 21 554 al., 2003; this study).For Similar Peer observations Review were made at Ambatovy, a montane 22 23 555 forest (near Mantadia NP) where MBB (unpubl. data) captured two juveniles in 24 25 26 556 late February (~2 months old), suggesting a distinct birth season in mid-late 27 28 557 December. An adult male captured in early March showed evidence of tail 29 30 558 fattening. At Tsinjoarivo (Table S2), another high-altitude forest, M. lehilahytsara 31 32 33 559 females were observed gestating or lactating in late November, December and 34 35 560 early January (Blanco, 2010). From these sites there was also evidence of 36 37 561 rebound polyestry, i.e., females undergo renewed estrus after the loss of 38 39 40 562 offspring or early abortions. The capture of juvenile mouse lemurs (2-3 months 41 42 563 old) in early February, however, suggested a main birth season at this site in 43 44 564 early-mid December. In sum, observations of tail fattening at Tsinjoarivo and 45 46 565 Ambatovy in early March suggest that, at least for a portion of the mouse lemur 47 48 49 566 population, reproductive season is over by this time of the year. 50 51 52 567 Ambavala, unlike Mantadia NP, Ambatovy and Tsinjoarivo, is a low elevation 53 54 568 rainforest site (250 m vs. >800 m a.s.l., respectively). Seasonal climatic 55 56 569 fluctuations are more pronounced in montane rainforests than in the lowland 57 58 59 570 rainforests of eastern Madagascar. For example, Mantadia NP has a lower 60

John Wiley & Sons American Journal of Primatology Page 26 of 66

Schüßler et al. page 25 1 2 3 4 571 mean annual temperature (19°C compared to 23°C), lower minimum 5 6 7 572 temperature (9°C compared to 15°C) and a higher annual temperature range 8 9 573 (19°C compared to 15°C; Fick & Hijmans, 2017) than Ambavala. Climatic 10 11 574 conditions have already been suggested as major determinants of reproductive 12 13 14 575 schedules for small-bodied mouse lemurs (Evasoa et al., 2018) with higher 15 16 576 mean ambient temperatures and smaller temperature fluctuations allowing a 17 18 577 less seasonal reproduction compared to the harsh conditions of the montane 19 20 578 rainforests (Evasoa et al., 2018; Randrianambinina et al., 2003). Taken 21 For Peer Review 22 23 579 together, the findings of this study and previous work suggest a considerable 24 25 580 degree of intraspecific variation concerning reproductive timing in M. 26 27 581 lehilahytsara. 28 29 30 582 Another pair of sympatric mouse lemur species from lowland rainforests 31 32 33 583 (Anjiahely: about 350-400 m a.s.l.) were characterized with regard to 34 35 584 reproductive activities in the northern part of our study region: M. macarthurii 36 37 585 and M. mittermeieri. The enlarged testes of M. macarthurii males from October 38 39 40 586 onwards and the presence of pre-estrous females in November indicate that the 41 42 587 mating season in these forests probably begins at that time (Figure 6). 43 44 588 Furthermore, the absence of juvenile and young mouse lemurs during our 45 46 589 sampling period may indicate one short mating season probably limited to 47 48 49 590 November-December. 50 51 52 591 Sympatric M. mittermeieri females were also found in estrus in November and 53 54 592 the respective males had enlarged testes in September. Moreover, juvenile M. 55 56 593 mittermeieri of about 20 g (~2 months old) were also found in November and 57 58 59 594 December. In analogy to the case of M. lehilahytsara (see above), we therefore 60

John Wiley & Sons Page 27 of 66 American Journal of Primatology

Schüßler et al. page 26 1 2 3 4 595 also predict some mating of M. mittermeieri to occur as early as in July - August 5 6 7 596 in Anjiahely. Reproductive observations of two M. mittermeieri females from 8 9 597 another low elevation forest (Marojejy NP east site, ~200 m a.s.l.; MBB, unpubl. 10 11 598 data) captured in mid-late March, also suggest an extended reproductive period 12 13 14 599 and evidence of polyestry in this species. One of the females showed vaginal 15 16 600 swelling (pre-estrus) and 3 pairs of developed nipples, suggesting she had 17 18 601 given birth earlier in the season and was getting ready to mate again. The 19 20 602 second female showed a vaginal opening (estrus) and well-developed nipples, 21 For Peer Review 22 23 603 also suggesting she had given birth earlier in the season and could, if mating 24 25 604 were successful, have another birth at the end of May or early June. 26 27 28 605 Interestingly, reproductive observations of M. mittermeieri from a montane 29 30 606 rainforest at Anjanaharibe Sud SR (980 m a.s.l; MBB, unpubl. data), were more 31 32 33 607 similar to those of M. lehilahytsara from other montane/high elevation forests 34 35 608 than to conspecifics from nearby low elevation forests: for instance, one adult 36 37 609 female captured in early May showed visible nipples but no signs of recent 38 39 40 610 lactation and evidence of tail fattening, and one adult male showed regressed 41 42 611 testes and tail fattening during the same capture period. Five captured young 43 44 612 mouse lemurs (~2-3 months old) showed no signs of reproductive activity and 45 46 613 no evidence of tail fattening. That a portion of the mouse lemur population is 47 48 49 614 displaying tail fattening in early May is indicative of a shorter reproductive 50 51 615 season, likely over by March. 52 53 54 616 In conclusion, and if supported by future studies, the M. lehilahytsara/M. 55 56 617 mittermeieri lineage in the warmer lowland rainforests of eastern Madagascar 57 58 59 618 seem to have more flexible and extended reproductive schedules compared to 60

John Wiley & Sons American Journal of Primatology Page 28 of 66

Schüßler et al. page 27 1 2 3 4 619 populations living in the harsher montane humid forest sites or to species 5 6 7 620 inhabiting the western dry deciduous forests (see Evasoa et al., 2018). In this 8 9 621 respect, these mouse lemurs show similarities to M. mamiratra and M. 10 11 622 margotmarshae that live in the lowland humid forests of northwestern 12 13 14 623 Madagascar (Sambirano region) and that are also reproductively active in the 15 16 624 first half of the dry season (Evasoa et al. 2018). Reproductive timing of 17 18 625 sympatric non-sister species was different, since the two microendemic species 19 20 626 (M. macarthurii and M. sp. #3) did not show extended reproductive schedules, 21 For Peer Review 22 23 627 while allopatric sister linages showed highly similar schedules in both cases. 24 25 26 628 27 28 29 629 Conclusion and implications for conservation 30 31 32 630 A total of five different Microcebus lineages were demonstrated to occur in a 33 34 631 geographically restricted region of northeastern Madagascar. All of these taxa 35 36 632 inhabit a 130 km wide stretch of lowland rainforest making this region one of the 37 38 39 633 most species-rich areas so far identified for mouse lemurs. These lineages can 40 41 634 be distinguished genetically (Poelstra et al., 2020) and morphologically as 42 43 635 shown in this study. The genomic and phenotypic differentiation between M. 44 45 macarthurii and M. sp. #3 provides sufficient support for the recognition of M. 46 636 47 48 637 sp. #3 as a distinct species which we describe below (see Species Description). 49 50 51 638 The studied taxa were found in a variety of habitat types, ranging from nearly 52 53 639 undisturbed to selectively logged forest, from shrubby secondary re-growth 54 55 640 vegetation to areas dominated by perennial plants (i.e., Aframomum spp.; Miller 56 57 58 641 et al., 2018; Radespiel et al., 2008; Schüßler et al., 2018). However, species 59 60 642 differed most substantially in their altitudinal range and the inferred Extent of

John Wiley & Sons Page 29 of 66 American Journal of Primatology

Schüßler et al. page 28 1 2 3 4 643 Occurrence. M. lehilahytsara and M. mittermeieri were found both in montane 5 6 7 644 and lowland forests, but M. sp. #3 and M. macarthurii occurred only in lowland 8 9 645 forests. Locally restricted so-called “lowland specialists” have been found in at 10 11 646 least three other cases along the Malagasy east coast, i.e., M. gerpi (Radespiel 12 13 14 647 et al., 2012), M. marohita (Rasoloarison et al., 2013) and M. jollyae (Louis et al., 15 16 648 2006). These taxa share a narrow altitudinal range and a small estimated 17 18 649 Extent of Occurrence. Alarmingly, lowland rainforest habitats have disappeared 19 20 650 from most of the east coast and our study region is no exception (Schüßler et 21 For Peer Review 22 23 651 al., 2020; Vieilledent et al., 2018). Under these circumstances, population 24 25 652 declines are unavoidable, and ongoing anthropogenic land use change and 26 27 653 forest cover loss (Schüßler et al., 2020) will probably accelerate and increase 28 29 30 654 this threat to the newly described species. M. sp. #3 occurs in one National 31 32 655 Park, Mananara-Nord NP, which is already isolated from surrounding forests. 33 34 656 Anticipated environmental disruptions due to future climatic changes and the 35 36 37 657 need for species to flexibly adapt to altered environments may be severely 38 39 658 compromised, if conservation planning does not generate and maintain possible 40 41 659 dispersal pathways for migrating species (Brown & Yoder, 2015; Schüßler et al., 42 43 2020). 44 660 45 46 661 47 48 662 5. Species Description 49 50 663 Systematics 51 52 53 664 Order: Primates (Linnaeus 1758) 54 55 56 665 Suborder: Strepshirrini (É. Geoffroy 1812) 57 58 59 666 Family: Cheirogaleidae (Gray 1873) 60

John Wiley & Sons American Journal of Primatology Page 30 of 66

Schüßler et al. page 29 1 2 3 4 667 Genus: Microcebus (É. Geoffroy 1828) 5 6 7 668 Species: Microcebus jonahi species nova 8 9 10 669 11 12 13 670 Holotype 14 15 16 671 B34, adult male, captured on 06 September 2017 by DS. Tissue samples, hair 17 18 19 672 samples as well as e-voucher photos of the animal are stored at the Institute of 20 21 673 Zoology, University ofFor Veterinary Peer Medicine Review Hanover, Germany. The animal itself 22 23 674 was released after field handling, sampling, and photographing, since its 24 25 675 taxonomic distinctiveness was not recognized at the time of capture. Field 26 27 28 676 measurements (all lengths measured in mm): ear length: 17.6, ear width: 13.7, 29 30 677 head length: 37.7, head width: 23.0, snout length: 10.0, intra-orbital distance: 31 32 678 8.2, inter-orbital distance: 26.0, lower leg length: 41.7, hind foot length: 24.5, 33 34 35 679 third toe length: 10.6, body length: 95.6, tail length: 130.0, body mass: 66 g. The 36 37 680 population around Ambavala is designated as the source population for physical 38 39 681 specimens in support of the holotype. 40 41 42 682 43 44 45 683 Type locality 46 47 48 684 Forest near the rural village of Ambavala (S 16° 12.307’, E 49° 35.371’), in a 49 50 685 community protected forest at about 342 m a.s.l. approx. 20 km west of 51 52 53 686 Mananara Avaratra (Mananara-Nord), Province of Analanjirofo, Madagascar. 54 55 56 687 57 58 59 688 Paratypes 60

John Wiley & Sons Page 31 of 66 American Journal of Primatology

Schüßler et al. page 30 1 2 3 4 689 (a) BD1, adult female, captured in the community protected forest of 5 6 7 690 Antsiradrano (near Ambavala) on 04 September 2017. Tissue and hair samples 8 9 691 as well as photographs and morphometric measurements are stored at the 10 11 692 Institute of Zoology, University of Veterinary Medicine Hanover, Germany. 12 13 14 693 (b) B13, adult male, captured in the community protected forest near Ambavala 15 16 17 694 on 11 September 2017. Tissue and hair samples as well as photographs and 18 19 695 morphometric measurements are stored at the Institute of Zoology of the 20 21 696 University of VeterinaryFor Medicine Peer Hanover Review in Germany. 22 23 24 697 It is planned that one physical specimen will be obtained as a further paratype 25 26 698 soon and that this specimen will then be deposited in the Museum of the 27 28 29 699 Zoology Department of the University of Antananarivo, Madagascar. Although 30 31 700 not being a standard procedure, this method is most appropriate for 32 33 701 endangered primates that should not be prematurely sacrificed if the taxonomic 34 35 36 702 assignment is not yet clear. The same procedure was used for the scientific 37 38 703 description of M. gerpi (Radespiel et al., 2012) for which a paratype individual 39 40 704 had been collected during a subsequent field mission and was then deposited 41 42 43 705 at the University of Antananarivo. 44 45 46 706 47 48 707 Description 49 50 51 708 Microcebus jonahi is a large-bodied, reddish-brown and small-eared mouse 52 53 54 709 lemur (Figure 7). This species has short and dense fur. The head is rufous 55 56 710 colored with a darker brownish area around the eyes which can slightly vary 57 58 711 among individuals. A distinct white stripe lies between the eyes ending at the 59 60

John Wiley & Sons American Journal of Primatology Page 32 of 66

Schüßler et al. page 31 1 2 3 4 712 forehead (Figure 7C). The ears are of the same rufous color as the head. The 5 6 7 713 cheeks are lighter brownish and less rufous than the head becoming even 8 9 714 lighter and almost white towards the throat. The ventrum is white with slightly 10 11 715 yellowish nuances (Figure 7D) which can vary in appearance among 12 13 14 716 individuals. The dorsum is rather uniformly brown than reddish (Figure 7E). A 15 16 717 darker dorsal stripe can be either present or absent. The ventrum and dorsum 17 18 718 are separated by a significant change in coloration with only marginal transition. 19 20 719 The coloration of the limbs shows the same pattern with a brownish dorsal and 21 For Peer Review 22 23 720 a white to slightly yellowish ventral side. The tail is densely furred and of the 24 25 721 same coloration as the dorsum. Hands and feet show only sparse but whitish- 26 27 722 gray hair. The skin on the palmar and plantar surfaces of hands and feet is 28 29 30 723 brownish pink. Males and females do not show any sexual dimorphism. 31 32 33 724 34 35 36 725 Habitat information 37 38 39 726 M. jonahi individuals were captured at altitudes between 42 and 356 m a.s.l. 40 41 727 Out of the 25 captured individuals, six were caught in near-primary forest (= low 42 43 728 degradation) with a rather continuous canopy and five were captured in highly 44 45 degraded forests with discontinuous canopy cover and strong regrowth of early 46 729 47 48 730 successional trees. The majority of individuals (N = 14) were captured in 2 - 4 m 49 50 731 high stands of the perennial Madagascar cardamom (Aframomum 51 52 732 angustifolium), sometimes intermixed with trees along the forest edges. At 53 54 55 733 Antanambe (Figure 1), a total of 39 individuals were sighted of which 35 were 56 57 734 found in forest habitats of different degradation stages. No M. jonahi were 58 59 735 sighted in treeless secondary vegetation except for dense Aframomum 60

John Wiley & Sons Page 33 of 66 American Journal of Primatology

Schüßler et al. page 32 1 2 3 4 736 angustifolium habitats. It currently inhabits one protected area (Mananara-Nord 5 6 7 737 NP) and a community managed forest area around the village of Ambavala 8 9 738 (Schüßler et al., 2018). 10 11 12 739 13 14 15 740 Diagnosis 16 17 18 741 M. jonahi can be distinguished from other taxa in northeastern Madagascar by 19 20 742 morphometric features and genomic distinctiveness. Compared to its closest 21 For Peer Review 22 relative, M. macarthurii, M. jonahi is longer, has a shorter tail, wider ears, a 23 743 24 25 744 larger head width and a shorter head length. In addition, M. jonahi can be 26 27 745 differentiated from M. macarthurii by its ventral coloration which is rather whitish 28 29 746 (Figure 6), but distinctly yellowish orange in M. macarthurii (Radespiel et al., 30 31 32 747 2008; Radespiel & Raveloson, unpubl. data). 33 34 35 748 Moreover, it can be easily distinguished from the sympatric, small-bodied M. 36 37 749 lehilahytsara (at Ambavala) by its higher body mass, larger body size and 38 39 750 longer tail length. Finally, M. jonahi can be differentiated from its southern 40 41 42 751 geographical neighbor, M. simmonsi, by its shorter ear length and its larger 43 44 752 inter- and intra-orbital distances. M. jonahi could be unambiguously 45 46 753 distinguished from the other four taxa in this study across all analyses of 47 48 754 nuclear RADseq data (Poelstra et al., 2020). However, it may not be reliably 49 50 51 755 distinguished from M. macarthurii based solely on mitochondrial sequences, 52 53 756 likely due to some introgression from M. jonahi into M. macarthurii in the past 54 55 757 (Poelstra et al., 2020). 56 57 58 758 59 60

John Wiley & Sons American Journal of Primatology Page 34 of 66

Schüßler et al. page 33 1 2 3 4 759 Etymology 5 6 7 760 M. jonahi is named in honor of Malagasy primatologist Professor Jonah 8 9 10 761 Ratsimbazafy. He has dedicated his life's work to the conservation of Malagasy 11 12 762 lemurs. With both national and international outreach to the scientific community 13 14 763 (e.g., GERP, IPS, LemursPortal), to the public of Madagascar (e.g., by initiating 15 16 17 764 the World Lemur Festival), and to the political leaders of Madagascar, he serves 18 19 765 as an inspirational role model for young Malagasy students and scientists. He 20 21 766 provides hope for theFor future ofPeer Madagascar Review and for its iconic lemurs during very 22 23 767 challenging times. 24 25 26 768 27 28 29 769 Vernacular name 30 31 32 770 English name: Jonah’s mouse lemur, French name: Microcèbe de Jonah, 33 34 35 771 German name: Jonah’s Mausmaki. 36 37 38 772 39 40 773 6. Acknowledgments 41 42 43 774 This study was conducted under the research permit No. 44 45 775 197/17/MEEF/SG/DGF/DSAP/SCB.Re (DS), 46 47 072/15/MEEMF/SG/DGF/DCB.SAP/SCB (MBB), 48 776 49 50 777 137/13/MEF/SG/DGF/DCB.SAP/SCB (DWR) and 51 52 53 778 175/14/MEF/SG/DGF/DCB.SAP/SCB (AM), kindly issued by the directeur du 54 55 779 système des aires protégées, Antananarivo and the regional authorities 56 57 780 (Direction Régional de l’Environnement, de l’Ecologie et de Forêts). Our biggest 58 59 60 781 thanks go to the local authorities who allowed us to conduct our study in their

John Wiley & Sons Page 35 of 66 American Journal of Primatology

Schüßler et al. page 34 1 2 3 4 782 forests and furthermore to the Wildlife Conservation Society Madagascar for 5 6 7 783 valuable support when working in and around Makira Natural Park. 8 9 10 784 We are indebted to J.H. Ratsimbazafy, N.V. Andriaholinirina, C. Misandeau, B. 11 12 785 Le Pors and S. Rasoloharijaona, for their help with administrative tasks and to 13 14 786 G. Besnard and G. Tiley for facilitating this study. We thank our field assistants 15 16 17 787 (I. Sitrakarivo, C. Hanitriniaina and T. Ralantoharijaona) and the ADAFAM 18 19 788 (Association Des Amis de la Forêt d'Ambodiriana-Manompana, in particular C 20 21 789 Misandeau) for theirFor valuable Peer help during Review sample collection. We warmly thank 22 23 790 the many local guides and cooks for sharing their incomparable expertise and 24 25 26 791 help in the field, misaotra anareo jiaby. We finally would like to thank the editor 27 28 792 and the two referees for their positive and constructive comments that helped to 29 30 793 clarify the manuscript and improve its quality. 31 32 33 794 Funding was granted by the Bauer Foundation and the Zempelin Foundation of 34 35 36 795 the "Deutsches Stiftungszentrum" under grant no. T237/22985/2012/kg and 37 38 796 T0214/32083/2018/sm to DS, Duke Tropical Conservation Initiative Grant to 39 40 797 ADY, and Duke Lemur Center/SAVA Conservation research funds to MBB, the 41 42 43 798 School of Animal Biology at The University of Western Australia to AM, the 44 45 799 Fundação para a Ciência e a Tecnologia, Portugal (PTDC/BIA- 46 47 800 BEC/100176/2008, PTDC/BIA-BIC/4476/2012, and SFRH/BD/64875/2009), the 48 49 801 Groupement de Recherche International (GDRI) Biodiversité et développement 50 51 52 802 durable – Madagascar, the Laboratoire d’Excellence (LABEX) TULIP (ANR-10- 53 54 803 LABX-41) and CEBA (ANR-10-LABX-25-01), the Instituto Gulbenkian de 55 56 804 Ciência, Portugal to LC and JS, the ERA-NET BiodivERsA project: 57 58 59 805 INFRAGECO (Inference, Fragmentation, Genomics, and Conservation, ANR- 60

John Wiley & Sons American Journal of Primatology Page 36 of 66

Schüßler et al. page 35 1 2 3 4 806 16-EBI3-0014 & FCT-Biodiversa/0003/2015) the LIA BEEG-B (Laboratoire 5 6 7 807 International Associé – Bioinformatics, Ecology, Evolution, Genomics and 8 9 808 Behaviour, CNRS) to LC and JS. Further financial support came from the 10 11 809 Institute of Zoology, University of Veterinary Medicine Hannover and UR 12 13 14 810 acknowledges the long-term support of the late Elke Zimmermann for her 15 16 811 research activities on Madagascar. ADY also gratefully acknowledges support 17 18 812 from the John Simon Guggenheim Memorial Foundation and the Alexander von 19 20 813 Humboldt Foundation. EELJ would like to acknowledge support from the 21 For Peer Review 22 23 814 Ahmanson Foundation for the field sample collection and subsequent data 24 25 815 generation. This work was performed in collaboration with the GeT core facility, 26 27 816 Toulouse, France (http://get.genotoul.fr), and was supported by France 28 29 30 817 Génomique National infrastructure, funded as part of “Investissement d’avenir” 31 32 818 program managed by Agence Nationale pour la Recherche (contract ANR-10- 33 34 819 INBS-09). All co-authors declare that they have no conflict of interest. 35 36 37 820 38 39 40 821 Data availability statement: 41 42 43 822 The data that support the findings of this study are added in the supplementary 44 45 files. 46 823 47 48 824 49 50 51 825 7. References 52 53 54 826 Abdi, H. & Williams, L.J. (2010). Principal component analysis. Wiley 55 56 57 827 Interdisciplinary Reviews: Computational Statistics, 2(4), 433–459. 58 59 828 https://doi.org/10.1002/wics.101 60

John Wiley & Sons Page 37 of 66 American Journal of Primatology

Schüßler et al. page 36 1 2 3 4 829 American Society of Primatologists (2001). Principles for the ethical treatment of 5 6 7 830 Non-Human primates. 8 9 831 https://www.asp.org/society/resolutions/EthicalTreatmentOfNonHumanPrim 10 11 832 ates.cfm 12 13 14 833 Anderson, M.J. (2017) Permutational multivariate analysis of variance 15 16 17 834 (PERMANOVA) Wiley statsref: statistics reference online, 1-15. 18 19 835 https://doi.org/10.1002/9781118445112.stat07841 20 21 For Peer Review 22 836 Andriaholinirina, N., Baden, A., Blanco, M., Chikhi, L., Cooke, A., Davies, N., … 23 24 837 Zaramody, A. (2014) Microcebus mittermeieri. The IUCN Red List of 25 26 838 Threatened Species 2014: e.T136272A16113275. 27 28 29 839 https://dx.doi.org/10.2305/IUCN.UK.2014-1.RLTS.T136272A16113275.en. 30 31 32 840 Balakrishnama, S. & Ganapathiraju, A. (1998). Linear discriminant analysis - a 33 34 841 brief tutorial. Institute for Signal and Information Processing, 18, 1–8. 35 36 37 842 Beaudrot, L., Struebig, M.J., Meijaard, E., van Balen, S., Husson, S. & Marshall, 38 39 843 A.J. (2013). Co-occurrence patterns of Bornean vertebrates suggest 40 41 42 844 competitive exclusion is strongest among distantly related species. 43 44 845 Oecologia, 173(3), 1053–1062. https://doi.org/10.1007/s00442-013-2679-7 45 46 47 846 Blanco, M.B. (2008). Reproductive schedules of female Microcebus rufus at 48 49 847 Ranomafana National Park, Madagascar. International Journal of 50 51 848 Primatology, 29(2), 323–338. https://doi.org/10.1007/s10764-008-9238-9 52 53 54 849 Blanco MB. 2010. Reproductive biology of mouse and dwarf lemurs of eastern 55 56 57 850 Madagascar, with an emphasis on brown mouse lemurs (Microcebus rufus) 58 59 60

John Wiley & Sons American Journal of Primatology Page 38 of 66

Schüßler et al. page 37 1 2 3 4 851 at Ranomafana National Park, a southeastern rainforest, Ph.D. dissertation, 5 6 7 852 University of Massachusetts, Amherst. 8 9 10 853 Dammhahn, M. & Kappeler, P.M. (2008). Comparative feeding ecology of 11 12 854 sympatric Microcebus berthae and M. murinus. International Journal of 13 14 855 Primatology, 29(6), 1567. https://doi.org/10.1007/s10764-008-9312-3 15 16 17 856 Estrada, A., Garber, P.A., Rylands, A.B., Roos, C., Fernandez-Duque, E., Di 18 19 20 857 Fiore, A., … Li, B. (2017). Impending extinction crisis of the worlds 21 For Peer Review 22 858 primates: Why primates matter. Science Advances, 3(1), e1600946. 23 24 859 https://doi.org/10.1126/sciadv.1600946 25 26 27 860 Evasoa, R.M., Radespiel, U., Hasiniaina, A.F., Rasoloharijaona, S., 28 29 861 Randrianambinina, B., Rakotondravony, R. & Zimmermann, E. (2018). 30 31 32 862 Variation in reproduction of the smallest-bodied primate radiation, the 33 34 863 mouse lemurs (Microcebus spp.): A synopsis. American Journal of 35 36 864 Primatology, e22874. https://doi.org/10.1002/ajp.22874 37 38 39 865 Fabre, A.C., Perry, J.M., Hartstone-Rose, A., Lowie, A., Boens, A. & Dumont, 40 41 42 866 M. (2018). Do muscles constrain skull shape evolution in strepsirrhines? 43 44 867 The Anatomical Record, 301(2), 291–310. https://doi.org/10.1002/ar.23712 45 46 47 868 Fick, S.E. & Hijmans, R.J. (2017). Worldclim 2: new 1-km spatial resolution 48 49 869 climate surfaces for global land areas. International Journal of Climatology, 50 51 870 37(12), 4302–4315. https://doi.org/10.1002/joc.5086 52 53 54 871 Fox, J. & Weisberg, S. (2011). An R Companion to Applied Regression. 55 56 57 872 Thousand Oaks CA: Sage, 2nd Edition. 58 59 873 http://socserv.socsci.mcmaster.ca/jfox/Books/Companion 60

John Wiley & Sons Page 39 of 66 American Journal of Primatology

Schüßler et al. page 38 1 2 3 4 874 Hafen, T., Neveu, H., Rumpler, Y., Wilden, I. & Zimmermann, E. (1998). 5 6 7 875 Acoustically dimorphic advertisement calls separate morphologically and 8 9 876 genetically homogenous populations of the grey mouse lemur (Microcebus 10 11 877 murinus). Folia Primatologica, 69(Suppl. 1), 342–356. 12 13 14 878 https://doi.org/10.1159/000052723 15 16 17 879 Hardin, G. (1960). The competitive exclusion principle. Science, 131(3409), 18 19 880 1292–1297 20 21 For Peer Review 22 881 Hotaling, S., Foley, M.E., Lawrence, N.M., Bocanegra, J., Blanco, M.B., 23 24 882 Rasoloarison, R.M., … Weisrock, D.W. (2016). Species discovery and 25 26 883 validation in a cryptic radiation of endangered primates: coalescent-based 27 28 29 884 species delimitation in Madagascar’s mouse lemurs. Molecular Ecology, 25, 30 31 885 2029–2045. https://doi.org/10.1111/mec.13604 32 33 34 886 IUCN (2012). IUCN Red List categories and criteria. IUCN, 2nd Edition. 35 36 37 887 Kamilar, J.M., Blanco, M.B. & Muldoon, K.M. (2016). Ecological niche modeling 38 39 888 of mouse lemurs (Microcebus spp.) and its implications for their species 40 41 42 889 diversity and biogeography. In S. Lehman, E. Zimmermann & U. Radespiel 43 44 890 (Editors), The Dwarf and Mouse Lemurs of Madagascar: Biology, Behavior, 45 46 891 and Conservation Biogeography of the Cheirogaleidae. (pp. 451–463). 47 48 892 Cambridge, UK: University Press Cambridge 49 50 51 893 Kappeler, P., Rasoloarison, R., Razafimanantsoa, L., Walter, L. & Roos, C. 52 53 54 894 (2005). Morphology, behavior and molecular evolution of giant mouse 55 56 895 lemurs (Mirza spp.) Gray, 1870, with description of a new species. Primate 57 58 896 Report, 71, 3–26 59 60

John Wiley & Sons American Journal of Primatology Page 40 of 66

Schüßler et al. page 39 1 2 3 4 897 Knoop, S., Chikhi, L. & Salmona, J. (2018). Mouse lemurs use of degraded 5 6 7 898 habitat. Lemur News, 21, 20–31 8 9 10 899 Kottek, M., Grieser, J., Beck, C., Rudolf, B. & Rubel, F. (2006). World map of 11 12 900 the Köppen-Geiger climate classification updated. Meteorologische 13 14 901 Zeitschrift, 15(3), 259–263. https://doi.org/10.1127/0941-2948/2006/0130 15 16 17 902 Kraus, C., Eberle, M. & Kappeler, P.M. (2008). The costs of risky male 18 19 20 903 behaviour: sex differences in seasonal survival in a small sexually 21 For Peer Review 22 904 monomorphic primate. Proceedings of the Royal Society B: Biological 23 24 905 Sciences, 275(1643), 1635–1644. https://doi.org/10.1098/rspb.2008.0200 25 26 27 906 Lei, R., Frasier, C.L., Hawkins, M.T., Engberg, S.E., Bailey, C.A., Johnson, 28 29 907 S.E., … Louis, E.E. (2016). Phylogenomic reconstruction of sportive lemurs 30 31 32 908 (genus Lepilemur) recovered from mitogenomes with inferences for 33 34 909 Madagascar biogeography. Journal of Heredity, 108(2), 107–119. 35 36 910 https://doi.org/10.1093/jhered/esw072 37 38 39 911 Louis, E.E., Coles, M.S., Andriantompohavana, R., Sommer, J.A., Engberg, 40 41 42 912 S.E., Zaonarivelo, J.R., … Brenneman, R.A. (2006). Revision of the mouse 43 44 913 lemurs (Microcebus) of eastern Madagascar. International Journal of 45 46 914 Primatology, 27(2), 347–389. https://doi.org/10.1007/s10764-006-9036-1 47 48 49 915 Louis, E.E., Engberg, S.E., McGuire, S.M., McCormick, M.J., 50 51 916 Randriamampionona, R., Ranaivoarisoa, J.F., … Lei, R. (2008). Revision of 52 53 54 917 the mouse lemurs, Microcebus (Primates, Lemuriformes), of northern and 55 56 918 northwestern Madagascar with descriptions of two new species at 57 58 59 60

John Wiley & Sons Page 41 of 66 American Journal of Primatology

Schüßler et al. page 40 1 2 3 4 919 Montagne d’Ambre National Park and Antafondro Classified Forest. Primate 5 6 7 920 Conservation, 23(1), 19–38. https://doi.org/10.1896/052.023.0103 8 9 10 921 Martin, R. (1972). A preliminary field-study of the lesser mouse lemur 11 12 922 (Microcebus murinus JF Miller 1777). Zeitschrift für Tierpsychologie, 9, 43– 13 14 923 89 15 16 17 924 Meloro, C., Cáceres, N.C., Carotenuto, F., Sponchiado, J., Melo, G.L., Passaro, 18 19 20 925 F. & Raia, P. (2015). Chewing on the trees: Constraints and adaptation in 21 For Peer Review 22 926 the evolution of the primate mandible. Evolution, 69(7), 1690–1700. 23 24 927 https://doi.org/10.1111/evo.12694 25 26 27 928 Miller, A., Mills, H., Ralantoharijaona, T., Volasoa, N.A., Misandeau, C., Chikhi, 28 29 929 L., Bencini, R. & Salmona, J. (2018). Forest type influences population 30 31 32 930 densities of nocturnal lemurs in Manompana, northeastern Madagascar. 33 34 931 International Journal of Primatology, 39(4), 646–669. 35 36 932 https://doi.org/10.1007/s10764-018-0055-5 37 38 39 933 Myers, N., Mittermeier, R.A., Mittermeier, C.G., Da Fonseca, G.A. & Kent, J. 40 41 42 934 (2000). Biodiversity hotspots for conservation priorities. Nature, 403(6772), 43 44 935 853–858. https://doi.org/10.1038/35002501 45 46 47 936 Oksanen, J., Blanchet, F.G., Friendly, M., Kindt, R., Legendre, P., McGlinn, D., 48 49 937 …, Wagner, H. (2019) vegan: Community Ecology Package. 50 51 938 https://CRAN.R-project.org/package=vegan 52 53 54 939 Olivieri, G., Zimmermann, E., Randrianambinina, B., Rasoloharijaona, S., 55 56 57 940 Rakotondravony, D., Guschanski, K. & Radespiel, U. (2007). The ever- 58 59 941 increasing diversity in mouse lemurs: three new species in north and 60

John Wiley & Sons American Journal of Primatology Page 42 of 66

Schüßler et al. page 41 1 2 3 4 942 northwestern Madagascar. Molecular Phylogenetics and Evolution, 43(1), 5 6 7 943 309–327. https://doi.org/10.1016/j.ympev.2006.10.026 8 9 10 944 Padial, J.M., Miralles, A., De la Riva, I. Vences, M. (2010). The integrative 11 12 945 future of taxonomy. Frontiers in Zoology, 7(1), 16. 13 14 946 https://doi.org/10.1186/1742-9994-7-16 15 16 17 947 Poelstra, J., Salmona, J., Tiley, G.P., Schüßler, D., Blanco, M.B., 18 19 20 948 Andriambeloson, J.B., … Yoder, A.D. (2020). Cryptic patterns of speciation 21 For Peer Review 22 949 in cryptic primates: microendemic mouse lemurs and the multispecies 23 24 950 coalescent. BioRxiv. https://doi.org/10.1101/742361 25 26 27 951 R Core Team (2019). R: A Language and Environment for Statistical 28 29 952 Computing. R Foundation for Statistical Computing, Vienna, Austria. URL 30 31 32 953 https://www.R-project.org/ 33 34 35 954 RStudio Team (2016). RStudio: Integrated Development Environment for R. 36 37 955 RStudio Inc., Boston MA, USA. URL: http://www.rstudio.com/ 38 39 956 Radespiel, U. (2016). Can behavioral ecology help to understand the divergent 40 41 42 957 geographic range size of mouse lemurs? In S.M. Lehman, U. Radespiel & 43 44 958 E. Zimmermann (Editors), The Dwarf and Mouse Lemurs of Madagascar: 45 46 959 Biology, Behavior and Conservation Biogeography of the Cheirogaleidae. 47 48 960 (pp. 498–519). Cambridge, UK: Cambridge University Press 49 50 51 961 Radespiel, U., Olivieri, G., Rasolofoson, D.W., Rakotondratsimba, G., 52 53 54 962 Rakotonirainy, O., Rasoloharijaona, S., … Randrianarison, R.M. (2008). 55 56 963 Exceptional diversity of mouse lemurs (Microcebus spp.) in the Makira 57 58 59 60

John Wiley & Sons Page 43 of 66 American Journal of Primatology

Schüßler et al. page 42 1 2 3 4 964 region with the description of one new species. American Journal of 5 6 7 965 Primatology, 70(11), 1033–1046. https://doi.org/10.1002/ajp.20592 8 9 10 966 Radespiel, U., Ratsimbazafy, J.H., Rasoloharijaona, S., Raveloson, H., 11 12 967 Andriaholinirina, N., Rakotondravony, R., … Randrianambinina, B. (2012). 13 14 968 First indications of a highland specialist among mouse lemurs (Microcebus 15 16 17 969 spp.) and evidence for a new mouse lemur species from eastern 18 19 970 Madagascar. Primates, 53(2), 157–170. https://doi.org/10.1007/s10329- 20 21 971 011-0290-2 For Peer Review 22 23 24 972 Rakotondravony, R. & Radespiel, U. (2009). Varying patterns of coexistence of 25 26 973 two mouse lemur species (Microcebus ravelobensis and M. murinus) in a 27 28 29 974 heterogeneous landscape. American Journal of Primatology, 71(11), 928– 30 31 975 938. https://doi.org/10.1002/ajp.20732 32 33 34 976 Randrianambinina, B. (2001). Contribution à l’etude comparative de 35 36 977 l’ecoethologie de deux microcebes rouges de Madagascar: Microcebus 37 38 39 978 ravelobensis (Zimmermann et al., 1998) Microcebus rufus (Lesson, 1840). 40 41 979 Ph.D. Thesis, University of Antananarivo 42 43 44 980 Randrianambinina, B., Rakotondravony, D., Radespiel, U. & Zimmermann, E. 45 46 981 (2003). Seasonal changes in general activity, body mass and reproduction 47 48 982 of two small nocturnal primates: a comparison of the golden brown mouse 49 50 51 983 lemur (Microcebus ravelobensis) in Northwestern Madagascar and the 52 53 984 brown mouse lemur (Microcebus rufus) in Eastern Madagascar. Primates, 54 55 985 44(4), 321–331. https://doi.org/ 10.1007/s10329-003-0046-8 56 57 58 59 60

John Wiley & Sons American Journal of Primatology Page 44 of 66

Schüßler et al. page 43 1 2 3 4 986 Rasoloarison, R.M., Goodman, S.M. & Ganzhorn, J.U. (2000). Taxonomic 5 6 7 987 revision of mouse lemurs (Microcebus) in the western portions of 8 9 988 Madagascar. International Journal of Primatology, 21(6), 963–1019. 10 11 989 https://doi.org/10.1023/A:1005511129475 12 13 14 990 Rasoloarison, R.M., Weisrock, D.W., Yoder, A.D., Rakotondravony, D. & 15 16 17 991 Kappeler, P.M. (2013). Two new species of mouse lemurs (Cheirogaleidae: 18 19 992 Microcebus) from eastern Madagascar. International Journal of 20 21 993 Primatology, 34(3),For 455–469. Peer https://doi.org/10.1007/s10764-013-9672-1 Review 22 23 24 994 Riley, E.P., MacKinnon, K.C., Fernandez-Duque, E., Setchell, J.M. & Garber, 25 26 995 P.A. (2014). Code of best practices for field primatology. International 27 28 29 996 Primatological Society and American Society of Primatologists Steering 30 31 997 Committee. URL 32 33 998 http://www.internationalprimatologicalsociety.org/docs/Code%20of_Best_Pr 34 35 36 999 actices%20Oct%202014.pdf 37 38 39 1000 Schmelting, B., Zimmermann, E., Berke, O., Bruford, M.W. & Radespiel, U. 40 41 1001 (2007). Experience-dependent recapture rates and reproductive success in 42 43 1002 male grey mouse lemurs (Microcebus murinus). American Journal of 44 45 Physical Anthropology, 133(1), 743–752. https://doi.org/10.1002/ajpa.20566 46 1003 47 48 1004 Schneider, N., Chikhi, L., Currat, M., & Radespiel, U. (2010). Signals of recent 49 50 51 1005 spatial expansions in the grey mouse lemur (Microcebus murinus). BMC 52 53 1006 Evolutionary Biology, 10(1), 105. https://doi.org/10.1186/1471-2148-10-105 54 55 56 1007 Schüßler, D., Mantilla-Contreras, J., Stadtmann, R., Ratsimbazafy, J.H. & 57 58 1008 Radespiel, U. (2020). Identification of crucial stepping stone habitats for 59 60

John Wiley & Sons Page 45 of 66 American Journal of Primatology

Schüßler et al. page 44 1 2 3 4 1009 biodiversity conservation in northeastern Madagascar using remote sensing 5 6 7 1010 and comparative predictive modeling. Biodiversity and Conservation. 8 9 1011 https://doi.org/10.1007/s10531-020-01965-z 10 11 12 1012 Schüßler, D., Radespiel, U., Ratsimbazafy, J.H. & Mantilla-Contreras, J. (2018). 13 14 1013 Lemurs in a dying forest: Factors influencing lemur diversity and distribution 15 16 17 1014 in forest remnants of north-eastern Madagascar. Biological Conservation, 18 19 1015 228, 17–26. https://doi.org/10.1016/j.biocon.2018.10.008 20 21 For Peer Review 22 1016 Sgarlata, G. M., Salmona, J., Le Pors, B., Rasolondraibe, E., Jan, F., 23 24 1017 Ralantoharijaona, T., ... & Chikhi, L. (2019). Genetic and morphological 25 26 1018 diversity of mouse lemurs (Microcebus spp.) in northern Madagascar: The 27 28 29 1019 discovery of a putative new species? American Journal of Primatology, 30 31 1020 e23070. https://doi.org/10.1002/ajp.23070 32 33 34 1021 Thorén, S., Linnenbrink, M. & Radespiel, U. (2011). Different competitive 35 36 1022 potential in two coexisting mouse lemur species in northwestern 37 38 39 1023 Madagascar. American Journal of Physical Anthropology, 145(1), 156–162. 40 41 1024 https://doi.org/10.1002/ajpa.21516 42 43 44 1025 Venables, W.N. & Ripley, B.D. (2002). Modern Applied Statistics with S. New 45 46 1026 York: Springer, 4th Edition. URL http://www.stats.ox.ac.uk/pub/MASS4, 47 48 1027 iSBN 0-387-95457-0 49 50 51 1028 Vieilledent, G., Grinand, C., Rakotomalala, F.A., Ranaivosoa, R., 52 53 54 1029 Rakotoarijaona, J.R., Allnutt, T.F. & Achard, F. (2018). Combining global 55 56 1030 tree cover loss data with historical national forest-cover maps to look at six 57 58 1031 decades of deforestation and forest fragmentation in Madagascar. 59 60

John Wiley & Sons American Journal of Primatology Page 46 of 66

Schüßler et al. page 45 1 2 3 4 1032 Biological Conservation, 222, 189–197. 5 6 7 1033 https://doi.org/10.1016/j.biocon.2018.04.008 8 9 10 1034 Viguier, B. (2004). Functional adaptations in the craniofacial morphology of 11 12 1035 Malagasy primates: shape variations associated with gummivory in the 13 14 1036 family Cheirogaleidae. Annals of Anatomy - Anatomischer Anzeiger, 186(5- 15 16 17 1037 6), 495–501. https://doi.org/10.1016/S0940-9602(04)80093-1 18 19 20 1038 Weisrock, D.W., Rasoloarison, R.M., Fiorentino, I., Ralison, J.M., Goodman, 21 For Peer Review 22 1039 S.M., Kappeler, P.M. & Yoder, A.D. (2010). Delimiting species without 23 24 1040 nuclear monophyly in Madagascar’s mouse lemurs. PLoS One, 5(3), e9883. 25 26 1041 https://doi.org/10.1371/journal.pone.0009883 27 28 29 1042 Wrogemann, D. & Zimmermann, E. (2001). Aspects of reproduction in the 30 31 32 1043 eastern rufous mouse lemur (Microcebus rufus) and their implications for 33 34 1044 captive management. Zoo Biology, 20(3), 157–167. 35 36 1045 https://doi.org/10.1002/zoo.1017 37 38 39 1046 Yoder, A.D., Campbell, C.R., Blanco, M.B., dos Reis, M., Ganzhorn, J.U., 40 41 42 1047 Goodman, S.M., … Weisrock, D.W. (2016). Geogenetic patterns in mouse 43 44 1048 lemurs (genus Microcebus) reveal the ghosts of Madagascar’s forests past. 45 46 1049 Proceedings of the National Academy of Sciences, 113(29), 8049–8056. 47 48 1050 https://doi.org/10.1073/pnas.1601081113 49 50 51 1051 Yoder, A.D., Rasoloarison, R.M., Goodman, S.M., Irwin, J.A., Atsalis, S., 52 53 54 1052 Ravosa, M.J. & Ganzhorn, J.U. (2000). Remarkable species diversity in 55 56 1053 Malagasy mouse lemurs (Primates, Microcebus). Proceedings of the 57 58 1054 National Academy of Sciences, 97(21), 11325–11330. 59 60

John Wiley & Sons Page 47 of 66 American Journal of Primatology

Schüßler et al. page 46 1 2 3 4 1055 Zimmermann, E., Cepok, S., Rakotoarison, N., Zietemann, V. & Radespiel, U. 5 6 7 1056 (1998). Sympatric mouse lemurs in north-west Madagascar: A new rufous 8 9 1057 mouse lemur species (Microcebus ravelobensis). Folia Primatologica, 69(2), 10 11 1058 106–114 12 13 14 1059 Zimmermann, E. & Radespiel, U. (2014). Species concepts, diversity, and 15 16 17 1060 evolution in primates: Lessons to be learned from mouse lemurs. 18 19 1061 Evolutionary Anthropology: Issues, News, and Reviews, 23(1), 11–14. 20 21 1062 https://doi.org/10.1002/evan.21388For Peer Review 22 23 24 1063 25 1064 26 27 1065 28 29 30 1066 31 32 1067 33 34 1068 35 36 1069 37 38 39 1070 40 41 1071 42 43 1072 44 45 1073 46 47 48 1074 49 50 1075 51 52 1076 53 54 1077 55 56 57 1078 58 59 1079 60

John Wiley & Sons American Journal of Primatology Page 48 of 66

Schüßler et al. page 47 1 2 3 4 1080 5 6 7 1081 8 9 1082 Tables: 10 11 1083 12 13 1084 Table 1: Age classes of Microcebus spp. based on their mean body mass in g 14 15 1085 (± standard deviation). Intraspecific differences between age classes were 16 17 1086 highly significant in all cases (two-tailed t-tests: P < 0.001). For M. macarthurii 18 19 1087 and M. simmonsi only adult individuals were caught. 20 Species M. sp. #3 M. lehilahytsara M. lehilahytsara M. mittermeieri 21 For Peer Review 22 (Ambavala- (Ambavala) (Mantadia NP) (Anjiahely) 23 Antanambe) 24 Capture time Aug.-Sept. Sept. May-Nov. Sept.-Dec. 25 26 Adults 57.4 ± 5.6 43.3 ± 5.5 43.8 ± 4.7 45.1 ± 6.2 27 (N = 18) (N = 3) (N = 30) (N = 22) 28 29 Young 45.9 ± 2.5 24.4 ± 5.3 31.2 ± 4.0 21.5 ± 1.9 30 (N = 7) (N = 5) (N = 12) (N = 4) 31 32 1088 33 34 1089 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

John Wiley & Sons Page 49 of 66 American Journal of Primatology

Schüßler et al. page 48 1 2 3 4 1090 Table 2: Occurrence locations, altitudinal range (m a.s.l.) and estimated Extent 5 6 1091 of Occurrence (EOO in km² as defined by the IUCN, 2012; see Figure 1) of 7 8 1092 Microcebus spp. in northeastern Madagascar. Combined EOO for M. 9 10 1093 lehilahytsara and M. mittermeieri is estimated with 66,800 km². Coordinates of 11 1094 the locations are given in Supplementary Table S2. 12 13 Taxon Locations Altitude EOO References 14 15 M. mittermeieri Marojejy NP, 350-1,056 9,250 Louis et al., 2006; Radespiel et 16 Anjanaharibe-Sud SR, al., 2008; Weisrock et al., 2010; 17 Anjiahely this study 18 19 M. macarthurii Anjiahely 350-400 560 Radespiel et al., 2008; this 20 study 21 M. lehilahytsara Ambavala,For Riamalandy, Peer 233-1,552 Review50,700 Kappeler et al., 2005; Weisrock 22 23 Mantadia NP, et al., 2010; Yoder et al., 2016; 24 Ambohitantely, this study 25 Ankofabe, Tsinjoarivo 26 M. sp. #3 Ambavala, Mananara- 42-356 1,500 This study 27 28 Nord NP, Antanambe 29 M. simmonsi Ambodiriana, Tampolo, 19-956 13,250 Louis et al., 2006; this study 30 Zahamena NP, 31 32 Betampona SNR 33 1095 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

John Wiley & Sons American Journal of Primatology Page 50 of 66

Schüßler et al. page 49 1 2 3 4 1096 Table 3: Morphometric measurements of Microcebus spp. in northeastern Madagascar (mean ± standard deviation). 5 6 1097 Comparisons are based on one-way ANOVA (P < 0.001 for all parameters) and grouping (letters after values) according to Tukey 7 8 1098 post-hoc tests. Sample sizes per site for M. sp. #3 are N = 5 at Ambavala, N = 7 at Mananara-Nord NP, N = 6 at Antanambe. §: 9 10 1099 N = 11 for M. sp. #3. 11 12 Variables M. sp. #3 M. macarthurii M. simmonsi M. mittermeieri M. lehilahytsara M. lehilahytsara 13 (in mm) (N = 18, at Ambavala, (N = 14, at Anjiahely, this (N = 8, at Ambodiriana, this (N = 22, at Anjiahely, (N = 29, at Mantadia (N = 3, at Ambavala, 14 Mananara-Nord NP and study andFor Radespiel et al.,Peer study) ReviewRadespiel et al., 2008) NP, this study) 15 Antanambe, this study) 2008) Randrinambinina, 16 2001) 17 Ear length 17.2 ± 0.8 a 18.0 ± 0.8 a,c 19.5 ± 2.7 b,c 18.3 ± 1.4 a,c 20.0 ± 1.1 b 17.9 ± 0.2 18 19 Ear width§ 12.7 ± 1.0 a 10.6 ± 1.3 b 12.6 ± 0.9 a 12.1 ± 0.8 a 12.3 ± 0.9 a 11.8 ± 1.1 20 21 Head length 36.2 ± 1.4 a 37.5 ± 1.3 b 35.3 ± 0.9 a,c 34.3 ± 0.8 c 33.2 ± 1.0 d 35.2 ± 1.3 22 Head width 21.5 ± 0.9 a 20.0 ± 1.5 b 20.4 ± 0.2 a,c 19.3 ± 0.7 b,c 18.6 ± 1.0 c 20.0 ± 1.2 23 24 Snout length 10.7 ± 1.2 a 9.6 ± 1.4 a 10.0 ± 1.8 a 7.8 ± 0.9 b 4.9 ± 0.6 c 9.6 ± 0.7 25 Interorbital dist.§ 24.7 ± 1.1 a 23.6 ± 1.5 a,b 22.6 ± 0.7 b 20.6 ± 1.1 c 20.1 ± 0.6 c 21.6 ± 2.0 26 27 Intraorbital dist.§ 7.7 ± 0.5 a 7.1 ± 0.6 a,b 6.5 ± 0.6 b 6.6 ± 0.7 b 5.8 ± 0.5 c 7.1 ± 0.6 28 Lower leg length§ 39.9 ± 1.9 a 39.9 ± 1.3 a 38.7 ± 3.0 a 35.4 ± 1.5 b 35.7 ± 1.3 b 37.5 ± 0.9 29 30 Hind foot length§ 22.7 ± 1.1 a 23.3 ± 1.5 a 23.4 ± 2.7 a 20.2 ± 1.3 b 20.3 ± 1.0 b 20.7 ± 0.5 31 Third toe length§ 9.8 ± 1.2 a 9.5 ± 0.9 a NA ± NA NA 7.9 ± 0.5 b 7.1 ± 0.6 c 8.0 ± 0.2 32 33 Tail length 132.7 ± 9.0 a 142.6 ± 5.5 b 130.6 ± 9.6 a 121.5 ± 8.3 c 115.2 ± 5.1 d 108.7 ± 19.7 34 § 35 Body length 93.3 ± 6.3 a 89.9 ± 7.0 b 89.2 ± 6.1 a,c 70.9 ± 4.5 c 71.3 ± 4.5 d 80.6 ± 6.4 36 Body mass (g) 57.4 ± 5.6 a 61.1 ± 7.8 a 51.8 ± 9.1 a,b 45.1 ± 6.2 b 43.8 ± 4.7 b 43.3 ± 5.5 37 38 39 40 41 42 43 John Wiley & Sons 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60 Page 51 of 66 American Journal of Primatology

Schüßler et al. page 50 1 2 3 1101 Figure legends: 4 5 6 1102 7 8 9 1103 Figure 1: Map depicting the study region with confirmed species occurrences 10 11 1104 (Hotaling et al., 2016; Kappeler et al., 2005; Louis et al., 2006; Radespiel et al., 12 13 14 1105 2008, 2012; Weisrock et al., 2010). New sampling locations for this study are 15 16 1106 indicated with “*” and forest cover in 2017/2018 was derived from Vieilledent et 17 18 1107 al. (2018) and Schüßler et al. (2020). NP = National Park; SNR = Special 19 20 21 1108 Nature Reserve; SRFor = Special Peer Reserve. Review 22 23 24 1109 25 26 27 1110 Figure 2: Maximum likelihood tree illustrating the phylogenetic relationships 28 29 1111 between Microcebus spp. in northeastern Madagascar as inferred by RAxML 30 31 1112 (based on nuclear sequence data). Sampling locations are indicated at the tips 32 33 of the branches. Illustration adapted from Poelstra et al. (2020). NP = National 34 1113 35 36 1114 Park; SNR = Special Nature Reserve; SR = Special Reserve. 37 38 39 1115 40 41 42 1116 Figure 3: Selected morphometric measurements of Microcebus spp. in 43 44 1117 northeastern Madagascar. Comparison based on one-way ANOVA (P < 0.001 45 46 1118 for all parameters) and grouping (letters above values) according to Tukey post- 47 48 49 1119 hoc tests. Plots for all parameters in supplementary Figure S1. Abbreviations: 50 51 1120 sp.3 = M. sp. #3 (at Ambavala and Antanambe); mac = M. macarthurii (at 52 53 1121 Anjiahely); sim = M. simmonsi (at Ambodiriana); mit = M. mittermeieri (at 54 55 56 1122 Anjiahely); leh = M. lehilahytsara (at Mantadia NP). 57 58 59 1123 60

John Wiley & Sons American Journal of Primatology Page 52 of 66

Schüßler et al. page 51 1 2 3 1124 Figure 4: Principal component analysis including all morphometric parameters 4 5 6 1125 (except third toe length) showing PC1/PC2 (left) and PC3/PC4 (right). Small (M. 7 8 1126 mittermeieri and M. lehilahytsara) and large (M. sp. #3 and M. macarthurii) 9 10 1127 lineages differ along PC1, with some differentiation of M. mittermeieri and M. 11 12 1128 lehilahytsara along both PC1 and PC2, whereas M. sp. #3 and M. macarthurii 13 14 15 1129 split along PC3. Clusters corresponding to the five lineages were significantly 16 17 1130 different from each other (PERMANOVA: F = 36.88, df = 77, P < 0.001). 18 19 1131 Abbreviations: dia: Mantadia NP, vala: Ambavala 20 21 For Peer Review 22 1132 23 24 25 1133 Figure 5: Linear discriminant analysis including all morphometric parameters 26 27 28 1134 (except third toe length). All five linages can be distinguished statistically (Wilk’s 29 30 1135 Lambda = 0.005, F = 10.338, P < 0.001) with a misclassification rate of 12.8%. 31 32 1136 Abbreviations: dia: Mantadia NP, vala: Ambavala 33 34 35 1137 36 37 38 1138 Figure 6: Reproductive records for adult males (A) and females (B) of 39 40 1139 Microcebus spp. in northeastern Madagascar and presence of juvenile 41 42 43 1140 individuals (C) in the population. 44 45 46 1141 47 48 49 1142 Figure 7: Outer morphology of Microcebus jonahi. (A) Drawing of an adult 50 51 1143 individual; (B) Habitus of adult female (paratype individual BD1); (C-E) Close- 52 53 1144 ups of adult male (holotype B34). Illustration copyright by Stephen D. Nash / 54 55 56 1145 IUCN SSC Primate Specialist Group; used with permission. Photos by D. 57 58 1146 Schüßler. 59 60

John Wiley & Sons Page 53 of 66 American Journal of Primatology

1 2 3 Research highlights 4 5 6  Two pairs of Microcebus species occur in partial sympatry 7 8 9  Morphological distinctiveness supports genomic species delimitation in cryptic 10 11 lemurs 12 13  High plasticity in reproductive schedules in a lineage of habitat generalists 14 15 16 detected 17 18 19 20 21 For Peer Review 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

John Wiley & Sons American Journal of PrimatologyPage 54 of 66

1 2 3 4 For Peer Review 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 John Wiley & Sons 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60 Page 55 of 66 American Journal of Primatology

1 2 3 4 5 6 7 8 For Peer Review 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 John Wiley & Sons 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60 American Journal of Primatology Page 56 of 66

1 2 3 4 5 6 7 8 9 For Peer Review 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 John Wiley & Sons 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60 Page 57 of 66 American Journal of Primatology

1 2 3 4 5 For Peer Review 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 John Wiley & Sons 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60 For PeerAmerican Journal Review of PrimatologyPage 58 of 66

1 2 3 4 5 6 7 8 9 10 11 12 13 14 John Wiley & Sons 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60 Page 59 of 66 American Journal of Primatology

1 2 3 4 For Peer Review 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 John Wiley & Sons 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60 American Journal of Primatology Page 60 of 66

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 For Peer Review 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 John Wiley & Sons 51 52 53 54 55 56 57 58 59 60 Page 61 of 66 American Journal of Primatology

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 For Peer Review 20 21 22 23 24 25 26 27 28 29 30 31 32 The newly described Jonah’s mouse lemur (Microcebus jonahi) is distributed in northeastern Madagascar 33 with an estimated Extent of Occurrence of about 1,500 km². This species has a mean head-body length of 34 13.0 cm, a mean tail length of 13.3 cm and weighs on average 57.4 g. 35 36 37 38 50x67mm (150 x 150 DPI) 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60 John Wiley & Sons American Journal of Primatology Page 62 of 66

A new species of mouse lemur - page 1 1 2 3 4 5 6 Supplementary material for: 7 8 Ecology and morphology of mouse lemurs (Microcebus spp.) 9 10 11 in a hotspot of microendemism in northeastern Madagascar, 12 13 14 with the description of a new species 15 16 17 18 19 20 21 For Peer Review 22 23 24 25 26 27 Dominik Schüßler, Marina B. Blanco, 28 29 Jordi Salmona, Jelmer Poelstra, Jean B. Andriambeloson, Alex Miller, Blanchard 30 31 Randrianambinina, David W. Rasolofoson, Jasmin Mantilla-Contreras, Lounès 32 33 Chikhi, Edward E. Louis Jr., Anne D. Yoder 34 35 & Ute Radespiel 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 Corresponding author: 57 58 59 Ute Radespiel, Institute of Zoology, University of Veterinary Medicine Hannover, 60 Buenteweg 17, 30559 Hannover, Germany; email: [email protected]

John Wiley & Sons Page 63 of 66 American Journal of Primatology

A new species of mouse lemur - page 2 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 For Peer Review 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 Supplementary Figure S1: Morphometric measurements of Microcebus spp. in 54 northeastern Madagascar. Comparisons are based on one-way ANOVA (P < 0.001 55 56 for all parameters) and grouping (letters after values) according to Tukey post-hoc 57 58 tests. Abbreviations: sp.3 = M. sp. #3 (at Ambavala and Antanambe); mac = M. 59 macarthurii (at Anjiahely); sim = M. simmonsi (at Ambodiriana); mit = M. mittermeieri 60 (at Anjiahely); leh = M. lehilahytsara (at Mantadia NP).

John Wiley & Sons American Journal of Primatology Page 64 of 66

A new species of mouse lemur - page 3 1 2 3 4 5 6 Supplementary Table S1: Morphometric and reproductive dataset provided as 7 supplementary xlsx-file. 8 9 10 11 12 13 Supplementary Table S2: Coordinates and altitude of sampling locations of 14 Microcebus spp. used in this study. 15 16 Location Longitude Latitude Altitude Reference 17 [m a.s.l.] 18 Marojejy NP +49º 50’ 21’’ -14º 28’ 02’’ 344 Weisrock et al., 2010 19 20 Anjanaharibe- +49º 27’ 53’’ -14º 47’ 27’’ 1056 Louis et al., 2006 21 Sud SR For Peer Review 22 Anjiahely +49° 29’ 38’’ -15° 24’ 45’’ 350-400 Radespiel et al., 23 24 2008, this study 25 Ambavala +49° 34’ 42’’ -16° 11’ 03’’ 233-462 This study 26 Mananara-Nord +49° 47’ 41’’ -16° 18’ 18’’ 277-377 This study 27 28 NP 29 Antanambe +49° 48’ 10’’ -16° 27’ 32’’ 42-273 This study 30 Riamalandy +48º 48’ 54’’ -16º 17’ 06’’ 833 Weisrock et al., 2010 31 32 Ambodiriana +49° 42’ 05’’ -16° 40’ 27’’ 42-222 This study 33 Tampolo +49º 25’ 00’’ -17º 16’ 60’’ 19 Louis et al., 2006 34 Zahamena NP +48º 44’ 30’’ -17º 29’ 13’’ 956 Louis et al., 2006 35 36 Betampona +49º 12’ 12’’ -17º 55’ 52’’ 300 Louis et al., 2006 37 SNR 38 Mantadia NP +48º 25’ 29’’ -18º 48’ 29’’ 969 Kappeler et al., 2005 39 40 Ankofabe +47° 11’ 15’’ -18° 05’ 27’’ 1459 Yoder et al., 2016 41 Ambohitantely +47° 16’ 26’’ -18° 28’ 33’’ 1271 Weisrock et al., 2010 42 Tsinjoarivo +47° 41’ 43’’ -19° 40’ 22’’ 1552 Yoder et al., 2016 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

John Wiley & Sons Page 65 of 66 American Journal of Primatology

A new species of mouse lemur - page 4 1 2 3 4 Supplementary Table S3: Test statistics from parameter-wise comparisons of 5 morphometric measurements using Analyses of Variance. For third toe length, 6 comparison excluding M. simmonsi individuals. 7 8 Morphometric F P 9 10 parameter 11 Ear length 13.80 < 0.001 12 Ear width 8.14 < 0.001 13 14 Head length 35.09 < 0.001 15 Head width 20.73 < 0.001 16 Snout length 91.84 < 0.001 17 Interorbital distance 48.20 < 0.001 18 19 Intraorbital distance 21.36 < 0.001 20 Lower leg length 26.70 < 0.001 21 Hind footFor length Peer 16.03Review < 0.001 22 23 Third toe length 42.65 < 0.001 24 Body length 53.08 < 0.001 25 Tail length 30.29 < 0.001 26 27 Body mass 23.89 < 0.001 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

John Wiley & Sons American Journal of Primatology Page 66 of 66

A new species of mouse lemur - page 5 1 2 3 4 Supplementary Table S4: Classification matrix of the Linear Discriminant Analysis (LDA). Misclassification rate is 12.8%. Correct 5 classifications on diagonal. 6 7 M. sp. #3 M. lehilahytsara M. lehilahytsara M. macarthurii M. mittermeieri M. simmonsi 8 (at Ambavala and (at Mantadia NP) (at Ambavala) (at Anjiahely) (at Anjiahely) (at Ambodiriana) 9 Antanambe) 10 M. lehilahytsara 11 29 0 0 0 0 0 12 (at Mantadia NP) 13 M. lehilahytsara 0 1 0 0 2 0 14 (at Ambavala) For Peer Review 15 M. macarthurii 16 0 0 7 0 0 2 (at Anjiahely) 17 M. mittermeieri 18 1 0 0 19 0 0 19 (at Anjiahely) 20 M. simmonsi 21 1 0 0 1 3 1 (at Ambodiriana) 22 23 M. sp. #3 24 (at Ambavala and 0 0 1 0 1 9 25 Antanambe) 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 John Wiley & Sons 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60 Page 67 of 66 American Journal of Primatology

1

2 Supplementary Table S1 for: Ecology and morphology of mouse lemurs (Microcebus spp.) 3 in a hotspot of microendemism in northeastern Madagascar, with the description of a new species 4

5 Dominik Schüßler, Marina B. Blanco, Jordi Salmona, Jelmer Poelstra, Jean B. Andriambeloson, Alex Miller, Blanchard Randrianambinina, David W. Rasolofoson, Jasmin Mantilla-Contreras, Lounès Chikhi, Edward E. Louis Jr., Anne D. Yoder & 6 Ute Radespiel 7 Corresponding author: Ute Radespiel, Institute of Zoology, University of Veterinary Medicine Hannover, Buenteweg 17, 30559 Hannover, Germany 8 email: [email protected] 9 Remarks: all values in mm except weight, this parameter is in g, NA: not available all measurements are according to Hafen et al. 1998 (Folia Primatologica 69:342-356) and Zimmermann et al. 1998 (Folia Primatologia 69:106- 10 114) 11 Reproductive state is provided for first-time capture

12 Individual.ID Location Age Sex Species weight ear.length ear.width head.length head.width interorbital.dist intraorbital.dist snout.length lower.leg.length hind.foot.length third.toe.length body.length tail.length testes_state female_reproduction Date Collector B-MF-BC2 Ambavala young F lehilahytsara 30 15,5 10,8 32,3 19,1 19,4 4,9 8,6 33,1 20,8 8,3 79,1 111,0 NA closed 8-Sep-2017 DS 13 B-MM-B12 Ambavala juvenile M lehilahytsara 19 14,4 9,3 30,4 15,5 19,8 5,9 8,1 29,6 18,4 7,9 63,2 81,0 regressed NA 9-Sep-2017 DS B-MM-B23 Ambavala adult M lehilahytsara 37 18,0 11,6 33,8 18,9 19,6 7,2 10,4 36,5 20,4 8,2 75,3 86,0 regressed NA 9-Sep-2017 DS B-MM-B14 Ambavala young M lehilahytsara 30 16,4 11,7 33,2 19,3 20,2 8,0 8,8 34,1 22,5 9,4 79,1 119,0 regressed NA 12-Sep-2017 DS 14 B-MM-C12 Ambavala adult M lehilahytsara 46 18,1 10,8 36,4 21,2 23,5 6,4 9,5 38,0 20,5 7,9 78,7 122,0 regressed NA 15-Sep-2017 DS B-MF-BC3 Ambavala adult F lehilahytsara 47 17,7 13,0 35,4 19,9 21,7 7,6 9,0 38,0 21,3 8,0 87,7 118,0 NA closed 19-Sep-2017 DS B-MM-C24 Ambavala juvenile M lehilahytsara 20 13,0 10,7 29,3 17,8 17,1 6,5 5,9 25,6 18,0 8,7 57,6 76,0 regressed NA 19-Sep-2017 DS 15 B-MM-C23 Ambavala juvenile M lehilahytsara 23 15,3 10,7 30,7 17,4 18,1 6,5 6,6 24,4 17,8 7,5 68,3 97,0 regressed NA 19-Sep-2017 DS F06-99 Mantadia adult F lehilahytsara 46 19,2 12,5 33,4 19,2 20,1 6,6 4,2 35,1 20,7 7,6 75,0 117,0 NA closed 5-May-1999 BR 16 F12-99 Mantadia adult F lehilahytsara 41 20,2 13,6 34,7 18,8 20,4 6,8 5,3 38,2 21,9 7,6 80,0 125,0 NA closed 9-May-1999 BR F14-99 Mantadia adult F lehilahytsara 45 20,4 11,2 33,8 19,2 20,0 5,5 4,1 34,8 19,0 7,3 70,0 110,0 NA closed 18-May-1999 BR F32-99 Mantadia adult F lehilahytsara 45 20,0 11,9 33,8 19,2 20,1 5,0 4,3 35,1 21,2 7,3 75,0 115,0 NA closed 4-Sep-1999 BR 17 F42-99 Mantadia adult F lehilahytsara 39 18,1 11,4 34,9 17,5 20,4 5,6 5,1 35,8 21,1 7,2 70,0 120,0 NA closed 24-Oct-1999 BR F45-99 Mantadia adult F lehilahytsara 46 19,8 11,6 34,1 19,1 20,3 5,8 4,3 37,2 22,6 7,4 70,0 115,0 NA open 9-Nov-1999 BR 18 F47-99 Mantadia adult F lehilahytsara 50 18,5 12,0 33,2 17,5 20,0 5,8 5,1 35,9 20,3 7,0 70,0 115,0 NA pregnant 7-Dec-1999 BR F48-99 Mantadia adult F lehilahytsara 49 21,7 12,7 33,8 18,9 19,9 4,7 4,8 37,2 20,2 6,8 70,0 115,0 NA pregnant 7-Dec-1999 BR F49-99 Mantadia adult F lehilahytsara 55 21,0 13,0 32,3 18,2 18,7 5,5 4,9 37,5 18,8 6,4 70,0 115,0 NA pregnant 7-Dec-1999 BR 19 M07-99 Mantadia adult M lehilahytsara 43 22,1 13,0 32,8 19,0 20,1 6,4 5,4 37,0 21,7 7,8 75,0 125,0 regressed NA 7-May-1999 BR M03-99 Mantadia adult M lehilahytsara 52 18,5 11,6 32,0 18,6 20,4 6,7 4,2 34,9 20,1 5,9 75,0 110,0 regressed NA 5-May-1999 BR 20 M10-99 Mantadia adult M lehilahytsara 37 21,2 12,3 33,1 19,1 18,9 5,4 4,1 35,8 19,9 8,0 67,0 120,0 regressed NA 9-May-1999 BR M02-99 Mantadia adult M lehilahytsara 46 18,8 11,3 32,5 17,9 21,9 5,2 4,3 32,5 19,4 6,7 70,0 120,0 regressed NA 11-Apr-1999 BR M04-99 Mantadia adult M lehilahytsara 38 18,8 11,0 33,3 19,6 19,6 5,0 5,4 37,3 21,1 6,5 65,0 115,0 regressed NA 5-May-1999 BR 21 M08-99 Mantadia adult M lehilahytsara 37 20,8 12,5For 35,0 18,5Peer19,6 6,0Review5,9 36,0 20,0 6,9 68,0 115,0 regressed NA 7-May-1999 BR M16-99 Mantadia adult M lehilahytsara 39 19,6 11,4 33,4 19,8 20,2 5,7 5,3 36,3 18,4 7,6 70,0 115,0 regressed NA 30-May-1999 BR M18-99 Mantadia adult M lehilahytsara 37 18,7 11,9 32,4 18,8 18,9 6,2 4,8 34,5 19,2 6,8 65,0 107,0 regressed NA 30-May-1999 BR 22 M31-99 Mantadia adult M lehilahytsara 48 20,6 14,7 33,0 19,4 21,3 5,0 5,6 36,9 20,8 7,0 80,0 115,0 enlarged NA 7-Aug-1999 BR M33-99 Mantadia adult M lehilahytsara 42 20,6 12,3 31,5 19,1 20,2 6,2 4,5 35,4 20,8 7,9 75,0 115,0 enlarged NA 7-Sep-1999 BR 23 M34-99 Mantadia adult M lehilahytsara 43 19,5 12,3 33,5 19,8 20,0 5,8 4,2 35,9 19,9 7,8 70,0 115,0 enlarged NA 7-Sep-1999 BR M35-99 Mantadia adult M lehilahytsara 47 20,2 12,8 34,1 18,8 20,5 6,1 4,8 35,3 19,5 7,8 70,0 110,0 enlarged NA 7-Sep-1999 BR M36-99 Mantadia adult M lehilahytsara 46 20,0 12,0 32,9 18,0 20,2 6,4 4,9 34,5 19,0 7,8 70,0 110,0 enlarged NA 7-Sep-1999 BR 24 M37-99 Mantadia adult M lehilahytsara 42 20,0 11,3 30,1 18,6 20,0 5,7 5,2 36,4 20,5 7,0 70,0 115,0 enlarged NA 7-Sep-1999 BR M38-99 Mantadia adult M lehilahytsara 46 20,8 13,8 33,3 18,2 20,4 5,7 4,2 36,4 20,7 6,8 70,0 120,0 enlarged NA 7-Sep-1999 BR 25 M39-99 Mantadia adult M lehilahytsara 39 19,4 12,0 33,7 18,0 20,4 5,9 5,3 34,1 21,0 6,6 75,0 120,0 enlarged NA 24-Sep-1999 BR M40-99 Mantadia adult M lehilahytsara 43 18,9 12,8 33,7 18,6 21,0 4,9 6,2 35,6 19,7 6,2 70,0 110,0 enlarged NA 24-Sep-1999 BR M41-99 Mantadia adult M lehilahytsara 45 21,7 13,5 33,8 15,6 20,0 5,7 5,3 36,6 21,3 6,5 75,0 116,0 enlarged NA 26-Sep-1999 BR 26 M43-99 Mantadia adult M lehilahytsara 35 20,6 12,4 31,6 16,7 19,6 6,3 5,0 33,5 20,3 7,3 60,0 115,0 enlarged NA 7-Nov-1999 BR M44-99 Mantadia adult M lehilahytsara 40 18,5 11,3 32,9 18,2 19,9 6,0 4,6 34,8 18,8 5,7 70,0 100,0 enlarged NA 9-Nov-1999 BR 27 M20-99 Mantadia adult M lehilahytsara NA 21,9 13,5 33,7 21,0 20,0 5,7 4,6 35,9 19,6 7,2 80,0 120,0 enlarged NA 31-May-1999 BR F01-19 Mantadia juvenile F lehilahytsara 20 NA NA NA NA NA NA NA NA NA NA NA NA NA closed 25-Mar-1999 BR F11-99 Mantadia young F lehilahytsara 33 NA NA NA NA NA NA NA NA NA NA NA NA NA closed 9-May-1999 BR 28 M17-99 Mantadia young M lehilahytsara 30 NA NA NA NA NA NA NA NA NA NA NA NA regressed NA 30-May-1999 BR M05-99 Mantadia young M lehilahytsara 30 NA NA NA NA NA NA NA NA NA NA NA NA regressed NA 5-May-1999 BR 29 M09-99 Mantadia young M lehilahytsara 36 NA NA NA NA NA NA NA NA NA NA NA NA regressed NA 7-May-1999 BR M13-19 Mantadia young M lehilahytsara 30 NA NA NA NA NA NA NA NA NA NA NA NA regressed NA 18-May-1999 BR M15-99 Mantadia young M lehilahytsara 32 NA NA NA NA NA NA NA NA NA NA NA NA regressed NA 18-May-1999 BR 30 M19-99 Mantadia young M lehilahytsara 32 NA NA NA NA NA NA NA NA NA NA NA NA regressed NA 31-May-1999 BR M27-99 Mantadia young M lehilahytsara 33 NA NA NA NA NA NA NA NA NA NA NA NA regressed NA 9-Aug-1999 BR M28-99 Mantadia young M lehilahytsara 34 NA NA NA NA NA NA NA NA NA NA NA NA regressed NA 5-Aug-1999 BR 31 M29-99 Mantadia young M lehilahytsara 31 NA NA NA NA NA NA NA NA NA NA NA NA regressed NA 7-Aug-1999 BR M30-99 Mantadia young M lehilahytsara 33 NA NA NA NA NA NA NA NA NA NA NA NA regressed NA 7-Aug-1999 BR 32 W04-06 Anjiahely adult F macarthurii 50 18,7 10,2 36,5 19,7 22,0 6,9 8,4 39,7 21,4 8,1 75,0 151,0 NA swollen 9-Nov-2006 DWR M03-06 Anjiahely adult M macarthurii 68 18,0 12,0 37,5 19,8 23,3 6,8 8,0 40,2 21,6 8,2 85,0 142,0 enlarged NA 9-Nov-2006 DWR W01-07 Anjiahely adult F macarthurii 53 19,5 11,1 35,1 19,7 21,6 6,4 8,3 38,7 23,5 8,2 85,0 NA NA closed 4-Sep-2007 DWR 33 M01-06 Anjiahely adult M macarthurii 58 18,8 10,2 36,6 18,4 21,1 6,2 8,1 38,2 21,5 8,8 82,0 142,0 enlarged NA 8-Nov-2006 DWR M05-08 Anjiahely adult M macarthurii 72 17,5 8,5 39,7 22,4 24,4 8,0 11,3 40,8 25,5 10,0 95,0 143,0 NA NA 12-Dec-2008 DWR 34 M0608 Anjiahely adult M macarthurii 68 17,2 9,6 37,2 20,1 24,7 6,7 10,0 41,6 23,2 9,8 94,0 144,0 enlarged NA 13-Dec-2008 DWR M07-08 Anjiahely adult M macarthurii 62 17,6 10,3 39,0 21,3 25,6 7,8 10,8 40,6 25,4 9,9 94,0 138,0 enlarged NA 13-Dec-2008 DWR M08-08 Anjiahely adult M macarthurii 55 17,2 9,6 37,1 22,3 25,0 7,6 9,3 39,9 22,5 10,0 96,0 145,0 enlarged NA 13-Dec-2008 DWR 35 F04-11 Anjiahely adult F macarthurii 72 19,4 10,3 38,7 22,0 23,8 8,1 12,0 42,7 24,5 10,9 99,0 146,0 NA closed 22-Nov-2011 DWR M03-11 Anjiahely adult M macarthurii 61 17,9 9,1 39,2 20,2 22,7 6,7 9,8 39,9 23,8 9,8 91,0 134,0 enlarged NA 22-Nov-2011 DWR 36 M01-13 Anjiahely adult M macarthurii 70 17,1 11,8 37,6 17,8 26,6 7,2 NA 38,4 NA 10,3 96,6 141,8 enlarged NA 25-Oct-2013 DWR F02-13 Anjiahely adult F macarthurii 61 17,9 11,3 37,4 19,6 23,7 7,5 NA 39,6 NA 9,3 94,9 141,5 NA closed 25-Oct-2013 DWR F03-13 Anjiahely adult F macarthurii 51 17,7 11,5 35,6 18,4 23,1 6,7 NA 37,8 NA NA 82,8 133,1 NA closed 25-Oct-2008 DWR 37 F04-13 Anjiahely adult F macarthurii 55 17,5 13,3 37,4 18,9 23,2 7,1 NA 39,8 NA 10,1 88,3 151,9 NA closed 26-Oct-2008 DWR W06-06 Anjiahely adult F mittermeieri 51 19,1 12,2 34,0 19,2 20,7 5,4 7,0 35,2 20,4 6,9 75,0 125,0 NA open 9-Nov-2006 DWR M01-06leng Anjiahely adult M mittermeieri 50 16,5 12,1 34,3 NA 22,4 NA 9,3 38,7 21,9 8,7 68,0 116,0 enlarged NA 13-Dec-2006 DWR 38 W02-06 Anjiahely adult F mittermeieri 49 19,7 13,4 35,4 18,7 21,4 6,7 8,8 36,2 22,9 7,7 80,0 119,0 NA closed 8-Nov-2006 DWR M07-06 Anjiahely adult M mittermeieri 40 17,7 12,7 32,5 18,8 22,6 6,4 6,3 33,8 20,0 7,6 72,0 107,0 enlarged NA 12-Nov-2006 DWR 39 M05-06 Anjiahely adult M mittermeieri 40 19,2 12,9 33,0 17,9 20,3 6,2 7,7 35,5 19,8 7,5 73,0 121,0 enlarged NA 9-Nov-2006 DWR M02-07 Anjiahely adult M mittermeieri 49 20,0 11,8 34,7 18,8 20,6 6,6 8,0 36,6 19,4 7,5 70,0 119,0 enlarged NA 4-Sep-2007 DWR M03-07 Anjiahely adult M mittermeieri 41 18,7 10,8 33,3 19,3 20,9 6,0 6,4 34,1 19,3 7,6 68,0 115,0 enlarged NA 5-Sep-2007 DWR 40 W04-07 Anjiahely adult F mittermeieri 49 18,9 11,0 34,8 20,0 20,2 6,6 7,6 36,4 20,8 8,4 70,0 135,0 NA closed 5-Sep-2007 DWR M05-07 Anjiahely adult M mittermeieri 38 18,2 11,8 34,4 20,0 19,8 6,5 7,2 34,9 20,0 8,0 65,0 134,0 enlarged NA 5-Sep-2007 DWR 41 M06-07 Anjiahely adult M mittermeieri 43 19,9 12,4 34,2 18,1 19,0 6,4 8,9 35,6 20,7 7,9 72,0 115,0 enlarged NA 5-Sep-2007 DWR M07-07 Anjiahely adult M mittermeieri 48 17,8 12,8 34,5 19,7 20,4 7,9 6,9 35,3 20,6 7,1 70,0 125,0 enlarged NA 5-Sep-2007 DWR W08-07 Anjiahely adult F mittermeieri 39 18,7 11,9 34,5 19,9 20,9 5,6 7,8 34,2 21,0 8,0 70,0 120,0 NA closed 5-Sep-2007 DWR 42 M09-07 Anjiahely adult M mittermeieri 40 17,5 11,6 35,3 19,0 20,9 6,1 7,7 35,8 19,2 7,5 67,0 128,0 enlarged NA 6-Sep-2007 DWR W10-07 Anjiahely adult F mittermeieri 38 18,6 11,0 33,8 20,0 20,5 7,0 8,7 34,6 21,0 8,0 65,0 127,0 NA closed 6-Sep-2007 DWR 43 M11-07 Anjiahely adult M mittermeieri 52 20,0 12,0 33,4 19,0 20,8 6,2 8,0 34,4 19,6 7,8 75,0 122,0 enlarged NA 6-Sep-2007 DWR M12-07 Anjiahely adult M mittermeieri 48 18,8 11,8 34,6 19,8 20,4 7,0 8,3 35,2 20,0 7,9 75,0 125,0 enlarged NA 6-Sep-2007 DWR M13-07 Anjiahely adult M mittermeieri 58 19,8 13,8 35,3 21,0 21,6 6,5 8,3 37,9 20,1 7,7 80,0 135,0 enlarged NA 6-Sep-2007 DWR 44 M14-07 Anjiahely adult M mittermeieri 46 16,5 11,0 35,7 19,0 20,2 7,1 9,1 37,1 20,0 8,0 65,0 125,0 enlarged NA 6-Sep-2007 DWR M15-07 Anjiahely adult M mittermeieri 42 15,0 12,1 34,7 19,0 18,7 7,1 8,2 33,7 18,1 8,6 70,0 100,0 enlarged NA 6-Sep-2007 DWR 45 W16-07 Anjiahely adult F mittermeieri 34 16,0 12,1 33,4 19,2 18,6 6,8 7,0 32,7 18,0 8,2 65,0 120,0 NA closed 6-Sep-2007 DWR W17-07 Anjiahely adult F mittermeieri 42 17,1 11,3 33,7 20,0 19,6 5,8 6,6 33,4 18,2 8,6 73,0 118,0 NA closed 6-Sep-2007 DWR W08-06 Anjiahely adult F mittermeieri 55 18,1 13,4 34,4 19,7 22,2 8,4 7,9 36,7 22,8 8,7 NA 123,0 NA open 17-Nov-2006 DWR 46 M01-08 Anjiahely juvenile M mittermeieri 20 11,3 9,8 27,3 18,0 15,4 4,0 9,0 25,9 17,2 6,3 51,0 85,0 regressed NA 11-Dec-2008 DWR M02-08 Anjiahely juvenile M mittermeieri 20 15,7 9,2 31,1 17,5 18,0 6,4 7,9 28,5 20,0 7,7 56,0 100,7 regressed NA 11-Dec-2008 DWR W03-08 Anjiahely juvenile F mittermeieri 22 14,9 8,7 32,6 18,0 18,7 7,6 7,9 31,1 20,8 8,2 74,8 93,5 NA closed 11-Dec-2008 DWR 47 W04-08 Anjiahely juvenile F mittermeieri 24 15,3 7,2 31,8 18,5 18,7 6,4 8,0 31,3 20,8 8,3 58,8 99,7 NA closed 11-Dec-2008 DWR A1 Ambodiriana adult F simmonsi 50 NA 12,9 35,5 20,4 22,2 7,5 11,9 NA 23,3 NA NA 148,0 NA closed 18-Jun-2014 AM 48 A2 Ambodiriana adult F simmonsi 46 NA 12,4 33,7 20,7 22,1 6,0 10,5 NA 29,8 NA NA 121,0 NA closed 18-Jun-2014 AM A3 Ambodiriana adult F simmonsi 43 15,5 11,5 35,1 20,2 22,2 6,8 11,0 37,8 22,7 NA 80,0 130,0 NA closed 21-Jun-2014 AM A4 Ambodiriana adult M simmonsi 70 20,7 12,8 36,8 20,5 23,0 6,4 11,5 41,3 22,3 NA 95,0 138,0 enlarged NA 2-Jul-2014 AM 49 A5 Ambodiriana adult M simmonsi 43 16,7 11,5 34,6 20,1 21,9 5,7 7,3 33,2 20,6 NA 85,0 120,0 enlarged NA 2-Jul-2014 AM A6 Ambodiriana adult M simmonsi 54 21,0 12,7 35,1 20,7 22,4 6,2 7,4 39,0 22,7 NA 93,0 123,0 enlarged NA 17-Jul-2014 AM 50 A7 Ambodiriana adult M simmonsi 58 22,0 14,4 36,0 20,2 23,7 7,1 10,2 40,1 23,1 NA 95,0 130,0 enlarged NA 17-Jul-2014 AM A8 Ambodiriana adult M simmonsi 50 20,9 12,4 35,8 20,2 23,4 6,4 10,5 40,8 22,3 NA 87,0 135,0 enlarged NA 17-Jul-2014 AM B-MF-BD1 Ambavala adult F sp3 64 17,0 12,4 39,7 20,0 26,7 7,5 10,4 41,1 24,0 10,4 103,0 151,0 NA closed 4-Sep-2017 DS 51 B-MM-B34 Ambavala adult M sp3 66 17,6 13,7 37,7 23,0 26,0 8,2 10,0 41,7 24,5 10,6 95,6 130,0 enlarged NA 6-Sep-2017 DS B-MM-B13 Ambavala adult M sp3 63 18,0 11,8 36,8 20,0 24,9 7,0 10,3 39,3 21,5 8,4 94,0 138,0 enlarged NA 11-Sep-2017 DS 52 B-MM-B24 Ambavala adult M sp3 62 15,3 11,5 35,8 21,3 23,7 7,6 9,0 37,4 20,8 9,9 92,6 135,0 enlarged NA 13-Sep-2017 DS B-MF-BC1 Ambavala adult F sp3 64 18,0 12,5 36,3 23,1 23,8 8,2 10,6 39,2 23,0 8,9 96,3 132,0 NA closed 17-Sep-2017 DS A-MM-A12 Antanambe adult M sp3 50 18,7 13,5 36,7 21,4 24,9 7,3 10,1 38,1 22,5 11,6 95,9 128,0 enlarged NA 13-Aug-2017 DS 53 A-MM-A23 Antanambe adult M sp3 54 18,0 13,5 35,9 21,6 23,2 7,0 11,4 38,4 22,9 10,4 94,0 137,0 enlarged NA 15-Aug-2017 DS A-MM-A34 Antanambe adult M sp3 63 17,6 14,7 34,5 22,0 24,7 8,1 11,0 44,0 23,0 8,7 100,1 133,0 enlarged NA 17-Aug-2017 DS A-MM-A24 Antanambe adult M sp3 53 17,0 12,1 32,8 21,3 25,4 8,1 9,1 40,1 22,6 10,9 82,8 146,0 enlarged NA 22-Aug-2017 DS 54 A-MM-A13 Antanambe adult M sp3 52 18,5 12,1 37,4 20,7 24,0 8,1 9,9 39,8 23,0 7,9 83,3 146,0 enlarged NA 24-Aug-2017 DS A-MF-AB1 Antanambe adult F sp3 56 17,0 12,4 36,3 20,1 24,2 7,5 9,1 39,4 21,5 10,4 88,7 135,0 NA closed 26-Aug-2017 DS 55 MBB 019 Mananara_Nord adult M sp3 55 17,4 NA 36,7 21,6 NA NA 13,8 NA NA NA NA 131,9 enlarged NA 13-Aug-2015 MBB MBB 020 Mananara_Nord adult M sp3 54 16,4 NA 35,7 21,4 NA NA 11,1 NA NA NA NA 131,0 enlarged NA 15-Aug-2015 MBB MBB 021 Mananara_Nord adult M sp3 54 16,5 NA 35,4 21,4 NA NA 10,0 NA NA NA NA 131,0 enlarged NA 15-Aug-2015 MBB 56 MBB 022 Mananara_Nord young F sp3 46 16,3 NA 36,5 22,1 NA NA 11,7 NA NA NA NA 122,0 NA closed 16-Aug-2015 MBB MBB 023 Mananara_Nord young M sp3 46 17,6 NA 35,2 21,6 NA NA 11,2 NA NA NA NA 119,0 enlarged NA 18-Aug-2015 MBB 57 MBB 024 Mananara_Nord young F sp3 47 15,4 NA 36,4 21,8 NA NA 11,6 NA NA NA NA 134,0 NA closed 18-Aug-2015 MBB MBB 025 Mananara_Nord adult M sp3 53 16,9 NA 35,5 22,6 NA NA 11,7 NA NA NA NA 121,0 enlarged NA 21-Aug-2015 MBB MBB 026 Mananara_Nord adult M sp3 57 17,0 NA 36,7 21,3 NA NA 12,4 NA NA NA NA 115,6 enlarged NA 22-Aug-2015 MBB 58 MBB 027 Mananara_Nord young F sp3 46 15,4 NA 35,6 22,0 NA NA 9,4 NA NA NA NA 123,3 NA closed 23-Aug-2015 MBB MBB 028 Mananara_Nord adult M sp3 64 17,1 NA 36,5 22,8 NA NA 11,2 NA NA NA NA 121,2 enlarged NA 23-Aug-2015 MBB 59 MBB 029 Mananara_Nord young M sp3 44 16,3 NA 35,9 20,9 NA NA 11,7 NA NA NA NA 127,0 enlarged NA 23-Aug-2015 MBB MBB 030 Mananara_Nord young F sp3 42 14,2 NA 35,7 20,4 NA NA 10,5 NA NA NA NA 131,0 NA closed 24-Aug-2015 MBB MBB 031 Mananara_Nord young M sp3 50 17,2 NA 36,4 20,5 NA NA 11,9 NA NA NA NA 128,7 enlarged NA 24-Aug-2015 MBB 60 MBB 032 Mananara_Nord adult M sp3 49 16,4 NA 35,5 21,6 NA NA 11,0 NA NA NA NA 126,0 enlarged NA 24-Aug-2015 MBB

John Wiley & Sons