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Abstract Background Pleistocene events have shaped the phylogeography of many taxa worldwide. Their genetic signatures in tropical species have been much less explored than in those living in temperate regions. We analysed the genetic structure of a Malagasy primate species, a mouse lemur with a wide distribution (M. murinus), in order to investigate such phylogeographic processes on a large tropical island. We also evaluated the effects of anthropogenic pressures (fragmentation/deforestation) and natural features (geographic distance, rivers) on genetic structure in order to complement our understanding of past and present processes of genetic differentiation. Results The analysis of the mitochondrial D-loop sequences of 195 samples from 15 study sites (10 from a continuous forest and five from isolated forest fragments) from two adjacent Inter-River-Systems (IRSs) revealed that forest fragmentation and the river restrict gene flow, thereby leading to an increased genetic differentiation between populations beyond the effect of isolation-by-distance. Demographic simulations detected signals of two successive spatial expansions that could be preliminarily dated to the late Pleistocene and early Holocene. The haplotype network revealed geographic structure and showed deep molecular divergences within and between the IRSs that would be congruent with a two-step colonization scenario. Conclusions This study supports the hypothesis of a relatively recent spatial expansion of the grey mouse lemur in northwestern Madagascar, which may also explain why this taxon, in contrast to its congeners, has not yet undergone allopatric speciation in the studied area and possibly across its presently wide range.

Abstract Background The lemurs of Madagascar provide an excellent mammalian radiation to explore mechanisms and processes favouring species diversity and evolution. Species diversity, in particular of nocturnal species, increased considerably during the last decade. However, the factors contributing to this high diversity are not well understood. We tested predictions derived from two existing biogeographic models by exploring the genetic and morphological divergence among populations of a widely distributed lemur genus, the sportive lemur (Lepilemur ssp.) along a 560 km long transect from western to northern Madagascar. Results By using the phylogenetic analyses of mtDNA sequence data, molecular diagnostic sites and phenotypic morphometric traits, we uncovered two previously undetected species whose distributions contradict the two existing biogeographic models. Brief species descriptions are provided and a new biogeographic model is proposed (the ”large river model“). Conclusion According to the ”large river model“, large rivers in north and northwestern Madagascar acted as geographical barriers for gene flow and facilitated speciation events on a much smaller spatial scale than previously thought. Thereby, this study does not only show that species diversity in nocturnal Malagasy primates is continuously underestimated but aims to emphasize the need for conservation actions if those species with small ranges shall not face extinction in the near future.

Abstract Background The number of species within the Malagasy genus Lepilemur and their phylogenetic relationships is disputed and controversial. In order to establish their evolutionary relationships, a comparative cytogenetic and molecular study was performed. We sequenced the complete mitochondrial cytochrome b gene (1140 bp) from 68 individuals representing all eight sportive lemur species and most major populations, and compared the results with those obtained from cytogenetic studies derived from 99 specimens. Results Interspecific genetic variation, diagnostic characters and significantly supported phylogenetic relationships were obtained from the mitochondrial sequence data and are in agreement with cytogenetic information. The results confirm the distinctiveness of Lepilemur ankaranensis, L. dorsalis, L. edwardsi, L. leucopus, L. microdon, L. mustelinus, L. ruficaudatus and L. septentrionalis on species level. Additionally, within L. ruficaudatus large genetic differences were observed among different geographic populations. L. dorsalis from Sahamalaza Peninsula and from the Ambanja/Nosy Be region are paraphyletic, with the latter forming a sister group to L. ankaranensis. Conclusion Our results support the classification of the eight major sportive lemur taxa as independent species. Moreover, our data indicate further cryptic speciation events within L. ruficaudatus and L. dorsalis. Based on molecular data we propose to recognize the sportive lemur populations from north of the Tsiribihina River, south of the Betsiboka River, and from the Sahamalaza Peninsula, as distinct species.

Although it is well known that humans substantially altered the Malagasy ecosystems, the timing of the human arrival as well as the extension of their environmental impact is yet not well understood. This research aims to study the influence of early human impact and climate change on rainforests and wildlife in northern Madagascar during the past millennia. Results obtained from the lake sediment in a montane environment showed significant changes in vegetation within the lake catchment associated with a major drought that started approximately 1100 years ago. Human impact, revealed by fires, began at roughly the same time and occurred outside the lake catchment. Although this does not dismiss the impacts that humans had at a regional scale, this result demonstrates that the late Holocene natural drought also significantly impacted the ecosystems independently of anthropogenic activities. At a regional scale, a review of species demographic history revealed a substantial number of population bottlenecks during the last millennia, probably resulting from this combination of human-related impact and natural climate changes. This research highlights the importance of a multi-site and multi-proxy comparison for deciphering the nature and succession of environmental changes., Competing Interests: We declare we have no competing interests., (© 2024 The Author(s).)

Madagascar's lemur populations are declining in dwindling habitats due to anthropogenic expansion and changing climatic conditions. Gastrointestinal parasites can be important indicators to assess the health status of threatened species. However, parasites, hosts and the environment are connected in complex interactions. The present study aimed to disentangle the impact of seasonal and several host-specific factors (sex, species, age, reproductive status, and body mass) on endoparasitism in two small-bodied, co-occurring lemur species ( Microcebus murinus and Microcebus ravelobensis ) in the Ankarafantsika National Park. Helminth prevalence and egg shedding intensity was investigated via copromicroscopic examination of 810 fecal samples that were obtained from 178 individuals across an 11-month period with a longitudinal approach via repeated captures in a 30.6 ha forest area. Both mouse lemur hosts shed seven morphologically distinct egg types (assigned to Subulura baeri , unidentified Enterobiinae, Spirura sp., Lemuricola sp., two Hymenolepididae spp., one unidentified ascarid). Postmortem examination of two deceased individuals enabled assignment of adult worms to egg morphotypes of S. baeri , Spirura sp. and one Hymenolepididae sp., supported by molecular analysis. A significant seasonal variation was observed in the occurrence of the three most common helminth species S . baeri (total prevalence 71%), unidentified Enterobiinae (46%) and Spirura sp. (38%), with a higher likelihood of infection with advancing dry season. Neither host species, sex nor reproductive status had a significant effect on gastrointestinal helminth infections. Host body mass showed pronounced seasonal changes but did not differ significantly between infected and non-infected individuals. The pathogenic effects of gastrointestinal helminths therefore likely remained within compensable limits in the studied mouse lemur populations. Our findings highlight the prominent influence of seasonal changes on helminth communities. The results of combined morphologic and genetic approaches can furthermore help to overcome limitations of parasite identification via copromicroscopy by linking egg morphology to DNA sequences., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2024 The Authors.)

Habitat loss and fragmentation are of concern to conservation biologists worldwide. However, not all organisms are affected equally by these processes; thus, it is important to study the effects of living in fragmented habitats on species that differ in lifestyle and habitat requirements. In this study, we examined the dispersal and connectivity patterns of rodents, one endemic ( Eliurus myoxinus ) and one invasive ( Rattus rattus ), in two landscapes containing forest fragments and adjacent continuous forest patches in northwestern Madagascar. We generated genetic (RADseq) data for 66 E. myoxinus and 81 R. rattus individuals to evaluate differences in genetic diversity as well as inbreeding and connectivity in two landscapes. We found higher levels of inbreeding and lower levels of genetic diversity in E. myoxinus compared with R. rattus . We observed related dyads both within and between habitat patches and positive spatial autocorrelation at lower distance classes for both species, with a stronger pattern of spatial autocorrelation in R. rattus . Across each site, we identified contrasting migration rates for each species, but these did not correspond to habitat-matrix dichotomies. The relatively low genetic diversity in the endemic E. myoxinus suggests ecological constraints that require further investigation.

Madagascar exhibits exceptionally high levels of biodiversity and endemism. Models to explain the diversification and distribution of species in Madagascar stress the importance of historical variability in climate conditions which may have led to the formation of geographic barriers by changing water and habitat availability. The relative importance of these models for the diversification of the various forest-adapted taxa of Madagascar has yet to be understood. Here, we reconstructed the phylogeographic history of Gerp's mouse lemur ( Microcebus gerpi ) to identify relevant mechanisms and drivers of diversification in Madagascar's humid rainforests. We used restriction site associated DNA (RAD) markers and applied population genomic and coalescent-based techniques to estimate genetic diversity, population structure, gene flow and divergence times among M. gerpi populations and its two sister species M. jollyae and M. marohita . Genomic results were complemented with ecological niche models to better understand the relative barrier function of rivers and altitude. We show that M. gerpi diversified during the late Pleistocene. The inferred ecological niche, patterns of gene flow and genetic differentiation in M. gerpi suggest that the potential for rivers to act as biogeographic barriers depended on both size and elevation of headwaters. Populations on opposite sides of the largest river in the area with headwaters that extend far into the highlands show particularly high genetic differentiation, whereas rivers with lower elevation headwaters have weaker barrier functions, indicated by higher migration rates and admixture. We conclude that M. gerpi likely diversified through repeated cycles of dispersal punctuated by isolation to refugia as a result of paleoclimatic fluctuations during the Pleistocene. We argue that this diversification scenario serves as a model of diversification for other rainforest taxa that are similarly limited by geographic factors. In addition, we highlight conservation implications for this critically endangered species, which faces extreme habitat loss and fragmentation., Competing Interests: The authors declare no competing interests., (© 2023 The Authors. Ecology and Evolution published by John Wiley & Sons Ltd.)

Sucking lice live in intimate association with their hosts and often display a high degree of host specificity. The present study investigated sucking lice of the genus Lemurpediculus from six mouse lemur ( Microcebus ) and two dwarf lemur ( Cheirogaleus ) species endemic to the island of Madagascar, considered a biodiversity hotspot. Louse phylogenetic trees were created based on cytochrome C oxidase subunit I (COI), elongation factor 1α (EF1α) and internal transcribed spacer 1 (ITS1) sequences. While clustering according to host species was generally observed for COI and ITS1, suggesting high host specificity of the examined lice, EF1α sequences alone did not distinguish between lice of different Microcebus species, possibly due to rather recent divergence. As bootstrap support for basal tree structure was rather low, further data are necessary to resolve the evolutionary history of louse-mouse lemur associations. Three new species of sucking lice are described: Lemurpediculus zimmermanni sp. Nov. From Microcebus ravelobensis , Lemurpediculus gerpi sp.nov. from Microcebus gerpi , and Lemurpediculus tsimanampesotsae sp. nov. from Microcebus griseorufus . These new species are compared with all known congeneric species and identifying features are illustrated for all known species of Lemurpediculus ., Competing Interests: None., (© 2023 The Authors.)

Madagascar's flora and fauna have evolved in relative isolation since the island split from the African and Indian continents. When the last common ancestors of lemurs left Africa between 40 and 70 million years ago, they carried a subset of the viral diversity of the mainland population within them, which continued to evolve throughout the lemur radiation. Relative to other primate radiations, we know very little about the past or present viral diversity of lemurs, particularly mouse lemurs. Using high-throughput sequencing, we identified two gammaretroviruses and three betaretroviruses in the genomes of four species of wild mouse lemurs. The two gammaretroviruses and two betaretroviruses have not previously been described. One betaretrovirus was previously identified. All identified viruses are present in both Lorisiformes and Lemuriformes but absent from haplorrhine primates. The estimated ages of these viruses are consistent with the estimated divergence dates of the host lineages, suggesting they colonized the lemur genome after the Haplorrhine-Strepsirrhine split, but before the Lorisiformes-Lemuriformes split and before the colonization of Madagascar. The viral phylogenies connect multiple lineages of retroviruses from non-lemur and non-Madagascar-native species, suggesting substantial cross-species transmission occurred deep in the primate clade prior to its geographic dispersal. These phylogenies provide novel insights into known retroviral clades. They suggest that the origin of gammaretroviruses in rodents or bats may be premature and that the Jaagsiekte sheep virus clade may be older and more broadly distributed among mammals than previously thought., (© The Author(s) 2023. Published by Oxford University Press.)

The occurrence of natural hybridization has been reported in a wide range of organisms, including primates. The present study focuses on the endemic lemurs of Madagascar, primates for which only a few species occur in sympatry or parapatry with congeners, thereby creating limited opportunity for natural hybridization. This study examines RADseq data from 480 individuals to investigate whether the recent expansion of Microcebus murinus towards the northwest and subsequent secondary contact with Microcebus ravelobensis has resulted in the occurrence of hybridization between the two species. Admixture analysis identified one individual with 26% of nuclear admixture, which may correspond to an F2- or F3-hybrid. A composite-likelihood approach was subsequently used to test the fit of alternative phylogeographic scenarios to the genomic data and to date introgression. The simulations yielded support for low levels of gene flow (2Nm0 = 0.063) between the two species starting before the Last Glacial Maximum (between 54 and 142 kyr). Since M. murinus most likely colonized northwestern Madagascar during the Late Pleistocene, the rather recent secondary contact with M. ravelobensis has likely created the opportunity for occasional hybridization. Although reproductive isolation between these distantly related congeners is not complete, it is effective in maintaining species boundaries.

A high prevalence of AA-amyloidosis was identified in a breeding colony of northern tree shrews ( Tupaia belangeri ) in a retrospective analysis, with amyloid deposits in different organs being found in 26/36 individuals (72%). Amyloid deposits, confirmed by Congo red staining, were detected in kidneys, intestines, skin, and lymph nodes, characteristic of systemic amyloidosis. Immunohistochemically, the deposited amyloid was intensely positive with anti-AA-antibody (clone mc4), suggesting AA-amyloidosis. The kidneys were predominantly affected (80%), where amyloid deposits ranged from mild to severe and was predominantly located in the renal medulla. In addition, many kidneys contained numerous cysts with atrophy of the renal parenchyma. There was no significant association between concurrent neoplastic or inflammatory processes and amyloidosis. The lack of distinctive predisposing factors suggests a general susceptibility of captive T. belangeri to develop amyloidosis. Clinical and laboratory findings of a female individual with pronounced kidney alterations were indicative of renal failure. The observed tissue tropism with pronounced kidney alterations, corresponding renal dysfunction, and an overall high prevalence suggests amyloidosis as an important disease in captive tree shrews.

Background: Quaternary climate fluctuations have been acknowledged as major drivers of the geographical distribution of the extraordinary biodiversity observed in tropical biomes, including Madagascar. The main existing framework for Pleistocene Malagasy diversification assumes that forest cover was strongly shaped by warmer Interglacials (leading to forest expansion) and by cooler and arid glacials (leading to forest contraction), but predictions derived from this scenario for forest-dwelling animals have rarely been tested with genomic datasets., Results: We generated genomic data and applied three complementary demographic approaches (Stairway Plot, PSMC and IICR-simulations) to infer population size and connectivity changes for two forest-dependent primate species (Microcebus murinus and M. ravelobensis) in northwestern Madagascar. The analyses suggested major demographic changes in both species that could be interpreted in two ways, depending on underlying model assumptions (i.e., panmixia or population structure). Under panmixia, the two species exhibited larger population sizes across the Last Glacial Maximum (LGM) and towards the African Humid Period (AHP). This peak was followed by a population decline in M. ravelobensis until the present, while M. murinus may have experienced a second population expansion that was followed by a sharp decline starting 3000 years ago. In contrast, simulations under population structure suggested decreasing population connectivity between the Last Interglacial and the LGM for both species, but increased connectivity during the AHP exclusively for M. murinus., Conclusion: Our study shows that closely related species may differ in their responses to climatic events. Assuming that Pleistocene climatic conditions in the lowlands were similar to those in the Malagasy highlands, some demographic dynamics would be better explained by changes in population connectivity than in population size. However, changes in connectivity alone cannot be easily reconciled with a founder effect that was shown for M. murinus during its colonization of the northwestern Madagascar in the late Pleistocene. To decide between the two alternative models, more knowledge about historic forest dynamics in lowland habitats is necessary. Altogether, our study stresses that demographic inferences strongly depend on the underlying model assumptions. Final conclusions should therefore be based on a comparative evaluation of multiple approaches., (© 2021. The Author(s).)

Quaternary climatic changes have been invoked as important drivers of species diversification worldwide. However, the impact of such changes on vegetation and animal population dynamics in tropical regions remains debated. To overcome this uncertainty, we integrated high-resolution paleoenvironmental reconstructions from a sedimentary record covering the past 25,000 years with demographic inferences of a forest-dwelling primate species (Microcebus arnholdi), in northern Madagascar. Result comparisons suggest that climate changes through the African Humid Period (15.2 - 5.5 kyr) strongly affected the demographic dynamics of M. arnholdi. We further inferred a population decline in the last millennium which was likely shaped by the combination of climatic and anthropogenic impacts. Our findings demonstrate that population fluctuations in Malagasy wildlife were substantial prior to a significant human impact. This provides a critical knowledge of climatically driven, environmental and ecological changes in the past, which is essential to better understand the dynamics and resilience of current biodiversity., (© 2021. The Author(s).)

Deleterious effects of habitat loss and fragmentation on biodiversity have been demonstrated in numerous taxa. Although parasites represent a large part of worldwide biodiversity, they are mostly neglected in this context. We investigated the effects of various anthropogenic environmental changes on gastrointestinal parasite infections in four small mammal hosts inhabiting two landscapes of fragmented dry forest in northwestern Madagascar. Coproscopical examinations were performed on 1,418 fecal samples from 903 individuals of two mouse lemur species, Microcebus murinus ( n  = 199) and M. ravelobensis ( n  = 421), and two rodent species, the native Eliurus myoxinus ( n  = 102) and the invasive Rattus rattus ( n  = 181). Overall, sixteen parasite morphotypes were detected and significant prevalence differences between host species regarding the most common five parasites may be explained by parasite-host specificity or host behavior, diet, and socioecology. Ten host- and habitat-related ecological variables were evaluated by generalized linear mixed modeling for significant impacts on the prevalence of the most abundant gastrointestinal parasites and on gastrointestinal parasite species richness (GPSR). Forest maturation affected homoxenous parasites (direct life cycle) by increasing Lemuricola , but decreasing Enterobiinae gen. sp. prevalence, while habitat fragmentation and vegetation clearance negatively affected the prevalence of parasites with heterogenic environment (i.e., Strongyloides spp.) or heteroxenous (indirect cycle with intermediate host) cycles, and consequently reduced GPSR. Forest edges and forest degradation likely change abiotic conditions which may reduce habitat suitability for soil-transmitted helminths or required intermediate hosts. The fragility of complex parasite life cycles suggests understudied and potentially severe effects of decreasing habitat quality by fragmentation and degradation on hidden ecological networks that involve parasites. Since parasites can provide indispensable ecological services and ensure stability of ecosystems by modulating animal population dynamics and nutrient pathways, our study underlines the importance of habitat quality and integrity as key aspects of conservation., Competing Interests: No conflict of interest has been declared by the authors., (© 2021 The Authors. Ecology and Evolution published by John Wiley & Sons Ltd.)

Background: Edge effects can influence species composition and community structure as a result of changes in microenvironment and edaphic variables. We investigated effects of habitat edges on vegetation structure, abundance and body mass of one vulnerable Microcebus species in northwestern Madagascar. We trapped mouse lemurs along four 1000-m transects (total of 2424 trap nights) that ran perpendicular to the forest edge. We installed 16 pairs of 20 m 2 vegetation plots along each transect and measured nine vegetation parameters. To determine the responses of the vegetation and animals to an increasing distance to the edge, we tested the fit of four alternative mathematical functions (linear, power, logistic and unimodal) to the data and derived the depth of edge influence (DEI) for all parameters., Results: Logistic and unimodal functions best explained edge responses of vegetation parameters, and the logistic function performed best for abundance and body mass of M. ravelobensis. The DEI varied between 50 m (no. of seedlings, no. of liana, dbh of large trees [dbh ≥ 10 cm]) and 460 m (tree height of large trees) for the vegetation parameters, whereas it was 340 m for M. ravelobensis abundance and 390 m for body mass, corresponding best to the DEI of small tree [dbh < 10 cm] density (360 m). Small trees were significantly taller and the density of seedlings was higher in the interior than in the edge habitat. However, there was no significant difference in M. ravelobensis abundance and body mass between interior and edge habitats, suggesting that M. ravelobensis did not show a strong edge response in the study region. Finally, regression analyses revealed three negative (species abundance and three vegetation parameters) and two positive relationships (body mass and two vegetation parameters), suggesting an impact of vegetation structure on M. ravelobensis which may be partly independent of edge effects., Conclusions: A comparison of our results with previous findings reveals that edge effects are variable in space in a small nocturnal primate from Madagascar. Such an ecological plasticity could be extremely relevant for mitigating species responses to habitat loss and anthropogenic disturbances.

Solitary species often employ chemocommunication to facilitate mate localization. In the solitarily foraging, nocturnal mouse lemurs (Microcebus spp.), females advertise their short period of estrus acoustically and by increased scent marking, whereas males search widely for receptive females. Both sexes can be trained by operant conditioning to discriminate conspecific from heterospecific urine scent. However, it is not known, if males during and outside the reproductive season show different spontaneous interest in conspecific female urine, and if urine from estrous females elicits a higher investigation response than that from diestrous females. We established a spontaneous discrimination paradigm and quantified olfactory investigation responses of 21 captive male mouse lemurs of M. lehilahytsara and M. murinus when presenting 1 conspecific and 1 heterospecific female urine odor sample simultaneously. Overall, M. murinus investigated stimuli significantly longer than M. lehilahytsara. Moreover, males of M. murinus showed significantly longer olfactory investigation at conspecific urine samples during but not outside the reproductive season. This indicates that female urinary cues are spontaneously discriminated by male M. murinus and that this discrimination is more relevant during the reproductive season. However, males of both species did not show different responses toward urine samples from estrous versus diestrous females. Finally, male age did not correlate with the overall duration of olfactory investigation, and investigation levels were similar when testing with fresh or frozen urine samples. In conclusion, this new spontaneous discrimination paradigm provides a useful additional tool to study olfactory communication of nocturnal primates from the receiver's perspective., (© The Author(s) 2020. Published by Oxford University Press. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Introduction: The gray mouse lemur (Microcebus murinus) is an important nonhuman primate model in biomedical research. Numerous studies investigated mouse lemur behavior and possible factors underlying interindividual variation in both, animal personality and cognitive performance. Some effects, such as an age-related decline in executive functioning, have robustly been found across laboratory colonies; however, little is known about the brain structural substrates in mouse lemurs., Methods: Here, we provide first exploratory data linking in vivo magnetic resonance imaging of 34 mouse lemurs to performance in a standardized, touchscreen-based task on object discrimination and reversal learning as well as to animal personality under different scenarios in an open field., Results: High interindividual variability in both brain morphometric and behavioral measurements was found, but only few significant correlations between brain structure and behavior were revealed: Object discrimination learning was linked to the volume of the hippocampus and to temporal lobe thickness, while reversal learning was linked to thalamic volume and the thickness of the anterior cingulate lobe. Emergence latency into the open field correlated with volume of the amygdala. General exploration-avoidance in the empty open-field arena correlated with thicknesses of the anterior cingulate lobe and fronto-parietal substructures. Neophilia, assessed as exploration of a novel object placed in the arena, among others, related to the volume of the caudate nucleus., Conclusion: In summary, our data suggest a prominent role of temporal structures (including the hippocampus) for learning capability, as well as thalamic and anterior cingulate structures for cognitive flexibility and response inhibition. The amygdala, the anterior cingulate lobe, and the caudate nucleus are particularly linked to animal personality in the open-field setting. These findings are congruent with the comparative psychological literature and provide a valuable basis for future studies elucidating aspects of behavioral variation in this nonhuman primate model., (© 2020 The Authors. Brain and Behavior published by Wiley Periodicals LLC.)

Background: Constraints in migratory capabilities, such as the disruption of gene flow and genetic connectivity caused by habitat fragmentation, are known to affect genetic diversity and the long-term persistence of populations. Although negative population trends due to ongoing forest loss are widespread, the consequence of habitat fragmentation on genetic diversity, gene flow and genetic structure has rarely been investigated in Bornean small mammals. To fill this gap in knowledge, we used nuclear and mitochondrial DNA markers to assess genetic diversity, gene flow and the genetic structure in the Bornean tree shrew, Tupaia longipes, that inhabits forest fragments of the Lower Kinabatangan Wildlife Sanctuary, Sabah. Furthermore, we used these markers to assess dispersal regimes in male and female T. longipes., Results: In addition to the Kinabatangan River, a known barrier for dispersal in tree shrews, the heterogeneous landscape along the riverbanks affected the genetic structure in this species. Specifically, while in larger connected forest fragments along the northern riverbank genetic connectivity was relatively undisturbed, patterns of genetic differentiation and the distribution of mitochondrial haplotypes in a local scale indicated reduced migration on the strongly fragmented southern riverside. Especially, oil palm plantations seem to negatively affect dispersal in T. longipes. Clear sex-biased dispersal was not detected based on relatedness, assignment tests, and haplotype diversity., Conclusion: This study revealed the importance of landscape connectivity to maintain migration and gene flow between fragmented populations, and to ensure the long-term persistence of species in anthropogenically disturbed landscapes.

Acoustic phenotypic variation is of major importance for speciation and the evolution of species diversity. Whereas selective and stochastic forces shaping the acoustic divergence of signaling systems are well studied in insects, frogs, and birds, knowledge on the processes driving acoustic phenotypic evolution in mammals is limited. We quantified the acoustic variation of a call type exchanged during agonistic encounters across eight distinct species of the smallest-bodied nocturnal primate radiation, the Malagasy mouse lemurs. The species live in two different habitats (dry forest vs. humid forest), differ in geographic distance to each other, and belong to four distinct phylogenetic clades within the genus. Genetically defined species were discriminated reliably on the phenotypic level based on their acoustic distinctiveness in a discriminant function analysis. Acoustic variation was explained by genetic distance, whereas differences in morphology, forest type, or geographic distance had no effect. The strong impact of genetics was supported by a correlation between acoustic and genetic distance and the high agreement in branching pattern between the acoustic and molecular phylogenetic trees. In sum, stochastic factors such as genetic drift best explained acoustic diversification in a social communication call of mouse lemurs., Competing Interests: We have no conflict of interest to declare., (© 2020 The Authors. Ecology and Evolution published by John Wiley & Sons Ltd.)

Olfactory communication is highly important for nocturnal mammals, especially for solitary foragers, but knowledge is still limited for nocturnal primates. Mouse lemurs (Microcebus spp.) are nocturnal solitary foragers with a dispersed lifestyle and frequently use chemo-sensory signalling behaviour for governing social interactions. Different mouse lemur species can co-occur in a given forest but it is unknown whether olfaction is involved in species recognition. We first screened 24 captive mouse lemurs (9 M. murinus, 15 M. lehilahytsara) for their olfactory learning potential in an experimental arena and then tested the species discrimination ability with urine odour in an operant conditioning paradigm in four individuals. The majority of the screened animals (75%) did not pass the screening criteria within a 2-week test period. However, all four final test animals, two M. murinus and two M. lehilahytsara, were successfully trained in a 5-step-conditioning process to reliably discriminate conspecific from heterospecific urine odour (requiring an overall median of 293 trials). Findings complement previous studies on the role of acoustic signalling and suggest that olfaction may be an important additional mechanism for species discrimination.

The dynamic relationship of vector-borne parasites, arthropod vectors and their hosts is prone to change under the influence of climate change, global integration, shifting demographics and deforestation. It is therefore essential to better understand parasitism in wildlife populations, including parasites transmitted by blood-feeding vectors, and explore host range and heterogeneity of parasitic infections. We investigated Giemsa stained blood smears of two sympatric Malagasy primate species ( Microcebus murinus : 184 samples from 69 individuals and M. ravelobensis : 264 samples from 91 individuals) for blood-stage parasites and tested for a potential influence of host species, sex, body mass and sampling month on blood-stage parasite prevalence and infection intensity. No protozoan parasites were detected in either host species. A host-specific difference was observed in filarial nematode infections, with higher risk of infection in M. murinus (prevalence 30.43%), than in M. ravelobensis (prevalence 6.59%), which may be explained by differences in host behavior and/or immune competence, linked to the period of host-parasite coevolution. Neither sex nor sampling month influenced infection prevalence or intensity significantly. We did not observe a negative effect of microfilarial infections on host fitness when taking body mass as a proxy. Our results support the hypothesis of a long-term evolutionary adaptation of hosts and parasites, leading to persistent infection with low morbidity. Morphological and molecular analyses indicate the finding of a new species, " Lemurfilaria lemuris ". Genetic analysis furthermore showed >99% sequence identity with microfilariae described from a sympatric, larger-bodied lemur species of a different genus, suggesting low host-specificity of the detected filariae and pathogen transmission across genus boundaries. Findings contribute to a more comprehensive picture of vector-borne diseases of Malagasy lemurs.

Background: Social tolerance strongly influences the patterns of affiliation and aggression in animal societies. However, not much is known about the variation of social tolerance in species living in dispersed social systems that combine solitary foraging activities with the need of coordinating social interactions with conspecifics on a regular basis. This study aims to investigate the sources of variation in social tolerance within a Malagasy primate radiation with dispersed social systems, the mouse lemurs (Microcebus spp.). Six mouse lemur species were selected as model species that belong to three different taxonomic clades, live in two types of forest environments (dry and humid), and differed in this study with respect to their reproductive activity. Six male-female and six male-male dyads of each species were tested temporarily in a standardized social encounter paradigm in Madagascar to collect data on joint use of space, non-agonistic body contacts, aggression rates, the number of conflicts and the establishment of intra- and intersexual dominance., Results: Male-female dyads of the six species differed significantly in the frequency of affiliative and agonistic behaviors. In contrast, the variations between male-male dyads could not be explained by one parameter only, but clade membership, forest type, reproductive state as well as species were all suggested to be partially influential. Only one species (Microcebus mamiratra) showed signals of unambiguous female dominance in all male-female dyads, whereas the others had no or only a few dyads with female dominance., Conclusions: Variations in social tolerance and its consequences are most likely influenced by two factors, ecology (via forest type) and physiology (via reproductive activity), and only to a lesser extent by clade membership. The study suggests that mouse lemur females have higher aggression rates and more agonistic conflicts with males when females in the population are reproducing, at least in resource-rich humid forests. The study confirms a high degree of social plasticity between species in these small solitary foragers that supports their taxonomic distinctiveness and requires further scientific attention.

Here we report a case of severe growth retardation and neurologic abnormalities in a female gray mouse lemur ( Microcebus murinus ), a small NHP species for which the genomic sequence recently became available. The female lemur we present here died on postnatal day 125. This lemur had impaired development of motor skills and showed severe ataxia and tremors. In addition, hearing seemed normal whereas ophthalmic examination revealed incipient bilateral cataracts, abnormal pigmentation in the lens of the left eye, and a missing optokinetic nystagmus, which indicated impaired vision. Most prominently, the lemur showed severe growth retardation. Necropsy revealed maldevelopment of the left reproductive organs and unilateral dilation of the right lateral ventricle, which was confirmed on brain MRI. Brain histology further revealed large, bilateral areas of vacuolation within the brainstem, but immunohistochemistry indicated no sign of pathologic prion protein deposition. Full genomic sequencing of the lemur revealed a probably pathologic mutation in LARGE2 of the LARGE gene family, which has been associated with congenital muscular dystrophies. However, potentially functional mutations in other genes were also present. The observed behavioral and motor signs in the presented animal might have been linked to spongiform degeneration and resulting brainstem dysfunction and progressive muscle weakness. The macroscopic developmental abnormalities and ophthalmic findings might be genetic in origin and linked to the mutation in LARGE2 .

Background: Ectoparasitic infections are of particular interest for endangered wildlife, as ectoparasites are potential vectors for inter- and intraspecific pathogen transmission and may be indicators to assess the health status of endangered populations. Here, ectoparasite dynamics in sympatric populations of two Malagasy mouse lemur species, Microcebus murinus and M. ravelobensis, were investigated over an 11-month period. Furthermore, the animals' body mass was determined as an indicator of body condition, reflecting seasonal and environmental challenges. Living in sympatry, the two study species experience the same environmental conditions, but show distinct differences in socioecology: Microcebus murinus sleeps in tree holes, either solitarily (males) or sometimes in groups (females only), whereas M. ravelobensis sleeps in mixed-sex groups in more open vegetation., Results: Both mouse lemur species hosted ticks (Haemaphysalis sp.), lice (Lemurpediculus sp.) and mites (Trombiculidae gen. sp. and Laelaptidae gen. sp.). Host species, as well as temporal variations (month and year), were identified as the main factors influencing infestation. Tick infestation peaked in the late dry season and was significantly more often observed in M. murinus (P = 0.011), while lice infestation was more likely in M. ravelobensis (P < 0.001) and showed a continuous increase over the course of the dry season. Genetic analyses identified Lemurpediculus sp. infesting both mouse lemur species. Ticks morphologically conform to H. lemuris, but genetic analysis showed a clear differentiation of the specimens collected in this study, suggesting a potentially new tick species. Host body mass decreased from the early to the late dry season, indicating nutritional stress during this period, which may render individuals more susceptible to parasitic infections., Conclusions: Seasonal differences and species-specific variations in sleeping site ecology in terms of sleeping site type and sociality were determined as key factors influencing ectoparasitism in M. murinus and M. ravelobensis. This needs to be taken into account when evaluating ectoparasite infestations at a given time point. The detection of the same parasite species on two closely related and sympatric host species furthermore indicates a potential pathway for disease transmission, not only within but also between lemur species.

Background: Various factors, such as climate, body size and sociality are often linked to parasitism. This constrains the identification of other determinants driving parasite infections. Here, we investigate for the first time intestinal parasites in two sympatric arboreal primate species, which share similar activity patterns, feeding ecology, body size and sociality, and cope with the same climate conditions, but differ in sleeping site ecology: the Milne-Edward's sportive lemur (Lepilemur edwardsi) and the Western woolly lemur (Avahi occidentalis). Comparison of these two species aimed to test whether differences in sleeping sites are related to differences in parasite infection patterns. Additionally, gender and seasonal factors were taken into account. Animals were radio-collared to record their sleeping site dynamics and to collect fecal samples to assess intestinal parasitism during both the dry and the rainy season., Results: Only low parasite diversity was detected, which is attributable to the strict arboreal lifestyle of these lemurs, limiting their contact with infective parasite stages. L. edwardsi, which sleeps in tree holes and repeatedly uses the same sleeping site, excreted eggs of strongyle and oxyurid nematodes, whereby strongyles always occurred in coinfection with oxyurids. In contrast, A. occidentalis, which sleeps on open branches and frequently changes sleeping sites, only excreted eggs of strongyle nematodes. This difference can be attributed to a potential favorable environment presented by tree holes for infective stages, facilitating parasitic transmission. Additionally, Strongylida in A. occidentalis were only observed in the rainy season, suggesting an arrested development during the dry season in the nematodes' life cycle. Males and females of both lemur species showed the same frequency of parasitism. No differences in body mass of infected and non-infected individuals were observed, indicating that the animals' body condition remains unaffected by the detected gastrointestinal parasites., Conclusions: The comparison of two primate hosts with a very similar lifestyle suggests an influence of the sleeping site ecology on intestinal parasites. In A. occidentalis there was a clear seasonal difference in strongyle egg excretion. These results improve our understanding of the parasite ecology in these endangered primate species, which may be critical in the light of species conservation.

Understanding determinants shaping infection risk of endangered wildlife is a major topic in conservation medicine. The proboscis monkey, Nasalis larvatus, an endemic primate flagship species for conservation in Borneo, is endangered through habitat loss, but can still be found in riparian lowland and mangrove forests, and in some protected areas. To assess socioecological and anthropogenic influence on intestinal helminth infections in N. larvatus, 724 fecal samples of harem and bachelor groups, varying in size and the number of juveniles, were collected between June and October 2012 from two study sites in Malaysian Borneo: 634 samples were obtained from groups inhabiting the Lower Kinabatangan Wildlife Sanctuary (LKWS), 90 samples were collected from groups of the Labuk Bay Proboscis Monkey Sanctuary (LBPMS), where monkeys are fed on stationary feeding platforms. Parasite risk was quantified by intestinal helminth prevalence, host parasite species richness (PSR), and eggs per gram feces (epg). Generalized linear mixed effect models were applied to explore whether study site, group type, group size, the number of juveniles per group, and sampling month predict parasite risk. At the LBPMS, prevalence and epg of Trichuris spp., strongylids, and Strongyloides spp. but not Ascaris spp., as well as host PSR were significantly elevated. Only for Strongyloides spp., prevalence showed significant changes between months; at both sites, the beginning rainy season with increased precipitation was linked to higher prevalence, suggesting the external life cycle of Strongyloides spp. to benefit from humidity. Higher prevalence, epgs, and PSR within the LBPMS suggest that anthropogenic factors shape host infection risk more than socioecological factors, most likely via higher re-infection rates and chronic stress. Noninvasive measurement of fecal parasite stages is an important tool for assessing transmission dynamics and infection risks for endangered tropical wildlife. Findings will contribute to healthcare management in nature and in anthropogenically managed environments.

Mouse lemurs, the world's smallest primates, inhabit forests in Madagascar. They are nocturnal, arboreal and dependent on vision for their everyday lives. In the last decades, the grey mouse lemur became increasingly important for biomedical research, in particular aging research. Experiments which require the combination of visual fitness and old age consequently depend on a solid knowledge of ocular pathologies. Although ocular diseases in mouse lemurs have been described as being common, they have not received much attention so far. Yet it is important to know when and why ocular diseases in captive mouse lemurs may occur. This review aims to provide a comprehensive overview of known ocular findings in mouse lemurs. It summarizes the frequency of ocular findings in captive mouse lemur colonies and points to their likely causes and treatment options based on the evidence available from other animals and humans. In addition, it shall be discussed whether age or genetic background may affect their development. This review may be used as a reference for future studies which require an assessment of visual performance in mouse lemurs and help to evaluate observed clinical signs and ocular diseases. Furthermore, the high incidence of specific diseases may provide new perspectives and set the groundwork for a new animal model for ocular research., Competing Interests: The authors declare that they have no conflict of interest., (Copyright: © 2017 Marko Dubicanac et al.)

Background: Large animal models of Huntington's disease (HD) may increase the reliability of translating preclinical findings to humans. Long live expectancy offers opportunities particularly for disease modifying approaches, but also challenges. The transgenic (tg) HD minipig model assessed in this study exhibits a high genetic homology with humans, similar body weight, and comparable brain structures. To test long-term safety, tolerability, and efficacy of novel therapeutic approaches in this model reliable assessments applicable longitudinally for several years are warranted for all phenotypical domains relevant in HD., Objective: To investigate whether the tests proposed assessing motor, cognitive and behavioral domains can be applied repetitively over a 3-year period in minipigs with acceptable variability or learning effects and whether tgHD minipigs reveal changes in these domains compared to wildtype (wt) minipigs suggesting the development of an HD phenotype., Methods: A cohort of 14 tgHD and 18 wt minipigs was followed for three years. Tests applied every six months included a tongue coordination and hurdle test for the motor domain, a color discrimination test for cognition, and a dominance test for assessing behavior. Statistical analyses were performed using repeated ANOVA for longitudinal group comparisons and Wilcoxon-tests for intra-visit differences between tgHD and wt minipigs., Results: All tests applied demonstrated feasibility, acceptable variance and good consistency during the three-year period. No significant differences between tgHD and wt minipigs were detected suggesting lack of a phenotype before the age of four years., Conclusions: The assessment battery presented offers measures in all domains relevant for HD and can be applied in long-term phenotyping studies with tgHD minipigs. The observation of this cohort should be continued to explore the timeline of phenotype development and provide information for future interventional studies.

Background: A central question in evolutionary parasitology is to what extent ecology impacts patterns of parasitism in wild host populations. In this study, we aim to disentangle factors influencing the risk of parasite exposure by exploring the impact of sleeping site ecology on infection with ectoparasites and vector-borne hemoparasites in two sympatric primates endemic to Madagascar. Both species live in the same dry deciduous forest of northwestern Madagascar and cope with the same climatic constraints, they are arboreal, nocturnal, cat-sized and pair-living but differ prominently in sleeping site ecology. The Western woolly lemur ( Avahi occidentalis ) sleeps on open branches and frequently changes sleeping sites, whereas the Milne-Edward's sportive lemur ( Lepilemur edwardsi ) uses tree holes, displaying strong sleeping site fidelity. Sleeping in tree holes should confer protection from mosquito-borne hemoparasites, but should enhance the risk for ectoparasite infestation with mites and nest-adapted ticks. Sex may affect parasite risk in both species comparably, with males bearing a higher risk than females due to an immunosuppressive effect of higher testosterone levels in males or to sex-specific behavior. To explore these hypotheses, ectoparasites and blood samples were collected from 22 individuals of A. occidentalis and 26 individuals of L. edwardsi during the dry and rainy season., Results: L. edwardsi, but not A. occidentalis , harbored ectoparasites, namely ticks ( Haemaphysalis lemuris [Ixodidae], Ornithodoros sp. [Argasidae]) and mites ( Aetholaelaps trilyssa , [Laelapidae]), suggesting that sleeping in tree holes promotes infestation with ectoparasites. Interestingly, ectoparasites were found solely in the hot, rainy season with a prevalence of 75% ( N  = 16 animals). Blood smears were screened for the presence and infection intensity of hemoparasites. Microfilariae were detected in both species. Morphological characteristics suggested that each lemur species harbored two different filarial species. Prevalence of microfilarial infection was significantly lower in L. edwardsi than in A. occidentalis. No significant difference in infection intensity between the two host species, and no effect of season, daytime of sampling or sex on prevalence or infection intensity was found. In neither host species, parasite infection showed an influence on body weight as an indicator for body condition., Conclusions: Our findings support that sleeping site ecology affects ectoparasite infestation in nocturnal, arboreal mammalian hosts in the tropics, whereas there is no significant effect of host sex. The influence of sleeping site ecology to vector-borne hemoparasite risk is less pronounced. The observed parasite infections did not affect body condition and thus may be of minor importance for shaping reproductive fitness. Findings provide first evidence for the specific relevance of sleeping site ecology on parasitism in arboreal and social mammals. Further, our results increase the sparse knowledge on ecological drivers of primate host-parasite interactions and transmission pathways in natural tropical environments.

Non-human primates of South-East Asia remain under-studied concerning parasite epidemiology and co-infection patterns. Simultaneously, efforts in conservation demand knowledge of parasite abundance and biodiversity in threatened species. The Endangered proboscis monkey, Nasalis larvatus , a primate flagship species for conservation in Borneo, was investigated in the present study. Habitat loss and fragmentation are among the greatest threats to bachelor and harem groups of this folivorous colobine. Designed as a follow-up study, prevalence and co-infection status of intestinal parasites from N. larvatus in a protected area in Malaysian Borneo were analyzed from fecal samples using a flotation method. For the first time, the intestinal parasite co-infection patterns were examined using quantitative analyses. Overall, 92.3% of fecal samples (N = 652) were positive for helminth eggs. Five helminth groups were detected: (1) trichurids (82.7% prevalence) including Trichuris spp. (82.1%) and Anatrichosoma spp. (1.4%), (2) strongyles (58.9%) including Trichostrongylus spp. (48.5%) and Oesophagostomum/Ternidens spp. (22.8%), (3) Strongyloides fuelleborni (32.7%), (4) Ascaris lumbricoides (8.6%), and (5) Enterobius spp. (5.5%). On average, an individual was co-infected with two different groups. Significant positive associations were found for co-infections of trichurids with strongyles and S. fuelleborni as well as S. fuelleborni with A. lumbricoides and strongyles. This study shows a high prevalence of various gastrointestinal helminths with potential transmission pathways primarily related to soil and with zoonotic relevance in wild proboscis monkeys in their remaining natural habitats. Observed positive associations of trichurids with strongyles and Strongyloides spp. may result from the high prevalence of trichurids. Similarly, positive associations between Strongyloides and Ascaris were found, both of which typically occur predominantly in juvenile hosts. These findings should be considered when proposing conservation actions in altered habitats nearby human settlements and when managing captive populations.

Background: A major effort is underway to use population genetic approaches to identify loci involved in adaptation. One issue that has so far received limited attention is whether loci that show a phylogenetic signal of positive selection in the past also show evidence of ongoing positive selection at the population level. We address this issue using vomeronasal receptors (VRs), a diverse gene family in mammals involved in intraspecific communication and predator detection. In mouse lemurs, we previously demonstrated that both subfamilies of VRs (V1Rs and V2Rs) show a strong signal of directional selection in interspecific analyses. We predicted that ongoing sexual selection and/or co-evolution with predators may lead to current directional or balancing selection on VRs. Here, we re-sequence 17 VRs and perform a suite of selection and demographic analyses in sympatric populations of two species of mouse lemurs (Microcebus murinus and M. ravelobensis) in northwestern Madagascar., Results: M. ravelobensis had consistently higher genetic diversity at VRs than M. murinus. In general, we find little evidence for positive selection, with most loci evolving under purifying selection and one locus even showing evidence of functional loss in M. ravelobensis. However, a few loci in M. ravelobensis show potential evidence of positive selection. Using mismatch distributions and expansion models, we infer a more recent colonisation of the habitat by M. murinus than by M. ravelobensis, which most likely speciated in this region earlier on., Conclusions: These findings suggest that the analysis of VR variation is useful in inferring demographic and phylogeographic history of mouse lemurs. In conclusion, this study reveals a substantial heterogeneity over time in selection on VR loci, suggesting that VR evolution is episodic.

The vomeronasal organ (VNO) is functional in most terrestrial mammals, though progressively reduced in the primate lineage, and is used for intraspecific communication and predator recognition. Vomeronasal receptor (VR) genes comprise two families of chemosensory genes (V1R and V2R) that have been considered to be specific for the VNO. However, recently a large number of VRs were reported to be expressed in the main olfactory epithelium (MOE) of mice, but there is little knowledge of the expression of these genes outside of rodents. To explore the function of VR genes in mammalian evolution, we analyzed and compared the expression of 64 V1R and 2 V2R genes in the VNO and the MOE of the gray mouse lemur (Microcebus murinus), the primate with the largest known VR repertoire. We furthermore compared expression patterns in adults of both sexes and seasons, and in an infant. A large proportion (83-97%) of the VR loci was expressed in the VNO of all individuals. The repertoire in the infant was as rich as in adults, indicating reliance on olfactory communication from early postnatal development onwards. In concordance with mice, we also detected extensive expression of VRs in the MOE, with proportions of expressed loci in individuals ranging from 29 to 45%. TRPC2, which encodes a channel protein crucial for signal transduction via VRs, was co-expressed in the MOE in all individuals indicating likely functionality of expressed VR genes in the MOE. In summary, the large VR repertoire in mouse lemurs seems to be highly functional. Given the differences in the neural pathways of MOE and VNO signals, which project to higher cortical brain centers or the limbic system, respectively, this raises the intriguing possibility that the evolution of MOE-expression of VRs enabled mouse lemurs to adaptively diversify the processing of VR-encoded olfactory information.

Introduction: Maternal kin selection is a driving force in the evolution of mammalian social complexity and it requires that kin are distinctive from nonkin. The transition from the ancestral state of asociality to the derived state of complex social groups is thought to have occurred via solitary foraging, in which individuals forage alone, but, unlike the asocial ancestors, maintain dispersed social networks via scent-marks and vocalizations. We hypothesize that matrilineal signatures in vocalizations were an important part of these networks. We used the solitary foraging gray mouse lemur (Microcebus murinus) as a model for ancestral solitary foragers and tested for matrilineal signatures in their calls, thus investigating whether such signatures are already present in solitary foragers and could have facilitated the kin selection thought to have driven the evolution of increased social complexity in mammals. Because agonism can be very costly, selection for matrilineal signatures in agonistic calls should help reduce agonism between unfamiliar matrilineal kin. We conducted this study on a well-studied population of wild mouse lemurs at Ankarafantsika National Park, Madagascar. We determined pairwise relatedness using seven microsatellite loci, matrilineal relatedness by sequencing the mitrochondrial D-loop, and sleeping group associations using radio-telemetry. We recorded agonistic calls during controlled social encounters and conducted a multi-parametric acoustic analysis to determine the spectral and temporal structure of the agonistic calls. We measured 10 calls for each of 16 females from six different matrilineal kin groups., Results: Calls were assigned to their matriline at a rate significantly higher than chance (pDFA: correct = 47.1%, chance = 26.7%, p = 0.03). There was a statistical trend for a negative correlation between acoustic distance and relatedness (Mantel Test: g = -1.61, Z = 4.61, r = -0.13, p = 0.058)., Conclusions: Mouse lemur agonistic calls are moderately distinctive by matriline. Because sleeping groups consisted of close maternal kin, both genetics and social learning may have generated these acoustic signatures. As mouse lemurs are models for solitary foragers, we recommend further studies testing whether the lemurs use these calls to recognize kin. This would enable further modeling of how kin recognition in ancestral species could have shaped the evolution of complex sociality.

Two classes of vomeronasal receptor genes, V1R and V2R, occur in vertebrates. Whereas, V1R loci are found in a wide variety of mammals, including primates, intact V2R genes have thus far only been described in rodents and marsupials. In primates, the V2R repertoire has been considered degenerate. Here, we identify for the first time two intact V2R loci in a strepsirrhine primate, the grey mouse lemur (Microcebus murinus), and demonstrate their expression in the vomeronasal organ. Putatively functional orthologues are present in two other strepsirrhines, whereas, both loci are pseudogenes in a range of anthropoid species. The functional significance of the loci is unknown, but positive selection on one of them is consistent with an adaptive role in pheromone detection. Finally, conservation of V2R loci in strepsirrhines is notable, given their high diversity and role in MUP and MHC detection in rodents.

Chemosensory genes are frequently the target of positive selection and are often present in large gene families, but little is known about heterogeneity of selection in these cases and its relation to function. Here, we use the vomeronasal-1 receptor (V1R) repertoire of mouse lemurs (Microcebus spp.) as a model system to study patterns of selection of chemosensory genes at several different levels. Mouse lemurs are small nocturnal strepsirrhine primates and have a large (~200 loci) repertoire of V1R loci that are likely important for intraspecific pheromonal communication and interspecific interactions, for example, recognition of predator cues. We investigated signals and patterns of positive selection among the 105 identified full length V1R loci in the gray mouse lemur and within 7 V1R loci amplified across multiple mouse lemur species. Phylogenetic reconstructions of published sequences revealed at least nine monophyletic clusters of V1Rs in gray mouse lemurs that have diversified since the split between lemurs and lorisoid primates. A large majority of clusters evolved under significant positive selection. Similar results were found in V1Rs of closely related greater galagos. Comparison with function of related V1R clusters in mice suggested a potential relationship between receptor function and strength of selection. Interestingly, most codons identified as being under positive selection are located in the extracellular domains of the receptors and hence likely indicate the position of residues involved in ligand binding. Positive selection was also detected within five V1R loci (=71% of analyzed loci) sequenced from 6 to 10 mouse lemur species, indicating ongoing selection within the genus, which may be related to sexual selection and, potentially, speciation processes. Variation in strength of positive selection on V1Rs showed no simple relationship to cluster size. The diversity of V1R loci in mouse lemurs reflects their adaptive evolution and is most likely related to the fundamental relevance of olfactory communication and predator recognition in these primates. Overall, adaptive evolution is the predominant mode of evolution of V1R loci at all levels, and the substantial heterogeneity in the strength of selection may be related to receptor function.

Lentiviruses, the genus of retrovirus that includes HIV-1, rarely endogenize. Some lemurs uniquely possess an endogenous lentivirus called PSIV ("prosimian immunodeficiency virus"). Thus, lemurs provide the opportunity to study the activity of host defense factors, such as TRIM5α, in the setting of germ line invasion. We characterized the activities of TRIM5α proteins from two distant lemurs against exogenous retroviruses and a chimeric PSIV. TRIM5α from gray mouse lemur, which carries PSIV in its genome, exhibited the narrowest restriction activity. One allelic variant of gray mouse lemur TRIM5α restricted only N-tropic murine leukemia virus (N-MLV), while a second variant restricted N-MLV and, uniquely, B-tropic MLV (B-MLV); both variants poorly blocked PSIV. In contrast, TRIM5α from ring-tailed lemur, which does not contain PSIV in its genome, revealed one of the broadest antiviral activities reported to date against lentiviruses, including PSIV. Investigation into the antiviral specificity of ring-tailed lemur TRIM5α demonstrated a major contribution of a 32-amino-acid expansion in variable region 2 (v2) of the B30.2/SPRY domain to the breadth of restriction. Data on lemur TRIM5α and the prediction of ancestral simian sequences hint at an evolutionary scenario where antiretroviral specificity is prominently defined by the lineage-specific expansion of the variable loops of B30.2/SPRY.

Background: Pleistocene events have shaped the phylogeography of many taxa worldwide. Their genetic signatures in tropical species have been much less explored than in those living in temperate regions. We analysed the genetic structure of a Malagasy primate species, a mouse lemur with a wide distribution (M. murinus), in order to investigate such phylogeographic processes on a large tropical island. We also evaluated the effects of anthropogenic pressures (fragmentation/deforestation) and natural features (geographic distance, rivers) on genetic structure in order to complement our understanding of past and present processes of genetic differentiation., Results: The analysis of the mitochondrial D-loop sequences of 195 samples from 15 study sites (10 from a continuous forest and five from isolated forest fragments) from two adjacent Inter-River-Systems (IRSs) revealed that forest fragmentation and the river restrict gene flow, thereby leading to an increased genetic differentiation between populations beyond the effect of isolation-by-distance. Demographic simulations detected signals of two successive spatial expansions that could be preliminarily dated to the late Pleistocene and early Holocene. The haplotype network revealed geographic structure and showed deep molecular divergences within and between the IRSs that would be congruent with a two-step colonization scenario., Conclusions: This study supports the hypothesis of a relatively recent spatial expansion of the grey mouse lemur in northwestern Madagascar, which may also explain why this taxon, in contrast to its congeners, has not yet undergone allopatric speciation in the studied area and possibly across its presently wide range.

Background: The lemurs of Madagascar provide an excellent mammalian radiation to explore mechanisms and processes favouring species diversity and evolution. Species diversity, in particular of nocturnal species, increased considerably during the last decade. However, the factors contributing to this high diversity are not well understood. We tested predictions derived from two existing biogeographic models by exploring the genetic and morphological divergence among populations of a widely distributed lemur genus, the sportive lemur (Lepilemur ssp.) along a 560 km long transect from western to northern Madagascar., Results: By using the phylogenetic analyses of mtDNA sequence data, molecular diagnostic sites and phenotypic morphometric traits, we uncovered two previously undetected species whose distributions contradict the two existing biogeographic models. Brief species descriptions are provided and a new biogeographic model is proposed (the "large river model")., Conclusion: According to the "large river model", large rivers in north and northwestern Madagascar acted as geographical barriers for gene flow and facilitated speciation events on a much smaller spatial scale than previously thought. Thereby, this study does not only show that species diversity in nocturnal Malagasy primates is continuously underestimated but aims to emphasize the need for conservation actions if those species with small ranges shall not face extinction in the near future.

It is widely accepted that natal philopatry is a prerequisite for the evolution of sociality. The life-history hypothesis maintains that longevity of adults results in extended territory tenure and thus limits breeding vacancies for offspring, which makes natal philopatry more likely. Here, we tested the importance of longevity for natal philopatry in females of a basal primate, the grey mouse lemur (Microcebus murinus). This species is regarded as being solitary due to its foraging habits but while males disperse, female offspring in this species forgo dispersal and form long-term sleeping groups with their mothers. We tested whether high adult survival could be a cause for natal philopatry of female offspring. In addition, we assessed costs and benefits associated with space sharing between mothers and daughters and whether mothers actively increase survival of daughters by beqeauthal of territories, information transfer about resources or thermoregulation. Contrary to our predictions, adult females had low-survival rates. Space sharing appeared to improve survival of both, mothers and daughters. This could be a result of information transfer about sleeping sites and thermoregulatory benefits. Our results cast doubt on the idea that longevity predisposes species for social traits and provide support for benefits of philopatry.