Non-structural genes of novel lemur adenoviruses reveal codivergence of virus and host.

Adenoviruses (AdVs) are important human and animal pathogens and are frequently used as vectors for gene therapy and vaccine delivery. Surprisingly, there are only scant data regarding primate AdV origin and evolution, especially in the most basal primate hosts. We detect and sequence AdVs from faeces of two Madagascan lemur species. Complete genome sequence analyses define a new AdV species with a particularly large gene encoding a protein of unknown function in the early gene region 3. Unexpectedly, the new AdV species is not most similar to human or other simian AdVs but to bat adenovirus C. Genome characterisation shows signals of virus-host codivergence in non-structural genes, which show lower diversity than structural genes. Outside a lemur species mixing zone, recombination less frequently separates structural genes, as in human adenovirus C. The evolutionary history of lemur AdVs likely involves both a host switch and codivergence with the lemur hosts.

RADseq data reveal a lack of admixture in a mouse lemur contact zone contrary to previous microsatellite results.

Microsatellites have been a workhorse of evolutionary genetic studies for decades and are still commonly in use for estimating signatures of genetic diversity at the population and species level across a multitude of taxa. Yet, the very high mutation rate of these loci is a double-edged sword, conferring great sensitivity at shallow levels of analysis (e.g. paternity analysis) but yielding considerable uncertainty for deeper evolutionary comparisons. For the present study, we used reduced representation genome-wide data (restriction site-associated DNA sequencing (RADseq)) to test for patterns of interspecific hybridization previously characterized using microsatellite data in a contact zone between two closely related mouse lemur species in Madagascar (Microcebus murinus and Microcebus griseorufus). We revisit this system by examining populations in, near, and far from the contact zone, including many of the same individuals that had previously been identified as hybrids with microsatellite data. Surprisingly, we find no evidence for admixed nuclear ancestry. Instead, re-analyses of microsatellite data and simulations suggest that previously inferred hybrids were false positives and that the program NewHybrids can be particularly sensitive to erroneously inferring hybrid ancestry. Combined with results from coalescent-based analyses and evidence for local syntopic co-occurrence, we conclude that the two mouse lemur species are in fact completely reproductively isolated, thus providing a new understanding of the evolutionary rate whereby reproductive isolation can be achieved in a primate.

Evidence of MHC class I and II influencing viral and helminth infection via the microbiome in a non-human primate.

Until recently, the study of major histocompability complex (MHC) mediated immunity has focused on the direct link between MHC diversity and susceptibility to parasite infection. However, MHC genes can also influence host health indirectly through the sculpting of the bacterial community that in turn shape immune responses. We investigated the links between MHC class I and II gene diversity gut microbiome diversity and micro- (adenovirus, AdV) and macro- (helminth) parasite infection probabilities in a wild population of non-human primates, mouse lemurs of Madagascar. This setup encompasses a plethora of underlying interactions between parasites, microbes and adaptive immunity in natural populations. Both MHC classes explained shifts in microbiome composition and the effect was driven by a few select microbial taxa. Among them were three taxa (Odoribacter, Campylobacter and Prevotellaceae-UCG-001) which were in turn linked to AdV and helminth infection status, correlative evidence of the indirect effect of the MHC via the microbiome. Our study provides support for the coupled role of MHC diversity and microbial flora as contributing factors of parasite infection.

Challenges of next-generation sequencing in conservation management: Insights from long-term monitoring of corridor effects on the genetic diversity of mouse lemurs in a fragmented landscape.

Long-term genetic monitoring of populations is essential for efforts aimed at preserving genetic diversity of endangered species. Here, we employ a framework of long-term genetic monitoring to evaluate the effects of fragmentation and the effectiveness of the establishment of corridors in restoring population connectivity and genetic diversity of mouse lemurs Microcebus ganzhorni. To this end, we supplement estimates of neutral genetic diversity with the assessment of adaptive genetic variability of the major histocompatibility complex (MHC). In addition, we address the challenges of long-term genetic monitoring of functional diversity by comparing the genotyping performance and estimates of MHC variability generated by single-stranded conformation polymorphism (SSCP)/Sanger sequencing with those obtained by high-throughput sequencing (next-generation sequencing [NGS], Illumina), an issue that is particularly relevant when previous work serves as a baseline for planning management strategies that aim to ensure the viability of a population. We report that SSCP greatly underestimates individual diversity and that discrepancies in estimates of MHC diversity attributable to the comparisons of traditional and NGS genotyping techniques can influence the conclusions drawn from conservation management scenarios. Evidence of migration among fragments in Mandena suggests that mouse lemurs are robust to the process of fragmentation and that the effect of corridors is masked by ongoing gene flow. Nonetheless, results based on a larger number of shared private alleles at neutral loci between fragment pairs found after the establishment of corridors in Mandena suggest that gene flow is augmented as a result of enhanced connectivity. Our data point out that despite low effective population size, M. ganzhorni maintains high individual heterozygosity at neutral loci and at MHC II DRB gene and that selection plays a predominant role in maintaining MHC diversity. These findings highlight the importance of long-term genetic monitoring in order to disentangle between the processes of drift and selection maintaining adaptive genetic diversity in small populations.

Tooth wear patterns in black rats (Rattus rattus) of Madagascar differ more in relation to human impact than to differences in natural habitats.

Dietary characteristics and environmental variables are important selective factors directing ecological diversification in rodents. On Madagascar, the introductions and spread of the commensal black rat (Rattus rattus) can be seen as example cases to study dietary niche occupation and dietary adaptation in an insular environment. We investigate how tooth wear as a measure of dietary adaptation of black rats differs between four distinct habitats (village, manioc fields, spiny forest, and rainforest) with different dietary resources. We use the 3D surface texture analysis (3DST, using 30 parameters according to ISO 25178) as a measure of dietary abrasiveness. 3DST is applied on the occlusal surface of the upper first molar of 37 black rat specimens. The rainforest sample displays less rough and less voluminous surface textures compared to the village samples as indicated by smaller values for height parameters (Sa, Sp, Sq), inverse areal material ratio (Smc), and volume parameters (Vm, Vmc, Vmp, Vv, and Vvc). We therefore rank sampling areas from highest to lowest abrasiveness (village>manioc fields/spiny forest>rainforest). The rats from villages and rainforest differ to such an extent that one could have interpreted them to belong to different species. This indicates a high degree of variability in terms of ingesta abrasiveness. Furthermore, the pronounced difference between rats from human habitations compared to rats from associated fields or natural vegetation is interpreted to clearly indicate shifts in dietary niche occupation in relation to human impact.

Hybridization of mouse lemurs: different patterns under different ecological conditions.

BACKGROUND: Several mechanistic models aim to explain the diversification of the multitude of endemic species on Madagascar. The island's biogeographic history probably offered numerous opportunities for secondary contact and subsequent hybridization. Existing diversification models do not consider a possible role of these processes. One key question for a better understanding of their potential importance is how they are influenced by different environmental settings. Here, we characterized a contact zone between two species of mouse lemurs, Microcebus griseorufus and M. murinus, in dry spiny bush and mesic gallery forest that border each other sharply without intermediate habitats between them. We performed population genetic analyses based on mtDNA sequences and nine nuclear microsatellites and compared the results to a known hybrid zone of the same species in a nearby wide gradient from dry spiny bush over transitional forest to humid littoral forest. RESULTS: In the spiny-gallery system, Microcebus griseorufus is restricted to the spiny bush; Microcebus murinus occurs in gallery forest and locally invades the dryer habitat of its congener. We found evidence for bidirectional introgressive hybridization, which is closely linked to increased spatial overlap within the spiny bush. Within 159 individuals, we observed 18 hybrids with mitochondrial haplotypes of both species. Analyses of simulated microsatellite data indicate that we identified hybrids with great accuracy and that we probably underestimated their true number. We discuss short-term climatic fluctuations as potential trigger for the dynamic of invasion and subsequent hybridization. In the gradient hybrid zone in turn, long-term aridification could have favored unidirectional nuclear introgression from Microcebus griseorufus into M. murinus in transitional forest. CONCLUSIONS: Madagascar's southeastern transitional zone harbors two very different hybrid zones of mouse lemurs in different environmental settings. This sheds light on the multitude of opportunities for the formation of hybrid zones and indicates an important influence of environmental factors on secondary contact and hybridization. Our findings suggest that hybridization could enhance the adaptability of mouse lemurs without necessarily leading to a loss of distinctiveness. They point to a potential role of hybridization in Madagascar's diversification history that requires further investigation.

Regional, seasonal and interspecific variation in 15N and 13C in sympatric mouse lemurs.

Madagascar provides some of the rare examples where two or more primate species of the same genus and with seemingly identical niche requirements occur in sympatry. If congeneric primate species co-occur in other parts of the world, they differ in size in a way that is consistent with Hutchinson's rule for coexisting species, or they occupy different ecological niches. In some areas of Madagascar, mouse lemurs do not follow these "rules" and thus seem to violate one of the principles of community ecology. In order to understand the mechanisms that allow coexistence of sympatric congeneric species without obvious niche differentiation, we studied food composition of two identical sized omnivorous mouse lemur species, Microcebus griseorufus and M. murinus with the help of stable isotope analyses (δ(15)N and δ(13)C). The two species are closely related sister species. During the rich season, when food seems abundant, the two species do not differ in their nitrogen isotope composition, indicating that the two species occupy the same trophic level. But they differ in their δ(13)C values, indicating that M. griseorufus feeds more on C(4) and CAM (Crassulacean-acid-metabolism) plants than M. murinus. During the lean season, M. murinus has lower δ(15)N values, indicating that the two species feed at different trophic levels during times of food shortage. Hybrids between the two species showed intermediate food composition. The results reflect subtle differences in foraging or metabolic adaptations that are difficult to quantify by traditional observations but that represent possibilities to allow coexistence of species.

Habitat separation of sympatric Microcebus spp. in the dry spiny forest of south-eastern Madagascar.

We investigated whether or not habitat structure contributes to the separation of two sister species of lemurs and their hybrids. For this, we studied Microcebus murinus and M. griseorufus along a continuous vegetation gradient where populations of the two species occur in sympatry or in allopatry. In allopatry, the two species are generalists without any sign of microhabitat selectivity. In sympatry, both species differed significantly and discriminated against certain habitat structures: M. murinus was found in microhabitats with larger trees than average while M. griseorufus utilized microhabitats with smaller trees. Hybrids between the two species did not show any significant discrimination for or against microhabitat structure and did not differ in their habitat utilization from either parent species. Both species can go into torpor and hibernation. M. griseorufus is seen more frequently during the cool dry season than M. murinus. We assume that M. murinus goes into extended torpor or hibernation more frequently than M. griseorufus. We interpret the different occurrence of large-sized trees in microhabitats of M. murinus as a prerequisite for M. murinus to be able to spend extended periods of time in tree holes that are isolated and allow hibernation at reduced temperature levels.