Isolation of Bacteria from Freeze-Dried Samples and the Functional Characterization of Species-Specific Lactic Acid Bacteria with a Comparison of Wild and Captive Proboscis Monkeys.

Previously, we isolated a novel lactic acid bacteria species (Lactobacillus nasalidis) from the fresh forestomach contents of a captive proboscis monkey (Nasalis larvatus) in a Japanese zoo. In this study, we isolated two strains of L. nasalidis from the freeze-dried forestomach contents of a wild proboscis monkey inhabiting a riverine forest in Malaysia. The samples had been stored for more than six years. Phenotypic analysis showed that strains isolated from the wild individual had more diverse sugar utilization and lower salt tolerance than strains previously isolated from the captive counterpart. These phenotypic differences are most likely induced by feeding conditions; wild individuals consume a wide variety of natural food, unlike their zoo-raised counterparts that consume formula feed with sodium sufficiency. Since 16s rRNA sequences of L. nasalidis were detected in the previously created 16S rRNA libraries of wild, provisioned, and captive proboscis monkeys in Malaysia and Japan, L. nasalidis may be an essential bacterium of the foregut microbial community of the proboscis monkey. The currently established method for the isolation of gut bacteria from freeze-dried samples under storage will be applicable to many already-stored precious samples.

Large male proboscis monkeys have larger noses but smaller canines.

The uniquely enlarged noses of male proboscis monkeys are prominent adornments, and a sexually selected male trait. A recent study showed significant correlations among nose, body, and testis sizes and clear associations between nose size and the number of females in a male's harem. However, to date, the analyses of other common male traits, i.e., canines, are lacking. Whereas male nose size had a positive correlation with body size, we unexpectedly found a negative correlation between body and canine sizes. We explain this by an interaction between sexual and natural selection. Larger noses in males may interfere with the use of canines, thereby reducing their effectiveness as weapons. Additionally, longer canines are opposed by natural selection because the larger gape it imposes upon its bearer reduces foraging efficiency, particularly in folivores. This unique case of decoupling of body and canine size reveals that large canines carry an ecological cost.

First report of foregut microbial community in proboscis monkeys: are diverse forests a reservoir for diverse microbiomes?

Foregut fermentation is well known to occur in a wide range of mammalian species and in a single bird species. Yet, the foregut microbial community of free-ranging, foregut-fermenting monkeys, that is, colobines, has not been investigated so far. We analysed the foregut microbiomes in four free-ranging proboscis monkeys (Nasalis larvatus) from two different tropical habitats with varying plant diversity (mangrove and riverine forests), in an individual from a semi-free-ranging setting with supplemental feeding, and in an individual from captivity, using high-throughput sequencing based on 16S ribosomal RNA genes. We found a decrease in foregut microbial diversity from a diverse natural habitat (riverine forest) to a low diverse natural habitat (mangrove forest), to human-related environments. Of a total of 2700 bacterial operational taxonomic units (OTUs) detected in all environments, only 153 OTUs were shared across all individuals, suggesting that they were not influenced by diet or habitat. These OTUs were dominated by Firmicutes and Proteobacteria. The relative abundance of the habitat-specific microbial communities showed a wide range of differences among living environments, although such bacterial communities appeared to be dominated by Firmicutes and Bacteroidetes, suggesting that those phyla are key to understanding the adaptive strategy in proboscis monkeys living in different habitats.