Microbial rewilding in the gut microbiomes of captive ring-tailed lemurs (Lemur catta) in Madagascar.

Microbial rewilding, whereby exposure to naturalistic environments can modulate or augment gut microbiomes and improve host-microbe symbiosis, is being harnessed as an innovative approach to human health, one that may also have significant value to animal care and conservation. To test for microbial rewilding in animal microbiomes, we used a unique population of wild-born ring-tailed lemurs (Lemur catta) that were initially held as illegal pets in unnatural settings and, subsequently, relocated to a rescue center in Madagascar where they live in naturalistic environments. Using amplicon and shotgun metagenomic sequencing of lemur and environmental microbiomes, we found multiple lines of evidence for microbial rewilding in lemurs that were transitioned from unnatural to naturalistic environments: A lemur's duration of exposure to naturalistic settings significantly correlated with (a) increased compositional similarly to the gut communities of wild lemurs, (b) decreased proportions of antibiotic resistance genes that were likely acquired via human contact during pethood, and (c) greater covariation with soil microbiomes from natural habitats. Beyond the inherent psychosocial value of naturalistic environments, we find that actions, such as providing appropriate diets, minimizing contact with humans, and increasing exposure to natural environmental consortia, may assist in maximizing host-microbe symbiosis in animals under human care.

Gut microbiota of ring-tailed lemurs (Lemur catta) vary across natural and captive populations and correlate with environmental microbiota.

BACKGROUND: Inter-population variation in host-associated microbiota reflects differences in the hosts' environments, but this characterization is typically based on studies comparing few populations. The diversity of natural habitats and captivity conditions occupied by any given host species has not been captured in these comparisons. Moreover, intraspecific variation in gut microbiota, generally attributed to diet, may also stem from differential acquisition of environmental microbes-an understudied mechanism by which host microbiomes are directly shaped by environmental microbes. To more comprehensively characterize gut microbiota in an ecologically flexible host, the ring-tailed lemur (Lemur catta; n = 209), while also investigating the role of environmental acquisition, we used 16S rRNA sequencing of lemur gut and soil microbiota sampled from up to 13 settings, eight in the wilderness of Madagascar and five in captivity in Madagascar or the U.S. Based on matched fecal and soil samples, we used microbial source tracking to examine covariation between the two types of consortia. RESULTS: The diversity of lemur gut microbes varied markedly within and between settings. Microbial diversity was not consistently greater in wild than in captive lemurs, indicating that this metric is not necessarily an indicator of host habitat or environmental condition. Variation in microbial composition was inconsistent both with a single, representative gut community for wild conspecifics and with a universal 'signal of captivity' that homogenizes the gut consortia of captive animals. Despite the similar, commercial diets of captive lemurs on both continents, lemur gut microbiomes within Madagascar were compositionally most similar, suggesting that non-dietary factors govern some of the variability. In particular, soil microbial communities varied across geographic locations, with the few samples from different continents being the most distinct, and there was significant and context-specific covariation between gut and soil microbiota. CONCLUSIONS: As one of the broadest, single-species investigations of primate microbiota, our study highlights that gut consortia are sensitive to multiple scales of environmental differences. This finding begs a reevaluation of the simple 'captive vs. wild' dichotomy. Beyond the important implications for animal care, health, and conservation, our finding that environmental acquisition may mediate aspects of host-associated consortia further expands the framework for how host-associated and environmental microbes interact across different microbial landscapes.

Lessons Learned and Paths Forward for Rabies Dog Vaccination in Madagascar: A Case Study of Pilot Vaccination Campaigns in Moramanga District.

Canine rabies causes an estimated 60,000 human deaths per year, but these deaths are preventable through post-exposure prophylaxis of people and vaccination of domestic dogs. Dog vaccination campaigns targeting 70% of the population are effective at interrupting transmission. Here, we report on lessons learned during pilot dog vaccination campaigns in the Moramanga District of Madagascar. We compare two different vaccination strategies: a volunteer-driven effort to vaccinate dogs in two communes using static point vaccination and continuous vaccination as part of routine veterinary services. We used dog age data from the campaigns to estimate key demographic parameters and to simulate different vaccination strategies. Overall, we found that dog vaccination was feasible and that most dogs were accessible to vaccination. The static-point campaign achieved higher coverage but required more resources and had a limited geographic scope compared to the continuous delivery campaign. Our modeling results suggest that targeting puppies through community-based vaccination efforts could improve coverage. We found that mass dog vaccination is feasible and can achieve high coverage in Madagascar; however, context-specific strategies and an investment in dog vaccination as a public good will be required to move the country towards elimination.

Genetic population structure of endangered ring-tailed lemurs (Lemur catta) from nine sites in southern Madagascar.

Madagascar's ring-tailed lemurs (Lemur catta) are experiencing rapid population declines due to ongoing habitat loss and fragmentation, as well as increasing exploitation for bushmeat and the illegal pet trade. Despite being the focus of extensive and ongoing behavioral studies, there is comparatively little known about the genetic population structuring of the species. Here, we present the most comprehensive population genetic analysis of ring-tailed lemurs to date from across their likely remaining geographic range. We assessed levels of genetic diversity and population genetic structure using multilocus genotypes for 106 adult individuals from nine geographically representative localities. Population structure and F ST analyses revealed moderate genetic differentiation with localities being geographically partitioned into northern, southern, western and also potentially central clusters. Overall genetic diversity, in terms of allelic richness and observed heterozygosity, was high in the species (AR = 4.74, H O = 0.811). In fact, it is the highest among all published lemur estimates to date. While these results are encouraging, ring-tailed lemurs are currently affected by ongoing habitat fragmentation and occur at lower densities in poorer quality habitats. The effects of continued isolation and fragmentation, coupled with climate-driven environmental instability, will therefore likely impede the long-term viability of the species.

Daily lettuce supplements promote foraging behavior and modify the gut microbiota in captive frugivores.

For captive primates, greater provisioning of leafy greens or foliage can promote natural foraging behavior while boosting fiber intake. Recalcitrant fiber, although minimally available to endogenous metabolism, is readily fermented into nutrients by gut microbes. Whereas most primates in captivity consume fiber-limited diets and harbor imbalanced gut microbiota compared to their wild conspecifics, the importance of fiber provisioning to primate gut microbiota has predominately been studied in folivores. We, therefore, determined if commercial lettuce could be used to encourage foraging behavior and modify the gut microbiota of captive frugivores. We provisioned ruffed lemurs (Varecia rubra and V. variegata) with romaine lettuce, on top of the standard dietary fare, for 10 consecutive days. Before and across the period of lettuce supplementation, we collected observational data of animal feeding and fecal samples for microbiome analysis, determined via amplicon sequencing. The ruffed lemurs and their gut microbes responded to lettuce provisioning. In particular, younger animals readily ate lettuce and showed no decline in consumption across study days. When controlling for the effects of host species and social-group membership, lettuce consumption shifted the composition of the gut microbiome away from each lemur's own baseline, an effect that became stronger as the study progressed. In the final study days, Ruminococcaceae UCG-008 and Akkermansia, microbes typically and respectively associated with fiber metabolism and host health, were significantly enriched in the consortia of lettuce-provisioned subjects. Ultimately, the routine offering of lettuce, leafy greens, or foliage to captive frugivores may benefit animal wellbeing.

A national survey of household pet lemur ownership in Madagascar.

Primates are extracted from the wild for the pet trade across the world. In Madagascar, lemurs are kept as illegal pets and an understanding of lemur pet ownership at the national level is lacking. In 2013 and 2016, we undertook a national survey in 11 of Madagascar's 22 administrative regions (n = 28 towns) with 1,709 households. To our knowledge, this is the first national survey of the household ownership of pet primates in a country where they are endemic. In the 1.5 years prior to being surveyed, 8% ± 4% (towns as replicates) of respondents had seen a captive lemur while a further 0.7% ± 0.5% of respondents had owned one personally. We estimate that 33,428 ± 24,846 lemurs were kept in Malagasy households in the six months prior to our survey efforts, with 18,462 ± 12,963 of these pet lemurs estimated in urban household alone. Rates of lemur ownership did not differ by province but increased with the human population of a town and with the popularity of the town on Flickr (a proxy indicator for tourism). We found that the visibility of pet lemur ownership did not differ across the country, but it did increase with the size of the town and popularity with tourists. Areas with visible pet lemurs were not always the areas with the highest rates of pet lemur ownership, highlighting that many pet lemurs are hidden from the general public. Our study highlights the need for conservation programs to consider both the proportion of inhabitants that own pet lemurs and the total number of lemurs that are potentially being kept as pets in those towns. We close by noting that for some species, even just a small amount of localized live extraction for pet ownership could be enough to cause localized population extinctions over time. Moreover, an urgent response is needed to combat a recent and alarming rise in illegal exploitation of biodiversity across Madagascar.

Illegal Trade of Wild-Captured Lemur catta within Madagascar.

Lemur catta is the most reported illegal captive lemur. We document 286 L. catta that were held in illegal captive conditions in Madagascar. Coastal tourist destinations are "hot spots" for sightings. Many of the L. catta reported were in businesses (49%) and were perceived to be held captive for the purpose of generating income (41%). Infant/juvenile L. catta were overwhelmingly observed annually in December (41%) and may suffer high mortality rates given that they are not weaned during this month of the year. Population growth modeling suggests that known capture rates may be sustainable in all but small populations of 500 individuals and when infants/juveniles are targeted. However, of the seven remaining populations of L. catta with more than 100 individuals, only one is known to contain more than 500 animals, and we present evidence here that infants/juveniles are targeted. Moreover L. catta face significant other threats including habitat loss, bushmeat hunting, and climate change. Several actions could reduce the illegal capture and ownership of L. catta in Madagascar such as tourist behavior change initiatives, enforcement of laws, and alternative livelihoods for local people. These interventions are urgently needed and could be adapted to protect other exploited wildlife in the future.

A viral video and pet lemurs on Twitter.

Content shared on social media platforms can impact public perceptions of wildlife. These perceptions, which are in part shaped by context (e.g. non-naturalistic setting, presence of a human), can influence people's desires to interact with or acquire wild animals as pets. However, few studies have examined whether this holds true for wild animals featured in viral videos. This study reports on opportunistic data collected on Twitter before, during, and after a video that featured a habituated ring-tailed lemur (Lemur catta), called "Sefo", in southern Madagascar went 'viral' (i.e. circulated rapidly on the internet). Our dataset of 13,953 tweets (from an 18.5-week time period in early 2016) referencing lemurs was collected using targeted keywords on the Twitonomy Service. We identified 613 individual tweets about people wanting a lemur as a pet. In addition, 744 tweets that were captured in our dataset linked to the Sefo viral video. We found that as the number of tweets about the viral video increased, so did the number of tweets where an individual wanted to have a lemur as a pet. Most tweets (91%) did not make reference to a specific species of lemur, but when they did, they often (82%) referenced ring-tailed lemurs (L. catta), ruffed lemurs (Varecia spp.), and mouse lemurs (Microcebus spp.). This study serves as a case study to consider how viral content can impact how wild animals are perceived. We close by noting that social media sites like Twitter, which are increasingly providing their users with news and information, should carefully consider how information about wild animals is shared on their platforms, as it may impact animal welfare.

Exploring the Role of Wealth and Religion on the Ownership of Captive Lemurs in Madagascar Using Qualitative and Quantitative Data.

Primates are kept as pets for various reasons including as indicators of wealth. Ownership of primates can also be influenced by religion. In Madagascar, thousands of lemurs are kept as pets, but the roles of wealth and religion in the ownership of captive lemurs have not been explored. We use quantitative and qualitative data to examine these aspects of ownership. Quantitative data were collected (July to August 2016) in households (n = 596) of 12 urban and rural towns in Madagascar using semi-structured interviews. International standards for research ethics were followed. Research was approved by an ethics oversight committee. We also opportunistically visited 13 religious facilities. Qualitative data were used to frame the context of the quantitative data. We found that pet lemur owners do not speak about their lemurs as a symbol of wealth, but non-owners associate pet lemurs with wealth. Therefore, status/wealth may be a motivating factor in the ownership of pet lemurs. We also found evidence that Catholic entities in Madagascar sometimes take in captive lemurs when the owner can no longer care for the animal (being viewed as animal-friendly institutions). However, we did not find evidence of religion (institutional or traditional) influencing the ownership of pet lemurs.

Rapid Decrease in Populations of Wild Ring-Tailed Lemurs (Lemur catta) in Madagascar.

Lemurs are the most threatened group of mammals on earth. Lemur catta (ring-tailed lemur) represents one of the most iconic lemur species and faces numerous anthropogenic threats in the wild. In this study, we present population estimates from 32 sites across the range of L. catta, collected from primary and secondary data sources, to assess the number of ring-tailed lemurs left in the wild. We estimate that there are approximately 2,220 individual L. catta remaining in the 32 sites considered. We note local extinctions of populations of L. catta in at least 12 of the 32 sites examined, and that significantly more extinctions occurred in areas without some form of protection. This decrease in extant populations could represent a decrease of more than 95% of all ring-tailed lemurs in Madagascar since the year 2000. While these results should be considered preliminary, we stress the rapid decline of the species and note that habitat loss, bushmeat hunting and the illegal pet trade are driving populations to local extinction. Based on the data presented here, urgent and immediate funding and conservation action are crucial to ensure the viability of the remaining wild populations of ring-tailed lemurs.

Genetic Diversity of the Ring-Tailed Lemur (Lemur catta) in South-Central Madagascar.

Madagascar's lemurs, now deemed the most endangered group of mammals, represent the highest primate conservation priority in the world. Due to anthropogenic disturbances, an estimated 10% of Malagasy forest cover remains. The endangered Lemur catta is endemic to the southern regions of Madagascar and now occupies primarily fragmented forest habitats. We examined the influence of habitat fragmentation and isolation on the genetic diversity of L. catta across 3 different forest fragments in south-central Madagascar. Our analysis revealed moderate levels of genetic diversity. Genetic differentiation among the sites ranged from 0.05 to 0.11. These data suggest that the L. catta populations within south-central Madagascar have not yet lost significant genetic variation. However, due to ongoing anthropogenic threats faced by ring-tailed lemurs, continued conservation and research initiatives are imperative for long-term viability of the species.