Gut microbiome transmission in lizards

Complex communities of microorganisms living in the gastrointestinal tracts of animal hosts are known to play critical roles in a variety of processes affecting host fitness including physiology, immunity, behaviour and development. These gut microbes, often collectively referred to as the 'gut microbiome', are becoming better characterized across animal hosts as high-throughput, next-generation sequencing (NGS) becomes more affordable. Due to this wave of NGS studies, we now have a better understanding of how host ecology and evolution influence gut microbial community structure of animal hosts (Shapira 2016). Specifically, we now understand that universally the genomes of symbiotic microbes, together with the genomes of natural hosts, form the holobiont on which evolution acts (Rosenberg & Zilber-Rosenberg 2016). The majority of these commensal microbes are found in the gut, and host diet is one of the strongest influences on gut microbial community structure (Ley et al. 2008; Kohl & Carey 2016). However, excluding humans and other placental mammals that obtain many of their commensal microbes via vertical transmission during the birthing process, little is known about how animal hosts acquire their gut microbial communities. Moreover, large-scale studies of nonhuman gut microbiomes often rely on samples collected from captive individuals to characterize microbial diversity of the gut (e.g. Ley et al. 2008). While this caveat has been raised before (Amato 2013), recent studies clearly show that the effects of captivity call into question the utility of microbiome data from captive individuals in elucidating natural patterns (e.g. Clayton et al. 2016). However, with so little known about how other animals acquire their gut microbial communities or to what extent those communities are influenced by diet or environmental factors, it is unknown whether captivity will cause significant shifts in the gut microbial communities of other animals. Reptiles, the second most specious clade of amniotic vertebrates, have received relatively little investigation into the characterization or acquisition of their gut microbiota and only a handful of studies have used NGS technologies to characterize the gut microbiomes of reptiles (Colston & Jackson 2016). In a benchmark study in this issue of Molecular Ecology, Kohl et al. (2017) elegantly test not only the influence of diet and the environment on the gut bacterial community composition of lizards, but also, whether captivity influences shifts in gut bacterial community structure and whether viviparous lizards gain their gut microbiota via vertical transmission during birth. Gut microbial communities are likely comprised of a mix of transient, environmentally acquired microbes, commensal 'core' microbial taxa and a flexible pool of beneficial microbes which may confer a selective advantage to the host during periods of stress or rapid adaptation (reviewed in Shapira 2016). There are several hypothesized routes by which organisms might obtain their gut microbiota (Fig. 1). Specifically within lizards, both diet and coprophagy have been suggested as mechanisms of core gut microbiome acquisition. By comparing faecal samples of wild lizards over multiple seasons with available prey items and environmental samples (soil and plant material), Kohl et al. 2017 were able to test the influence of diet and the environment on gut bacterial community structure. In a comparison across three closely related host species in the family Liolaemidae, they found that neither host sex nor season had a significant effect on bacterial community membership or structure, but both diet (e.g. omnivorous vs. herbivorous host) and host species had a significant effect on both metrics. This is quite interesting as all three host species are syntopic; therefore, both omnivorous species should have the same potential prey availability, implying a role of host genetics on structuring the gut bacterial community. In a highly unexpected result, the authors found that the gut bacterial communities of the two omnivorous species overlapped with that of communities found on local plant material by 46%, while the single herbivorous species shared only 26% of its gut bacterial community with plant communities. Furthermore, the herbivorous species shared significantly more bacterial taxa with bacterial samples of potential invertebrate prey items. This is quite interesting and may be a result of both herbivorous lizards and local invertebrates utilizing the same gut microbiota to process local plant material. Bacteria found in local soil samples did not overlap significantly with bacterial taxa found in the gut of any host species. While it has been suggested that exposure to local microbial species underlies the variation in gut bacterial community composition, Kohl and colleagues have elegantly shown that this might be an oversimplification and it is unlikely that these lizards acquire commensal bacteria from soil. Captivity has been shown to alter microbial communities of a variety of vertebrate taxa, and gut bacterial community structure can influence reintroduction success in conservation programmes (Redford et al. 2012). Many squamate reptile species are considered threatened or in decline by the IUCN, and reptiles are considered particularly vulnerable to changing environmental conditions under projected climate change scenarios. To test the effects of captivity on the gut bacterial structure of lizards, Kohl et al. 2017 brought wild-caught individuals into the laboratory and maintained them for 8 weeks on artificial diets appropriate for omnivorous and herbivorous lizard species. At the end of the 8 weeks, the authors collected faecal samples as they had previously from lizards in the wild, as well as bacterial samples directly from the GI tract of euthanized individuals. Kohl and colleagues found that captivity significantly altered both bacterial community membership and structure in all three host species, with bacterial phyla and genera present being altered by 23% and 25%, respectively. Notably, they showed an invasion of the gut community by Enterobacter, Salmonella and Trabulsiella, all potentially pathogenic bacterial genera which were not detected in any wild-collected samples but were present in >75% of captive individuals after 8 weeks. However, it is important to note that while captivity altered the gut bacterial composition in all host species, the majority of gut bacteria from captive samples still overlapped with wild-collected samples, thus affirming that most individual lizards maintained their core gut microbial communities during the 8 weeks. The effects of captivity were shown to be decoupled from the artificial diet as <1% of bacteria found in laboratory meals were present in faecal samples. Additionally, bacterial communities sampled along the GI tract of euthanized individuals revealed that similar to other vertebrates lizards have distinct bacterial communities in the stomach, small intestine and hindgut with the hindgut communities bing most similar to faecal bacterial communities. This finding not only has implications for the distinct roles of these microbes in specific physiological processes of these gut regions, but also highlights the utility of using faecal samples as a proxy for hindgut bacterial communities. Lastly, samples were collected from gravid females prior to birth and from neonates post parturition; to test whether vertical transmission of microbes occurs in viviparous lizards (Fig. 1). Analyses of these samples revealed that a significant portion (34%) of faecal bacterial communities was shared between mothers and offspring. This is consistent with the hypothesis of vertical transmission via birth or common rearing. Interestingly, one bacterial genus, Trabulsiella, which was shared between mothers and young, had been shown earlier to be an introduction and result of captivity. However, juvenile lizards still had distinct bacterial communities to those of their mothers and these communities were less diverse. While viviparous lizards likely acquire gut microbiota via vertical transmission and communal rearing, these results suggest that the gut microbiome likely changes throughout host development and the core gut bacterial community may not stabilize until maturity. The (Kohl et al. 2017) study is admirable in its experimental design and framing of results in the context of the influence of gut microbiomes on the evolution of herbivory in lizards. While herbivory is rare in most reptile taxa, the authors have identified several bacteria taxa which were enriched in herbivorous host species and are involved in plant polysaccharide degradation and fibre fermentation. It is quite possible that adaptation to herbivory in lizards must be preceded or concurrent with the acquisition and utilization of beneficial microbes that can aid specific metabolic processes. As more studies seek to characterize the gut microbiome in a wide variety of animal hosts and investigate factors influencing microbiome composition, this study provides an excellent example upon which to model experimental design. T.J.C. wrote this perspective.