Beyond the theory: From holobiont concept to microbiome engineering.

Holobiont research has increasingly moved from descriptive studies to sophisticated field- and laboratory-based manipulations; however, the extent to which changes in the holobiont persist remains largely unknown. In this Burning Question, we ask whether the underlying principles of the holobiont concept, whereby an externally applied evolutionary pressure can lead to a beneficial change in host-associated microbial community composition, could be used to facilitate microbiome engineering and thereby addition of a new ecosystem service that persists across generations. The answer to this question has potential implications for diverse fields including symbiosis, conservation and biotechnology.

16S rRNA gene-based meta-analysis of the reptile gut microbiota reveals environmental effects, host influences and a limited core microbiota.

An animal's gut microbiota plays an important role in host health, reproduction and digestion. However, many studies focus on only a few individuals or a single species, limiting our ability to recognize emergent patterns across a wider taxonomic grouping. Here, we compiled and reanalysed published 16S rRNA gene sequence data for 745 gut microbiota samples from 91 reptile species using a uniform bioinformatics pipeline to draw broader conclusions about the taxonomy of the reptile gut microbiota and the forces shaping it. Our meta-analysis revealed the significant differences in alpha- and beta-diversity across host order, environment, diet, habitat and conservation status, with host diet and order contributing the most to these differences. We identified the principal bacterial phyla present in the reptile gut microbiota as Bacteroidota, Proteobacteria (mostly Gamma class), and Firmicutes, and detected the bacterial genus Bacteroides in most reptile individuals, thus representing a putative 'core' microbiota. Our study provides novel insights into key drivers of the reptile gut microbiota, highlights existing knowledge gaps and lays the groundwork for future research on these fascinating hosts and their associated microbes.

Problem based review: the pregnant woman presenting to the AMU with palpitations.

Pregnant patients commonly present to the acute medical team with symptoms requiring further investigation. Palpitations are a common reason for presentation on the acute medical take, and most acute physicians will be familiar with the process of investigation. The combination of pregnancy and palpitations raises a broad differential diagnosis and can complicate the management pathway. This problem based review is designed to summarise the key issues which may arise during the management of a typical patient presenting in this way.

Gut microbiome of the sole surviving member of reptile order Rhynchocephalia reveals biogeographic variation, influence of host body condition and a substantial core microbiota in tuatara across New Zealand.

Tuatara are the sole extant species in the reptile order Rhynchocephalia. They are ecologically and evolutionarily unique, having been isolated geographically for ~84 million years and evolutionarily from their closest living relatives for ~250 million years. Here we report the tuatara gut bacterial community for the first time. We sampled the gut microbiota of translocated tuatara at five sanctuaries spanning a latitudinal range of ~1000 km within Aotearoa New Zealand, as well as individuals from the source population on Takapourewa (Stephens Island). This represents a first look at the bacterial community of the order Rhynchocephalia and provides the opportunity to address several key hypotheses, namely that the tuatara gut microbiota: (1) differs from those of other reptile orders; (2) varies among geographic locations but is more similar at sites with more similar temperatures and (3) is shaped by tuatara body condition, parasitism and ambient temperature. We found significant drivers of the microbiota in sampling site, tuatara body condition, parasitism and ambient temperature, suggesting the importance of these factors when considering tuatara conservation. We also derived a 'core' community of shared bacteria across tuatara at many sites, despite their geographic range and isolation. Remarkably, >70% of amplicon sequence variants could not be assigned to known genera, suggesting a largely undescribed gut bacterial community for this ancient host species.

Gut microbiota predicts body fat change following a low-energy diet: a PREVIEW intervention study.

BACKGROUND: Low-energy diets (LEDs) comprise commercially formulated food products that provide between 800 and 1200 kcal/day (3.3-5 MJ/day) to aid body weight loss. Recent small-scale studies suggest that LEDs are associated with marked changes in the gut microbiota that may modify the effect of the LED on host metabolism and weight loss. We investigated how the gut microbiota changed during 8 weeks of total meal replacement LED and determined their associations with host response in a sub-analysis of 211 overweight adults with pre-diabetes participating in the large multicentre PREVIEW (PREVention of diabetes through lifestyle intervention and population studies In Europe and around the World) clinical trial. METHODS: Microbial community composition was analysed by Illumina sequencing of the hypervariable V3-V4 regions of the 16S ribosomal RNA (rRNA) gene. Butyrate production capacity was estimated by qPCR targeting the butyryl-CoA:acetate CoA-transferase gene. Bioinformatics and statistical analyses, such as comparison of alpha and beta diversity measures, correlative and differential abundances analysis, were undertaken on the 16S rRNA gene sequences of 211 paired (pre- and post-LED) samples as well as their integration with the clinical, biomedical and dietary datasets for predictive modelling. RESULTS: The overall composition of the gut microbiota changed markedly and consistently from pre- to post-LED (P = 0.001), along with increased richness and diversity (both P < 0.001). Following the intervention, the relative abundance of several genera previously associated with metabolic improvements (e.g., Akkermansia and Christensenellaceae R-7 group) was significantly increased (P < 0.001), while flagellated Pseudobutyrivibrio, acetogenic Blautia and Bifidobacterium spp. were decreased (all P < 0.001). Butyrate production capacity was reduced (P < 0.001). The changes in microbiota composition and predicted functions were significantly associated with body weight loss (P < 0.05). Baseline gut microbiota features were able to explain ~25% of variation in total body fat change (post-pre-LED). CONCLUSIONS: The gut microbiota and individual taxa were significantly influenced by the LED intervention and correlated with changes in total body fat and body weight in individuals with overweight and pre-diabetes. Despite inter-individual variation, the baseline gut microbiota was a strong predictor of total body fat change during the energy restriction period. TRIAL REGISTRATION: The PREVIEW trial was prospectively registered at ClinicalTrials.gov ( NCT01777893 ) on January 29, 2013.

Integrity of the Human Faecal Microbiota following Long-Term Sample Storage.

In studies of the human microbiome, faecal samples are frequently used as a non-invasive proxy for the study of the intestinal microbiota. To obtain reliable insights, the need for bacterial DNA of high quality and integrity following appropriate faecal sample collection and preservation steps is paramount. In a study of dietary mineral balance in the context of type 2 diabetes (T2D), faecal samples were collected from healthy and T2D individuals throughout a 13-day residential trial. These samples were freeze-dried, then stored mostly at -20°C from the trial date in 2000/2001 until the current research in 2014. Given the relative antiquity of these samples (~14 years), we sought to evaluate DNA quality and comparability to freshly collected human faecal samples. Following the extraction of bacterial DNA, gel electrophoresis indicated that our DNA extracts were more sheared than extracts made from freshly collected faecal samples, but still of sufficiently high molecular weight to support amplicon-based studies. Likewise, spectrophotometric assessment of extracts revealed that they were of high quality and quantity. A subset of bacterial 16S rRNA gene amplicons were sequenced using Illumina MiSeq and compared against publicly available sequence data representing a similar cohort analysed by the American Gut Project (AGP). Notably, our bacterial community profiles were highly consistent with those from the AGP data. Our results suggest that when faecal specimens are stored appropriately, the microbial profiles are preserved and robust to extended storage periods.

Presence of antibodies to Salmonella in tuatara (Sphenodon punctatus) sera.

Colonisation of a host by pathogenic microorganisms is a near constant threat to the health of all vertebrates and most species have evolved an efficient adaptive immune response which produces antibodies following exposure to a specific antigen. The strength of this response can be influenced by many factors including sex and season. Tuatara are exposed to Salmonella through contact with infected skinks and soil; however, no gastrointestinal colonisation of tuatara with Salmonella has been found. Using Western blot and flow cytometry we have demonstrated that tuatara possess antibodies which recognise Salmonella antigens, but many of these antibodies are not specific and are cross-reactive with two closely related and ubiquitous bacteria, Escherichia coli and Citrobacter koseri. Our study describes the anti-Salmonella immune responses in tuatara and will help to inform decisions around maintaining wildlife health, as well as providing important insights into the role and development of adaptive immunity in reptilian species.

Reptile reservoirs and seasonal variation in the environmental presence of Salmonella in an island ecosystem, Stephens Island, New Zealand.

A wide variety of Salmonella serotypes occurs within reptilian hosts, but their ecology is poorly understood. We collected cloacal swabs from tuatara (Sphenodon punctatus), fairy prions (Pachyptila turtur), and skinks (Oligosoma spp.) on Stephens Island, New Zealand, to screen for Salmonella. Soil samples were also collected from inside burrows of tuatara and fairy prions and tested for Salmonella. We sampled repeatedly from October 2009 to October 2011. Cloacal swabs were collected from 620 tuatara, and no intestinal shedding of Salmonella was detected. Similarly, no Salmonella was detected in fairy prions. In contrast, we isolated Salmonella from 6.5% of skinks and 8.4% of soil samples. We identified two serovars of Salmonella from 52 isolates, Salmonella Saintpaul and Salmonella Mississippi. Salmonella Mississippi was isolated from skinks only and S. Saintpaul was found in skinks and soil samples. Salmonella persists in this ecosystem with skinks as the main wildlife reservoir, and an environmental reservoir exists in the soil from burrows used by skinks, tuatara, and fairy prions. Salmonella was absent from skinks and the soil in winter, raising the question of bacteria persisting through winter.