Non-Lethal Detection of Frog Virus 3-Like (RUK13) and Common Midwife Toad Virus-Like (PDE18) Ranaviruses in Two UK-Native Amphibian Species.

Ranaviruses have been involved in amphibian mass mortality events worldwide. Effective screening to control this pathogen is essential; however, current sampling methods are unsuitable for the detection of subclinical infections. Non-lethal screening is needed to prevent both further spread of ranavirus and losses of at-risk species. To assess non-lethal sampling methods, we conducted two experiments: bath exposing common frogs to RUK13 ranavirus at three concentrations, and exposing common toads to RUK13 or PDE18. Non-lethal sampling included buccal, digit, body and tank swabs, along with toe clips and stool taken across three time-points post-exposure. The presence/load of ranavirus was examined using quantitative PCR in 11 different tissues obtained from the same euthanised animals (incl. liver, gastro-intestinal tract and kidney). Buccal swab screening had the highest virus detection rate in both species (62% frogs; 71% toads) and produced consistently high virus levels compared to other non-lethal assays. The buccal swab was effective across multiple stages of infection and differing infection intensities, though low levels of infection were more difficult to detect. Buccal swab assays competed with, and even outperformed, lethal sampling in frogs and toads, respectively. Successful virus detection in the absence of clinical signs was observed (33% frogs; 50% toads); we found no difference in detectability for RUK13 and PDE18. Our results suggest that buccal swabbing could replace lethal sampling for screening and be introduced as standard practice for ranavirus surveillance.

Heritage-specific oral microbiota in Indigenous Australian dental calculus.

Background and objectives: Aboriginal Australians and Torres Strait Islanders (hereafter respectfully referred to as Indigenous Australians) experience a high burden of chronic non-communicable diseases (NCDs). Increased NCD risk is linked to oral diseases mediated by the oral microbiota, a microbial community influenced by both vertical transmission and lifestyle factors. As an initial step towards understanding the oral microbiota as a factor in Indigenous health, we present the first investigation of oral microbiota in Indigenous Australian adults. Methodology: Dental calculus samples from Indigenous Australians with periodontal disease (PD; n = 13) and non-Indigenous individuals both with (n = 19) and without PD (n = 20) were characterized using 16S ribosomal RNA gene amplicon sequencing. Alpha and beta diversity, differentially abundant microbial taxa and taxa unique to different participant groups were analysed using QIIME2. Results: Samples from Indigenous Australians were more phylogenetically diverse (Kruskal-Wallis H = 19.86, P = 8.3 × 10-6), differed significantly in composition from non-Indigenous samples (PERMANOVA pseudo-F = 10.42, P = 0.001) and contained a relatively high proportion of unique taxa not previously reported in the human oral microbiota (e.g. Endomicrobia). These patterns were robust to stratification by PD status. Oral microbiota diversity and composition also differed between Indigenous individuals living in different geographic regions. Conclusions and implications: Indigenous Australians may harbour unique oral microbiota shaped by their long relationships with Country (ancestral homelands). Our findings have implications for understanding the origins of oral and systemic NCDs and for the inclusion of Indigenous peoples in microbiota research, highlighting the microbiota as a novel field of enquiry to improve Indigenous health.

Effectiveness of decontamination protocols when analyzing ancient DNA preserved in dental calculus.

Ancient DNA analysis of human oral microbial communities within calcified dental plaque (calculus) has revealed key insights into human health, paleodemography, and cultural behaviors. However, contamination imposes a major concern for paleomicrobiological samples due to their low endogenous DNA content and exposure to environmental sources, calling into question some published results. Decontamination protocols (e.g. an ethylenediaminetetraacetic acid (EDTA) pre-digestion or ultraviolet radiation (UV) and 5% sodium hypochlorite immersion treatments) aim to minimize the exogenous content of the outer surface of ancient calculus samples prior to DNA extraction. While these protocols are widely used, no one has systematically compared them in ancient dental calculus. Here, we compare untreated dental calculus samples to samples from the same site treated with four previously published decontamination protocols: a UV only treatment; a 5% sodium hypochlorite immersion treatment; a pre-digestion in EDTA treatment; and a combined UV irradiation and 5% sodium hypochlorite immersion treatment. We examine their efficacy in ancient oral microbiota recovery by applying 16S rRNA gene amplicon and shotgun sequencing, identifying ancient oral microbiota, as well as soil and skin contaminant species. Overall, the EDTA pre-digestion and a combined UV irradiation and 5% sodium hypochlorite immersion treatment were both effective at reducing the proportion of environmental taxa and increasing oral taxa in comparison to untreated samples. This research highlights the importance of using decontamination procedures during ancient DNA analysis of dental calculus to reduce contaminant DNA.

Post-epizootic microbiome associations across communities of neotropical amphibians.

Microbiome-pathogen interactions are increasingly recognized as an important element of host immunity. While these host-level interactions will have consequences for community disease dynamics, the factors which influence host microbiomes at larger scales are poorly understood. We here describe landscape-scale pathogen-microbiome associations within the context of post-epizootic amphibian chytridiomycosis, a disease caused by the panzootic chytrid fungus Batrachochytrium dendrobatidis. We undertook a survey of Neotropical amphibians across altitudinal gradients in Ecuador ~30 years following the observed amphibian declines and collected skin swab-samples which were metabarcoded using both fungal (ITS-2) and bacterial (r16S) amplicons. The data revealed marked variation in patterns of both B. dendrobatidis infection and microbiome structure that are associated with host life history. Stream breeding amphibians were most likely to be infected with B. dendrobatidis. This increased probability of infection was further associated with increased abundance and diversity of non-Batrachochytrium chytrid fungi in the skin and environmental microbiome. We also show that increased alpha diversity and the relative abundance of fungi are lower in the skin microbiome of adult stream amphibians compared to adult pond-breeding amphibians, an association not seen for bacteria. Finally, stream tadpoles exhibit lower proportions of predicted protective microbial taxa than pond tadpoles, suggesting reduced biotic resistance. Our analyses show that host breeding ecology strongly shapes pathogen-microbiome associations at a landscape scale, a trait that may influence resilience in the face of emerging infectious diseases.

Biocultural Drivers of Salivary Microbiota in Australian Aboriginal and Torres Strait Islander Children.

Australian Aboriginal and Torres Strait Islander children experience unacceptably high rates of dental caries compared to their non-Indigenous Australian counterparts. Dental caries significantly impacts the quality of life of children and their families, particularly in remote communities. While many socioeconomic and lifestyle factors impact caries risk, the central role of the oral microbiota in mediating dental caries has not been extensively investigated in these communities. Here, we examine factors that shape diversity and composition of the salivary microbiota in Aboriginal and Torres Strait Islander children and adolescents living in the remote Northern Peninsula Area (NPA) of Far North Queensland. We employed 16S ribosomal RNA amplicon sequencing to profile bacteria present in saliva collected from 205 individuals aged 4-17 years from the NPA. Higher average microbial diversity was generally linked to increased age and salivary pH, less frequent toothbrushing, and proxies for lower socioeconomic status (SES). Differences in microbial composition were significantly related to age, salivary pH, SES proxies, and active dental caries. Notably, a feature classified as Streptococcus sobrinus increased in abundance in children who reported less frequent tooth brushing. A specific Veillonella feature was associated with caries presence, while features classified as Actinobacillus/Haemophilus and Leptotrichia were associated with absence of caries; a Lactobacillus gasseri feature increased in abundance in severe caries. Finally, we statistically assessed the interplay between dental caries and caries risk factors in shaping the oral microbiota. These data provide a detailed understanding of biological, behavioral, and socioeconomic factors that shape the oral microbiota and may underpin caries development in this group. This information can be used in the future to improve tailored caries prevention and management options for Australian Aboriginal and Torres Strait Islander children and communities.

Cross-Disciplinary Genomics Approaches to Studying Emerging Fungal Infections.

Emerging fungal pathogens pose a serious, global and growing threat to food supply systems, wild ecosystems, and human health. However, historic chronic underinvestment in their research has resulted in a limited understanding of their epidemiology relative to bacterial and viral pathogens. Therefore, the untargeted nature of genomics and, more widely, -omics approaches is particularly attractive in addressing the threats posed by and illuminating the biology of these pathogens. Typically, research into plant, human and wildlife mycoses have been largely separated, with limited dialogue between disciplines. However, many serious mycoses facing the world today have common traits irrespective of host species, such as plastic genomes; wide host ranges; large population sizes and an ability to persist outside the host. These commonalities mean that -omics approaches that have been productively applied in one sphere and may also provide important insights in others, where these approaches may have historically been underutilised. In this review, we consider the advances made with genomics approaches in the fields of plant pathology, human medicine and wildlife health and the progress made in linking genomes to other -omics datatypes and sets; we identify the current barriers to linking -omics approaches and how these are being underutilised in each field; and we consider how and which -omics methodologies it is most crucial to build capacity for in the near future.

Response of Salivary Microbiota to Caries Preventive Treatment in Aboriginal and Torres Strait Islander Children.

A once-annual caries preventive (Intervention) treatment was offered to Aboriginal and Torres Strait Islander schoolchildren-a population with disproportionately poorer oral health than non-Indigenous Australian children-in the Northern Peninsula Area (NPA) of Far North Queensland (FNQ), which significantly improved their oral health. Here, we examine the salivary microbiota of these children (mean age = 10 ± 2.96 years; n = 103), reconstructing the bacterial community composition with high-throughput sequencing of the V4 region of bacterial 16S rRNA gene. Microbial communities of children who received the Intervention had lower taxonomic diversity than those who did not receive treatment (Shannon, p < 0.05). Moreover, the Intervention resulted in further decreased microbial diversity in children with active carious lesions existing at the time of saliva collection. Microbial species associated with caries were detected; Lactobacillus salivarius, Lactobacillus reuteri, Lactobacillus gasseri, Prevotella multisaccharivorax, Parascardovia denticolens, and Mitsuokella HMT 131 were significantly increased (p < 0.05) in children with severe caries, especially in children who did not receive the Intervention. These insights into microbial associations and community differences prompt future considerations to the mechanisms behind caries-preventive therapy induced change;  important for understanding  the long-term implications of like treatment to improve oral health disparities within Australia. Trial registration: ANZCTR, ACTRN12615000693527. Registered 3 July 2015, https://www.anzctr.org.au/Trial/Registration/TrialReview.aspx?id=368750&isReview=true.

Consequences of colonialism: A microbial perspective to contemporary Indigenous health.

Nearly all Indigenous populations today suffer from worse health than their non-Indigenous counterparts, and despite interventions against known factors, this health "gap" has not improved. The human microbiome-the beneficial, diverse microbial communities that live on and within the human body-is a crucial component in developing and maintaining normal physiological health. Disrupting this ecosystem has repercussions for microbial functionality, and thus, human health. In this article, we propose that modern-day Indigenous population health may suffer from disrupted microbial ecosystems as a consequence of historical colonialism. Colonialism may have interrupted the established relationships between the environment, traditional lifeways, and microbiomes, altering the Indigenous microbiome with detrimental health consequences.