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Extensive research in well-studied animal models underscores the importance of commensal gastrointestinal (gut) microbes to animal physiology. Gut microbes have been shown to impact dietary digestion, mediate infection, and even modify behavior and cognition. Given the large physiological and pathophysiological contribution microbes provide their host, it is reasonable to assume that the vertebrate gut microbiome may also impact the fitness, health and ecology of wildlife. In accordance with this expectation, an increasing number of investigations have considered the role of the gut microbiome in wildlife ecology, health, and conservation. To help promote the development of this nascent field, we need to dissolve the technical barriers prohibitive to performing wildlife microbiome research. The present review discusses the 16S rRNA gene microbiome research landscape, clarifying best practices in microbiome data generation and analysis, with particular emphasis on unique situations that arise during wildlife investigations. Special consideration is given to topics relevant for microbiome wildlife research from sample collection to molecular techniques for data generation, to data analysis strategies. Our hope is that this article not only calls for greater integration of microbiome analyses into wildlife ecology and health studies but provides researchers with the technical framework needed to successfully conduct such investigations. 

Drinking water is one of numerous sources of human exposure to microscale and nanoscale plastic particles. Here, using a mouse model, we tested enteric and hepatic cellular responses to nanoplastic ingestion. At 1.5 or 25.5 h after an oral dose of 70 mg polystyrene nanospheres (PSNS)/kg (nominal diameters of 20 and 200 nm) in aqueous suspension female mice exhibit no overt signs of toxicity. Routine histopathology on small intestine and liver reveals no acute toxicity. Immunohistochemistry detects an increase in the number of enterocytes with cleaved caspase-3 (active form) after PSNS exposure ( p ≤ 0.05) indicating progression toward lytic cell death via a proinflammatory pathway. This is not evident in liver after PSNS exposure. Transmission electron microscopy detects lytic cell death in enterocytes at 25.5 h after 200 nm PSNS exposure. Putative endosomes in liver appear to sequester 20 and 200 nm particles 25.5 h after exposure. Both 20 and 200 nm PSNS appear in putative perinuclear autolysosomes 25.5 h after treatment. No significant changes in gene expression in the small intestine or liver 25.5 h were observed after dosing, but there was a trend toward altered expression of cyp1b1 in the liver. Analysis of the fecal microbiome shows loss of diversity after exposure to both 20 and 200 nm particles after 25.5 h. Taken together, these results suggest risk from ingestion of nanoscale plastic particles from drinking water, which deserves systematic investigation. 

Microbiomes are essential features of holobionts, providing their hosts with key metabolic and functional traits like resistance to environmental disturbances and diseases. In scleractinian corals, questions remain about the microbiome's role in resistance and resilience to factors contributing to the ongoing global coral decline and whether microbes serve as a form of holobiont ecological memory. To test if and how coral microbiomes affect host health outcomes during repeated disturbances, we conducted a large‐scale (32 exclosures, 200 colonies, and 3 coral species sampled) and long‐term (28 months, 2018–2020) manipulative experiment on the forereef of Mo'orea, French Polynesia. In 2019 and 2020, this reef experienced the two most severe marine heatwaves on record for the site. Our experiment and these events afforded us the opportunity to test microbiome dynamics and roles in the context of coral bleaching and mortality resulting from these successive and severe heatwaves. We report unique microbiome responses to repeated heatwaves inAcropora retusa,Porites lobata, andPocilloporaspp., which included: microbiome acclimatization inA. retusa, and both microbiome resilience to the first marine heatwave and microbiome resistance to the second marine heatwave inPocilloporaspp. Moreover, observed microbiome dynamics significantly correlated with coral species‐specific phenotypes. For example, bleaching and mortality inA. retusaboth significantly increased with greater microbiome beta dispersion and greater Shannon Diversity, whileP. lobatacolonies had different microbiomes across mortality prevalence. Compositional microbiome changes, such as changes to proportions of differentially abundant putatively beneficial to putatively detrimental taxa to coral health outcomes during repeated heat stress, also correlated with host mortality, with higher proportions of detrimental taxa yielding higher mortality inA. retusa. This study reveals evidence for coral species‐specific microbial responses to repeated heatwaves and, importantly, suggests that host‐dependent microbiome dynamics may provide a form of holobiont ecological memory to repeated heat stress.

 

ABSTRACT Shotgun metagenomic sequencing has transformed our understanding of microbial community ecology. However, preparing metagenomic libraries for high-throughput DNA sequencing remains a costly, labor-intensive, and time-consuming procedure, which in turn limits the utility of metagenomes. Several library preparation procedures have recently been developed to offset these costs, but it is unclear how these newer procedures compare to current standards in the field. In particular, it is not clear if all such procedures perform equally well across different types of microbial communities or if features of the biological samples being processed (e.g., DNA amount) impact the accuracy of the approach. To address these questions, we assessed how five different shotgun DNA sequence library preparation methods, including the commonly used Nextera Flex kit, perform when applied to metagenomic DNA. We measured each method’s ability to produce metagenomic data that accurately represent the underlying taxonomic and genetic diversity of the community. We performed these analyses across a range of microbial community types (e.g., soil, coral associated, and mouse gut associated) and input DNA amounts. We find that the type of community and amount of input DNA influence each method’s performance, indicating that careful consideration may be needed when selecting between methods, especially for low-complexity communities. However, the cost-effective preparation methods that we assessed are generally comparable to the current gold-standard Nextera DNA Flex kit for high-complexity communities. Overall, the results from this analysis will help expand and even facilitate access to metagenomic approaches in future studies. IMPORTANCE Metagenomic library preparation methods and sequencing technologies continue to advance rapidly, allowing researchers to characterize microbial communities in previously underexplored environmental samples and systems. However, widely accepted standardized library preparation methods can be cost-prohibitive. Newly available approaches may be less expensive, but their efficacy in comparison to standardized methods remains unknown. In this study, we compared five different metagenomic library preparation methods. We evaluated each method across a range of microbial communities varying in complexity and quantity of input DNA. Our findings demonstrate the importance of considering sample properties, including community type, composition, and DNA amount, when choosing the most appropriate metagenomic library preparation method. 

The intestinal nematodePseudocapillaria tomentosain zebrafish (Danio rerio) causes profound intestinal lesions, emaciation and death and is a promoter of a common intestinal cancer in zebrafish. This nematode has been detected in zebrafish from about 15% of the laboratories. Adult worms are readily detected about 3 weeks after exposure by either histology or wet mount preparations of the intestine, and larval worms are inconsistently observed in fish before this time. A quantitative PCR (qPCR) test was recently developed to detect the worm in fish and water, and here we determined that the test on zebrafish intestines was effective for earlier detection. Four lines of zebrafish (AB, TU, 5D and Casper) were experimentally infected and evaluated by wet mounts and qPCR at 8, 15‐, 22‐, 31‐ and 44‐day post‐exposure (dpe). At the first two time points, only 8% of the wet mounts from exposed fish were identified as infected, while the same intestines screened by qPCR showed 78% positivity, with low and consistent cycle threshold (Ct) values at these times. Wet mounts at later time points showed a high prevalence of infection, but this was still surpassed by qPCR.

 

ABSTRACT A growing body of research has established that the microbiome can mediate the dynamics and functional capacities of diverse biological systems. Yet, we understand little about what governs the response of these microbial communities to host or environmental changes. Most efforts to model microbiomes focus on defining the relationships between the microbiome, host, and environmental features within a specified study system and therefore fail to capture those that may be evident across multiple systems. In parallel with these developments in microbiome research, computer scientists have developed a variety of machine learning tools that can identify subtle, but informative, patterns from complex data. Here, we recommend using deep transfer learning to resolve microbiome patterns that transcend study systems. By leveraging diverse public data sets in an unsupervised way, such models can learn contextual relationships between features and build on those patterns to perform subsequent tasks (e.g., classification) within specific biological contexts. 

Microbes and sunlight convert terrigenous dissolved organic matter (DOM) in surface waters to greenhouse gases. Prior studies show contrasting results about how biological and photochemical processes interact to contribute to the degradation of DOM. In this study, DOM leached from the organic layer of tundra soil was exposed to natural sunlight or kept in the dark, incubated in the dark with the natural microbial community, and analyzed for gene expression and DOM chemical composition. Microbial gene expression (metatranscriptomics) in light and dark treatments diverged substantially after 4 hours. Gene expression suggested that sunlight exposure of DOM initially stimulated microbial growth by (a) replacing the function of enzymes that degrade higher molecular weight DOM such as enzymes for aromatic carbon degradation, oxygenation, and decarboxylation, and (b) releasing low molecular weight compounds and inorganic nutrients from DOM. However, growth stimulation following sunlight exposure of DOM came at a cost. Sunlight depleted the pool of aromatic compounds that supported microbial growth in the dark treatment, ultimately causing slower growth in the light treatment over 5 days. These first measurements of microbial metatranscriptomic responses to photo-alteration of DOM provide a mechanistic explanation for how sunlight exposure of terrigenous DOM alters microbial processing and respiration of DOM. 

ABSTRACT Plasmids harbor transferable genes that contribute to the functional repertoire of microbial communities, yet their contributions to metagenomes are often overlooked. Environmental plasmids have the potential to spread antibiotic resistance to clinical microbial strains. In soils, high microbiome diversity and high variability in plasmid characteristics present a challenge for studying plasmids. To improve the understanding of soil plasmids, we present RefSoil+, a database containing plasmid sequences from 922 soil microorganisms. Soil plasmids were larger than other described plasmids, which is a trait associated with plasmid mobility. There was a weak relationship between chromosome size and plasmid size and no relationship between chromosome size and plasmid number, suggesting that these genomic traits are independent in soil. We used RefSoil+ to inform the distributions of antibiotic resistance genes among soil microorganisms compared to those among nonsoil microorganisms. Soil-associated plasmids, but not chromosomes, had fewer antibiotic resistance genes than other microorganisms. These data suggest that soils may offer limited opportunity for plasmid-mediated transfer of described antibiotic resistance genes. RefSoil+ can serve as a reference for the diversity, composition, and host associations of plasmid-borne functional genes in soil, a utility that will be enhanced as the database expands. Our study improves the understanding of soil plasmids and provides a resource for assessing the dynamics of the genes that they carry, especially genes conferring antibiotic resistances. IMPORTANCE Soil-associated plasmids have the potential to transfer antibiotic resistance genes from environmental to clinical microbial strains, which is a public health concern. A specific resource is needed to aggregate the knowledge of soil plasmid characteristics so that the content, host associations, and dynamics of antibiotic resistance genes can be assessed and then tracked between the environment and the clinic. Here, we present RefSoil+, a database of soil-associated plasmids. RefSoil+ presents a contemporary snapshot of antibiotic resistance genes in soil that can serve as a reference as novel plasmids and transferred antibiotic resistances are discovered. Our study broadens our understanding of plasmids in soil and provides a community resource of important plasmid-associated genes, including antibiotic resistance genes. 

ABSTRACT Here, we report the draft genome sequence of Pseudomonas sp. strain DrBHI1. The total assembly length is 5,649,751 bp in 146 contigs. This strain was isolated from zebrafish ( Danio rerio ) feces. 

Aggression is a serious behavioral disorder in domestic dogs that endangers both dogs and humans. The underlying causes of canine aggression are poorly resolved and require illumination to ensure effective therapy. Recent research links the compositional diversity of the gut microbiome to behavioral and psychological regulation in other mammals, such as mice and humans. Given these observations, we hypothesized that the composition of the canine gut microbiome could associate with aggression. We analyzed fecal microbiome samples collected from a small population of pit bull type dogs seized from a dogfighting organization. This population included 21 dogs that displayed conspecific aggressive behaviors and 10 that did not. Beta-diversity analyses support an association between gut microbiome structure and dog aggression. Additionally, we used a phylogenetic approach to resolve specific clades of gut bacteria that stratify aggressive and non-aggressive dogs, including clades withinLactobacillus,Dorea,Blautia,Turicibacter,andBacteroides. Several of these taxa have been implicated in modulating mammalian behavior as well as gastrointestinal disease states. Although sample size limits this study, our findings indicate that gut microorganisms are linked to dog aggression and point to an aggression-associated physiological state that interacts with the gut microbiome. These results also indicate that the gut microbiome may be useful for diagnosing aggressive behaviors prior to their manifestation and potentially discerning cryptic etiologies of aggression.