Effects of life stage, site, and species on the dragonfly gut microbiome

© 2020 by the authors. Licensee MDPI, Basel, Switzerland. Insects that undergo metamorphosis from juveniles to adults provide an intriguing opportunity to examine the effects of life stage, species, and the environment on their gut microbiome. In this study, we surveyed the gut microbiomes of 13 species of dragonfly collected from five different locations subject to different levels of human impact. Juveniles were collected as nymphs from aquatic habitats while airborne adults were caught at the same locations. The gut microbiome was characterized by next generation sequencing of the bacterial 16S rRNA gene. Life stage was an important factor, with the gut microbiomes of dragonfly nymphs differing from those of adult dragonflies. Gut microbiomes of nymphs were influenced by sample site and, to a lesser extent, host species. Neither sample location nor host species had a strong effect on the gut microbiome of dragonfly adults. Regardless of life stage, gut microbiomes were dominated by members of the Proteobacteria, with members of the Bacteroidetes (especially in adults), Firmicutes, and Acidobacteria (especially in nymphs) also being proportionally abundant. These results demonstrate that different life stages of metamorphosing insects can harbor very different gut microbiomes and differ in how this microbiome is influenced by the surrounding environment

Evaluating Methods of Preserving Aquatic Invertebrates for Microbiome Analysis

Research on the microbiomes of animals has increased substantially within the past decades. More recently, microbial analyses of aquatic invertebrates have become of increased interest. The storage method used while collecting aquatic invertebrates has not been standardized throughout the scientific community, and the effects of common storage methods on the microbial composition of the organism is unknown. Using crayfish and dragonfly nymphs collected from a natural pond and crayfish maintained in an aquarium, the effects of two common storage methods, preserving in 95% ethanol and freezing at −20◦C, on the invertebrate bacterial microbiome was evaluated. We found that the bacterial community was conserved for two sample types (gut and exoskeleton) of field-collected crayfish stored either in ethanol or frozen, as was the gut microbiome of aquarium crayfish. However, there were significant differences between the bacterial communities found on the exoskeleton of aquarium crayfish stored in ethanol compared to those that were frozen. Dragonfly nymphs showed significant differences in gut microbial composition between species, but the microbiome was conserved between storage methods. These results demonstrate that preserving field-collected specimens of aquatic invertebrates in 95% ethanol is likely to be a simple and effective sample preservation method for subsequent gut microbiome analysis but is less reliable for the external microbiome

Phylogenetic analysis of bacterial and archaeal arsC gene sequences suggests an ancient, common origin for arsenate reductase

BACKGROUND: The ars gene system provides arsenic resistance for a variety of microorganisms and can be chromosomal or plasmid-borne. The arsC gene, which codes for an arsenate reductase is essential for arsenate resistance and transforms arsenate into arsenite, which is extruded from the cell. A survey of GenBank shows that arsC appears to be phylogenetically widespread both in organisms with known arsenic resistance and those organisms that have been sequenced as part of whole genome projects. RESULTS: Phylogenetic analysis of aligned arsC sequences shows broad similarities to the established 16S rRNA phylogeny, with separation of bacterial, archaeal, and subsequently eukaryotic arsC genes. However, inconsistencies between arsC and 16S rRNA are apparent for some taxa. Cyanobacteria and some of the γ-Proteobacteria appear to possess arsC genes that are similar to those of Low GC Gram-positive Bacteria, and other isolated taxa possess arsC genes that would not be expected based on known evolutionary relationships. There is no clear separation of plasmid-borne and chromosomal arsC genes, although a number of the Enterobacteriales (γ-Proteobacteria) possess similar plasmid-encoded arsC sequences. CONCLUSION: The overall phylogeny of the arsenate reductases suggests a single, early origin of the arsC gene and subsequent sequence divergence to give the distinct arsC classes that exist today. Discrepancies between 16S rRNA and arsC phylogenies support the role of horizontal gene transfer (HGT) in the evolution of arsenate reductases, with a number of instances of HGT early in bacterial arsC evolution. Plasmid-borne arsC genes are not monophyletic suggesting multiple cases of chromosomal-plasmid exchange and subsequent HGT. Overall, arsC phylogeny is complex and is likely the result of a number of evolutionary mechanisms

Microbiomes of Caribbean Octocorals Vary Over Time but Are Resistant to Environmental Change

The bacterial microbiome is an essential component of many corals, although knowledge of the microbiomes in scleractinian corals far exceeds that for octocorals. This study characterized the bacterial communities present in shallow water Caribbean gorgonian octocorals over time and space, in addition to determining the bacterial assemblages in gorgonians exposed to environmental perturbations. We found that seven shallow water Caribbean gorgonian species maintained distinct microbiomes and predominantly harbored two bacterial genera, Mycoplasma and Endozoicomonas. Representatives of these taxa accounted for over 70% of the sequences recovered, made up the three most common operational taxonomic units (OTUs), and were present in most of the gorgonian species. Gorgonian species sampled in different seasons and/or in different years, exhibited significant shifts in the abundances of these bacterial OTUs, though there were few changes to overall bacterial diversity, or to the specific OTUs present. These shifts had minimal impact on the relative abundance of inferred functional proteins within the gorgonian corals. Sequences identified as Escherichia were ubiquitous in gorgonian colonies sampled from a lagoon but not in colonies sampled from a back reef. Exposure to increased temperature and/or ultraviolet radiation (UVR) or nutrient enrichment led to few significant changes in the gorgonian coral microbiomes. While there were some shifts in the abundance of the prevalent bacteria, more commonly observed was “microbial switching” between different OTUs identified within the same bacterial genus. The relative stability of gorgonian coral bacterial microbiome may potentially explain some of the resistance and resilience of Caribbean gorgonian corals against changing environmental conditions

The gut microbiome of freshwater Unionidae mussels is determined by host species and is selectively retained from filtered seston

© 2019 Weingarten et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Freshwater mussels are a species-rich group of aquatic invertebrates that are among the most endangered groups of fauna worldwide. As filter-feeders that are constantly exposed to new microbial inoculants, mussels represent an ideal system to investigate the effects of species or the environment on gut microbiome composition. In this study, we examined if host species or site exerts a greater influence on microbiome composition. Individuals of four co-occurring freshwater mussel species, Cyclonaias asperata, Fusconaia cerina, Lampsilis ornata, and Obovaria unicolor were collected from six sites along a 50 km stretch of the Sipsey River in Alabama, USA. High throughput 16S rRNA gene sequencing revealed that mussel gut bacterial microbiota were distinct from bacteria on seston suspended in the water column, and that the composition of the gut microbiota was influenced by both host species and site. Despite species and environmental variation, the most frequently detected sequences within the mussel microbiota were identified as members of the Clostridiales. Sequences identified as the nitrogen-fixing taxon Methylocystis sp. were also abundant in all mussel species, and sequences of both bacterial taxa were more abundant in mussels than in water. Site physicochemical conditions explained almost 45% of variation in seston bacterial communities but less than 8% of variation in the mussel bacterial microbiome. Together, these findings suggest selective retention of bacterial taxa by the freshwater mussel host, and that both species and the environment are important in determining mussel gut microbiome composition

Ranking the biases: The choice of OTUs vs. ASVs in 16S rRNA amplicon data analysis has stronger effects on diversity measures than rarefaction and OTU identity threshold

Advances in the analysis of amplicon sequence datasets have introduced a methodological shift in how research teams investigate microbial biodiversity, away from sequence identity-based clustering (producing Operational Taxonomic Units, OTUs) to denoising methods (producing amplicon sequence variants, ASVs). While denoising methods have several inherent properties that make them desirable compared to clustering-based methods, questions remain as to the influence that these pipelines have on the ecological patterns being assessed, especially when compared to other methodological choices made when processing data (e.g. rarefaction) and computing diversity indices. We compared the respective influences of two widely used methods, namely DADA2 (a denoising method) vs. Mothur (a clustering method) on 16S rRNA gene amplicon datasets (hypervariable region v4), and compared such effects to the rarefaction of the community table and OTU identity threshold (97% vs. 99%) on the ecological signals detected. We used a dataset comprising freshwater invertebrate (three Unionidae species) gut and environmental (sediment, seston) communities sampled in six rivers in the southeastern USA. We ranked the respective effects of each methodological choice on alpha and beta diversity, and taxonomic composition. The choice of the pipeline significantly influenced alpha and beta diversities and changed the ecological signal detected, especially on presence/absence indices such as the richness index and unweighted Unifrac. Interestingly, the discrepancy between OTU and ASV-based diversity metrics could be attenuated by the use of rarefaction. The identification of major classes and genera also revealed significant discrepancies across pipelines. Compared to the pipeline’s effect, OTU threshold and rarefaction had a minimal impact on all measurements

The saltpan microbiome is structured by sediment depth and minimally influenced by variable hydration

© 2020 by the authors. Licensee MDPI, Basel, Switzerland. Saltpans are a class of ephemeral wetland characterized by alternating periods of inundation, rising salinity, and desiccation. We obtained soil cores from a saltpan on the Mississippi Gulf coast in both the inundated and desiccated state. The microbiomes of surface and 30 cm deep sediment were determined using Illumina sequencing of the V4 region of the 16S rRNA gene. Bacterial and archaeal community composition differed significantly between sediment depths but did not differ between inundated and desiccated states. Well-represented taxa included marine microorganisms as well as multiple halophiles, both observed in greater proportions in surface sediment. Functional inference of metagenomic data showed that saltpan sediments in the inundated state had greater potential for microbial activity and that several energetic and degradation pathways were more prevalent in saltpan sediment than in nearby tidal marsh sediment. Microbial communities within saltpan sediments differed in composition from those in adjacent freshwater and brackish marshes. These findings indicate that the bacterial and archaeal microbiomes of saltpans are highly stratified by sediment depth and are only minimally influenced by changes in hydration. The surface sediment community is likely isolated from the shallow subsurface community by compaction, with the microbial community dominated by marine and terrestrial halophiles

Gut microbiomes of freshwater mussels (Unionidae) are taxonomically and phylogenetically variable across years but remain functionally stable

Freshwater mussels perform essential ecosystem functions, yet we have no information on how their microbiomes fluctuate over time. In this study, we examined temporal variation in the microbiome of six mussel species (Lampsilis ornata, Obovaria unicolor, Elliptio arca, Fusconaia cerina, Cyclonaias asperata, and Tritogonia verrucosa) sampled from the same river in 2016 and 2019. We examined the taxonomic, phylogenetic, and inferred functional (from 16S rRNA sequences) facets of their microbiome diversity. Significant differences between the two years were identified in five of the six species sampled. However, not all species that exhibited a temporally variable microbiome were functionally distinct across years, indicating functional redundancy within the mussel gut mi-crobiome. Inferred biosynthesis pathways showed temporal variation in pathways involved in deg-radation, while pathways involved in cellular metabolism were stable. There was no evidence for phylosymbiosis across any facet of microbiome biodiversity. These results indicate that temporal variation is an important factor in the assembly of the gut microbiomes of freshwater mussels and provides further support that the mussel gut microbiome is involved in host development and ac-tivity

Unintended and accidental medical radiation exposures in radiology: guidelines on investigation and prevention

This paper sets out guidelines for managing radiation exposure incidents involving patients in diagnostic and interventional radiology. The work is based on collation of experiences from representatives of international and national organizations for radiologists, medical physicists, radiographers, regulators, and equipment manufacturers, derived from an International Atomic Energy Agency Technical Meeting. More serious overexposures can result in skin doses high enough to produce tissue reactions, in interventional procedures and computed tomography, most notably from perfusion studies. A major factor involved has been deficiencies in training of staff in operation of equipment and optimization techniques. The use of checklists and time outs before procedures commence, and dose alerts when critical levels are reached during procedures can provide safeguards to reduce risks of these effects occurring. However, unintended and accidental overexposures resulting in relatively small additional doses can take place in any diagnostic or interventional X-ray procedure and it is important to learn from errors that occur, as these may lead to increased risks of stochastic effects. Such events may involve the wrong examinations, procedural errors, or equipment faults. Guidance is given on prevention, investigation and dose calculation for radiology exposure incidents within healthcare facilities. Responsibilities should be clearly set out in formal policies, and procedures should be in place to ensure that root causes are identified and deficiencies addressed. When an overexposure of a patient or an unintended exposure of a foetus occurs, the foetal, organ, skin and/or effective dose may be estimated from exposure data. When doses are very low, generic values for the examination may be sufficient, but a full assessment of doses to all exposed organs and tissues may sometimes be required. The use of general terminology to describe risks from stochastic effects is recommended rather than calculation of numerical values, as these are misleading when applied to individuals