The bacterial genus Endozoicomonas is prevalent and abundant in the microbiome of many corals, but the combination of environmental and host factors shaping its distribution remains uncertain. In this study, we used 16S rRNA gene sequencing to examine the distribution of Endozoicomonas across hard (Porites astreoides, Montastraea cavernosa, Agaricia spp.) and soft coral (Muricea spp., Gorgoniidae, Cladiellidae, Malacalcyonacea) microbiomes across a depth gradient from shallow (4.5 m) to upper mesophotic zones (36.6 m) at four sites around the Little Cayman Island, including two marine protected areas. Endozoicomonas was abundant across all coral microbiomes, with soft corals exhibiting near-total dominance (relative abundance range 0.1–99

Biodiversity loss threatens ecosystems worldwide, but we lack a thorough understanding of how genotypic diversity-as opposed to species diversity-impacts threatened foundation species, such as corals on tropical reefs. Corals of the genus Pocillopora have been important to natural reef recovery and human-assisted restoration in the Pacific. They consistently perform better in polycultures with other coral species than in monocultures, but whether genotypic diversity within a species enhances performance remains untested. We assessed whether increased genotypic diversity of the coral Pocillopora verrucosa (ie the number of genetically distinct source colonies per plot) improved growth, reduced tissue mortality, or increased resistance to invasion of competing seaweeds over 3.5 years in experimental plots on reefs in Mo'orea, French Polynesia. We could detect no significant differences in any of these performance metrics among plots comprised of one, three, six, or nine genotypes. Moreover, when we quantified mitochondrial genetic divergence among genotypes within each plot using mean pairwise nucleotide distance, performance was unrelated to the degree of genetic differentiation represented. Thus, we detected no effect of genotypic diversity on Pocillopora performance. Pocillopora acclimates rapidly to variable physical and biological conditions, which may make genotypic diversity effects less critical for these corals.

Sea cucumbers have been overharvested world-wide, making assessments of their ecological effects challenging, but recent research demonstrated that sea cucumbers increased coral survival via disease suppression and were therefore important for facilitating reef health. The mechanisms underpinning the sea cucumber-coral interaction are not well understood but are likely mediated through sea cucumber grazing of microbes from reef sediments. We explored how sea cucumber grazing alters the sediment microbiome by leveraging a healthy sea cucumber population on a reef in French Polynesia. We used quantitative PCR, 16S rRNA gene sequencing, and shotgun metagenomics to compare the sediment microbiome in cages placed in situ with or without sea cucumbers. We hypothesized that grazing would lower microbial biomass, change sediment microbiome composition, and deplete sediment metagenomes of anaerobic metabolisms, likely due to aeration of the sediments. Sea cucumber grazing resulted in a 75% reduction in 16S rRNA gene abundances and reshaped microbiome composition, causing a significant decrease of cyanobacteria and other phototrophs relative to ungrazed sediments. Grazing also resulted in a depletion of genes associated with cyanotoxin synthesis, suggesting a potential link to coral health. In contrast to expectations, grazed sediment metagenomes were enriched with marker genes of diverse anaerobic or microaerophilic metabolisms, including those encoding high oxygen affinity cytochrome oxidases. This enrichment differs from patterns linked to other bioturbating invertebrates. We hypothesize that grazing enriches anaerobic processes in sediment microbiomes through removal of oxygen-producing autotrophs, fecal deposition of sea cucumber gut-associated anaerobes, or modification of sediment diffusibility. These results suggest that sea cucumber harvesting influences biogeochemical processes in reef sediments, potentially mediating coral survival by altering the sediment microbiome and its production of coral-influencing metabolites.

The gastrointestinal (GI) microbiome plays a critical role in animal health and fitness, yet it remains understudied in many species—particularly those inhabiting freshwater environments affected by anthropogenic activity. This study investigates the gut microbiomes of two benthic fish species, Rocky Mountain Sculpin (Cottus bondi) and suckerfish (Catostomus spp.), collected upstream and downstream of a wastewater treatment plant (WWTP) effluent outflow into the East Gallatin River in southwestern Montana. Prokaryotic and eukaryotic microbiome diversity in the fish GI tract was assessed using 16S and 18S rRNA gene sequencing, respectively, from samples collected in the summer and fall of 2022 and 2023. While alpha diversity only had insignificant and small shifts across samples, beta diversity (taxonomic composition) differed significantly across sites and collection dates. Notably, the composition of eukaryotic sequences shifted markedly from upstream to downstream locations, suggesting that WWTP effluents may influence both prokaryotic and eukaryotic microbial communities. By establishing baseline GI microbiome characteristics for these species, this study provides important insights into the potential ecological effects of wastewater discharge on freshwater systems and supports conservation efforts aimed at mitigating pollutant impacts.

The marine invertebrate-associated microbiome has garnered significant interest in recent years due to its wealth of novel genes that can be explored for biomining. By combining genomics with untargeted data-dependent mass spectrometry (MS) and molecular networking, we characterized the secreted metabolome of Streptomyces sp. In a previous study, we isolated and characterized a strain of Streptomyces, designated as strain 34, from the nudibranch Chromodoris quadricolor, collected by SCUBA diving in the Red Sea near El Tor in the Gulf of Suez, Egypt. In the present study, the Streptomyces isolate was identified as Streptomyces tunisiensis GCF 039538125 1 (p-value: 0). Genomic and metabolomic analysis reveal 36 predicted biosynthetic gene clusters. A total of 569 metabolites were detected in the culture, with 86 of these being identified based on standards and public spectral libraries. Moreover, a single lassopeptide synthesis gene cluster was found in both the genome and the metabolic extract, along with various sets of siderophores identified in the metabolic extract. Since the metabolic processes of marine invertebrate microbiomes are poorly understood, our findings are a significant addition to the research on metabolism in host microbiomes.

Seaweed–coral competition is increasingly important as reef communities degrade, with algal turfs being the most common competitor. However, experiments assessing the impacts and mechanisms involved in turf–coral competition under field conditions are rare. We evaluated turf–coral interactions and their impacts relative to those of macroalgae by placing corals (Acropora pulchra and Porites rus) in contact with turf communities from territories of two species of damselfishes, with two common macroalgae, and with inert algal mimics as physical controls. After 13 d, turfs reduced coral photosynthesis by 31–59

Hundreds of studies now document positive relationships between biodiversity and critical ecosystem processes, but as ecological communities worldwide shift toward new species configurations, less is known regarding how the biodiversity of undesirable species will shape the functioning of ecosystems or foundation species. We manipulated macroalgal species richness in experimental field plots to test whether and how the identity and diversity of competing macroalgae affected the growth, survival, and microbiome of a common coral in Mo'orea, French Polynesia. Compared to controls without algal competitors, coral growth was significantly suppressed across three macroalgal monocultures, a polyculture of the same three macroalgae, and plots containing inert seaweed mimics; coral mortality was limited and did not differ significantly among treatments. One macroalga suppressed coral growth significantly less than the other two, but none differed from the inert mimic in terms of coral suppression. The composition, dispersion, and diversity of coral microbiomes in treatments with live macroalgae or inert plastic mimics did not differ from controls experiencing no competition. Microbiome composition differed between two macroalgal monocultures and a monoculture versus plastic mimics, but no other microbiome differences were observed among macroalgal or mimic treatments. Together, these findings suggest that algal diversity does not alter harmful impacts of macroalgae on coral performance, which could be accounted for by physical structure alone in these field experiments. While enhancing biodiversity is a recognized strategy for promoting desirable species, it would be worrisome if biodiversity also enhanced the negative impacts of undesirable species. We documented no such effects in this investigation.

Coral reefs are in global decline with coral diseases playing a significant role. This is especially true for Acroporid corals that represent ~25% of all Pacific coral species and generate much of the topographic complexity supporting reef biodiversity. Coral diseases are commonly sediment-associated and could be exacerbated by overharvest of sea cucumber detritivores that clean reef sediments and may suppress microbial pathogens as they feed. Here we show, via field manipulations in both French Polynesia and Palmyra Atoll, that historically overharvested sea cucumbers strongly suppress disease among corals in contact with benthic sediments. Sea cucumber removal increased tissue mortality of Acropora pulchra by ~370% and colony mortality by ~1500%. Additionally, farmerfish that kill Acropora pulchra bases to culture their algal gardens further suppress disease by separating corals from contact with the disease-causing sediment—functioning as mutualists rather than parasites despite killing coral bases. Historic overharvesting of sea cucumbers increases coral disease and threatens the persistence of tropical reefs. Enhancing sea cucumbers may enhance reef resilience by suppressing disease.

Animals under managed care in zoos and aquariums are ideal surrogate study subjects for endangered species that are difficult to obtain in the wild. We compared the fecal and oral microbiomes of healthy, managed African penguins ( Spheniscus demersus ) to those of other domestic and wild vertebrate hosts to determine how host identity, diet, and environment shape the penguin microbiome. The African penguin oral microbiome was more similar to that of piscivorous marine mammals, suggesting that diet and a marine environment together play a strong role in shaping the oral microbiome. Conversely, the penguin cloaca/fecal microbiome was more similar to that of other birds, suggesting that host phylogeny plays a significant role in shaping the gut microbiome. Although the penguins were born under managed care, they had a gut microbiome more similar to that of wild bird species compared to domesticated (factory-farmed) birds, suggesting that the managed care environment and diet resemble those experienced by wild birds. Finally, the microbiome composition at external body sites was broadly similar to that of the habitat, suggesting sharing of microbes between animals and their environment. Future studies should link these results to microbial functional capacity and host health, which will help inform conservation efforts.

Over the past decade, an abundance of 16S rRNA gene surveys have provided microbiologists with data regarding the prokaryotes present in a coral-associated microbial community. Functional gene studies that provide information regarding what those microbes might do are fewer, particularly for non-tropical corals. Using the GeoChip 5.0S microarray, we present a functional gene study of microbiomes from five species of cold-water corals collected from depths of 296–1567 m. These species included two octocorals, Acanthogorgia aspera and Acanthogorgia spissa , and three stony corals: Desmophyllum dianthus , Desmophyllum pertusum (formerly Lophelia pertusa ), and Enallopsammia profunda . A total of 24,281 gene sequences (representing different microbial taxa) encoding for 383 functional gene families and representing 9 metabolic gene categories were identified. Gene categories included metabolism of carbon, nitrogen, phosphorus, and sulfur, as well as virulence, organic remediation, metal homeostasis, secondary metabolism and phylogeny. We found that microbiomes from Acanthogorgia spp. were the most functionally distinct but also least diverse compared against those from stony corals. Desmophyllum spp. microbiomes were more similar to each other than to E. profunda . Of 383 total gene families detected in this study, less than 20% were significantly different among these deep-water coral species. Similarly, out of 59 metabolic sub-categories for which we were able to make a direct comparison to microbiomes of tropical corals, only 7 were notably different: anaerobic ammonium oxidation (anammox), chitin degradation, and dimethylsulfoniopropionate (DMSP) degradation, all of which had higher representations in deep-water corals; and chromium homeostasis/resistance, copper homeostasis/resistance, antibiotic resistance, and methanogenesis, all of which had higher representation in tropical corals. This implies a broad-scale convergence of the microbial functional genes present within the coral holobiont, independent of coral species, depth, symbiont status, and morphology.

Coastal waters are a major source of marine methane to the atmosphere. Particularly high concentrations of this potent greenhouse gas are found in anoxic waters, but it remains unclear if and to what extent anaerobic methanotrophs mitigate the methane flux. Here we investigate the long-term dynamics in methanotrophic activity and the methanotroph community in the coastal oxygen minimum zone (OMZ) of Golfo Dulce, Costa Rica, combining biogeochemical analyses, experimental incubations and 16S rRNA gene sequencing over 3 consecutive years. Our results demonstrate a stable redox zonation across the years with high concentrations of methane (up to 1.7 μmol L-1 ) in anoxic bottom waters. However, we also measured high activities of anaerobic methane oxidation in the OMZ core (rate constant, k, averaging 30 yr-1 in 2018 and 8 yr-1 in 2019-2020). The OPU3 and Deep Sea-1 clades of the Methylococcales were implicated as conveyors of the activity, peaking in relative abundance 5-25 m below the oxic-anoxic interface and in the deep anoxic water respectively. Although their genetic capacity for anaerobic methane oxidation remains unexplored, their sustained high relative abundance indicates an adaptation of these clades to the anoxic, methane-rich OMZ environment, allowing them to play major roles in mitigating methane fluxes.

Background Animal-associated microbiomes can be influenced by both host and environmental factors. Comparing wild animals to those in zoos or aquariums can help disentangle the effects of host versus environmental factors, while also testing whether managed conditions foster a ‘natural’ host microbiome. Focusing on an endangered elasmobranch species—the whitespotted eagle ray Aetobatus narinari —we compared the skin, gill, and cloaca microbiomes of wild individuals to those at Georgia Aquarium. Whitespotted eagle ray microbiomes from Georgia Aquarium were also compared to those of cownose rays ( Rhinoptera bonasus ) in the same exhibit, allowing us to explore the effect of host identity on the ray microbiome. Results Long-term veterinary monitoring indicated that the rays in managed care did not have a history of disease and maintained health parameters consistent with those of wild individuals, with one exception. Aquarium whitespotted eagle rays were regularly treated to control parasite loads, but the effects on animal health were subclinical. Microbiome α- and β-diversity differed between wild versus aquarium whitespotted eagle rays at all body sites, with α-diversity significantly higher in wild individuals. β-diversity differences in wild versus aquarium whitespotted eagle rays were greater for skin and gill microbiomes compared to those of the cloaca. At each body site, we also detected microbial taxa shared between wild and aquarium eagle rays. Additionally, the cloaca, skin, and gill microbiomes of aquarium eagle rays differed from those of cownose rays in the same exhibit. Potentially pathogenic bacteria were at low abundance in all wild and aquarium rays. Conclusion For whitespotted eagle rays, managed care was associated with a microbiome differing significantly from that of wild individuals. These differences were not absolute, as the microbiome of aquarium rays shared members with that of wild counterparts and was distinct from that of a cohabitating ray species. Eagle rays under managed care appear healthy, suggesting that their microbiomes are not associated with compromised host health. However, the ray microbiome is dynamic, differing with both environmental factors and host identity. Monitoring of aquarium ray microbiomes over time may identify taxonomic patterns that co-vary with host health.

BACKGROUND:Sharks play essential roles in ocean food webs and human culture, but also face population declines worldwide due to human activity. The relationship between sharks and the microbes on and in the shark body is unclear, despite research on other animals showing the microbiome as intertwined with host physiology, immunity, and ecology. Research on shark-microbe interactions faces the significant challenge of sampling the largest and most elusive shark species. We leveraged a unique sampling infrastructure to compare the microbiomes of two apex predators, the white (Carcharodon carcharias) and tiger shark (Galeocerdo cuvier), to those of the filter-feeding whale shark (Rhincodon typus), allowing us to explore the effects of feeding mode on intestinal microbiome diversity and metabolic function, and environmental exposure on the diversity of microbes external to the body (on the skin, gill).RESULTS:The fecal microbiomes of white and whale sharks were highly similar in taxonomic and gene category composition despite differences in host feeding mode and diet. Fecal microbiomes from these species were also taxon-poor compared to those of many other vertebrates and were more similar to those of predatory teleost fishes and toothed whales than to those of filter-feeding baleen whales. In contrast, microbiomes of external body niches were taxon-rich and significantly influenced by diversity in the water column microbiome.CONCLUSIONS:These results suggest complex roles for host identity, diet, and environmental exposure in structuring the shark microbiome and identify a small, but conserved, number of intestinal microbial taxa as potential contributors to shark physiology.

Data collected during annual sampling campaigns to the coastal oxygen minimum zone of Golfo Dulce, carried out in January-February 2018, 2019 and 2020. Methods and results are presented and discussed in Steinsdóttir et al. 2022. Anaerobic methane oxidation in a coastal oxygen minimum zone: spatial and temporal dynamics. Environmental Microbiology, in press, doi: 10.1111/1462-2920.16003 The content of files is as follows: nutrient_and_methane_concentrations.csv - Concentrations of methane, nitrite, nitrate, and ammonium. methane_oxidation_rates.csv - Rates of anaerobic methane oxidation. kinetics_of_anaerobic_methane_oxidation.csv - Kinetics of anaerobic methane oxidation, carried out in 2019. methylococcales.fa - Methylococcales 16S rRNA amplicon sequences methanofastidiosa.fa - Methanofastidiosa 16S rRNA amplicon sequences

All animals are host to a multitude of microorganisms that are essential to the animal’s health. Host-associated microbes have been shown to defend against potential pathogens, provide essential nutrients, interact with the host’s immune system, and even regulate mood. However, it can be difficult to preserve and obtain nucleic acids from some host-associated microbiomes, making studying their microbial communities challenging. Corals are an example of this, in part due to their potentially remote, underwater locations, their thick surface mucopolysaccharide layer, and various inherent molecular inhibitors. This study examined three different preservatives (RNAlater, DNA/RNA Shield, and liquid nitrogen) and two extraction methods (the Qiagen PowerBiofilm kit and the Promega Maxwell RBC kit with modifications) to determine if there was an optimum combination for examining the coral microbiome. These methods were employed across taxonomically diverse coral species, including deep-sea/shallow, stony/soft, and zooxanthellate/azooxanthellate: Lophelia pertusa, Paragorgia johnsoni, Montastraea cavernosa, Porites astreoides, and Stephanocoenia intersepta. Although significant differences were found between preservative types and extraction methods, these differences were subtle, and varied in nature from coral species to coral species. Significant differences between coral species were far more profound than those detected between preservative or extraction method. We suggest that the preservative types presented here and extraction methods using a bead-beating step provide enough consistency to compare coral microbiomes across various studies, as long as subtle differences in microbial communities are attributed to dissimilar methodologies. Additionally, the inclusion of internal controls such as a mock community and extraction blanks can help provide context regarding data quality, improving downstream analyses.

Abstract Background In adults undergoing allogeneic hematopoietic cell transplantation (HCT), higher gut microbiome diversity is associated with reduced bloodstream infections (BSI) and improved overall survival (OS). Rifaximin prophylaxis in adult HCT helps to maintain microbiome diversity. We examine changes in microbiome in a cohort of pediatric patients undergoing HCT. Methods Patients were enrolled in an institutional biorepository (n=82) with a subset enrolled in an ongoing trial using rifaximin (n=21) between 2013–2020. All patients had HCT for a hematologic malignancy, using myeloablative conditioning. Patients in the rifaximin trial received rifaximin starting 7 days before HCT (D-7) through D+28, otherwise, no prophylactic antibiotics were used. Systemic antibiotic timing was categorized as none, early (≤ Day 0, day of HCT), and late (> D0). We performed 16s rRNA sequencing from stool for 73 subjects, at baseline (D-7), and weekly through D+28 (engraftment). Microbiome diversity was assessed by Shannon index. Results Median age was 9 years (range 1–20), 59% male, 41% Caucasian and 29% Black. There were no differences in BSI or mortality by age, sex, or race. Microbiome diversity changed significantly over time (p=0.008). Drop in diversity was most notable in patients who had early antibiotics (Mean=1.4, CI -0.15, 2.94, p=0.077). Higher diversity was seen when patients received none or late versus early antibiotics, but this was not statistically significant (Figure 1, p=0.23). Piperacillin-tazobactam was used empirically in 91% of patients. OS at 1 year was 88.5% (CI 68.4%, 96.1%) for patients with high (≥ median) D+28 diversity compared to 60% (CI 38.4%, 76.1%) for patients with low diversity (Figure 2, p=0.018) Only 1 of 21 (4.8%) in the rifaximin group developed a BSI with a gut bacterium compared to 8 of 61 (13.1%) not on rifaximin within the first 30 days (trial enrollment ongoing). Figure 1. Effect of systemic antibiotic timing on microbiome diversity over time. Figure 2. One-year overall survival of patients with high (>2.77) versus low (<2.77) diversity defined by median Shannon-Index. Conclusion We have shown a significant correlation between engraftment microbiome diversity and 1-year OS. Early antibiotic exposure was detrimental to microbiome diversity. Approaches to preserve microbiome diversity and prevent BSI are likely to improve HCT outcomes. Our ongoing trial using rifaximin will provide preliminary data regarding this approach. Disclosures All Authors: No reported disclosures

Elasmobranchs (sharks, skates and rays) are of broad ecological, economic, and societal value. These globally important fishes are experiencing sharp population declines as a result of human activity in the oceans. Research to understand elasmobranch ecology and conservation is critical and has now begun to explore the role of body-associated microbiomes in shaping elasmobranch health. Here, we review the burgeoning efforts to understand elasmobranch microbiomes, highlighting microbiome variation among gastrointestinal, oral, skin, and blood-associated niches. We identify major bacterial lineages in the microbiome, challenges to the field, key unanswered questions, and avenues for future work. We argue for prioritizing research to determine how microbiomes interact mechanistically with the unique physiology of elasmobranchs, potentially identifying roles in host immunity, disease, nutrition, and waste processing. Understanding elasmobranch–microbiome interactions is critical for predicting how sharks and rays respond to a changing ocean and for managing healthy populations in managed care.

The deep ocean hosts a large diversity of azooxanthellate cold-water corals whose associated microbiomes remain to be described. While the bacterial genus Endozoicomonas has been widely identified as a dominant associate of tropical and temperate corals, it has rarely been detected in deep-sea corals. Determining microbial baselines for these cold-water corals is a critical first step to understanding the ecosystem services their microbiomes contribute, while providing a benchmark against which to measure responses to environmental change or anthropogenic effects. Samples of Acanthogorgia aspera , A. spissa , Desmophyllum dianthus , and D. pertusum ( Lophelia pertusa ) were collected from western Atlantic sites off the US east coast and from the northeastern Gulf of Mexico. Microbiomes were characterized by 16S rRNA gene amplicon surveys. Although D. dianthus and D. pertusum have recently been combined into a single genus due to their genetic similarity, their microbiomes were significantly different. The Acanthogorgia spp. were collected from submarine canyons in different regions, but their microbiomes were extremely similar and dominated by Endozoicomonas . This is the first report of coral microbiomes dominated by Endozoicomonas occurring below 1000 m, at temperatures near 4°C. D. pertusum from 2 Atlantic sites were also dominated by distinct Endozoicomonas , unlike D. pertusum from other sites described in previous studies, including the Gulf of Mexico, the Mediterranean Sea and a Norwegian fjord.

The microbiome of the temperate coral Astrangia poculata was first described in 2017 using next-generation Illumina sequencing to examine the coral’s bacterial and archaeal associates across seasons and among hosts of differing symbiotic status. To assess the impact of methodology on the detectable diversity of the coral’s microbiome, we obtained near full-length Sanger sequences from clone libraries constructed from a subset of the same A. poculata samples. Eight samples were analyzed: two sets of paired symbiotic (brown) and aposymbiotic (white) colonies collected in the fall (September) and two sets collected in the spring (April). Analysis of the Sanger sequences revealed that the microbiome of A. poculata exhibited a high level of richness; 806 OTUs were identified among 1390 bacterial sequences. While the Illumina study revealed that A. poculata’s microbial communities did not significantly vary according to symbiotic state, but did vary by season, Sanger sequencing did not expose seasonal or symbiotic differences in the microbiomes. Proteobacteria dominated the microbiome, forming the majority (55% to 80%) of classifiable bacteria in every sample, and the five bacterial classes with the highest mean relative portion (5% to 35%) were the same as those determined by prior Illumina sequencing. Sanger sequencing also captured the same core taxa previously identified by next-generation sequencing. Alignment of all sequences and construction of a phylogenetic tree revealed that both sequencing methods provided similar portrayals of the phylogenetic diversity within A. poculata’s bacterial associates. Consistent with previous findings, the results demonstrated that the Astrangia microbiome is stable notwithstanding the choice of sequencing method and the far fewer sequences generated by clone libraries (46 to 326 sequences per sample) compared to next-generation sequencing (3634 to 48481 sequences per sample). Moreover, the near-full length 16S rRNA sequences produced by this study are presented as a resource for the community studying this model system since they provide necessary information for designing primers and probes to further our understanding of this coral’s microbiome.