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1. Evaluating Methods of Preserving Aquatic Invertebrates for Microbiome Analysis

   https://doi.org/10.3390/microorganisms10040811  

   Vaughn, Stephanie N. ; Jackson, Colin R. ( April 2022 , Microorganisms)

   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 microbiomemore »analysis but is less reliable for the external microbiome.« less
2. Community structure of coral microbiomes is dependent on host morphology

   https://doi.org/10.1186/s40168-022-01308-w  

   Morrow, Kathleen M. ; Pankey, M. Sabrina ; Lesser, Michael P. ( December 2022 , Microbiome)

   Abstract Background The importance of symbiosis has long been recognized on coral reefs, where the photosynthetic dinoflagellates of corals (Symbiodiniaceae) are the primary symbiont. Numerous studies have now shown that a diverse assemblage of prokaryotes also make-up part of the microbiome of corals. A subset of these prokaryotes is capable of fixing nitrogen, known as diazotrophs, and is also present in the microbiome of scleractinian corals where they have been shown to supplement the holobiont nitrogen budget. Here, an analysis of the microbiomes of 16 coral species collected from Australia, Curaçao, and Hawai’i using three different marker genes (16S rRNA, nif H, and ITS2) is presented. These data were used to examine the effects of biogeography, coral traits, and ecological life history characteristics on the composition and diversity of the microbiome in corals and their diazotrophic communities. Results The prokaryotic microbiome community composition (i.e., beta diversity) based on the 16S rRNA gene varied between sites and ecological life history characteristics, but coral morphology was the most significant factor affecting the microbiome of the corals studied. For 15 of the corals studied, only two species Pocillopora acuta and Seriotopora hystrix , both brooders, showed a weak relationship between the 16S rRNAmore »gene community structure and the diazotrophic members of the microbiome using the nif H marker gene, suggesting that many corals support a microbiome with diazotrophic capabilities. The order Rhizobiales , a taxon that contains primarily diazotrophs, are common members of the coral microbiome and were eight times greater in relative abundances in Hawai’i compared to corals from either Curacao or Australia. However, for the diazotrophic component of the coral microbiome, only host species significantly influenced the composition and diversity of the community. Conclusions The roles and interactions between members of the coral holobiont are still not well understood, especially critical functions provided by the coral microbiome (e.g., nitrogen fixation), and the variation of these functions across species. The findings presented here show the significant effect of morphology, a coral “super trait,” on the overall community structure of the microbiome in corals and that there is a strong association of the diazotrophic community within the microbiome of corals. However, the underlying coral traits linking the effects of host species on diazotrophic communities remain unknown.« less
3. Mink ( Neovison vison ) fecal microbiomes are influenced by sex, temperature, and time postdefecation

   https://doi.org/10.1093/jmammal/gyab140  

   Lafferty, Diana J. R. ; Gillman, Sierra J. ; Jeakle, Lane K. ; Roell, Brian J. ; McKenney, Erin A. ; Hayes, ed., Loren ( January 2022 , Journal of Mammalogy)

   Gut microbiomes encode myriad metabolic functions critical to mammalian ecology and evolution. While fresh fecal samples provide an efficient, noninvasive method of sampling gut microbiomes, collecting fresh feces from elusive species is logistically challenging. Nonfresh feces, however, may not accurately represent the gut microbiome of the host due to succession of gut microbial consortia postdefecation as well as colonization by microbes from the surrounding environment. Using American mink (Neovison vison) as a model species, we examined postdefecation microbial community succession to learn how ambient temperature and temporal sampling constraints influence the reliability of nonfresh feces to represent host gut microbiomes. To achieve our goal, we analyzed fresh mink feces (n = 5 females; n = 5 males) collected at the time of defecation from captive mink at a farm in the Upper Peninsula of Michigan and we subsequently subsampled each fecal specimen to investigate microbial community succession over five days, under both warm (21°C) and cold (–17°C to –1°C) temperature treatments. We found that both temperature and time influenced fecal microbiome composition; and we also detected significant sexual dimorphism in microbial community structures, with female mink microbiomes exhibiting significantly greater variation than males’ when exposed to the warm temperaturemore »treatment. Our results demonstrate that feces from unknown individuals can be a powerful tool for examining carnivore gut microbiomes, though rigorous study design is required because sex, ambient temperature, and time since defecation drive significant microbial variation and the sample size requirements necessary for detecting statistically significant differences between target populations is an important consideration for future ecologically meaningful research.

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4. Impacts of the Marine Hatchery Built Environment, Water and Feed on Mucosal Microbiome Colonization Across Ontogeny in Yellowtail Kingfish, Seriola lalandi

   https://doi.org/10.3389/fmars.2021.676731  

   Minich, Jeremiah J. ; Nowak, Barbara ; Elizur, Abigail ; Knight, Rob ; Fielder, Stewart ; Allen, Eric E. ( May 2021 , Frontiers in Marine Science)

   The fish gut microbiome is impacted by a number of biological and environmental factors including fish feed formulations. Unlike mammals, vertical microbiome transmission is largely absent in fish and thus little is known about how the gut microbiome is initially colonized during hatchery rearing nor the stability throughout growout stages. Here we investigate how various microbial-rich surfaces from the built environment “BE” and feed influence the development of the mucosal microbiome (gill, skin, and digesta) of an economically important marine fish, yellowtail kingfish, Seriola lalandi , over time. For the first experiment, we sampled gill and skin microbiomes from 36 fish reared in three tank conditions, and demonstrate that the gill is more influenced by the surrounding environment than the skin. In a second experiment, fish mucous (gill, skin, and digesta), the BE (tank side, water, inlet pipe, airstones, and air diffusers) and feed were sampled from indoor reared fish at three ages (43, 137, and 430 dph; n = 12 per age). At 430 dph, 20 additional fish were sampled from an outdoor ocean net pen. A total of 304 samples were processed for 16S rRNA gene sequencing. Gill and skin alpha diversity increased while gut diversity decreased withmore »age. Diversity was much lower in fish from the ocean net pen compared to indoor fish. The gill and skin are most influenced by the BE early in development, with aeration equipment having more impact in later ages, while the gut “allochthonous” microbiome becomes increasingly differentiated from the environment over time. Feed had a relatively low impact on driving microbial communities. Our findings suggest that S. lalandi mucosal microbiomes are differentially influenced by the BE with a high turnover and rapid succession occurring in the gill and skin while the gut microbiome is more stable. We demonstrate how individual components of a hatchery system, especially aeration equipment, may contribute directly to microbiome development in a marine fish. In addition, results demonstrate how early life (larval) exposure to biofouling in the rearing environment may influence fish microbiome development which is important for animal health and aquaculture production.« less
5. Microbiomes of the Sydney Rock Oyster are acquired through both vertical and horizontal transmission

   https://doi.org/10.1186/s42523-022-00186-9  

   Unzueta-Martínez, Andrea ; Scanes, Elliot ; Parker, Laura M. ; Ross, Pauline M. ; O’Connor, Wayne ; Bowen, Jennifer L. ( May 2022 , Animal Microbiome)

   The term holobiont is widely accepted to describe animal hosts and their associated microorganisms. The genomes of all that the holobiont encompasses, are termed the hologenome and it has been proposed as a unit of selection in evolution. To demonstrate that natural selection acts on the hologenome, a significant portion of the associated microbial genomes should be transferred between generations. Using the Sydney Rock Oyster (Saccostrea glomerata) as a model, we tested if the microbes of this broadcast spawning species could be passed down to the next generation by conducting single parent crosses and tracking the microbiome from parent to offspring and throughout early larval stages using 16S rRNA gene amplicon sequencing. From each cross, we sampled adult tissues (mantle, gill, stomach, gonad, eggs or sperm), larvae (D-veliger, umbo, eyed pediveliger, and spat), and the surrounding environment (water and algae feed) for microbial community analysis.

   We found that each larval stage has a distinct microbiome that is partially influenced by their parental microbiome, particularly the maternal egg microbiome. We also demonstrate the presence of core microbes that are consistent across all families, persist throughout early life stages (from eggs to spat), and are not detected in the microbiomes ofmore »the surrounding environment. In addition to the core microbiomes that span all life cycle stages, there is also evidence of environmentally acquired microbial communities, with earlier larval stages (D-veliger and umbo), more influenced by seawater microbiomes, and later larval stages (eyed pediveliger and spat) dominated by microbial members that are specific to oysters and not detected in the surrounding environment.

   Our study characterized the succession of oyster larvae microbiomes from gametes to spat and tracked selected members that persisted across multiple life stages. Overall our findings suggest that both horizontal and vertical transmission routes are possible for the complex microbial communities associated with a broadcast spawning marine invertebrate. We demonstrate that not all members of oyster-associated microbiomes are governed by the same ecological dynamics, which is critical for determining what constitutes a hologenome.

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