Biology subjects

Gomez-Chiarri, M.

Publications and source records attributed to Gomez-Chiarri, M..

7 recordsLinked to original sources

bioRxiv2026

Phylogenetically distinct Vibrio mediterranei lineages confer robust protection under thermal stress against oyster pathogens

Marine bivalve mortality events cause substantial economic losses in aquaculture and threaten global food security. While pathogenic Vibrio species are frequently implicated, growing evidence suggests that loss of beneficial microbes can increase host susceptibility to disease. We previously observed that Vibrio mediterranei was consistently isolated from healthy oysters but systematically disappeared prior to mortality events, coinciding with proliferation of pathogenic Vibrio species. Here, we test whether this pattern reflects a protective functional role. Pre-colonization with V. mediterranei strain Vm02 increased Crassostrea virginica oyster larval survival from 10-19% (pathogen-only controls) to 94-97% when challenged with V. harveyi or V. coralliilyticus, representing near-complete protection from pathogen-induced mortality. Protection was maintained at both ambient (28{degrees}C) and thermal stress (32{degrees}C) temperatures where pathogen virulence is enhanced, rapid (effective from co-inoculation), and durable (maintained for >96 hours). Fluorescence microscopy confirmed stable colonization of larval digestive tissues by fluorescently-tagged Vm02. Larval colonization by eleven V. mediterranei strains revealed three distinct phenotypes - protective, pathogenic, and intermediate - corresponding to monophyletic clades with 97.1-97.8% average nucleotide identity between protective and pathogenic lineages. Pangenome analysis identified 230 protective-specific versus 80 pathogenic-specific orthogroups. Protective strains encode unique regulatory systems, stress tolerance mechanisms, and metabolic versatility while lacking Type I and Type VI secretion system variants associated with pathogenicity. Together, these findings demonstrate that beneficial versus pathogenic phenotypes are phylogenetically constrained within distinct V. mediterranei lineages. This supports reports that V. mediterranei acts as both a pathogen and potential symbiont in marine hosts and reveals a clade that provides robust protection against oyster pathogens. ImportanceAquaculture disease management has traditionally emphasized either prophylactic treatment using antibiotics to avoid disease and dysbiosis or has focused entirely on pathogen detection. Both of these approaches have overlooked the potential contributions of beneficial microbes to host defense and grow-out performance. Developing beneficial probiotic tools for disease prevention represents an emerging opportunity for sustainable aquaculture management. This study demonstrates that specific lineages of Vibrio mediterranei function as protective symbionts capable of rescuing oyster larvae from near-complete pathogen-induced mortality. By integrating field observations of microbial succession during mortality events with experimental validation and comparative genomics, we show that protective versus pathogenic phenotypes are phylogenetically constrained within V. mediterranei clades separated by 97.1-97.8% average nucleotide identity. This resolution of strain-level functional variation provides fundamental insights into how host-microbe mutualisms evolve within species complexes that also harbor pathogens. The unique genomic markers identified here enable reliable screening for protective symbionts, while the temperature-stable and durable protection demonstrated in this study highlights the potential for biological control strategies in shellfish hatcheries increasingly affected by warming oceans and Vibrio-driven mortality events.

microbiology↗

bioRxiv2025

Hemocyte Viability Assay as an Alternative Method for Testing Bacterial Pathogenicity in Bivalves

Bacterial pathogens cause disease outbreaks in hatcheries, leading to larval mortality and significant economic losses. One of the major challenges in studying bacterial pathogenesis in larvae is the limited availability of healthy larvae throughout the year. This seasonal constraint delays testing and impedes progress in understanding bacterial infections, disease management, and the development of effective treatments for bivalve aquaculture. To address this challenge, a hemocyte viability assay was developed as an alternative for testing bacterial pathogenesis. A resazurin-based assay was optimized using eastern oyster hemocytes, and lethal concentration (LC)50 values were determined for the bacterial pathogen Vibrio coralliilyticus RE22 (121 {+/-} 2 multiplicity of infection, MOI), probiont Phaeobacter inhibens S4 (2,157 {+/-} 56 MOI), and several bacterial isolates from shellfish hatcheries. The most virulent isolates had LC50 values ranging from 53 to 195 MOI. To validate the assay, the isolates were tested on oyster larvae, confirming that bacterial isolates with LC50 <200 MOI against hemocytes also caused larval mortality, while isolates exhibiting LC50 values [≥]690 MOI showed no adverse effects on oyster larvae. This high-throughput hemocyte viability assay provides a rapid and reliable alternative to larval assays, particularly during the off-season for hatcheries, facilitating year-round research on bacterial pathogenicity in bivalves.

immunology↗

bioRxiv2022

Development and evaluation of a formulation of probiont Phaeobacter inhibens S4 for themanagement of vibriosis in bivalve hatcheries

Larval eastern oysters (Crassostrea virginica) grown in shellfish hatcheries are susceptible to bacterial diseases, particularly vibriosis. Probiotics are microbes that confer health benefits to the host and have been identified as promising tools to manage diseases in aquaculture. The marine bacterium Phaeobacter inhibens S4 (S4) protects larval eastern oysters against challenge with the bacterial pathogen Vibrio coralliilyticus RE22 (RE22). A concentrated liquid formulation of probiont S4 that maintained high cell viability after long-term storage was developed for commercial use in shellfish hatcheries. The safety and efficacy of the formulation was tested in six different trials in two hatcheries. The S4 formulation was added to C. virginica larvae culture tanks daily at 104 colony forming units (CFU)/mL from day 1 post fertilization until day 6, 12, or 14, depending on the trial. Treatment of larvae in the hatchery with the S4 formulation did not significantly affect the survival and growth of the larvae. Formulated probiont S4 treatment in the hatchery led to a significant increase in Relative Percent Survival (RPS) when larvae were subsequently challenged with the pathogen RE22 (105 CFU/mL) for 24 hours in a laboratory challenge, as compared to probiotic-untreated RE22-challenged larvae (Relative Percent Survival increase of 46 - 74%, p < 0.05). These results suggest that this novel S4 formulation is a safe, easy to use, and effective tool in preventing larval losses due to vibriosis in hatcheries.

microbiology↗

bioRxiv2022

Nucleotide and structural polymorphisms of the eastern oyster genome paint a mosaic of divergence, selection, and human impacts

The eastern oyster, Crassostrea virginica, is a valuable fishery and aquaculture species that provides critical services as an ecosystem engineer. Oysters have a life-history that promotes high genetic diversity and gene flow while also occupying a wide range of habitats in variable coastal environments from the southern Gulf of Mexico to the southern waters of Atlantic Canada. To understand the interplay of genetic diversity, gene flow, and intense environmental selection, we used whole genome re-sequencing data from 90 individuals across the eastern United States and Gulf of Mexico, plus 5 selectively bred lines. Our data confirmed a large phylogeographic break between oyster populations in the Gulf of Mexico and the Atlantic coast of the USA. We also demonstrated that domestication has artificially admixed genetic material between the two ocean basins, and selected lines with admixed ancestry continue to maintain heterozygosity at these sites through several generations post admixture, possibly indicating relevance to desirable aquaculture traits. We found that genetic and structural variation are high in both wild and selected populations, but we also demonstrated that, when controlling for domestication admixture across ocean basins, wild populations do have significantly higher levels of nucleotide diversity and copy number variation than selected lines. Within the Atlantic coast, we detected subtle but distinct population structure, introgression of selected lines within wild individuals, an interaction between structural variation and putatively adaptive population structure, and evidence of candidate genes responding to selection from salinity. Our study highlights the potential for applying whole genome sequencing to highly polymorphic species and provides a road map for future work examining the genome variation of eastern oyster populations.

evolutionary biology↗

bioRxiv2022

A second unveiling: haplotig masking of the eastern oyster genome improves population-level inference

Genome assembly can be challenging for species that are characterized by high amounts of polymorphism, heterozygosity, and large effective population sizes. High levels of heterozygosity can result in genome mis-assemblies and a larger than expected genome size due to the haplotig versions of a single locus being assembled as separate loci. Here, we describe the first chromosome-level genome for the eastern oyster, Crassostrea virginica. Publicly released and annotated in 2017, the assembly has a scaffold N50 of 54 mb and is over 97.3% complete based on BUSCO analysis. The genome assembly for the eastern oyster is a critical resource for foundational research into molluscan adaptation to a changing environment and for selective breeding for the aquaculture industry. Subsequent resequencing data suggested the presence of haplotigs in the original assembly, and we developed a post hoc method to break up chimeric contigs and mask haplotigs in published heterozygous genomes and evaluated improvements to the accuracy of downstream analysis. Masking haplotigs had a large impact on SNP discovery and estimates of nucleotide diversity and had more subtle and nuanced effects on estimates of heterozygosity, population structure analysis, and outlier detection. We show that haplotig-masking can be a powerful tool for improving genomic inference, and we present an open, reproducible resource for the masking of haplotigs in any published genome.

genomics↗

bioRxiv2021

Two Type VI Secretion Systems in Vibrio coralliilyticus RE22Sm exhibit differential target specificity for bacteria prey and oyster larvae

Vibrio coralliilyticus is an extracellular bacterial pathogen and a causative agent of vibriosis in larval oysters. Host mortality rates can quickly reach 100% during vibriosis outbreaks in oyster hatcheries. Type VI Secretion Systems (T6SS) are rapidly polymerizing, contact dependent injection apparatus for prey cell intoxication and play important roles in pathogenesis. DNA sequencing of V. coralliilyticus RE22Sm indicated the likely presence of two functional T6SSs with one on each of two chromosomes. Here, we investigated the antibacterial and anti-eukaryotic roles of the two T6SSs (T6SS1 and T6SS2) against E. coli Sm10 cells and Crassostrea virginica larvae, respectively. Mutations in hcp and vgrG genes were created and characterized for their effects upon bacterial antagonism and eukaryotic host virulence. Mutations in hcp1 and hcp2 resulted in significantly reduced antagonism against E. coli Sm10, with the hcp2 mutation demonstrating the greater impact. In contrast, mutations in vgrG1 or vgrG2 had little effect on E. coli killing. In eastern oyster larval challenge assays, T6SS1 mutations in either hcp1 or vgrG1 dramatically attenuated virulence against C. virginica larvae. Strains with restored wild type hcp or vgrG genes reestablished T6SS-mediated killing to that of wild type V. coralliilyticus RE22Sm. These data suggest that the T6SS1 of V. coralliilyticus RE22Sm principally targets eukaryotes and secondarily bacteria, while the T6SS2 primarily targets bacterial cells and secondarily eukaryotes. Attenuation of pathogenicity was observed in all T6SS mutants, demonstrating the requirement for proper assembly of the T6SS systems to maintain maximal virulence. ImportanceVibriosis outbreaks lead to large-scale hatchery losses of oyster larvae (product and seed) where Vibrio sp. associated losses of 80 to 100 percent are not uncommon. Practical and proactive biocontrol measures can be taken to help mitigate larval death by Vibrio sp. by better understanding the underlying mechanisms of virulence in V. coralliilyticus. In this study, we demonstrate the presence of two Type VI Secretion Systems (T6SS) in V. coralliilyticus RE22Sm and interrogate the roles of each T6SS in bacterial antagonism and pathogenesis against a eukaryotic host. Specifically, we show that the loss of T6SS1 function results in the loss of virulence against oyster larvae.

microbiology↗

bioRxiv2020

Diversity and Function of the Eastern Oyster (Crassostrea virginica) Microbiome

Marine invertebrate microbiomes play important roles in various host and ecological processes. However, a mechanistic understanding of host-microbe interactions is so far only available for a handful of model organisms. Here, an integrated taxonomic and functional analysis of the microbiome of the eastern oyster, Crassostrea virginica, was performed using 16S rRNA gene amplicon profiling, shotgun metagenomics, and genome-scale metabolic reconstruction. A relatively low number of amplicon sequence variants (ASVs) were observed in oyster tissues compared to water samples, while high variability was observed across individual oysters and among different tissue types. Targeted metagenomic sequencing of the gut microbiota led to further characterization of a dominant bacterial taxon, the class Mollicutes, which was captured by the reconstruction of a metagenome-assembled genome (MAG). Genome-scale metabolic reconstruction of the oyster Mollicutes MAG revealed a reduced set of metabolic functions and a high reliance on the uptake of host-derived nutrients. A chitin degradation and an arginine deiminase pathway were unique to the MAG as compared to other closely related Mycoplasma genomes, indicating a distinct mechanism of carbon and energy acquisition by the oyster- associated Mollicutes. A systematic reanalysis of public eastern oyster-derived microbiome data revealed the Mollicutes as a ubiquitous taxon among adult oysters despite their general absence in larvae and biodeposit samples, suggesting potential horizontal transmission via an unknown mechanism. IMPORTANCEDespite well-documented biological significance of invertebrate microbiomes, a detailed taxonomic and functional characterization is frequently missing from many non-model marine invertebrates. By using 16S rRNA gene-based community profiling, shotgun metagenomics, and genome-scale metabolic reconstruction, this study provides an integrated taxonomic and functional analysis of the microbiome of the eastern oyster, Crassostrea virginica. Community profiling revealed a surprisingly low richness, as compared to surrounding seawater, and high variability among different tissue types and individuals. Reconstruction of a Mollicutes MAG enabled the phylogenomic positioning and functional characterization of the oyster-associated Mollicutes. Comparative analysis of the adult oyster gut, biodeposits, and oyster larvae samples indicated the potentially ubiquitous associations of the Mollicutes taxon with adult oysters. To the best of our knowledge, this study represented the first metagenomics derived functional inference of the eastern oyster microbiome. An integrated analytical procedure was developed for the functional characterization of microbiomes in other non-model host species.

bioinformatics↗