Microplastic (MP) contamination in the aquatic environment is a cause of concern worldwide since MP can be taken up by different organisms, altering different biological functions. In particular, evidence is accumulating that MP can affect the relationship between the host and its associated microbial communities (the microbiome), with potentially negative health consequences. Synthetic microfibers (MFs) represent one of the main MPs in the marine environment, which can be accumulated by filter-feeding invertebrates, such as bivalves, with consequent negative effects and transfer through the food chain. In the mussel Mytilus galloprovincialis, polyethylene terephthalate (PET) MFs, with a size distribution resembling that of an MF released from textile washing, have been previously shown to induce multiple stress responses. In this work, in the same experimental conditions, the effects of exposure to PET-MF (96 h, 10, and 100 μg/L) on mussel hemolymph microbiome were evaluated by 16S rRNA gene amplification and sequencing. The results show that PET-MF affects the composition of bacterial communities at the phylum, family and genus level, with stronger effects at the lowest concentration tested. The relationship between MF-induced changes in hemolymph microbial communities and responses observed at the whole organism level are discussed.

Introduction: Marine bivalves are continuously exposed to a variety of environmental stressors, including different Vibrio species potentially involved in disease outbreaks that have severely impacted farmed populations over the past two decades. Vibrio ostreicida was firstly identified as a lethal pathogen for oyster larvae ( Ostrea edulis ), but its interactions with the immune system of the bivalve host are largely unexplored. Methods: In this study, we investigated the pathogenic potential of a V. ostreicida strain r172, isolated from a 2022 mortality event in adult Mytilus galloprovincialis in the Ebro Delta (Spain), focusing on mussel early larval development and hemolymph immune responses in in vitro short-term exposure experiments. Immune responses were compared with those of the oyster Magallana gigas . Results: Both live and heat-killed V. ostreicida significantly impaired normal larval development with a dose-concentration effect (EC ₅₀ ≈ 10 ³ - 10 ₄ CFU/mL), with live bacteria inducing shell malformations and heat-killed Vibrio causing developmental arrest. In the hemocytes of adult mussels, exposure to heat-killed V. ostreicida led to dose-dependent lysosomal destabilization, reduced phagocytic activity, and increased intracellular ROS production. Similar lysosomal destabilization was observed with live V. ostreicida , which also stimulated extracellular ROS and nitric oxide release, but only in the presence of hemolymph serum (HS). Mussel HS displayed a strong bactericidal activity towards V. ostreicida , highlighting a key role for soluble immune effectors. In oyster hemocytes, V. ostreicida induced similar lysosomal stress; however, neither hemocytes nor serum showed any bactericidal activity towards this strain. Discussion: This data represents the first attempt to elucidate the mechanisms underlying the interactions of an environmental strain of V. ostreicida with marine bivalves. The species-specific differences observed in immune responses highlight the complexity of host-pathogen interactions in these organisms and emphasize the need for further investigation into immune responses of different aquacultured species.

Introduction: Vibrio species are widespread in coastal environments, where they are being increasingly associated with mortality episodes of farmed bivalves. This is the case for Vibrio ostreicida strain r172 isolated from a mortality event of adult mussels Mytilus galloprovincialis in Spain in 2022. Methods and Results: In this study, the infection dynamics and immune responses of adult M. galloprovincialis challenged with V. ostreicida r172 were investigated using different in vivo experimental approaches. First, a virulence assay by injection (1 x 108 CFU/100 µL) was performed at different temperatures (12, 18 and 24 °C). The results showed that mussel mortality (about 50 % within 8 days) was independent of increasing temperatures. Subsequently, an injection/cohabitation experiment was carried out placing together in the same tank V. ostreicida -injected (Donors, 108 CFU/mL) with un-injected mussels (Recipients). The time course of infection was then followed by evaluating positivity to V. ostreicida by PCR, responses of haemolymph components (haemocyte lysosomal membrane stability and serum lysozyme activity) and tissue histopathology (gills and digestive gland). The results showed a partial horizontal transfer of V. ostreicida from infected to uninfected mussels, with transient effects on haemolymph responses and histopathological lesions in both groups. Finally, in order to mimic more realistic environmental conditions, a bath infection experiment was carried out, exposing mussels to V. ostreicida in seawater (105 CFU/mL). This condition resulted in lower stress in haemocytes; moreover, no lysozyme release or histopathological alterations were observed. Discussion: Overall, the results show that M. galloprovincialis is able to cope with challenge with V. ostreicida , indicating that this Vibrio species is moderately pathogenic to adult mussels under the established experimental conditions.