Perkinsus olseni infections in Chinese mollusks have historically shown high prevalence but low intensity with no associated mortality. In August 2025, mass mortality (~50%) occurred in cultured Manila clams (Ruditapes philippinarum) in Nantong, China. Affected clams exhibited significantly higher infection intensity (mean >2.0 vs. 1.0; P < 0.01) and lower condition index (P < 0.0001) than healthy clams. Histopathology confirmed trophozoite aggregation in vital tissues. ITS sequencing identified P. olseni as the causative agent. Bacterial loads were significantly lower in affected clams, ruling out bacterial pathogens as the primary cause. This first report of Perkinsus-induced mortality in China challenges the benign parasite paradigm.

Whether zinc (Zn) and copper (Cu) bioaccumulation in the Pacific oyster (Magallana gigas) reflects toxicological stress or is an incidental consequence of growth remains unclear. We cultured three commercial triploid M. gigas stocks for approximately one year, sampling gill and hepatopancreas at the start and end of this period, when Zn/Cu burden was naturally low and high, respectively. Pooled samples from both time points were profiled by whole-transcriptome sequencing, enzyme activity and oxidative damage assays, qPCR validation, and protein-protein interaction network analysis. Transcriptome-wide changes in both tissues tracked the culture period, but growth and Zn/Cu burden were too highly collinear (r = 0.92-0.98) to separate statistically. Critically, of the four metals measured (Zn, Cu, iron [Fe], and manganese [Mn]), only Zn and Cu increased with growth, whereas Fe and Mn did not, indicating metal-specific rather than generalized accumulation. Superoxide dismutase (SOD) activity and the transcript abundance of its copper/zinc isoform (Cu/Zn-SOD) increased with growth in both tissues, whereas catalase (CAT) activity was unchanged and glutathione peroxidase (GPX) activity rose only in gill. Malondialdehyde (MDA), a marker of oxidative damage, increased in both tissues. Gill mounted a broader response than hepatopancreas, including upregulation of KEAP1 alongside downregulation of detoxification, proteostasis, and ribosome-related genes. Stock-level qPCR further revealed stock-dependent regulation of antioxidant genes in hepatopancreas. Together, these results indicate that Zn/Cu bioaccumulation in M. gigas co-varies with growth in a metal-specific manner, consistent with cofactor demand for Cu/Zn-SOD. The accompanying oxidative and proteostatic changes therefore more plausibly reflect growth physiology than an independent pollutant signal.

During summer 2025, mass mortalities (20%-80%) affected juvenile triploid Pacific oysters (Magallana gigas) across major mariculture zones of Northern China. Epidemiological investigations of 941 individuals from 58 batches confirmed a strong association with high Ostreid herpesvirus 1 (OsHV-1) loads. Multivariable analysis revealed that each log₁₀ increase in viral load conferred a 55% increase in mortality odds (OR = 1.55, p < 0.001), with loads >106 GE/mg associated with 94.0% mortality. A significant viral load-temperature interaction (p = 0.027) demonstrated temperature modulates pathogenicity. After accounting for this interaction, host species showed no significant effect (p = 0.564), indicating apparent differences between M. gigas and M. angulata reflect thermal habitats rather than genetic resistance. Histopathological examination revealed characteristic tissue damage, including hemocyte infiltration and connective tissue lysis. This event marks the first documented transition of the pathogen to open-sea juvenile populations. Genomic reconstruction showed that tree topology was primarily shaped by host species. Within the oyster-associated clade, the 2025 sea-farm variants grouped with a 2023 northern hatchery variant (SD23L), separately from European μVars-a pattern consistent with hatchery-to-sea spillover. One southern variant clustered with European μVars, raising the possibility of international introduction. Collectively, these findings suggest that the outbreak associated with changes in aquaculture practices-particularly the "North-South Relay" and year-round seedling production-rather than with emergence of a novel variant. This study underscores the urgent need for enhanced biosecurity and viral screening in China's oyster industry.

Ostreid herpesvirus 1 (OsHV-1) poses a major threat to bivalve aquaculture, but its impact on blood clams (Anadara broughtonii) remains poorly understood. This study integrated epidemiological and genomic data from 72 batches (1358 samples) collected across Chinese production systems (2019-2025). OsHV-1 was detected in hatcheries, holding facilities, and wild populations, with high viral loads linked to mass mortalities. Prevalence was significantly higher in land-based facilities (72%-74%) than in open-water systems (0%-50%; p < 0.001). Asymptomatic carriers, including wild populations and clams imported from the Republic of Korea, harbored lower viral loads than diseased individuals, acting as cryptic reservoirs. Temperature analysis identified disease onset at 13 degrees C and a viral load peak at 18.9 degrees C (95% CI: 16.4 degrees C-22.3 degrees C), approximately 3 degrees C-5 degrees C lower than those reported for oysters, reflecting host-specific viral adaptation. Effluent from land-based facilities tested virus-positive, indicating a potential transmission route. Random forest analysis identified source type as the strongest predictor of infection (AUC = 0.911). Mortality risk was strongly associated with temperature (OR = 2.65, p = 0.017) and viral load (OR = 4.23, p = 0.006). Genomic analysis revealed a distinct, host-specific lineage with no clear temporal progression. These findings demonstrate that effluent discharge, asymptomatic wild carriers, and international trade sustain OsHV-1 endemicity in blood clam aquaculture, highlighting the need for enhanced biosecurity measures calibrated to this cold-adapted pathosystem.

Bacterial disease is a major constraint on Magallana gigas larval production, yet hatchery microbiology has been characterised almost exclusively through the lens of Vibrio, and rarely across the whole production chain. We tracked bacterial communities throughout an entire larval rearing cycle at a commercial hatchery in northern China. A total of 78 samples were collected from the full production chain, from water intake to larvae. Bacterial communities were characterised using 16S rRNA (V4–V5) amplicon sequencing and culture-based isolation, with larvae sampled from three replicate tanks at six developmental stages. A protracted mortality event began at the D-veliger stage, with cumulative losses of roughly 40% before sinking ceased; ostreid herpesvirus 1 was not detected. The only taxon that rose above its healthy baseline during larval mortality was a single undescribed a single undescribed amplicon sequence variant (ASV) of the family Cryomorphaceae. Its relative abundance increased to a mean of 41% (with a tank-to-tank range of 33.7–51.6%) before disappearing once the mortality event concluded. No described species exceeds 92.2% 16S rRNA gene sequence identity to it, whereas its closest environmental relatives (97–98%) are, without exception, uncultured bacteria associated with marine invertebrates. It was an order of magnitude more abundant in larvae than in the surrounding water. Vibrio rose transiently at the onset of mortality but fell below its healthy-stage abundance while larvae were still dying. Tenacibaculum, by contrast, was the dominant genus of the rearing water yet was never recovered on Vibrio-selective medium and showed no association with mortality. Chlorination of the feed-room supply removed Tenacibaculum while leaving Vibrio uncontrolled. These findings reveal the limitations of the Vibrio-targeted, culture-based paradigm that has long dominated hatchery microbiology, as it overlooks both prevailing water-column organisms and mortality-linked taxa. Our results highlight the need for whole-community, culture-independent surveillance to better understand larval disease.

Ostreid herpesvirus 1 (OsHV-1) infection is the primary viral disease responsible for large-scale mortality in bivalve mollusks worldwide, and effective strategies to control the outbreaks of this disease are still lacking. Berberine (BBR), a plant-derived alkaloid, has demonstrated antiviral activity against various vertebrate viruses, while its potential antiviral effects on molluscan herpesviruses remain to be fully elucidated. Therefore, the present study sought to investigate the potential of berberine hydrochloride (BBH) against OsHV-1 infection in blood clams (Anadara broughtonii). The most optimal BBH concentration was figured out according to virus replication and mortality rates during in vivo experimental infection. Quantitative PCR and reverse transcription quantitative PCR were utilized to monitor the OsHV-1 genomic copy numbers and viral gene transcription levels during the development of OsHV-1 infection in the BBH-treated and control groups. The results demonstrated that a 3 mg/L BBH bath immersion significantly suppressed OsHV-1 replication in blood clams. During the early stage of infection (24 h), BBH treatment significantly reduced the expression of OsHV-1 open reading frames (ORFs) related to early enzymes, putative membrane proteins, and nucleocapsid proteins. At 96 h post-infection, all untreated blood clams died, whereas the survival rate of BBH-treated individuals increased to 46.67%. This study provides preliminary evidence for the inhibitory effects of BBH on OsHV-1, paving the way for the development of pharmacological control technologies for OsHV-1 infections.

Ostreid herpesvirus 1 (OsHV-1) is an infectious viral disease of bivalve mollusks with a wide geographic distribution across five continents. Since its initial identification in 1991, the disease has been identified as the cause of mortality in more than ten species of bivalve mollusks. The transmission of OsHV-1 is thought to be facilitated by water. In 2019 and 2021, mass mortalities of blood cockle (Anadara kagoshimensis) hatchery broodstock associated with OsHV-1 infection were reported. The presence of OsHV-1 DNA was confirmed in all hatchery samples with levels exceeding 106 copies per milligram of DNA. OsHV-1 was not detected in the wild populations from which the broodstock was collected, indicating that the virus was introduced into the hatchery with seawater from the open ocean. The results of the histopathological examination indicated that tissue lesions and associated hemocyte infiltration were most prevalent in the connective tissues of the digestive tract, mantle, gills and foot of the diseased A. kagoshimensis. In situ hybridization (ISH) signals of OsHV-1 DNA were frequently observed in infiltrated hemocytes, putative fibroblasts, and on occasion, in muscle cells with weak staining. Transmission electron microscopy analysis demonstrated the presence of herpes-like viral particles within the nucleus of infected cells. The genomes of the OsHV-1 variants from cases 2019 and 2021 were successfully resolved through direct high-throughput sequencing of infected tissues. A phylogenetic analysis of the complete genome sequences of 32 OsHV-1 variants revealed that the segregation of the virus was primarily structured by host species and geographic origin, with some exceptions. We propose that the sympatric distribution of different host species and the potential occurrence of cross-species transmission of OsHV-1 are responsible for the inconsistency between the phylogenetic segregation and their host species. This is the first report on the pathogenic and genomic characterization of OsHV-1 infecting A. kagoshimensis. The epidemiological and phylogenetic analyses offer new insights into the evolution of the virus and provide valuable information for the prevention and control of the disease.

Haliotid herpesvirus 1 (HAHV-1) causes significant damage to the abalone aquaculture industry. Knowledge of HAHV-1 invasion and host defense mechanisms is limited due to the lack of stable molluscan cell lines. The present study established an in vitro infection model of HAHV-1 using the primary suspension cultures of hemocytes from Haliotis diversicolor supertexta and Haliotis discus hannai. The cytopathic effects of HAHV-1 on adherent-cultured hemocytes of both species were also investigated. The HAHV-1 DNA loads were firstly monitored by means of quantitative PCR during the development of viral infection, and subsequently the mechanism of interaction between HAHV-1 and hemocytes was explored by means of a transcriptome analysis. H. diversicolor supertexta hemocytes exhibited a high degree of susceptibility to HAHV-1, with viral loads reaching a peak of 4.0 × 10⁷ copies/ng DNA. In contrast, no significant replication was observed in H. discus hannai hemocytes. Transcriptome analysis revealed that HAHV-1 evades the host immune response in the early stages of infection, and hijacks the host’s energy and redox metabolism to promote its replication at the late stages. Consequently, this study provides a valuable reference point for the investigation of virus−host interaction between HAHV-1 and abalone in vitro.

China is the most productive country in the world in terms of shellfish farming,with seawater shellfish occupying a dominant position in China's shellfish farming industry.In the past two decades,China's marine shellfish culture has been enriched in terms of species and culture methods.Moreover,its scale has expanded,with annual production remaining above 10 million tons and the total economic output exceeding 220 billion yuan.However,the economic losses caused by epidemic diseases in the shellfish aquaculture industry are also increasing annually,exceeding 10 billion yuan in 2021.These diseases have become some of the primary limiting factors for the healthy development of the shellfish aquaculture industry.Epidemiological surveys in recent years have shown that vibriosis is the most prevalent bacterial disease and the leading cause of mass mortality in shellfish farming.Vibrio alginolyticus is one of the most common Vibrio pathogens in shellfish diseases,posing a grave threat to the healthy development of the shellfish farming industry.However,effective methods for preventing and controlling V alginolyticus are still lacking.The pathogenicity of V alginolyticus is frequently closely related to its virulence factors and biological characteristics,and it is unclear how the virulence factors and biological characteristics of V alginolyticus vary according to different sources and regions.Therefore,in this study,we aimed to gain a deeper understanding of the pathogenicity and scientific control of V alginolyticus by analyzing the genetic variation and distribution patterns of V alginolyticus in different aquaculture environments and shellfish tissues.The study was conducted to observe the external morphological characteristics and conduct 16S identification of twelve V alginolyticus isolates collected from four regions:Qingdao,Weifang,Weihai,and Yantai.These were isolated and purified in TCBS selective medium;MLST typing of the strains by four housekeeping genes;the distribution of thirteen Vibrio virulence genes in V alginolyticus;and the resistance of V alginolyticus to 10 common antibiotics.The findings showed that all V alginolyticus colonies were yellow or yellowish in color,round and transparent in shape with a raised center and smooth edges,moist,and difficult to harvest.16S rRNA sequencing showed a homology of greater than 99％with V alginolyticus,which was initially verified as V alginolyticus and was consistent with the initial identification results.The ST typing of the twelve V algolyticus strains differed from each other.Seven strains(A2,A3,B2,B3,C3,D2,and D3)contained ST types already included in the PubMLST database,with ST types 45,87,156,56,125,96,and 57;five strains(A1,B1,C1,C2,and D1)formed new ST types owing to allelic locus changes in housekeeping genes,and four of the new ST types were isolated from shellfish tissue.The Qingdao isolate(A1)has similar ST types 38,131,134,46,and 56 in the database;the Weihai isolate(C1,C2)has similar ST types 111,322,and 61 in the database;and the Yantai isolate(D1)has similar ST types 268,275,341,344,351,and 358 in the database.These results suggested that V alginolyticus in the shellfish culture environment had a high genetic diversity and that V alginolyticus of shellfish origin might be more easily typed than V alginolyticus from water sources.The MLST typing phylogenetic tree showed that there were four distinct branches:Group 1,Group 2,Group 3,and Group 4.All V alginolyticus from shellfish tissues were predominantly observed in Group 1;isolates from Group 2 and Group 3 were mainly from marine environments and had closer evolutionary relationships with V alginolyticus from the same region of the aquatic environment.The evolutionary relationships between V alginolyticus from different areas of the aquatic environment and V alginolyticus from shellfish tissues showed different characteristics.All V alginolyticus strains carried three virulence genes:tlh,fur,and collagenase;VscB,Ompw,FlaA,and toxS virulence genes were present in most strains;UreB and AspA virulence genes were only present in a few strains;and tdh,trh,toxR,and tcpA virulence genes were not detected in any of the strains.The variety and number of virulence factors carried by V alginolyticus were influenced by factors such as regional distribution.V alginolyticus of different origins were characterized by multiple drug resistance interactions,but there were differences in the types of antibiotics to which resistance was developed.All V alginolyticus species showed high susceptibility to cotrimoxazole and chloramphenicol,while they were resistant to penicillin and ampicillin.The majority of V alginolyticus developed intermediate or high susceptibility to antibiotics such as butyraminecarbana,gentamicin,erythromycin,norfloxacin,and ciprofloxacin.Among all V alginolyticus,one strain was resistant to ceftolozoline,five strains were intermediated,and six strains were highly sensitive.In addition,five strains were intermediated,and six were highly sensitive to butyraminecarbana,gentamicin,and erythromycin.Two strains were intermediated,and ten were highly sensitive to norfloxacin.Four strains were intermediated,and eight were highly sensitive to ciprofloxacin.Combining the MLST typing results,drug resistance results,and virulence genes in this study,no significant correlation between the three was found at this time.This study showed that V alginolyticus in a shellfish culture environment was characterized by complex populations and high genetic diversity.There were large differences in virulence gene carriage and drug resistance among strains from different sources.The study provides a theoretical reference framework for understanding the pathogenicity of V alginolyticus and assisting in the effective control of V alginolyticus of shellfish origin by investigating the genetic variation and drug resistance of V alginolyticus from different sources in various regions.

Ostreid herpesvirus 1 (OsHV-1), a member of the family Malacoherpesviridae (order Herpesvirales), is a major pathogen of bivalves. However, the molecular details of the malacoherpesvirus infection cycle and its overall similarity to the replication of mammalian herpesviruses (family Orthoherpesviridae) remain obscure. Here, to gain insights into the OsHV-1 biology, we performed long-read sequencing of infected blood clams, Anadara broughtonii, which yielded over one million OsHV-1 long reads. These data enabled the annotation of the viral genome with 78 gene units and 274 transcripts, of which 67 were polycistronic mRNAs, 35 ncRNAs, and 20 natural antisense transcripts (NATs). Transcriptomics and proteomics data indicate preferential transcription and independent translation of the capsid scaffold protein as an OsHV-1 capsid maturation protease isoform. The conservation of this transcriptional architecture across Herpesvirales likely indicates its functional importance and ancient origin. Moreover, we traced RNA editing events using short-read sequencing and supported the presence of inosine nucleotides in native OsHV-1 RNA, consistent with the activity of adenosine deaminase acting on dsRNA 1 (ADAR1). Our data suggest that, whereas RNA hyper-editing is concentrated in specific regions of the OsHV-1 genome, single-nucleotide editing is more dispersed along the OsHV-1 transcripts. In conclusion, we reveal the existence of conserved pan-Herpesvirales transcriptomic architecture of the capsid maturation module and uncover a transcription-based viral counter defence mechanism, which presumably facilitates the evasion of the host ADAR antiviral system.

Perkinsosis has been recognized as one of the major threats to natural and farmed bivalve populations, many of which are of commercial as well as environmental significance. Three Perkinsus species have been identified in China, and the Manila clam (Ruditapes philippinarum) was the most frequently infected species in northern China. Although the occurrence and seasonal variation of Perkinsus spp. have previously been examined, the pathological characteristics of these infections in wild Manila clams and sympatric species in China have seldom been reported. In the present study, the prevalence and intensity of Perkinsus infection in wild populations of Manila clams and 10 sympatric species from three sites were investigated by Ray's fluid thioglycolate medium (RFTM) assay seasonally across a single year. Perkinsus infection was only identified in Manila clams, with a high prevalence (274/284 = 96.48 %) and low intensity (89.8 % with a Mackin value <= 2, suggesting generally lowintensity infections) throughout the year. Heavily infected clams were mainly identified in Tianheng in January, which displayed no macroscopic signs of disease. An overview of the whole visceral mass section showed that the trophozoites mostly aggregated in gills and connective tissue of the digestive tract, to a lesser extent in the mantle and foot, and even less frequently in adductor muscle and connective tissues of the gonad. PCR and ITS-5.8S rRNA sequencing of 93 representative RFTM-positive samples revealed a 99.69 to 100 % DNA sequence identity to Perkinsus olseni. Unexpectedly, significantly higher infection intensities were usually identified in January and April when the Condition Index (CI) was relatively high. We propose that factors associated with the anthropogenic harvesting pressure and irregular disturbances should be responsible for the uncommon seasonal infection dynamics of perkinsosis observed in the present study.

OsHV-1 caused detrimental infections in a variety of bivalve species of major importance to aquaculture worldwide. Since 2012, there has been a notable increase in the frequency of mass mortality events of the blood clam associated with OsHV-1 infection. The pathological characteristics, tissue and cellular tropisms of OsHV-1 in A. broughtonii remain unknown. In this study, we sought to investigate the distribution of OsHV-1 in five different organs (mantle, hepatopancreas, gill, foot, and adductor muscle) of A. broughtonii by quantitative PCR, histopathology and in situ hybridization (ISH), to obtain insight into the progression of the viral infection. Our results indicated a continuous increase in viral loads with the progression of OsHV-1 infection, reaching a peak at 48 h or 72 h post-infection according to different tissues. Tissue damage and necrosis, as well as colocalized OsHV-1 ISH signals, were observed primarily in the connective tissues of various organs and gills. Additionally, minor tissue damage accompanied by relatively weak ISH signals was detected in the foot and adductor muscle, which were filled with muscle tissue. The predominant cell types labeled by ISH signals were infiltrated hemocytes, fibroblastic-like cells, and flat cells in the gill filaments. These results collectively illustrated the progressive alterations in pathological confusion and OsHV-1 distribution in A. broughtonii, which represent most of the possible responses of cells and tissues to the virus.

The ferritin secreted by mammals has been well documented, with the protein capable of localizing to cell membranes and facilitating the delivery of iron to cells through endocytosis. However, the presence of ferritin in the circulatory fluid of mollusks and its functions remain largely unknown. In this study, we aimed to investigate the potential interacting proteins of ferritin in the ark clam (SbFn) through the use of a pull-down assay. Our findings revealed the presence of an insulin-like growth factor type 1 receptor (IGF-1R) in ark clams, which was capable of binding to SbFn and was named SbIGF-1R. SbIGF-1R was found to be composed of two leucine-rich repeat domains (L domain), a cysteine-rich domain, three fibronectin type III domains, a transmembrane domain, and a tyrosine kinase domain. The ectodomain of SbIGF-1R was observed to form a symmetrical antiparallel homodimer in the shape of the letter ‘A’, with the fibronectin type III domains serving as its ‘legs’. The mRNA expression of SbIGF-1R gene was detected ubiquitously in various tissues of the ark clam, with the highest expression levels found in hemocytes, as determined by qRT-PCR. Using a confocal microscopic and yeast two-hybrid assays, the interaction between SbIGF-1R and SbFn was further verified. The results showed that SbFn co-localized with SbIGF-1R on the cell membrane, and their interaction was expected to occur on the FNIII domains of the SbIGF-1R. In conclusion, our findings highlight the identification of a putative receptor, SbIGF-1R, for SbFn, demonstrating the versatility of IGF-1R in ark clams.

In aquaculture, high-density seaweed farming brings higher economic benefits but also increases outbreaks of diatom felt.The effective control of diatom felt in high-density seaweed farming has always been a research hotspot.This study selected two potential allelochemicals 2-hydroxycinnamic acid and quinic acid to explore their effects on a diatom Nitzschia closterium and an economic seaweed Monostroma nitidum.The results showed that 2-hydroxycinnamic acid had better inhibitory effects than quinic acid on the growth, pigment content and photosynthetic efficiency of N. closterium.Their half-maximal inhibitory concentrations at 120 h (IC 50-120 h ) were 0.9000 and 1.278 mM, respectively.Additionally, these allelochemicals had limited inhibitory effects on the growth, pigment content and photosynthetic efficiency of M. nitidum before 24 h.To further explore the allelopathic effect of these chemicals, this study focused on the photosystem II energy fluxes of N. closterium.It was found that 3 mM 2-hydroxycinnamic acid could destroy the whole photosynthetic system by devastating the PSII reaction centre (RC) before 24 h; however, the same concentration of quinic acid could only down-regulate the electron transport efficiency by changing the effective antenna size of an active RC and downregulating the PSII reaction centre density.These experimental results are expected to provide a new strategy to control diatom felt blooms on the high-density seaweed farming areas.

Global warming causes great thermal stress to macroalgae and those species that can adapt to it are thought to be better able to cope with warmer oceans. Gracilaria bailinae, a macroalgae with high economic and ecological values, can survive through the hot summer in the South China Sea, but the molecular mechanisms underlying its adaptation to high temperatures are unclear. To address this issue, the present study analyzed the growth and transcriptome of G. bailinae after a 7-day exposure to 15°C (LT: low temperature), 25°C (MT: middle temperature), and 35°C (HT: high temperature). Growth analysis showed that the HT group had the highest relative growth rate (RGR = 2.1%) with the maximum photochemical quantum yield of PSII (Fv/Fm = 0.62) remaining within the normal range. Transcriptome analysis showed more differentially expressed genes (DEGs) in the comparison between MT and HT groups than in that between MT and LT, and most of these DEGs tended to be downregulated at higher temperatures. The KEGG pathway enrichment analysis showed that the DEGs were mainly enriched in the carbohydrate, energy, and lipid metabolisms. In addition, the genes involved in NADPH and ATP synthesis, which are associated with photosynthesis, the Calvin cycle, pyruvate metabolism, and the citrate cycle, were downregulated. Downregulation was also observed in genes that encode enzymes involved in fatty acid desaturation and alpha-linolenic acid metabolism. In summary, G. bailinae regulated the synthesis of NADPH and ATP, which are involved in the above-mentioned processes, to reduce unnecessary energy consumption, and limited the synthesis of enzymes in the metabolism of unsaturated fatty acids and alpha-linolenic acid to adapt to high environmental temperatures. The results of this study improve our understanding of the molecular mechanisms underlying the adaptation of G. bailinae to high temperatures.

The Pacific oyster (Crassostrea gigas) aquaculture industry increased rapidly in China with the introduction and promotion of triploid oysters in recent years. Mass mortalities affecting different life stages of Pacific oysters emerged periodically in several important production areas of Northern China. During 2020 and 2021, we conducted a passive two-year investigation of infectious pathogens linked to mass mortality. Ostreid herpesvirus-1 (OsHV-1) was detected to be associated with mass mortalities of hatchery larvae, but not juveniles and adults in the open sea. Protozoan parasites, such as Marteilia spp., Perkinsus spp. and Bonamia spp. were not detected. Bacterial isolation and identification revealed that Vibrio natriegens and Vibrio alginolyticus were the most frequently (9 out of 13) identified two dominant bacteria associated with mass mortalities. Pseudoalteromonas spp. was identified as the dominant bacteria in three mortality events that occurred during the cold season. Further bacteriological analysis was conducted on two representative isolates of V. natriegens and V. alginolyticus, designated as CgA1-1 and CgA1-2. Multisequence analysis (MLSA) showed that CgA1-1 and CgA1-2 were closely related to each other and nested within the Harveyi clade. Bacteriological investigation revealed faster growth, and more remarkable haemolytic activity and siderophore production capacity at 25 °C than at 15 °C for both CgA1-1 and CgA1-2. The accumulative mortalities of experimental immersion infections were also higher at 25 °C (90% and 63.33%) than at 15 °C (43.33% and 33.33%) using both CgA1-1 and CgA1-2, respectively. Similar clinical and pathological features were identified in samples collected during both naturally and experimentally occurring mortalities, such as thin visceral mass, discolouration, and connective tissue and digestive tube lesions. The results presented here highlight the potential risk of OsHV-1 to hatchery production of larvae, and the pathogenic role of V. natriegens and V. alginolyticus during mass mortalities of all life stages of Pacific oysters in Northern China.

The Pacific oyster Crassostrea gigas is one of the most important cultured marine species around the world. Production of Pacific oysters in China has depended primarily on hatchery produced seeds since 2016, with the successful introduction and development of triploid oysters. However, the seed supply of Pacific oysters is threatened by recurring mass mortality events in recent years. Vibriosis is the most commonly encountered disease associated with intensive oyster culture in hatcheries and nurseries. Vibrio alginolyticus and Bacillus hwajinpoensis were the two strains with pathogenic and probiotic effects, respectively, identified during the Pacific oyster larvae production. To monitor their colonization process in Pacific oyster larvae, green fluorescent protein (GFP) and red fluorescent protein (RFP) were labeled to the pathogenic V. alginolyticus and the probiotic B. hwajinpoensis stain, respectively. The pathogenic and probiotic effects of the two strains during the colonization process were then assessed. Stabile expression of GFP and RFP were observed in corresponding stains, and the capabilities of growth, biofilm formation and in vitro adhesion of GFP- and RFP- tagged stains were not significantly different from those of the wild-type strains. Usage of probiotics of 105 CFU/mL significantly inhibited the growth of pathogenic V. alginolyticus and reduced the mortality of D-sharped larvae. Both the pathogenic and probiotic strains employed a similar route to enter and colonize the oyster larvae, which indicates that competing with pathogens for binding and spreading sites were one of the mechanisms of B. hwajinpoensis to provide the probiotic effects to oyster larvae. In summary, employment of fluorescence-tagged pathogenic and probiotic strains simultaneously provides us with an excellent bioassay model to investigate the potential mechanisms of probiotics.

Elemental iron is an indispensable prosthetic group of DNA replication relative enzymes. The upregulation of ferritin translation by iron regulatory proteins (IRP1) in host cells is a nutritional immune strategy to sequester available iron to pathogens. The efficient replication of Ostreid herpesvirus 1 (OsHV-1), a lethal dsDNA virus among bivalves, depends on available iron. OsHV-1 infection was found to trigger iron limitation in ark clams; however, it is still an enigma how OsHV-1 successfully conducted rapid replication, escaping host iron limitations. In this study, we identified the IRP1 protein (designated as SbIRP-1) in the ark clam (Scapharca broughtonii) and found it could bind to the iron-responsive element (IRE) of ferritin (SbFn) mRNA based on electrophoretic mobility shift assay (EMSA). Knockdown of SbIRP-1 expression (0.24 ± 1.82-fold of that in NC group, p < 0.01) by RNA interference resulted in the accumulation of SbFn in hemocytes (1.79 ± 0.01-fold, p < 0.01) post-24 h of enhanced RNA interference injection. During OsHV-1 infection, SbFn mRNA was significantly upregulated in hemocytes from 24 h to 60 h, while its protein level was significantly reduced from 24 h to 48 h, with the lowest value at 36 h post-infection (0.11 ± 0.01-fold, p < 0.01). Further analysis by RNA immunoprecipitation assays showed that OsHV-1 could enhance the binding of SbIRP-1 with the SbFn IRE, which was significantly increased (2.17 ± 0.25-fold, p < 0.01) at 36 h post-infection. Consistently, SbIRP-1 protein expression was significantly increased in hemocytes from 12 h to 48 h post OsHV-1 infection (p < 0.01). In conclusion, the results suggest that OsHV-1 infection could suppress post-transcriptional translation of SbFn through the regulation of SbIRP-1, which likely contributes to OsHV-1 evasion of SbFn-mediating host iron limitation.

The genera Monostroma and Gayralia belong to the order of monostromatic green algae; however, their taxonomic delimitation remains controversial at the genus level. This study attempts to address this issue through the combined analysis of the morphology and nuclear-encoded Internal Transcribed Spacer region sequences of monostromatic green algal samples collected in the South China Sea. Our phylogenetic data revealed that the monostromatic specimens were separated into the M. nitidum clade, G. brasiliensis clade, and a single Monostroma sp. clade, and that the inter-genera genetic distance between the Monostroma and Gayralia genera was lower than that observed within the Monostroma genus. All the specimens presented similar morphology in their single cell-layered thallus, with irregularly arranged cells, rounded cell corners, a parietal chloroplast, and predominantly one (>90%) pyrenoid. Their most obvious morphological difference was in thallus thickness and size. Moreover, the monostromatic specimens of the M. nitidum clade corresponded to the morphological description of the M. nitidum-type specimens. The genus Monostroma was erected earlier than the genus Gayralia. Therefore, we propose to assign the genus Gayralia to Monostroma based on the morphological and phylogenetic analysis and genetic distance data presented here.

Macroalgae can accumulate a wide array of metals, leading to their appliance as biomonitors of aquatic environments. With the rapid development of industrial and agricultural-based activities, Cd pollution in aquatic environments is considered an increasingly severe problem worldwide. Although La could alleviate the Cd stress in higher terrestrial plants, the response mechanisms of macroalgae to Cd and La are unknown. Along these lines, in this work, Cd significantly affected the growth, internal cellular structure, photosynthesis, pigment content, antioxidant enzyme activity, and lipid peroxidation level of G. bailiniae. However, the presence of La alleviated these adverse effects from Cd. Furthermore, the response mechanism of G. bailiniae to Cd was attributed to the self-antioxidant ability enhancement, membrane defense, and programmed-cellular regulation. However, the presence of La mediated the biosynthesis of both flavonoids and lipids, which inhibited the Cd accumulation, modulated algal stress signalling networks, renewed the impaired chlorophyll molecule, maintained the activity of the crucial enzyme, enhanced antioxidant ability, and maintained the stabilization of redox homeostasis, alleviating the adverse impact from Cd and improve the growth of G. bailiniae. The experimental results successfully demonstrate a new detoxicant to alleviate Cd stress, promoting a more comprehensive array of macroalgal applications.