Lymphocystis Disease Virus Research Articles

Preliminary studies on the detection and presence of Lymphocystis disease virus (LCDV) in sea breams (Sparus aurata) raised in the Aegean Sea

Preliminary studies on the detection and presence of Lymphocystis disease virus (LCDV) in sea breams (Sparus aurata) raised in the Aegean Sea

Susceptibility and Target Organ of Lymphocystis Disease Virus Infection in Giant Gourami (Osphronemus goramy), Hybrid Tilapia (Oreochromis sp.), Siamese Fighting Fish (Betta splendens), and Hybrid Catfish (Clarias sp.)

Highlight Research 1. Lymphocystis disease is reported to infect seawater and freshwater fishes 2. The four important freshwater fish species in Indonesia are evaluated on their susceptibility to Lymphocystis Disease Virus 3. Lymphocystis infection causes behavioural changes and mortality with different onset times after infection on the four fish species 4. LCDV load is varied in quantity among different organs. Abstract Lymphocystis disease has a broad host range and has been reported to enter Indonesia. However, information regarding its susceptibility and predilection organs in fish is lacking. This study examined the susceptibility of four important fish species in Indonesia, namely, giant gourami (Osphronemus goramy), hybrid tilapia (Oreochromis sp.), Siamese fighting fish (Betta splendens), and hybrid catfish (Clarias sp.). The fish were infected with virus filtrate by intraperitoneal injection and immersion. The postinfection observation period was 60 days. Viral load was quantified by qPCR and expressed as major capsid protein (MCP) copy number/mg tissue. Mortality was observed in all fish species, with the highest recorded in hybrid catfish and the lowest in Siamese fighting fish. All the fish species showed changes in their clinical symptoms, such as anorexia and separation from schools. However, only giant gourami showed internal change seven days after injection (dpi), with white lesion detected in the liver. Viral load quantification showed that LCDV had different predilection organs in the four fish species. The highest viral load of giant gourami (1.7 x 104) was observed in the liver at 7 dpi, hybrid tilapia (7.5 x 103) was observed in the fins at 21 dpi, Siamese fighting fish (8.4 x 103) was observed in the fins at 14 dpi, and hybrid catfish (1.2 x 103) were observed in the fins and gills at 7 and 14 dpi. The findings indicated that giant gourami, hybrid tilapia, Siamese fighting fish, and hybrid catfish were susceptible to LCDV infection with different predilection organs.

Molecular characterization of lymphocystis disease virus in Indian glass fish: first report from the Andaman Islands.

Here, we report the first detection of lymphocystis disease virus (LCDV) in Indian glass fish in the Andaman Islands, India. Microscopic examination revealed the presence of whitish clusters of nodules on the fish's skin, fins, and eyes. The histopathology of the nodules revealed typical hypertrophied fibroblasts. Molecular characterization of the major capsid protein (MCP) gene of the virus showed a significant resemblance to known LCDV sequences from Korea and Iran, with 98.92% and 97.85% sequence identity, respectively. Phylogenetic analysis confirmed that the MCP gene sequence of the virus belonged to genotype V. This study represents the first documented case of LCDV in finfish from the Andaman Islands, emphasizing the necessity for continued monitoring and research on the health of aquatic species in this fragile ecosystem.

Extracellular traps in skin lesions infected with lymphocystis disease virus in black rockfish (Sebastes schlegelii)

The lymphocystis disease (LCD), caused by Lymphocystis disease virus (LCDV), is a benign and self-limiting disease described in a many freshwater and marine fish species. Hypertrophic fibroblasts and extensive aggregation of inflammatory cells are characteristics of LCD. In the present study, small animal imaging and ultrastructural investigations were carried out on the lymphocystis nodules of black rockfish (Sebastes schlegelii) naturally infected with lymphocystis iridovirus, to assess pathology, and the exudate with particular attention to the formation of extracellular traps (ETs) in vivo. Ex vivo were examined by nodules sections and primary cells stimulation. By histopathological analysis, the nodules contained infiltrated inflammatory cells and extensive basophilic fibrillar filaments at the periphery of the hypertrophied fibroblasts. ETs were assessed in nodules samples using indirect immunofluorescence to detect DNA and myeloperoxidase. Moreover, LCDV was able to infect peritoneal cells of black rockfish in vitro and induce the formation of ETs within 4 h. In summary, this study proved that ETs are involved in the response to LCDV infection and may be involved in formation of lymphoid nodules. Taken together, the findings provide a new perspective to determine the impact factors on the growth of nodules.

Comprehensive surveillance and molecular profiling of viral infections in cultured olive flounder (Paralichthys olivaceus) across Jeju Island, South Korea

To effectively control viral diseases, ongoing research into virus detection and their genetic characteristics is imperative. We conducted an analysis of the prevalence of five species of viruses in olive flounder (Paralichthys olivaceus) farms in Jeju, specifically viral haemorrhagic septicaemia virus (VHSV), lymphocystis disease virus (LCDV), marine birnavirus (MABV), nervous necrosis virus (NNV), and flounder iridovirus (FLIV). Between July and December 2022, a total of 3282 olive flounders were monitored. VHSV and LCDV were detected in 116 and 173 olive flounders, respectively, confirming detection rates of 3.5% and 5.4%. The highest detection rate for VHSV occurred in December (12.5%), while for LCDV, it was in June (13.8%). MABV, NNV, and FLIV were not detected. Phylogenetic analysis of 19 VHSV isolates collected from 2010 to 2022 revealed that all VHSVs belonged to the IVa genotype based on the nucleoprotein gene. Mutations were identified, indicating genetic variability extending to protein mutations. Additionally, continuous and specific nucleotide mutations of VHSV were confirmed on Jeju Island, a central hub for the aquaculture of olive flounder. These observed mutations in VHSV are considered valuable markers for tracking future mutations in subsequent studies. LCDV isolates were classified under genotype II. The results of the phylogenetic analyses of VHSV, a negative-sense single-stranded RNA virus, and LCDV, a double-stranded DNA virus, provide insights into the mutation patterns of marine viruses.

Characterization and function of Japanese flounder (Paralichthys olivaceus) slc2a6 in response to lymphocystis disease virus infection

Slc2a6 is a member of the slc2 family (solute carrier 2 family) and previous reports have indicated its involvement in the inflammatory response. Slc2a6 is regulated by the NF-ĸB signaling pathway. This study investigated the differential expression of slc2a6 in the early embryonic development of Japanese flounder, revealing that the early gastrula stage had the highest level of slc2a6 expression. Moreover, slc2a6 expression was increased in vitro after stimulation by lymphocystis disease virus (LCDV), and in vivo experiments also showed significantly elevated levels in the spleen and muscle tissues following LCDV stimulation. Subcellular localization revealed that Slc2a6 was expressed in both the nucleus and cytoplasm of cells. The pcDNA3.1-slc2a6 overexpression plasmid was successfully constructed; the si-slc2a6 interfering strand was screened and samples were collected. The expression of NF-ĸB signaling pathway-related genes il-1β, il-6, nf-ĸb, and tnf-α was evaluated in overexpressed, silenced, and LCDV-stimulated samples. The results showed that slc2a6 is involved in viral regulation in Japanese flounder by regulating innate immune responses.

Transcriptome analysis reveals molecular mechanisms of lymphocystis formation caused by lymphocystis disease virus infection in flounder (Paralichthys olivaceus).

Lymphocystis disease is frequently prevalent and transmissible in various teleost species worldwide due to lymphocystis disease virus (LCDV) infection, causing unsightly growths of benign lymphocystis nodules in fish and resulting in huge economic losses to aquaculture industry. However, the molecular mechanism of lymphocystis formation is unclear. In this study, LCDV was firstly detected in naturally infected flounder (Paralichthys olivaceus) by PCR, histopathological, and immunological techniques. To further understand lymphocystis formation, transcriptome sequencing of skin nodule tissue was performed by using healthy flounder skin as a control. In total, RNA-seq produced 99.36%-99.71% clean reads of raw reads, of which 91.11%-92.89% reads were successfully matched to the flounder genome. The transcriptome data showed good reproducibility between samples, with 3781 up-regulated and 2280 down-regulated differentially expressed genes. GSEA analysis revealed activation of Wnt signaling pathway, Hedgehog signaling pathway, Cell cycle, and Basal cell carcinoma associated with nodule formation. These pathways were analyzed to interact with multiple viral infection and tumor formation pathways. Heat map and protein interaction analysis revealed that these pathways regulated the expression of cell cycle-related genes such as ccnd1 and ccnd2 through key genes including ctnnb1, lef1, tcf3, gli2, and gli3 to promote cell proliferation. Additionally, cGMP-PKG signaling pathway, Calcium signaling pathway, ECM-receptor interaction, and Cytokine-cytokine receptor interaction associated with nodule formation were significantly down-regulated. Among these pathways, tnfsf12, tnfrsf1a, and tnfrsf19, associated with pro-apoptosis, and vdac2, which promotes viral replication by inhibiting apoptosis, were significantly up-regulated. Visual analysis revealed significant down-regulation of cytc, which expresses the pro-apoptotic protein cytochrome C, as well as phb and phb2, which have anti-tumor activity, however, casp3 was significantly up-regulated. Moreover, bcl9, bcl11a, and bcl-xl, which promote cell proliferation and inhibit apoptosis, were significantly upregulated, as were fgfr1, fgfr2, and fgfr3, which are related to tumor formation. Furthermore, RNA-seq data were validated by qRT-PCR, and LCDV copy numbers and expression patterns of focused genes in various tissues were also investigated. These results clarified the pathways and differentially expressed genes associated with lymphocystis nodule development caused by LCDV infection in flounder for the first time, providing a new breakthrough in molecular mechanisms of lymphocystis formation in fish.

PCR detection and phylogenetic analysis of the major capsid protein gene of Lymphocystis disease virus from cultured Asian seabass (Lates calcarifer Bloch, 1790) along the west coast of Kerala, India

PCR detection and phylogenetic analysis of the major capsid protein gene of Lymphocystis disease virus from cultured Asian seabass (Lates calcarifer Bloch, 1790) along the west coast of Kerala, India

Immune response of DNA vaccinated-gilthead seabream (Sparus aurata) against LCDV-Sa infection: relevance of the inflammatory process.

Lymphocystis disease is one of the main viral pathologies affecting cultured gilthead seabream (Sparus aurata) in the Mediterranean region. Recently, we have developed a DNA vaccine based on the major capsid protein (MCP) of the Lymphocystis disease virus 3 (LCDV-Sa). The immune response triggered by either LCDV-Sa infection or vaccination have been previously studied and seem to be highly related to the modulation of the inflammatory and the IFN response. However, a comprehensive evaluation of immune-related gene expression in vaccinated fish after viral infection to identify immunogenes involved in vaccine-induced protection have not been carried out to date. The present study aimed to fulfill this objective by analyzing samples of head-kidney, spleen, intestine, and caudal fin from fish using an OpenArray® platform containing targets related to the immune response of gilthead seabream. The results obtained showed an increase of deregulated genes in the hematopoietic organs between vaccinated and non-vaccinated fish. However, in the intestine and fin, the results showed the opposite trend. The global effect of fish vaccination was a significant decrease (p<0.05) of viral replication in groups of fish previously vaccinated, and the expression of the following immune genes related to viral recognition (tlr9), humoral and cellular response (rag1 and cd48), inflammation (csf1r, elam, il1β, and il6), antiviral response (isg15, mx1, mx2, mx3), cell-mediated cytotoxicity (nccrp1), and apoptosis (prf1). The exclusive modulation of the immune response provoked by the vaccination seems to control the progression of the infection in the experimentally challenged gilthead seabream.

VPIF-1 is an insulin-like antiferroptotic viral peptide

Iridoviridae, such as the lymphocystis disease virus-1 (LCDV-1) and other viruses, encode viral insulin-like peptides (VILPs) which are capable of triggering insulin receptors (IRs) and insulin-like growth factor receptors. The homology of VILPs includes highly conserved disulfide bridges. However, the binding affinities to IRs were reported to be 200- to 500-fold less effective compared to the endogenous ligands. We therefore speculated that these peptides also have noninsulin functions. Here, we report that the LCDV-1 VILP can function as a potent and highly specific inhibitor of ferroptosis. Induction of cell death by the ferroptosis inducers erastin, RSL3, FIN56, and FINO2 and nonferroptotic necrosis produced by the thioredoxin-reductase inhibitor ferroptocide were potently prevented by LCDV-1, while human insulin had no effect. Fas-induced apoptosis, necroptosis, mitotane-induced cell death and growth hormone-releasing hormone antagonist-induced necrosis were unaffected, suggesting the specificity to ferroptosis inhibition by the LCDV-1 VILP. Mechanistically, we identified the viral C-peptide to be required for inhibition of lipid peroxidation and ferroptosis inhibition, while the human C-peptide exhibited no antiferroptotic properties. In addition, the deletion of the viral C-peptide abolishes radical trapping activity in cell-free systems. We conclude that iridoviridae, through the expression of insulin-like viral peptides, are capable of preventing ferroptosis. In analogy to the viral mitochondrial inhibitor of apoptosis and the viral inhibitor of RIP activation (vIRA) that prevents necroptosis, we rename the LCDV-1 VILP a viral peptide inhibitor of ferroptosis-1. Finally, our findings indicate that ferroptosis may function as a viral defense mechanism in lower organisms.

An outbreak of lymphocystis disease virus infection in cultured juvenile John's snapper (Lutjanus johnii) in Malaysia

This study reports an outbreak of lymphocystis disease virus infection in cultured juvenile John's snapper (Lutjanus johnii) in Malaysia. In mid-January 2021, approximately 19,000 juvenile John's snapper exhibited signs of lethargy and isolating themselves from the schooling group, developed lymphocysts all over the body and fins, and died. The outbreak started at day 3 after the introduction of fish into the hatchery, and the severity of the disease was observed between day 7 and 14, resulting in 53% mortality. No bacteria was isolated, while a low prevalence of protozoan parasite was observed at 5% of fish samples. Histopathology examination revealed focal nodular dermal fibroblast hypertrophy with karyomegaly and hyaline capsule formation. Basophilic intracytoplasmic inclusions were observed in the diseased fish. PCR and sequence analyses revealed that all affected fish were positive to lymphocystis disease virus (LCDV), which was absent from the clinically healthy fish. Phylogenetic analysis revealed that the isolates from this outbreak were from the cluster of LCDV genotype 2. This paper reports the first documentation of LCDV infection in cultured John's snapper in Malaysia. Farmers and authorities should beware of the impacts LCDV infection, as it can cause high mortality and economic losses in cultured fish.

DETECTION AND GENOTYPE DETERMINATION OF LYMPHOCYSTIS DISEASE VIRUS FROM ORANGE CLOWNFISH Amphiprion percula IN BATAM

The orange clownfish, Amphiprion percula, is a popular fish in the global marine ornamental trade. In March 2021, several orange clown fish at Batam Mariculture Development Center exhibited lethargic behavior with wart-like nodules on the fins suggesting a viral disease infection. This study aimed to detect the suspected viral disease agent from the clownfish and determine its genotype based on the major capsid protein gene. The fish tissues with wart-like nodules were collected and fixed in 96% ethanol. The DNA was extracted from the tissues and used as the template for the amplification of the major capsid protein (MCP) and myristylated membrane protein (MMP) genes using polymerase chain reaction (PCR). The nucleotide sequences of the PCR products were analyzed for their homology using the Basic Local Alignment Search Tool (BLAST). Multiple alignments of the amino acid sequence of MCP were performed using MEGA-X to determine the genotype. The PCR amplification produced the expected bands for detecting MCP, MMP and DNAPol genes. These results indicated the presence of Lymphocystis disease virus (LCDV), designated as LCDV-Oc-Btm. The sequencing of MCP and MMP genes produced the 1221 and 407 nucleotides, respectively. The BLAST analysis showed the highest identity was obtained with the species of LCDV-1 (LCDV-Sa strain) at 91.04 % and 88.19 % for MCP and MMP, respectively. The UPGMA phylogenic tree showed LCDV-Oc-Btm differs from the existing genotype and can be assigned as a new genotype. This study concludes that LCDVOc-Btm is a novel species of lymphocystis disease virus.

Lymphocystis Disease Virus Infection in Clownfish Amphiprion ocellaris and Amphiprion clarkii in Taiwan.

Lymphocystic disease affects over 150 species of marine and freshwater fish worldwide. In this study, the lymphocystis pathogen was found in 2 (Amphiprion ocellaris and Amphiprion clarkii) of the 9 species of clownfish. Detection of lymphocystis disease virus (LCDV) was based on histopathological study, electron microscope observation of virus particles and gene sequence analysis from the MCP region. Infected A. ocellaris hosts showed sparse, multifocal, white, stiff, papilloma-like nodules on the body, skin, gills and fins; while, on A. clarkia, nodules were found on the operculum skin. Histopathologic study showed lymphocystic cells with an irregular nucleus, enlarged cytoplasm and intracytoplasmic inclusion bodies surrounded by the cell membrane. The viral particle presents virions 180-230 nm in diameter, hexagonal in shape with an inner dense nucleoid under transmission electron micrographs (TEM). From the ML polygenetic tree, the clownfish LCVD genotype was closely related to the LCDV strain from paradise fish, Macropodus opercularis (KJ408271) (pairwise distance: 92.5%) from China, then followed by the strain from Spain (GU320726 and GU320736) (pairwise distance: 90.8-90.5%), Korea (AB299163, AB212999, AB213004, and AB299164) (pairwise distance: 91.5-80.5%) and lastly Canada (GU939626) (pairwise distance: 83%). This is the first report of lymphocystis disease in A. clarkii in Taiwan.

New Insights into Lymphocystis Disease Virus Genome Diversity.

Lymphocystis disease viruses (LCDVs) are viruses that infect bony fish which has been found in different locations across the globe. Four virus species have been classified by the International Committee on Taxonomy of Viruses (ICTV), despite remarkable discrepancies in genome size. Whole genome sequencing and phylogenetic analysis of LCDVs from wild fish from the North Sea and partial sequences from gilthead sea bream of an aquafarm located in the Aegean Sea in Turkey confirm that the LCDV1 genome at 100 kb is approximately half the size of the genomes of LCDV2-4. Since the fish species, of which LCDV1 was isolated, differ taxonomically at the order level, co-speciation can be excluded as the driver of the adaptation of the genome of this nucleocytoplasmic large DNA virus, but may represent an adaptation to the lifestyle of this demersal fish in the northeast Atlantic.

A New Cell Line Derived from the Spleen of the Japanese Flounder (Paralichthys olivaceus) and Its Application in Viral Study.

A new cell line Japanese flounder spleen (JFSP) derived from the spleen of Japanese flounder (Paralichthys olivaceus) was established and characterized in this study. The JFSP cells grew rapidly at 29 °C, and the optimum fetal bovine serum concentration in the L-15 medium was 15%. Cells were subcultured for more than 80 passages. The JFSP cells have a diploid chromosome number of 2n = 68, which differs from the chromosome number of normal diploid Japanese flounder. The established cells were susceptible to Bohle virus (BIV), Viral hemorrhagic septicemia virus (VHSV), Hirame rhabdovirus (HIRRV), Infectious hematopoietic necrosis virus (IHNV), and Lymphocystis disease virus (LCDV), as evidenced by varying degrees of cytopathic effects (CPE). Replication of the virus in JFSP cells was confirmed by qRT-PCR and transmission electron microscopy. In addition, the expression of four immune-related genes, TRAF3, IL-1β, TNF-α, and TLR2, was differentially altered following viral infection. The results indicated that the cells underwent an antiviral immune response. JFSP cell line is an ideal tool in vitro for virology. The use of fish cell lines to study the immune genes and immune mechanism of fish and to clarify the immune mechanism of fish has important theoretical significance and practical application value for the fundamental prevention and treatment of fish diseases.

Interaction of a viral insulin-like peptide with the IGF-1 receptor produces a natural antagonist

Lymphocystis disease virus-1 (LCDV-1) and several other Iridoviridae encode viral insulin/IGF-1 like peptides (VILPs) with high homology to human insulin and IGFs. Here we show that while single-chain (sc) and double-chain (dc) LCDV1-VILPs have very low affinity for the insulin receptor, scLCDV1-VILP has high affinity for IGF1R where it can antagonize human IGF-1 signaling, without altering insulin signaling. Consequently, scLCDV1-VILP inhibits IGF-1 induced cell proliferation and growth hormone/IGF-1 induced growth of mice in vivo. Cryo-electron microscopy reveals that scLCDV1-VILP engages IGF1R in a unique manner, inducing changes in IGF1R conformation that led to separation, rather than juxtaposition, of the transmembrane segments and hence inactivation of the receptor. Thus, scLCDV1-VILP is a natural peptide with specific antagonist properties on IGF1R signaling and may provide a new tool to guide development of hormonal analogues to treat cancers or metabolic disorders sensitive to IGF-1 without affecting glucose metabolism.

Molecular characterisation of Lymphocystis Disease Virus detected from Sea Bream in Turkey

Lymphocystis disease is a common, chronic and benign infection of fish. The disease has a worldwide distribution and has been reported from more than 125 marine and freshwater sites. In this study, molecular diagnosis and characterisation of Lymphocystis Disease Virus were performed from samples collected from sea bream fry showing lymphocystis disease symptoms in a farm located in the Aegean Sea. As a result of the phylogenetic analysis, the virus was classified as genotype VII. According to the similarity of nucleotide sequence, SA1.ETun.2011 virus reported from Tunisia was the closest related virus. The rate of amino acid substitution between genotypes was quite high, while amongst the genotype VII isolates substitution rates were low. When the similarity between the genotypes was investigated using the nucleotide sequences, the closest and most distant genotypes to the genotype VII were V and I, respectively.

Detection of lymphocystis disease virus (LCDV) from yellowbar angelfish (Pomacanthus maculosus Forsskål, 1775) in Iran: Histopathological and phylogenetic analysis

Lymphocystis disease (LCD) is a benign viral disease characterized by the appearance of hypertrophic fibroblastic cells in the connective tissues of fish. In December 2021, a group of yellowbar angelfish (Pomacanthus maculosus Forsskål, 1775) which had significant mortalities (10%) and showed whitish nodules on the fins and skin, were packed in water-filled plastic bags, supplied with oxygen, and transported to the Ornamental Fish Clinic, Faculty of Veterinary Medicine, University of Tehran (Tehran, Iran). Following the clinical inspection and observation of whitish clusters of nodules mostly on the fins and skin by stereomicroscopy, wet smears of skin and fins revealed numerous hypertrophic lymphocystis cells. Histopathological study of the whitish nodules in skin and fins revealed many hypertrophic cells with thick hyaline capsules, irregular nucleus, and multiple intracytoplasmic inclusion bodies. In addition to the extension of hypertrophic lymphocystis cells' growth until the superficial muscle layer, infiltration of inflammatory cells and hemorrhage between hypertrophic cells are also observed. The hyaline capsule of hypertrophic fibroblasts stained with the PAS. Molecular characterization (PCR) confirmed the presence of Lymphocystis disease virus (LCDV) in the external nodules and the internal organs homogenate of the infected fish. Sequence analysis of the MCP gene of the detected virus (IR_LCDV_RGZIS) assigned it to genogroup 5. This is the first detection and characterization of LCDV in yellowbar angelfish (Pomacanthus maculosus) in Iran.

Establishment and characterization of a cell line from the brain of the Japanese flounder (Paralichthys olivaceus) and its application in the study of viral infection

Fish cell lines are important tools in many biological studies. Here, we established a new Japanese flounder brain (JFB) cell line from the brain tissue of the Japanese flounder (Paralichthys olivaceus), which was successfully sub-cultured for over 120 passages, and consisted predominantly of fibroblast-like cells. Sequencing of the actin gene confirmed that JFB cells were derived from the Japanese flounder, and JFB was also identified as a neural stem cell line on the basis of the mRNA expression of nestin gene. The JFB cells could grow at a temperature ranging from 17 to 29 °C, and 23 °C was the optimum temperature. Karyotype analysis showed that the chromosome mode number was 48, demonstrating a normal diploid chromosome number. The transfection efficiency of pEGFP-N1 in JFB cells was up to 30%. The JFB cells showed susceptibility to Hirame rhabdovirus (HIRRV), Lymphocystis disease virus (LCDV), Bohle virus (BIV) and Chinese giant salamander iridovirus (GSIV), which was demonstrated by varying degrees of cytopathic effects. Viral infection or Poly (I:C) stimulation upregulated the expression of multiple immune-related genes, including IL-1β, IL8, Mx and TNF. These results showed that the newly established JFB cell line is an ideal tool for studying gene manipulation, host-virus interaction and potential vaccines development.

Peripheral Blood B-Lymphocytes Are Involved in Lymphocystis Disease Virus Infection in Flounder (Paralichthys olivaceus) via Cellular Receptor-Mediated Mechanism.

Previous studies imply that peripheral blood leukocytes (PBLs) may play an important role in systemic lymphocystis disease virus (LCDV) dissemination, but whether the PBLs are susceptible and permissive to LCDV infection and the dissemination mechanism need to be clarified. In this study, LCDV was firstly confirmed to infect the PBLs in flounder (Paralichthys olivaceus) in vivo, and to replicate in PBLs in vitro. Subsequently, the 27.8 kDa receptor protein (27.8R), a functional receptor mediating LCDV infection in flounder gill cells, was shown to locate on the cell membrane of PBLs and co-localize with LCDV in PBLs, while blocking of the 27.8R via pre-incubation of anti-27.8R MAb with the PBLs could obviously inhibit LCDV infection, revealing the 27.8R as a receptor for LCDV entry into PBLs. Multicolor fluorescence imaging studies verified that IgM+ and IgD+ B-lymphocyte were involved in LCDV infection. In the sorted IgM+ B-cells, 27.8R+ and LCDV+ signals were simultaneously observed, and LCDV copy numbers increased with time, indicating that IgM+ B-cells expressed the 27.8R and were permissive to LCDV infection. Furthermore, the dynamic changes of IgM+, 27.8R+, LCDV+ and LCDV+/IgM+ PBLs were monitored during the early phase of LCDV infection. It was found that the percentage of IgM+ B-cells in PBLs clearly declined first and then increased, suggesting LCDV infection facilitated damage to B-cells, whereas the amounts of 27.8R+ and LCDV+ PBLs, as well as LCDV-infected IgM+ B-cells, showed an opposite trend. These results proved that IgM+ B-lymphocytes could be infected by LCDV via a receptor-mediated mechanism and support viral replication, which provided novel insights for the first time into the role of B-lymphocytes in LCDV dissemination and pathogenesis in teleost fish.