Evaluation of protective immune response of immersion inactivated vaccine against Singapore grouper iridovirus.

Singapore grouper iridovirus (SGIV) always causes high transmission efficiency and mortality in the larval and juvenile stages of grouper in aquaculture industry. Although inactivated virus and recombinant DNA vaccines administered via intraperitoneal injection have shown efficacy in protection against SGIV, their potential applications in field testing were limited due to the vaccine delivery methods. Here, we developed an immersion vaccine containing inactivated virus and Montanide IMS 1312 adjuvant (IMS 1312) and evaluated its protective efficacy against SGIV infection. Compared to the PBS group, fish vaccinated with immersion inactivated vaccine with or without IMS 1312 were significantly protected against SGIV, with a relative percent survival (RPS) of 57.69 % and 38.47 %, respectively. Furthermore, the transcripts of viral core genes were reduced, and the histopathological severity caused by SGIV were relatively mild in multiple tissues of the IMS + V group. The immersion vaccine activated the AKP and ACP activities and increased the mRNA levels of IFN and inflammation-associated genes. The transcriptome analysis showed that a total of 731 and 492 genes were significantly regulated in the spleen and kidney from the IMS + V group compared to the PBS group, respectively. Among them, 129 DEGs were co-regulated, and enriched in the KEGG pathways related to immune and cell proliferation, including MAPK signaling, JAK-STAT signaling and PI3K-Akt signaling pathways. Similarly, the DEGs specially regulated in the kidney and spleen upon vaccine immunization were significantly enriched in the KEGG pathways related to interferon and inflammation response. Together, our results elucidated that the immersion vaccine of inactivated SGIV with IMS 1312 induced a protective immune response of grouper against SGIV.

Mandarin fish von Hippel-Lindau protein regulates the NF-κB signaling pathway via interaction with IκB to promote fish ranavirus replication.

The von Hippel-Lindau tumor suppressor protein (VHL), an E3 ubiquitin ligase, functions as a critical regulator of the oxygen-sensing pathway for targeting hypoxia-inducible factors. Recent evidence suggests that mammalian VHL may also be critical to the NF-κB signaling pathway, although the specific molecular mechanisms remain unclear. Herein, the roles of mandarin fish ( Siniperca chuatsi) VHL ( scVHL) in the NF-κB signaling pathway and mandarin fish ranavirus (MRV) replication were explored. The transcription of scVHL was induced by immune stimulation and MRV infection, indicating a potential role in innate immunity. Dual-luciferase reporter gene assays and reverse transcription quantitative PCR (RT-qPCR) results demonstrated that scVHL evoked and positively regulated the NF-κB signaling pathway. Treatment with NF-κB signaling pathway inhibitors indicated that the role of scVHL may be mediated through scIKKα, scIKKß, scIκBα, or scp65. Co-immunoprecipitation (Co-IP) analysis identified scIκBα as a novel target protein of scVHL. Moreover, scVHL targeted scIκBα to catalyze the formation of K63-linked polyubiquitin chains to activate the NF-κB signaling pathway. Following MRV infection, NF-κB signaling remained activated, which, in turn, promoted MRV replication. These findings suggest that scVHL not only positively regulates NF-κB but also significantly enhances MRV replication. This study reveals a novel function of scVHL in NF-κB signaling and viral infection in fish.

Singapore grouper iridovirus VP128 inhibits STING-TBK1 mediated signaling to evade antiviral immunity.

Singapore grouper iridovirus (SGIV) belongs to the family Iridoviridae and the genus Ranavirus, which is a large cytoplasmic DNA virus. Infection of grouper with SGIV can cause hemorrhage and swelling of the spleen of the fish. Previous work on genome annotation demonstrated that SGIV contained numerous uncharacterized or hypothetical open reading frames (ORFs), whose functions remained largely unknown. In the present study, the protein encoded by SGIV ORF128 (VP128) was identified. VP128 is predominantly localized within the endoplasmic reticulum (ER). Overexpression of VP128 significantly promoted SGIV replication. VP128 inhibited the interferon (IFN)-3 promoter activity and mRNA level of IFN-related genes induced by poly(I:C), Epinephelus coioides cyclic GMP/AMP synthase (EccGAS)/stimulator of IFN genes (EcSTING), and TANK-binding kinase 1 (EcTBK1). Moreover, VP128 interacted with EcSTING and EcTBK1. The interaction between VP128 and EcSTING was independent of any specific structural domain of EcSTING. Together, our results demonstrated that SGIV VP128 negatively regulated the IFN response by inhibiting EcSTING-EcTBK1 signaling for viral evasion.

Epinephelus coioides Sec3 promotes Singapore grouper iridovirus infection by negatively regulates immune response.

Exocyst, a protein complex, plays a crucial role in various cellular functions, including cell polarization, migration, invasion, cytokinesis, and autophagy. Sec3, known as Exoc1, is a key subunit of the Exocyst complex and can be involved in cell survival and apoptosis. In this study, two subtypes of Sec3 were isolated from Epinephelus coioides, an important marine fish in China. The role of E. coioides Sec3 was explored during Singapore grouper iridovirus (SGIV) infection, an important pathogen of marine fish which could induce 90 % mortality. E. coioides Sec3 sequences showed a high similarity with that from other species, indicating the presence of a conserved Sec3 superfamily domain. E. coioides Sec3 mRNA could be detected in all examined tissues, albeit at varying expression levels. SGIV infection could upregulate E. coioides Sec3 mRNA. Upregulated Sec3 significantly promoted SGIV-induced CPE, and the expressions of viral key genes. E. coioides Sec3 could inhibit the activation of NF-κB and AP-1, as well as SGIV-induced cell apoptosis. The results illustrated that E. coioides Sec3 promotes SGIV infection by regulating the innate immune response.

In vitro antiviral activity of eugenol on Singapore grouper iridovirus.

The high mortality rate of Singapore grouper iridovirus (SGIV) posing a serious threat to the grouper aquaculture industry and causing significant economic losses. Therefore, finding effective drugs against SGIV is of great significance. Eugenol (C10H12O2) is a phenolic aromatic compound, has been widely studied for its anti-inflammatory, antioxidant and antiviral capacity. In this study, we explored the effect of eugenol on SGIV infection and its possible mechanisms using grouper spleen cells (GS) as an in vitro model. We found that treatment of GS cells with 100 µM eugenol for 4 h exhibited the optimal inhibitory effect on SGIV. Eugenol was able to reduce the expression level of inflammatory factors by inhibiting the activation of MAPK pathway and also inhibited the activity of NF-κB and AP-1 promoter. On the other hand, eugenol attenuated cellular oxidative stress by reducing intracellular ROS and promoted the expression of interferon-related genes. Therefore, we conclude that eugenol inhibits SGIV infection by enhancing cellular immunity through its anti-inflammatory and antioxidant functions.

Ingestion of polyethylene terephthalate microplastic water contaminants by Xenopus laevis tadpoles negatively affects their resistance to ranavirus infection and antiviral immunity.

Small plastic debris (0.1 µm-5 mm) or microplastics (MPs) have become major pollutants of aquatic ecosystems worldwide and studies suggest that MPs exposure can pose serious threats to human and wildlife health. However, to date the potential biological impacts of MPs accumulating in low amount in tissues during early life remains unclear. Here, for a more realistic assessment, we have used environmentally representative, mildly weathered, polyethylene terephthalate microplastics (PET MPs), cryomilled (1-100 µm) and fluorescently labelled. We leveraged the amphibian Xenopus laevis tadpoles as an animal model to define the biodistribution of PET MPs and determine whether exposure to PET MPs induce perturbations of antiviral immunity. Exposure to PET MPs for 1-14 days resulted in detectable PET MPs biodistribution in intestine, gills, liver, and kidney as determined by fluorescence microscopy on whole mount tissues. PET MPs accumulation rate in tissues was further evaluated via a novel in situ enzymatic digestion and subsequent filtration using silicon nanomembranes, which shows that PET MPs rapidly accumulate in tadpole intestine, liver and kidneys and persist over a week. Longer exposure (1 month) of tadpoles to relatively low concentration of PET MPs (25 µg/ml) significantly increased susceptibility to viral infection and altered innate antiviral immunity without inducing overt inflammation. This study provides evidence that exposure to MPs negatively impact immune defenses of aquatic vertebrates.

Cytokinins Reduce Viral Replication and Alter Plaque Morphology of Frog Virus 3 In Vitro.

Cytokinins (CKs) are a group of N6-substituted signaling molecules whose biosynthesis and metabolism have been documented in all kingdoms of life, including vertebrates. While their biological relevance in vertebrate systems continues to be elucidated, they have broadly been documented with therapeutic effects in exogenous applications. In this study, we evaluated the virostatic potential of four types of CKs including, N6-isopentenyladenine (iP), N6-isopentenyladenosine (iPR), N6-isopentenyladenosine-5'monophosphate (iPMP), and 2-methylthiol-N6-isopentenyladenosine (2MeSiPR) against the ranavirus type species, frog virus 3 (FV3). Following concurrent treatment and infection, iP and iPR reduced viral replication by 33.8% and 59.6%, respectively, in plaque formation assays. A decrease in viral replication was also observed when CK exposure was limited to 12 h prior to infection, where iP and iPR reduced viral replication by 31% and 23.75%, respectively. Treatment with iP and iPR was also marked by 48% and 60% decreases in viral load over 72 h, respectively, as measured in single step growth curves. Plaque morphology was altered in vitro, as iP and iPR treatment increased plaque area by 83% and 112% with lytic zone formation also becoming more prevalent in corresponding treatments. Treatment with iPMP and 2MeSiPR resulted in no effect on viral kinetics in vitro. The results of this study are the first to provide evidence of CK antiviral activity against a DNA virus and highlight the importance of their structure for therapeutic investigations.

Identification and functional characterization of interleukin-22 (IL-22) in orange-spotted grouper (Epinephelus coioides).

In mammals, IL-22 is considered as a critical cytokine regulating of immunity and homeostasis at barrier surfaces. Although IL-22 have been functional characterization in different species of fish, the studies about distinct responses of IL-22 in different organs/tissues/cell types is rather limited. Here, we identified and cloned IL-22 gene (named as Ec-IL-22) from grouper (Epinephelus coioides). Ec-IL-22 gene was detected in all orangs/tissues examined, and was induced in intestine, gill, spleen, head kidney, and primary head kidney/intestine leukocytes following the stimulation of LPS and poly (I:C), as well as Vibrio harveyi and Singapore grouper iridovirus infection (SGIV). In addition, the stimulation of DSS could induce the expression of Ec-IL-22 in intestine and primary leukocytes from intestine. Importantly, the treatment of recombinant Ec-IL-22 induced the mRNA level of proinflammatory cytokines in primary intestine/head kidney leukocytes. The present results improve the understanding of expression patterns and functional characteristics of fish IL-22 in different organs/tissues/cell types.

SGIV VP82 inhibits the interferon response by degradation of IRF3 and IRF7.

During virus-host co-evolution, viruses have developed multiple strategies to dampen IFN response and prevent its antiviral activity in host cells. To date, the interactions between host IFN response and the immune evasion strategies exploited by fish iridoviruses still remain largely uncertain. Here, a potential immune evasion protein candidate of Singapore grouper iridovirus (SGIV), VP82 (encoded by SGIV ORF82) was screened and its roles during viral replication were investigated in detail. Firstly, VP82 overexpression dramatically decreased IFN or ISRE promoter activity and the transcription levels of IFN stimulated genes (ISGs) stimulated by grouper cyclic GMP-AMP synthase (EccGAS)/stimulator of interferon genes (EcSTING), TANK-binding kinase 1 (EcTBK1), IFN regulatory factor 3 (EcIRF3)and EcIRF7. Secondly, Co-IP assays indicated that VP82 interacted with EcIRF3 and EcIRF7, but not EcSTING and EcTBK1, which was consistent with the co-localization between VP82 and EcIRF3 or EcIRF7. Furthermore, VP82 promoted the degradation of EcIRF3 and EcIRF7 in a dose-dependent manner via the autophagy pathway. Finally, VP82 overexpression accelerated SGIV replication, evidenced by the increased transcriptions of viral core genes and viral production. Moreover, the antiviral action of EcIRF3 or EcIRF7 was significantly depressed in VP82 overexpressed cells. Together, VP82 was speculated to exert crucial roles for SGIV replication by inhibiting the IFN response via the degradation of IRF3 and IRF7. Our findings provided new insights into understanding the immune evasion strategies utilized by fish iridovirus through IFN regulation.

Study of the antivirus function mediated by STING in Micropterus salmoides.

Stimulator of interferon genes (STING) has been demonstrated as a critical mediator in the innate immune response to cytosolic DNA and RNA derived from different pathogens. While the role of Micropterus salmoides STING (MsSTING) in largemouth bass virus is still unknown. In this study, RT-qPCR assay and Western-blot assay showed that the expression levels of MsSTING and its downstream genes were up-regulated after LMBV infection. Pull down experiment proved that a small peptide called Fusion peptide (FP) that previously reported to target to marine and human STING as a selective inhibitor also interacted with MsSTING in vitro. Comparing with the RNA-seq of Largemouth bass infected with LMBV singly, 326 genes were significantly up-regulated and 379 genes were significantly down-regulated in the FP plus LMBV group in which Largemouth bass was treatment with FP before LMBV-challenged. KEGG analysis indicated that the differentially expressed genes (DEGs) were mainly related to signaling transduction, infectious disease viral, immune system and endocrine system. Besides, the survival rate of LMBV-infected largemouth bass was highly decreased following FP treatment. Taken together, our study showed that MsSTING played an important role in immune response against LMBV infection.

Berberine inhibits SGIV replication by suppressing inflammatory response and oxidative stress.

Singapore grouper iridovirus (SGIV) is one of the major infectious diseases responsible for high mortality and huge economic losses in the grouper aquaculture industry. Berberine (BBR), a naturally occurring plant alkaloid, is a phytochemical having a variety of biological properties, such as antiviral, antioxidant, and anti-inflammatory effects. In this work, we used an in vitro model based on Western blot, ROS fluorescence probe, and real-time quantitative PCR (qRT-PCR) to examine the antiviral qualities of BBR against SGIV. The outcomes demonstrated that varying BBR concentrations could significantly inhibit the replication of SGIV. In addition, BBR greatly inhibited the production of genes associated with pro-inflammatory cytokines in SGIV-infected or SGIV-uninfected GS cells based on qRT-PCR data. Subsequent investigations demonstrated that BBR suppressed the expression of the promoter activity of NF-κB and NF-κB-p65 protein. Additionally, BBR reduced the phosphorylation of ERK 1/2, JNK, and p38. Furthermore, BBR also inhibits SGIV-induced ROS production by upregulating the expression of antioxidant-related genes. In conclusion, BBR is a viable therapy option for SGIV infection due to its antiviral properties.

The antiviral role of largemouth bass STING against iridovirus infection.

Stimulator of interferon gene (STING) plays a crucial role in the innate immune response against viral and bacterial pathogens. However, its function in largemouth bass iridovirus (LMBV) infection remains uncertain. Here, a STING homolog (MsSTING) from largemouth bass (Micropterus salmoides) was cloned and characterized. MsSTING encoded a 407-amino-acid polypeptide, which shared 84.08% and 41.45% identity with golden perch (Perca flavescens) and human (Homo sapiens) homologs, respectively. MsSTING contained four transmembrane domains and a conserved C-terminal domain. The mRNA level of MsSTING was significantly increased in response to LMBV infection in vitro. Subcellular localization observation indicated that MsSTING encoded a cytoplasmic protein, which co-localized predominantly with endoplasmic reticulum (ER) and partially with mitochondria. Moreover, its accurate localization was dependent on the N-terminal transmembrane motif (TM) domains. MsSTING was able to activate interferon (IFN) response, evidenced by the activation of IFN1, IFN3 and ISRE promoters by its overexpression in vitro. Mutant analysis showed that both the N-terminal and C-terminal domain of MsSTING were essential for its activation on IFN response. In addition, overexpression of MsSTING inhibited the transcription and protein levels of viral core genes, indicating that MsSTING exerted antiviral action against LMBV. Consistently, the inhibitory effects were significantly attenuated when the N-terminal or C-terminal domains of MsSTING was deleted. Furthermore, MsSTING overexpression upregulated the transcriptions of interferon-related genes and pro-inflammatory factors, including TANK-binding kinase 1(TBK1), interferon regulatory factor 3 (IRF3), interferon regulatory factor 7 (IRF7), interferon stimulated exonuclease gene 20 (ISG20), interferon-induced transmembrane protein 1(IFITM1), interferon γ (IFN-γ), tumor necrosis factor α (TNF-α), interleukin 1ß (IL-1ß), and interleukin 6 (IL-6). Together, MsSTING exerted antiviral action upon LMBV infection through positive regulation the innate immune response.

Singapore grouper iridovirus VP20 interacts with grouper TBK1 and IRF3 to attenuate the interferon immune response.

Singapore grouper iridovirus (SGIV), belonging to genus Ranavirus, family Iridoviridae, is a highly pathogenic agent and causes heavy economic losses in the global grouper aquaculture. Recent studies demonstrated that SGIV infection attenuated antiviral immune and inflammatory response induced by poly (I:C) in vitro. However, little was known about the potential functions of the immune regulatory proteins encoded by SGIV. Here, we identified the detailed roles of VP20 and clarified the potential mechanism underlying its immune regulatory function during SGIV infection. Our results showed that VP20 was an IE gene, and partially co-localized with Golgi apparatus and lysosomes in grouper cells. Overexpression of VP20 enhanced SGIV replication, demonstrated by the increase in the transcription levels of viral core genes and the protein synthesis of MCP. Reporter gene assays showed that SGIV VP20 overexpression significantly reduced the IFN promoter activity induced by poly (I:C), grouper stimulator of interferon genes (EcSTING) and TANK-binding kinase 1 (EcTBK1). Consistently, the transcription levels of IFN related genes were significantly decreased in VP20 overexpressing cells compared to those in control cells. Co-IP assay and confocal microscopy observations indicated that VP20 co-localized and interacted with EcTBK1 and EcIRF3, but not EcSTING. In addition, VP20 was able to degrade EcIRF3 and attenuate the antiviral action of EcIRF3, while had no effect on EcTBK1. Together, SGIV VP20 was speculated to promote viral replication through attenuating the IFN response mediated by TBK1-IRF3 in vitro. Our findings provided new insights into the immune regulatory function of SGIV encoded unknown proteins.

Antiviral Effect and Mechanism of Edaravone against Grouper Iridovirus Infection.

Singapore grouper iridovirus (SGIV) is a virus with high fatality rate in the grouper culture industry. The outbreak of SGIV is often accompanied by a large number of grouper deaths, which has a great impact on the economy. Therefore, it is of great significance to find effective drugs against SGIV. It has been reported that edaravone is a broad-spectrum antiviral drug, most widely used clinically in recent years, but no report has been found exploring the effect of edaravone on SGIV infections. In this study, we evaluated the antiviral effect of edaravone against SGIV, and the anti-SGIV mechanism of edaravone was also explored. It was found that the safe concentration of edaravone on grouper spleen (GS) cells was 50 µg/mL, and it possessed antiviral activity against SGIV infection in a dose-dependent manner. Furthermore, edaravone could significantly disrupt SGIV particles and interference with SGIV binding to host cells, as well as SGIV replication in host cells. However, edaravone was not effective during the SGIV invasion into host cells. This study was the first time that it was determined that edaravone could exert antiviral effects in response to SGIV infection by directly interfering with the processes of SGIV infecting cells, aiming to provide a theoretical basis for the control of grouper virus disease.

Largemouth bass Rel exerts antiviral role against fish virus and regulates the expression of interleukin-10.

Nuclear factor-κB (NF-κB)/Rel is a group of transcription factors that can be activated and regulates various aspects of innate and adaptive immune functions, which play a crucial role in mediating inflammatory responses. Interleukin-10 (IL-10) is a highly pleiotropic cytokine that has a central role in limiting the immune response to pathogens during infection and thereby alleviating damage to the host. This study aims to investigate the function of the Rel gene in virus infection and its regulatory effect on IL-10 in the largemouth bass (Micropterus salmoides). The ORF sequence of MsRel was 1941 bp, containing 646 amino acids with two conserved functional domains, including RHD and IPT domain. In healthy largemouth bass, the mRNA of MsRel was detected in all the tested tissues, including gill, liver, kidney, heart, spleen, intestine, stomach, skin, brain, fin and muscle. The expression of MsRel was induced by challenge with largemouth bass virus (LMBV) or red grouper nervous necrosis virus (RGNNV), as well as treatment with lipopolysaccharide (LPS) or poly (I:C) in vivo. As evidenced by the detection of viral gene mRNA levels, the infectivity of LMBV and morphological cytopathic effect (CPE), we found that overexpression of MsRel inhibited the infection and replication of LMBV, suggesting its antiviral roles in fish. Besides, the promoter analysis was carried out to determine whether MsRel was a regulator of MsIL-10. The results of the luciferase reporter assay indicated that MsRel has a positive regulatory role in MsIL-10 expression. Further analysis revealed that the potential binding sites of MsIL-10 may be located in the MsIL10-5-M (-42 to +8 bp) region of the MsIL-10 promoter. Furthermore, we observed that MsRel enhanced IFN-I and IFN-III promoter activities. Taken together, our findings demonstrated that MsRel affect LMBV infection by regulating the immune responses, and providing a new idea of the mechanisms how Rel regulate the expression of IL-10 in bony fish.

Antiviral role of grouper FoxO1 against RGNNV and SGIV infection.

As a key regulator of the innate immune system, FoxO1 has a variety of activities in biological organisms. In the present study, grouper FoxO1 (EcFoxO1) was cloned and the antiviral activity in red grouper neuron necrosis virus (RGNNV) and Singapore grouper iridescent virus (SGIV) was examined. The open reading frame (ORF) of EcFoxO1 contains 2,034 base pairs that encode a protein of 677 amino acids with a predicted molecular weight of 73.21 kDa. EcFoxO1 was shown to be broadly distributed in healthy grouper tissues, and was up-regulated in vitro in response to stimulation by RGNNV and SGIV. EcFoxO1 has a whole-cell distribution in grouper spleen (GS) cells. EcFoxO1 decreased the replication of RGNNV and SGIV, and activated interferon (IFN) 3, IFN-stimulated response element (ISRE), and nuclear factor-κB (NF-κB) promoter activities. EcFoxO1 could interact with EcIRF3. Together, the results demonstrated that EcFoxO1 might be an important regulator of grouper innate immune response against RGNNV and SGIV infection.

Grouper Atg14 promotes Singapore grouper iridovirus (SGIV) replication by inhibiting the host innate immune response.

As one of the important members of the autophagy-related protein family, Atg14 plays a key role in the formation and maturation of autophagosomes. However, little is known about the potential roles of fish Atg14 and its roles in virus infection. In the present study, the homolog of Atg14 (EcAtg14) from the orange-spotted grouper (Epinephelus coioides) was cloned and characterized. The open reading frame (ORF) of EcAtg14 consists of 1530 nucleotides, encoding 509 amino acids, with a predicted molecular weight of 56.9 kDa. EcAtg14 was distributed in all tested tissues, with higher expression in liver, blood and spleen. The expression of EcAtg14 was increased in grouper spleen (GS) cells after Singapore grouper iridovirus (SGIV) infection. EcAtg14 was distributed in the cytoplasm of GS cells. Overexpression of EcAtg14 promoted SGIV replication in GS cells and inhibited IFN3, ISRE and NF-κB promoter activities. Co-immunoprecipitation results showed that there was an interaction between EcAtg14 and EcBeclin. EcAtg14 also promoted the synthesis of LC3-II in GS cells. These findings provide a basis for understanding the innate immune mechanism of grouper against viral infection.

Singapore grouper iridovirus VP122 targets grouper STING to evade the interferon immune response.

Singapore grouper iridovirus (SGIV) is a highly pathogenic Iridoviridae that causes hemorrhage and spleen enlargement in grouper. Despite previous genome annotation efforts, many open reading frames (ORFs) in SGIV remain uncharacterized, with largely unknown functions. In this study, we identified the protein encoded by SGIV ORF122, now referred to as VP122. Notably, overexpression of VP122 promoted SGIV replication. Moreover, VP122 exhibited antagonistic effects on the natural antiviral immune response through the cGAS-STING signaling pathway. It specifically inhibited the cGAS-STING-triggered transcription of various immune-related genes, including IFN1, IFN2, ISG15, ISG56, PKR, and TNF-α in GS cells. Additionally, VP122 significantly inhibited the activation of the ISRE promoter mediated by EccGAS and EcSTING but had no effect on EccGAS or EcSTING alone. Immunoprecipitation and Western blotting experiments revealed that VP122 specifically interacts with EcSTING but not EccGAS. Notably, this interaction between VP122 and EcSTING was independent of any specific domain of EcSTING. Furthermore, VP122 inhibited the self-interaction of EcSTING. Interestingly, VP122 did not affect the recruitment of EcTBK1 and EcIRF3 to the EcSTING complex. Collectively, our results demonstrate that SGIV VP122 targets EcSTING to evade the type I interferon immune response, revealing a crucial role for VP122 in modulating the host-virus interaction.

Widespread amphibian Perkinsea infections associated with Ranidae hosts, cooler months and Ranavirus co-infection.

Amphibians suffer from large-scale population declines globally, and emerging infectious diseases contribute heavily to these declines. Amphibian Perkinsea (Pr) is a worldwide anuran pathogen associated with mass mortality events, yet little is known about its epidemiological patterns, especially in comparison to the body of literature on amphibian chytridiomycosis and ranavirosis. Here, we establish Pr infection patterns in natural anuran populations and identify important covariates including climate, host attributes and co-infection with Ranavirus (Rv). We used quantitative (q)PCR to determine the presence and intensity of Pr and Rv across 1234 individuals sampled throughout central Florida in 2017-2019. We then implemented random forest ensemble learning models to predict infection with both pathogens based on physiological and environmental characteristics. Perkinsea infected 32% of all sampled anurans, and Pr prevalence was significantly elevated in Ranidae frogs, cooler months, metamorphosed individuals and frogs co-infected with Rv, while Pr intensity was significantly higher in ranid frogs and individuals collected dead. Ranavirus prevalence was 17% overall and was significantly higher in Ranidae frogs, metamorphosed individuals, locations with higher average temperatures, and individuals co-infected with Pr. Perkinsea prevalence was significantly higher than Rv prevalence across months, regions, life stages and species. Among locations, Pr prevalence was negatively associated with crayfish prevalence and positively associated with relative abundance of microhylids, but Rv prevalence did not associate with any tested co-variates. Co-infections were significantly more common than single infections for both pathogens, and we propose that Pr infections may propel Rv infections because seasonal Rv infection peaks followed Pr infection peaks and random forest models found Pr intensity was a leading factor explaining Rv infections. Our study elucidates epidemiological patterns of Pr in Florida and suggests that Pr may be under-recognized as a cause of anuran declines, especially in the context of pathogen co-infection.

Oral immunizations with Bacillus subtilis spores displaying VP19 protein provide protection against Singapore grouper iridovirus (SGIV) infection in grouper.

Disease caused by Singapore grouper iridovirus (SGIV) results in major economic losses in the global grouper aquaculture industry. Vaccination is considered to be the most effective way to protect grouper from SGIV. In this study, the spores of Bacillus subtilis (B.subtilis) WB600 were utilized as the vehicle that the VP19 protein was displayed on the spores surface. To further investigate the effect of oral vaccination, the grouper were orally immunized with B.s-CotC-19 spores. After challenged, the survival rate of grouper orally vaccinated with B.s-CotC-19 spores was 34.5% and the relative percent survival (RPS) was 28.7% compared to the PBS group. Moreover, the viral load in the tissues of the B.s-CotC-19 group was significantly lower than that of the PBS group. The histopathological sections of head kidney and liver tissue from the B.s-CotC-19 group showed significantly less histopathology compared to the PBS group. In addition, the specific IgM levels in serum in the B.s-CotC-19 group was higher than those in the PBS group. In the hindgut tissue, the immune-related gene expression detected by quantitative real-time PCR (qRT-PCR) exhibited an increasing trend in different degrees in the B.s-CotC-19 group, suggesting that the innate and adaptive immune responses were activated. These results indicated that the oral administration of recombinant B.subtilis spores was effective for preventing SGIV infection. This study provided a feasible strategy for the controlling of fish virus diseases.