• Journal of Veterinary Science
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• v.22 no.5
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• pp.73.1-73.11
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• 2021

Background: Feline calicivirus (FCV) is a common pathogen of felids, and FCV vaccination is regularly practiced. The genetic variability and antigenic diversity of FCV hinder the effective control and prevention of infection by vaccination. Improved knowledge of the epidemiological characteristics of FCV should assist in the development of more effective vaccines. Objectives: This study aims to determine the prevalence of FCV in a population of cats with FCV-suspected clinical signs in Hangzhou and to demonstrate the antigenic and genetic relationships between vaccine status and representative isolated FCV strains. Methods: Cats (n = 516) from Hangzhou were investigated between 2018 and 2020. The association between risk factors and FCV infection was assessed. Phylogenetic analyses based on a capsid coding sequence were performed to identify the genetic relationships between strains. In vitro virus neutralization tests were used to assess antibody levels against isolated FCV strains in client-owned cats. Results: The FCV-positive rate of the examined cats was 43.0%. Risk factors significantly associated with FCV infection were vaccination status and oral symptoms. Phylogenetic analysis revealed a radial phylogeny with no evidence of temporal or countrywide clusters. There was a significant difference in the distribution of serum antibody titers between vaccinated and unvaccinated cats. Conclusions: This study revealed a high prevalence and genetic diversity of FCV in Hangzhou. The results indicate that the efficacy of FCV vaccination is unsatisfactory. More comprehensive and refined vaccination protocols are an urgent and unmet need.

• Korean Journal of Pharmacognosy
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• v.49 no.4
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• pp.291-297
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• 2018

Coxsackievirus B3 (CVB3) is a very well-known causative agent for viral myocarditis and meningitis in human. However, the effective vaccine and therapeutic drug are not developed yet. CVB3 infection activates host cell AKT signaling. Inhibition of AKT signaling pathway may attenuate CVB3 replication and prevent CVB3-mediate viral myocarditis. In this study, we determined antiviral effect of the selected natural plant extract to develop a therapeutic drug for CVB3 treatment. We screened several chemically extracted natural compounds by using HeLa cell-based cell survival assay. Among them, Linum usitatissimum L. extract was selected for antiviral drug candidate. L. usitatissimum extract significantly decreased CVB3 replication and cell death in CVB3 infected HeLa cells with no cytotoxicity. CVB3 protease 2A induced eIF4G1 cleavage and viral capsid protein VP1 production were dramatically decreased by L. usitatissimum extract treatment. In addition, virus positive and negative strand genome amplification were significantly decreased by 1 mg/ml L. usitatissimum extract treatment. Especially, L. usitatissimum extract was associated with inhibition of AKT signal and maintain mTOR activity. In contrast, Atg12 and LC3 expression were not changed by L. usitatissimum extract treatment. In this study, the potential AKT signal inhibitor, L. usitatissimum extract, was significantly inhibited viral genome replication and protein production by inhibition of AKT signal. These results suggested that L. usitatissimum extract is a novel therapeutic agent for treatment of CVB3-mediated diseases.

• Journal of Apiculture
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• v.34 no.1
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• pp.39-46
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• 2019

BQCV multi-point PCR was developed as a rapid multiplex detection method for BQCV, one of the viral pathogens of honeybees. It could detect BQCV specific genes qualitative as well as quantitative detection based on ultra-rapid PCR. Three primer pairs (RNA dependent RNA polymerase, capsid protein, 3C like protease) were specifically designed for accurate the detection and were optimized for minimizing the detection time and increasing the sensitivity. Our advanced diagnostic system have the accuracy by lowering the concern about the variation in the BQCV detection site. In addition, it should be an opportunity to identify mutations that are mixed with other viruses.

• BMB Reports
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• v.55 no.4
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• pp.175-180
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• 2022

Peptides are gaining substantial attention as therapeutics for human diseases. However, they have limitations such as low bioavailability and poor pharmacokinetics. Periostin, a matricellular protein, can stimulate the repair of ischemic tissues by promoting angiogenesis. We have previously reported that a novel angiogenic peptide (amino acids 142-151) is responsible for the pro-angiogenic activity of periostin. To improve the in vivo delivery efficiency of periostin peptide (PP), we used proteins self-assembled into a hollow cage-like structure as a drug delivery nanoplatform in the present study. The periostin peptide was genetically inserted into lumazine synthase (isolated from Aquifex aeolicus) consisting of 60 identical subunits with an icosahedral capsid architecture. The periostin peptide-bearing lumazine synthase protein cage nanoparticle with 60 periostin peptides multivalently displayed was expressed in Escherichia coli and purified to homogeneity. Next, we examined angiogenic activities of this periostin peptide-bearing lumazine synthase protein cage nanoparticle. AaLS-periostin peptide (AaLS-PP), but not AaLS, promoted migration, proliferation, and tube formation of human endothelial colony-forming cells in vitro. Intramuscular injection of PP and AaLS-PP increased blood perfusion and attenuated severe limb loss in the ischemic hindlimb. However, AaLS did not increase blood perfusion or alleviate tissue necrosis. Moreover, in vivo administration of AaLS-PP, but not AaLS, stimulated angiogenesis in the ischemic hindlimb. These results suggest that AaLS is a highly useful nanoplatform for delivering pro-angiogenic peptides such as PP.

• Fisheries and Aquatic Sciences
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• v.24 no.11
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• pp.351-359
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• 2021

The red sea bream iridovirus (RSIV) belonging to genus Megalocytivirus is responsible for red sea bream iridoviral disease (RSIVD) in marine and freshwater fishes. Although several diagnostic assays for RSIV have been developed, diagnostic sensitivity (DSe) and specificity (DSp) of real-time polymerase chain reaction (PCR) assays are not yet evaluated. In this study, we developed a TaqMan probe-based real-time PCR method and evaluated its DSe and DSp. To detect RSIV, the probe and primers were designed based on consensus sequences of the major capsid protein (MCP) genes from megalocytiviruses including RSIV, infectious spleen and kidney necrosis virus (ISKNV), and turbot reddish body iridovirus (TRBIV). The probe and primers were shown to be specific for RSIV, ISKNV, and TRBIV-types megalocytiviruses. A 95% limit of detection (LOD_95%) was determined to be 5.3 viral genome copies/µL of plasmid DNA containing the MCP gene from RSIV. The DSe and DSp of the developed real-time PCR assay for field samples (n = 112) were compared with those of conventional PCR assays and found to be 100% and 95.2%, respectively. The quantitative results for SYBR Green and TaqMan probe-based real-time PCR were not significantly different. The TaqMan probe-based real-time PCR assay for RSIV may be used as an appropriate diagnostic tool for qualitative and quantitative analysis.

• Journal of Microbiology and Biotechnology
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• v.32 no.10
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• pp.1335-1343
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• 2022

COVID-19 is an emerging disease that poses a severe threat to global public health. As such, there is an urgent demand for vaccines against SARS-CoV-2, the virus that causes COVID-19. Here, we describe a virus-like nanoparticle candidate vaccine against SARS-CoV-2 produced by an E. coli expression system. The fusion protein of a truncated ORF2-encoded protein of aa 439~608 (p170) from hepatitis E virus CCJD-517 and the receptor-binding domain of the spike protein from SARS-CoV-2 were expressed, purified and characterized. The antigenicity and immunogenicity of p170-RBD were evaluated in vitro and in Kunming mice. Our investigation revealed that p170-RBD self-assembled into approximately 24 nm virus-like particles, which could bind to serum from vaccinated people (p < 0.001) and receptors on cells. Immunization with p170-RBD induced the titer of IgG antibody vaccine increased from 14 days post-immunization and was significantly enhanced after a booster immunization at 28 dpi, ultimately reaching a peak level on 42 dpi with a titer of 4.97 log₁₀. Pseudovirus neutralization tests showed that the candidate vaccine induced a strong neutralizing antibody response in mice. In this research, we demonstrated that p170-RBD possesses strong antigenicity and immunogenicity and could be a potential candidate for use in future SARS-CoV-2 vaccine development.

• Journal of Veterinary Science
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• v.22 no.6
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• pp.84.1-84.10
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• 2021

Background: Enteritis of an infectious origin is a major cause of productivity and economic losses to cattle producers worldwide. Several pathogens are believed to cause or contribute to the development of calf diarrhea. Astroviruses (AstVs) are neglected enteric pathogens in ruminants, but they have recently gained attention because of their possible association with encephalitis in humans and various animal species, including cattle. Objectives: This paper describes a large outbreak of neonatal diarrhea in buffalo calves (Bubalus bubalis), characterized by high mortality, which was associated with an AstV infection. Methods: Following an enteritis outbreak characterized by high morbidity (100%) and mortality (46.2%) in a herd of Mediterranean buffaloes (B. bubalis) in Italy, 16 samples from buffalo calves were tested with the molecular tools for common and uncommon enteric pathogens, including AstV, kobuvirus, and torovirus. Results: The samples tested negative for common enteric viral agents, including Rotavirus A, coronavirus, calicivirus, pestivirus, kobuvirus, and torovirus, while they tested positive for AstV. Overall, 62.5% (10/16) of the samples were positive in a single round reverse transcription polymerase chain reaction (PCR) assay for AstV, and 100% (16/16) were positive when nested PCR was performed. The strains identified in the outbreak showed a clonal origin and shared the closest genetic relationship with bovine AstVs (up to 85% amino acid identity in the capsid). Conclusions: This report indicates that AstVs should be included in a differential diagnosis of infectious diarrhea in buffalo calves.

• Korean Journal of Veterinary Service
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• v.45 no.2
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• pp.87-99
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• 2022

A novel porcine circovirus 4 (PCV4) was recently emerged in Chinese and Korean pig herds, which provided epidemiological situation where three pathogenic PCVs, PCV2, PCV3, and newly emerged PCV4, could co-infect pig herds in these countries. In this study, a new triplex quantitative real-time polymerase chain reaction (tqPCR) method was developed for the rapid and differential detection of these viruses. The assay specifically amplified each viral capsid gene, whereas no other porcine pathogenic genes were detected. The detection limit of the assay was below 10 copies/µL and the assay showed high repeatability and reproducibility. In the clinical evaluation using 1476 clinical samples from 198 Korean pig farms, the detection rates of PCV2, PCV3 and PCV4 by the tqPCR assay were 13.8%, 25.4%, and 3.8%, respectively, which were 100% agreement with those of previously reported monoplex qPCR assays for PCV2, PCV3, and PCV4, with a κ value (95% CI) of 1 (1.00~1.00). The prevalence of PCV2, PCV3, and PCV4 at the farm levels were 46.5%, 63.6%, and 19.7%, respectively. The co-infection analysis for tested pig farms showed that single infection rates for PCV2, PCV3, and PCV4 were 28.8%, 44.4%, and 9.6%, respectively, the dual infection rates of PCV2 and PCV3, PCV2 and PCV4, and PCV3 and PCV4 were 12.6%, 3.5%, and 5.1%, respectively, and the triple infection rate for PCV2, PCV3, and PCV4 was 1.5%. These results demonstrate that three pathogenic PCVs are widely spread, and their co-infections are common in Korean pig herds, and the newly developed tqPCR assay will be useful for etiological and epidemiological studies of these pathogenic PCVs.

• Journal of Animal Science and Technology
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• v.65 no.4
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• pp.698-719
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• 2023

Postweaning multisystemic wasting syndrome (PMWS) is caused by a systemic inflammation after porcine circovirus type 2 (PCV2) infection. It was one of the most economically important pathogens affecting pig production worldwide before PCV2 vaccine was first introduced in 2006. After the development of a vaccine against PCV2a type, pig farms gradually restored enormous economic losses from PMWS. However, vaccine against PCV2a type could not be fully effective against several different PCV2 genotypes (PCV2b - PCV2h). In addition, PCV2a vaccine itself could generate antigenic drift of PCV2 capsid. Therefore, PCV2 infection still threats pig industry worldwide. PCV2 infection was initially found in local tissues including reproductive, respiratory, and digestive tracks. However, PCV2 infection often leads to a systemic inflammation which can cause severe immunosuppression by depleting peripheral lymphocytes in secondary lymphoid tissues. Subsequently, a secondary infection with other microorganisms can cause PMWS. Eleven putative open reading frames (ORFs) have been predicted to encode PCV2 genome. Among them, gene products of six ORFs from ORF1 to ORF6 have been identified and characterized to estimate its functional role during PCV2 infection. Acquiring knowledge about the specific interaction between each PCV2 ORF protein and host protein might be a key to develop preventive or therapeutic tools to control PCV2 infection. In this article, we reviewed current understanding of how each ORF of PCV2 manipulates host cell signaling related to immune suppression caused by PCV2.

• Journal of fish pathology
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• v.36 no.2
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• pp.213-228
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• 2023

We evaluated the transcriptional response of interferon (IFN)-related genes in rock bream iridovirus (RBIV)-infected rock bream under high-, low-, or no-mortality conditions induced by different stocking water temperatures. Under the high susceptibility condition (group A, water temperature 26℃, 100% mortality), only the Mx gene was expressed early, with prolonged expression, and with heavy viral loads of approximately 10⁶~10⁷ major capsid protein gene copies/μL from 4 to 10 days post infection (dpi). However, IRF1, IRF3, IRF8, STAT1, ISG15, PKR, Viperin, GVIN1, IFI44, and ISG56 were activated at later time points (8 dpi) and then quickly decreased (10 dpi). For the low susceptibility condition, the water temperature was set at 23℃ for 7 days (group B) and then reduced to 17℃. Group B exhibited a 28% mortality rate, in which persistent and effective antiviral responses were observed for long periods of time. In particular, at 20 and 22 dpi, when virus replication was peaked at approximately 10⁷/μL, the expressions of most of the IFN-related genes (IRF1, IRF3, IRF8, Mx, STAT1, ISG15, PKR, Viperin, GVIN1, IFI44, and ISG56) were significantly higher in group B than in the control group. Moreover, prolonged and higher levels of IRF3 (at least 30 dpi), IRF8 (at least 30 dpi), ISG15 (at least 30 dpi), PKR (at least 28 dpi), Viperin (at least 30 dpi), and IFI44 (at least 30 dpi) were also observed in the recovery stage of infection. Under the no-susceptibility condition at 17℃ (0% mortality), significantly elevated levels of IRF3, Mx, ISG15, and PKR were observed mostly until 20 dpi. The findings indicate that RBIV infection can induce an efficient IFN-mediated antiviral immune response in low- and no-susceptibility conditions. The findings could be valuable for effective control of viral pathogens in fish.