Journal of virological methods最新文献

This study describes the development of a Western blot (WB) assay employing native non-structural proteins (NSPs) from foot-and-mouth disease virus (FMDV)-infected ZZ-R 127 cell lysates to support confirmatory serodiagnosis within DIVA (differentiation of infected from vaccinated animals) frameworks. Native antigens were generated from six FMDV isolates representing serotypes A, O, and Asia-1 and were separated by denaturing SDS-PAGE. Antigen identity was verified by co-migration with recombinant counterparts and detection with monoclonal antibodies (MAbs). Using one NSP-positive and one NSP-negative polyclonal bovine serum, the assay demonstrated consistent identification of 2C, 3D, and an ~25.8kDa 3AB-consistent precursor as the most reliable diagnostic targets. While MAb profiling confirmed the presence of 3C in the lysates, it showed no reactivity with the polyclonal serum, indicating limited utility for serology. Pixel-based quantification revealed predictable dilution-dependent signal behavior within gels (R² > 0.91 on log-log plots), though inter-gel variability was substantial, favoring qualitative over quantitative application. Absence of signals in mock-infected cell lysates and in the negative serum supported specificity. By preserving the native processing context of immunodominant NSPs, this platform is well-suited as a second-tier tool to adjudicate equivocal ELISA results in vaccinated populations.

Chicken infectious anemia virus (CIAV) is a major immunosuppressive pathogen that causes huge economic losses in the poultry industry. The virus persists in the host for an extended period, making it difficult to eliminate and highlighting the need for rapid and accurate detection methods. In this study, we developed a recombinase polymerase amplification-lateral flow dipstick (RPA-LFD) kit for on-site detection of CIAV. Optimal primers and probes were designed and screened based on conserved regions of the full-gene sequences of 251 CIAV strains, ensuring high specificity, with no cross-reactions with 12 other common avian viruses. It demonstrated high sensitivity with a minimum detection limit of 10 copies of CIAV DNA, achieving efficient detection in just 15min. The kit also showed good repeatability and stability, with 100% concordance with qPCR in detecting 50 field samples. The developed CIAV RPA-LFD kit provides a practical tool for the rapid detection of CIAV, suitable for both on-site and laboratory use.

Background: Zika virus is a member of flaviviruses group and is primarily transmitted by infected mosquito. Most of the Zika virus infections are subclinical or self limiting illness like dengue fever, but certain cases are manifested with Guillain-Barre syndrome in adults and microcephaly in neonates from infected mothers. Since 2016, India has considerably reported the Zika virus infection from different states (Gujarat, Tamil Nadu, Madhya Pradesh, Rajasthan, Kerala, Maharashtra and Uttar Pradesh). Similarly there was a case suspecting arboviral infection in Shivamogga city in Mid June, 2024 and found positive for Zika virus.

Method-: A serum sample of patient with hemorrhagic fever syndrome from private super speciality hospital suspecting for Zika virus infection was collected and sent to our laboratory for diagnosis of Zika virus. The sample was screened by real time reverse transcription polymerase chain reaction (RT-PCR) and confirmed by nucleotide sequencing. Further, the sample was examined for presence of anti-Zika IgM antibody. Finding- The real time RT-PCR result revealed the presence Zika virus RNA in the sample (CT value- 27) and it was further confirmed by sequencing the partial envelope (E) gene. Additionally, the subsequent serum sample was positive for anti-Zika IgM antibody. The phylogenetic analysis evidenced that Zika virus identified in this study belongs to the Asian lineage and was homogenous to the previously reported strains from India.

Interpretation: The patient without any travel history admitted for hemorrhagic fever was found positive with Zika virus. This is the first case of laboratory-confirmed Zika virus infection in human from Shivamogga, South Karnataka. It was further confirmed by phylogenetic analysis of envelope coding region and detecting anti-Zika virus antibody. However further studies have to be transacted to know the source of infection and systematic community based surveillance is need of the hour for early detection of Zika virus transmission.

Piscine orthoreoviruses (PRV) are often associated with pathologies affecting various salmonid species, especially farmed species. Both surveillance and research purposes require diagnostic molecular-based tools able to detect the most frequent genotypes, namely PRV-1 and PRV-3. A new one-step reverse-transcription real-time PCR (RT-qPCR) was set up targeting a conserved region of the L1 segment. Based on the French standard NF U47-600, this PCR method proved to be specific to PRV, excluding other salmonid viruses. The assay can detect 1000 copies of a synthetic transcript of PRV-3, pure or mixed with nucleic acids from healthy fish. This assay produced signals similar to those observed using a previously published generic assay targeting a different viral segment. The new assay is reliable and can be used for the detection of PRV-1 and PRV-3 in rainbow trout and Atlantic salmon.

Piglets rely on passive immunity acquired from maternal-derived antibodies obtained by ingesting colostrum and milk from immunised sows. This study evaluated the differences in colostrum antibody profiles (IgA, IgG, and neutralising antibodies) according to the timing, frequency, and administration route of the porcine epidemic diarrhoea virus (PEDV) vaccination in pigs with confirmed PEDV infection, and to assess the corresponding clinical protective effects of PEDV vaccination in neonatal piglets. Piglets of sows that received two or more vaccinations, including an oral administration of a live attenuated vaccine 1 week before farrowing (3K1L and 5K3K1L groups), showed disappearance of diarrhoea and growth retardation, and exhibited a significantly higher weaning weight. Sows vaccinated with an oral live vaccine approximately 1 week before farrowing had significantly higher IgA levels than those vaccinated 2 weeks before farrowing. Neutralising antibody titres differed among the groups (p = 0.023): the thrice vaccination group (5K3K1L) showed significantly higher titres than the pre-vaccination, once (1K1L and 2K2L), and twice (3K1L) vaccination groups. IgG levels did not differ substantially between the vaccinated and unvaccinated groups. Overall, this study provides scientific evidence linking clinical protection against PEDV to antibody levels, especially IgA, in sow colostrum.

Background: Viruses, especially those of the Herpesviridae family, are increasingly linked to the pathogenesis of periapical lesions. Human cytomegalovirus (CMV), capable of lifelong latency and reactivation under stress or infection, can enhance proinflammatory cytokine release, stimulate osteoclast activity, and drive tissue destruction.

Objective: To evaluate the prevalence of CMV DNA and/or RNA in periapical lesions and to evaluate its potential role in their pathogenesis.

Materials and methods: A comprehensive search was performed using MEDLINE via PubMed, Google Scholar, Scopus, and ResearchGate up to April 2025, following PRISMA guidelines. Original studies that explored the role of CMV in periapical lesions were included. The risk of bias was evaluated with the modified Newcastle-Ottawa Scale, with results generated in Review Manager 5.4. Meta-analyses were carried out using OpenMeta Analyst.

Results: Thirty-one studies were included for qualitative analysis, of which 17 met eligibility for meta-analysis. The pooled prevalence of CMV in periapical lesions was 28 % (95 % CI: 17 %-40 %). Symptomatic lesions showed significantly higher CMV detection compared to asymptomatic cases (OR = 2.67, 95 % CI: 1.44-4.95). Subgroup analysis showed that CMV prevalence varied by detection method, with higher estimates using RT-PCR, indicating assay sensitivity as a major source of heterogeneity.

Conclusion: This review highlights that CMV was significantly associated with symptomatic and larger periapical lesions, likely contributing to disease progression via immune modulation, cytokine induction, and viral-bacterial interactions. Further research is warranted to clarify its pathogenic role and explore antiviral strategies in endodontic management.

We have generated two reporter plasmids that either monitor expression of human papillomavirus type 16 (HPV16) E2 mRNAs using splice site SA2709 or HPV16 E1C mRNAs utilizing splice site SA2582. In plasmid pE2sLuc, the entire E2 open reading frame was replaced by the secreted luciferase (sLuc) reporter gene and in the pE1nLuc plasmid the E1/E1C open reading frame was fused in frame with the nanoluciferase (nLuc) reporter gene. We demonstrate that these reporter plasmids can be used to investigate cis-acting RNA elements and trans-acting factor that regulate splicing to the E2 splice site SA2709 and the alternative E1C- splice site SA2582. Furthermore, we generated a stable C33A cell line carrying HPV16 reporter plasmid pE1nLuc. Using this cell line, we showed that splicing to the HPV16 E2 3'-splice site SA2709 could be perturbed by over expression of SR-proteins or by incubation with a pan-Akt kinase inhibitor. These proteins and substances shifted splicing from E2 splice site SA2709 to the upstream HPV16 3'-splice site SA2582 and enhanced nLuc production, thereby demonstrating the utility of the pE1nLuc cell line as a bioassay for HPV16 E2 mRNA splicing.