Journal of virological methods最新文献

{"title":"Monkeypox in the 21st century: Insights into its pathogenesis and public health implications","authors":"Duong Thuy Linh ,&nbsp;Muhammad Nadir Shabbir","doi":"10.1016/j.jviromet.2025.115223","DOIUrl":"10.1016/j.jviromet.2025.115223","url":null,"abstract":"<div><div>Monkeypox, a zoonotic illness caused by the Monkeypox virus (MPXV), is a re-emerging disease that has resurfaced as a prominent global public health issue in the 21st century. The 2022 outbreak, previously limited to Central and West Africa, exhibited extraordinary global dissemination, prompting critical inquiries over the virus's transmission mechanisms and containment strategies. This paper analyzes monkeypox's pathophysiology, epidemiological trends, and public health ramifications, emphasizing diagnostic difficulties, underreporting, and the inequitable access to vaccines and therapies. The discussion encompasses the significance of genomic surveillance, One Health strategies, and the necessity for research on effective antivirals and vaccines. Substantial knowledge deficiencies, encompassing virus mutation hazards and a restricted comprehension of animal reservoirs, impede preventative initiatives. Mitigating future epidemics necessitates addressing these problems through improved diagnoses, equitable vaccination distribution, and enhanced surveillance. This paper examines the present status of Monkeypox research and delineates prospective directions to inform global public health policies.</div></div>","PeriodicalId":17663,"journal":{"name":"Journal of virological methods","volume":"338 ","pages":"Article 115223"},"PeriodicalIF":2.2,"publicationDate":"2025-07-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144656479","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Retraction notice to \"Monitoring of human cytomegalovirus glycoprotein B genotypes using real-time quantitative PCR in immunocompromised Chinese patients\" [J. Virol. Methods, 160 (2009) 74-77].","authors":"J Fan, X Zhang, X M Chen, H N Gao, M F Yang, H Zhao, J H Hu, W H Ma","doi":"10.1016/j.jviromet.2025.115221","DOIUrl":"10.1016/j.jviromet.2025.115221","url":null,"abstract":"","PeriodicalId":17663,"journal":{"name":"Journal of virological methods","volume":" ","pages":"115221"},"PeriodicalIF":2.2,"publicationDate":"2025-07-15","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144649834","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Establishment of an indirect ELISA for feline coronavirus antibody detection and serotype discrimination","authors":"Xiaoman Lu ,&nbsp;Yanxuan Hu ,&nbsp;Xiaoyuan Chen ,&nbsp;Yongyi Shen","doi":"10.1016/j.jviromet.2025.115224","DOIUrl":"10.1016/j.jviromet.2025.115224","url":null,"abstract":"<div><div>Feline coronavirus (FCoV) is a highly contagious pathogen that is endemic to feline populations and is classified into two serotypes, I and II. Current diagnostic techniques are insufficient for distinguishing between these serotypes, which impedes effective surveillance and prevention efforts. In response to this limitation, we have developed an indirect enzyme-linked immunosorbent assay (ELISA) utilizing recombinant nucleocapsid (N) protein for the broad detection of FCoV, alongside receptor-binding domain (RBD) proteins specific to serotypes I (I-RBD) and II (II-RBD) for the purpose of serotype differentiation. The assay underwent systematic optimization, achieving high sensitivity (with detection limits of 1:64,000 dilution for N and I-RBD, and 1:32,000 for II-RBD) and specificity, exhibiting no cross-reactivity with feline calicivirus (FCV), feline herpesvirus (FHV), or feline parvovirus (FPV). The reproducibility of the assay was validated, with intra- and inter-assay coefficients of variation remaining below 10 %. Clinical validation conducted on 123 feline serum samples indicated a seroprevalence of 73.17 %, with the serotype distribution comprising 91.11 % serotype I, 1.11 % serotype II, 2.22 % mixed infections, and 5.56 % cases that could not be typed. This ELISA represents a rapid, cost-effective, and field-deployable method for large-scale FCoV surveillance and serotyping, thereby contributing to enhanced feline health management and epidemiological research.</div></div>","PeriodicalId":17663,"journal":{"name":"Journal of virological methods","volume":"338 ","pages":"Article 115224"},"PeriodicalIF":2.2,"publicationDate":"2025-07-13","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144632225","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Development and characterization of a mouse-panda chimeric antibody with neutralizing activity against canine distemper virus","authors":"Zhengguo Wang ,&nbsp;Guishan Ye ,&nbsp;Kuijing He ,&nbsp;Cong Cai ,&nbsp;Anding Zhang ,&nbsp;Long Li ,&nbsp;Li Han","doi":"10.1016/j.jviromet.2025.115222","DOIUrl":"10.1016/j.jviromet.2025.115222","url":null,"abstract":"<div><div>Canine distemper (CD) poses as the primary virulent infectious disease threatening the safety of panda populations, and monoclonal antibodies (mAbs) represent a potential therapeutic option for the treatment of CD in pandas. To avoid immune rejection of mouse-derived mAbs in panda clinical treatment, this study utilized a mouse-derived mAb targeting the CDV-H protein to develop a mouse-panda chimeric antibody and established a CHO cell line with stable high-expression of this chimeric antibody. SDS-PAGE results indicated that the chimeric antibody expressed by CHO cells was correctly expressed. Both indirect ELISA and indirect immunofluorescence experiments demonstrated that the chimeric antibody specifically recognized the CDV-H protein and CDV virus. Furthermore, neutralization experiments confirmed its neutralizing effect on CDV. This study successfully prepared the panda-derived chimeric antibody, providing a potential candidate drug reserve for the treatment of CD in pandas.</div></div>","PeriodicalId":17663,"journal":{"name":"Journal of virological methods","volume":"338 ","pages":"Article 115222"},"PeriodicalIF":2.2,"publicationDate":"2025-07-11","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144626668","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"A high-sensitivity qPCR method for detecting residual Vero cell DNA in rabies vaccine production","authors":"María P. Almario ,&nbsp;Jorge Rivera ,&nbsp;Catalina Páramo,&nbsp;Valentina Jaramillo,&nbsp;Yanira Chaparrro,&nbsp;Zulma Rocío Suárez-Moreno","doi":"10.1016/j.jviromet.2025.115217","DOIUrl":"10.1016/j.jviromet.2025.115217","url":null,"abstract":"<div><div>Residual host cell DNA in biological products requires precise quantification to meet regulatory safety standards. This study presents the development and validation of a qPCR assay for detecting and quantifying residual Vero cell DNA in rabies vaccine formulations. A SYBR Green-based quantitative PCR (qPCR) assay targeting the Vero cell-specific alpha-satellite DNA sequence was developed and validated in accordance with regulatory guidelines. The method was evaluated for specificity, sensitivity, linearity, precision, and robustness using a certified genomic DNA standard. The assay exhibited high specificity with no amplification from non-target DNA. The method demonstrated a linear dynamic range from 0.064 to 1000 ng/mL (R² &gt; 0.999) with an amplification efficiency of 96.3 %. Sensitivity analysis confirmed reliable detection well below the regulatory threshold of 10 ng per dose, with a calculated limit of quantification of 0.31 ng/mL. Intra- and inter-assay precision tests showed low variability (CV &lt; 20 %), supporting assay reproducibility. Accuracy was confirmed through recovery experiments (93.33–117.33 %) and concordance with a reference method (kappa = 1). This validated qPCR method offers a sensitive, specific, and reproducible tool for monitoring residual Vero cell DNA in rabies vaccine production, ensuring regulatory compliance through robust quality control.</div></div>","PeriodicalId":17663,"journal":{"name":"Journal of virological methods","volume":"338 ","pages":"Article 115217"},"PeriodicalIF":2.2,"publicationDate":"2025-07-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144588234","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Rapid and efficient generation of viral genome knock-in cell lines using the CRISPR-Cas9 system to produce infectious virus","authors":"Pooja Bhatia ,&nbsp;Aas Mohd ,&nbsp;Ishika Agrawal ,&nbsp;Harshita Katiyar ,&nbsp;Amit Goel ,&nbsp;Meghali Aich ,&nbsp;Debojyoti Chakraborty ,&nbsp;Naga Suresh Veerapu","doi":"10.1016/j.jviromet.2025.115219","DOIUrl":"10.1016/j.jviromet.2025.115219","url":null,"abstract":"<div><div>Several medically significant viruses are difficult to propagate with conventional laboratory host systems, limiting their availability for detailed characterization, antiviral screening, and functional studies. A range of methods can be used to generate viruses, such as creating sophisticated cell lines, organoid cultures, and the utilization of animal models. Here, we report the generation and characterization of CRISPR-Cas9 edited Huh7 stable cell lines engineered to carry and express overlength HBV genotypes A, B, C and D and full HEV genomes in the AAVS1 site. Viral polymerase inhibitors and IFN-α significantly reduced the production of viral genomes and proteins from the edited cells. The virus released by the edited cells was infectious in permissive cell lines and could be blocked by neutralizing antibodies. This approach can extend to other viruses, like HCV genotype 3, that are hard to culture or to culturable viruses, like Dengue, for vaccine production.</div></div>","PeriodicalId":17663,"journal":{"name":"Journal of virological methods","volume":"338 ","pages":"Article 115219"},"PeriodicalIF":2.2,"publicationDate":"2025-07-08","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144605866","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Development and validation of oligonucleotide sets for real time RT-PCR detection and next generation sequencing, of re-emerged sylvatic Dengue virus 2 strains","authors":"Ndeye Aminata Dia ,&nbsp;Mignane Ndiaye ,&nbsp;Diamilatou Balde ,&nbsp;Mouhamed Kane ,&nbsp;Agathe Shella Efire ,&nbsp;Gerald Mboowa ,&nbsp;Fatou Thiam ,&nbsp;Yahya Dieye ,&nbsp;Moussa Dia ,&nbsp;Gamou Fall ,&nbsp;Ndongo Dia ,&nbsp;Amadou Alpha Sall ,&nbsp;Ousmane Faye ,&nbsp;Oumar Faye ,&nbsp;Moussa Moïse Diagne ,&nbsp;Manfred Weidmann ,&nbsp;Idrissa Dieng","doi":"10.1016/j.jviromet.2025.115218","DOIUrl":"10.1016/j.jviromet.2025.115218","url":null,"abstract":"<div><div>Dengue virus (DENV) is one of the most prevalent arboviral threats worldwide. The virus is associated with a high health and economic burden mainly in tropical and subtropical regions. Available molecular tools however fail to correctly serotype and sequence sylvatic DENV-2 (DENV-2/GVI) which in known to circulate in forests in West Africa and Malaysia. The recent emergence of human case linked to this virus variant in Southern Senegal raises concerns about the correct detection and characterization of the virus for public health purposes. Here we developed and validate new sets of oligonucleotides for DENV-2 (DENV-2/GVI) detection, and next generation sequencing. Validations were carried out using epidemic DENV and sylvatic DENV-2 strains from the biobank of the WHO collaborating Center for Arboviruses and Haemorrhagic fevers. The presented approaches showed good performance to specifically detect sylvatic DENV-2 in both singleplex and multiplex PCR with other DENV serotypes respectively with a limit of detection of 68.85 and 133.21 RNA copies/reaction at 0.95 probability in a probit analysis. Additionally, tilling PCR primers were developed which yield a better genome coverage ranging from 93.9 % to 95.1 % for all processed DENV-2/GVI strains both on Illumina and Nanopore platforms and outperform previous schemes to efficiently amplified DENV-2/GVI strains. In summary the developed oligonucleotides will contribute to improving DENV surveillance and genomic epidemiology in endemic areas.</div></div>","PeriodicalId":17663,"journal":{"name":"Journal of virological methods","volume":"338 ","pages":"Article 115218"},"PeriodicalIF":2.2,"publicationDate":"2025-07-05","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144584273","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Detection of cross-reactivity of antibodies to the N proteins of feline morbillivirus and canine distemper virus in Japanese cat plasma samples","authors":"Shwe Thiri Maung Maung Khin ,&nbsp;Sheikhi Mohammad Jafar ,&nbsp;Minglin Ju ,&nbsp;Fumi Murakoshi ,&nbsp;Tetsuya Furuya","doi":"10.1016/j.jviromet.2025.115216","DOIUrl":"10.1016/j.jviromet.2025.115216","url":null,"abstract":"<div><div>Feline morbillivirus (FeMV) is a globally emerging virus that has been linked to chronic kidney disease (CKD) in infected cats. Immunological assays, such as enzyme-linked immunosorbent assay (ELISA), are important for studying the virus and monitoring its prevalence. A study using rabbit antiserum demonstrated antigenic cross-reactivity between nucleocapsid (N) proteins of FeMV and canine distemper virus (CDV), suggesting not only the risk of false-positive anti-FeMV antibody detection in ELISAs but also potentially false-positive FeMV antigen detection in Western blotting. To examine whether such cross-reactivity occurs in tests using cat plasma samples, we developed ELISAs using affinity-purified recombinant N proteins of FeMV and CDV with expression in <em>Escherichia coli</em> and tested 100 cat plasma samples collected from veterinary clinics in Japan. Twenty samples were found to be positive for anti-FeMV antibodies, while 6 were positive for anti-CDV antibodies. All these latter 6 samples were double-positive for anti-FeMV antibodies. Western blotting with the purified proteins confirmed the specificity of these antibodies to their target viral antigens. A reverse transcription-quantitative PCR assay with a detection limit of 100 copies failed to detect CDV genomic RNA in these 6 double-positive samples. These results strongly suggest the cross-reactivity between anti-FeMV N protein antibodies in cat plasma samples and the CDV N protein. This antigenic cross-reactivity should be considered in future studies using immunological methods employing FeMV or CDV N proteins, or antibodies targeting them.</div></div>","PeriodicalId":17663,"journal":{"name":"Journal of virological methods","volume":"338 ","pages":"Article 115216"},"PeriodicalIF":2.2,"publicationDate":"2025-07-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144549253","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Comparison of plant- and mammalian cell-produced human papillomavirus pseudovirions","authors":"Albertha R. van Zyl ,&nbsp;Sarah Lindsay ,&nbsp;Georgia Schäfer ,&nbsp;Edward P. Rybicki ,&nbsp;Inga I. Hitzeroth","doi":"10.1016/j.jviromet.2025.115215","DOIUrl":"10.1016/j.jviromet.2025.115215","url":null,"abstract":"<div><div>High-risk human papillomaviruses (HPVs) are the primary etiological agents of cervical, anal and oropharyngeal cancers. While existing vaccines are effective in preventing infection, their impact in low-and middle-income countries (LMICs) is limited by type coverage, high costs and uptake. To address this gap, there is a critical need for next-generation vaccines that are both regionally tailored and cost-effective, along with efficient and accessible tools for evaluating their efficacy. HPV pseudovirions (PsVs), which encapsidate a reporter plasmid, are widely used in pseudovirion-based neutralisation assays (PBNAs) and <em>in vivo</em> murine models to assess vaccine-induced immunity – and have potential for use as DNA vaccine delivery systems. Traditionally, PsVs are produced in mammalian cells, which remain the gold standard due to their high infectivity and structural fidelity. However, recent studies have demonstrated the feasibility of producing PsVs in plants, a platform that offers lower infrastructure and reagent costs, scalability, and biosafety advantages. Although plant-derived PsVs have shown promise in PBNAs, their performance in <em>in vivo</em> models had not been evaluated prior to this study. Here, we compared mammalian cell-derived PsVs encapsidating either Gaussia or firefly luciferase reporter plasmids and found that firefly luciferase provided more consistent and robust signals in both <em>in vitro</em> and <em>in vivo</em> assays. Building on this, we generated PsVs encapsidating the firefly luciferase gene using both mammalian and plant expression systems, and assessed their infectivity. While plant-derived PsVs were capable of infecting HeLa cells and mice in a cervicovaginal challenge model, mammalian-derived PsVs exhibited significantly higher infectivity overall. These findings represent the first demonstration of <em>in vivo</em> infectivity of plant-produced HPV PsVs and highlight their potential as a cost-effective alternative for immunogenicity testing and potentially as vaccines. Although further optimization is needed, particularly in capsid assembly and purification, plant-based PsV production holds promise for expanding access to HPV research tools and supporting vaccine development in resource-limited settings.</div></div>","PeriodicalId":17663,"journal":{"name":"Journal of virological methods","volume":"338 ","pages":"Article 115215"},"PeriodicalIF":2.2,"publicationDate":"2025-06-30","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144517584","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Development of rapid and simple FCoV RNA detection systems using RT-PCR and RT-RPA combined with STH-PAS to diagnose FIP in cats","authors":"Tomoyoshi Doki,&nbsp;Misaki Ohno,&nbsp;Mihoko Kaku,&nbsp;Saori Odani,&nbsp;Kaito To,&nbsp;Tomomi Takano","doi":"10.1016/j.jviromet.2025.115214","DOIUrl":"10.1016/j.jviromet.2025.115214","url":null,"abstract":"<div><div>Feline infectious peritonitis (FIP) is a fatal disease in cats that is caused by feline coronavirus (FCoV). FCoV RT-qPCR is widely used to diagnose FIP due to its high sensitivity and ability to quantify FCoV RNA. However, its convenience is limited by the need for expensive equipment and/or processing at external laboratories. We herein developed two rapid and simple FCoV RNA detection systems: one combining conventional RT-PCR with the Single Tag Hybridization-Printed Array Strip (STH-PAS) method (the RT-PCR and STH-PAS system) and the other combining RT-RPA, an isothermal nucleic acid amplification method, with STH-PAS (the RT-RPA and STH-PAS system). Evaluations using FCoV RNA standards showed that the limit of detection for the RT-PCR and STH-PAS system was 10^4.2 copies/reaction, while that for the RT-RPA and STH-PAS system was 10⁴ copies/reaction. The clinical performance of these systems was also examined using clinical samples from cats suspected of having FIP and compared to the conventional FCoV RT-qPCR genetic test. The results obtained showed a sensitivity of 66.7 % (95 % CI: 41.0–86.7) and specificity of 100 % (95 %CI: 9.4–100). These systems are a faster and simpler alternative to conventional methods, suggesting their potential in point-of-care testing in veterinary clinics.</div></div>","PeriodicalId":17663,"journal":{"name":"Journal of virological methods","volume":"338 ","pages":"Article 115214"},"PeriodicalIF":2.2,"publicationDate":"2025-06-28","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144518303","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}