Journal of virological methods最新文献

{"title":"Considerations and limitations for establishing an Intact Proviral DNA Assay (IPDA) on a chip-based digital PCR system for HIV-1 reservoir quantification","authors":"Jasmin Tschumi ,&nbsp;Kathrin Neumann ,&nbsp;Dominique L. Braun ,&nbsp;Huldrych F. Günthard ,&nbsp;Karin J. Metzner ,&nbsp;the Swiss HIV Cohort Study","doi":"10.1016/j.jviromet.2025.115205","DOIUrl":"10.1016/j.jviromet.2025.115205","url":null,"abstract":"<div><div>The intact proviral DNA assay (IPDA) has become a gold standard for HIV-1 reservoir quantification in HIV-1 latency research, as well as in the evaluation of HIV-1 cure strategies. In this work, we adjusted the IPDA assay to a chip-based digital PCR (dPCR) system, established the use of CCR5 as a different reference gene, and evaluated the performance on cell lines, clinical samples from people with HIV (PWH) off and on antiretroviral therapy (ART), and different HIV-1 subtypes. Our adapted IPDA performs well on the chip-based dPCR system with no false positive intact HIV-1 in negative controls and with less hands-on time compared to droplet-based dPCR systems. Undetectable intact HIV-1 DNA is common in clinical samples on ART and correlated with total HIV-1 reservoir size and sample input concentration for HIV-1 subtype B samples. Performance on non-B subtypes varies depending on the subtype and should be improved with subtype specific primer/probe combinations.</div></div>","PeriodicalId":17663,"journal":{"name":"Journal of virological methods","volume":"338 ","pages":"Article 115205"},"PeriodicalIF":2.2,"publicationDate":"2025-06-17","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144330465","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":"IC-ELISAs for the quantitative detection of monkeypox virus cross-immunogenic modified vaccinia virus Ankara antigens L1 and A33","authors":"Mingchan Liang ,&nbsp;Feixia Gao ,&nbsp;Min Liu ,&nbsp;Linya Zhang ,&nbsp;Yongshan Zheng ,&nbsp;Jinhui Miao ,&nbsp;Cheng He ,&nbsp;Zhewen Chen","doi":"10.1016/j.jviromet.2025.115204","DOIUrl":"10.1016/j.jviromet.2025.115204","url":null,"abstract":"<div><div>Vaccination is always the most effective approach to control and prevent monkeypox epidemics. The efficacy testing of vaccines is an important component of vaccine quality control. Based on the 3Rs principles, measuring the content of effective proteins in vaccines to evaluate vaccine efficacy is an excellent alternative to traditional <em>in vivo</em> assays. In this study, we reported two indirect competitive ELISA (IC-ELISA) methods based on the use of M1- and A35-specific antibodies that recognize the L1 and A33 proteins of the modified vaccinia virus Ankara (MVA) vaccine, which are cross-immunogenic to monkeypox virus (MPXV), respectively. M1-IC-ELISA was shown to be linear over the range of 31.25–2000 ng/mL with an LOD value of 48.36 ng/mL. A35-IC-ELISA was shown to be linear over the range of 3.90–1000 ng/mL with an LOD value of 3.74 ng/mL. These two methods were found to be specific, precise, accurate and robust in the quantification of cross-immunogenic L1 and A33 in final MVA vaccine and showed a strong correlation with viral titer, MPXV cross-antibody titer and MVA neutralizing antibody titer. The developed methods may be an alternative to the <em>in vivo</em> assay in quality control testing of MVA vaccine.</div></div>","PeriodicalId":17663,"journal":{"name":"Journal of virological methods","volume":"338 ","pages":"Article 115204"},"PeriodicalIF":2.2,"publicationDate":"2025-06-17","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144371302","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":"Design and feasibility of a novel personal protection device against airborne pathogens for everyday use","authors":"Gary Leschinsky ,&nbsp;W. Andrew Dexter ,&nbsp;Barbara Leschinsky ,&nbsp;Jeffery Trolinger","doi":"10.1016/j.jviromet.2025.115203","DOIUrl":"10.1016/j.jviromet.2025.115203","url":null,"abstract":"<div><div>Airborne viruses, such as SARS-CoV-2, influenza, and potentially emerging avian influenza, are a major cause of disease spread. Airborne disease outbreaks have increased by 35 times in the past 20 years. These viruses are particularly a concern for immunocompromised individuals worldwide. ActivAir is a patented novel reusable personal protection device designed to provide a pathogen-free personal air supply. Ambient air is passed under UVC light inside a small, enclosed chamber before being released as a gentle breeze over the face of the user. Feasibility tests with a prototype disinfection chamber of the device showed a 99.99 % efficiency in a single-pass deactivation of aerosolized Phi X 174 bacteriophage, and over 96 % efficiency in deactivating aerosolized MS2 bacteriophage, both are known surrogates for SARS-COV-2, influenza, and other airborne viruses. High-efficiency UVC LEDs consume low enough energy to make this battery-powered device wearable and easy to use, lasting several hours on a single charge. This device holds a promise of providing efficient everyday protection for immunocompromised patients today, as well as a new line of defense for everyone in case of a future pandemic.</div></div>","PeriodicalId":17663,"journal":{"name":"Journal of virological methods","volume":"338 ","pages":"Article 115203"},"PeriodicalIF":2.2,"publicationDate":"2025-06-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144313605","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":"Illumina MiSeq and iSeq platforms yield comparable results for viral genomic sequencing","authors":"Darlene D. Wagner ,&nbsp;Grace Nabakooza ,&nbsp;Nehalraza Momin ,&nbsp;Rachel L. Marine","doi":"10.1016/j.jviromet.2025.115202","DOIUrl":"10.1016/j.jviromet.2025.115202","url":null,"abstract":"<div><h3>Summary</h3><div>Illumina MiSeq and iSeq are widely used short-read next-generation sequencing (NGS) platforms. The MiSeq instrument is specialized for mid-range throughput, while the iSeq instrument has lower throughput but is less expensive and simpler to operate. Several studies have compared Illumina platforms for sequencing of a variety of specimen types, but very few have quantified differences in sequencing quality, particularly for viruses. This study compared read quality, single nucleotide polymorphism (SNP) calling, and assembly metrics for SARS-CoV-2, norovirus, and poliovirus samples sequenced on both platforms. MiSeq and iSeq trimmed reads exhibited equivalent percentage of bases ≥ Q30 (% ≥ Q30) and equivalent reference mapping percentages. SNP concordance rates between the two platforms were 41 out of 43 (95.3 %) for SARS-CoV-2, 1628 out of 1633 (99.7 %) for norovirus, and 9 out of 11 (81.8 %) for poliovirus. Within each of the three virus groups, MiSeq and iSeq assemblies had equivalent N50 and maximum contig lengths. Despite platform-specific differences in the sequencing chemistry and flow cell design, MiSeq and iSeq offered comparable data quality, indicating that using either platform should produce concordant results for viral genomic sequencing.</div></div>","PeriodicalId":17663,"journal":{"name":"Journal of virological methods","volume":"338 ","pages":"Article 115202"},"PeriodicalIF":2.2,"publicationDate":"2025-06-13","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144302403","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":"Corrigendum to “Rapid and sensitive detection of shrimp infectious myonecrosis virus using a reverse transcription loop-mediated isothermal amplification and visual colorogenic nanogold hybridization probe assay” [J. Virol. Methods, Vol. 193 (2013) 542–547]","authors":"Narong Arunrut ,&nbsp;Jantana Kampeera ,&nbsp;Rungkarn Suebsing ,&nbsp;Wansika Kiatpathomchai","doi":"10.1016/j.jviromet.2025.115199","DOIUrl":"10.1016/j.jviromet.2025.115199","url":null,"abstract":"","PeriodicalId":17663,"journal":{"name":"Journal of virological methods","volume":"338 ","pages":"Article 115199"},"PeriodicalIF":1.6,"publicationDate":"2025-06-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144266530","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 Novel Cell‐free Direct Neutralization Assay for SARS-CoV-2","authors":"Ji Youn Lim ,&nbsp;Alyssa Fiore ,&nbsp;Bruce Le ,&nbsp;Corinne Minzer ,&nbsp;Halle White ,&nbsp;Krystle Burinski ,&nbsp;Humaira Janwari ,&nbsp;David Wright ,&nbsp;Sasha Perebikovsky ,&nbsp;Ralph Davis ,&nbsp;David Okrongly ,&nbsp;Aravind Srinivasan","doi":"10.1016/j.jviromet.2025.115201","DOIUrl":"10.1016/j.jviromet.2025.115201","url":null,"abstract":"<div><div>Neutralizing antibody titer elicited through infection or vaccination is widely accepted as a reliable surrogate for protection from SARS-CoV-2 infection. The gold standard for measuring these titers viz. live-virus neutralization assay suffers from the following drawbacks: the need for a BSL-3 laboratory, limited reproducibility, and difficulty adapting to emerging variants. Using the Ancestral and BA.5 SARS-CoV-2 strains as model systems, we developed the Q-NAb IgG Test for the quantitative determination of neutralizing antibodies against SARS-CoV-2 variants, traceable to WHO International Standards. The test utilizes a novel Fusion Protein that mimics the Spike receptor binding domain docked to the human ACE2 protein and effectively blocks non-neutralizing antibodies in the sample. After pre-blocking samples with Fusion Protein, direct binding of the residual neutralizing antibodies to variant RBDs coated in the wells of the microtiter plate is measured with a fluorescent secondary antibody. Results of the Q-NAb IgG Test agree with a live-virus microneutralization assay for both the Ancestral strain (WA1–2020) and the Omicron BA.5 (COR-22-063113/2022) variant (Spearman’s correlation, ρ = 0.87 and 0.92, respectively). The analytical performance (sensitivity, linearity, precision, and interference) of the Q-NAb IgG Test was established along with specificity using a panel of monoclonal neutralizing and non-neutralizing anti-SARS-CoV-2 antibodies. Clinical sensitivity and specificity using pre-pandemic, convalescent, and vaccinated serum and plasma samples is also reported. The advantages of the Q-NAb IgG Test are its strong correlation to live virus neutralization tests, traceability to WHO International Standards, convenient microtiter plate format, low sample volume requirements, and suitability for a BSL-2 laboratory.</div></div>","PeriodicalId":17663,"journal":{"name":"Journal of virological methods","volume":"338 ","pages":"Article 115201"},"PeriodicalIF":2.2,"publicationDate":"2025-06-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144271499","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 an ORF154-DIVA ELISA for serological differentiation of LSDV-infected and vaccinated animals","authors":"Alka Nokhwal ,&nbsp;Ram Kumar ,&nbsp;Yogesh Chander ,&nbsp;Nitin Khandelwal ,&nbsp;Assim Verma ,&nbsp;Thachamvally Riyesh ,&nbsp;Bhupendra N. Tripathi ,&nbsp;Naveen Kumar","doi":"10.1016/j.jviromet.2025.115200","DOIUrl":"10.1016/j.jviromet.2025.115200","url":null,"abstract":"<div><div>Mass vaccination to achieve disease-free status requires a serological test to differentiate infected and vaccinated animals (DIVA) towards the end of the control program. The lumpy skin disease (LSD) vaccines currently in use (licensed) are not DIVA-compatible. India recently approved a new live-attenuated LSD vaccine derived from the local LSD virus (LSDV) strain (Ranchi). Unlike field LSDV strains, the Ranchi strain has a distinct 801-nucleotide deletion in its inverted terminal repeat (ITR) region, affecting ORF003/ORF154. In this study, we successfully cloned and expressed LSDV ORF154 into pET28a and purified the His-tagged protein using Ni-NTA affinity chromatography. SDS-PAGE and Western blot analyses confirmed the presence of a ∼28 kDa protein, consistent with the predicted molecular weight. An optimized antigen concentration of 500 ng/well and serum dilution of 1:50, and at a positivity cut-off of 22.63 %, the assay showed high sensitivity (95.77 %) and specificity (95.42 %), effectively distinguishing infected from vaccinated cattle. These findings demonstrate the potential of an ORF154-based ELISA as a reliable serological diagnostic tool for LSDV surveillance and disease control programs, making the Ranchi strain-based LSDV vaccine the first DIVA-compatible vaccine.</div></div>","PeriodicalId":17663,"journal":{"name":"Journal of virological methods","volume":"338 ","pages":"Article 115200"},"PeriodicalIF":2.2,"publicationDate":"2025-06-08","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144266531","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 evaluation of a capillary electrophoresis-based multiplex RT-PCR kit set for detection of upper respiratory tract pathogens","authors":"Yangqing Gong ,&nbsp;Yong Luo ,&nbsp;Zhi Xu ,&nbsp;Xinjiang Lu ,&nbsp;Miaomiao Niu ,&nbsp;Dijun Zhang ,&nbsp;Yong Wu ,&nbsp;Ying Yu","doi":"10.1016/j.jviromet.2025.115191","DOIUrl":"10.1016/j.jviromet.2025.115191","url":null,"abstract":"<div><div>Respiratory tract infections (RTIs) are a major global public health problem, impacting both health and economics significantly, and have long been a concern for governments and research institutions worldwide. Although numerous related products are available, many of them only offer a limited panel of targets. In this study, we describe a set of multiplex RT-PCR kits based on capillary electrophoresis. Kit A can simultaneously detect ten viruses, two atypical pathogens, and one bacterium, covering most upper respiratory tract pathogens. Kit B can classify eight pathogens into a total of 22 subtypes. To obtain suitable primers, we used bioinformatics methods and applied different conditions to assess the conservation of primers and amplification products. For the 10 targets included in Kit A, the minimum concentration that could be reliably detected did not exceed 0.5 copies/μL. Compared to the commercial kit (Kit Health), it offers broader assay coverage and higher positivity rates. Kit B can differentiate subtypes at the detection limit concentration of Kit A and Kit B showed no cross-interference between subtypes during supersaturated concentration testing. Using both kits in combination can enhance their performance, offering better support for pathogen research and clinical applications.</div></div>","PeriodicalId":17663,"journal":{"name":"Journal of virological methods","volume":"338 ","pages":"Article 115191"},"PeriodicalIF":2.2,"publicationDate":"2025-05-31","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144208891","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 antibodies against infectious bronchitis virus strain QX (GI-19) by commercial ELISA kits and virus neutralization test","authors":"Lana Ljuma Skupnjak,&nbsp;Davor Janković,&nbsp;Marina Gabrić Dragović,&nbsp;Anto Vrdoljak,&nbsp;Katarina Huić Babić,&nbsp;Anamarija Slović","doi":"10.1016/j.jviromet.2025.115192","DOIUrl":"10.1016/j.jviromet.2025.115192","url":null,"abstract":"<div><div>The study assessed the suitability of four commercially available IBV ELISA kits for detecting antibodies against the QX strain of IBV in poultry. Two experiments were conducted: one in SPF layers under laboratory conditions and another in broiler chickens under field conditions. Chickens were vaccinated with either a candidate vaccine or existing commercially available vaccines against QX strain of IBV. Sera and tear samples were collected post-vaccination and analysed using various ELISA and virus neutralization tests.</div><div>Results show that all ELISA kits have detected antibodies induced by the candidate vaccine, with antibody levels comparable to or higher than those induced by existing vaccines. The BioChek indirect ELISA kit detected the highest antibody levels in the first experiment, while the ID Screen indirect ELISA kit detected the highest antibody levels in the second experiment. Virus-neutralizing antibodies were found in most vaccinated groups, but not in the unvaccinated control group. Tear samples indicated local antibody development peaking at 28 days post-vaccination.</div><div>Although the antibody levels in the sera of vaccinated chickens do not always correlate with protection against infection, and the role of local antibodies remains controversial, measuring antibodies by ELISA provides an indication of successful vaccination or field challenge. In conclusion, this study highlights the importance of establishing a baseline standard for expected titer values through regular serological monitoring.</div></div>","PeriodicalId":17663,"journal":{"name":"Journal of virological methods","volume":"338 ","pages":"Article 115192"},"PeriodicalIF":2.2,"publicationDate":"2025-05-31","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144208890","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":"Tupaia orthoreovirus: The first successful isolation of a novel Mammalian orthoreovirus from treeshrews in Malaysia","authors":"Zhen Yun Siew ,&nbsp;Zi Ni Ngai ,&nbsp;Siew Tung Wong ,&nbsp;Xin Yi Chew ,&nbsp;Boon Shing Tan ,&nbsp;Chee-Onn Leong ,&nbsp;Rhun Yian Koh ,&nbsp;Pooi Pooi Leong ,&nbsp;Kenny Voon","doi":"10.1016/j.jviromet.2025.115189","DOIUrl":"10.1016/j.jviromet.2025.115189","url":null,"abstract":"<div><div>Treeshrews, which share a closer common ancestor with primates and flying lemurs than with rodents, have been proposed as a better animal model for biomedical research compared to traditional rodent models. Zoonotic viruses like <em>Mammalian orthoreovirus</em> (MRV) pose potential public health risks, but their prevalence and characteristics in treeshrews remain underexplored. This study reports the first isolation of a novel MRV from common treeshrews (<em>Tupaia glis</em>) in Malaysia and investigates its biological properties. Faecal and urine samples from common treeshrews were collected and processed through centrifugation and sterile filtration. The processed samples were inoculated into mammalian cell cultures for virus isolation using a novel virus isolation protocol. The isolated virus was sequenced and analysed phylogenetically. A novel MRV, designated MRV1UNM, was isolated and identified as MRV serotype 3 (MRV3) with a unique genomic profile. Syncytial formation was observed in Vero cells during early passages only. MRV1UNM infected a broad spectrum of cell lines, including some cancer cells, causing cell lysis within days without establishing persistent infections. Moreover, increased caspase 3/7 activity was also observed in the infected A549 cells. These findings provide evidence of the oncolytic potential of MRV1UNM. In conclusion, this study reports the first isolation of MRV from Malaysian treeshrews, highlighting their role as potential reservoirs for zoonotic viruses. The promising potential of MRV1UNM as a candidate for oncolytic virotherapy warrants further investigation into its therapeutic applications.</div></div>","PeriodicalId":17663,"journal":{"name":"Journal of virological methods","volume":"337 ","pages":"Article 115189"},"PeriodicalIF":2.2,"publicationDate":"2025-05-27","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144170594","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}