Journal of virological methods最新文献

D-cycloserine, a potential candidate for reducing Hepatitis B virus cccDNA in vitro d -环丝氨酸：体外减少乙型肝炎病毒cccDNA的潜在候选物

IF 2.2 4区医学

Journal of virological methods Pub Date : 2025-04-28 DOI: 10.1016/j.jviromet.2025.115172

Yongwook Choi , Yong Kwang Park , Wonhee Hur , Gahee Kim , Songmee Bae

{"title":"D-cycloserine, a potential candidate for reducing Hepatitis B virus cccDNA in vitro","authors":"Yongwook Choi , Yong Kwang Park , Wonhee Hur , Gahee Kim , Songmee Bae","doi":"10.1016/j.jviromet.2025.115172","DOIUrl":"10.1016/j.jviromet.2025.115172","url":null,"abstract":"<div><div>Hepatitis B virus (HBV) is a 3.2 kb hepatotropic DNA that possesses a unique episomal DNA form known as covalently closed circular DNA (cccDNA). cccDNA is the major risk factor for persistent HBV infection and consequently causes chronic liver diseases such as hepatitis, cirrhosis, and hepatocellular carcinoma. To prevent the progression of liver disease, eradication of HBV, especially cccDNA, is essential. In this study, we established a drug screening system using artificial recombinant HBV cccDNA (rcccDNA), which is regulated by a loxP-HBV genome and CRE expression. To identify potential drugs targeting cccDNA, a total of 379 antiviral reagents were tested. Among them, several chemicals including danoprevir, L- and D-cycloserine, phenytoin sodium, amantadine, and germacrone showed a decrease in cccDNA levels. Especially, D-cycloserine diminished the secretion of HBV antigens and induced cccDNA degradation in the HBV infection system. This screening system helps to develop the therapeutic drug target to cccDNA This screening system may help develop therapeutic drugs targeting cccDNA.</div></div>","PeriodicalId":17663,"journal":{"name":"Journal of virological methods","volume":"336 ","pages":"Article 115172"},"PeriodicalIF":2.2,"publicationDate":"2025-04-28","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143899920","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}

引用次数: 0

Strictly screening of HTLV-1/2 peptides can drive the development of rapid point-of-care tests 严格筛选HTLV-1/2肽可以推动快速护理点检测的发展

IF 2.2 4区医学

Journal of virological methods Pub Date : 2025-04-28 DOI: 10.1016/j.jviromet.2025.115170

Laura Jorge Cox , Vanessa Gomes Fraga , Ricardo Toshio Fujiwara , Danielle S.O. Daian e Silva , Gabriela Melo Franco , Anderson Santos Rocha , Tatyane M. Cirilo , Marina L. Martins , Agostinho G. Viana , Adele Caterino-de-Araujo , Antonio C.R. Vallinoto , Edel F. Barbosa-Stancioli

{"title":"Strictly screening of HTLV-1/2 peptides can drive the development of rapid point-of-care tests","authors":"Laura Jorge Cox , Vanessa Gomes Fraga , Ricardo Toshio Fujiwara , Danielle S.O. Daian e Silva , Gabriela Melo Franco , Anderson Santos Rocha , Tatyane M. Cirilo , Marina L. Martins , Agostinho G. Viana , Adele Caterino-de-Araujo , Antonio C.R. Vallinoto , Edel F. Barbosa-Stancioli","doi":"10.1016/j.jviromet.2025.115170","DOIUrl":"10.1016/j.jviromet.2025.115170","url":null,"abstract":"<div><div>The <em>Human T-cell lymphotropic virus</em> (HTLV-1/2) cause neglected infections that drives life-threatening diseases and the numbers of infected people around the world are underestimated. Point-of-care tests (POCT) are useful to identifying carriers, to controlling the infection’s spread with timely and cost-effectiveness, to include the most affected areas and susceptible populations, and the establishment of public health policies, including the control of vertical transmission. After in silico analysis of Env, Gag and Tax proteins of HTLV-1 and HTLV-2, we synthetized and characterized peptides to screening antibodies anti-HTLV-1/2 with high sensitivity and specificity. The 173 peptides chosen were screened by immunoblot, and by indirect in-house ELISA. Peptides that had best performed in recognize both, HTLV-1 or HTLV-2 sera from infected individual, were Gag-HTLV-1 and Gag-HTLV-2 showing to be very good candidates for screening tests. Peptides of Tax-HTLV-1, and Env-HTLV-2 had discriminated sera from HTLV-1 and HTLV-2 with high sensitivity and specificity. The screening of HTLV-1/2 peptides showed here, including the use of sera from HIV-infected individuals along with seronegative ones were crucial to avoid the use of peptides with unspecific reaction in the final pilot tests, and to reach the Point-of-care test that is under registration at regulatory Brazilian agency.</div></div>","PeriodicalId":17663,"journal":{"name":"Journal of virological methods","volume":"336 ","pages":"Article 115170"},"PeriodicalIF":2.2,"publicationDate":"2025-04-28","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143899921","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}

引用次数: 0

Laboratory confirmation of measles cases in GREECE, February-March 2024 2024年2月至3月希腊麻疹病例的实验室确认

IF 2.2 4区医学

Journal of virological methods Pub Date : 2025-04-27 DOI: 10.1016/j.jviromet.2025.115171

Elina Horefti , Theano Georgakopoulou , Sofia Makka , Emmanouil Angelakis

引用次数: 0

Clinical performance assessment of the sansure HIV-1 quantitative test system 无计量HIV-1定量检测系统临床性能评价

IF 2.2 4区医学

Journal of virological methods Pub Date : 2025-04-18 DOI: 10.1016/j.jviromet.2025.115167

Grace E. Kushemerewa , Wei-Min Miao , Xu Fan , Peng Yin , Peng Hu , Andrea Battola , Juan Liu , Isaac Ssewanyana , Li-Zhong Dai

{"title":"Clinical performance assessment of the sansure HIV-1 quantitative test system","authors":"Grace E. Kushemerewa , Wei-Min Miao , Xu Fan , Peng Yin , Peng Hu , Andrea Battola , Juan Liu , Isaac Ssewanyana , Li-Zhong Dai","doi":"10.1016/j.jviromet.2025.115167","DOIUrl":"10.1016/j.jviromet.2025.115167","url":null,"abstract":"<div><div>The significant burden of Human Immunodeficiency Virus 1 (HIV-1) infections, particularly in low- and middle-income countries (LMICs), emphasizes the need for industries to develop cost-effective and reliable solutions for the detection and management of HIV. This study presents the findings from a clinical evaluation of the Sansure HIV-1 Quantitative test, applied alongside a fully automated nucleic acid extraction process. A total of 503 blood samples from HIV infected or high-risk people were tested using both the Sansure HIV-1 Quantitative test and the Cobas HIV-1 Quantitative test (the reference method), and the diagnostic accuracy was assessed by comparing the results. The Sansure HIV-1 Quantitative test showed a sensitivity of 98.3 % and a specificity of 98.5 %. Misclassification occurred in 1.6 % (8/503) of cases, which remains below the acceptable threshold of 5 %. The correlation and agreement with the reference method fell within acceptable parameters, with a correlation coefficient of 0.93. These data establish the suitability and accuracy of the Sansure HIV-1 Quantitative test for quantifying HIV-1 RNA (viral load) in clinical samples.</div></div>","PeriodicalId":17663,"journal":{"name":"Journal of virological methods","volume":"336 ","pages":"Article 115167"},"PeriodicalIF":2.2,"publicationDate":"2025-04-18","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143888123","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}

引用次数: 0

Disassembly mediated multimodal chromatography based purification of HPV-VLPs produced in Pichia pastoris 毕赤酵母中HPV-VLPs的分离纯化

IF 2.2 4区医学

Journal of virological methods Pub Date : 2025-04-16 DOI: 10.1016/j.jviromet.2025.115168

Rashmi Sharma , Pragya Prakash , Lukas Gerstweiler , Anurag S. Rathore

引用次数: 0

Developing Zika virus-transduced hACE2 expression models for severe acute respiratory syndrome coronavirus 2 infection in vitro and in vivo 开发寨卡病毒转导的 hACE2 表达模型，用于严重急性呼吸系统综合征冠状病毒 2 的体外和体内感染

IF 2.2 4区医学

Journal of virological methods Pub Date : 2025-04-14 DOI: 10.1016/j.jviromet.2025.115166

Joh-Sin Wu , Ju-Ying Kan , Young-Sheng Chang , Uyen Nguyen Phuong Le , Wen-Chi Su , Hsueh-Chou Lai , Cheng-Wen Lin

{"title":"Developing Zika virus-transduced hACE2 expression models for severe acute respiratory syndrome coronavirus 2 infection in vitro and in vivo","authors":"Joh-Sin Wu , Ju-Ying Kan , Young-Sheng Chang , Uyen Nguyen Phuong Le , Wen-Chi Su , Hsueh-Chou Lai , Cheng-Wen Lin","doi":"10.1016/j.jviromet.2025.115166","DOIUrl":"10.1016/j.jviromet.2025.115166","url":null,"abstract":"<div><div>To address the human ACE2 dependence for SARS-CoV-2 infection, this study presents a novel strategy for generating ZIKV-hACE2 single-round infectious particles (SRIPs) by incorporating the hACE2 gene into a Zika virus (ZIKV) mini-replicon. SARS-CoV-2 SRIP infection was significantly enhanced in HEK293T cells pre-infected with ZIKV-hACE2, as evidenced by increased cytopathic effects and elevated mRNA and protein levels of the SARS-CoV-2 nucleocapsid (N) protein. A mouse model was also developed with this approach to investigate SARS-CoV-2 infection. Immunohistochemical and real-time RT-PCR analyses confirmed the presence of the SARS-CoV-2 N protein in the lungs of mice injected with ZIKV-hACE2 SRIPs, indicating successful infection. The mouse model displayed COVID-19-like pathological changes, including increased macrophages in BALF, severe lung damage, and elevated pro-inflammatory cytokines (IL-6 and IL-1β). These features mimic severe COVID-19 cases in humans. Additionally, treatment with nirmatrelvir resulted in a 6.2-fold reduction in viral load and a marked decrease in N protein levels. Overall, this ZIKV mini-replicon-mediated hACE2 expression model, both in vitro and in vivo, is a valuable tool for studying SARS-CoV-2 infection and evaluating therapeutic interventions. The mouse model’s pathological features further underscore its relevance for in vivo research on SARS-CoV-2.</div></div>","PeriodicalId":17663,"journal":{"name":"Journal of virological methods","volume":"336 ","pages":"Article 115166"},"PeriodicalIF":2.2,"publicationDate":"2025-04-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143828544","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}

引用次数: 0

Establishment of triple-RPA-LFS detection method for three common shrimp viruses 三种常见对虾病毒的三重rpa - lfs检测方法的建立

IF 2.2 4区医学

Journal of virological methods Pub Date : 2025-04-12 DOI: 10.1016/j.jviromet.2025.115156

Quanling Mu, Cunbao Ding, Ying Xie, Xi Zhen, Jiaming Zhang, Yakun Yu

{"title":"Establishment of triple-RPA-LFS detection method for three common shrimp viruses","authors":"Quanling Mu, Cunbao Ding, Ying Xie, Xi Zhen, Jiaming Zhang, Yakun Yu","doi":"10.1016/j.jviromet.2025.115156","DOIUrl":"10.1016/j.jviromet.2025.115156","url":null,"abstract":"<div><div>This study focuses on three viruses affecting farmed shrimp, including <em>White Spot Syndrome Virus</em> (WSSV), <em>Infectious Hypodermal and Hematopoietic Necrosis Virus</em> (IHHNV), and <em>Taura Syndrome Virus</em> (TSV). Specific primers and probes were designed by their respective conserved gene fragments to establish a triple-RPA-LFS detection method that simultaneously detects WSSV, IHHNV, and TSV. Seven pathogens and healthy shrimp tissues were collected to conduct specificity tests. This method can specifically amplify the gene fragments of WSSV, IHHNV, and TSV, while no fragments were amplified from the muscle tissues of healthy shrimp or other pathogens, indicating strong specificity. The reaction system was optimized, and specificity and sensitivity were validated. Sensitivity tests were conducted using a continuous dilution plasmid method, determining that the detection sensitivity of this method is 10<sup>1</sup> copies/reaction. Compared with the sensitivity of the qPCR detection method recommended by the World Organization for Animal Health (WOAH, formerly OIE), the triple-RPA-LFS method established in this study is faster and simpler to operate. When applied to test 110 samples simultaneously with the laboratory standard testing method, the results of the qPCR detection matched the results of the laboratory standard method with a 100 % concordance rate. These experimental results indicate that the triple-RPA-LFS detection method established in this study has the characteristics of high specificity, high sensitivity, short detection time, and high accuracy. It can be used for rapid on-site detection and diagnosis of the three pathogens: WSSV, IHHNV, and TSV.</div></div>","PeriodicalId":17663,"journal":{"name":"Journal of virological methods","volume":"336 ","pages":"Article 115156"},"PeriodicalIF":2.2,"publicationDate":"2025-04-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143852330","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}

引用次数: 0

A simple detection method for potato viruses combining template preparation by heat treatment of homogenate with primers and one-step multiplex RT-PCR 用引物热处理匀浆制备模板和一步多重RT-PCR相结合的简易马铃薯病毒检测方法

IF 2.2 4区医学

Journal of virological methods Pub Date : 2025-04-10 DOI: 10.1016/j.jviromet.2025.115165

Tomo Suzuki , Takehiro Ohki

{"title":"A simple detection method for potato viruses combining template preparation by heat treatment of homogenate with primers and one-step multiplex RT-PCR","authors":"Tomo Suzuki , Takehiro Ohki","doi":"10.1016/j.jviromet.2025.115165","DOIUrl":"10.1016/j.jviromet.2025.115165","url":null,"abstract":"<div><div>Potatoes are susceptible to viral diseases, and widespread viral infections lead to reduced production yields. Potato leaf roll virus, potato virus S, potato virus X, and potato virus Y are important viruses predominant in potato fields and cause significant economic damage in Japan. In this study, to reduce the labor and cost of virus testing in seed certification program, a simple molecular diagnostic assay for these potato viruses was developed. This method consists of template preparation by heat-treating a mixture of leaf homogenate and a newly designed primer set, followed by one-step conventional multiplex RT-PCR (mRT-PCR). In particular, to simplify RNA extraction, a procedure was adopted in which leaf homogenates in PBST were diluted 100-fold in RNase-free water, heat-treated with primers for 5 min at 70°C and subjected to one-step mRT-PCR. Interestingly, heat treatment of a mixture of the homogenate and primers improved the detection sensitivity for the virus. This new method’s detection sensitivity of each virus was 10–100 times higher than that of ELISA using commercial kits and 1–10 times higher than that of one-step mRT-PCR with filter paper-based RNA extraction. The heat-treated homogenate with primers was also applicable for detecting eight other potato viruses by one-step conventional mRT-PCR and four potato viruses by real-time mRT-PCR which our research group previously developed. New direct RT-PCR method for potato viruses can be applied to seed certification programs, where a large number of samples need to be tested.</div></div>","PeriodicalId":17663,"journal":{"name":"Journal of virological methods","volume":"336 ","pages":"Article 115165"},"PeriodicalIF":2.2,"publicationDate":"2025-04-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143847699","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}

引用次数: 0

Development of neutralization tests using single-round infectious particles and cytopathic effect as an alternative method for measuring antibody titers against Japanese encephalitis virus in national epidemiological surveillance program of vaccine-preventable diseases in Japan 在日本疫苗可预防疾病的国家流行病学监测规划中，发展单轮感染颗粒中和试验和细胞病变效应作为测定乙型脑炎病毒抗体滴度的替代方法

IF 2.2 4区医学

Journal of virological methods Pub Date : 2025-04-08 DOI: 10.1016/j.jviromet.2025.115163

Shunsuke Yazawa , Yumiko Saga , Mami Matsuda , Ryosuke Suzuki , Shigeru Tajima , Chang-Kweng Lim , Hideki Tani

{"title":"Development of neutralization tests using single-round infectious particles and cytopathic effect as an alternative method for measuring antibody titers against Japanese encephalitis virus in national epidemiological surveillance program of vaccine-preventable diseases in Japan","authors":"Shunsuke Yazawa , Yumiko Saga , Mami Matsuda , Ryosuke Suzuki , Shigeru Tajima , Chang-Kweng Lim , Hideki Tani","doi":"10.1016/j.jviromet.2025.115163","DOIUrl":"10.1016/j.jviromet.2025.115163","url":null,"abstract":"<div><div>The “National Epidemiological Surveillance of Vaccine-Preventable Diseases” is conducted annually in Japan for various infectious diseases. A susceptibility survey for Japanese encephalitis was conducted by measuring antibody titers using a focus-forming assay. However, this method is complicated to operate and hard to count small focuses; therefore, the development of new measurement methods is required. In this study, we developed and evaluated a single-round infectious particles (SRIPs) assay method and a cytopathic effect (CPE) assay method. The SRIPs assay showed a strong correlation (R = 0.94) with the focus-forming assay. The SRIPs assay method is not only simple and can be expected to improve accuracy, but also has the advantages of extremely low pathogenicity and safety for the operator. The CPE assay also showed a strong correlation (R = 0.80) with the focus-forming assay. It has the advantage of requiring almost no reagents and is easy to interpret. Both SRIPs and CPE assay methods can be used as alternatives to the focus-forming assay method, but further characterization of each method is necessary before one is selected for routine use.</div></div>","PeriodicalId":17663,"journal":{"name":"Journal of virological methods","volume":"336 ","pages":"Article 115163"},"PeriodicalIF":2.2,"publicationDate":"2025-04-08","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143807649","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}

引用次数: 0

High-dimensional droplet digital PCR of multiple hepatitis B splice variants in serum, liver, and tissue culture 血清、肝脏和组织培养中多种乙型肝炎剪接变异体的高维液滴数字PCR

IF 2.2 4区医学

Journal of virological methods Pub Date : 2025-04-08 DOI: 10.1016/j.jviromet.2025.115155

Tanner Grudda , Chenkai Jiang , Che-Min Lo , Nicole E. Skinner , Mark S. Sulkowski , Ashwin Balagopal , Chloe L. Thio

{"title":"High-dimensional droplet digital PCR of multiple hepatitis B splice variants in serum, liver, and tissue culture","authors":"Tanner Grudda , Chenkai Jiang , Che-Min Lo , Nicole E. Skinner , Mark S. Sulkowski , Ashwin Balagopal , Chloe L. Thio","doi":"10.1016/j.jviromet.2025.115155","DOIUrl":"10.1016/j.jviromet.2025.115155","url":null,"abstract":"<div><div>Hepatitis B virus (HBV) affects approximately 2 billion individuals; 254 million have chronic infection. The roles of spliced HBV (spHBV) RNAs in human infection are poorly understood partly due to limitations in quantitative methods. We designed and multiplexed long amplicon (0.3–2kbp) ddPCR assays that simultaneously quantify the most frequently observed spHBV isoforms (sp1, sp2, sp3, sp8, sp9) alongside unspliced pregenomic RNA (pgRNA). Our innovations include the use of ddPCR, long amplicons, and high-dimensional multiplexing. We tested primers and probes using qPCR on synthetic DNA targets to identify a high-fidelity combination recognizing HBV genotype A viruses. Using common forward and reverse primers we accounted for PCR efficiency differences between amplicons of differing size. We altered denaturation and annealing temperatures to enrich spHBV, the minority population of total HBV RNA. To achieve higher-order multiplexing, we increased the number of cycles to 80, added a post-reverse transcription cleanup step, and tested > 360 oligo concentration combinations. We performed spatial transcriptomic sequencing on liver tissue to confirm splice junctions measured by ddPCR. We successfully quantified spHBV and pgRNA in serum, liver, and HBV-infected HepG2<sup>NTCP</sup> cells. Our assay enables direct, sensitive quantification of spHBV and full-length pgRNA from serum, liver, and tissue culture samples, which will facilitate studies on the impact of spHBV on HBV infection outcomes in people. This technique has potential applications in HBV research and therapy development, with broader implications in virology and oncology.</div></div>","PeriodicalId":17663,"journal":{"name":"Journal of virological methods","volume":"336 ","pages":"Article 115155"},"PeriodicalIF":2.2,"publicationDate":"2025-04-08","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143859103","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}

引用次数: 0