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Journal of Virological Methods

Syed Shariq Nazir Qadiri, Soo-Jin Kim, Rahul Krishnan, Jae-Ok Kim, Wi-Sik Kim, Myung-Joo Oh
An in situ hybridization (RNA-ISH) assay has been developed and optimized to detect viral haemorrhagic septicemia virus (VHSV), an OIE listed piscine rhabdovirus, in infected fish cells using fathead minnow (FHM) as a model cell line. Two antisense riboprobes (RNA probes) targeting viral transcripts from a fragment of nucleoprotein (N) and glycoprotein (G) genes were generated by reverse transcription polymerase chain reaction (RT-PCR) using VHSV specific primers followed by a transcription reaction in the presence of digoxigenin dUTP...
November 8, 2018: Journal of Virological Methods
Nikolay Yu Saushkin, Jeanne V Samsonova, Alexander P Osipov, Sergey E Kondakov
We recently proposed a new so-called strip-dried format aimed for convenient use of dried biomaterial in diagnostic purposes. In this work, 334 blood samples obtained in strip-dried form were used for bovine leucosis analysis with ELISA and real-time PCR methods. High percentage of seropositive animals (18.3%) let us estimate both indirect (serological) and direct methods applicability for the analysis of strip-dried blood samples and also to compare them (PCR results concurred with ELISA in 93.4% cases). Parallel analysis of native and corresponding strip-dried samples approved the proposed format as a reliable analytical way of sampling being in 100% concordance with conventional serum/whole blood ELISA and PCR analysis...
November 6, 2018: Journal of Virological Methods
Maëlle Bochud, Wolfram Schäfer, Nathan J Roth, Carlos Ros
Hepatitis E virus (HEV) is an emerging concern for the safety of plasma-derived medicinal products. The lack of an efficient cell culture system hampers the studies on HEV biology as well as validation studies to test the capacity of virus reduction steps to clear HEV. Hence, a surrogate hepevirus that can efficiently replicate in cell culture is needed. Cutthroat trout virus (CTV) is a non-pathogenic fish hepevirus, which can replicate in cell culture to high titers. Under interferon inhibition, CTV replication reached up to 5 × 107 genome equivalents per µL in 4-5 days...
November 3, 2018: Journal of Virological Methods
Chong Zhao, Feng Sun, Xuejuan Li, Ying Lan, Linlin Du, Tong Zhou, Yijun Zhou
Rice black-streaked dwarf virus (RBSDV) infects rice plants, a major crop, and is transmitted via the small brown planthopper (SBPH: Laodelphax striatellus Fallén), causing significant economic loss in China. To rapidly diagnose RBSDV, a reverse transcription-recombinase polymerase amplification (RT-RPA) method was developed using P10 virus-specific primers and probes. Detection of terminally labeled amplification products was achieved with the lateral flow strip method. Our results demonstrate that RT-RPA and RT-PCR assays offer similar sensitivity and specificity in RBSDV detection using cDNA as template...
November 2, 2018: Journal of Virological Methods
Bingyu Yao, Yongcan Lu, Liu Li, Yanxiang Wang, Qingyu Li, Guoping Wang, Ni Hong
A highly sensitive nested multiplex reverse transcription-polymerase chain reaction (nmRT-PCR) assay was developed for the simultaneous detection of Apple chlorotic leaf spot virus (ACLSV), Apple stem grooving virus (ASGV) and Apple stem pitting virus (ASPV) infecting pear trees. In the assay, a set of three forward primers specific to each of the three viruses and a universal reverse primer was used as external primers in the first-round PCR, which was followed by a second-round PCR developed previously. The nmRT-PCR assay was 104 times more sensitive than conventional mRT-PCR assay in detecting the three viruses in in vitro pear plantlets...
October 29, 2018: Journal of Virological Methods
Christopher C Marohnic, Larry G Birkenmeyer, M Felicia Bogdan, Edwin C Frias, Kathy S Otis, Mary Ann P Palafox, Troy D McSherry, Svetoslava D Gregory, Cheng Zhao, Robin A Gutierrez, John C Prostko, A Scott Muerhoff
Hepatitis C Virus c33, a recombinant protein comprising residues 1192-1457 of NS3 helicase, has been a mainstay of HCV serology for decades. With seven unpaired cysteines, seroreactivity of E. coli expressed c33 is dependant on reductants. While engineering a c33 replacement for new anti-HCV serological tests, we sought to reduce oxidation sensitivity, a liability for immunodiagnostic reagent stability. A series of cysteine-to-serine substituted variants of a c33-like antigen was constructed and evaluated for reactivity against a panel of HCV-positive sera...
October 28, 2018: Journal of Virological Methods
Zhiyun Wen, Mike Citron, Andrew J Bett, Amy S Espeseth, Kalpit A Vora, Lan Zhang, Daniel J DiStefano
Viral plaque assays are important tools in the development and evaluation of new antiviral drugs or vaccines in both preclinical and clinical research. While plaque assays are the standard tools to measure infectious virus, the methodology is time-consuming and requires experience in recognizing plaques. The assays are also prone to variation among analysts due to plaque recognition and manual counting errors. Here we describe the development of two simplified plaque assays for measuring RSV virus titers and anti-RSV antibody neutralization titers using 96 well plate formats...
October 28, 2018: Journal of Virological Methods
Sören Hansen, Veronika Dill, Mohamed A Shalaby, Michael Eschbaumer, Susanne Böhlken-Fascher, Bernd Hoffmann, Claus-Peter Czerny, Ahmed Abd El Wahed
Foot-and-mouth disease virus (FMDV), belonging to the family of Picornaviridae, infects mostly cloven-hoofed animals and leads to huge economic losses. Since there is no cross-protection between the seven serotypes of FMDV, effective vaccination relies on the knowledge of the serotype causing the outbreak. The most common methods of serotyping are antigen ELISAs and amplification-based sequencing. Serotype-specific PCR methods exist but have limitations due to emerging mutants within serotypes. Sequencing is a promising technology, but currently suffers from cumbersome procedures and long turnaround times...
October 26, 2018: Journal of Virological Methods
Vu Duc Canh, Hideki Osawa, Kentaro Inoue, Ikuro Kasuga, Satoshi Takizawa, Hiroaki Furumai, Hiroyuki Katayama
Molecular assays are currently the most widely used method to quantify pathogenic viruses in water; however, their performance is often disrupted by matrices present in environmental samples. The present study used ferrihydrite (Fh) treatment to mitigate inhibition of RT-qPCR virus detection from environmental water concentrates. Fh treatment was performed to improve the detection of spiked Aichi virus 1 (AiV) and Qβ bacteriophage (Qβ) in commercial humic acids and seawater concentrates. The optimal Fh doses were found to be 200, 500, and 1000 mgFe/L for humic acid concentrates at UV254 of 1...
October 25, 2018: Journal of Virological Methods
Wildeberg Cál Moreira, Jéssica F S Freitas, Nathalia S Machado, Antônio Eugênio Castro Cardoso Almeida, Wlamir Corrêa de Moura
It is mandatory to ensure the quality of biological products used in the prevention of rabies, a zoonosis with nearly 100% lethality. Fifteen million people receive post-exposure prophylaxis yearly. The vaccine batches are assessed by the National Institutes of Health (NIH) test which has several disadvantages such as significant variability and animal welfare issues. The estimation of immunogenicity based on titration of neutralizing antibodies (NA) is not applied to the human vaccine yet. Despite this, a satisfactory concentration of NA (0...
October 25, 2018: Journal of Virological Methods
M M M Ferro, R Ramos-Sobrinho, C A D Xavier, F M Zerbini, G S A Lima, T Nagata, I P Assunção
Viruses belonging to the genus Begomovirus (family Geminiviridae) have circular single-strand DNA genomes encapsidated into quasi-icosahedral particles, and are transmitted by whiteflies of the Bemisia tabaci complex. Biological and molecular properties of begomoviruses have been studied efficiently with infectious clones containing dimeric genomic components. However, current approaches employing enzymatic digestion and ligation to binary vectors are laborious, mostly due to many cloning steps or partial digestion by restriction enzyme...
October 23, 2018: Journal of Virological Methods
Prabhu Narayan Meena, Lalit Laxman Kharbikar, Rajeev Singh Rana, Subrata Satpathy, Arti Shanware, Palaiyur Nanjappan Sivalingam, Shweta Nandanwar
A loop-mediated isothermal amplification (LAMP) assay was optimized for the detection of Mesta yellow vein mosaic virus (MeYVMV) in diseased plants of mesta (Hibiscus sabdariffa L.& H. cannabinus L.). The LAMP assay was optimized using a set of six primers targeting the MeYVMV genome and could be completed in 30-60 min at 63 °C. The LAMP amplification results were visualized by adding 1 μl of hydroxy naphthol blue (HNB) dye in a 25 μl LAMP reaction mixture prior to amplification as well as by electrophoresis...
October 22, 2018: Journal of Virological Methods
Fan Ming, Yanqing Cheng, Chenwei Ren, Sizhu Suolang, Hongbo Zhou
H9N2 avian influenza virus is threatening animals and public health systems. Effective diagnosis is imperative to control the disease. Thus, we developed a panel of monoclonal antibodies (Mabs) against the H9N2 avian influenza virus (AIV) and implemented a double-antibody sandwich enzyme-linked immunosorbent assay (DAS-ELISA) to detect the H9 viral antigen. Hybridomas 4D10 and 5G2 were screened to secrete immunoglobulin G (IgG) and IgA, respectively. Antibody 4D10 was used as the capture antibodies and HRP labeled 5G2 as the detector antibody...
October 21, 2018: Journal of Virological Methods
Ramya Bhatia, June Stewart, Sharon Moncur, Heather Cubie, Kavanagh Kim, Kevin Gj Pollock, Camille Busby-Earle, Alistair Rw Williams, Sarah Howie, Kate Cuschieri
Measuring anti-HPV antibody levels is important for surveillance of the immunological response to both natural infection and vaccination. Here, an ELISA test for measurement of HPV-16L1 antibodies was developed and validated in sera and dried blood spots. An in-house ELISA was developed for measuring anti-HPV-16L1 IgA and IgG levels. The assay was standardized against WHO international standard serum and validated on serum, dried blood spots and cervical liquid based cytology samples from women attending colposcopy clinics in Scotland...
October 20, 2018: Journal of Virological Methods
Karen Delgadillo-Gutiérrez, Rosa María Ribas-Aparicio, Alicia Jiménez-Alberto, Gerardo Aparicio-Ozores, Juan A Castelán-Vega
Retroviral pseudotypes are broadly used as safe instruments to mimic the structure and surface of highly pathogenic viruses. They have been employed for the discovery of new drugs, as diagnostic tools in vaccine studies, and part of serological assays. Because of their widespread use in research and their potential as tools for quality control, it is important to know their shelf life, stability, and best storage conditions. In this study, we produced pseudotypes carrying the lacZ reporter gene and the hemagglutinin (HA) of avian influenza virus subtypes H5 and H7 to investigate their stability under various storage conditions...
October 19, 2018: Journal of Virological Methods
Veera Kainulainen, Sonja Elf, Petri Susi, Minna Mäki, Anne Pitkäranta, Janne O Koskinen, Riitta Korpela, Kevin E Eboigbodin
BACKGROUND: Rhinovirus (RV), a major cause of respiratory infection in humans, imposes an enormous economic burden due to the direct and indirect costs associated with the illness. Accurate and timely diagnosis is crucial for deciding the appropriate clinical approach and minimizing unnecessary prescription of antibiotics. Diagnosis of RV is extremely challenging due to genetic and serological variability among its numerous types and their similarity to enteroviruses. OBJECTIVE: We sought to develop a rapid nucleic acid test that can be used for the detection of Rhinovirus within both laboratory and near patient settings...
October 19, 2018: Journal of Virological Methods
Anne J Jääskeläinen, Essi M Korhonen, Eili Huhtamo, Maija Lappalainen, Olli Vapalahti, Hannimari Kallio-Kokko
The laboratory confirmation of Zika virus (ZIKV) infection, and the differential diagnosis from other flavivirus infections such as dengue virus (DENV), often requires the use of several diagnostic test types. Cross-reactions and secondary infections complicate the serological diagnosis and specific viral RNA detection assays are often needed for confirming the diagnosis. The aim of this study was to validate serological and molecular methods for diagnosing ZIKV infection. This included the evaluation of a ZIKV RT-qPCR assay for diagnostics that was previously set up for research use and to compare the ZIKV, DENV and TBEV EIA methods...
October 18, 2018: Journal of Virological Methods
Gaopeng Liu, Yonghou Jiang, Tanja Opriessnig, Keda Gu, Hongli Zhang, Zongqi Yang
Porcine viral diarrhea, mainly caused by porcine epidemic diarrhea virus (PEDV), transmissible gastroenteritis virus (TGEV), porcine group A rotaviruses (RVA), porcine group C rotaviruses (RVC) and porcine circovirus 2 (PCV2), is a serious global problem, resulting in substantial economic losses to the swine industry. For fast and reliable diagnosis of the causative agent associated with viral diarrhea in pigs, an inexpensive and easy to perform gel-based multiplex PCR assay was developed in this study to detect and differentiate the different viruses by amplicon size...
October 15, 2018: Journal of Virological Methods
Alamira Marzouk Fouad, Hatem Soliman, Ebtsam S H Abdallah, Sherif Ibrahim, Mansour El-Matbouli, Ahmad A Elkamel
Spring viremia of carp, a fatal viral disease, is caused by the spring viremia of carp virus (SVCV) and can result in up to 70% mortalities in common carps and significant economic losses in several other cyprinid aquaculture. The present study aimed to investigate the possible control of SVCV replication in Epithelioma papulosum cyprini (EPC) cells using the RNA interference technology targeting the RNA-dependent RNA polymerase (L) gene of the SVCV that is essential for its replication. Three stealth small interfering RNA (siRNA) sequences were designed to target three different regions on the SVCV-L gene...
October 15, 2018: Journal of Virological Methods
Jordan Thomas, Alessandra Ruggiero, Francesco A Procopio, Giuseppe Pantaleo, William A Paxton, Georgios Pollakis
HIV-1 infection cannot be cured due to the presence of the latent reservoir (LR). Novel cure or treatment strategies, such as "shock and kill" or therapeutic vaccination, aim to reduce or eradicate the LR. Cure strategies utilise robust DNA quantification assays to measure the change in the LR in low copy scenarios. No standard assay exists, which impedes the reliable comparison of results from different therapy and vaccine trials and HIV-1 total DNA quantification methods have not been previously compared...
October 13, 2018: Journal of Virological Methods
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