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Journal of Virological Methods

M J Ankcorn, S Ijaz, B Haywood, J Neuberger, A M ElSharkawy, J Maggs, R S Tedder
Genotype 3 hepatitis E virus (HEV) can lead to persistent infections in immunocompromised hosts. A recently available commercial assay for the detection of HEV antigen (HEV-Ag ELISA, Wantai diagnostics) may enable the study of HEV-Ag dynamics in such persistent infections, however currently there is no confirmatory test available. We generated a putative neutralising reagent from a pool of four convalescent blood donor samples and explored neutralising activity against HEV antigens from clinical samples, HEV tissue-culture and virus-like particles...
November 17, 2017: Journal of Virological Methods
Viska I Iskandar, Yutaka Sasaki, Naoto Yoshino, Raden Z R Abubakar, Shigehiro Sato, Yasushi Muraki
A cell-based vaccine production method for influenza virus may be an effective and more rapid alternative to egg-based systems. For high-yield virus production, the effect of bovine, porcine, fungal, and recombinant trypsins on influenza A/H1N1 virus replication in MDCK SI-6 cells (SI-6 cells), a novel MDCK cell line developed by our research group, was examined. SI-6 cells infected with influenza A/H1N1 virus were incubated in the presence of four trypsin types at various concentrations, and virus yields in the culture medium were evaluated by a hemagglutination (HA) assay...
November 16, 2017: Journal of Virological Methods
Cédric Charretier, Aure Saulnier, Loïc Benair, Corinne Armanet, Isabelle Bassard, Sandra Daulon, Bertrand Bernigaud, Emanuel Rodrigues de Sousa, Clémence Gonthier, Edouard Zorn, Emmanuelle Vetter, Claire Saintpierre, Patrice Riou, David Gaillac
The classical cell-culture methods, such as cell culture infectious dose 50% (CCID50) assays, are time-consuming, end-point assays currently used during the development of a viral vaccine production process to measure viral infectious titers. However, they are not suitable for handling the large number of tests required for high-throughput and large-scale screening analyses. Impedance-based bio-sensing techniques used in real-time cell analysis (RTCA) to assess cell layer biological status in vitro, provide real-time data...
November 14, 2017: Journal of Virological Methods
Michelle Wille, Hong Yin, Åke Lundkvist, Juan Xu, Shaman Muradrasoli, Josef D Järhult
Surveillance of wild birds is critical in monitoring for highly pathogenic avian influenza A viruses (AIVs). However, a successful surveillance regime requires proper treatment of samples in the field - rapid placement of samples in -80°C and subsequent maintenance of cold-chain. Given the logistical difficulties of this, many avian taxa and/or geographic locations are not sampled, or, when sampled may result in false negatives due to poor sample treatment in the field. Here, we assessed the utility of RNAlater(®) as a stabilization agent for AIV sampling...
November 10, 2017: Journal of Virological Methods
Junli Liu, Lu Yao, Feifei Zhai, Yuqing Chen, Jing Lei, Zhenwei Bi, Jianhua Hu, Qian Xiao, Suquan Song, Liping Yan, Jiyong Zhou
Avian influenza virus (AIV), especially subtypes H5, H7 and H9, has contributed to enormous economic losses and poses a potential pandemic threat to global human public health. Early screening of suspected cases is key to controlling the spread of AIVs. In this study, an accurate, rapid, and triplex real-time polymerase chain reaction (PCR) assay was developed for the simultaneous detection of AIV subtypes H5, H7 and H9. The sensitivity of the real-time PCR was at least 100 times higher than that of the conventional PCR, with a detection limit of 50 copies and an EID50 of 1 (50% egg infections dose) for the H5, H7, and H9 subtypes...
November 9, 2017: Journal of Virological Methods
Frank Gillam, Jianqiang Zhang, Chenming Zhang
Porcine epidemic diarrhea Virus (PEDV) is the causative agent of porcine epidemic diarrhea, which is a devastating viral disease and causes severe economic loss to the swine industry. Current vaccine options for PEDV include modified live viruses and killed live viruses. Though these vaccines have shown efficacy, some have side effects including viral shedding. This report details an E. coli based expression and purification process of multiple vaccine candidates for PEDV using Hepatitis B Core Antigen (HBcAg) as a backbone protein...
November 9, 2017: Journal of Virological Methods
Wayne Dimech, Liza M Cabuang, Hans-Peter Grunert, Vanessa Lindig, Vivienne James, Brigitte Senechal, Giuseppe A Vincini, Heinz Zeichhardt
Quantification of Cytomegalovirus (CMV) DNA is required for the initiation and monitoring of anti-viral treatment and the detection of viral resistance. However, due to the lack of standardisation of CMV DNA nucleic acid tests, it is difficult to set universal thresholds. In 2010, the 1st WHO International Standard for Human Cytomegalovirus for Nucleic Acid Amplification Techniques was released. Since then CMV DNA viral load assays have been calibrated using this standard. Three external quality assessment (EQA) providers sent the same five samples to their participants and analysed the results to determine the equivalence of reporting CMV DNA results in international units per millilitre (IU/mL), and compared the difference in results reported in IU/mL with those reported in copies per millilitre (c/mL), and to determine the rate of adoption of IU/mL...
November 7, 2017: Journal of Virological Methods
Meruyert A Saduakassova, Akhmetzhan A Sultanov, Lespek B Kutumbetov, Jemma Wadsworth, Britta A Wood, Nick J Knowles, Donald P King, Katarzyna Bachanek-Bankowska
A new lineage of foot-and-mouth disease virus (FMDV), called A/ASIA/G-VII, emerged from the Indian subcontinent in 2015 and continues to spread in Western Asia. Currently, the distribution of viruses belonging to this lineage is defined using sequencing approaches, but other cheaper and faster diagnostic methods are urgently needed. Thus, this study describes the development and validation of a novel A/ASIA/G-VII lineage-specific real-time RT-PCR (rRT-PCR). Diagnostic sensitivity and specificity were evaluated using representative field specimens and isolates from the A/ASIA/G-VII lineage, as well as samples comprising other FMDV lineages that co-circulate in Asia (n=54)...
November 4, 2017: Journal of Virological Methods
Tuck-Weng Kok, Maurizio Costabile, Gregory A Tannock, Peng Li
Inhibition of viral replication by icIgA antibodies has only been observed with in vitro studies using epithelial cell lines in transwell cultures. This effect appears to involve an interaction between polymeric immunoglobulin A (pIgA) and viral particles within an intracellular compartment, since IgA is transported across polarized cells. Polyclonal guinea pig antisera against purified influenza A virus and mouse antisera prepared against Influenza A/H3N2 hemagglutinin (HA0) cleavage loop peptides, were used in confocal fluorescence microscopy to show specific staining of wild-type influenza H1N1 and H3N2 viruses in clinical specimens...
November 2, 2017: Journal of Virological Methods
Wesley Jun, Ran Hu, Laura Hyland, Darren Crandall, Pradeep Ramachandran, Chinmay Pangarkar, Sharada Sivaraman, Babak Haghiri
Herpes simplex virus type-2 (HSV-2) specific glycoprotein G (gG-2) is widely used as the antigen of choice for serodiagnosis of HSV-2. In order to develop an ELISA for serodetection of HSV-2 IgG in patient sera, the soluble form of the mature gG-2 antigen (mgG-2), gG283-649, was expressed using a baculovirus expression system. gG283-649 contains the complete extracellular domain of mgG-2 including the C-terminal region, which despite homology to gG-1, does not cross-react with HSV-1 antibodies present in HSV-1 positive patient sera...
October 28, 2017: Journal of Virological Methods
Li Mao, Wenliang Li, Tianci Zhou, Leilei Yang, Fei Hao, Jizong Li, Wenwen Zhang, Xuenong Luo, Jieyuan Jiang
Caprine parainfluenza virus type 3 (CPIV3) is a novel pathogen mainly causing respiratory diseases in goats. At present, there are no high throughput and rapid testing methods available for epidemiological investigation. In this study, we designed a modified method for selection of hybridomas that secrete monoclonal antibodies (mAb) specific for CPIV3. The monoclonal antibodies were obtained by combination of indirect enzyme-linked immunosorbent assay (iELISA) and blocking ELISA (bELISA). The technique was efficient to determine each mAb with specificity and sensitivity...
September 29, 2017: Journal of Virological Methods
Haciba Moudjahed, Claire Pinçon, Kazali Alidjinou, Anny Dewilde, Anne Goffard
Three molecular assays (FTD(®) Viral GE from Fast-track diagnostics, RIDA(®)GENE VSP1 from R-Biopharm, and Xpert Norovirus from Cepheid) were compared for virus detection in acute diarrhea samples. RIDA(®)GENE and FTD(®) Viral GE showed perfect/almost perfect agreement for Rotavirus, Sapovirus and Norovirus, substantial agreement for Adenovirus, and moderate agreement for Astrovirus.
September 29, 2017: Journal of Virological Methods
Dan Rao, Miaoli Wu, Jing Wang, Wen Yuan, Yujun Zhu, Feng Cong, Fengjiao Xu, Yuexiao Lian, Bihong Huang, Qiwen Wu, Meili Chen, Yu Zhang, Ren Huang, Pengju Guo
Murine parvovirus is one of the most prevalent infectious pathogens in mouse colonies. A specific primer pair targeting the VP2 gene of minute virus of mice (MVM) and mouse parvovirus (MPV) was utilized for high resolution melting (HRM) analysis. The resulting melting curves could distinguish these two virus strains and there was no detectable amplification of the other mouse pathogens which included rat parvovirus (KRV), ectromelia virus (ECT), mouse adenovirus (MAD), mouse cytomegalovirus (MCMV), polyoma virus (Poly), Helicobactor hepaticus (H...
September 23, 2017: Journal of Virological Methods
Charith Raj Adkar-Purushothama, Pierrick Bru, Jean-Pierre Perreault
5' RNA ligase-mediated rapid amplification of cDNA ends (5' RLM-RACE) is a widely-accepted method for the validation of direct cleavage of a target gene by a microRNA (miRNA) and viroid-derived small RNA (vd-sRNA). However, this method cannot be used if cleavage takes place in the 3' extremity of the target RNA, as this gives insufficient sequence length to design nested PCR primers for 5' RLM RACE. To overcome this hurdle, we have developed 3' RNA ligase-mediated rapid amplification of cDNA ends (3' RLM RACE)...
September 22, 2017: Journal of Virological Methods
Eva Guijarro-Pardo, Silvia Gómez-Sebastián, José M Escribano
Trichoplusia ni insect larvae infected with vectors derived from the Autographa californica multiple nucleopolyhedrovirus (AcMNPV), are an excellent alternative to insect cells cultured in conventional bioreactors to produce recombinant proteins because productivity and cost-efficiency reasons. However, there is still a lot of work to do to reduce the manual procedures commonly required in this production platform that limit its scalability. To increase the scalability of this platform technology, a current bottleneck to be circumvented in the future is the need of injection for the inoculation of larvae with polyhedrin negative baculovirus vectors (Polh-) because of the lack of oral infectivity of these viruses, which are commonly used for production in insect cell cultures...
September 22, 2017: Journal of Virological Methods
Hye-Ryung Kim, Yu-Ri Park, Da-Rae Lim, Min-Ji Park, Ji-Young Park, Seong-Hee Kim, Kyoung-Ki Lee, Young S Lyoo, Choi-Kyu Park
A multiplex quantitative real-time polymerase chain reaction (mqPCR) assay was developed for the rapid and differential detection of porcine circovirus 2 (PCV2) and PCV3. Each of the capsid genes of PCV2 and PCV3 were amplified using specific primers and probe sets, while no other porcine pathogen genes were detected. Limit of detection of the assay was below 50 copies of the target genes of PCV2 and PCV3, and was comparable to that of previously described methods The assay showed high repeatability and reproducibility, with coefficients of intra-assay and inter-assay variation of less than 4...
September 21, 2017: Journal of Virological Methods
Ashish Warghane, Pragati Misra, Sumit Bhose, Kajal Kumar Biswas, Ashwani Kumar Sharma, M Krishna Reddy, Dilip Kumar Ghosh
Tristeza is a devastating disease of citrus and reported to be present in almost all countries where it is cultivated as a commercial crop. The etiological agent of this disease is Citrus tristeza virus (CTV), a member of the genus Closterovirus with in the family Closteroviridae. The pathogen is restricted to the phloem tissue of the infected citrus plant and has a monopartite ss (+) RNA genome of ∼20kb size. Till date, there is no effective control measure available for this virus. Management of tristeza depends on destruction of CTV infected field plants, production of virus-free planting material for new orchard establishment and controlling viruliferous aphid vectors responsible for field spread of the pathogen...
September 21, 2017: Journal of Virological Methods
J E Burton, L Easterbrook, J Pitman, D Anderson, S Roddy, D Bailey, R Vipond, C B Bruce, A D Roberts
The 2014 Ebola outbreak in West Africa required the rapid testing of clinical material for the presence of potentially high titre Ebola virus (EBOV). Safe, fast and effective methods for the inactivation of such clinical samples are required so that rapid diagnostic tests including downstream analysis by RT-qPCR or nucleotide sequencing can be carried out. One of the most commonly used guanidinium - based denaturing agents, AVL (Qiagen) has been shown to fully inactivate EBOV once ethanol is added, however this is not compatible with the use of automated nucleic acid extraction systems...
September 20, 2017: Journal of Virological Methods
Emily S Bailey, Matthew Price, Lisa M Casanova, Mark D Sobsey
Somatic and F+ coliphages have been identified and validated as virus indicators of fecal contamination in ground water by US EPA and more recently they are being considered for use in managing both marine and fresh recreational water and wastewater discharges. Studies documenting their usefulness as viral indicator in reclaimed water sources in the USA are limited. However, simultaneous detection of both somatic and F+ coliphages on a single E. coli host is preferred over their separate analysis because both are abundant in wastewater, they may respond differently to wastewater reclamation treatment processes, and separate analysis for each group in separate host bacteria adds complexity and cost...
September 20, 2017: Journal of Virological Methods
Yang Liu, Andrea L Cathcart, William E Delaney, Kathryn M Kitrinos
The COBAS TaqMan assay has a lower limit of quantification (LLOQ) of 169 HBV copies/mL and a lower limit of detection (LLOD) of 58 copies/mL. HBV DNA below the TaqMan LLOQ is classified as target not detected (TND) (<58 copies/mL) or target detected (TD) (between 58 and 169 copies/mL). Here we have developed a more sensitive digital droplet PCR (ddPCR) assay to evaluate the impact of long-term tenofovir disoproxil fumarate (TDF) treatment in patients that did or did not achieve HBsAg seroconversion. A ddPCR assay was developed to detect HBV DNA to 8 copies/mL...
September 18, 2017: Journal of Virological Methods
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