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Journal of Virological Methods

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https://www.readbyqxmd.com/read/27915037/quantification-of-pea-enation-mosaic-virus-1-and-2-during-infection-of-pisum-sativum-by-one-step-real-time-rt-pcr
#1
Juliette Doumayrou, Melissa Sheber, Bryony C Bonning, W Allen Miller
Pea enation mosaic virus 1 (PEMV1) and Pea enation mosaic virus 2 (PEMV2) are two viruses in an obligate symbiosis that cause pea enation mosaic disease mainly in plants in the Fabaceae family. This virus system is a valuable model to investigate plant virus replication, movement and vector transmission. Thus, here we describe growth conditions, virus detection methods, and virus accumulation behavior. To measure the accumulation and movement of PEMV1 and PEMV2 in plants during the course of infection, we developed a quantitative real-time one-step reverse transcription PCR procedure using the SYBR-green(®) technology...
November 30, 2016: Journal of Virological Methods
https://www.readbyqxmd.com/read/27915036/a-rapid-assay-for-detection-of-rose-rosette-virus-using-reverse-transcription-recombinase-polymerase-amplification-using-multiple-gene-targets
#2
Binoy Babu, Brian K Washburn, Steven H Miller, Kristina Poduch, Tulin Sarigul, Gary W Knox, Francisco M Ochoa-Corona, Mathews L Paret
Rose rosette disease caused by Rose rosette virus (RRV; genus Emaravirus) is the most economically relevant disease of Knock Out(®) series roses in the U.S. As there are no effective chemical control options for the disease, the most critical disease management strategies include the use of virus free clean plants for propagation and early detection and destruction of infected plants. The current diagnostic techniques for RRV including end-point reverse transcription-polymerase chain reaction (RT-PCR) and real-time PCR (RT-qPCR) are highly sensitive, but limited to diagnostic labs with the equipment and expertise; and is time consuming...
November 30, 2016: Journal of Virological Methods
https://www.readbyqxmd.com/read/27899288/one-step-reverse-transcription-loop-mediated-isothermal-amplification-for-the-detection-of-maize-chlorotic-mottle-virus-in-maize
#3
Ling Chen, Zhiyuan Jiao, Dongmei Liu, Xingliang Liu, Zihao Xia, Congliang Deng, Tao Zhou, Zaifeng Fan
Maize chlorotic mottle virus (MCMV) is spreading in many regions worldwide, causing maize lethal necrosis when co-infected with a potyvirid. In this study, one-step reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay was developed to detect MCMV in maize. A set of four specific primers was designed based on the conserved coat protein gene sequences of MCMV. The RT-LAMP could be completed within 60min under isothermal condition at 63°C. The sensitivity test showed that the RT-LAMP was about 10-fold more sensitive than RT-PCR and no cross-reactivity was detected with other viral pathogens infecting maize in China...
November 26, 2016: Journal of Virological Methods
https://www.readbyqxmd.com/read/27894862/hiv-drug-resistance-testing-among-patients-failing-second-line-antiretroviral-therapy-comparison-of-in-house-and-commercial-sequencing
#4
Benjamin Chimukangara, Bhavini Varyani, Tinei Shamu, Junior Mutsvangwa, Justen Manasa, Elizabeth White, Cleophas Chimbetete, Ruedi Luethy, David Katzenstein
INTRODUCTION: HIV genotyping is often unavailable in low and middle-income countries due to infrastructure requirements and cost. We compared genotype resistance testing in patients with virologic failure, by amplification of HIV pol gene, followed by "in-house" sequencing and commercial sequencing. METHODS: Remnant plasma samples from adults and children failing second-line ART were amplified and sequenced using in-house and commercial di-deoxysequencing, and analyzed in Harare, Zimbabwe and at Stanford, U...
November 25, 2016: Journal of Virological Methods
https://www.readbyqxmd.com/read/27894861/development-of-a-competitive-double-antibody-lateral-flow-assay-for-the-detection-of-antibodies-specific-to-glycoprotein-b-of-aujeszky-s-disease-virus-in-swine-sera
#5
V V Vrublevskaya, V N Afanasyev, A A Grinevich, Yu Y Skarga, P P Gladyshev, S A Ibragimova, D V Krylsky, O S Morenkov
Three lateral flow assays (LFAs) for the detection of antibodies against glycoprotein B (gB) of Aujeszky's disease virus (ADV) in swine sera: a competitive double antibody sandwich LFA without a preincubation step (CDAS-gB-LFA), a CDAS-gB-LFA with a preincubation step (pCDAS-gB-LFA), and a competitive direct gB-LFA have been developed and were compared with each other and with a gB-ELISA. The assays are based on monoclonal antibodies to immunodominant epitopes of ADV gB. The pCDAS-gB-LFA proved to be the most specific and sensitive assay to detect antibodies directed to ADV gB...
November 25, 2016: Journal of Virological Methods
https://www.readbyqxmd.com/read/27889565/a-terminal-antibody-method-based-on-multiple-factors-that-influence-elisa-results-for-measurement-of-antibody-affinity-in-clinical-specimens
#6
Congcong Shang, Zhen Wang, Hui Liu
OBJECTIVE: To establish a new method for the measurement of antibody affinity in clinical samples. METHODS: Serial dilutions of antiserum samples were prepared to find the threshold concentration of antibody separating detectable from negative ELISA results. This threshold concentration was defined as the terminal antibody (TA) concentration, and a new method for measuring antibody affinity based on the effect of multiple factors that influence ELISA results at TA concentration was established, which we called the TA method...
November 23, 2016: Journal of Virological Methods
https://www.readbyqxmd.com/read/27867046/efficient-production-of-an-avian-adeno-associated-virus-vector-using-insect-cell-baculovirus-expression-system
#7
Anping Wang, Yongjuan Wang, Shuang Wu, Weiyong Zuo, Changming Guo, Weiming Hong, Shanyuan Zhu
Recombinant avian adeno-associated virus (rAAAV) is a promising gene transfer vector for avian cells. Although rAAAV can be produced by co-transfection of HEK293 cells with three plasmids, both scalability and productivity of the transient transfection method can not meet the demand for large-scale in vivo experiments. In this study, a scalable rAAAV production method was established by using insect cell/baculovirus expression system. Three recombinant baculoviruses, namely BacARep, BacAVP and BacAGFP, were generated by transfection of Sf9 cells with the three plasmids expressing AAAV Rep genes, modified VP gene or the inverted terminal repeats-flanked green fluorescent protein (GFP) gene...
November 17, 2016: Journal of Virological Methods
https://www.readbyqxmd.com/read/27867045/minvar-a-rapid-and-versatile-tool-for-hiv-1-drug-resistance-genotyping-by-deep-sequencing
#8
Michael Huber, Karin J Metzner, Fabienne D Geissberger, Cyril Shah, Christine Leemann, Thomas Klimkait, Jürg Böni, Alexandra Trkola, Osvaldo Zagordi
Genotypic monitoring of drug-resistance mutations (DRMs) in HIV-1 infected individuals is strongly recommended to guide selection of the initial antiretroviral therapy (ART) and changes of drug regimens. Traditionally, mutations conferring drug resistance are detected by population sequencing of the reverse transcribed viral RNA encoding the HIV-1 enzymes target by ART, followed by manual analysis and interpretation of Sanger sequencing traces. This process is labor intensive, relies on subjective interpretation from the operator, and offers limited sensitivity as only mutations above 20% frequency can be reliably detected...
November 17, 2016: Journal of Virological Methods
https://www.readbyqxmd.com/read/27865749/in-vitro-functional-assessment-of-natural-hiv-1-group-m-vpu-sequences-using-a-universal-priming-approach
#9
Asa Rahimi, Gursev Anmole, Maribel Soto-Nava, Tania Escamilla-Gomez, Tristan Markle, Steven W Jin, Guinevere Q Lee, P Richard Harrigan, David R Bangsberg, Jeffrey Martin, Santiago Avila-Rios, Gustavo Reyes-Teran, Mark A Brockman, Zabrina L Brumme
The HIV-1 accessory protein Vpu exhibits high inter- and intra- subtype genetic diversity that may influence Vpu function and possibly contribute to HIV-1 pathogenesis. However, scalable methods to evaluate genotype/phenotype relationships in natural Vpu sequences are limited, particularly those expressing the protein in CD4+ T-cells, the natural target of HIV-1 infection. A major impediment to assay scalability is the extensive genetic diversity within, and immediately upstream of, Vpu's initial 5' coding region, which has necessitated the design of oligonucleotide primers specific for each individual HIV-1 isolate (or subtype)...
November 16, 2016: Journal of Virological Methods
https://www.readbyqxmd.com/read/27865748/a-novel-reverse-genetics-system-for-production-of-infectious-west-nile-virus-using-homologous-recombination-in-mammalian-cells
#10
Shintaro Kobayashi, Kentaro Yoshii, Minato Hirano, Memi Muto, Hiroaki Kariwa
Reverse genetics systems facilitate investigation of many aspects of the life cycle and pathogenesis of viruses. However, genetic instability in Escherichia coli has hampered development of a reverse genetics system for West Nile virus (WNV). In this study, we developed a novel reverse genetics system for WNV based on homologous recombination in mammalian cells. Introduction of the DNA fragment coding for the WNV structural protein together with a DNA-based replicon resulted in the release of infectious WNV...
November 16, 2016: Journal of Virological Methods
https://www.readbyqxmd.com/read/27856212/development-of-a-novel-rapid-micro-neutralization-elisa-for-the-detection-of-neutralizing-antibodies-against-chandipura-virus
#11
R G Damle, A A Patil, V S Bhide, S D Pawar, G N Sapkal, V P Bondre
BACKGROUND: Chandipura virus (CHPV) is a leading cause of acute encephalitis with high mortality in paediatric population in India. A micro-neutralization ELISA (MN-ELISA) assay was developed for the detection of neutralizing antibodies (Nab) against CHPV. This novel method gives read-out in the form of ELISA optical density (OD) values and has a shorter turn-around time (TAT) as compared to the conventional cytopathic effect (CPE)-based neutralization assay (MN-CPE). The assay was developed using an Indian strain of CHPV...
November 14, 2016: Journal of Virological Methods
https://www.readbyqxmd.com/read/27825854/establishment-and-validation-of-an-enzyme-linked-immunosorbent-assay-for-igg-antibody-against-cytomegalovirus-based-on-pp150-antigen
#12
Huang Xi, Li Jinjie, Ge Shengxiang, Li Tingdong, Wang Han, Guo Xiaoyi, Fu Tong-Ming, Zhang Jun
The development of HCMV vaccines for the prevention of congenital HCMV has been identified as a top priority by the Institute of Medicine (USA), and virus infection is an important endpoint for the efficacy evaluation of the candidate vaccines. However, it is technically difficult to capture the HCMV viremia or uremia in infected individuals because the viremia or uremia can be detected only transiently during acute infection, and most people who are infected are asymptomatic. Thus, it is much desired to develop a serological assay for effectively distinguishing anti-HCMV antibodies as a result of natural infection from those elicited by subunit antigen vaccination...
November 4, 2016: Journal of Virological Methods
https://www.readbyqxmd.com/read/27793644/rapid-and-sensitive-detection-of-zika-virus-by-reverse-transcription-loop-mediated-isothermal-amplification
#13
Xuan Wang, Fenggui Yin, Yuhai Bi, Gong Cheng, Jing Li, Lidan Hou, Yunlong Li, Baozhi Yang, Wenjun Liu, Limin Yang
BACKGROUND: Zika virus (ZIKV) is an arbovirus that recently emerged and has expanded worldwide, causing a global threat and raising international concerns. Current molecular diagnostics, e.g., real-time PCR and reverse transcription PCR (RT-PCR), are time consuming, expensive, and can only be deployed in a laboratory instead of for field diagnostics. OBJECTIVES: This study aimed to develop a one-step reverse transcription loop-mediated isothermal amplification (RT-LAMP) platform showing sensitivity, specificity, and more convenience than previous methods, being easily distributed and implemented...
October 25, 2016: Journal of Virological Methods
https://www.readbyqxmd.com/read/27756548/evaluation-of-an-integrated-cell-culture-rt-pcr-assay-to-detect-and-quantify-infectious-lymphocystis-disease-virus
#14
Estefania J Valverde, Juan J Borrego, Dolores Castro
The lymphocystis disease virus (LCDV), a member of the Iridoviridae family, infects a wide range of fish species including gilthead seabream (Sparus aurata L.), the most important species cultured in the Mediterranean. LCDV is difficult to propagate in cell culture and does not produce clear and consistent cytopathic effects (CPE), especially in samples collected from subclinically infected fish. An integrated cell culture reverse transcription-polymerase chain reaction (ICC-RT-PCR) assay, followed by dot-blot hybridization of the RT-PCR products, was developed to improve the detection of infectious LCDV...
October 15, 2016: Journal of Virological Methods
https://www.readbyqxmd.com/read/27751950/preparation-of-viral-samples-within-biocontainment-for-ultrastructural-analysis-utilization-of-an-innovative-processing-capsule-for-negative-staining
#15
Mitchell K Monninger, Chrystal A Nguessan, Candace D Blancett, Kathleen A Kuehl, Cynthia A Rossi, Scott P Olschner, Priscilla L Williams, Steven L Goodman, Mei G Sun
Transmission electron microscopy can be used to observe the ultrastructure of viruses and other microbial pathogens with nanometer resolution. In a transmission electron microscope (TEM), the image is created by passing an electron beam through a specimen with contrast generated by electron scattering from dense elements in the specimen. Viruses do not normally contain dense elements, so a negative stain that places dense heavy metal salts around the sample is added to create a dark border. To prepare a virus sample for a negative stain transmission electron microscopy, a virus suspension is applied to a TEM grid specimen support, which is a 3mm diameter fragile specimen screen coated with a few nanometers of plastic film...
October 14, 2016: Journal of Virological Methods
https://www.readbyqxmd.com/read/27751949/first-international-collaborative-study-to-evaluate-rabies-antibody-detection-method-for-use-in-monitoring-the-effectiveness-of-oral-vaccination-programmes-in-fox-and-raccoon-dog-in-europe
#16
M Wasniewski, I Almeida, A Baur, T Bedekovic, D Boncea, L B Chaves, D David, P De Benedictis, M Dobrostana, P Giraud, P Hostnik, I Jaceviciene, S Kenklies, M König, K Mähar, M Mojzis, S Moore, S Mrenoski, T Müller, E Ngoepe, M Nishimura, T Nokireki, N Pejovic, M Smreczak, B Strandbygaard, E Wodak, F Cliquet
The most effective and sustainable method to control and eliminate rabies in wildlife is the oral rabies vaccination (ORV) of target species, namely foxes and raccoon dogs in Europe. According to WHO and OIE, the effectiveness of oral vaccination campaigns should be regularly assessed via disease surveillance and ORV antibody monitoring. Rabies antibodies are generally screened for in field animal cadavers, whose body fluids are often of poor quality. Therefore, the use of alternative methods such as the enzyme-linked immunosorbent assay (ELISA) has been proposed to improve reliability of serological results obtained on wildlife samples...
October 14, 2016: Journal of Virological Methods
https://www.readbyqxmd.com/read/27751948/a-loop-mediated-isothermal-amplification-assay-for-rapid-and-sensitive-detection-of-bovine-papular-stomatitis-virus
#17
Yohei Kurosaki, Sayaka Okada, Sayuri Nakamae, Jiro Yasuda
Bovine papular stomatitis virus (BPSV) causes pustular cutaneous disease in cattle worldwide. This paper describes the development of a specific loop-mediated isothermal amplification (LAMP) assay to detect BPSV which did not cross-react with other parapoxviruses. To assess analytical sensitivity of this LAMP assay, DNA was extracted from serially diluted BPSV from which the infectious titer was determined by a novel assay based on calf kidney epithelial cells. The LAMP assay had equivalent analytical sensitivity to quantitative PCR, and could detect as few as 86 copies of viral DNA per reaction...
October 14, 2016: Journal of Virological Methods
https://www.readbyqxmd.com/read/27744093/evaluation-of-propidium-monoazide-and-long-amplicon-qpcr-as-an-infectivity-assay-for-coliphage
#18
Nicole L McLellan, Hung Lee, Marc B Habash
Standardized and rapid assays for viable viral pathogens are needed to inform human health risk assessments. Conventional qPCR is designed to enumerate the gene copies of an organism in a sample, but does not identify those that originated from a viable pathogen. This study was undertaken to evaluate modified qPCR methods as infectivity assays for the enumeration of infectious MS2 coliphage. Propidium monoazide (PMA) treatment coupled with long-amplicon qPCR assays were assessed for their ability to quantify infectious MS2 in pure cultures and following inactivation by a range of UV light exposures and chlorine doses...
October 12, 2016: Journal of Virological Methods
https://www.readbyqxmd.com/read/27737784/rapid-and-sensitive-detection-of-lily-symptomless-virus-by-reverse-transcription-loop-mediated-isothermal-amplification
#19
Xiangfeng He, Fei Xue, Shufa Xu, Wenhe Wang
Lily symptomless virus (LSV) is one of the most prevalent viruses that infect lily plants worldwide. A rapid and sensitive reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay was developed for detection of LSV, using two primer pairs that specifically amplified the conserved sequence of LSV coat protein. The optimum reaction conditions were as follows: 4mM MgCl2 and 0.8M betaine with incubation at 64°C for 30min. The limit of detection of LSV from infected lily leaves was 10-fold higher for RT-LAMP than for conventional RT-PCR...
October 11, 2016: Journal of Virological Methods
https://www.readbyqxmd.com/read/27737783/development-of-a-phenotypic-susceptibility-assay-for-hiv-1-integrase-inhibitors
#20
Eva Heger, Alexandra Andrée Theis, Klaus Remmel, Hauke Walter, Alejandro Pironti, Elena Knops, Veronica Di Cristanziano, Björn Jensen, Stefan Esser, Rolf Kaiser, Nadine Lübke
Phenotypic resistance analysis is an indispensable method for determination of HIV-1 resistance and cross-resistance to novel drug compounds. Since integrase inhibitors are essential components of recent antiretroviral combination therapies, phenotypic resistance data, in conjunction with the corresponding genotypes, are needed for improving rules-based and data-driven tools for resistance prediction, such as HIV-Grade and geno2pheno[integrase]. For generation of phenotypic resistance data to recent integrase inhibitors, a recombinant phenotypic integrase susceptibility assay was established...
October 11, 2016: Journal of Virological Methods
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