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Journal of Virological Methods

Yanmin Guo, Weiwei Zeng, Qing Wang, Yingying Wang, Yingying Li, Jiyuan Yin, Yan Ren, Cunbin Shi
Grass carp reovirus (GCRV) is the causative agent of a hemorrhagic disease that causes severe economic losses in the grass carp (Ctenopharyngodon idella) farming industry in China. Discrimination between wild-type field and vaccine strains of GCRV is crucial for meaningful disease diagnosis and epidemiological investigation, yet current detection methods do not discriminate between these different viruses. This study exploited sequence differences between vaccine viruses and the virulent and field strains in the S6 gene that is present in all GCRV strains to develop a high resolution melting curve assay to differentiate between virus strains...
March 17, 2018: Journal of Virological Methods
Jinfeng Wang, Jianchang Wang, Ruoxi Zhang, Libing Liu, Ruihan Shi, Qingan Han, Wanzhe Yuan
A rapid and specific real-time reverse-transcription recombinase polymerase amplification assay (RT-RPA) was developed to detect the transmissible gastroenteritis virus (TGEV) in this study. The primers and exo probe were designed to be specific for a portion of spike (S) gene conserved in TGEV, but absent in the closely related porcine respiratory coronavirus (PRCV). The amplification was performed at 40°C for 20min. The assay could only detect the TGEV, and there was no cross-reaction with other pathogens tested...
March 14, 2018: Journal of Virological Methods
Cynthia Chapel, Nathalie Rocher-Hélion, Nathalie Mantel, Sandy Imbert, Eric Deshaies, Véronique Barban, Alain Sabouraud, Jean-Philippe Barbe, Laurent Mallet
Absence of 'live' residual poliovirus in Inactivated Poliovirus Vaccine (IPV) is routinely checked using Primary Monkey Kidney Cells (PMKC). However, the increasing demand for IPV and the ethical, technical and safety issues associated with the use of non-human primates in research and quality control, has made the replacement of primary cells with an established cell line a priority, in line with the principles of the 3Rs (Replacement, Reduction and Refinement in animal testing). As an alternative to PMKC, we evaluated the L20B cell line; a mouse cell-line genetically engineered to express human poliovirus receptor, CD155...
March 13, 2018: Journal of Virological Methods
Sonja Günther, Sandra Felten, Gerhard Wess, Katrin Hartmann, Karin Weber
Feline infectious peritonitis (FIP) is a fatal disease in cats worldwide. The aim of this study was to test two commercially available reaction mixtures in a reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay to detect feline Coronavirus (FCoV) in body cavity effusions of cats with and without FIP, in order to minimize the time from sampling to obtaining results. RNA was extracted from body cavity effusion samples of 71 cats, including 34 samples from cats with a definitive diagnosis of FIP, and 37 samples of control cats with similar clinical signs but other confirmed diseases...
March 11, 2018: Journal of Virological Methods
C A Rajabu, G G Kennedy, J Ndunguru, E M Ateka, F Tairo, L Hanley-Bowdoin, J T Ascencio-Ibáñez
Geminiviruses are devastating single-stranded DNA viruses that infect a wide variety of crops in tropical and subtropical areas of the world. Tomato, which is a host for more than 100 geminiviruses, is one of the most affected crops. Developing plant models to study geminivirus-host interaction is important for the design of virus management strategies. In this study, "Florida Lanai" tomato was broadly characterized using three begomoviruses (Tomato yellow leaf curl virus, TYLCV ; Tomato mottle virus, ToMoV; Tomato golden mosaic virus, TGMV) and a curtovirus (Beet curly top virus, BCTV)...
March 9, 2018: Journal of Virological Methods
Moyra Machado Portilho, Ana Carolina da Fonseca Mendonça, Cristianne Bezerra da Silva, Márcia Paschoal do Espirito-Santo, Vanessa Salete de Paula, Leticia Cancella Nabuco, Cristiane Alves Villela-Nogueira, Lia Laura Lewis-Ximenez, Elisabeth Lampe, Livia Melo Villar
For quantification of hepatitis B virus DNA (HBV DNA), commercial assays are used with serum or plasma samples, but oral fluid samples could be an alternative for HBV diagnosis due to ease of collection. This study aims to develop in-house real time PCR using synthetic curve for HBV DNA quantification for serum and oral fluid samples. Samples were collected from 103 individuals (55 HBsAg reactive and HBV DNA reactive by commercial assay and 48 without HBV markers) and submitted to two in-house real time PCR assays for HBV pre-S/S region with different standard curves: qPCR plasmidial and qPCR synthetic...
March 4, 2018: Journal of Virological Methods
Junwei Ge, Yunjia Shi, Xingyang Cui, Shanshan Gu, Lili Zhao, Hongyan Chen
To date, the pathogenic role of mink circovirus (MiCV) remains unclear, and its prevalence and economic importance are unknown. Therefore, a rapid and sensitive molecular diagnosis is necessary for disease management and epidemiological surveillance. However, only PCR methods can identify MiCV infection at present. In this study, we developed a nested PCR and established a novel recombinase polymerase amplification (RPA) assay for MiCV detection. Sensitivity analysis showed that the detection limit of nested PCR and RPA assay was 101 copies/reaction, and these methods were more sensitive than conventional PCR, which has a detection limit of 105 copies/reaction...
February 28, 2018: Journal of Virological Methods
Hui Hu, Kwonil Jung, Qiuhong Wang, Linda J Saif, Anastasia N Vlasova
Coronaviruses (CoVs) are critical human and animal pathogens because of their potential to cause severe epidemics of respiratory or enteric diseases. In pigs, the newly emerged porcine deltacoronavirus (PDCoV) and re-emerged porcine epidemic diarrhea virus (PEDV) reported in the US and Asia, as well as the discovery of novel CoVs in wild bats or birds, has necessitated development of improved detection and control measures for these CoVs. Because the previous pancoronavirus (panCoV) RT-PCR established in our laboratory in 2007-2011 did not detect deltacoronaviruses (δ-CoVs) in swine fecal and serum samples, our goal was to develop a new panCoV RT-PCR assay to detect known human and animal CoVs, including δ-CoVs...
February 27, 2018: Journal of Virological Methods
Rosana P Rota, Carlos A Palacios, C Facundo Temprana, Marcelo H Argüelles, Marcelo G Mandile, Nora Mattion, Andrea S Laimbacher, Cornell Fraefel, Alejandro A Castello, Graciela Glikmann
Group C Rotavirus (RVC) has been associated globally with sporadic outbreaks of gastroenteritis in children and adults. RVC also infects animals, and interspecies transmission has been reported as well as its zoonotic potential. Considering its genetic diversity and the absence of effective vaccines, it is important and necessary to develop new generation vaccines against RVC for both humans and animals. The aim of the present study was to develop and characterize an HSV-1-based amplicon vector expressing a human RVC-VP6 protein and evaluate the humoral immune response induced after immunizing BALB/c mice...
February 26, 2018: Journal of Virological Methods
Ye Feng, Muyang Wang, Tingfang Liu, Yan Zhang, Zhongzhong Tu, Huancheng Guo, Cuijuan Zhang, Renying Zhu, Wenlin Ren, Le Sun, Weidi Xu, Yuyang Wang, Maohua Li, Changchun Tu
Rabies is a major public health problem in developing countries in Asia and Africa. Although a number of laboratory diagnoses can be used for rabies control, the WHO and OIE recommended gold standard for rabies diagnosis is the direct fluorescent antibody test (FAT). However, FAT is not widely used in developing countries because of deficient financial sources to procure fluorescent microscope. Recently the direct rapid immunohistochemical test (dRIT) has been developed and has a worldwide promising application, particularly in developing countries, since its result can be read by inexpensive light microscopy, in addition to be consistent with that of FAT...
February 25, 2018: Journal of Virological Methods
Long Feng, Wuhao Lu, Yunyun Ma, Wentao Guo, Yuanyuan Wang, Qianqian Sun, Jianbo Wu, Guoqiang Zhao, Xiaoli Zhang
Acquired immunodeficiency syndrome (AIDS) is a serious worldwide disease caused by infection with the human immunodeficiency virus (HIV). C-C chemokine receptor 5 (CCR5) and C-X-C chemokine receptor 4 (CXCR4) are important coreceptors mediating HIV-1 cell entry. Many new anti-HIV drugs are currently in preclinical and clinical trials; however, drug development has proceeded slowly partly because of the lack of a high-throughput system to screen these drugs. Here, we describe the development of a novel dual-luciferase assay using a CCR5/CXCR4 promoter-driven firefly and Renilla luciferase vector (pGL4...
February 23, 2018: Journal of Virological Methods
James Logue, Kaylie Tuznik, Dean Follmann, Greg Grandits, Jonathan Marchand, Cavan Reilly, Yeya Dit Sadio Sarro, James Pettitt, Eric J Stavale, Mosoka Fallah, Gene G Olinger, Fatorma K Bolay, Lisa E Hensley
As part of the scientific community's development of medical countermeasures against Ebola virus disease, optimization of standardized assays for product evaluation is paramount. The recent outbreak heightened awareness to the scarcity of available assays and limited information on performance and reproducibility. To evaluate the immunogenicity of vaccines entering Phase I-III trials and to identify survivors, two enzyme-linked immunosorbent assays, the Filovirus Animal Non-Clinical Group assay and the Alpha Diagnostics International assay, were evaluated for detection of immunoglobulin G against Ebola virus glycoprotein...
February 23, 2018: Journal of Virological Methods
Ana Del Cerro, Ana Balseiro, Rosa Casais, Kevin Dalton, Luis Salinas, Juan Carlos Saiz, Luis J Royo
Louping ill-like virus (LI) has been recently detected in two different locations in the north of Spain and separated by only around 400 km. Using molecular approaches, the viruses causing both outbreaks have been shown to be different to LI virus, but also different to each other. They have been called SSEV (Spanish sheep encephalitis virus) and SGEV (Spanish goat encephalitis virus) taking into account the species from which they were isolated. The aim of this paper was to design a quantitative TaqMan real-time RT-PCR protocol, for the specific diagnostic and quantitation of SGEV...
February 23, 2018: Journal of Virological Methods
HyeSoon Song, YouChan Bae, SeokChan Park, HyukMan Kwon, HeeSoo Lee, SeongJoon Joh
Loop-mediated isothermal amplification (LAMP) methods to detect chicken infectious anemia virus (CIAV), reticuloendotheliosis virus (REV), and Marek's disease virus (MDV), and a reverse transcription (RT)-LAMP assay to detect infectious bursal disease virus (IBDV), were developed. The CIAV-LAMP, REV-LAMP, MDV-LAMP, and IBDV-RT-LAMP methods were performed using four sets of six primers targeting the VP1 gene of CIAV, the gp90 gene of REV, the Meq gene of MDV, and the VP2 gene of IBDV. The results (a change in color) were observed visually...
February 21, 2018: Journal of Virological Methods
Vaea Richard, Maite Aubry
Experimental studies on Zika virus (ZIKV) may require improvement of infectious titers in viral stocks obtained by cell culture amplification. We highlight here the use of centrifugal filter devices to increase infectious titers of ZIKV from cell-culture supernatants. A mean gain of 2.33 ± 0.12 log10 DICT50 /mL was easily and rapidly obtained with this process. This efficient method of ultrafiltration may be applied to other viruses and be useful in various experimental studies requiring high viral titers...
February 20, 2018: Journal of Virological Methods
Kelly A Curtis, Daphne Morrison, Donna L Rudolph, Anupama Shankar, Laura S P Bloomfield, William M Switzer, S Michele Owen
Isothermal nucleic acid amplification techniques, such as reverse-transcription loop-mediated isothermal amplification (RT-LAMP), exhibit characteristics that are suitable for the development of a rapid, low-cost NAT that can be used at the POC. For demonstration of utility for global use, studies are needed to validate the performance of RT-LAMP for the detection of divergent subtypes. In this study, we designed and evaluated multiplexed HIV-1 integrase RT-LAMP primers to detect subtypes within group M, along with an RNase P positive internal processing and amplification control...
February 20, 2018: Journal of Virological Methods
Michael Versoza, Wonseok Jung, Mona Loraine Barabad, Yongil Lee, Kyomin Choi, Duckshin Park
The inactivation of viruses that retain their infectivity when transmitted through the air is challenging. To address this issue, this study used a non-contact ultrasound transducer (NCUT) to generate shock waves in the air at specific distances, input voltages, and exposure durations, targeting bacteriophage virus aerosols captured on to H14 HEPA filters. Initially, a frequency of 27.56 kHz (50V) at 25-mm distance was used, which yielded an inactivation efficiency of up to 32.69 ± 12.10%. Other frequencies at shorter distances were investigated, where 29...
February 20, 2018: Journal of Virological Methods
Massimo Giangaspero, Claudio Apicella
Pestivirus bovine viral diarrhea virus type 1 species is responsible for cosmopolitan diseases affecting cattle and other ruminants, presenting a wide range of clinical manifestations, with relevant impact on zootechnic production. Understanding genomic characteristic and virus taxonomy is fundamental in order to sustain control and prophylactic programs. Given the recent various studies reporting a relatively high number of new strains, in particular from Asian countries, in the present study, four hundred-eighty-two genomic sequences have been evaluated applying the palindromic nucleotide substitutions method for genotyping...
February 17, 2018: Journal of Virological Methods
Ningning Han, Zishu Chen, Hu Wan, Guohua Huang, Jianhong Li, Byung Rae Jin
Ascoviruses are a recently described family and the traditional plaque assay and end-point PCR assay have been used for their titration. However, these two methods are time-consuming and inaccurate to titrate ascoviruses. In the present study, a quick method for the determination of the titer of ascovirus stocks was developed based on ascovirus-induced apoptosis in infected insect cells. Briefly, cells infected with serial dilutions of virus (10 -2 to 10 -10 ) for 24 h were stained with trypan blue. The stained cells were counted, and the percentage of nonviable cells was calculated...
February 14, 2018: Journal of Virological Methods
Parvapan Bhattarakosol, Kamolwan Plaignam, Amornpun Sereemaspun
HPV-16 infection is the most common cause of cervical cancer. As HPV-16 transforms the cell, E6 oncoprotein is over-expressed. Therefore, molecular detection of HPV-16 E6 mRNA is now being used for diagnosis and prediction of cancer development. Besides detecting E6 mRNA, a rapid lateral flow detecting the E6 protein using enzyme immunoassay is also now on market with a sensitivity of 53.5% for cervical intraepithelial neoplasia (CIN)-3 or more severe (CIN-3 + ). Here, an immunogold-agglutination assay was developed to detect not only HPV-16 E6 protein but also L1, a major capsid protein found in the productive stage of the virus...
February 12, 2018: Journal of Virological Methods
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