scholarly journals Pan-Piscine Orthoreovirus (PRV) Detection Using Reverse Transcription Quantitative PCR

Pathogens ◽  
2021 ◽  
Vol 10 (12) ◽  
pp. 1548
Author(s):  
Julie Zhao ◽  
Niccolò Vendramin ◽  
Argelia Cuenca ◽  
Mark Polinski ◽  
Laura M. Hawley ◽  
...  
Keyword(s):  
Quantitative Pcr ◽  
Reverse Transcription ◽  
High Sensitivity ◽  
Test Results ◽  
Reverse Transcription Quantitative Pcr ◽  
Highly Sensitive ◽  
Surveillance Programs ◽  
America South ◽  
Universal Detection ◽  
Quantitative Pcr Assay

Piscine orthoreovirus (PRV) infects farmed and wild salmon and trout species in North America, South America, Europe, and East Asia. PRV groups into three distinct genotypes (PRV-1, PRV-2, and PRV-3) that can vary in distribution, host specificity, and/or disease potential. Detection of the virus is currently restricted to genotype specific assays such that surveillance programs require the use of three assays to ensure universal detection of PRV. Consequently, herein, we developed, optimized, and validated a real-time reverse transcription quantitative PCR assay (RT-qPCR) that can detect all known PRV genotypes with high sensitivity and specificity. Targeting a conserved region at the 5′ terminus of the M2 segment, the pan-PRV assay reliably detected all PRV genotypes with as few as five copies of RNA. The assay exclusively amplifies PRV and does not cross-react with other salmonid viruses or salmonid host genomes and can be performed as either a one- or two-step RT-qPCR. The assay is highly reproducible and robust, showing 100% agreement in test results from an inter-laboratory comparison between two laboratories in two countries. Overall, as the assay provides a single test to achieve highly sensitive pan-specific PRV detection, it is suitable for research, diagnostic, and surveillance purposes.

scholarly journals Development of a Reverse Transcription-Quantitative PCR Assay for Detection of Salivirus/Klassevirus

2013 ◽  
Vol 79 (11) ◽  
pp. 3529-3532 ◽  
Author(s):  
Eiji Haramoto ◽  
Masaaki Kitajima ◽  
Mikie Otagiri
Keyword(s):  
Quantitative Pcr ◽  
Reverse Transcription ◽  
Water Samples ◽  
Maximum Concentration ◽  
Pcr Assay ◽  
River Water Samples ◽  
Treated Sewage ◽  
Reverse Transcription Quantitative Pcr ◽  
Highly Sensitive ◽  
Quantitative Pcr Assay

ABSTRACTA broadly reactive and highly sensitive reverse transcription-quantitative PCR assay to detect salivirus/klassevirus was developed. By means of the developed assay, salivirus/klassevirus was detected in 13 (93%) raw sewage, 4 (29%) secondary-treated sewage, and 9 (16%) river water samples, with a maximum concentration of 9.7 × 106copies/liter.

Screening of Fruit Products for Norovirus and the Difficulty of Interpreting Positive PCR Results

2011 ◽  
Vol 74 (3) ◽  
pp. 425-431 ◽  
Author(s):  
AMBROOS STALS ◽  
LEEN BAERT ◽  
VICKY JASSON ◽  
ELS VAN COILLIE ◽  
MIEKE UYTTENDAELE
Keyword(s):  
Quantitative Pcr ◽  
Reverse Transcription ◽  
Food Samples ◽  
Murine Norovirus ◽  
Virus Particles ◽  
Viral Particles ◽  
Reverse Transcription Quantitative Pcr ◽  
Fruit Samples ◽  
Positive Results ◽  
Quantitative Pcr Assay

Despite recent norovirus (NoV) outbreaks related to consumption of fruit products, little is known regarding the NoV load on these foods. Therefore, 75 fruit products were screened for NoV presence by using an evaluated in-house NoV detection methodology consisting of a NoV extraction method and a reverse transcription quantitative PCR assay. Additionally, the fruit samples were screened for bacterial pathogens and bacterial hygiene indicators. Results of the NoV screening showed that 18 of 75 samples tested positive for GI and/or GII NoV despite a good bacteriological quality. The recovery of murine norovirus 1 virus particles acting as process control was successful in 31 of 75 samples with a mean recovery efficiency of 11.32% plusmn; 6.08%. The level of detected NoV genomic copies ranged between 2.5 and 5.0 log per 10 g. NoV GI and/or GII were found in 4 of 10, 7 of 30, 6 of 20, and 1 of 15 of the tested raspberries, cherry tomatoes, strawberries, and fruit salad samples, respectively. However, confirmation of the positive quantitative PCR results by sequencing genotyping regions in the NoV genome was not possible. Due to the nature of the method used (reverse transcription quantitative PCR) for detection of genomic material, no differentiation was possible between infectious and noninfectious viral particles. No NoV outbreaks related to the tested fruit product types were reported during the screening period, which hampers a conclusion as to whether these unexpected high numbers of NoV-positive results should be perceived as a public health threat. These results, however, may indicate a prior NoV contamination of the tested food samples throughout the fresh produce chain.

scholarly journals Establishment and Application of Rapid Diagnosis for Reverse Transcription-Quantitative PCR of Newly Emerging Goose-Origin Nephrotic Astrovirus in China

mSphere ◽  
2018 ◽  
Vol 3 (6) ◽  
Author(s):  
Xiaoyuan Yuan ◽  
Kai Meng ◽  
Yuxia Zhang ◽  
Lihong Qi ◽  
Wu Ai ◽  
...  
Keyword(s):  
Quantitative Pcr ◽  
Clinical Application ◽  
Reverse Transcription ◽  
Test Sample ◽  
Sensitivity Testing ◽  
Content Type ◽  
Reverse Transcription Quantitative Pcr ◽  
Emerging Virus ◽  
New Type ◽  
And Control

ABSTRACT In 2017, a new type of goose-origin astrovirus (GoAstV) that is completely different from previously identified avian astroviruses (which have only 30.0% to 50.5% homology with GoAstV) has been isolated from diseased geese in China. This disease can cause joint swelling in sick geese, and the anatomy shows a clear precipitation of urate in the kidney. The rate of death and culling can reach more than 30%, revealing the disease’s severe pathogenicity. To quickly and accurately diagnose the newly emerging disease, we established a highly specific reverse transcription-quantitative PCR (RT-qPCR) method of detecting GoAstV. Sensitivity testing showed that the minimum amount of test sample for this method is 52.5 copies/μl. Clinical application confirmed that this method can quickly and effectively detect GoAstV, providing a diagnostic platform for the prevention and control of goose disease. IMPORTANCE Goose-origin astrovirus (GoAstV), as a newly emerging virus in 2017, is different from previously known astroviruses in the genus Avastrovirus. So far, few studies have focused on the novel virus. Considering the infectious development of astrovirus (AstV), we established a reverse transcription-quantitative PCR (RT-qPCR) assay with a strong specificity to quickly and accurately diagnose GoAstV. Confirmed by clinical application, this method can quickly and accurately detect prevalent GoAstV. The assay is thus convenient for clinical operation and is applicable to the monitoring of GoAstV disease.

scholarly journals Selection of Valid Reference Genes for Reverse Transcription Quantitative PCR Analysis inHeliconius numata(Lepidoptera: Nymphalidae)

2016 ◽  
Vol 16 (1) ◽  
pp. 50 ◽  
Author(s):  
Florence Piron Prunier ◽  
Mathieu Chouteau ◽  
Annabel Whibley ◽  
Mathieu Joron ◽  
Violaine Llaurens
Keyword(s):  
Quantitative Pcr ◽  
Reverse Transcription ◽  
Reference Genes ◽  
Pcr Analysis ◽  
Reverse Transcription Quantitative Pcr ◽  
Selection Of
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