scholarly journals Use of Three Air Samplers for the Detection of PRRSV-1 under Experimental and Field Conditions

Animals ◽  
2018 ◽  
Vol 8 (12) ◽  
pp. 233 ◽  
Author(s):  
Heiko Stein ◽  
Jochen Schulz ◽  
Rebecca Morgenstern ◽  
Thomas Voglmayr ◽  
Georg Freymüller ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Field Conditions ◽  
Respiratory Syndrome Virus ◽  
Chain Reaction ◽  
Airborne Transmission ◽  
Experimental Conditions ◽  
Qrt Pcr ◽  
Active Virus ◽  
Long Time ◽  
Polymerase Chain

Airborne transmission of porcine reproductive and respiratory syndrome virus (PRRSV) has been known for a long time. Most experiments were performed using PRRSV-2 strains and fairly little information is available on the airborne spread of PRRSV-1. The aim of this study was to assess three different air samplers for their ability to detect PRRSV-1 under experimental and field conditions. All three devices were able to detect PRRSV-1 by quantitative reverse trascription polymerase chain reaction (qRT-PCR) under experimental conditions. However, the detection of PRRSV-1 in a PRRSV-positive farm with active virus circulation was not successful.

scholarly journals Analysis and validation of a highly sensitive one-step nested quantitative real-time polymerase chain reaction assay for specific detection of severe acute respiratory syndrome coronavirus 2

2020 ◽  
Vol 17 (1) ◽  
Author(s):  
Yang Zhang ◽  
Chunyang Dai ◽  
Huiyan Wang ◽  
Yong Gao ◽  
Tuantuan Li ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Real Time ◽  
Quantitative Polymerase Chain Reaction ◽  
Clinical Samples ◽  
Chain Reaction ◽  
Pcr Assay ◽  
Qrt Pcr ◽  
Highly Sensitive ◽  
One Step ◽  
Polymerase Chain

Abstract Background Coronavirus disease 2019 (COVID-19), caused by SARS-CoV-2, is posing a serious threat to global public health. Reverse transcriptase real-time quantitative polymerase chain reaction (qRT-PCR) is widely used as the gold standard for clinical detection of SARS-CoV-2. Due to technical limitations, the reported positive rates of qRT-PCR assay of throat swab samples vary from 30 to 60%. Therefore, the evaluation of alternative strategies to overcome the limitations of qRT-PCR is required. A previous study reported that one-step nested (OSN)-qRT-PCR revealed better suitability for detecting SARS-CoV-2. However, information on the analytical performance of OSN-qRT-PCR is insufficient. Method In this study, we aimed to analyze OSN-qRT-PCR by comparing it with droplet digital PCR (ddPCR) and qRT-PCR by using a dilution series of SARS-CoV-2 pseudoviral RNA and a quality assessment panel. The clinical performance of OSN-qRT-PCR was also validated and compared with ddPCR and qRT-PCR using specimens from COVID-19 patients. Result The limit of detection (copies/ml) of qRT-PCR, ddPCR, and OSN-qRT-PCR were 520.1 (95% CI: 363.23–1145.69) for ORF1ab and 528.1 (95% CI: 347.7–1248.7) for N, 401.8 (95% CI: 284.8–938.3) for ORF1ab and 336.8 (95% CI: 244.6–792.5) for N, and 194.74 (95% CI: 139.7–430.9) for ORF1ab and 189.1 (95% CI: 130.9–433.9) for N, respectively. Of the 34 clinical samples from COVID-19 patients, the positive rates of OSN-qRT-PCR, ddPCR, and qRT-PCR were 82.35% (28/34), 67.65% (23/34), and 58.82% (20/34), respectively. Conclusion In conclusion, the highly sensitive and specific OSN-qRT-PCR assay is superior to ddPCR and qRT-PCR assays, showing great potential as a technique for detection of SARS-CoV-2 in patients with low viral loads.

scholarly journals Histopathological and virological findings in emaciated pigs from Mexico: an exploratory study

2018 ◽  
Vol 87 (3) ◽  
pp. 213-217
Author(s):  
Aide Alpízar ◽  
Joaquim Segalés ◽  
Simón Martínez ◽  
Atalo Martínez ◽  
Guadalupe Socci ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Body Condition ◽  
Porcine Circovirus Type ◽  
Porcine Circovirus ◽  
Economic Losses ◽  
Respiratory Syndrome Virus ◽  
Chain Reaction ◽  
Polymerase Chain ◽  
Histopathological Studies ◽  
Weaning Piglets

The objective of this work was to detect the presence of three main pig respiratory viral agents (porcine rubulavirus [PorPV], porcine circovirus type 2 [PCV-2], and porcine reproductive and respiratory syndrome virus [PRRSV]) in tissues of emaciated piglets from the Baj'o Region (Mexico). Necropsies and histopathological studies of 37 pigs with poor body condition were performed; viruses were detected by molecular biology methods and PCV-2 was further assessed by immunohistochemistry (IHC). Histopathologically, interstitial pneumonia was observed in 25/37 (68%) of the piglets. Also, a varying degree of lymphocyte depletion in lymphoid organs was found in 14/37 (38%) animals. Through polymerase chain reaction (PCR) and/or reverse transcription polymerase chain reaction (RT-PCR), from the 37 pigs, 16 were positive for PCV-2, 18 for PRRSV and 1 for PorPV. In accordance with these results, the infection and/or co-infection with PCV-2 and PRRSV were fairly frequent findings in piglets with poor body condition in Mexico, while the infection by PorPV was apparently negligible. Wasting of post-weaning piglets is a global pig farming problem that causes great economic losses and has been associated with diverse factors: microbial agents, environmental factors, nutritional factors, and management. When the Blue Eye Disease was first reported in Mexico, it was associated with severe wasting in post-weaning piglets. This study demonstrated that this disease does not seem to play such an important role in the wasting as was previously thought.

scholarly journals Quantitation of 35S Promoter in Maize DNA Extracts from Genetically Modified Organisms Using Real-Time Polymerase Chain Reaction, Part 2: Interlaboratory Study

2005 ◽  
Vol 88 (2) ◽  
pp. 558-573 ◽  
Author(s):  
Max Feinberg ◽  
Sophie Fernandez ◽  
Sylvanie Cassard ◽  
Chrystèle Charles-Delobel ◽  
Yves Bertheau ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Real Time ◽  
Genetically Modified Organisms ◽  
Genetically Modified ◽  
Interlaboratory Study ◽  
Performance Criteria ◽  
Tolerance Interval ◽  
Chain Reaction ◽  
Qrt Pcr ◽  
Polymerase Chain

Abstract The European Committee for Standardization (CEN) and the European Network of GMO Working Laboratories have proposed development of a modular strategy for stepwise validation of complex analytical techniques. When applied to the quantitation of genetically modified organisms (GMOs) in food products, the instrumental quantitation step of the technique is separately validated from the DNA extraction step to better control the sources of uncertainty and facilitate the validation of GMO-specific polymerase chain reaction (PCR) tests. This paper presents the results of an interlaboratory study on the quantitation step of the method standardized by CEN for the detection of a regulatory element commonly inserted in GMO maize-based foods. This is focused on the quantitation of P35S promoter through using the quantitative real-time PCR (QRT-PCR). Fifteen French laboratories participated in the interlaboratory study of the P35S quantitation operating procedure on DNA extract samples using either the thermal cycler ABI Prism® 7700 (Applied Biosystems, Foster City, CA) or Light Cycler® (Roche Diagnostics, Indianapolis, IN). Attention was focused on DNA extract samples used to calibrate the method and unknown extract samples. Data were processed according to the recommendations of ISO 5725 standard. Performance criteria, obtained using the robust algorithm, were compared to the classic data processing after rejection of outliers by the Cochran and Grubbs tests. Two laboratories were detected as outliers by the Grubbs test. The robust precision criteria gave values between the classical values estimated before and after rejection of the outliers. Using the robust method, the relative expanded uncertainty by the quantitation method is about 20% for a 1% Bt176 content, whereas it can reach 40% for a 0.1% Bt176. The performances of the quantitation assay are relevant to the application of the European regulation, which has an accepted tolerance interval of about ±50%. These data were fitted to a power model (r2 = 0.96). Thanks to this model, it is possible to propose an estimation of uncertainty of the QRT-PCR quantitation step and an uncertainty budget depending on the analytical conditions.

scholarly journals Aerosol-generating procedures in thoracic surgery in the COVID-19 era in Malaysia

2020 ◽  
Vol 28 (8) ◽  
pp. 495-499
Author(s):  
Narasimman Sathiamurthy ◽  
Narendran Balasubbiah ◽  
Benedict Dharmaraj
Keyword(s):  
Polymerase Chain Reaction ◽  
Thoracic Surgery ◽  
Reverse Transcriptase ◽  
Personal Protective Equipment ◽  
Healthcare Workers ◽  
Polymerase Chain Reaction Method ◽  
Protective Equipment ◽  
Chain Reaction ◽  
Long Time ◽  
Polymerase Chain

Background The Covid-19 pandemic has caused changes in the surgical treatment of non-Covid patients, especially in thoracic surgery because most procedures are aerosol generating. Hospital Kuala Lumpur, where thoracic procedures are performed, was badly affected. We describe our experience in performing aerosol generating procedures safely in thoracic surgery during the Covid-19 era. Methods Medical records of patients who underwent thoracic surgery from March 18, 2020 to May 17, 2020 were reviewed retrospectively. All patients undergoing thoracic surgery were tested for Covid-19 using the reverse transcriptase polymerase chain reaction method. Patients with malignancy were observed for 10 to 14 days in the ward after testing negative. The healthcare workers donned personal protective equipment for all the cases, and the number of healthcare workers in the operating room was limited to the minimum required. Results A total of 44 procedures were performed in 26 thoracic surgeries. All of these procedures were classified as aerosol generating, and the mean duration of the surgery was 130 ± 43 minutes. None of the healthcare workers involved in the surgery were exposed or infected by Covid-19. Conclusion Covid-19 will be a threat for a long time and thoracic surgeons must continue to provide their services, despite having to deal with aerosol generating procedures, in the new normal. Covid-19 testing of all surgical candidates, using the reverse transcriptase polymerase chain reaction, donning full personal protective equipment for healthcare workers, and carefully planned procedures are among the measures suggested to prevent unnecessary Covid-19 exposure in thoracic surgery.

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