scholarly journals Development of Real-Time PCR Based on A137R Gene for the Detection of African Swine Fever Virus

2021 ◽  
Vol 8 ◽  
Author(s):  
Dan Yin ◽  
Renhao Geng ◽  
Hui Lv ◽  
Chunhui Bao ◽  
Hongxia Shao ◽  
...  
Keyword(s):  
Real Time ◽  
Real Time Pcr ◽  
African Swine Fever Virus ◽  
African Swine Fever ◽  
Fever Virus ◽  
Economic Losses ◽  
Clinical Samples ◽  
Pcr Assay ◽  
Taqman Pcr ◽  
Control And Eradication

African swine fever virus (ASFV) can infect domestic pigs and wild boars and causes huge economic losses in global swine industry. Therefore, early diagnosis of ASFV is important for the control and eradication of African swine fever (ASF). In this study, a SYBR Green-based real-time polymerase chain reaction (PCR) assay targeting the viral encoded A137R gene was established for the detection of ASFV infection. For the evaluation of the established real-time PCR, 34 clinical samples were assessed by both the A137R gene-based real-time PCR and OIE-recommended TaqMan PCR. The results showed that 85.29% (29/34) were detected by A137R gene-based real-time PCR, but only 79.41% (27/34) positive using OIE-recommended TaqMan PCR. Moreover, no cross-reaction with other common swine pathogens was found in the A137R gene-based real-time PCR. These results demonstrated that the established real-time PCR assay in this study showed better performance than the OIE-recommended method in detecting ASFV from clinical samples, which could be applied for control and eradication programs of ASF.

Development of a real‐time PCR assay for detection of African swine fever virus with an endogenous internal control

2020 ◽  
Vol 67 (6) ◽  
pp. 2446-2454 ◽  
Author(s):  
Yin Wang ◽  
Lizhe Xu ◽  
Lance Noll ◽  
Colin Stoy ◽  
Elizabeth Porter ◽  
...  
Keyword(s):  
Real Time ◽  
Internal Control ◽  
Real Time Pcr ◽  
African Swine Fever Virus ◽  
African Swine Fever ◽  
Fever Virus ◽  
Pcr Assay

Development of a triplex real-time PCR assay for detection and differentiation of gene-deleted and wild-type African swine fever virus

2020 ◽  
Vol 280 ◽  
pp. 113875
Author(s):  
Yanxing Lin ◽  
Chenfu Cao ◽  
Weijun Shi ◽  
Chaohua Huang ◽  
Shaoling Zeng ◽  
...  
Keyword(s):  
Real Time ◽  
Real Time Pcr ◽  
African Swine Fever Virus ◽  
African Swine Fever ◽  
Fever Virus ◽  
Wild Type ◽  
Pcr Assay

scholarly journals Development and evaluation of duplex TaqMan real-time PCR assay for detection and differentiation of wide-type and MGF505-2R gene-deleted African swine fever viruses

2020 ◽  
Author(s):  
zhenhua Guo ◽  
Kunpeng Li ◽  
Songlin Qiao ◽  
Xinxin Chen ◽  
Ruiguang Deng ◽  
...  
Keyword(s):  
Real Time ◽  
Real Time Pcr ◽  
Large Scale ◽  
Limit Of Detection ◽  
African Swine Fever ◽  
Economic Losses ◽  
Clinical Samples ◽  
Diagnosis Method ◽  
Pcr Method ◽  
And Control

Abstract Background: African swine fever (ASF) is the most important disease to the pigs and cause serious economic losses to the countries with large-scale swine production. Vaccines are recognized as the most useful tool to prevent and control ASF virus (ASFV) infection. Currently, the MGF505 and MGF360 gene-deleted ASFVs or combined with CD2v deletion were confirmed to be the most promising vaccine candidates. Thus, it is essential to develop a diagnosis method to discriminate wide-type strain from the vaccines used.Results: In this study, we established a duplex TaqMan real-time PCR based on the B646L gene and MGF505-2R gene. The sequence alignment showed that the targeted regions of primers and probes are highly conserved in the genotype II ASFVs. The duplex real-time assay can specifically detect B646L and MGF505-2R gene single or simultaneously without cross-reaction with other porcine viruses tested. The limit of detection was 5.8 copies and 3.0 copies for the standard plasmids containing B646L and MGF505-2R genes, respectively. Clinical samples were tested in parallel by duplex real-time PCR and a commercial ASFV detection kit. The detection results of these two assays against B646L gene were well consistent.Conclusion: We successfully developed and evaluated a duplex TaqMan real-time PCR method which can effectively distinguish the wide type and MGF505 gene-deleted ASFVs. It would be a useful tool for the clinical diagnosis and control of ASF.

scholarly journals Rapid Extraction and Detection of African Swine Fever Virus DNA Based on Isothermal Recombinase Polymerase Amplification Assay

Viruses ◽  
2021 ◽  
Vol 13 (9) ◽  
pp. 1731
Author(s):  
Arianna Ceruti ◽  
Rea Maja Kobialka ◽  
Judah Ssekitoleko ◽  
Julius Boniface Okuni ◽  
Sandra Blome ◽  
...  
Keyword(s):  
Real Time ◽  
Real Time Pcr ◽  
Dna Isolation ◽  
African Swine Fever Virus ◽  
African Swine Fever ◽  
Fever Virus ◽  
Recombinase Polymerase Amplification ◽  
Reference Laboratory ◽  
Analytical Sensitivity ◽  
Lysis Buffer

African swine fever virus (ASFV) is the causative agent of a deadly disease in pigs and is spread rapidly across borders. Samples collected from suspected cases must be sent to the reference laboratory for diagnosis using polymerase chain reaction (PCR). In this study, we aimed to develop a simple DNA isolation step and real-time recombinase polymerase amplification (RPA) assay for rapid detection of ASFV. RPA assay based on the p72 encoding B646L gene of ASFV was established. The assays limit of detection and cross-reactivity were investigated. Diagnostic performance was examined using 73 blood and serum samples. Two extraction approaches were tested: silica-column-based extraction method and simple non-purification DNA isolation (lysis buffer and heating, 70 °C for 20 min). All results were compared with well-established real-time PCR. In a field deployment during a disease outbreak event in Uganda, 20 whole blood samples were tested. The assay’s analytical sensitivity was 3.5 DNA copies of molecular standard per µL as determined by probit analysis on eight independent assay runs. The ASFV RPA assay only detected ASFV genotypes. Compared to real-time PCR, RPA diagnostic sensitivity and specificity were 100%. Using the heating/lysis buffer extraction procedure, ASFV-RPA revealed better tolerance to inhibitors than real-time PCR (97% and 38% positivity rate, respectively). In Uganda, infected animals were identified before the appearance of fever. The ASFV-RPA assay is shown to be as sensitive and specific as real-time PCR. Moreover, the combination of the simple extraction protocol allows its use at the point of need to improve control measures.

scholarly journals Highly Sensitive PCR Assay for Routine Diagnosis of African Swine Fever Virus in Clinical Samples

2003 ◽  
Vol 41 (9) ◽  
pp. 4431-4434 ◽  
Author(s):  
M. Aguero ◽  
J. Fernandez ◽  
L. Romero ◽  
C. Sanchez Mascaraque ◽  
M. Arias ◽  
...  
Keyword(s):  
African Swine Fever Virus ◽  
African Swine Fever ◽  
Fever Virus ◽  
Clinical Samples ◽  
Pcr Assay ◽  
Routine Diagnosis ◽  
Highly Sensitive
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