Peer-reviewed veterinary case report
Development and evaluation of a loop-mediated isothermal amplification method for rapid detection and differentiation of two genotypes of porcine circovirus type 2.
- Journal:
- Journal of microbiology, immunology, and infection = Wei mian yu gan ran za zhi
- Year:
- 2014
- Authors:
- Wang, Chun et al.
- Affiliation:
- Animal Health Research Institute
Plain-English summary
Researchers have developed a new testing method called LAMP to quickly identify two types of a virus known as porcine circovirus type 2 (PCV2), which affects pigs and can lead to significant economic losses for farmers. This method can differentiate between the two main types of the virus, PCV2a and PCV2b, and it works by using specific DNA markers. In tests with various pig virus samples, LAMP showed high accuracy, detecting the target virus without mixing up with other viruses. The results indicated that LAMP is both fast and reliable, making it a promising tool for diagnosing these viral infections in pigs.
Abstract
BACKGROUND: Porcine circovirus type 2 (PCV2) is one of the major swine viral diseases and has caused significant economic loss to pig producers. PCV2 has been divided into two major genotypes: PCV2a, PCV2b. A loop-mediated isothermal amplification (LAMP) method was developed for the detection and differentiation of PCV2a and PCV2b in clinical samples. METHODS: LAMP-specific primer sets were designed based on six PCV2a and six PCV2b reference isolates. To determine the analytical specificity of LAMP, DNA samples extracted from 36 porcine virus isolates were tested by LAMP, including eight PCV2a, 11 PCV2b, four PCV type 1, two porcine parvovirus, three pseudorabies virus, and eight porcine reproductive and respiratory virus. To evaluate the analytical sensitivity of the assay, 10-fold serial dilutions of PCV2a and PCV2b recombinant plasmids were performed to prepare the dilutions at concentration from 10(6) to 1 copy(ies)/μL, and each dilution was tested by both LAMP and nested polymerase chain reaction (nested PCR). A total of 168 clinical samples were analyzed by both LAMP and nested PCR, and the relative sensitivity and specificity of LAMP compared to nested PCR were calculated. RESULTS: Using different primer sets of LAMP, LAMP could be completed within 50 minutes. This method was found to be highly analytically specific for PCV2a and PCV2b; only the target gene was detected without cross-reaction. The analytical sensitivity of LAMP for PCV2a and PCV2b were 10 copies/μL, demonstrating analytical sensitivity comparable to that obtained using nested PCR. In addition, the sensitivity and specificity of LAMP relative to those of nested PCR were 97.7% and 100.0%, respectively. The percentage of observed agreement was 98.2%, and the κ statistic was 0.949. CONCLUSION: LAMP is a rapid, specific, and sensitive diagnostic method for the detection and differentiation of PCV2a and PCV2b in clinical samples.
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Search related cases →Original publication: https://pubmed.ncbi.nlm.nih.gov/23845855/