Development and Implementation of a Quadruple RT-qPCR Method for the Identification of Porcine Reproductive and Respiratory Syndrome Virus Strains

Ruan, Shengnan; Ren, Wenhui; Yu, Bin; Yu, Xuexiang; Wu, Hao; Li, Wentao; Jiang, Yunbo; He, Qigai

doi:10.3390/v15091946

Cited by 4 publications

(5 citation statements)

References 34 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…The compromised immunity resulting from PRDC pathogen infections in fattening pigs renders them susceptible to additional pathogens, resulting in heightened economic losses [ 18 ]. Furthermore, secondary infections from pathogens such as Actinobacillus pleuropneumoniae, S. suis, G. parasuis, and Pasteurella multocida are associated with PRDC [ 3 , 8 , 19 ]. Conventional diagnostic methods for PRDC-related viruses involve virus isolation in cell culture, antigen detection through direct fluorescent antibody staining, enzyme immunoassays, and bacterial isolation [ 3 , 20 ].…”

Section: Discussionmentioning

confidence: 99%

“…Moreover, bacterial pathogen detection relies on culture-based approaches, which may take days to yield results [ 23 ]. Due to the high sensitivity and ease of use of PCR and real-time PCR testing, several assays for PRDC-related agents have been developed, but these tests are often specific to individual pathogens [ 8 , 24 , 25 ]. Although conventional PCR methods are economical, they are less sensitive and require intricate post-PCR processing, limiting their application in sample analysis [ 21 , 26 ].…”

Section: Discussionmentioning

confidence: 99%

“…Although conventional PCR methods are economical, they are less sensitive and require intricate post-PCR processing, limiting their application in sample analysis [ 21 , 26 ]. Different from the conventional PCR, real-time PCR monitors target amplification in real-time, offering heightened sensitivity and rapidity [ 8 ]. Recently, Lung et al reported a novel automated microarray prototype for streamlining post-PCR processing for simultaneous detection and genotyping of bacteria ( Mycoplasma hyopneumonia , A. pleuropneumoniae, Salmonella choleraesuis , and S. suis ) and viruses (PRRSV, SIV, PCV2, PRCV) [ 27 ].…”

Section: Discussionmentioning

confidence: 99%

“…PCV2 and PCV3 often co-infect swine, and their clinical symptoms and pathological changes are similar, making it challenging to differentiate between them. This has led to an increased incidence and severity of swine respiratory diseases, complicating diagnoses [ 7 , 8 ]. Single-pathogen infections may not always result in symptoms, but complex infections involving multiple pathogens can lead to severe diseases [ 9 ].…”

Section: Introductionmentioning

confidence: 99%

“…To minimize the economic losses associated with PRDC, itis essential to detect a variety of PRDC pathogens in the herd promptly, accurately, and comprehensively [ 10 , 11 ]. Currently, diagnostic methods for PRRSV, PCV2, PCV3, and SS infections include pathogen isolation, conventional or real-time RT-PCR, and other molecular diagnostic techniques [ 8 ]. While pathogen isolation is considered the diagnostic confirmation of infection, it has limitations in terms of sensitivity, duration, technical equipment, and expertise, making rapid diagnosis impractical.…”

Section: Introductionmentioning

confidence: 99%

See 4 more Smart Citations

Establishment and Application of a Quadruplex Real-Time Reverse-Transcription Polymerase Chain Reaction Assay for Differentiation of Porcine Reproductive and Respiratory Syndrome Virus, Porcine Circovirus Type 2, Porcine Circovirus Type 3, and Streptococcus suis

Wang,

Zhu,

Zhan

et al. 2024

Microorganisms

View full text Add to dashboard Cite

Respiratory illnesses present a significant threat to porcine health, with co-infections involving Porcine Reproductive and Respiratory Syndrome Virus (PRRSV), Streptococcus suis (SS), Porcine Circovirus Type 2 (PCV2), and Porcine Circovirus Type 3 (PCV3) acting as the primary causative agents. As a result, the precise diagnosis of PRRSV, PCV2, PCV3 and SS is of paramount importance in the prevention and control of respiratory diseases in swine. Therefore, we conducted a molecular bioinformatical analysis to concurrently detect and differentiate PRRSV, PCV2, PCV3 and SS. We selected the ORF6 gene of PRRSV, the ORF2 gene of PCV2 and PCV3, and the glutamate dehydrogenase (GDH) gene of SS as targets. Specific primers and probes were designed for each pathogen, and following meticulous optimization of reaction conditions, we established a multiple TaqMan fluorescence quantitative PCR detection method. Subsequently, we subjected this method to a comprehensive assessment, evaluating its specificity, sensitivity, and repeatability. The research results demonstrated that the established multiple TaqMan fluorescence quantitative PCR detection method displays displayed exemplary specificity, with no instances of cross-reactivity with other pathogens. The method’s minimum detection concentrations for PRRSV, PCV2, PCV3, and SS were 2.80 × 101 copies/µL, 1.96 × 102 copies/µL, 2.30 × 102 copies/µL, and 1.75 × 103 copies/µL, respectively. When applied to the analysis of 30 clinical samples, the results closely mirrored those obtained through Chinese standard uniplex real-time qPCR detection method for PRRSV, as well as the general PCR methods for SS, PCV2, and PCV3. This study underscores the robust specificity, high sensitivity, and consistent stability of the multiple TaqMan fluorescence quantitative PCR detection method that we have developed. It is ideally suited to the clinical monitoring of PRRSV, PCV2, PCV3, and SS, and it carries significant importance in ongoing efforts to prevent and manage respiratory diseases in porcine populations.

show abstract