TaqMan-based real-time polymerase chain reaction assay for specific detection of bocavirus-1 in domestic cats

Abstract

Feline bocavirus-1 (FBoV-1) was first discovered in Hong Kong in 2012, and studies have indicated that the virus may cause feline hemorrhagic enteritis. Currently, there is a lack of an effective and quantitative method for FBoV-1 detection. In this study, a TaqMan-based quantitative real-time PCR (qPCR) for FBoV-1 detection was established. Primers and probes were designed to target the conserved region of the FBoV-1 NS1 gene. The sensitivity analysis indicated that the minimum detection limit was 4.57 × 10 1 copies/μL. The specificity test revealed no cross-reaction with seven other common feline viruses, including the same species—FBoV-2 and FBoV-3. The sensitivity of this method was 100 times higher than that of conventional PCR (cPCR). The established method showed good repeatability, with the intra-assay and inter-assay coefficients of variation of 0.18%–1.00% and 0.27%–0.45%, respectively. Furthermore, the analysis of feline feces revealed that the detection rate by qPCR was 7.0% (9/128), whereas that by cPCR was 4.7% (6/128). In conclusion, the established qPCR assay can quantitatively detect FBoV-1 with a high sensitivity, high specificity, and good reproducibility, making it a promising technique for the clinical detection of and basic and epidemiological research on FBoV-1.

Development of SYBR Green I-based polymerase chain reaction for feline bocavirus 1 detection

“…The amplification efficiency of the method was 95.0% and correlation coefficient was 0.999 ( R 2 = 0.999). The detection limit of SYBR Green I qPCR was 3.87 × 10 1 copies/μL, 100 times that of cPCR, and similar to that of the TaqMan method (4.57 × 10 1 copies/μL) established in our laboratory (Wang et al 2020 ). In addition, FBoV-2, FBoV-3, and four other common feline viruses were tested and the results showed that the established assay had good specificity for FBoV-1.…”

“…An efficient identification method for FBoV can provide a powerful tool for the clinical diagnosis of cats with diarrhea. So far, there are conventional polymerase chain reaction (cPCR) methods and Taqman probe-based methods to detect FBoV (Wang et al 2020 ); however, they are not without limitations. cPCR methods are associated with low sensitivity and are time-consuming, while the Taqman probe-based methods require the design and synthesis of specific probes, resulting in high detection prices (Wang et al 2017 ).…”

Establishment of SYBR green I-based quantitative real-time polymerase chain reaction for the rapid detection of a novel Chaphamaparvovirus in cats

“…Previous studies on FeChPV have not provided any methods with the ability to rapidly detect the target virus. The TaqMan method has been described in articles related to enteritis and upper respiratory tract diseases (Wang et al 2020a , b ). The qPCR assay has a sensitivity and specificity that are greater than those obtained with the cPCR assay (Di Martino et al 2020 ; Wang et al 2020a , b , 2021a , b ; Zheng et al 2020 ).…”

“…Hamaparvovirinae has a broad host range, including vertebrates and invertebrates. ChPV is one of the genera in Hamaparvovirinae and has been reported to infect various animals, such as dogs, cats, rats, barn owls, peafowls, mice, tilapias, red-crowned cranes, pigs, ducks, and chickens (Di Profio et al 2021 ; Yang et al 2016 ; Hargitai et al 2021 ; Liu et al 2020 ; Ge et al 2020 ; Du et al 2019 ; Wang et al 2019 , 2020a , b ; Vibin et al 2020 ; Lima et al 2019 ; Mohamed et al, 2013 ). Feline Chaphamaparvovirus (FeChPV) was first identified in fecal samples of cats from animal shelters, where outbreaks of diarrhea and vomiting occurred in Canada in 2019 (Li et al 2020 ).…”

Emerging Parvoviruses in Domestic Cats

“…FBoV-1 infection could pose a concrete health risk to cats since it has been associated with enteritis [104,105]. Several studies have shown that FBoV-1 is more likely to be detected in cats with diarrhea than in healthy cats [93,[102][103][104].…”