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

Abstract

Feline parvovirus causes infectious diseases, and Chaphamaparvovirus is a novel type of feline parvovirus. The present study aims to establish a method that can be used in clinical rapid detection of feline Chaphamaparvovirus (FeChPV), for facilitate the timely and effective diagnosis and treatment of sick animals and shorten the diagnosis time of clinical diseases. The experimental samples in this study are from 20 cats undergoing physical examination in Hefei Xin’an Animal Hospital. An SYBR Green I-based qPCR assay was performed to detect FeChPV. A pair of specific primers was designed based on the VP1 gene to perform the assay. The detection assay showed high sensitivity with a detection limit of 1.07 × 10 1 copies/μL and high specificity for detection of only the target virus. The coefficients of C t value variation were calculated to assess the reproducibility of the qPCR assay, and the inter- and intra-assay ranged from 0.21 to 0.67% and 0.10 to 0.56%, respectively. The result of clinical sample detection showed that the infection rate of FeChPV in 124 samples detected using qPCR assay was higher than that with conventional PCR. The established qPCR assay could be a low-cost, convenient, and reliable method to detect FeChPV in clinical practice.

Detection of FeChPV in a cat shelter outbreak of upper respiratory tract disease in China

“…To isolate viral DNA from tissue samples, each tissue sample (1 g; including heart, liver, spleen, lung, kidney, brain, cervical lymph node, nasal turbinate, trachea, stomach, duodenum, jejunum, ileum, cecum, and colon tissues) was first homogenized in phosphate-buffered saline (1 ml) using a homogenizer. Referring to previous studies ( Liu et al, 2022 ), the virus DNA loads of FeChaPV were determined using the SYBR Green I method (AbColonal, China) on an LC480 instrument (Roche). The primers were synthesized by Sangon Biotech (Shanghai, China) and are shown in Table 1 .…”

“…An increasing number of novel parvoviruses have been identified, and thus far, the Parvoviridae family comprises three subfamilies: Parvovirinae , Hamaparvovirinae , and Densovirinae ( Penzes et al, 2020 ; Abayli and Can-Sahna, 2022 ). Hamaparvovirinae has a broad host range, including vertebrates and invertebrates ( Liu et al, 2022 ). Chaphamaparvovirus (ChPV), a genus in Hamaparvovirinae , has been reported to have a large host range, including mammals, such as dogs, cats, black bears, rats, mice, sheep, and pigs ( Yang et al, 2016 ; Alex et al, 2020 ; Ge et al, 2020 ; Wang et al, 2020 ; Di Profio et al, 2022 ); birds, such as barn owls, peafowl, paradise tanagers, red-crowned cranes, parrots, ducks, and chickens ( Lima et al, 2019 ; Wang et al, 2019 ; Liu X. et al, 2020 ; Vibin et al, 2020 ; Hargitai et al, 2021 ; Sarker, 2021 ); and even fish, such as tilapia ( Du et al, 2019 ).…”