Dataset Information

Isolation and identification of feline herpesvirus type 1 from a South China tiger in China.

ABSTRACT: In 2012, an FHV-1-like virus was isolated from a tiger that presented with clinical signs of sialorrhea, sneezing and purulent rhinorrhea. Isolation was performed with the FK81 cell line, and the virus was identified by PCR, transmission electron microscopy (TEM), and the phylogenetic analysis of the partial thymidine kinase (TK) and glycoprotein B (gB) genes. A total of 253 bp of the TK gene and 566 bp of the gB gene were amplified from the trachea of the tiger by PCR/RT-PCR method. Phylogenetic analysis showed that the isolate belonged to the same cluster with other FHV-1 strains obtained from GenBank. Herpes-like viruses with an envelope and diameters of approximately 200 nm were observed in the culture supernatants of FK81 cells inoculated with samples from the tiger. The FHV-1 infection was confirmed by an animal challenge experiment in a cat model. Our finding extends the host range of FHV-1 and has implications for FHV-1 infection and South China tiger conservation.

SUBMITTER: Sun H

PROVIDER: S-EPMC3970135 | biostudies-literature | 2014

REPOSITORIES: biostudies-literature

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Isolation and identification of feline herpesvirus type 1 from a South China tiger in China.

Sun Heting H Li Yuanguo Y Jiao Weiyi W Liu Cunfa C Liu Xiujuan X Wang Haijun H Hua Fuyou F Dong Jianxiu J Fan Shengtao S Yu Zhijun Z Gao Yuwei Y Xia Xianzhu X

Viruses 20140228 3

In 2012, an FHV-1-like virus was isolated from a tiger that presented with clinical signs of sialorrhea, sneezing and purulent rhinorrhea. Isolation was performed with the FK81 cell line, and the virus was identified by PCR, transmission electron microscopy (TEM), and the phylogenetic analysis of the partial thymidine kinase (TK) and glycoprotein B (gB) genes. A total of 253 bp of the TK gene and 566 bp of the gB gene were amplified from the trachea of the tiger by PCR/RT-PCR method. Phylogeneti ...[more]

PMID: 24590411

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Project description:BackgroundFeline herpesvirus type 1 (FHV) and Feline calicivirus (FCV) are the primary co-infecting pathogens that cause upper respiratory tract disease in cats. However, there are currently no visual detection assays available for on-site testing. Here, we develop an ultrasensitive and visual detection method based on dual recombinase polymerase amplification (dRPA) reaction and the hybrid Cas12a/Cas13a trans-cleavage activities in a one-tube reaction system, referred to as one-tube dRPA-Cas12a/Cas13a assay.ResultsThe recombinant plasmid DNAs, crRNAs, and RPA oligonucleotides targeting the FCV ORF1 gene and FHV-1 TK gene were meticulously prepared. Subsequently, dual RPA reactions were performed followed by screening of essential reaction components for hybrid CRISPR-Cas12a (targeting the FHV-1 TK gene) and CRISPR-Cas13a (targeting the FCV ORF1 gene) trans-cleavage reaction. As a result, we successfully established an ultra-sensitive and visually detectable method for simultaneous detection of FCV and FHV-1 nucleic acids using dRPA and CRISPR/Cas-powered technology in one-tube reaction system. Visual readouts were displayed using either a fluorescence detector (Fluor-based assay) or lateral flow dipsticks (LDF-based assay). As expected, this optimized assay exhibited high specificity towards only FHV-1 and FCV without cross-reactivity with other feline pathogens while achieving accurate detection for both targets with limit of detection at 2.4 × 10- 1 copies/μL for the FHV-1 TK gene and 5.5 copies/μL for the FCV ORF1 gene, respectively. Furthermore, field detection was conducted using the dRPA-Cas12a/Cas13a assay and the reference real-time PCR methods for 56 clinical samples collected from cats with URTD. Comparatively, the results of Fluor-based assay were in exceptional concordance with the reference real-time PCR methods, resulting in high sensitivity (100% for both FHV-1 and FCV), specificity (100% for both FHV-1 and FCV), as well as consistency (Kappa values were 1.00 for FHV-1 and FCV). However, several discordant results for FHV-1 detection were observed by LDF-based assay, which suggests its prudent use and interpretaion for clinical detection. In spite of this, incorporating dRPA-Cas12a/Cas13a assay and visual readouts will facilitate rapid and accurate detection of FHV-1 and FCV in resource-limited settings.ConclusionsThe one-tube dRPA-Cas12a/Cas13a assay enables simultaneously ultrasensitive and visual detection of FHV-1 and FCV with user-friendly modality, providing unparalleled convenience for FHV-1 and FCV co-infection surveillance and decision-making of URTD management.

Dataset Information

Isolation and identification of feline herpesvirus type 1 from a South China tiger in China.

Publications

Isolation and identification of feline herpesvirus type 1 from a South China tiger in China.

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