Abstract
The aim of this study was to identify potential ligands and develop a novel diagnostic test to pathogenic bovine rotavirus (BRV) using phage display technology. The viruses were used as an immobilized target followed by incubation with a 12-mer phage display random peptide library. After five rounds of biopanning, phages had a specific binding activity to BRV were isolated. DNA sequencing indicated that phage displayed peptides HVHPPLRPHSDK, HATNHLPTPHNR or YPTHHAHTTPVR were potential ligands to BRV. Using the specific peptide-expressing phages, we developed a phage-based ELISA to differentiate BRV from other viruses. Compared with quantitative real-time PCR (qPCR), the phage-mediated ELISA was more suitable for the capture of BRV and the detection limitation of this approach was 0.1 µg/ml of samples. The high sensitivity, specificity and low cross-reactivity for the phage-based ELISA were confirmed in receiver operating characteristics (ROC) analysis.
Highlights
Neonatal calf diarrhea (NCD) is a common gastroenteritis infection worldwide, threatening the cattle production and causing substantial economic losses
Some diagnostic procedures such as viral culture, RT-PCR and serology have been proved to be useful for detection of the pathogens, the World Health Organization recommends the use of enzyme immunoassays for the diagnosis of rotavirus infections [7]
Phage display technology is a good approach for ligand identification of targets and phage-displayed peptides from combinatorial libraries that interacting with hepatitis B virus, adenovirus type 2, Andes virus, Sin Nombre virus, Hantaan virus, Avian H5N1 Virus and coronavirus have been selected [17,18,19,20,21]
Summary
Neonatal calf diarrhea (NCD) is a common gastroenteritis infection worldwide, threatening the cattle production and causing substantial economic losses. Group A BRV, the major viral pathogen for NCD can further be differentiated into 23 G- (glycoprotein) and 32 P- (protease sensitive protein) types, based on the VP7 and VP4 antigens, respectively [4]. The clinical course following rotavirus infection is typically short, virus can be detected in faeces for up to 3 weeks post-infection [6]. Some diagnostic procedures such as viral culture, RT-PCR and serology have been proved to be useful for detection of the pathogens, the World Health Organization recommends the use of enzyme immunoassays for the diagnosis of rotavirus infections [7]
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