title: Genotype-specific fluorogenic RT-PCR assays for the detection and quantitation of canine coronavirus type I and type II RNA in faecal samples of dogs.
abstract: Two genotype-specific fluorogenic RT-PCR assays were developed for the detection and quantitation of canine coronavirus (CCoV) type I and type II RNA in the faeces of dogs with diarrhoea. Both the fluorogenic assays showed high specificity, sensitivity and reproducibility, allowing a precise quantitation of CCoV type I and type II RNA over a linear range of about eight orders of magnitude (from 10(1) to 10(8) copies of standard RNA). Comparison with genotype-specific gel-based RT-PCR assays revealed that the fluorogenic assays were more sensitive and more rapid than conventional amplifications, with a large increase in throughput. The genotype-specific fluorogenic assays were then used to detect and measure viral loads in the faecal samples collected from dogs naturally or experimentally infected with type I, type II, or both genotypes. Of 174 samples collected from naturally infected dogs, 77 were positive for CCoV type I and 46 for CCoV type II. Thirty-eight dogs were found to be infected naturally by both genotypes, with viral RNA titres generally higher for type I in comparison to type II. At the same time, dogs infected experimentally shed type I RNA with higher titres with respect to type II.
authors: Department of Animal Health and Well-being, Faculty of Veterinary Medicine of Bari, Strada prov per Casamassima Km 3, 70010 Valenzano, Bari, Italy. n.decaro@veterinaria.uniba.it
authors: Nicola
authors: N
authors: Decaro
authors: Vito
authors: V
authors: Martella
authors: Dominga
authors: D
authors: Ricci
authors: Gabriella
authors: G
authors: Elia
authors: Costantina
authors: C
authors: Desario
authors: Marco
authors: M
authors: Campolo
authors: Nicola
authors: N
authors: Cavaliere
authors: Livia
authors: L
authors: Di Trani
authors: Maria
authors: M
authors: Tempesta
authors: Canio
authors: C
authors: Buonavoglia
mesh_terms: D000818:Animals; D018352:Coronavirus Infections / Q000175:diagnosis / Q000662:veterinary*; D017939:Coronavirus, Canine / Q000235:genetics / Q000302:isolation & purification*; D003967:Diarrhea / Q000175:diagnosis / Q000662:veterinary*; D004283:Dog Diseases / Q000175:diagnosis*; D004285:Dogs; D005243:Feces / Q000821:virology; D005453:Fluorescence; D007558:Italy; D012367:RNA, Viral / Q000032:analysis*; D015203:Reproducibility of Results; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012680:Sensitivity and Specificity; D013045:Species Specificity
chemical_list: D012367:RNA, Viral