Detection and strain differentiation of feline calicivirus in conjunctival swabs by RT-PCR of the hypervariable region of the capsid protein gene.
Reverse transcription-polymerase chain reaction (RT-PCR) was used to direct the amplification of a 670 to 680 base pair segment that included the hypervariable regions of the capsid protein gene of feline calicivirus (FCV). The segment was amplified from 13/13 cultivated FCV strains including 12 field isolates collected over 18 years. The sensitivities of culture and this RT-PCR for the detection of FCV in conjunctival swabs over the course of infection were then compared and correlated with clinical signs in 5 vaccinated and 3 unvaccinated experimentally-infected cats. Conjunctival swabs were taken daily from days 0 to 14 and on days 16, 19, 21 and 24 after challenge. FCV was detected in 19/144 swabs by RT-PCR and 16/144 swabs by culture. Virus detection correlated poorly with clinical signs regardless of the assay used. The Sau3AI restriction profiles of the RT-PCR products amplified from both cultivated strains and clinical samples were compared. All 13 cultivated isolates, including the 12 field isolates, exhibited different profiles, whereas all profiles from the experimentally-infected cats were identical, and matched the profile of the challenge/vaccine strain. This study has established that the RT-PCR assay described is as sensitive as culture for detection of FCV in conjunctival swabs, that a broad range of field isolates can be detected and rapidly differentiated by restriction endonuclease digestion, and that the assay thus meets the requirements for large scale epidemiological studies of FCV infections in cats. However, it has also shown that even the use of RT-PCR on conjunctival swabs alone is insufficient for accurate diagnosis of FCV infection in cats with conjunctivitis.
Department of Veterinary Science, University of Melbourne, Parkville, Victoria, Australia.
J E
JE
Sykes
V P
VP
Studdert
G F
GF
Browning
D000818:Animals; D001483:Base Sequence; D017250:Caliciviridae Infections / Q000175:diagnosis / Q000662:veterinary / Q000821:virology; D017927:Calicivirus, Feline / Q000145:classification / Q000235:genetics* / Q000302:isolation & purification*; D002213:Capsid / Q000235:genetics*; D002371:Cat Diseases / Q000175:diagnosis / Q000821:virology; D002415:Cats; D003228:Conjunctiva / Q000821:virology; D017931:DNA Primers / Q000235:genetics; D015252:Deoxyribonucleases, Type II Site-Specific; D005260:Female; D005814:Genes, Viral; D008297:Male; D016133:Polymerase Chain Reaction / Q000379:methods* / Q000706:statistics & numerical data; D012367:RNA, Viral / Q000235:genetics / Q000302:isolation & purification; D012680:Sensitivity and Specificity; D013045:Species Specificity; D014776:Virus Cultivation / Q000706:statistics & numerical data
D017931:DNA Primers; D012367:RNA, Viral; D015252:Deoxyribonucleases, Type II Site-Specific; C024405:GATC-specific type II deoxyribonucleases