The detection and quantification of feline immunodeficiency provirus in peripheral blood mononuclear cells using the polymerase chain reaction.
The polymerase chain reaction method (PCR) was used to detect feline immunodeficiency virus proviral DNA in peripheral blood mononuclear cells (PBMC) of a group of 8 experimentally infected cats. The proportion of PBMC containing provirus was determined from 6 to 32 weeks post inoculation (p.i.) by performing PCR on serially diluted samples of PBMC. Primers from the p15 and p24 regions of the gag gene were used and Southern hybridization using an end-labelled probe was required to confirm primer-specific products. Provirus was detected in 5 of 8 cats by 6 weeks p.i. in 50000 PBMC, and in all 8 infected cats by 8 weeks p.i. Provirus was not detected in PBMC from any of 3 FIV negative cats. The proportion of PBMC containing provirus in individual cats ranged from 1 in 70 to 1 in 99600 PBMC. There was no significant decline over time in the proportion of PBMC containing provirus. Sequencing of a segment (287 bases) of the gag region of a West Australian FIV isolate (T90) revealed only slight nucleotide divergence from the North American Petaluma and PPR isolates and wider divergence from the Japanese TM2 clone.
Lawson
M
M
School of Biological and Environmental Sciences, Murdoch University, Australia.
Meers
J
J
Blechynden
L
L
Robinson
W
W
Greene
W
W
Carnegie
P
P
D000818:Animals; D001483:Base Sequence; D015139:Blotting, Southern / Q000662:veterinary; D002415:Cats; D002460:Cell Line; D016384:Consensus Sequence; D017931:DNA Primers / Q000737:chemistry; D004279:DNA, Viral / Q000032:analysis / Q000737:chemistry; D016181:Feline Acquired Immunodeficiency Syndrome / Q000382:microbiology*; D015683:Gene Products, gag / Q000235:genetics; D015750:Genes, gag; D016089:Immunodeficiency Virus, Feline / Q000235:genetics / Q000254:growth & development / Q000302:isolation & purification*; D007963:Leukocytes, Mononuclear / Q000382:microbiology*; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction / Q000662:veterinary; D011533:Proviruses / Q000235:genetics / Q000254:growth & development / Q000302:isolation & purification*; D012367:RNA, Viral / Q000737:chemistry / Q000235:genetics; D011897:Random Allocation; D012680:Sensitivity and Specificity; D014158:Transcription, Genetic; D014766:Viremia / Q000382:microbiology*
D017931:DNA Primers; D004279:DNA, Viral; D015683:Gene Products, gag; D012367:RNA, Viral