Typing of human group A rotavirus with alkaline phosphatase-labeled oligonucleotide probes.
Rotavirus (RV) in stools of children less than 1 year of age with diarrhea in Bangkok in 1989 were serotyped by monoclonal enzyme immunoassay (MEIA). RNA extracted from these specimens was tested for hybridization with alkaline phosphatase (AP) and 32P-labeled oligonucleotides constructed from the nucleotide sequences of VP7 of human G types 1 (HuG1Ac), 2 (HuG2Ac), 3 (HuG3Ac), and 4 (HuG4Ac). Of 148 specimens that contained RV, 72% (106/148) hybridized with RV G type specific AP-labeled oligonucleotides compared to 47% (70/148) that were serotyped by MEIA (P less than 0.001). Of 68 specimens that contained only one VP7 serotype (G-type), as identified by MEIA, 94% (16/17) of G1, 90% (27/30) of G2, 57% (4/7) of G3, and 36% (5/14) of G4 RV hybridized with the AP-labeled HuG1Ac, HuG2Ac, HuG3Ac, and HuG4Ac oligonucleotides, respectively. The probes for G1, 2, 3, and 4 RV were specific for each G type. The results of hybridizing specimens with 32P- and AP-labeled oligonucleotides were similar. After transcription and amplification of cDNA of gene 9, AP-labeled RV G type specific oligonucleotides hybridized with 90% (134/148) of RV specimens. The high sensitivity of these nonimmunological techniques could be of value in identifying G types of RV during vaccine trials.
Sethabutr
O
O
Armed Forces Research Institute of Medical Sciences, Children's Hospital, Bangkok, Thailand.
Hanchalay
S
S
Lexomboon
U
U
Bishop
R F
RF
Holmes
I H
IH
Echeverria
P
P
D000469:Alkaline Phosphatase; D006801:Humans; D007223:Infant; D015336:Molecular Probe Techniques / Q000706:statistics & numerical data; D015345:Oligonucleotide Probes; D012401:Rotavirus / Q000145:classification* / Q000235:genetics* / Q000276:immunology; D012400:Rotavirus Infections / Q000382:microbiology / Q000517:prevention & control; D012680:Sensitivity and Specificity; D012703:Serotyping / Q000379:methods; D013785:Thailand; D014765:Viral Vaccines / Q000302:isolation & purification
D015345:Oligonucleotide Probes; D014765:Viral Vaccines; D000469:Alkaline Phosphatase