Nested reverse transcription polymerase chain reaction for the detection of rubella virus in clinical specimens.
OBJECTIVE
Rubella virus (RV) is one of the leading causes of childhood blindness in India. In this study we applied an optimized nested reverse transcription polymerase chain reaction (nRT-PCR) to detect RV in clinical specimens.


METHODS
nRT-PCR was optimized using total RNA extracted from standard strain of RV using nested sets of primers specific for E1 open reading frame. nRT-PCR was applied onto 30 lens aspirates and corresponding peripheral blood leucocytes of 30 infants with congenital (29)/ developmental (01) cataract. Serology for anti-RV IgG and IgM antibodies was done. RV isolation was attempted using Vero and SIRC cell cultures.


RESULTS
Optimized nRT-PCR was specific for RV and sensitive to detect 10 fg of RV RNA. Among 30 patients, nRT-PCR detected presence of RV in lens aspirates of 6 (20%) and 4 corresponding leucocytes. RV was isolated from 3 (10%) lens aspirates (nRT-PCR positive) of the 30 patients. Sera of these 6 patients showed presence of anti-RV IgG and IgM in one, only anti-RV IgG in 3 others and none in the other two. Of the remaining 24 patients, anti-RV IgG was detected in 3 and no anti-RV IgM antibodies in others.


CONCLUSIONS
Findings of our study showed that the nRT-PCR was a more sensitive and rapid technique to detect RV from lens aspirates compared to conventional methods of virus isolation and serology.
Shyamala
G
G
L & T Microbiology Research Centre, Vision Research Foundation, Sankara Nethralaya, Chennai, India.
Malathi
J
J
Moses
Y S
YS
Therese
K L
KL
Madhavan
H N
HN
D000818:Animals; D002386:Cataract / Q000151:congenital / Q000821:virology; D002522:Chlorocebus aethiops; D006801:Humans; D007223:Infant; D007231:Infant, Newborn; D007908:Lens, Crystalline / Q000821:virology*; D016133:Polymerase Chain Reaction / Q000379:methods*; D012367:RNA, Viral / Q000032:analysis; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*; D012409:Rubella / Q000151:congenital / Q000821:virology; D012412:Rubella virus / Q000235:genetics / Q000302:isolation & purification*; D014709:Vero Cells
D012367:RNA, Viral