Detection of adenovirus DNA in clinical samples by SYBR Green real-time polymerase chain reaction assay.
BACKGROUND
Adenoviruses are associated with a variety of diseases including upper respiratory tract infections, acute conjunctivitis, cystitis and gastroenteritis. Adenoviruses can also cause fatal disseminated infections in patients undergoing stem cell transplantation. Measurement of adenovirus load in clinical samples from localized adenovirus infections or disseminated adenovirus infections may provide important information for analyzing the pathogenesis of various adenovirus infections. The purpose of the present study was to develop and optimize a highly sensitive real-time polymerase chain reaction (PCR) assay to detect a wide range of adenoviruses and to detect adenovirus DNA in clinical samples from immunocompetent children.


METHODS
Clinical samples of throat swabs and blood were collected from 111 patients suspected of having adenovirus infection. The copy number of adenovirus DNA was measured by real-time PCR assay.


RESULTS
SYBR Green real-time PCR assay is able to detect 10-10(6) copies of standard adenovirus DNA per run. Adenovirus DNA was detected in all culture-positive samples serotyped as 1, 2, 3, 4, 5, 6, 8 and 11. Viral loads on throat swabs from immunocompetent children with adenovirus infection ranged from 10(5) to 10(11) copies/mL. Adenovirus DNA was detected in 60% of blood samples and copy number ranged from 10(3) to 10(5) copies/mL.


CONCLUSIONS
SYBR Green real-time PCR is a useful quantitative tool for analysis of adenovirus DNA. The present results for immunocompetent children with adenovirus infections provided basic data for comparison with data obtained from immunocompromised patients.
Watanabe
Masahiro
M
Suzuka Kodomo Clinic, Kobe City General Hospital, Kobe City, Japan.
Kohdera
Urara
U
Kino
Minoru
M
Haruta
Tsunekazu
T
Nukuzuma
Syoichi
S
Suga
Tomoko
T
Akiyoshi
Kyoko
K
Ito
Masahiro
M
Suga
Shigeru
S
Komada
Yoshihiro
Y
D000258:Adenovirus Infections, Human / Q000175:diagnosis*; D000260:Adenoviruses, Human / Q000235:genetics / Q000302:isolation & purification*; D052160:Benzothiazoles; D002675:Child, Preschool; D004279:DNA, Viral / Q000097:blood / Q000302:isolation & purification; D003959:Diamines; D005260:Female; D005456:Fluorescent Dyes; D006801:Humans; D008297:Male; D009930:Organic Chemicals; D010614:Pharynx / Q000821:virology; D016133:Polymerase Chain Reaction; D011804:Quinolines
D052160:Benzothiazoles; D004279:DNA, Viral; D003959:Diamines; D005456:Fluorescent Dyes; D009930:Organic Chemicals; D011804:Quinolines; C098022:SYBR Green I