Comparison of primer sets for detection of fecal and ocular adenovirus infection using the polymerase chain reaction

Abstract

Adenoviruses of subgenus F (types 40 and 41) cause infantile gastroenteritis and adenoviruses principally of types 1-7 are found in feces during respiratory or generalized infections. Adenoviruses (mostly types 3, 4, 8, 19, or 37) are also linked with follicular or epidemic conjunctivitis. The diagnostic efficiency of the polymerase chain reaction (PCR) for adenoviruses was assessed using genus-reactive primers H1 and H2 or JCH1 and JCH2 or subgenus F-specific primers F1a and F2a. With diarrheal stool specimens containing subgenus F adenoviruses, F1a/F2a PCR achieved at least as high a positivity rate (75/76 [99%]) as electron microscopy (72/76 [95%]) and was more sensitive than polyclonal antibody-based immune electron microscopy (IEM) for subgenus identification (75/76 [99%] vs. 66/76 [87%], P = 0.008). Twenty-three subgenus F strains untypeable by monoclonal antibody-based IEM were typed as 40 (n = 4) or 41 (n = 19) by Hha I digestion of the PCR product. The genus-reactive primer pairs provided DNA amplification assays of generally equal efficiency on conjunctival swab specimens though possible nucleic acid degradation in DNA extracts during storage could have meant that JCH1/JCH2 PCR was truly the more sensitive. The use of either genus-reactive primer set on fecal specimens cannot be recommended because, although the positivity rates with subgenus F PCR positive specimens were high (70/75 [93%] for H1 and H2, 14/15 [93%] for JCH1 and JCH2), the detection rates were disappointing with similar specimens yielding nonsubgenus F adenoviruses.

Circulation of a Novel Pattern of Infections by Enteric Adenovirus Serotype 41 among Children below 5 Years of Age in Kolkata, India

“…The primers gave an amplicon of 668 bp. The partial shaft region of the fiber genes was also amplified using primers AdFfib(ϩ) (5Ј-ACTTAATGCTGACACGGG CAC-3Ј) and AdFfib(Ϫ) (5Ј-TAATGTTTGTGTTACTCCGCTC-3Ј) for phylogenetic analysis (16,53). PCR was performed in a 0.2-ml reaction tube in a total volume of 25 l containing 5 l of 5ϫ Go Taq reaction buffer, 0.5 l of a 10 mM deoxynucleoside triphosphate (dNTP) mixture, 0.2 l Go Taq polymerase (5 U/l), 1 l of each primer at 10 pM, and 2 l of nucleic acid extract, according to the manufacturer's instructions (Promega Corporation, Madison, WI).…”

Dexamethasone/Povidone Eye Drops versus Artificial Tears for Treatment of Presumed Viral Conjunctivitis: A Randomized Clinical Trial

“…In addition, each patient was asked to report their opinions on the usefulness of the treatment in relieving their symptoms on the following four-point scale: the treatment did not help (0), I'm unsure if the treatment helped (1), I think the treatment helped (2) and the treatment definitely helped (3). Both the questionnaires and the criteria for selecting patients were similar to a previously published conjunctivitis clinical trial.…”

“…An American study on adults and children in an ophthalmic emergency room found that 62% of acute infective conjunctivitis cases had an adenoviral cause. [1][2][3] Correspondence: Roberto Damian Pacheco Pinto. E-mail: rpacheco42@gmail.com Generally, a diagnosis of viral conjunctivitis is made on the clinical features alone.…”

Identification and characterization of enteric adenoviruses in infants and children hospitalized for acute gastroenteritis

“…PCR was carried out in 50 ml reaction mix containing 2 ml of DNA, 10Â Taq DNA polymerase buffer, dNTPmix (10 mM), adenovirus hexon gene primers (30 pmol/ml) 19,136-19,158 bp) [Allard et al, 1990], and Fiber gene primers (25 pmol/ml) F1-5 0 -CTG CCA GTG TTA AGC ATA ATA TG-3 0 (2,689-2,711 bp), F2-5 0 -GTC ACG TTC AGA GTT GCT GGA TAT-3 0 (3,053-3,077 bp) [Hussain et al, 1996], 0.25 ml of Taq DNA Polymerase (3 U/ml, Bangalore Genei Pvt. Ltd., Banglore, India).…”