Molecular Diagnosis of Human Adenoviruses D and E by a Phylogeny-Based Classification Method Using a Partial Hexon Sequence

Abstract

Human adenoviruses (HAdVs) are the major causes of a variety of acute illnesses. Virus isolation and neutralization tests are usually done to identify the causative virus, but these tests are labor-intensive and time-consuming, and standardized antisera are in limited supply. This study investigated a rapid and reliable method of virus identification based on PCR and phylogenetic analysis. The phylogenetic tree constructed by neighbor joining on the basis of the newly determined partial hexon sequences from 33 prototypes of HAdV-D and -E, along with 11 available prototypes of HAdV-A to -C and -F from GenBank, allowed HAdVs to be grouped into six distinct clusters. These clusters correspond closely to the six newly designated species, HAdV-A to -F. The partial hexon sequences of 57 isolates from patients with acute conjunctivitis obtained over 20 years plus those of 44 prototype strains were analyzed. Each isolate formed a monophyletic cluster along with its respective prototype strain, allowing serotype identification. Partial-hexon-based classification appears to be an effective tool for studying the molecular epidemiology of HAdVs.

Epidemic Keratoconjunctivitis Due to the Novel Hexon-Chimeric-Intermediate 22,37/H8 Human Adenovirus

“…We have recently determined the partial hexon sequences of all 51 prototype strains and developed a rapid and reliable method for diagnosis based on phylogenetic analysis (22,28). This method successfully classified the 51 prototype strains of HAdVs into the six designated species, as approved by the International Committee on Taxonomy of Viruses; we have applied molecular diagnosis to identify hundreds of isolates or swabs obtained over the last 30 years from patients with EKC and lower respiratory tract infections from different parts of the world (5,6,18,22,28). Through such study, we found another novel HAdV from nosocomially infected patients with EKC (17).…”

“…Serotyping of HAdVs is generally performed by virus isolation followed by a neutralization test (NT) and hemagglutination inhibition test (HI) using type-specific antiserum (29). To date, amplification of the genome by PCR and determination of the nucleotide sequences followed by phylogenetic analysis have become the general procedure for both the classification and identification of HAdVs (21,22,28). In 2003, we encountered an EKC outbreak in Japan (Infectious Agents Surveillance Report [Weekly reports of adenovirus isolation/detection from epidemic keratoconjunctivitis cases, [2001][2002][2003][2004][2005][2006][2007]; http://idsc.nih.go.jp/iasr/prompt/graph/ad3.gif]) and detected HAdV DNA in 67 swabs from EKC patients who visited five eye clinics in different parts of Japan.…”

Molecular Epidemiology of Adenoviral Conjunctivitis in Hanoi, Vietnam

“…2 RT-PCR was performed to amplify enterovirus genome including the entire VP4 region as described. 3 …”

Development of a PCR-Based Assay for Detection, Quantification, and Genotyping of Human Adenoviruses

“…Today, in many laboratories, genotyping by sequencing or restriction enzyme analysis of PCR products has replaced serotyping of a virus isolate by neutralization assays (9,14,20,23,25 ). These molecular methods are much faster, but they still take a few days until a diagnosis is confirmed.…”