Ngan Thi Kim Pham

Isolation and Molecular Characterization of Aichi Viruses from Fecal Specimens Collected in Japan, Bangladesh, Thailand, and Vietnam

ABSTRACT Aichi virus is a new member of the family Picornaviridae, genus Kobuvirus, and is associated with human gastroenteritis. This study detected Aichi virus in 28 of 912 fecal specimens which were negative for rotavirus, adenovirus, norovirus, sapovirus, and astrovirus and were collected in Japan, Bangladesh, Thailand, and Vietnam during 2002 to 2005.

Diversity of Human Parechoviruses Isolated from Stool Samples Collected from Thai Children with Acute Gastroenteritis

ABSTRACT A total of 82 fecal specimens which were known to be negative for rotavirus, adenovirus, norovirus, sapovirus, and astrovirus and which were collected from infants and children with acute gastroenteritis in Chiang Mai, Thailand, from January to December 2005 were screened for human parechovirus (HPeV). HPeV was detected by reverse transcription-PCR with a primer pair that amplified the 5′ untranslated region of its genome and was genotyped by sequencing of the VP1 region. HPeV was detected in 12 of 82 specimens tested, and the detection rate was found to be 14.6%. The capsid VP1 gene was successfully sequenced from nine of the HPeV strains detected. The HPeV strains studied clustered into four different genotypes, HPeV genotype 1 (HPeV1) to HPeV4, and the majority of the strains studied (five strains) belonged to HPeV1. This is the first finding of HPeV from children with acute gastroenteritis in Thailand. In addition, the diversity of the Thai HPeV strains was also noted.

Human Parechovirus Infection in Children Hospitalized with Acute Gastroenteritis in Sri Lanka

ABSTRACT Of 362 fecal specimens collected from infants and children hospitalized with acute gastroenteritis in Sri Lanka from September 2005 to August 2006, 30 (8.3%) were positive for human parechovirus (HPeV). Six different HPeV genotypes, including HPeV1, -3, -4, -5, -10, and -11, were identified, of these, HPeV11 was reported for the first time.

A novel RT-multiplex PCR for detection of Aichi virus, human parechovirus, enteroviruses, and human bocavirus among infants and children with acute gastroenteritis.

A novel reverse transcription-multiplex polymerase chain reaction assay was developed to detect Aichi virus, human parechovirus, enteroviruses, and human bocavirus. A mixture of four pairs of published specific primers, 6261 and 6779, ev22(+) and ev22(-), F1 and R1, 188F and 542R, was used to amplify the viral genomes and specifically generate four different amplicon sizes of 519, 270, 440, and 354 bp for Aichi virus, human parechovirus, enteroviruses, and human bocavirus, respectively. A total of 247 fecal specimens previously screened for rotavirus, adenovirus, norovirus, sapovirus, and astrovirus-negative, collected from infants and children with acute gastroenteritis in Japan from July 2007 to June 2008, were tested further for the presence of the four viruses, Aichi virus, human parechovirus, enteroviruses, and human bocavirus, by RT-multiplex PCR. The total detection rate of these viruses was 26.7% (66 out of 247 samples). Of these, HPeV, EVs, and HBoV were identified in 20, 41, and 5 specimens. No Aichi virus was found among these subjects. The sensitivity and specificity of RT-multiplex PCR were assessed and demonstrated a strong validation against RT-monoplex PCR. This is the first report of detecting these types of viruses in fecal samples from infants and children with acute gastroenteritis by RT-multiplex PCR.

Novel Human Parechovirus, Sri Lanka

Of 362 fecal samples collected from children with acute gastroenteritis in Sri Lanka during 2005–2006, 30 (8.3%) were positive for human parechovirus (HPeV) by reverse transcription–PCR. A novel HPeV, designated as HPeV10, was identified in 2 samples by sequence analysis of the viral protein 1 gene of the detected HPeVs.

Detection of human parechovirus in stool samples collected from children with acute gastroenteritis in Japan during 2007–2008

Of 477 stool specimens, which had been screened for rotavirus, adenovirus, norovirus, sapovirus and astrovirus, collected from infants and children with acute gastroenteritis in pediatric clinics encompassing five localities (Sapporo, Tokyo, Maizuru, Osaka, and Saga) in Japan from July 2007 to June 2008, 247 negative samples (51.7%) were subjected to screening for human parechovirus. Human parechovirus (HPeV) was detected by RT‐PCR using a primer pair to amplify 5′UTR region of its genome and was genotyped by sequencing of the VP1 gene. HPeV was detected in 20 of 247 specimens tested, and the detection rate was found to be 8.1%. Seventeen of the 20 strains that tested positive for HPeV were sequenced successfully the VP1 gene. The majority of the HPeV strains (n = 15) could be identified as HPeV1, and the remaining 2 strains could be typed as HPeV3. By phylogenetic and identical matrix analyses of HPeV VP1 sequences, HPeV1 should be divided into two lineages, and all of the Japanese studied HPeV1 strains belong to the lineage 2 accordingly. This is the first report of the circulation of HPeV, especially HPeV1 in Japan. J. Med. Virol. 83:331–336, 2011.

Human bocavirus infection in children with acute gastroenteritis in Japan and Thailand.

A total of 329 fecal specimens, which had been known to be negative for rotavirus, adenovirus, norovirus, sapovirus, and astrovirus, and which were collected from infants and children with acute gastroenteritis in Japan and Thailand during 2005–2008 were screened for human bocavirus (HBoV). HBoV was detected by PCR with a primer pair that amplified the NP1 region of its genome and was genotyped by sequencing of the VP1/VP2 region. Of the 329 samples tested, 6 (1.8%) were positive for HBoV. Of these, five samples were collected from Japan and one sample was from Thailand, and the detection rates of HBoV in each country were 2% and 1.2%, respectively. For the detected HBoV, the capsid VP1/VP2 gene of all HBoV strains was successfully sequenced. Four Japanese HBoV strains studied were clustered into group 1, while the remaining Japanese strain and a unique Thai strain belonged to group 2. No severe acute gastroenteritis associated with HBoV was noted. This study provides better understanding on the epidemiology of HBoV infections in children with acute gastroenteritis in Japan and Thailand. J. Med. Virol. 83:286–290, 2011.

Sequence Analysis of the VP7 Gene of Human Rotaviruses G2 and G4 Isolated in Japan, China, Thailand, and Vietnam During 2001-2003

Sequence and phylogenetic analyses of the rotavirus VP7 gene were performed on 52 human G2 and G4 strains isolated in Japan, China, Thailand, and Vietnam during 2001–2003. All genotype G2 strains included in the study clustered into lineage II of the phylogenetic tree, together with the majority of global G2 strains detected since 1995. The amino acid substitution at position 96 from aspartic acid to asparagine was noted among the emerging or re‐emerging G2 rotavirus strains in Japan, Thailand, and Vietnam during 2002–2003. Genotype G4 strains detected in Vietnam grouped into lineage Ia of the phylogenetic tree, whereas Japanese G4 strains clustered in lineage Ic which included emerging G4 strains from Argentina, Italy, Paraguay, and Uruguay. It is noteworthy that an insertion of asparagine was found at position 76 in all the Japanese strains and that its presence might be involved in the emergence of G4 rotavirus in Japan during 2002–2003. J. Med. Virol. 82: 878–885, 2010.

Sequence analysis of the capsid gene of Aichi viruses detected from Japan, Bangladesh, Thailand, and Vietnam.

Sequence analysis of the capsid gene of Aichi viruses was performed on 12 strains detected in Japan, Bangladesh, Thailand, and Vietnam during 2002–2005. The phylogenetic tree constructed from 17 nucleotide sequences of the capsid gene of the strains studied and reference strains demonstrated that Aichi virus strains clustered into two branches. A classification of Aichi viruses based on the capsid gene was proposed, in which lineage I consists of the Aichi virus strains detected from Japan, Thailand, Vietnam, and Germany, and lineage II includes Bangladeshi strains and a Brazilian strain. J. Med. Virol. 80: 1222–1227, 2008.

Phylogenetic analysis of rubella viruses in vietnam during 2009–2010†

Rubella virus (RV) usually causes a mild disease. However, infection during the first trimester of pregnancy often leads to severe birth defects known as congenital rubella syndrome (CRS). Although wild‐type RVs exist and circulate worldwide, their genotypes remain unknown in many countries. The aim of this study was to identify the molecular characteristics of RVs found in Vietnam during the years 2009–2010 and to provide the first data concerning RV genotypes in this country. Throat swab samples were collected between 2009 and 2010 from four CRS cases and nine rubella infection cases visiting one Childrens Hospital and one outpatient clinic in Ho Chi Minh City. The 739‐nucleotide coding region of the RV E1 gene recommended by the World Health Organization was amplified by reverse transcriptase PCR, and the resulting DNA fragments were then sequenced. Sequences were assigned to genotypes by phylogenetic analysis with RV reference strains. RV RNA was detected in 11 clinical specimens. Phylogenetic analysis of the sequences showed that all 11 strains belonged to 2B genotype. Several variations in amino acids were found, among which five changes were involved in the B and T cell epitopes. These data indicate that viruses of genotype 2B were circulating in Vietnam. The increasing information about RV genotype in Vietnam should aid in the control of rubella infection and CRS in this country. J. Med. Virol. 84:705–710, 2012.