Antibiotic Resistance of Campylobacter jejuni and C. coli Isolated from Children with Diarrhea in Thailand and Japan.

A total of 29 Campylobacter jejuni and C. coli strains were isolated from Thai and Japanese children with diarrhea using the Loop-mediated Isothermal Amplification method. The samples were evaluated for mutations in gyrA and 23S rRNA in order to assess resistance against fluoroquinolones and macrolides, respectively. Among the isolated strains, 9 (8 C. jejuni and 1 C. coli) were from Thai children, and the other 20 (C. jejuni) were isolated from Japanese children. High fluoroquinolone resistance rates were observed in Thai (66.7%) and Japanese (90%) children. Macrolide resistance was not observed in Japanese children but was observed at a considerable rate of 12.5% of C. jejuni isolated in the Thai cohort. The results indicate that continuous monitoring of resistance of Campylobacter strains to fluoroquinolones and macrolides is definitely necessary.

Rapid diagnostic tests apply for pediatric infections at outpatient clinic setting.

BACKGROUND: Early identification of the etiology of infection is beneficial. Most infections are treated as outpatients. However, facilities for rapid diagnosis are not available in clinic settings. METHODS: We applied Immunochromatography (IC) and Loop-mediated Isothermal Amplification (LAMP) methods to rapidly diagnose pathogens among 31 children with respiratory infection and 12 with gastroenteritis at a clinic in Saitama prefecture, Japan. Pathogens were then screened by multiplex conventional and real-time PCRs and bacterial culture. RESULTS: Respiratory pathogens were found in 64.5%. Despite the narrow spectrum, rapid tests identified pathogens in 28.6% of cases with a high agreement rate of 89.3% with PCR. Gastroenteritis pathogens were found in 66.7%. E. coli was positive in 3 cases and all were negative for verotoxin by LAMP. The agreement rate of IC and PCR assay was high, 100%. CONCLUSIONS: IC and LAMP are reliable and suitable methods in limited-resource settings for early pathogenic identification, which will help appropriate management, avoid unnecessary intervention, and cost saving.

Molecular epidemiology of influenza A virus infection among hospitalized children in Vietnam during post-pandemic period.

Genetic variability makes influenza virus to escape the immunity and causes yearly epidemics. Monitoring those changes is necessary for vaccine selection. In addition, H3N2 viruses were considered to be seeded from Southeast Asia before spreading globally. This study described the molecular epidemiology of influenza A during the post-pandemic season 2010-2011 in Vietnam. Nasopharyngeal samples were collected from children with respiratory infections at Children's Hospital 2, Ho Chi Minh City. The HA, NA, M genes were amplified, sequenced and analyzed. Thirty-five of 1,082 (3.2%) patients were positive for influenza A, including 14 pandemic H1N1 2009 (H1N1pdm09) and 21 H3N2 infections. H3N2 was dominant in the rainy season (May-October 2010) while H1N1pdm09 was dominant in the dry season (November 2010-April 2011). Phylogenetic analysis showed that Vietnamese H1N1pdm09 sequences in 2010-2011 formed the distinct cluster, with other contemporary Asian and 2012-American sequences, suggesting a possible common ancestor. All were oseltamivir-sensitive except two strains carrying S247N and D199N in NA which reduced the neuraminidase inhibitor susceptibility. The Vietnamese H3N2 viruses in mid-2010 belonged to the emerging subclade Perth10/2010, which then spread worldwide in 2011. The Vietnamese influenza viruses were well matched with the Southern Hemisphere vaccine formulation. Mutations at antigenic sites were also identified in these viruses. Surveillance of influenza viruses in tropical countries is important not only for development of their prevention and control strategies but also for earlier identification of the newly emerged strains that may be selected for future vaccine.

Sensitive and rapid detection of campylobacter species from stools of children with diarrhea in Japan by the loop-mediated isothermal amplification method.

We detected Campylobacter spp. in 5% (20/380) of diarrheal stool samples collected at an outpatient clinic in Kyoto using a commercial loop-mediated isothermal amplification (LAMP) kit with a fluorescent detection reagent after DNA extraction. The sensitivity and specificity were 100% in comparison with those of semi-nested PCR for the differentiation of Campylobacter jejuni and Campylobacter coli. Fourteen of the 20 samples were already determined as C. jejuni by the culture method. All 20 samples were also positive for C. jejuni by the PCR method. Among the 58 cultured samples, the sensitivity of the culture method against the LAMP method was 93.3% (14/15) and the specificity was 100% (43/43). The detection rate of Campylobacter spp. from the heated supernatants by the LAMP method was lower than that from the supernatant after DNA extraction. In total, 25% (5/20) of the Campylobacter-positive samples by the LAMP method were co-infected with norovirus (3/20), rotavirus (1/20), and human parechovirus (1/20), although no other bacterial co-infection was identified by the culture method. C. jejuni was mostly detected in children aged >5 years throughout the year. Based on these results, we concluded that care should be taken while diagnosing Campylobacter infection in children. Our newly modified LAMP method is a rapid, easy, and useful method for this diagnosis.

Human bocavirus in children with acute respiratory infections in Vietnam.

Acute respiratory infections are the major cause of morbidity and mortality globally. Human bocavirus (HBoV), a novel virus, is recognized to increasingly associate with previously unknown etiology respiratory infections in young children. In this study, the epidemiological, clinical, and molecular characteristics of HBoV infections were described in hospitalized Vietnamese pediatric patients. From April 2010 to May 2011, 1,082 nasopharyngeal swab samples were obtained from patients with acute respiratory infections at the Children's Hospital 2, Ho Chi Minh City, Vietnam. Samples were screened for HBoV by PCR and further molecularly characterized by sequencing. HBoV was found in 78 (7.2%) children. Co-infection with other viruses was observed in 66.7% of patients infected with HBoV. Children 12-24 months old were the most affected age group. Infections with HBoV were found year-round, though most cases occurred in the dry season (December-April). HBoV was possible to cause severe diseases as determined by higher rates of hypoxia, pneumonia, and longer hospitalization duration in patients with HBoV infection than in those without (P-value <0.05). Co-infection with HBoV did not affect the disease severity. The phylogenetic analysis of partial VP1 gene showed minor variations and all HBoV sequences belonged to species 1 (HBoV1). In conclusion, HBoV1 was circulating in Vietnam and detected frequently in young children during dry season. Acute respiratory infections caused by HBoV1 were severe enough for hospitalization, which implied that HBoV1 may have an important role in acute respiratory infections among children.

Molecular epidemiology and disease severity of human respiratory syncytial virus in Vietnam.

Respiratory syncytial virus (RSV) is a major cause of acute respiratory infections (ARIs) in children worldwide and can cause high mortality, especially in developing countries. However, information on the clinical and molecular characteristics of RSV infection in developing countries is limited. From April 2010 to May 2011, 1,082 nasopharyngeal swabs were collected from children with ARI admitted to the Children's Hospital 2, Ho Chi Minh City, Vietnam. Samples were screened for RSV and genotyped by reverse transcription-PCR and sequencing. Demographic and clinical data was also recorded. RSV was found in 23.8% (257/1,082) of samples. RSV A was the dominant subgroup, accounting for 91.4% (235/257), followed by RSV B, 5.1% (13/257), and 9 cases (3.5%) were mixed infection of these subgroups. The phylogenetic analysis revealed that all group A strains belonged to the GA2 genotype. All group B strains belonged to the recently identified BA genotype, and further clustered into 2 recently described subgenotypes BA9 and BA10. One GA2 genotype strain had a premature stop codon which shortened the G protein length. RSV infection was significantly associated with younger age and higher severity score than those without. Co-infection with other viruses did not affect disease severity. RSV A caused more severe disease than RSV B. The results from this study will not only contribute to the growing database on the molecular diversity of RSV circulating worldwide but may be also useful in clinical management and vaccine development.

Evaluation and comparison of the efficiency of immunochromatography methods for norovirus detection.

BACKGROUND: The efficiency of a commercial immunochromatography kit (IP-NoV) was evaluated for rapid detection of norovirus in Japanese fecal specimens and its sensitivity and specificity were compared with two other commercial kits. METHODS: A total of 70 samples collected from children who suffered from acute gastroenteritis in 2009 - 2010 were tested for norovirus by three different immunochromatography kits. RT-PCR was employed as the gold standard method. RESULTS: The sensitivity of IP-NoV, QuickEx-Norovirus, and QuickNavi-Norovirus kits were 85.2%, 63.9%, and 55.7%, respectively. The IP-NoV kit could detect various genotypes of norovirus with higher efficiency as compared to QuickEx-Norovirus and QuickNavi-Norovirus kits. In addition, the phylogenetic analysis revealed that the majority of noroviruses circulating in Japan during 2009 - 2010 belonged to GII/4 variant 2006b and 2008a, which were responsible for the outbreaks worldwide. CONCLUSIONS: The findings indicated that the IC kits could be used as an alternative method for direct detection of norovirus in clinical specimens covering a wide range of norovirus genotypes circulating in Japan.

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 Children's 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.

Viral molecular characterization of the first congenital rubella syndrome case in Vietnam.

BACKGROUND: Rubella virus (RV) infection during the first trimester of pregnancy often leads to severe birth defects known as congenital rubella syndrome (CRS). METHODS: A premature newborn male was clinically diagnosed as CRS with cataracts, congenital heart defects, microcephaly, hepatosplenomegaly, and thrombocytopenia. The infection was confirmed serologically and molecularly. RESULTS: The RV was characterized and clustered with the 2B genotype. CONCLUSIONS: The integrated description of clinical features, serological and molecular data provide a baseline for diagnosis and control of rubella and CRS in Vietnam. This is the first report of molecular investigation of wildtype RV strain in Vietnam, thus contributing to the documentation of RV's worldwide distribution.

Comparison of the rapid methods for screening of group a rotavirus in stool samples.

In this study, the sensitivity and specificity of three immunochromatography (IC) test kits for rapid detection of group A rotavirus were compared and evaluated with stool samples collected from children, who suffered from acute gastroenteritis during February to June, 2009 in Japan. A total of 86 stool samples were tested and compared with a reference RT-PCR method. The sensitivity among IP-Rota V, Dipstick Eiken ROTA and ROTA-ADENO test kits were 97.2, 95.8 and 88.7%, while the specificity were 100, 93.3 and 100%, respectively. It was demonstrated that the IC kits evaluated in this study could be used as an alternative method for the rapid screening of group A rotavirus in fecal specimens, especially during acute gastroenteritis outbreak season.

Human parechovirus infection in children hospitalized with acute gastroenteritis in Sri Lanka.

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.