Prevalence and molacular genetic analysis of human respiratory syncytial virus and enterovirus 68 among children with acute lower respiratory tract infection in Thailand

Respiratory syncytial virus (RSV) causes acute lower respiratory tract infection in infants and young children worldwide. A clear description of local RSV molecular epidemiology, evolution, and transmission requires detailed sequence data and can inform new strategies for virus control and vaccine development. To investigate the RSV burden in Thailand over six consecutive years (January 2012 to December 2017). There were two different methods to detect RSV in the present study. First, we screened 3306 samples obtained from children ≤5 years old with acute respiratory tract infection using semi-nested reverse-transcription polymerase chain reaction (RT-PCR) during January 2012 to December 2015. Second, we screened 8842 samples using real-time RT-PCR to determine the burden of RSV, including hMPV infections in patients in all age groups, who presented with influenza-like illnesses in Bangkok, Thailand, during January 2016 to December 2017. In 2012-2015, 8.4% (277/3,306) of the specimens tested positive for RSV, and then genotyped RSV by sequencing the G glycoprotein gene and performed phylogenetic analysis to determine the RSV antigenic subgroup. The result revealed that NA1, ON1 and BA (BA9, BA10 and BA-C) were the circulating RSV genotypes. Of 8842 specimens to testing between 2016 and 2017, RSV was detected in 1011 (11.4%) specimens and hMPV was detected in 318 (3.6%) specimens. The most commonly RSV and hMPV infections were observed in young children, but both virus infections can sporadically occur in adult groups. For RSV and hMPV strains, ON1 and BA were the circulating RSV genotypes, while A2, B1, and B2 were the circulating hMPV genotypes. The current work described RSV genome evolution and transmission, we have generated 10 complete or nearly complete genomes of ON1 in Thailand, over a 7-year period using a RT-PCR. Comparison of local versus global strains demonstrated that most ON1 variants observed locally in Thailand were also seen in other parts of the world. The nucleotide substitution rates for the individual open reading frames (ORFs) were highest in the regions encoding the attachment (G) glycoprotein and the NS2 protein. The analysis of RSV full genomes, compared to subgenomic regions, provided more precise estimates of the RSV sequence changes and revealed important patterns of RSV genomic variation and global movement. The data presented expand our understanding of the epidemiology of RSV infection in Thailand. Moreover, the new RSV genomic sequences reported here expand our knowledge base for the further study on transmission of virus at the local community level. The outcome of these studies can support new strategies for RSV control and vaccine use and development. Enterovirus D68 (EV-D68) is associated with severe lower respiratory tract infection and neurological abnormalities including acute myelitis and cranial nerve dysfunction. To determine whether an increased incidence of EV-D68 occurs in Southeast Asia, we retrospectively tested specimens collected from Thai pediatric patients who were less than 5 years of age and presented with acute respiratory tract infections between 2012 and 2014. RT-PCR and nucleotide sequencing of the 5'-UTR/VP2 region were used to identify EV-D68. We also examined the epidemiological pattern of EV-D68 since 2009, when it was first identified in Thailand, and compiled records of clinical manifestations in children with confirmed EV-D68 infection. From 837 samples, 5 samples (0.6%) tested positive for EV-D68. All patients presented with viral pneumonia and required hospitalization. Phylogenetic analysis of the VP4/VP2 regions revealed that EV-D68 strains circulating in Thailand between 2012 and 2014 were closely related to strains reported in Japan, United Kingdom, China, and France. Continued surveillance of probable EV-D68-associated severe respiratory tract infection and the development of a rapid diagnostic test for EV-D68 are essential in supporting awareness and facilitating disease prevention and control.