Epidemic of influenza A(H1N1)pdm09 analyzed by full genome sequences and the first case of oseltamivir-resistant strain in Myanmar 2017

Abstract

A community outbreak of human influenza A(H1N1)pdm09 virus strains was observed in Myanmar in 2017. We investigated the circulation patterns, antigenicity, and drug resistance of 2017 influenza A(H1N1)pdm09 viruses from Myanmar and characterized the full genome of influenza virus strains in Myanmar from in-patients and out-patients to assess the pathogenicity of the viruses. Nasopharyngeal swabs were collected from out-patients and in-patients with acute respiratory tract infections in Yangon and Pyinmana City in Myanmar during January-December 2017. A total of 215 out-patients and 18 in-patients infected with A(H1N1)pdm09 were detected by virus isolation and real-time RT-PCR. Among the positive patients, 90.6% were less than 14 years old. Hemagglutination inhibition (HI) antibody titers against A(H1N1)pdm09 viruses in Myanmar were similar to the recommended Japanese influenza vaccine strain for 2017-2018 seasons (A/Singapore/GP1908/2015) and WHO recommended 2017 southern hemisphere vaccine component (A/Michigan/45/2015). Phylogenetic analysis of the hemagglutinin sequence showed that the Myanmar strains belonged to the genetic subclade 6B.1, possessing mutations of S162N and S164T at potential antigenic sites. However, the amino acid mutation at position 222, which may enhance the severity of disease and mortality, was not found. One case with no prior history of oseltamivir treatment possessed H275Y mutated virus in neuraminidase (NA), which confers resistance to oseltamivir and peramivir with elevated IC50 values. The full genome sequence of Myanmar strains showed no difference between samples from in-patients and out-patients, suggesting no additional viral mutations associated with patient severity. Several amino acid changes were observed in PB2, PB1, and M2 of Myanmar strains when compared to the vaccine strain and other Asian strains. However, no mutations associated with pathogenicity were found in the Myanmar strains, suggesting that viral factors cannot explain the underlying reasons of the massive outbreak in Myanmar. This study reported the first detection of an oseltamivir-resistant influenza virus in Myanmar, highlighting the importance of continuous antiviral monitoring and genetic characterization of the influenza virus in Myanmar.

Fig 1. Monthly distribution of influenza virus by region in Myanmar during the 2017 influenza season and the location of sample collection sites.

The total number of collected samples is shown in the black dashed line. Influenza A(H1N1) pdm09 is shown as red bar; A(H3N2), blue bar; B/Victoria, purple bar and B/Yamagata, green bar. The seasonal peak of A(H1N1)pdm09 was observed in June-July, during the rainy season, and influenza A(H1N1)pdm09 was the predominant subtype in Myanmar during the 2017 influenza season. The map of Myanmar and the green circles represent the two study sites, Yangon and Pyinmana. This map is generated by the software ArcMap 10.0 (ESRI Japan Corporation, Tokyo, Japan). Boundaries are generated by using a world basemap supplied by ESRI Japan Corporation.

Fig 2. HA and NA phylogeny of influenza A(H1N1) 2009 pandemic isolates in Myanmar in 2017.

Trees were constructed by the maximum likelihood method using MEGA software (version 6.06). Bootstrap value was determined for 1000 iterations; only values greater than 70% are shown. Myanmar out-patient is shown in red fonts, and in-patient is shown in dark blue fonts. The HA and NA sequences of Myanmar strains belonged to clade 6B.1. Red circle (●) represents oseltamivir- and peramivir-resistant strain exhibiting NA H275Y substitution. Amino acid changes were based on A/California/07/2009.