sites

The sequence sites selected to display amino acid substitutions on the phylogenetic tree of H3 influenza viruses

NAG_inHA1

The predicted number of N-linked glycosylation sites in the hemagglutinin proteins of H3 influenza viruses

MCC_FluA_H3

Time-scaled phylogenetic tree of H3 influenza viruses inferred by BEAST using molecular clock model

rst

Output file of CODEML inferring ancestral sequences of H3 influenza viruses using codon model

mlc

Output file of CODEML estimating selection pressure using codon model

Genotype

Genotype table of the H3 influenza viruses

Maximum clade credibility (MCC) tree reconstruction of Phuket/3073-like subclade.

<p>(A) HA gene and (B) NA gene. The 95% highest posterior density (HPD) for the ancestral node is indicated. Timescale is shown at the bottom of the tree. MCC tree with complete taxa identity is shown in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0136254#pone.0136254.s005" target="_blank">S5</a> and <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0136254#pone.0136254.s006" target="_blank">S6</a> Figs.</p

Evolutionary rate and age of influenza B Phuket/3073-like subclade.

<p>tMRCA: time of the most recent common ancestor; HPD: highest posterior density</p><p>Evolutionary rate and age of influenza B Phuket/3073-like subclade.</p

Epidemiological and Evolutionary Dynamics of Influenza B Viruses in Malaysia, 2012-2014

<div><p>Epidemiological and evolutionary dynamics of influenza B Victoria and Yamagata lineages remained poorly understood in the tropical Southeast Asia region, despite causing seasonal outbreaks worldwide. From 2012–2014, nasopharyngeal swab samples collected from outpatients experiencing acute upper respiratory tract infection symptoms in Kuala Lumpur, Malaysia, were screened for influenza viruses using a multiplex RT-PCR assay. Among 2,010/3,935 (51.1%) patients infected with at least one respiratory virus, 287 (14.3%) and 183 (9.1%) samples were tested positive for influenza A and B viruses, respectively. Influenza-positive cases correlate significantly with meteorological factors—total amount of rainfall, relative humidity, number of rain days, ground temperature and particulate matter (PM10). Phylogenetic reconstruction of haemagglutinin (HA) gene from 168 influenza B viruses grouped them into Yamagata Clade 3 (65, 38.7%), Yamagata Clade 2 (48, 28.6%) and Victoria Clade 1 (55, 32.7%). With neuraminidase (NA) phylogeny, 30 intra-clade (29 within Yamagata Clade 3, 1 within Victoria Clade 1) and 1 inter-clade (Yamagata Clade 2-HA/Yamagata Clade 3-NA) reassortants were identified. Study of virus temporal dynamics revealed a lineage shift from Victoria to Yamagata (2012–2013), and a clade shift from Yamagata Clade 2 to Clade 3 (2013–2014). Yamagata Clade 3 predominating in 2014 consisted of intra-clade reassortants that were closely related to a recent WHO vaccine candidate strain (B/Phuket/3073/2013), with the reassortment event occurred approximately 2 years ago based on Bayesian molecular clock estimation. Malaysian Victoria Clade 1 viruses carried H274Y substitution in the active site of neuraminidase, which confers resistance to oseltamivir. Statistical analyses on clinical and demographic data showed Yamagata-infected patients were older and more likely to experience headache while Victoria-infected patients were more likely to experience nasal congestion and sore throat. This study describes the evolution of influenza B viruses in Malaysia and highlights the importance of continuous surveillance for better vaccination policy in this region.</p></div

Phylogenetic analysis of the HA gene of influenza B viruses in Malaysia from 2012 to 2014.

<p>HA sequences of Malaysian influenza B viruses were compared with WHO recommended candidate vaccine and reference strains. The phylogeny was reconstructed using maximum likelihood (ML) method. Bootstrap values (>60%) and amino acid substitutions are mapped to key branches. Intra-and inter-clade reassortants are indicated as boxes. Yamagata Clade 3 (Yam-3) (blue), Yamagata Clade 2 (Yam-2) (orange) and Victoria Clade 1 (Vic-1) (green) are indicated. Scale bar represents a genetic distance of 0.008 substitutions/site. Phylogenetic tree for HA gene with complete taxa identity is shown in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0136254#pone.0136254.s001" target="_blank">S1 Fig</a>.</p