Genetic diversity of hemagglutinin gene of A(H1N1)pdm09 influenza strains isolated in Taiwan and its potential impact on HA-neutralizing epitope interaction

Figures & data

Table 1. Sample information and clinical symptoms of flu infection among A(H1N1)pdm09 Taiwan patients

Sample number	Age	Sex	Date of isolation	Geographical location of isolation	Clinical symptoms
2009–02826	5	Male	2009–07–27	Central Taiwan	cough, fever (>38 °C), pneumonia
2009–00940	70	Male	2009–09–10	Northern Taiwan	cough, fever (>38 °C)
2009–08542	45	Male	2009–09–28	Eastern Taiwan	cough, sore throat, dyspnea, pneumonia
2010–03994	4	Male	2010–08–11	Southern Taiwan	cough, fever (>38 °C), dyspnea, pneumonia, respiratory failure
2010–00842	63	Male	2010–12–20	Northern Taiwna	dyspnea, sore throat, fever, pneumonia
2010–06031	42	Female	2010–08–18	Northern Taiwan	unavailable
2011–02054	60	Male	2011–01–20	Central Taiwan	myalgia, cough, dyspnea, pneumonia, sore throat, fever (>38 °C)
2011–00623	56	Female	2011–02–09	Northern Taiwan	cough, fever (>38 °C), dyspnea
2011–01219	53	Male	2011–02–10	Northern Taiwan	cough, fever (>38 °C), pneumonia
2009–06078	10	Male	2009–08–27	Central Taiwan	cough, fever (>38 °C), pneumonia
2009–04909	58	Female	2009–09–28	Eastern Taiwan	myalgia, dyspnea, cough, sore throat
2009–00937	83	Male	2009–09–09	Northern Taiwan	dyspnea, cough, fever (>38 °C), pneumonia
2009–08575	22	Female	2009–09–20	Eastern Taiwan	myalgia, dyspnea, cough, sore throat, pneumonia
2010–01164	15	Female	2010–05–05	Northern Taiwan	unavailable
2010–05270	37	Male	2010–09–06	Northern Taiwan	cough, sore throat, fever (>38 °C), dyspnea, pneumonia
2010–05347	56	Male	2010–08–12	Northern Taiwan	cough, fever (>38 °C), pneumonia
2011–04512	38	Female	2011–01–17	Eastern Taiwan	myalgia, dyspnea, cough, sore throat, fever (>38 °C)
2011–02068	4	Male	2011–01–24	Central Taiwan	cough, fever (>38 °C), vomiting
2011–04611	55	Male	2011–02–08	Northern Taiwan	dyspnea, cough

Figure 1. New genetic variants among A(H1N1)pdm09 isolates collected at Taiwan between 2009–2011 detected by MSSCP genotyping. RT-PCR products of hemagglutinin gene obtained from pandemic Taiwan A(H1N1)pdm09 virus isolates, as well as reference seasonal (s) and pandemic (p) strains of influenza virus A(H1N1)pdm09 were denatured and ssDNA’s were separated on a 9% polyacrylamide gel using MSSCP method under optimum electrophoretic conditions. DNA bands were visualized with silver stain. Strains are indicated as follows: s – reference seasonal strain, p – reference pandemic strains. Taiwan isolates are described with symbols listed in Table 1. Note the presence of five distinct MSSCP electrophoretic profiles (arrows at the bottom of the figure) (samples number: 2010–03994, 2011–01219, 2010–01164, 2010–05270, 2010–05347) among samples, which based on RT-PCR assay, were classified as A(H1N1)pdm09 strain. Dividing lines indicate grouping of images from different parts of the same gel.

Table 2. Genetic diversity of HA gene fragment in Taiwan A(H1N1)pdm09 isolates

Figure 2. Schematic representation of genetic diversity of hemagglutinin (HA) sequence in five Taiwan isolates of A(H1N1)v pan09 strain. Black arrows above and below A(H1N1)v pdm09 reference sequence indicate modified DNA codons. Red letters show nucleotide changes. Altered amino acids in HA protein sequence are also shown. Blue star over the description indicates amino acids localized in the epitope E of HA. Additionally, red arrows underline three point mutations, which are translated to amino acids in the epitope E of HA.

Figure 3. Structure of HA-Ab complex. Hemagglutinin protein monomer (blue color) interacting which neutralizing antibody (red color). Amino acid 66, localized on the HA-Ab interface is marked in green.

Figure 4. Phylogenetic tree of nucleotide sequences of hemagglutinin gene of Taiwan isolates of influenza A(H1N1)pdm09 virus. All sequences included in phylogenetic analysis were 935 nucleotides long. All Taiwan isolates deposited from 2009 through 2011 were retrieved from the EpiFlu Database. Each isolate (EPI210250 HA, EPI382332 HA, EPI382342 HA, EPI382415 HA, EPI190807 HA, EPI382397 HA, EPI382345 HA, EPI382352 HA, EPI382407 HA, EPI382412 HA, EPI382365 HA, EPI382375 HA) represents a number of identical sequences used in alignment. Isolates studied in this paper are described in Table 2. Numbers on branches indicate number of nucleotide substitutions per site.