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Large-scale evolutionary surveillance of the 2009 H1N1 influenza A virus using resequencing arrays.

Lee CW, Koh CW, Chan YS, Aw PP, Loh KH, Han BL, Thien PL, Nai GY, Hibberd ML, Wong CW, Sung WK - Nucleic Acids Res. (2010)

Bottom Line: The accompanying base-calling software (EvolSTAR) introduces novel methods that utilize neighbourhood hybridization intensity profiles and substitution bias of probes on the microarray for mutation confirmation and recovery of ambiguous base queries.Our results demonstrate that EvolSTAR is highly accurate and has a much improved call rate.The high throughput and short turn-around time from sample to sequence and analysis results (30 h for 24 samples) makes this kit an efficient large-scale evolutionary biosurveillance tool.

View Article: PubMed Central - PubMed

Affiliation: Genome Institute of Singapore, Genome, 60 Biopolis Street, Singapore.

ABSTRACT
In April 2009, a new influenza A (H1N1 2009) virus emerged that rapidly spread around the world. While current variants of this virus have caused widespread disease, particularly in vulnerable groups, there remains the possibility that future variants may cause increased virulence, drug resistance or vaccine escape. Early detection of these virus variants may offer the chance for increased containment and potentially prevention of the virus spread. We have developed and field-tested a resequencing kit that is capable of interrogating all eight segments of the 2009 influenza A(H1N1) virus genome and its variants, with added focus on critical regions such as drug-binding sites, structural components and mutation hotspots. The accompanying base-calling software (EvolSTAR) introduces novel methods that utilize neighbourhood hybridization intensity profiles and substitution bias of probes on the microarray for mutation confirmation and recovery of ambiguous base queries. Our results demonstrate that EvolSTAR is highly accurate and has a much improved call rate. The high throughput and short turn-around time from sample to sequence and analysis results (30 h for 24 samples) makes this kit an efficient large-scale evolutionary biosurveillance tool.

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Related in: MedlinePlus

Observed neighbourhood hybridization intensity profiles for true-non-mutation calls. A representative set of observed NHIPs for true-non-mutation calls from patient sample 380. This representative set consists of five true-non-mutation calls randomly selected from each segment. Each line represents the NHIP (±6 bp from query base position) of a true-non-mutation call.
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Figure 3: Observed neighbourhood hybridization intensity profiles for true-non-mutation calls. A representative set of observed NHIPs for true-non-mutation calls from patient sample 380. This representative set consists of five true-non-mutation calls randomly selected from each segment. Each line represents the NHIP (±6 bp from query base position) of a true-non-mutation call.

Mentions: Figure 3 shows the NHIPs of a representative set of 40 randomly selected query bases that result in true-non-mutation calls (wild-type calls). We observed that in these NHIPs, the PM probe of the query base together with neighbouring PM probes, have hybridization intensities significantly higher (>1.4-fold) than that of their MM probes in general. We also identified 10 mutations using capillary sequencing in the patient sample. The NHIPs of these 10 true-mutation calls (Figure 4) are very different from NHIPs of wild-type calls. The presence of a mutation at the query base created a MM in neighbouring PM probes and caused a drop in their hybridization intensities. The closer this mutation is to the centre of a neighbouring PM probe, the bigger the drop in hybridization intensity. This results in a distinctive dip to the immediate left and right of the centre of the NHIP where the mutation is.Figure 3.


Large-scale evolutionary surveillance of the 2009 H1N1 influenza A virus using resequencing arrays.

Lee CW, Koh CW, Chan YS, Aw PP, Loh KH, Han BL, Thien PL, Nai GY, Hibberd ML, Wong CW, Sung WK - Nucleic Acids Res. (2010)

Observed neighbourhood hybridization intensity profiles for true-non-mutation calls. A representative set of observed NHIPs for true-non-mutation calls from patient sample 380. This representative set consists of five true-non-mutation calls randomly selected from each segment. Each line represents the NHIP (±6 bp from query base position) of a true-non-mutation call.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2874996&req=5

Figure 3: Observed neighbourhood hybridization intensity profiles for true-non-mutation calls. A representative set of observed NHIPs for true-non-mutation calls from patient sample 380. This representative set consists of five true-non-mutation calls randomly selected from each segment. Each line represents the NHIP (±6 bp from query base position) of a true-non-mutation call.
Mentions: Figure 3 shows the NHIPs of a representative set of 40 randomly selected query bases that result in true-non-mutation calls (wild-type calls). We observed that in these NHIPs, the PM probe of the query base together with neighbouring PM probes, have hybridization intensities significantly higher (>1.4-fold) than that of their MM probes in general. We also identified 10 mutations using capillary sequencing in the patient sample. The NHIPs of these 10 true-mutation calls (Figure 4) are very different from NHIPs of wild-type calls. The presence of a mutation at the query base created a MM in neighbouring PM probes and caused a drop in their hybridization intensities. The closer this mutation is to the centre of a neighbouring PM probe, the bigger the drop in hybridization intensity. This results in a distinctive dip to the immediate left and right of the centre of the NHIP where the mutation is.Figure 3.

Bottom Line: The accompanying base-calling software (EvolSTAR) introduces novel methods that utilize neighbourhood hybridization intensity profiles and substitution bias of probes on the microarray for mutation confirmation and recovery of ambiguous base queries.Our results demonstrate that EvolSTAR is highly accurate and has a much improved call rate.The high throughput and short turn-around time from sample to sequence and analysis results (30 h for 24 samples) makes this kit an efficient large-scale evolutionary biosurveillance tool.

View Article: PubMed Central - PubMed

Affiliation: Genome Institute of Singapore, Genome, 60 Biopolis Street, Singapore.

ABSTRACT
In April 2009, a new influenza A (H1N1 2009) virus emerged that rapidly spread around the world. While current variants of this virus have caused widespread disease, particularly in vulnerable groups, there remains the possibility that future variants may cause increased virulence, drug resistance or vaccine escape. Early detection of these virus variants may offer the chance for increased containment and potentially prevention of the virus spread. We have developed and field-tested a resequencing kit that is capable of interrogating all eight segments of the 2009 influenza A(H1N1) virus genome and its variants, with added focus on critical regions such as drug-binding sites, structural components and mutation hotspots. The accompanying base-calling software (EvolSTAR) introduces novel methods that utilize neighbourhood hybridization intensity profiles and substitution bias of probes on the microarray for mutation confirmation and recovery of ambiguous base queries. Our results demonstrate that EvolSTAR is highly accurate and has a much improved call rate. The high throughput and short turn-around time from sample to sequence and analysis results (30 h for 24 samples) makes this kit an efficient large-scale evolutionary biosurveillance tool.

Show MeSH
Related in: MedlinePlus