Prospective surveillance and molecular characterization of seasonal influenza in a university cohort in Singapore.

BACKGROUND: Southeast Asia is believed to be a potential locus for the emergence of novel influenza strains, and therefore accurate sentinel surveillance in the region is critical. Limited information exists on sentinel surveillance of influenza-like illness (ILI) in young adults in Singapore in a University campus setting. The objective of the present study was to determine the proportion of ILI caused by influenza A and B viruses in a university cohort in Singapore. METHODOLOGY/PRINCIPAL FINDINGS: We conducted a prospective surveillance study from May through October 2007, at the National University of Singapore (NUS). Basic demographic information and nasopharyngeal swabs were collected from students and staff with ILI. Reverse-transcriptase PCR (RT-PCR) and viral isolation were employed to detect influenza viruses. Sequencing of hemagglutinin (HA) and neuraminidase (NA) genes of some representative isolates was also performed. Overall proportions of influenza A and B virus infections were 47/266 (18%) and 9/266 (3%) respectively. The predominant subtype was A/H3N2 (55%) and the rest were A/H1N1 (45%). The overall sensitivity difference for detection of influenza A viruses using RT-PCR and viral isolation was 53%. Phylogenetic analyses of HA and NA gene sequences of Singapore strains showed identities higher than 98% within both the genes. The strains were more similar to strains included in the WHO vaccine recommendation for the following year (2008). Genetic markers of oseltamivir resistance were not detected in any of the sequenced Singapore isolates. CONCLUSIONS/SIGNIFICANCE: HA and NA gene sequences of Singapore strains were similar to vaccine strains for the upcoming influenza season. No drug resistance was found. Sentinel surveillance on university campuses should make use of molecular methods to better detect emerging and re-emerging influenza viral threats

Surveillance and clinical characterization of influenza in a university cohort in Singapore.

Southeast Asia is a potential locus for the emergence of novel influenza strains. However, information on influenza within the region is limited.This study was to determine the proportion of influenza-like illness (ILI) caused by influenza A and B viruses in a university cohort in Singapore, identify important distinctive clinical features of influenza infection and potential factors associated with influenza infection compared with other causes of ILI.A surveillance study was conducted from 2007 to 2009, at the University Health and Wellness Centre, National University of Singapore (NUS). Basic demographic information and nasopharyngeal swabs were collected from consenting students and staff with ILI, with Influenza A and B identified by both culture and molecular methods.Proportions of influenza A and B virus infections in subjects with ILI were 153/500 (30.6%) and 11/500 (2.2%) respectively. The predominant subtype was A/H1N1, including both the seasonal strain (20/153) and the pandemic strain (72/153). The clinical symptom of fever was more common in subjects with laboratory confirmed influenza than other ILIs. On-campus hostel residence and being a student (compared with staff) were associated with increased risk of laboratory confirmed influenza A/H1N1 2009 infection.This study provides a baseline prevalence of influenza infection within young adults in Singapore in a university setting. Potential risk factors, such as hostel residence, were identified, allowing for more targeted infection control measures in the event of a future influenza pandemic

Number (%) of samples positive for influenza A virus infection by RT-PCR and viral isolation (shown in bold).

1<p><b>Present</b> means sample was positive for influenza A infection by either or both the methods.</p>2<p><b>Absent</b> means sample was negative for influenza A infection by both methods.</p

Primer sets for sequencing HA and NA genes.

<p>*R =  A/G, Y =  C/T, M =  A/C, N =  A/C/G/T.</p

Influenza A Neuraminidase (NA) protein sequence Analysis for Oseltamivir Resistance.

<p>(A) The consensus sequence of neuraminidase (N1) gene from residues 251 to 300. Substitution of amino acid Histidine (H) to Tyrosine (Y) at position 275 in N1 gene (shown in bold) and at position 274 in N2 gene confers resistance to oseltamivir. Protein sequences of (B) N1 and (C) N2 genes of Singapore isolates.</p

Monthly distribution of total number of samples and influenza A positive samples detected by RT-PCR and viral isolation during the study period.

<p>Monthly distribution of total number of samples and influenza A positive samples detected by RT-PCR and viral isolation during the study period.</p

Phylogenetic trees of Seasonal influenza viruses circulating in a university cohort in Singapore, 2007.

<p>The phylogenetic trees of (A) 6 HA and (B) 6 (NA) genes of A/H3N2 viruses; (C) 5 HA and (D) 5 NA genes of A/H1N1 viruses with WHO vaccine strains and other 2007 sequences from GenBank constructed using neighbor-joining method. Bootstrap values 90 and over are shown. Singapore isolates are in bold and vaccine strains in rectangles.</p

Number (%) of Subjects positive for Influenza Virus Infection.

<p>Number (%) of Subjects positive for Influenza Virus Infection.</p

Symptom distribution in subjects and prevalence rate ratios with predictive values.

<p>OR: Odds Ratio, PPV: Positive Predictive Value, NPV: Negative Predictive Value</p><p>Symptom distribution in subjects and prevalence rate ratios with predictive values.</p