Specimen collection and laboratory analysis:
Trained staff collected nasopharyngeal and oropharyngeal swabs at
recruitment within 72 hours of date and time of AMI onset. Specimen
collection within the aforementioned timeframe was conducted in order to
maximize the sensitivity of the study to detect influenza RNA in the
swabs prior to the cessation of viral shedding. The specimens were
transported daily in Viral Transport Medium (VTM) at 2-
8o C to virology laboratory of International Centre
for Diarrheal Disease Research, Bangladesh (icddr,b), in Dhaka. The
specimens were aliquoted in a BSL-2 (bio-safety level-2) safety cabinet
and were stored in freezers at or below -70 o C until analysis. Viral
RNA was extracted from the swab specimen, and Real time Reverse
Transcription Polymerase Chain Reaction (qRT-PCR) was performed using
primers and probes specific for influenza A and B viruses provided by
Influenza Division at CDC. Hemagglutinin subtyping of type A and B
viruses was performed to detect subtypes, A/H3, A/H1N1pdm09, B/Victoria,
B/Yamagata. The laboratory used validated protocols
(31) and followed standard quality
control procedures.