This is the reference influenza detection method and has the highest sensitivity and specificity [34, 35] . Several modalities of RT-PCR have been designed: conventional gel-based PCR (cRT-PCR), multiplex PCR (mRT-PCR), and real-time RT-PCR (rRT-PCR). They can differentiate between influenza types (A or B) and subtypes (including pandemic H1N1 influenza and avian H5N1 influenza), and results are available in 2-6 h (although due to transportation of batched specimens to reference centers for processing, it may take longer for results to be available). During the 2009 H1N1 influenza A pandemic, it became clear that rapid case identification was essential for timely management of patients and for adequate public health actions to be taken. To answer to this threat, the CDC optimized the previously developed rRT-PCR procedures for detection of the A/H1N1 2009 pandemic influenza virus [36• ]. Yang et al. compared the performance of 12 rRT-PCR primer-probe sets designed for detecting the hemagglutinin (HA) or the neuraminidase (NA) gene of the pandemic influenza A/H1N1 2009 virus, using the primer-probe set developed at the CDC as reference. They found that although all primer-probe sets had specificity levels as high as 98.4 % to 100 %, some of the primer-probe sets had better specificity, sensitivity, and amplification efficiency than others, and that a combination of primer-probe sets targeted to the HA and NA genes had higher detection sensitivity than those targeting HA or NA individually [37] . In another study, Lam et al. showed that although rRT-PCR assays can be 10-fold more sensitive than cRT-PCR, newly developed cRT-PCR assays targeting the HA gene are a reliable alternative for laboratories where a rRT-PCR machine is not available [38] . Sensitivity and specificity of these assays also depend on the type of respiratory sample employed. For example, in a Spanish study that assessed the utility of rRT-PCR for the diagnosis of the novel influenza A/H1N1 virus, the authors reported that the diagnostic yield of combined nose and throat swabs was higher than that of nasopharyngeal aspirates [39] .