This study shows that, using the described swabs, material obtained from the nasopharynx contains more influenza virus than that recovered from the oropharynx. The combined calculated diagnostic sensitivity for influenza B and influenza A(H3N2)pdm09 for NP and OP samples were 78 % and 63 %, respectively (χ2 < 0.01).
Our finding that the NP samples contain more virus than the OP samples are in contrast to a prospective study from Kenya, where influenza A(H1N1) were found significantly more often in OP samples and influenza B virus more often in NP samples, whereas with influenza A (H3N2), no significant difference between the two sampling sites were observed. However, the Kenyan study compared a flocked swab used in the nasopharynx with a polyester swab used in the oropharynx. As flocked swabs contain, on average, 4.8 times more virus than rayon swabs, a comparison based on different swab types is difficult to interpret . Lieberman et al. performed NP and OP sampling by rigid cotton swabs as well as by NP washes to detect eight different respiratory virus types . NP washes were found to be superior to NP swabbing, and OP swabbing was the least sensitive of the three sampling methods. As NP washes are difficult to obtain, NP swabbing samples appear to be the most appropriate method, especially in frail individuals .
In this selected material of initially PCR-positive individuals, our sampling method reached a sensitivity of 78 % for all observed influenza virus and 71 % for influenza A(H1N1)pdm09 alone. As the initial viral material was collected, on average, 1.5 days before the collection of swabs for the current study, natural viral clearance in some patients would be expected to reduce the sensitivity of the test. On the other hand, the use of absorptive swabs and specially trained technicians probably contributed to the high sensitivity of 78 % for all influenza viruses and 71 % for the H1N1 strain.
In a retrospective cohort study of 25 patients hospitalized for pandemic influenza with community-acquired pneumonia, a comparison between lower respiratory tract specimens and NP swabs revealed a sensitivity of only 63 % for the NP swabs and nearly 98 % for the lower respiratory samples . A lower respiratory tract infection could possibly delay viral clearance in the lungs, leading to a prolonged viral excretion in the lower airways. Therefore, specimens from the bronchial tree could be more suitable than NP samples when investigating patients with pneumonia. However, at least in patients without lower respiratory tract affection, NP swabs represent a low-cost, easily accessible, and quick diagnostic method, and provided a proper swabbing technique.
The viral load is reduced in a nonlinear fashion and different types of influenza virus seem to follow different time lines for viral clearance [3–5, 8, 9, 13]. Samples from influenza A(H1N1)pdm09 patients display a lower viral load than patients suffering from seasonal influenza A (H1N1 and H3N2) . The majority of studies on viral load and clearance have been performed on influenza A subgroups. Although less is known about influenza B in this regard, one study reported continuous viral shedding in 70 % of influenza B patients compared to 33 % among those suffering from influenza A disease after 7 days of illness . We did not find a significant difference in viral load when comparing samples from patients with influenza B and influenza A(H1N1)pdm09. Among our patients, more of the initially influenza A(H1N1)pdm09-positive than influenza B-positive individuals had turned PCR-negative on average 4.8 days after onset of the disease. However, the small numbers involved do not allow any general conclusions be made. The time of sampling, seriousness of the infection, comorbidity, lower respiratory tract infection, sampling method, and vaccination status may all influence the viral load at the sampling site [4, 8, 9, 14]. An assessment of the relative impact on the viral load of such factors would require careful examination of a large group of comparable patients.
One might speculate that the H1N1 dominance of the current study is explained by sampling bias towards detecting influenza A(H1N1)pdm09 rather than influenza A(H3N2).
However, recent subtyping of influenza strains in a comparable study  conducted during the influenza season 2008–2009 provided additional H3N2 isolates (Olav Hungnes, Norwegian Institute of Public Health, personal communication). The NP samples for these patients were positive at a lower CT value than the OP samples, with a mean difference in CT value of 10.9 (95 % CI 7.1–14.7, p < 0.001), corresponding to a mean difference in the calculated viral load of 1,911 times (95 % CI 137.2–26,615.9) higher in the nasopharynx than the oropharynx. H1N1 and H3N2 shared the higher affinity for the nasopharynx compared to the oropharynx. This finding makes it unlikely that the H1N1 dominance of the current study is explained by methodological issues.
A strength of our study is the prospective design and the low number of specially trained staff responsible for the swabbing procedure, reducing the possibility of suboptimal techniques. A limitation is that the sensitivity of the diagnostic procedure is evaluated, on average, 1.5 days after admission, and not at the time of hospitalization. The genetic variation in influenza A(H1N1)pdm09 is still limited, with most isolates genetically similar to influenza A/California/07/2009 (H1N1). Therefore, future studies need to be performed when influenza A(H1N1)pdm09 has been thoroughly established as a seasonal pathogen.
To our knowledge, this is the first study comparing the load of influenza viruses in swabbed samples from the oropharynx and nasopharynx of adult patients. We show that, in influenza patients, the etiological diagnosis by PCR is achieved more efficiently by NP than by OP swabbing.