Effective control of any outbreak of a novel influenza strain will rely on the ability of available testing modalities to detect and, critically, to rule-out infection with the novel strain. During the recent epidemic of novel H1N1 disease in Boston, Massachusetts, we found that we were able to effectively rule out novel H1N1 infection in symptomatic adults using DFA testing--a rapid, relatively low-cost, and commercially-available method. Even prior to optimization of criteria for specimen adequacy (below), the NPV of our DFA tests was 96%.
Our specimens were predominantly nasopharyngeal swab specimens collected using flocked swabs (which contain brush-like nylon fibers to improve cell collection), and this collection method produced excellent yield of columnar epithelial cells. Any discrepant results (DFA-negative/PCR-positive) that we did see were explained by one or more of the following factors: 1) borderline insufficient numbers of respiratory columnar epithelial cells in the specimen, or 2) misinterpretation of neutrophils as columnar epithelial cells and therefore incorrect assessment of specimens as having adequate numbers of cells.
In response to these findings, we have now increased our laboratory's cell threshold (for considering a sample adequate for DFA testing) from ≥30 to ≥60 columnar epithelial cells per test well. Past laboratory data (internal data from 2007–8 influenza season, not shown) indicated that use of this more stringent criterion for specimen adequacy would not significantly increase the frequency of specimen rejections. Furthermore, we have emphasized with our technologists the need to reliably distinguish respiratory columnar epithelial cells (the cell type infected by influenza virus) from neutrophils and squamous epithelial cells in assessing specimen adequacy.
We expect that with these changes we will have very few false negative DFA results going forward. Importantly, immunofluorescent staining (e.g., DFA testing) is a standard method for identifying viruses in clinical virology laboratories. Therefore, clinical virology laboratories should be readily able to apply this prior experience to implementation of highly sensitive DFA testing for novel H1N1. It should be noted that our study examined test performance primarily for adult inpatients and healthcare workers (outpatients) who met CDC criteria for influenza-like illness. Therefore, our results may not similarly apply to other patient populations, e.g., pediatric patients or other specific outpatient groups, in whom the performance characteristics of the DFA test may differ. Furthermore, our NPV is specific to the disease prevalence associated with this period of the H1N1 pandemic; if the prevalence of H1N1 in adults increases in the future, the NPV of the DFA tests may decrease to some degree. Nevertheless, our results demonstrate that DFA testing can be used to effectively exclude infection with the novel H1N1 influenza strain in a hospital setting, allowing resources and infection control efforts to be focused on those who test positive.