There were 2,220 specimens processed by the Diagnostic Virology Laboratory for respiratory viral culture during the study period. A total of 479 specimens were tested by the neuraminidase assay performed by a single technician (B.C.) during daylight, weekday hours, and constitute the study samples. Figure shows results from viral cultures for all specimens processed during the study period. There were 130 specimens that had the neuraminidase assay performed by other technicians and are not included in the study sample.
Results of respiratory viral cultures performed at the Diagnostic Virology Laboratory during the study period (1 January 1999 to 20 March 1999). The numbers are the numbers of specimens in the different categories.
The 479 study samples were obtained from patients with a mean age of 3.8 years (range, 7 days to 27 years). Of the 479 patients, 293 (61.1%) were male and 186 (38.8%) were female. A total of 423 (88.3%) of these samples were also processed for IFA and 417 (87%) had an EIA for influenza A detection performed. All three rapid tests were performed on 374 (78%) specimens. Of the 479 study specimens, 201 (41.9%) had a virus isolated (102 influenza virus type A, 22 influenza virus type B, 32 rhinovirus, 17 parainfluenza virus, 17 respiratory syncytial virus, 6 adenovirus, 7 cytomegalovirus, 2 enterovirus, and 1 herpes simplex virus; in 5 specimens, a dual infection was detected).
Table shows the performance of all three rapid detection assays compared to viral isolation by culture. There were 114 samples positive by the neuraminidase detection assay; the sensitivity was 70.1%, and the specificity was 92.4%. The sensitivity for detection of influenza A virus alone was 76.4%, and for influenza B virus alone, it was 40.9%. When tests categorized only as strong positive were analyzed as positive, the specificity for detection of influenza A and B virus was higher (95.8%), with a lower sensitivity (64.2%). When performance of 130 additional tests carried out by other technicians (not B.C.) less experienced with the kit was analyzed, lower sensitivity (45.7%) and specificity (76.2%) for detection of all influenza viruses compared to culture were observed.
TABLE 1 Performance of rapid diagnostic tests for the detection of influenza virus infection in nasal wash samples compared to viralculture
Of the 479 specimens, 423 also were tested by IFA. Microscopic analysis showed that 102 (24.1%) samples were inadequate for IFA (<20 cells), while 60 had sparse cellularity, and 261 had abundant cells. Of the 321 specimens adequate for IFA, there were 48 (14.9%) positive for influenza A virus and 11 (3.4%) positive for influenza B virus. The sensitivity and specificity of IFA compared to viral culture for detection of any influenza virus was 59.8 and 97%, respectively. Influenza A EIA was performed in 417 of the 479 specimens. There were 93 (22.3%) positive and 324 (77.6%) negative test results, providing a sensitivity of 89.7% and specificity of 98.1% for detection of influenza A virus by EIA.
There was a decreased ability to recover viruses by culture in specimens that were considered to have sparse or inadequate cellularity during IFA compared to those with abundant cellularity (Table ). However, when isolation of influenza viruses alone was compared between the groups, the difference did not attain statistical significance. There was no difference in the mean time for a culture to become positive between the three different IFA cellularity groups (mean times of 4.5, 4, and 4.2 days). The sensitivity for detection of influenza A or B viruses in specimens with abundant cellularity was 77% and the specificity of 89.4%, compared to sensitivity of 57.1% and specificity of 96.8% for the combination of the groups with sparse and inadequate cellularity. Sample cellularity also adversely influenced the performance of the IFA and EIA rapid tests.
TABLE 2 Effect of quality of nasal wash specimen on the ability to isolate viruses and detect influenza virus by rapid diagnosis or byculturea
There were 27 samples for which there was a positive neuraminidase test result, but no virus was recovered (Table ). In six of the specimens, influenza virus RNA was detected by RT-PCR (influenza A virus detected in five and influenza B virus detected in one). In addition, 5 of the 27 samples were positive by EIA and/or IFA, providing further corroboration that the neuraminidase assay results represented true-positive results in these samples.
TABLE 3 Results of additional tests for 27 nasal wash specimens that tested positive by neuraminidase detection assay but tested negative for influenza virus byculture
When the performance of the neuraminidase assay was analyzed again, considering all positive results obtained from any additional diagnostic tests as an influenza infection, the specificity of the neuraminidase assay increased to 94.2%, with a positive predictive value of 82.5%, while the sensitivity and negative predictive value were 68.6 and 88.2%, respectively.
There were 37 samples negative by the neuraminidase assay from which influenza virus was grown (24 samples for influenza A virus and 13 for influenza B virus). Of the 37 samples, 36 were tested by EIA and 14 were positive. A total of 24 samples were tested by IFA; 18 had abundant cellularity, while 6 and 9 had sparse and inadequate cellularity, respectively. In six samples, the IFA result was positive. Influenza virus strains isolated from these 37 samples (false-negative result by the neuraminidase assay) were tested directly with the neuraminidase assay to confirm the affinity of the substrate for the neuraminidase enzyme produced by each strain, and all strains tested positive.