Detection of respiratory syncytial virus in clinical specimens by viral culture, direct and indirect immunofluorescence, and enzyme immunoassay.

We evaluated prospectively the detection of respiratory syncytial virus (RSV) by culture and by direct antigen detection using an indirect immunofluorescence assay (IFA), a direct monoclonal immunofluorescence assay (DFA), and a monoclonal enzyme immunoassay (EIA). Of 221 specimens, 95 (43%) were culture positive for RSV, 4 (1.8%) contained more than one virus, and 17 (7.6%) contained a virus other than RSV. Overall, HEp-2 and Flow 6000 cells grew significantly more RSV isolates (82 and 72%, respectively) than A549 cells, which grew only 29% of the isolates. The mean time for RSV detection with HEp-2 cells was 2.9 days. This was significantly less than the mean time for RSV detection with either Flow 6000 cells (6.1 days) or A549 cells (6.4 days). Of 221 specimens, 129 were tested simultaneously by culture, IFA, and DFA. Of these 129 specimens, 62 (48%) were positive by culture, 69 (53%) were positive by IFA, and 70 (54%) were positive by DFA. For 92 specimens screened simultaneously by culture, IFA, and EIA, positive results were obtained for 33 (36%) of the specimens by both culture and IFA and for 29 (32%) of the specimens by EIA. Of 126 culture-negative specimens, 21 (17%) were positive for RSV when determined by IFA. Conversely, 14 (15%) of 95 RSV culture-positive specimens were negative by IFA, whereas DFA missed 19% of the culture-positive specimens. Compared with culture, the Kallestad EIA kit had a sensitivity and specificity of 73 and 92% respectively, but missed 9 (27%) of 33 culture-positive specimens. These data demonstrate that isolation by culture continues to be important for viral diagnosis of REV infections and that for valid comparative studies between viral isolations and rapid detection methods, both sensitive host cells and appropriate test conditions are required.     

Investigation of atypical western blot (immunoblot) reactivity involving core proteins of human immunodeficiency virus type.

Serum specimens which originally exhibited a narrow (indeterminate) 24-kilodalton core protein (p24) or p24/p55 pattern of reactivity with human immunodeficiency virus (HIV) in the Western blot (immunoblot) test were studied to gather information on antibody specificity. A total of 12 specimens were initially reevaluated with an indirect immunofluorescence assay (IFA), three enzyme-linked immunosorbent assays (ELISAs), and Western blot analyses. Five of the specimens were IFA positive and contained anti-gp160/gp120 antibodies which were observed only when an HIV Western blot antigen rich in gp160 and gp120 was used. The remaining seven serum specimens were nonreactive by IFA and showed variable reactivity in HIV antibody ELISAs. The specimens did not cross-react with core antigens for human T-cell leukemia virus types 1 and 2 or contain detectable levels of HIV p24 antigen. The p24/p55 reactivity of six of the seven indeterminate specimens could be reduced or eliminated by preincubating the specimens with disrupted, HIV-infected H9 cells but not with uninfected H9 cells. The six specimens also exhibited discernible reactivity with recombinant HIV p24 antigen. When an additional 23 indeterminate specimens were assayed, all of the serum specimens were nonreactive by IFA while 65% (15 of 23) showed various degrees of reactivity with the recombinant p24 protein. There was no indication that any of the HIV core antibody reactivity was caused by HIV infection. Indeterminate results for five patients with specific p24 reactivity, who were retested after a period of weeks or months, remained indeterminate for HIV antibody with no significant change in ELISA or Western blot reactivity.     

Rapid 24-well plate centrifugation assay for detection of influenza A virus in clinical specimens

Two methods for detection of influenza virus in 234 clinical respiratory specimens were compared: (i) a 24-well plate-centrifugation assay using Madin Darby canine kidney (MDCK) cells and staining with monoclonal antibody pools to influenza A and B (Centers for Disease Control, Atlanta, GA) after incubation for 16 h and 40 h, and (ii) conventional tube cell culture using MDCK cells and primary rhesus monkey kidney cells. Influenza A was identified in 23 specimens (10%). No influenza B was recovered. The rapid centrifugation and tissue culture methods were positive for influenza A in 21 (91%) and 16 (70%) of the 23 specimens, respectively. Fourteen specimens were positive by both methods, 2 were positive by tissue culture alone, and 7 were positive by rapid centrifugation only. Of the 21 specimens positive by rapid centrifugation, 16 (76%) were detected after overnight incubation, and 5 (24%) were positive only after incubation for 40 h. Cytopathic effect was observed in 13 (81%) of the 16 isolates identified by tissue culture after an average of 6 days, and 3 (19%) were identified only by hemadsorption and staining with monoclonal antibodies at day 10. Compared with conventional tissue culture, the 24-well plate centrifugation assay is a more rapid and more sensitive method for detecting influenza virus in clinical specimens.     

Comparative study of respiratory syncytial virus in nasopharyngeal aspirates using conventional cell culture, shell viral centrifugation culture, immunofluorescence and biotin-avidin enzyme linked immunosorbent assays.

133 nasopharyngeal aspirates (NPA) were simultaneously tested for the presence of respiratory syncytial virus (RSV) by conventional cell culture (CCC), shell vial centrifugation culture (SVC), immunofluorescence assay (IFA) and biotin-avidin enzyme linked immunosorbent assay (B-A ELISA). These yielded positive results in 32(24%), 45(33.8%), 36(27%) and 40(30%) of specimens, respectively. Specimens positive by IFA and B-A ELISA were all also positive by SVC. The sensitivity of CCC, IFA, and B-A ELISA comparing to SVC was 71%, 80%, and 88.9%, respectively. For rapid detection of RSV, we recommend the SVC method where a cell culture laboratory is available and the B-A ELISA method where a cell culture laboratory is not available.     

Evaluation of two rapid methods for the detection of herpes simplex virus antigen in patient specimens

An indirect immunofluorescent antibody procedure (IFA) for the detection and typing of herpes simplex virus (HSV) and an enzyme-linked immunosorbent assay (ELISA) procedure were compared with conventional viral culture. Specimens for culture and ELISA were inoculated into serum free viral transport medium (VTM) and, for IFA, onto slides provided in the kit. Tissue cultures (MRC-5 and primary rabbit kidney) were inoculated and examined daily for cytopathogenic effect (CPE). The remaining VTM was frozen at -70 degrees C until tested by the ELISA system. Slides for IFA were stained with HSV common and HSV-2 specific monoclonal antibodies. Of 155 specimens, 47 (30 percent) were unsatisfactory for the IFA test owing to an inadequate number of epithelial cells on the slides. Of 108 adequate specimens, 45 were culture positive; 39 were positive by the IFA test with a sensitivity of 87 percent and a specificity of 90 percent. Of the 39 positives, 29 (75 percent) were correctly classified as type 1 or type 2, six (15 percent) were typed incorrectly, and four (10 percent) were inadequate for typing by the IFA test. All 155 specimens were suitable for testing by the ELISA procedure. Of 55 specimens positive by culture, only 25 (sensitivity 45 percent) were positive by ELISA. However, the specificity was 100 percent. After incubation of two, three, and six days, the tissue cultures detected 71 percent, 89 percent, and 100 percent of the positives, respectively.     

Rapid detection of respiratory viruses by shell vial culture and direct staining by using pooled and individual monoclonal antibodies.

The Bartels respiratory virus panel detection kit is an indirect fluorescent-antibody (IFA) method that uses pooled and individual antisera for tissue culture confirmation of seven respiratory viruses. We evaluated these reagents for detecting viral antigen in shell vial cultures and by direct staining of cells from respiratory specimens. The isolation from 254 specimens of respiratory viruses in shell vial cultures compared with standard tube cultures was highly sensitive (94%) and specific (97.3%). The numbers of viral isolates detected in three consecutive years of testing with shell vial cultures were 68 of 254 (26.8%), 101 of 381 (26.5%), and 122 of 430 (28.4%). IFA direct staining of all 1,065 specimens resulted in 183 (17.2) being uninterpretable because of inadequate numbers of cells or interfering fluorescence. The sensitivity and specificity of the interpretable IFA direct stains in comparison with shell vial cultures were 85.9 and 87.1%, respectively. For detection of 881 adequate specimens, Bartels respiratory syncytial virus IFA direct staining compared with an Ortho Diagnostics Systems direct fluorescent-antibody test for respiratory syncytial virus RSV was highly sensitive (95.5%) and specific (97%). Shell vial cultures combined with Bartels IFA reagents are a rapid alternative to standard tube cultures. Bartels IFA direct staining with individual antisera provides useful same-day screening of respiratory specimens, but the antiserum pool was not effective in screening for positive specimens because of excessive amounts of nonspecific fluorescence.     

Diagnosis of herpes simplex virus infection by immunofluorescence.

The utility of the indirect immunofluorescent antibody (IFA) technique for diagnosis of herpes simplex virus (HSV) infection was examined by testing specimens for this agent from 31 patients with encephalitis or meningitis, 17 with conjunctivitis, 19 with genital disease, and 1 with genital disease and meningitis. Brain biopsy tissue from four patients with encephalitis was positive by IFA and virus culture for HSV. Leukocytes in cerebrospinal fluid from these four patients and one with HSV meningitis were also positive by IFA, but virus isolation attempts on the fluid were all negative. Conjunctival scrapings from two patients with conjunctivitis were positive for HSV by both IFA and virus culture. Eleven of 12 culture-positive lesions of herpes progenitalis were positive by IFA, and 1 dark field-positive syphilitic chancre was also positive for HSV by both IFA and culture. Evidence for specificity of the results was provided by internal controls in each test and negative results from patients with other diagnoses. Thus, the IFA technique constituted a rapid, sensitive, and specific diagnostic method for the diagnosis of HSV infections.     

Comparison of an enzyme immunoassay to an indirect fluorescent immunoassay for the detection of antinuclear antibodies

The standard method for detecting antinuclear antibodies (ANAs) is by immunofluorescence assay (IFA), a method that is labor intensive and subjective. In an attempt to overcome these limitations, several commercial enzyme immunoassays (EIAs) have been developed. We report the results of our evaluation of the ANA Microplate EIA (Sanofi Diagnostics Pasteur, Chaska, MN). For the evaluation, 808 serum samples were tested by EIA and IFA; 52 specimens were positive by both assays, 561 were negative by both assays, 91 were positive by EIA only, and 3 were positive by IFA only. Borderline results (not positive or negative) were obtained for 101 specimens, which were excluded when calculating the sensitivity, specificity, and positive and negative predictive values of this assay, which were 94.6%, 86.0%, 36.4%, and 99.5%, respectively. Because of its high negative predictive value, this assay can be used reliably to detect ANA-negative samples; however, the low positive predictive value indicates that EIA-positive specimens should be retested by an IFA to determine the final result.     

Survey of antibody against chicken anaemia agent (CAA) by an indirect immunofluorescent antibody technique in breeder flocks in Japan

Specific immunofluorescent antigens were detected in MDCC-MSB1 cells, a lymphoblastoid cell line from Marek's disease lymphoma, infected with chicken anaemia agent (CAA) by an indirect immunofluorescent antibody (IFA) technique. The large and small granular antigens were first recognised in a small proportion of the cells at 12 hours postinoculation (PI) with CAA. The antigen-positive cells increased gradually thereafter, and they were misshaped and stained irregularly, showing the occurrence of cytopathic effect, after 24 hours PI. Antibody against CAA was examined by an IFA test, in which the MDCC-MSB1 cells infected with CAA were used as a source of antigen, in breeder flocks in Japan. Thirty-nine out of 40 commercial breeder flocks (97.5%) were found to possess the antibody. Of the 381 individual serum samples, 357 (93.7%) were positive. Two positive flocks were found among the 19 specific-pathogen-free flocks examined, which have been maintained at our and six private research laboratories in Japan. The IFA test had the same sensitivity as the neutralisation test (NT) for detecting the antibody against CAA and could be performed more quickly and easily.     

Use of monoclonal antibodies in an epidemiological marker system: a retrospective study of lung specimens from the 1976 outbreak of Legionnaires disease in Philadelphia by indirect fluorescent-antibody and enzyme-linked immunosorbent assay methods.

Autopsy specimens of lung tissues from 15 patients that contracted legionellosis during the 1976 Philadelphia outbreak of Legionnaires disease were examined for the presence of Legionella organisms and soluble antigens by indirect fluorescent-antibody (IFA) testing and by an enzyme-linked immunosorbent assay (ELISA) with both polyclonal and monoclonal antibodies. In all 15 cases, at least one specimen was positive for Legionella pneumophila serogroup 1 (Lp-1) antigens by a polyclonal antibody ELISA system. Of the 15 cases tested for Lp-1, 9 were positive by a polyclonal antibody IFA test. Nine mouse monoclonal antibodies to Lp-1 gave essentially the same reactivity pattern with extracts from lung tissue homogenates as that obtained with a Philadelphia 1 culture extract by using a monoclonal antibody ELISA system. The same monoclonal antibody panel gave similar results when used in the IFA system with tissue homogenates. Monoclonal antibodies can be used as epidemiological marker systems with both IFA and ELISA testing. This study provides evidence that the 1976 common source outbreak in Philadelphia was probably caused by a single Lp-1 strain. ELISA testing of the soluble antigen of Lp-1 from lung tissue homogenate supernatants was more sensitive than IFA testing of the homogenates and should be extremely useful as either a primary test or as an adjunct to fluorescent antibody testing for legionellosis.     

Rapid and sensitive detection of metapneumovirus in clinical specimens by indirect fluorescence assay using a monoclonal antibody

Human metapneumovirus, with two known genotypes named A and B, is associated with mild respiratory symptoms to severe LRTI in children, high-risk adults and the elderly. Rapid and reliable methods of hMPV detection in clinical samples are essential to implement appropriate care, to better understand the pathology of hMPV and to determine its epidemiology. Respiratory samples from 1,386 patients collected during 2 consecutive years were screened for hMPV using indirect immunofluorescence (IFA) assay with a monoclonal antibody. Forty-three patients tested positive for hMPV by the IFA method. In parallel, the samples were examined with RT-PCR on the F gene. Of these, 41 specimens were RT-PCR positive. The remaining two IF positives were cultured and the cultures were subsequently RT-PCR positive. IFA showed therefore a sensitivity of 100%. No false positive signals were obtained with the influenza virus, respiratory syncytial virus or parainfluenza. When tested by RT-PCR, all IFA-negative samples (n = 204)were found negative. Therefore the specificity of IFA was 100%, IC95 [98-100%], with a negative predictive value of 100%. Based upon phylogenetic analysis of the fusion gene, both subgroups of hMPV were efficiently detected by IFA, and the viral aetiology could be given in 2 hr. These results demonstrate the potential usefulness of immunofluorescence with our monoclonal antibody for the rapid detection of hMPV in clinical specimens in the management of therapy and the control of nosocomial diffusion.     

Comparison of nasopharyngeal aspirate and nasopharyngeal swab specimens for respiratory syncytial virus diagnosis by cell culture, indirect immunofluorescence assay, and enzyme-linked immunosorbent assay.

Paired nasopharyngeal aspirate (NPA) and nasopharyngeal swab (NPS) specimens obtained from each of 32 hospitalized infants with X-ray-confirmed pneumonia (91%) or bronchiolitis were tested for respiratory syncytial virus (RSV) infection by virus culture, the indirect immunofluorescent-antibody (IFA) technique, enzyme-linked immunosorbent assay (ELISA; Ortho Diagnostic Systems, Inc.), and spot hybridization with a human genomic probe to quantitate cellular DNA. RSV was isolated in cell cultures from 72% (23 of 32) of patients by using NPA specimens compared with 47% (15 of 32) by using NPS specimens. With tissue culture positivity as the reference test, the sensitivities of the ELISA on NPA and NPS specimens were found to be 69% (16 of 23) and 61% (14 of 23), respectively, with a specificity and a positive predictive value from both sites of 100%. The sensitivities of the IFA technique compared with the cell culture on NPA and NPS specimens were 61% (14 of 23) and 52% (12 of 23) with specificities of 89 and 78% and positive predictive values of 96 and 92%, respectively. Despite the recovery of significantly more cells (as shown by detection of more cellular DNA by using NPA specimens), virus was detected by the IFA technique or ELISA at similar frequencies in paired specimens. However, virus was recovered more often from NPA than NPS specimens by cell culture, and ELISA optical density readings and the number of RSV-positive fluorescing cells were greater for NPA specimens. NPA specimen collection was less traumatic for the patient, was an easier procedure for the physician to perform, and provided a superior laboratory specimen for RSV diagnosis than the NPS technique.     

Detection of adenovirus by rapid 24-well plate centrifugation and conventional cell culture with dexamethasone

Two methods for rapid detection of adenovirus were tested: (i) 24-well plate centrifugation followed by staining with a monoclonal antibody after incubation for 24 h and 48 h, and (ii) pretreatment of A549 cells used in conventional cell culture and 24-well plate centrifugation with 10(-5)M dexamethasone. Twenty-seven clinical isolates of adenovirus and 12 specimens from which adenovirus had been recovered were included in the analysis. Both isolates and specimens had been frozen at -70 degrees C for up to 6 months. By 24-well plate centrifugation both with and without dexamethasone, 21 (78%) and 27 (100%) isolates were positive for adenovirus at 24 h and 48 h, respectively. Of the specimens, 6 (50%) and 8 (67%) were positive by 24-well plate centrifugation without dexamethasone at 24 h and 48 h, respectively, whereas with dexamethasone 3 (25%) were positive at 24 h and 7 (58%) were positive at 48 h. Overall, combining isolates and specimens, the sensitivity of 24-well plate centrifugation for detection of adenovirus at 24 h was 69% without dexamethasone and 62% with dexamethasone, and at 48 h the sensitivity was 90% without dexamethasone and 87% with dexamethasone. The specificity under all conditions tested was 100%. In conventional tissue culture dexamethasone inhibited recovery of adenovirus. Without dexamethasone, adenovirus was recovered from all 39 samples within 7 days after inoculation; however with dexamethasone pretreatment, the virus was detected in only 31 (79%) of the samples tested in the same period of time.     

Serology of culture-confirmed cases of human granulocytic ehrlichiosis

We evaluated the antibody responses in the sera of 24 patients with culture-confirmed human granulocytic ehrlichiosis (HGE). Antibody titers were measured by an indirect immunofluorescent-antibody assay (IFA) by using a local human isolate as the source of antigen. All patients received appropriate antimicrobial treatment. One hundred five serum specimens collected at baseline and at periodic intervals for up to 14 months were included in the study. Seroconversion was observed in 21 of 23 patients (91.3%) from whom convalescent-phase sera were obtained. Antibodies were first detected at an average of 11.5 days after onset of symptoms. Peak titers (>/=2,560 for 71.4% of patients and >/=640 for 95.2% of patients) were obtained an average of 14.7 days after onset of symptoms. Eleven of 13 patients (84.6%) from whom sera were collected between 6 and 10 months after onset of symptoms were still seropositive, and sera from 5 of 10 (50%) patients tested positive between 11 and 14 months after onset of symptoms. For a subset of 71 serum specimens from 17 patients with culture-confirmed HGE also tested by IFA by using either a human isolate from Wisconsin or an Ehrlichia equi isolate from a horse, there was qualitative agreement for 62 serum specimens (87. 3%). Peak titers were higher, however, with the local human HGE isolate, but the difference was not statistically significant. In summary, most patients with culture-confirmed HGE develop antibodies within 2 weeks of onset of symptoms. Antibodies reach high titers during the first month and remain detectable in about one-half of patients at 1 year after onset of symptoms.     

Detection of human metapneumovirus in respiratory secretions by reverse-transcriptase polymerase chain reaction, indirect immunofluorescence, and virus isolation in human bronchial epithelial cells

Over two winters in Newcastle upon Tyne, respiratory secretions, negative by immunofluorescence staining for other respiratory viruses, were tested for the presence of human metapneumovirus (HMPV) by RT-PCR. In the second winter, specimens were also tested by immunofluorescence staining with an anti-HMPV polyclonal rabbit antiserum and immunofluorescence positive specimens were inoculated into a line of human bronchiolar cells, 16HBE140. Overall, 55 of 549 (10%) specimens tested were positive for HMPV by RT-PCR. Of 162 specimens tested by both RT/PCR and immunofluorescence staining, 23 were positive by both techniques. Of five specimens positive by RT-PCR alone, only one was confirmed with a second set of primers. Of three specimens positive by immunofluorescence alone, only one was confirmed by virus culture. All four previously recognized sub-genotypes of the virus were identified by both RT-PCR and immunofluorescence staining. Sub-genotype A1 was prevalent in the first winter and B1 prevalent in the second. HMPV replication and virus isolation rates were higher in 16HBE140 cells than in monkey kidney cells and did not require exogenous trypsin. Low passage isolates of both sub-genotypes A2 and B1 replicated slowly reaching peak titers only 12 days after inoculation. In summary, single round RT/PCR and immunofluorescence staining with a polyclonal rabbit antiserum proved of equal sensitivity in the diagnosis of HMPV infection in respiratory secretions both detecting 96% of confirmed positive specimens. 16HBE40 cells provided a significant improvement on monkey kidney cells for the isolation and propagation of the virus.     

Rapid detection of respiratory syncytial virus in nasopharyngeal aspirates by a commercial enzyme immunoassay.

A commercial enzyme immunoassay (EIA) for the rapid detection of respiratory syncytial virus (RSV) in respiratory secretions was evaluated by comparison with both virus isolation in HEp-2 cells and indirect immunofluorescence (IFA) staining of exfoliated respiratory cells. Initial examination of 80 nasopharyngeal aspirates collected from infants with acute respiratory illness showed that the RSV EIA was positive for 21 of 24 specimens positive by virus isolation or IFA (87.5% sensitivity) and negative for 53 of 56 specimens negative by virus isolation and IFA (95% specificity). The EIA appears to be an acceptable and more rapid test than virus isolation for the detection of RSV, especially for laboratories in which prompt inoculation of specimens is not always possible. IFA staining with commercial bovine anti-RSV serum was found to be the most sensitive and rapid test for the detection of RSV. However, three of four specimens positive by IFA and negative by virus isolation were not cultured under optimal conditions. In addition, the IFA test requires a highly trained technologist to interpret the staining results.     

Comparison between indirect immunofluorescence assay and shell vial culture for detection of mumps virus from clinical samples

We report a prospective comparison of the efficacies of an indirect immunofluorescence assay (IFA) and shell vial culture (SVC) of throat swab and urine samples from patients with mumps. Throat swab samples were used for the IFA; the urine samples and throat swabs were inoculated into vials of Vero cells. We studied 62 patients by using 62 throat swabs and 50 urine samples (50 patients with both samples). Sixty (96.7%) throat samples were positive in the SVC, and 61 (98.3%) were positive in the IFA. For the 50 patients from whom both samples were available, the IFA was positive in 50 (100%) cases, the urine sample was positive in 49 (98%) cases, and the throat swab was positive in 48 (96%) cases (P > 0.05). This comparison of throat swabs and urine samples has shown that the two clinical samples are similar in efficacy.     

Production of monoclonal antibodies against parainfluenza 3 virus and their use in diagnosis by immunofluorescence.

Monoclonal antibodies were produced against parainfluenza virus type 3 (PI-3) and used to identify PI-3 clinical isolates in cell culture and PI-3 antigen in cells obtained from nasopharyngeal (NP) washes of patients. Two (2E9 and 4G5) of the three monoclonal antibodies characterized reacted by immunoblotting with a 67,000-dalton PI-3 protein, and one antibody (4E5) reacted with two viral proteins in the range of 29,000 to 31,000 daltons. The three monoclonal antibodies did not cross-react by indirect immunofluorescence (IFA) with PI-1 or PI-2 and identified by IFA 18 isolates of PI-3 in cell culture. The 2E9 antibody reacted with PI-3 antigen in cells of 8 NP wash specimens that also yielded PI-3 in cell culture. Cells from 12 specimens reactive by IFA for respiratory syncytial virus, 1 specimen yielding adenovirus in cell culture, and 5 specimens yielding influenza virus were not reactive.     

Utility of direct immunofluorescence and virus culture for detection of varicella-zoster virus in skin lesions.

A direct immunofluorescence assay (DFA) with a monoclonal antibody from Ortho Diagnostic Systems was compared with conventional cell culture for the rapid detection of varicella-zoster virus (VZV) in 140 dermal lesions from 133 patients. A total of 79 (56%) specimens were positive for VZV: 40 (51%) by DFA alone, 2 (3%) by culture only, and 37 (47%) by both culture and DFA. After discordant analysis, the sensitivities and negative predictive values, respectively, were 97.5% (77 of 79) and 96.8% (61 of 63) for DFA and 49.4% (39 of 79) and 60.4% (61 of 101) for viral culture. Of the 39 positive viral cultures, VZV was isolated from 38 (97%) cultures in A549 cells, 23 (59%) in primary rhesus monkey kidney cells, and only 16 (41%) in MRC-5 cells. We conclude that DFA is the optimal method for rapid identification of VZV. In addition, better recovery of VZV in culture may be achieved by using A549 cells.     

Diagnostic utility of PCR-enzyme immunoassay, culture, and serology for detection of Chlamydia pneumoniae in symptomatic and asymptomatic patients.

To assess the utility of PCR-enzyme immunoassay (EIA) for diagnosis of acute infection with Chlamydia pneumoniae, we compared tissue culture, PCR-EIA, direct fluorescent-antibody (DFA) stain, and serology in studies with 56 patients with respiratory symptoms and 80 asymptomatic persons. Thirty-five patients were positive by either culture or PCR-EIA, and 101 were negative by both assays. Thirty specimens from symptomatic patients and one from an asymptomatic patient were culture positive; 23 of these were also PCR-EIA positive. Of the eight culture-positive, PCR-EIA-negative specimens, five were DFA negative and three were DFA positive. Four additional specimens were culture negative and PCR-EIA positive; of these, three were DFA positive and one was DFA negative. When we used culture- and/or DFA-positive results as a reference or "gold standard," the sensitivity and specificity of PCR were 76.5 and 99.0%, respectively. When we used PCR- and/or DFA-positive results as the reference, the sensitivity of culture was 87.5%. On the basis of single acute serum specimens, only 8 of these 35 patients had diagnostic antibody titers. Of the asymptomatic patients, 75% had immunoglobulin G or immunoglobulin M antibody to C. pneumoniae; 15 (18.8%) of these had antibody levels considered to be diagnostic of acute infection. This multicenter study indicates that culture and/or PCR-EIA is more reliable for prompt diagnosis of C. pneumoniae infection than single-point serology alone.