Comparison of the Sofia and Veritor direct antigen detection assay systems for identification of influenza viruses from patient nasopharyngeal specimens.

Influenza antigen detection assays (Sofia fluorescent immunoassay [FIA] and Veritor) yield objective results, which are potentially useful for point-of-care testing. The assays were evaluated with reverse transcriptase PCR (RT-PCR) using 411 nasopharyngeal swab specimens. Sensitivity and specificity values (percentages) of 79.0/99.0 and 64.0/99.4 for influenza A and 92.9/96.7 and 78.6/98.7 for influenza B were obtained for the Sofia and Veritor assays, respectively.

Evaluation of respiratory syncytial virus (RSV) direct antigen detection assays for use in point-of-care testing.

RSV infections cause lower respiratory tract infections and result in surges in physician's office, emergency department visits and hospitalizations, especially in infants and toddlers. Point-of-care (POC) testing reduces healthcare costs and permits informed decisions on treatment, however, optimal POC assays must be sensitive, easy to perform and provide rapid results. A prospective study tested 230 patient nasopharyngeal specimens using 4 RSV direct antigen detection assays (Directigen, Quickvue, Sofia and Veritor) and RT-PCR. A RSV dilution study was also performed to evaluate sensitivity. RSV fluorescent antibody testing in 46/230 patients was also evaluated. Sensitivity values obtained for the Sofia, Veritor, Directigen and Quickvue assays (%) were 85, 72.5, 70 and 57.5, respectively. Fluorescent result interpretation may account for Sofia's enhanced sensitivity. Specificity (%) was 97-100 among assays. Sensitivity data were confirmed in the dilution studies. Fluorescent antibody testing demonstrated 64% sensitivity compared with RT-PCR. Objective result reporting, walk away testing and high sensitivity make the Sofia a valuable choice for POC testing. Veritor's sensitivity may also render it acceptable in POC. Lack of objective results by Directigen and the poor sensitivity observed by Quickvue may preclude their value in diagnostic testing.

Evaluation of verification assays in EIA specimens presumptively positive for Chlamydia trachomatis.

Verification of specimens positive for Chlamydia trachomatis by enzyme immunoassay (EIA) has been recommended when testing low prevalence populations. This study compared direct fluorescent antibody (DFA) and blocking antibody (BLA) verification assays in specimens presumptively positive for C. trachomatis by the Syva Microtrak II EIA. Of 1785 specimens originally tested by EIA, 96 were presumptively positive for C. trachomatis. Verification assays were concordant in 86 specimens (69 positive, 17 negative); nine of the remaining samples gave positive results in a second EIA and one was unresolved. Both verification assays gave some false-negative results. When initial EIA absorbance values were correlated with verified results, all EIA false positive results had absorbances in the low range (less than a three-fold increase over assay cut-off values). Verification of EIA results by both DFA and BLA was effective in detecting false positive results, but confining verification to low-value positive specimens could be considered for cost effective C. trachomatis testing.

Use of continuous human lung cell culture for adenovirus isolation.

Adenovirus sensitivity, TCD50 value and mean time to cytopathic effect (CPE) were compared in Madin-Darby canine kidney (MDCK), rhesus monkey kidney (RhMK), human lung carcinoma (A549) and in newly identified continuous human lung (CHL) cell cultures. High-titer stock and positive original specimens were inoculated in culture. Patients specimens were also placed in shell-vial cultures. A549 and CHL cells demonstrated comparable sensitivity, however the former generally produced CPE earlier. Sensitivity was respectively reduced and poor in RhMK and MDCK cultures, as expected in cells of nonhuman origin. The performance of CHL for adenoviruses and other pathogens makes them a useful addition in diagnostic virology laboratories.

Comparison of rapid detection methods for influenza A virus and their value in health-care management of institutionalized geriatric patients.

Respiratory specimens from 160 geriatric patients with suspected influenza illness were used to evaluate the abilities of two enzyme immunoassays (EIAs; Directigen FLU-A [Becton Dickinson Microbiology Systems, Cockeysville, Md.] and Prima EIA [Baxter/Bartels Diagnostics, Inc., Issaquah, Wash.]) and direct immunofluorescence testing (immunofluorescence assay [IFA]) to identify influenza A virus. In comparison with culture isolation, the sensitivities and specificities of the IFA, Directigen FLU-A, and Prima EIA were 92.5 and 97.2%, 86.8 and 99.1%, and 92.5 and 98.1%, respectively. In contrast to EIA, IFA was labor intensive and required a high degree of technical expertise, and the results of IFA were difficult to interpret. These factors may preclude the use of IFA for testing large numbers of specimens. A retrospective epidemiologic survey of influenza infection was done in six geriatric institutions which had used either rapid and culture testing or culture alone. Preventable cases of influenza A virus infection ranged from 9 to 38% of all cases in facilities which used culture testing only and which had not instituted amantadine prophylaxis. The use of direct specimen testing is recommended as an adjunct to culture isolation for the identification of influenza A virus. Use of a combination of these methods permits the timely administration of appropriate antiviral therapy and infection control measures, while it also permits the antigenic surveillance of circulating influenza strains, which is necessary for present vaccine efficacy evaluations and the creation of future effective vaccine formulations.

Echovirus type 30 infection associated with aseptic meningitis in Nassau County, New York, USA.

An outbreak of echovirus type 30, primarily affecting infant under 1 year old, was documented in Nassau County, N.Y., USA. Twenty-four of the 40 infected patients were definitively diagnosed with aseptic meningitis. A positive correlation was found between the proportion of polymorphonuclear leukocytes and the overall cerebrospinal fluid white cell count.

Evaluation of three immunoassays for detection of Chlamydia trachomatis in urine specimens from asymptomatic males.

The performances of three commercially available immunoassays (Chlamydiazyme/Antibody Blocking Assay [Abbott Diagnostics, Abbott Park, Ill.], IDEIA [Analytab Products, Plainview, N.Y.], and Microtrak EIA [Syva Co. Palo Alto, Calif.]) were evaluated for the detection of Chlamydia trachomatis in urine specimens from asymptomatic males. Assay results were compared with direct specimen immunofluorescence (DFA) analysis of urine sediment (Syva Microtrak; Syva Co.), which was chosen as the study confirmation assay. An overall Chlamydia prevalence of 7% (24 of 340) was found in our study population, with peak incidences occurring in the adolescent (8 of 93 specimens) and young adult (11 of 146 specimens) age groups. Sensitivity and specificity data among the Chlamydiazyme, IDEIA, and Microtrak enzyme immunoassays (EIAs) were determined to be 79.1 and 99%, 91.7 and 98%, and 95.8 and 99%, respectively. The Microtrak EIA and IDEIA products demonstrated sensitivities and specificities equal to or greater than those claimed for urine specimens. The diagnostic accuracies of these assays on asymptomatic subjects, along with the ease of this collection method, suggest a role for these products as screening tools. The sensitivity of the Chlamydiazyme assay was lower than that claimed previously in symptomatic patients, with 5 of 24 positive specimens demonstrating false-negative results. In those cases, centrifugation of the original immunoassay aliquot material and then DFA examination confirmed specimen positivity. Urine immunoassay screening in combination with DFA confirmation (which was chosen because it has antibody epitopic specificity different from that of the primary assay) provides a high degree of diagnostic precision. The use of noninvasive collection methods could result in greater testing compliance among asymptomatic males and, subsequently, could reduce the incidences of both symptomatic and silent chlamydial infections.

Enhanced detection of cytomegalovirus in shell vial culture following MRC-5 monolayer pretreatment with glucocorticoids.

The effect of cortisol and its synthetic derivative, dexamethasone on the infectivity of cytomegalovirus (CMV) was investigated. Twenty-four to 48 h pretreatment of MCR-5 shell vials by 100 micrograms % (compared with medium supplemented with 10 or 25 micrograms %) cortisol affected the greatest increase in virus infectivity (measured in fluorescent focus units/0.2 ml inoculum). This enhancement effect was more pronounced in the CMV strain AD-169 than in a clinical CMV isolate (NCMC 7125). Attempts to reisolate CMV from 32 frozen positive original specimens in the presence or absence of cortisol demonstrated an enhanced infectivity of 8 specimens in hormone pretreated cultures compared with two specimens showing an increased activity in untreated controls. Three specimens were reisolated only in hormone pretreated shell vials. Pretreatment of shell vials maintained in maintenance medium plus 2% FBS with cortisol or dexamethasone (10(-5) M) increased CMV infectivity to ca. 300%. This hormonal mediated effect was more pronounced in 4 rather than 16 day old shell vials. Infectivity in 16 day old shell vials maintained in 10% FBS but without hormone equalled the infectivity levels achieved in 4 day old shell vials with cortisol and either a 2 or 10% FBS supplement. This study denotes the applicability of glucocorticoid pretreatment of MRC-5 monolayers but more importantly, identifies the association between FBS concentration and glucocorticoid-mediated CMV detection using shell vial technology.

Evaluation of five monoclonal antibody-based kits or reagents for the identification and culture confirmation of herpes simplex virus.

Five immunofluorescence (IF) kits or reagents (Bartels [Bartels Immunodiagnostic Supplies, Inc., Bellevue, Wash.], Imagen [CellTech Diagnostics, Ltd., distributed by Analytab Products, Plainview, N.Y.], Ortho [Ortho Diagnostics Systems, Inc., Raritan, N.J.], Syva [Syva Co., Palo Alto, Calif.], Whittaker [Whittaker Bioproducts, Walkersville, Md.]) were evaluated for typing and laboratory confirmation of herpes simplex virus (HSV). Of 101 clinical isolates tested by each kit or reagent, results for 97 of them were in agreement. Identification of the four isolates with discordant results was performed by restriction endonuclease analysis of the viral DNA. The sensitivity and specificity of the Imagen and Bartels kits were 100%. For the Ortho, Syva, and Whittaker kits or reagents, the HSV type 1 (HSV-1) and HSV type 2 (HSV-2) sensitivities were 97.4 and 100%, 100 and 100%, and 97.4 and 100%, respectively, and the specificities were 100 and 97.4%, 100 and 92.4%, and 100 and 97.4%, respectively. There was one false-positive HSV-2 isolate identified by each of the Ortho and Whittaker kits or reagents. Three false-positive HSV-2 isolates occurred by staining with Syva, giving the erroneous indication of dual isolates. Several isolates stained with Imagen and Whittaker reagents displayed dull IF patterns. A dull green background occurred in ca. one-third of the HSV-2 isolates tested with the Ortho kit. The intensities of IF staining by the Bartels and Syva kits were satisfactory; however, the latter displayed a specificity of 92.7%. A total of 38 and 63 specimens were finally designated as HSV-1 and HSV-2, respectively. Identification of each isolate with the Bartels kit was consistently interpretable and is recommended as the typing and confirmatory assay of choice.

Occurrence of nonspecific reactions among stool specimens tested by the Abbott TestPack rotavirus enzyme immunoassay.

Sixty-five stool specimens obtained from children suffering from gastroenteritis were tested for the presence of antigen to rotavirus by the Abbott TestPack Rotavirus (TestPack) enzyme immunoassay kit. The Kallestad Pathfinder enzyme immunoassay, polyacrylamide gel electrophoresis, immune electron microscopy, and virus isolation were utilized as reference assays. Fifty-four specimens were in accord by TestPack and Kallestad Pathfinder. Among 11 discordant specimens positive with TestPack but negative by Kallestad Pathfinder, rotavirus was not identified by polyacrylamide gel electrophoresis, immune electron microscopy, or isolation in primary African green monkey kidney cell cultures. TestPack displayed a performance specificity of 83%. The inordinately high number of stool specimens reported as false-positive by TestPack precludes the incorporation of this antigen detection kit into our routine regimen of diagnostic virologic testing.

Morphometric analysis of bone marrow sinus cell elements after induction of monomyelocytic leukemia in BALB/c mice.

Morphometric measurements of bone marrow sinus wall adventitial and endothelial cells were made in normal and leukemic samples by using a Bioquant II computer image analysis program (R & M Biometrics, Nashville TN). Electron micrographs of bone marrow from at least eight mice per group were studied in normal and day-2, day-4, and day-6 leukemic postinoculation groups. The adventitial cells abuting the sinus endothelium was found to be significantly reduced with disease progression. In addition, adventitial cell area and perimeter were found to be drastically reduced with disease progression whereas endothelial cell area and perimeter showed no significant differences. Other investigators have suggested that the marrow sinus adventitial cell cover is drastically reduced in rodent leukemias (Chen et al.; Blood, 39:99-112, 1972; Campbell; Am. J. Anat., 142:319-334, 1975). Our findings give quantitative data to support this hypothesis.

Sustained hypertransfusion and induction of a transplantable myeloid leukemia in RLV-A-infected BALB/c mice.

Infection of BALB/c mice with the RLV-A virus typically results in an erythropoietic dysplasia characterized by hepatosplenomegaly, erythroblastosis, erythroblastemia and severe anemia without reticulocytosis. Mice hypertransfused weekly with 75%-packed red cells for 42 days prior to RLV-A infection and viral potency controls manifested this typical RLV-A response. Mice that were hypertransfused prior to and following RLV-A infection never developed the "typical" RLV-A pathogenesis. Instead, a transplantable myeloid leukemia was established. Although the reason for altered pathogenesis remains uncertain, it seems plausible that continued hypertransfusion, presumably after establishment of an altered granulopoietic microenvironment, resulted in a completely different viral expression and development of the transplantable myeloid leukemia.

Effect of sustained hypertransfusion on Rauscher leukemia virus-variant A (RLV-A) infection in BALB/c mice.

Infection of BALB/c mice with the RLV-A virus normally induces an erythropoietic dysplasia characterized by hepatosplenomegaly, erythroblastosis, erythroblastemia and severe anemia without reticulocytosis. Time to death varies between 20-30 weeks. Mice were inoculated with RLV-A after being hypertransfused with 75% packed red cells for 42 days which has been shown to eliminate erythropoiesis and modify the microenvironment to favor granulopoiesis. Following RLV-A inoculation, one group did not receive further transfusion (short-term) and another group continued with hypertransfusion weekly (long-term). The pathogenesis of RLV-A in the short-term group paralleled the characteristic RLV-A response. In the long-term group however, the characteristic RLV-A response was never observed. Instead, a granulocytic leukemia was developed. Continued hypertransfusion presumably after establishment of an altered microenvironment resulted in a completely different viral pathogenesis and the development of a transplantable myeloid leukemia.

Renal and extrarenal production of erythropoietin following exchange transfusion with plasma or a perfluorocarbon-polyol.

The extrarenal production of erythropoietin (Ep) was measured in young adolescent rats following exchange-transfusion with normal rat plasma or the whole blood replacement fluid, Fluosol-43. In the plasma-transfused anephric groups, the peak extrarenal Ep response to the severe degree of anemic hypoxia represented 23% of that evoked in renal-intact groups. In contrast, the peak extrarenal production of Ep in Fluosol-transfused renoprival groups was almost equal to those serum Ep levels produced in renal-intact groups. Histological studies revealed significant fluorocarbon uptake by hepatic macrophages that coincided with the time of peak production of Ep. Thus, while intense levels of hypoxia are required to stimulate the extrarenal production of Ep, these data suggest that as Fluosol-43 is taken up by the hepatic macrophages, Ep production by these cells is stimulated.