Biomedical subjects

C A Gleaves

Publications and source records attributed to C A Gleaves.

At least 19 recordsLinked to original sources

PubMed1994

Recognition and rapid diagnosis of upper gastrointestinal cytomegalovirus infection in marrow transplant recipients. A comparison of seven virologic methods.

This is a retrospective study of 54 consecutive upper gastrointestinal endoscopies in marrow graft recipients performed to determine the incidence and distribution of CMV infection in symptomatic patients and to compare the sensitivities of 7 CMV detection techniques. At each endoscopy, 3 biopsies were obtained from the esophagus, stomach, and duodenum. Each of the 3 biopsies was assayed for CMV by different techniques. Enteric CMV was identified by one or more techniques in 52 of 486 (11%) biopsies from 14 infected patients. All patients infected with CMV initially had nausea and vomiting. In 13 of these patients, there was esophageal CMV, often associated with stomach (10 patients) and duodenal (7 patients) involvement. CMV infection of the esophagus was never identified cytologically in esophageal imprints or histologically, immunohistologically, or by DNA hybridization in esophageal epithelial cells. The most sensitive diagnostic methods were conventional and centrifugation cultures, which each identified CMV in 17 of the 30 (57%) organs positive by any technique. Indirect fluorescent antibody (IFA) staining for a late CMV antigen detected 53%, followed by in situ DNA hybridization (40%), IFA and immunoperoxidase (IP) staining for an early CMV antigen (37% and 43%), and routine histology (30%). Although no single detection technique is completely adequate for the rapid identification of CMV in small endoscopic biopsies, centrifugation culture is the method of choice, with supplementary immunohistology and in situ hybridization of archival tissue if needed.

Adult↗

PubMed1993

An enzyme immunoassay for the direct detection of adenovirus in clinical specimens.

A total of 414 clinical samples were tested by an enzyme immunoassay (EIA) for direct detection of adenovirus as well as cultured in Graham 293, A549, and human diploid foreskin fibroblast cells. Adenovirus was detected in 69 (16.7%) of 414 clinical specimens. The EIA detected adenovirus in 51 (12.3%) of 414 specimens; two were EIA positive only. Adenovirus was isolated from 67 (16.2%) of 414 specimens; 18 were culture positive only. Compared with viral isolation, EIA had a sensitivity of 73.4% and a specificity of 99.4%, with a positive predictive value of 96% and a negative predictive value of 95%.

Adenovirus Infections, Human↗

PubMed1993

Diagnosis of cytomegalovirus pneumonia by the polymerase chain reaction with archived frozen lung tissue and bronchoalveolar lavage fluid.

The authors developed a polymerase chain reaction (PCR) procedure to detect cytomegalovirus (CMV) in archived, frozen lung tissue and bronchoalveolar lavage (BAL) fluid preparations. The procedure incorporated an internal beta-globin control to assess the adequacy of the sample. Twenty-nine lung tissue and 96 archived BAL specimens from marrow transplant recipients were tested. One of the lung tissue specimens and 46 of the BAL specimens had insufficient tissue for PCR analysis because they did not show a beta-globin band. In lung tissue, the PCR had a sensitivity of 100% and specificity of 89% compared with conventional tube culture. In BAL specimens, the PCR had a sensitivity of 87% and specificity of 90% compared with conventional tube culture or centrifugation culture. Provided tissue was sufficient, the results suggest that the PCR can be used to diagnose CMV interstitial pneumonia effectively from frozen specimens of lung tissue or BAL fluid.

Base Sequence↗

PubMed1993

Ganciclovir sensitivity of cytomegalovirus at diagnosis and during treatment of cytomegalovirus pneumonia in marrow transplant recipients.

Cytomegalovirus (CMV) often persists in the lungs of marrow transplant patients with CMV pneumonia, despite ganciclovir (GCV) treatment. To determine whether GCV resistance contributes to viral persistence, the susceptibilities of CMV isolates from diagnostic bronchoalveolar lavage samples and CMV isolates obtained during treatment or from autopsy lung tissue from 12 patients were compared by DNA hybridization. Resistance (50% effective dose, > 12 microM) was detected in an isolate from only one patient who had also received several courses of GCV. GCV resistance did not explain the persistence of CMV in the lung.

Bone Marrow Transplantation↗

PubMed1992

Hepatitis due to herpes simplex virus in marrow-transplant recipients.

Eight cases of hepatitis due to herpes simplex virus (HSV) (five "confirmed," three "possible") were identified among marrow-transplant recipients at the Fred Hutchinson Cancer Research Center between 1975 and 1988. The clinical and pathological characteristics of these patients are described and compared with published findings for non-marrow-transplant recipients with HSV hepatitis. Clinical syndromes in the marrow-transplant recipients ranged from fever associated with abdominal pain and elevations in serum aminotransferase levels to fulminant hepatitis. Evidence of hepatic infection appeared before day 20 following transplantation unless prophylaxis with acyclovir was given, in which case HSV hepatitis did not appear until after day 40. All patients died, although the one patient who was given early, empirical high-dose acyclovir therapy clearly improved with antiviral therapy and may have died of other causes. Involvement of multiple organs with HSV was found in three of four patients with confirmed HSV hepatitis who underwent autopsy. In all cases, reactivation of latent HSV was the presumed source of the HSV hepatitis. HSV hepatitis should be considered when HSV-seropositive recipients of marrow transplants develop abdominal pain, fever, and elevations in aminotransferase levels.

Adult↗

PubMed1992

An outbreak of respiratory syncytial virus in a bone marrow transplant center.

An outbreak of respiratory syncytial virus (RSV) infection occurred among 31 patients in a marrow transplant center over a 13-week period beginning in January 1990. RSV infection was also documented in 35 family members and employees. Of 18 patients with pneumonia, 14 (78%) died. None of 13 with upper respiratory infection died. Preengraftment patients tended to develop pneumonia more frequently than did engrafted patients. Early administration of ribavirin may have had a beneficial effect in patients with pneumonia. Antigenic and genomic analysis of 14 available isolates suggested that at least four different viral strains were responsible for the outbreak. One group of patients and 1 employee in spatial proximity were infected with the same strain and likely acquired their infections nosocomially. RSV infection in marrow transplant patients is a serious and life-threatening infection with a high mortality rate once pneumonia develops.

Adolescent↗

PubMed1992

Quantification of cytomegalovirus in bronchoalveolar lavage fluid after allogeneic marrow transplantation by centrifugation culture.

A technique to quantify cytomegalovirus (CMV) by centrifugation culture of bronchoalveolar lavage fluid from marrow transplant recipients was developed. This technique was used to assess the CMV response to antiviral treatment and the relationship between viral load, asymptomatic excretion versus symptomatic infection, and prognosis. Relative to tube cell culture, centrifugation culture of bronchoalveolar lavage fluid was more sensitive than direct fluorescent-antibody staining. It was also a rapid, replicable method for detecting and measuring the amount of CMV. There was no significant difference between viral load at diagnosis and after 9 days of treatment with ganciclovir and intravenous immunoglobulin. Viral load was not predictive of outcome, and there was no difference in amount of virus between patients with asymptomatic CMV excretion and those with CMV pneumonia. The amount of CMV may not be as important as other factors (e.g., host immune response) in the pathogenesis of CMV pneumonia.

Bone Marrow Transplantation↗

PubMed1992

Evaluation of an automated immunodiagnostic assay system for direct detection of herpes simplex virus antigen in clinical specimens.

The Vitek ImmunoDiagnostic Assay System (VIDAS) is a 2 1/3-h automated qualitative enzyme-linked fluorescent immunoassay developed for the direct detection of herpes simplex virus (HSV) antigen in clinical specimens. A total of 356 clinical specimens submitted for HSV isolation were prospectively evaluated with the VIDAS, and the results of the technique were compared with those of both HSV isolation in cell culture and Herpchek, a nonautomated enzyme immunoassay. Compared to cell culture, VIDAS had a sensitivity of 91.6% and a specificity of 89.3%, with positive and negative predictive values of 82.6 and 95.0%, respectively. In comparison to Herpchek, VIDAS had a sensitivity of 93.7% and a specificity of 93.0%, with positive and negative predictive values of 89.4 and 95.9%, respectively. The results demonstrated that the VIDAS required minimal manipulation in order to produce results comparable to those of Herpchek and HSV isolation in cell culture.

Antigens, Viral↗

PubMed1992

Detection of human cytomegalovirus in clinical specimens by centrifugation culture with a nonhuman cell line.

The sensitivities of MRC-5 and mink lung (ML) cells in centrifugation culture were compared simultaneously for the detection of cytomegalovirus (CMV) IE antigen (immediate-early antigen) from clinical specimens. Of 413 samples assayed, 51 (12%) were positive for CMV by both centrifugation and standard cell culture. At 20 h postinoculation (p.i.), 46 of 51 (90.2%) CMV-positive specimens were detected in ML cells. At 40 h p.i., 50 of 51 (98.0%) CMV-positive specimens were detected in ML cells, compared with 48 of 51 (94.0%) in MRC-5 cells. There was no significant difference in the detection of CMV in either cell line by centrifugation culture. However, in 19 of 23 positive samples that had countable foci at 20 h p.i., there was a 25% increase in the number of positive foci observed for ML cells compared with MRC-5 cells. Less toxicity was also noted for ML cells than for MRC cells, particularly in viral blood specimens. These data suggest that ML cells are comparable to MRC-5 cells for the rapid detection of CMV by centrifugation culture.

Animals↗

PubMed1991

Early treatment with ganciclovir to prevent cytomegalovirus disease after allogeneic bone marrow transplantation.

BACKGROUND: Cytomegalovirus (CMV) infection is a major cause of morbidity and mortality after allogeneic bone marrow transplantation. We conducted a controlled trial of ganciclovir for the early treatment of CMV infection in asymptomatic recipients of bone marrow transplants whose surveillance cultures for CMV became positive. METHODS: Bone marrow--allograft recipients who were seropositive for CMV antibodies or who received seropositive marrow were screened for CMV excretion by culture of throat swabs, blood, urine, or bronchoalveolar-lavage fluid. In this double-blind trial, 72 patients who had marrow engraftment and were excreting virus were randomly assigned to receive either placebo or ganciclovir (5 mg per kilogram of body weight twice a day for one week, followed by 5 mg per kilogram per day) for the first 100 days after transplantation. Patients were followed for the development of biopsy-confirmed CMV disease, ganciclovir-related toxicity, and survival. RESULTS: Between assignment to the study drug and day 100 after transplantation, CMV disease developed in only 1 of the 37 patients assigned to receive ganciclovir (3 percent), but in 15 of the 35 patients assigned to receive placebo (43 percent, P less than 0.00001). The ganciclovir recipients had rapid suppression of virus excretion; 85 percent had negative cultures after one week of treatment, as compared with 44 percent of the placebo group (P = 0.001). The principal toxic reaction was neutropenia; 11 ganciclovir recipients had an absolute neutrophil count below 0.75 x 10(9) per liter, as compared with 3 placebo recipients (P = 0.052). Treatment was discontinued in 11 ganciclovir recipients and 1 placebo recipient because of neutropenia (P = 0.003). After treatment was stopped, the neutrophil count recovered in all patients. Overall survival was significantly greater in the ganciclovir group than in the placebo group both 100 days and 180 days after transplantation (P = 0.041 and 0.027, respectively). CONCLUSIONS: Early treatment with ganciclovir in patients with positive surveillance cultures reduces the incidence of CMV disease and improves survival after allogeneic bone marrow transplantation.

Adolescent↗

PubMed1991

Comparison of MRC-5 and U-373MG astrocytoma cells for detection of cytomegalovirus in shell vial centrifugation cultures.

U-373MG astrocytoma cells are susceptible to human cytomegalovirus (CMV) infection and offer the advantage of a continuous cell line for clinical laboratory use. U-373MG to MRC-5 cells for detection of CMV by centrifugation culture were therefore compared. At 20 h, 10 (6.1%) versus 12 (7.4%) of 163 clinical specimens were positive for CMV, and at 40 h, 12 (7.4%) versus 17 (10.4%) were positive in U-373MG and MRC-5 cells, respectively. Substantial toxicity was found in U-373MG cells (84%) when inoculated with blood specimens. For detection of CMV in centrifugation culture, MRC-5 cells are superior due both to higher sensitivity and lesser toxicity.

Astrocytoma↗

PubMed1990

Evaluation of an enzyme immunoassay for the detection of herpes simplex virus (HSV) antigen from clinical specimens in viral transport media.

A total of 301 clinical specimens (229 culture positive and 72 culture negative) were assayed retrospectively by an enzyme immunoassay. Specimens were transported to the virus lab in viral transport media (VTM), and were inoculated into HF and A549 cell culture tubes for viral isolation with the remainder of the sample being saved at -70 degrees C. Specimens were thawed, vortexed and resuspended in 10 x Herptran concentrate and each sample was then added in duplicate to designated wells of a microtiter plate for the EIA assay. The EIA detected 147/150 (98%) culture positive specimens from symptomatic patients, 63/79 (79.7%) culture positive specimens from patients considered asymptomatic and 210/229 (91.7%) culture positive specimens overall. The EIA was negative for 70/72 (97.2%) culture negative specimens. These data suggest that the EIA test can be used with clinical specimens submitted in conventional VTM. However, VTM samples which are EIA negative, particularly with EIA values close to the EIA positive cutoff value, need to be cultured.

Antigens, Viral↗

PubMed1990

Use of serum substitutes for the growth of four cell lines commonly used for virus isolation.

Four serum extenders (Opti-MEM I, BioRich, Excyte I and Excyte III) and two serum substitutes (Omni Serum and Serum Plus) were evaluated for the reduction or replacement of FBS for the growth and maintenance of four representative cell lines used for virus isolation. MRC-5, human fetal foreskin fibroblast (HF), A549 and Hep-2 cells were seeded into culture flasks containing MEM with 10% FBS or with the serum extenders or substitutes and subcultured into 16 x 125 mm glass tubes and 1-dram shell vials. Only cells cultured with Opti-MEM I and Omni Serum grew consistently in tubes and vials and these reagents were compared to FBS for viral isolation and detection. Laboratory stocks of CMV, HSV, VZV, adenovirus (Ad) and RSV were tested. In HF and MRC-5 cells, CMV was isolated in cells cultured in either Opti-MEM I or Omni Serum before CPE appeared in cultures containing FBS. Ad and VZV CPE were observed first in HF and MRC-5 cells containing Omni Serum. HSV CPE was seen at the same time in HF and MRC-5 cells with all 3 media. HSV and Ad CPE in A549 cells were also seen at the same time in all 3 media. RSV and Ad CPE in Hep-2 cells were observed first in FBS containing media. Both Opti-MEM I and Omni Serum performed well as substitutes for FBS for growth of stock viruses without loss of cell sensitivity.

Animals↗

PubMed1990

Evaluation of the CMV-CUBE assay for detection of cytomegalovirus serologic status in marrow transplant patients and marrow donors.

We evaluated a new membrane dot immunobinding assay (CMV-CUBE; Difco Laboratories) for the detection of cytomegalovirus (CMV) antibody in marrow transplant patients and donors. The CMV-CUBE assay was compared with a commercially available enzyme immunoassay (EIA; CMV STAT) and a latex agglutination (LA; CMVScan) test. Serum samples were collected from 311 transplant patients and donors prior to transplantation. A total of 164 serum specimens were positive for CMV antibody by one or more of the three assays, with 153 of 164 samples (93.3%) positive by all three tests. A total of 147 serum specimens were CMV antibody negative. CMV-CUBE detected 154 of 164 (94%) of the positive samples, EIA detected 160 of 164 (97.5%), and LA detected 157 of 164 (95.7%) CMV-positive samples. Compared with EIA, CMV-CUBE had a sensitivity of 95.6% and a specificity of 99.3%. Compared with LA, CMV-CUBE had a sensitivity of 97.5% and a specificity of 99.4%. CMV-CUBE is a simple and rapid visual assay which can be used for the qualitative detection of antibody to CMV in patient serum.

Antibodies, Viral↗

PubMed1989

Rapid detection of cytomegalovirus in bronchoalveolar lavage specimens from marrow transplant patients: evaluation of a direct fluorescein-conjugated monoclonal antibody reagent.

An FITC-conjugated monoclonal antibody reagent containing three CMV-specific monoclonal antibodies was evaluated for the rapid detection of CMV in bronchoalveolar lavage (BAL) cytospin preparations by direct IF (DFA). Eighty-six BAL samples from 72 marrow transplant patients were inoculated into both centrifugation and standard cell culture. CMV was detected in 49/86 (57%) BAL samples. DFA detected 37/46 (80%) samples which were positive in centrifugation culture. While DFA staining lacked the sensitivity (overall sensitivity 38/49, 78%) to replace either standard or centrifugation culture, the total laboratory time needed to complete the DFA was only 1.5 h and its concurrent use with centrifugation culture can provide rapid specific diagnosis of CMV pneumonia.

Antibodies, Monoclonal↗

PubMed1989

Detection of cytomegalovirus from clinical specimens in centrifugation culture by in situ DNA hybridization and monoclonal antibody staining.

An in situ DNA hybridization kit for cytomegalovirus (CMV) was evaluated for the detection of CMV in centrifugation culture. Of 61 clinical specimens, 17 (27.8%) were positive for CMV by monoclonal antibody staining following centrifugation. Of the 17 positive specimens, 15 were detected by DNA hybridization (24.5%). However, the earliest that CMV could be detected by DNA hybridization was 58 h as compared with 16 h with monoclonal antibodies following centrifugation. DNA hybridization remains of great interest for the study and detection of CMV infection. However, current DNA hybridization techniques are not sufficiently rapid to replace the use of monoclonal antibodies in centrifugation culture.

Antibodies, Monoclonal↗

PubMed1989

Direct detection of cytomegalovirus from bronchoalveolar lavage samples by using a rapid in situ DNA hybridization assay.

An in situ DNA hybridization assay was compared with centrifugation culture for rapid detection of cytomegalovirus (CMV) from bronchoalveolar lavage (BAL) samples. Eighty BAL samples were inoculated into both centrifugation culture and standard culture. Cytospin preparations of the BAL samples were studied in a 75-min in situ DNA hybridization assay using the PathoGene CMV kit (Enzo Biochem, Inc., New York, N.Y.). Of the 80 samples, 39 (49%) were positive for CMV; 37 of 39 (95%) were positive by centrifugation culture, 34 of 39 (87%) were positive in standard culture, 24 of 39 (62%) were positive by in situ hybridization, and 20 of 39 (56%) were positive by histologic and/or immunofluorescence techniques. The in situ hybridization assay detected 23 of the 37 samples positive in centrifugation culture, for a sensitivity of 62% and a specificity of 98%. We conclude that the in situ hybridization assay is a specific and more rapid test than centrifugation culture and standard culture for diagnosis of CMV pulmonary infection. For the clinical laboratory, however, current hybridization methods are not sufficiently sensitive to replace centrifugation culture for detection of CMV in BAL specimens.

Bronchoalveolar Lavage Fluid↗

PubMed1989

Evaluation of a latex particle agglutination assay for the detection of cytomegalovirus antibody in patient serum.

The Virogen CMV Antibody Test is a simple and rapid latex agglutination assay for the detection of cytomegalovirus antibody in human serum and plasma. Evaluation of this assay with respect to enzyme immunoassay yielded a sensitivity of 98% with a specificity of 100%. In comparison to CMVScan, the Virogen CMV Antibody Test had a sensitivity of 98.4% and a specificity of 100%.

Antibodies, Viral↗