Association of Epstein-Barr virus with primary central nervous system lymphoma in AIDS.
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Acquired Immunodeficiency Syndrome↗
Biomedical subjects
P Charache
Publications and source records attributed to P Charache.
Epstein-Barr virus in AIDS-related primary central nervous system lymphoma.
Primary central nervous system lymphoma occurs more often in patients with AIDS. Epstein-Barr virus (EBV) has been detected in these tumours, but the degree of association has not been defined because of both the highly restricted expression of EBV in malignant tissue and the lack of a technique that is reliable in formalin-fixed paraffin-embedded specimens. EBV-transformed lymphocytes contain short non-protein coding EBV transcripts (EBERs), which are expressed in much higher quantity than other EBV-latency transcripts. We describe a new strategy for detection of latent EBV with these transcripts as targets for in-situ hybridisation. 18 cases of AIDS-related primary CNS lymphoma from a consecutive necropsy series together with specimens from 3 further cases were studied. In each case, a strong positive signal over tumour cells indicated abundant expression of the EBV-EBER1 transcript. This 100% association suggests that the pathogenesis of these AIDS-associated lymphomas may differ from the systemic disease in which only 30-50% of tumours are associated with EBV. A pathogenetic role for EBV was further supported by showing expression of a viral protein (the latent membrane protein) that is implicated as an effector for EBV-associated lymphomagenesis. EBV might have a role as a tumour marker in the diagnosis and management of AIDS-related primary CNS lymphoma.
Central Nervous System Neoplasms↗
The effect of tetracycline on levels of oral contraceptives.
Despite the widespread use of tetracycline for treatment of dermatologic disorders and sexually transmitted diseases, the pharmacodynamics of oral contraceptive use in the presence of tetracycline has not been studied. Seven normal women ingested an oral contraceptive containing ethinyl estradiol 35 micrograms and norethindrone 1 mg in the follicular phase of the menstrual cycle. On day 0 baseline ethinyl estradiol and norethindrone levels were obtained at 0, 1/2, 3/4, 1, 2, 4, 12, and 24 hours after oral contraceptive administration. On day 1 tetracycline 500 mg was given orally every 6 hours while the oral contraceptive was continued. Tetracycline, ethinyl estradiol, and norethindrone levels were determined at the same time intervals as on day 0. Oral contraceptive and tetracycline were continued for up to 10 days, and additional concentrations of ethinyl estradiol, norethindrone, and tetracycline were determined between days 5 and 10. Four additional normal women ingested tetracycline for 5 to 10 days. Tetracycline levels were determined at the time intervals noted above on day 1 and days 5 to 10. No significant decrease in plasma ethinyl estradiol or norethindrone concentration was seen with either short-term (24 hours) or long-term (5 to 10 days) ingestion of tetracycline. Similarly, levels of tetracycline do not significantly decrease with ingestion of a low-dose oral contraceptive.
Adolescent↗
Culture confirmation of cytomegalovirus and herpes simplex virus by direct enzyme-labeled DNA probes and in situ hybridization.
Using probes consisting of horseradish peroxidase (HRP) directly attached to DNA, scrapings or trypsinized cells from 217 adequate clinical samples were cultured and analyzed in 3 blind studies by in situ hybridization for the presence of cytomegalovirus (CMV) and herpes simplex virus (HSV). Sixty samples were judged inadequate due to insufficient cell numbers; however, this problem was significantly decreased during the course of the study. One hundred and eighteen samples were found positive and 70 samples were found negative for CMV. Scrapings of cultured cells from 29 clinical samples revealed 9 samples which were positive and 20 samples which were negative for HSV. Forty-two additional samples, containing either uninfected cells or cells infected with various strains of CMV, were analyzed for the ability of the HRP-DNA CMV probe to detect such isolates. Twenty samples were positive and 22 negative for CMV. No false-negatives or false-positives were observed for either CMV or HSV. In addition to the specificity noted above neither the CMV nor the HSV DNA probe hybridized to potential contaminants found in clinical specimens.
Cell Line↗
Prevalence of infection with human immunodeficiency virus in elective surgery patients.
An anonymous survey of elective surgery patients was performed to assess prevalence of antibody to human immunodeficiency virus (HIV) in a large urban hospital. Of 4087 patients evaluated, 18 (0.4%) were found to be infected with HIV as confirmed by a positive Western blot antibody test. Assessment of risk factors demonstrated that patients with a history of a blood transfusion did not differ in demographics or rate of infection from the population as a whole. Of the 18 HIV infected patients, 13 gave an admission history of one or more risk factors, including 10 with a history of a prior positive test. Only five, or 0.12% of the patients, provided no history of a risk factor or a history of transfusion only. The authors conclude that the prevalence of HIV infection among elective surgery patients is low, and that there would not be any substantial benefit from screening such patients for antibody against HIV.
AIDS Serodiagnosis↗
Abundant expression of EBER1 small nuclear RNA in nasopharyngeal carcinoma. A morphologically distinctive target for detection of Epstein-Barr virus in formalin-fixed paraffin-embedded carcinoma specimens.
The highly restricted expression of the Epstein-Barr virus (EBV) genome in malignancy has limited the use of EBV detection methods applicable to formalin-fixed paraffin-embedded carcinoma specimens. In EBV-transformed lymphocytes very short nonprotein coding EBV transcripts (EBERs) are expressed in much higher abundance (10(7) copies per cell) than other EBV latency transcripts. Using a 3H riboprobe, the authors demonstrated EBER1 expression in NASOPHARYNGEAL CARCINOMAS (NPCs) as well as in parotid salivary gland. Recognition of EBER1 expression was facilitated by the intensity of hybridization and its characteristic morphology (nuclear with nucleolar sparing). EBER1 expression was not demonstrated in other epithelial malignancies arising from mucosal surfaces (oropharynx, uterine cervix) from which EBV shedding has been detected. Repeat study of the NPC specimens with digoxigenin-labeled probe yielded hybridization signal with subcellular morphologic detail and without background in a 12-hour procedure. Thus the EBER1 transcript is an appropriate target for in situ hybridization detection of EBV in formalin-fixed paraffin-embedded carcinoma specimens.
Carcinoma↗
Detection of EBV gene expression in Reed-Sternberg cells of Hodgkin's disease.
EBV DNA has been detected by Southern blot hybridization in 20-25% of Hodgkin's disease tumor specimens and localized to the Reed-Sternberg cells by in situ hybridization. In the present investigation we used a 3H-labelled EBER I anti-sense RNA for in situ hybridization of archival formalin-fixed paraffin-embedded Hodgkin's disease specimens previously shown by Southern Blot hybridization to be EBV-positive. In 6 of 8 specimens neoplastic cells showed an intense signal in virtually all of the tumor cells. The background lymphocytes, eosinophils, plasma cells and histiocytes did not demonstrate significant hybridization. In each case hybridization tended to spare the nucleoli. Hybridization was detected in specimens with histologies of mixed cellularity and nodular sclerosis. The intensity of signal relative to background is better than that in previous studies utilizing 35S-labelled large internal repeat probes for EBV in Reed-Sternberg cells. The exposure time is one week in contrast to the 4-5 weeks reported by others for detection of EBV in Hodgkin's disease. Both increased relative intensity and shorter exposure requirements may be attributed to the very high number of EBER transcripts in the target cells. The demonstration that EBER I is expressed is consistent with a role for EBV in growth regulation of Reed-Sternberg cells and suggests that the virus is not merely a silent passenger in Hodgkin's disease.
Blotting, Southern↗
Inappropriate antimicrobial use in patients with positive blood cultures.
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Anti-Bacterial Agents↗
Oligonucleotides for polymerase chain reaction amplification and hybridization detection of Epstein-Barr virus DNA in clinical specimens.
We designed synthetic oligonucleotide primers and hybridization probe for use in polymerase chain reaction (PCR) amplification and hybridization detection of Epstein-Barr virus (EBV) nucleic acid sequences. Primer sequences were chosen from the coding region for the Epstein-Barr virus nuclear antigen-1 (EBNA-1). PCR amplification and hybridization with these oligonucleotides was carried out on standard laboratory cell lines including African Burkitt's lymphoma and infectious mononucleosis derived cell lines, as well as cell lines recently established from clinical EBV isolates from bone marrow transplant recipients. All EBV cell lines tested were positive. No false-positives were detected with uninfected cell lines, human placental DNA or with other viruses. The sensitivity of the detection procedure was such that four copies of the EBV genome could consistently be detected in a background of 1 microgram of placental DNA. EBV was detected in DNA extracts from the peripheral blood mononuclear cells of two patients with infectious mononucleosis and one patient with viral-associated hemophagocytic syndrome. Three of 18 EBV seropositive patients without known ongoing EBV-associated illness undergoing ambulatory surgery also had EBV detected in DNA extracts from their peripheral blood mononuclear cells. EBV was detected in DNA extracts from lymphoma tissue from two patients with post-transplant lymphomas and two AIDS patients with primary CNS lymphomas. EBV was not detected in 12 B-cell lymphoma specimens from patients without history of immunocompromise. DNA extracts from formalin-fixed paraffin-embedded Hodgkin's tissues previously shown to be EBV positive by Southern blot were also demonstrated to be EBV positive by PCR. Thus, with the oligonucleotides described, PCR is applicable to the detection of EBV in a spectrum of clinical isolates. The broad specificity of these oligonucleotides for all strains of EBV tested is probably a function of the highly conserved sequence of the EBNA-1 DNA binding domain.
Antigens, Viral↗
Rapid detection of Pneumocystis carinii using a direct fluorescent monoclonal antibody stain.
A collaborative study was undertaken at two institutions to assess the performance of a direct fluorescent-antibody stain for the detection of Pneumocystis carinii in respiratory specimens from patients with known or suspected human immunodeficiency virus type 1 infections. A total of 163 specimens (125 induced sputa, 37 bronchoalveolar lavage fluids, and 1 tracheal aspirate) from 124 patients were examined by using modified Giemsa (Diff-Quik; Baxter American Scientific Products, Chicago, Ill.) and direct fluorescent-antibody stains. A total of 73 specimens contained P. carinii, which was detected in 66 (92%) of the specimens by using the modified Giemsa and in 71 (97%) of the specimens by using the fluorescent-antibody stain. One bronchoalveolar lavage fluid specimen in which P. carinii was detected only with the fluorescent-antibody stain was determined to be a false-positive based on subsequent clinical evaluation of the patient. Although the overall time for processing and examining specimens stained with either stain was not significantly different for those specimens containing P. carinii, considerably less time was required for microscopic examination of those fluorescent-antibody-stained specimens lacking P. carinii.
Antibodies, Monoclonal↗
Clinical application of novel sample processing technology for the identification of salmonellae by using DNA probes.
Two hundred and fifty clinical fecal specimens collected over a 7-month period were analyzed for the presence of salmonellae by a rapid DNA hybridization procedure. Hybridizations were performed by using a novel specimen processing protocol called wicking and a previously unreported 1,600-base-pair probe cloned from Salmonella enteritidis DNA. The probe was shown to be reactive with all 70 Salmonella serotypes tested and not reactive with 101 stock strains of other enteric bacteria. Southern analysis of 30 Salmonella isolates representing 22 serotypes suggested that the probe sequence was highly conserved, appearing as a 1,600-base-pair band in a BglII digest of isolate DNA in 29 of 30 isolates and as a 2,300-base-pair fragment in 1 of the isolates. The probe correctly identified all salmonellae (nine isolates) among 47 H2S-producing colonies tested from among 250 clinical specimens cultured on xylose-lysine-desoxycholate medium. Salmonellae grown on xylose-lysine-desoxycholate medium gave consistently higher hybridization values than did those grown on either MacConkey or Hektoen enteric agar. In addition, of eight gram-negative broth enrichments in which salmonellae were identified by conventional means, seven were probe positive. The use of this nucleic acid probe and hybridization technique provides a simple and rapid identification of Salmonella species.
Bacteriological Techniques↗
Detection of surgical pathogens by in vitro DNA amplification. Part I. Rapid identification of Candida albicans by in vitro amplification of a fungus-specific gene.
The management of candidemia and disseminated candidiasis depends on rapid, unambiguous identification of Candida. Such identification is retarded by the slow growth of Candida from clinical specimens. Administration of effective but potentially toxic antifungal therapy is often withheld pending identification. To circumvent this slow growth and thus to expedite diagnosis and therapy, the polymerase chain reaction (PCR) was used to amplify a segment of fungal DNA coding for the cytochrome P450L1A1 (lanosterol-14 alpha-demethylase) in vitro. The technique provides unambiguous evidence of C. albicans in as few as 6 hours with a detection threshold of 10 organisms in a 100 mu specimen. Clinical specimens of urine (n = 4), sputum (n = 6), wound fluid (n = 1, and blood (n = 2) were collected from patients, and C. albicans was conventionally documented at these sites; in each case, PCR was confirmed. Of 17 additional specimens that were culture negative, PCR suggested the presence of yeast in two of the specimens. PCR-based detection of surgical pathogens may have broad application in rapid screening for the presence of organisms either indigenous to a particular surgical intensive care unit or peculiar to selected patient populations.
Candida albicans↗
Comparison of nasal brush and nasopharyngeal aspirate techniques in obtaining specimens for detection of respiratory syncytial viral antigen by immunofluorescence.
Diagnosis of respiratory syncytial virus by antigen detection is dependent on obtaining adequate respiratory epithelial cells. Two specimen collection methods, nasopharyngeal aspiration (NPA) and nasal brushing (NB), were compared. Thirty-two pediatric patients with presumed viral pneumonia or bronchiolitis (34 episodes) had both NPA and NB performed. Of 34 specimens 15 were culture-positive for respiratory syncytial virus. Of these 12 NPA samples and 10 NB samples had viral inclusions by immunofluorescent antibody staining (IFA). Of culture-negative samples, 1 of 17 NB was positive by IFA. One specimen obtained by NB had too few cells to read by the IFA method. Sensitivity and specificity were 80 and 100% for NPA and 67 and 94% for NB. Total respiratory cells and IFA-positive cells (classified as few, moderate, or many) were greater with NPA; however, NB was also an effective procedure and was better tolerated by children, less expensive and easier to perform.
Antigens, Viral↗
Comparison of in situ hybridization and monoclonal antibodies for early detection of cytomegalovirus in cell culture.
The abilities of each of four diagnostic tests--direct fluorescent monoclonal antibody (direct FA) staining, indirect fluorescent monoclonal antibody (indirect FA) staining, in situ hybridization with biotinylated DNA probes, and in situ hybridization with DNA probes directly linked to enzymatically active horseradish peroxidase-to detect cytomegalovirus soon after culture were compared. Only the indirect FA test and the in situ hybridization method with directly linked HRP-DNA probes provided consistent and reliable cytomegalovirus detection as early as 15 h postinfection.
Antibodies, Monoclonal↗
BK and JC virus infections in recipients of bone marrow transplants.
Fifty-five recipients of bone marrow transplants were monitored prospectively for urinary excretion of BK (BKV) and JC (JCV) viruses and for infections with other viruses. For both BKV and JCV, viruria occurred exclusively in patients who were seropositive at transplantation, a finding indicating that shedding of virus was very likely the result of reactivation. BKV reactivation, which occurred in 26 (55%) of 47 BKV-seropositive patients, was far more common than JCV reactivation, which was detected in only two (7%) of 30 JCV-seropositive patients (P less than .0001). In most patients, BK viruria began two to eight weeks after transplantation and resolved spontaneously after a two- to three-week period. Posttransplantation, there was a temporal pattern in the onsets of infection with the different viruses; herpes simplex virus infections occurred first (mean, 7 d), followed by BKV infections (mean, 33 d) and then cytomegalovirus infections (mean, 51 d). BK viruria was associated with hemorrhagic cystitis.
Antibodies, Viral↗
Incidence of cytomegalovirus disease in cyclosporine-treated renal transplant recipients based on donor/recipient pretransplant immunity.
We retrospectively reviewed the clinical data of all renal transplant patients treated with cyclosporine as their main chronic immunosuppressive agent between 12/83 and 11/85 to identify cytomegalovirus-negative patients at our institutions who received cytomegalovirus (CMV)-positive kidneys. Using a latex agglutination test, twenty-two such patients were identified, of whom 2 were excluded due to early death and lack of posttransplant follow-up serology. Of the remaining 20 patients, 12 developed CMV antibody in the first 4 months posttransplant, and of these, 11 were hospitalized with complications related to primary CMV disease. Two of these seroconverting patients eventually died, and one lost her kidney. Of the 8 persistantly CMV-negative patients, 1 lost his kidney soon after transplantation, and one had a febrile illness 4 months posttransplant caused by a bacterial pneumonia. Concomitantly, 145 renal transplants (CMV-negative recipient receiving a CMV-negative kidney or CMV-positive recipient receiving either positive or negative kidneys) given to 142 patients functioned for at least 4 weeks. Only 3 cases of CMV reactivation disease occurred in previously antibody-positive patients. We conclude that the transplantation of a cytomegalovirus-positive kidney into a CMV-negative recipient carries a high risk of mortality/morbidity from primary cytomegalovirus disease. On the other hand, reactivation of CMV disease was uncommon early in the posttransplant course of cyclosporine-treated patients.
Antibodies, Viral↗
Morbid outcome of cytomegalovirus-negative transplant recipients receiving cytomegalovirus-positive kidneys.
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Adolescent↗
Empiric use of vancomycin during prolonged treatment-induced granulocytopenia. Randomized, double-blind, placebo-controlled clinical trial in patients with acute leukemia.
Because gram-positive infections cause morbidity following intensive antileukemic chemotherapy, the effects of vancomycin versus placebo were evaluated in a randomized, double-blind, placebo-controlled trial in 60 adult patients with acute leukemia and first infectious fever during prolonged (mean of 32 days) granulocytopenia. Gram-positive sepsis was associated with first fever in 17 (28 percent) of the 60 patients. None of 31 patients randomly assigned to receive vancomycin demonstrated gram-positive infection, whereas 16 of 22 patients randomly assigned to receive placebo subsequently had gram-positive infection (seven had sepsis, and nine had local infections; p less than 0.005). All patients with gram-positive infection were then given vancomycin, and all showed prompt clinical responses. The predominant gram-positive organism causing infection was beta-lactam-resistant Staphylococcus epidermis (19 of 44 isolates). Patients randomly assigned to receive vancomycin had more rapid resolution of first infectious fever and fewer total febrile days during the granulocytopenic course than did patients randomly assigned to receive placebo. Although vancomycin had no effect on the presence or absence of documented fungal infection, patients treated with vancomycin received empiric amphotericin B for recurrent or persistent fever later (mean of 14 days after initial antibiotic coverage was begun) than did patients receiving placebo (mean of 9.9 days; p less than 0.005), and thus received fewer total days of empiric amphotericin B therapy (mean of 16.3 days) than did patients given placebo (mean of 24.6 days; p less than 0.01). These data demonstrate that empiric use of vancomycin reduces the morbidity of gram-positive infections following intensive antileukemic therapy and decreases the need for empiric use of toxic amphotericin B.
Adult↗