Oropharyngeal samples for detection of Pneumocystis carinii by DNA amplification.
ABSTRACT Pneumocystis carinii pneumonia is a major complication of T-lymphocyte immune deficiency. Restriction of the disease to the alveolar spaces and failure to culture R. carinii has hindered simple diagnostic methods. We have developed a specific DNA amplification method for P. carinii and shown diagnostic sensitivity and specificity exceeding 95% for pneumocystis pneumonia when applied to bronchoscopic lavage and hypertonic saline induced sputum. We here report application of DNA amplification to simple oropharyngeal samples in 31 HIV-positive patients with respiratory illness. P. carinii-specific DNA was detected in 10 of 18 (56%) patients with pneumocystis pneumonia by ethidium bromide stained gels and 14 of 18 (78%) patients by the more sensitive technique of oligoblotting. P. carinii DNA was not detected in samples from 13 patients with other respiratory diagnoses. An oropharyngeal sample offers a simple specimen for detecting P. carinii by DNA amplification; refinements of technique and calibration may allow its development for accurate diagnostic and epidemiological work.
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ABSTRACT: Pneumocystis pneumonia (PCP) is a major cause of hospitalization and mortality in HIV-infected African children. Microbiologic diagnosis relies predominantly on silver or immunofluorescent staining of a lower respiratory tract (LRT) specimens which are difficult to obtain in children. Diagnosis on upper respiratory tract (URT) specimens using PCR has been reported useful in adults, but data in children are limited. The main objectives of the study was (1) to compare the diagnostic yield of PCR with immunofluorescence (IF) and (2) to investigate the usefulness of upper compared to lower respiratory tract samples for diagnosing PCP in children. Children hospitalised at an academic hospital with suspected PCP were prospectively enrolled. An upper respiratory sample (nasopharyngeal aspirate, NPA) and a lower respiratory sample (induced sputum, IS or bronchoalveolar lavage, BAL) were submitted for real-time PCR and direct IF for the detection of Pneumocystis jirovecii. A control group of children with viral lower respiratory tract infections were investigated with PCR for PCP. 202 children (median age 3.3 [inter-quartile range, IQR 2.2 - 4.6] months) were enrolled. The overall detection rate by PCR was higher than by IF [180/349 (52%) vs. 26/349 (7%) respectively; p < 0.0001]. PCR detected more infections compared to IF in lower respiratory tract samples [93/166 (56%) vs. 22/166 (13%); p < 0.0001] and in NPAs [87/183 (48%) vs. 4/183 (2%); p < 0.0001]. Detection rates by PCR on upper (87/183; 48%) compared with lower respiratory tract samples (93/166; 56%) were similar (OR, 0.71; 95% CI, 0.46 - 1.11). Only 2/30 (6.6%) controls were PCR positive. Real-time PCR is more sensitive than IF for the detection of P. jirovecii in children with PCP. NPA samples may be used for diagnostic purposes when PCR is utilised. Wider implementation of PCR on NPA samples is warranted for diagnosing PCP in children.BMC Infectious Diseases 11/2011; 11:329. · 3.03 Impact Factor
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ABSTRACT: Pneumocystis jirovecii is an atypical fungus exhibiting pulmonary tropism and a highly defined host specificity. It is generally regarded as an opportunistic microorganism causing serious pneumonia in AIDS patients. However, with the currently rising number of patients receiving immunosuppressive therapies for malignancies, allogeneic organ transplantations and autoimmune diseases, Pneumocystis pneumonia is becoming more and more recognized in non-HIV immunosuppressed individuals. The clinical presentation in HIV-infected patients may differ from that in other immunocompromised patients and its diagnosis continues to be challenging as there are no specific symptoms or signs. Cotrimoxazole is the drug of choice for prophylaxis and therapy of any form or severity of Pneumocystis pneumonia, but there are only a few options for other alternative treatments. The management of this pneumonia remains a major challenge for all physicians caring for immunosuppressed patients.Expert Review of Anticancer Therapy 06/2010; 8(6):683-701. · 3.22 Impact Factor
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ABSTRACT: Detection of Pneumocystis jiroveci colonization in lungs or oral samples due to high sensitivity of PCR methods results in undue treatment of patients without any symptoms of Pneumocystis pneumonia. The aim of the present study is to demonstrate Pneumocystis carinii in rats, immune suppressed by oral and subcutaneous administration of dexamethasone. Blood, oral, nasal and eye swabs were collected prior to immune suppression and 2, 6, 12 weeks after administration of dexamethasone. Also, samples were collected from lung, heart, liver, kidney, diaphragm, brain, spleen, tongue, muscle, eye, intestine, and feces. Cysts and trophozoites were investigated in stained slides and MSG gene was detected by PCR. The results showed that weight loss is significantly higher in rats administered oral dexamethasone (P<0.05). Microscopy was positive only in lungs of rats orally administered dexamethasone. PCR was positive in lungs and oral swabs of rats prior to the administration of dexamethasone. After the administration of dexamethasone, the MSG gene was detected in oral swabs, lungs, spleen, kidney and (for the first time) in nasal swabs. PCR was positive in nasal swabs during the second and sixth weeks of oral and subcutaneous administration of dexamethasone, respectively. Presence of P. jiroveci in nasopharyngeal aspirate, oropharyngeal wash, oral swab, induced sputum or BAL, and absence in nasal swab in a patient without symptoms of PCP may support clinician's decision regarding colonization. Overall, detection of P. carinii in nasal swabs of rats by PCR demonstrated that nasal sampling can be used for the diagnosis of Pneumocystis pneumonia.Medical science monitor basic research. 01/2013; 19:62-7.