Etiologic Diagnosis of Adult Bacterial Pneumonia by Culture and PCR Applied to Respiratory Tract Samples

Abstract

Respiratory culture and multiplex PCR for Streptococcus pneumoniae, Haemophilus influenzae, Mycoplasma pneumoniae, and Chlamydophila pneumoniae were applied to sputum, nasopharyngeal swabs, and nasopharyngeal aspirates from 235 adult patients with community-acquired pneumonia and 113 controls. Both culture and multiplex PCR performed well with the different samples and appear to be useful as diagnostic tools.As an etiologic agent can rarely be identified in more than 50% of patients with community-acquired pneumonia (CAP) (9), further development of diagnostic methods has been encouraged (4, 10).The aim of the present study was to estimate the diagnostic accuracy of respiratory culture and a single-run multiplex PCR (mPCR) for specific genes of Streptococcus pneumoniae (lytA), Haemophilus influenzae (16S rRNA, with verification by PCR for P6), Mycoplasma pneumoniae (P1), and Chlamydophila pneumoniae (ompA) (12) applied to respiratory samples in CAP patients.(The study was approved by the ethics committee of the Ö rebro County Council [868-1999; 556-2000].)In a prospective study described previously (14), 235 hospitalized CAP patients with X-ray infiltrates were enrolled. Their median age was 71 years (age range, 18 to 96 years), 40% belonged to severity risk classes IV and V, 14% had chronic obstructive pulmonary disease, and 22% were smokers. In 82% of the patients, the interval from the onset of illness to admission was 0 to 7 days.During the study period, 113 adult controls (median age, 69 years) without respiratory symptoms were enrolled. They were hospitalized for skin infection (n ϭ 14), urinary tract infection (n ϭ 14), arthritis or spondylitis (n ϭ 6), or planned orthopedic or urological surgery (n ϭ 79).The results of the respiratory cultures and mPCR analyses performed with specimens from the patients and the controls are shown in Table 1. Sputum samples were analyzed if there were more than five leukocytes per squamous epithelial cell (6). To obtain a nasopharyngeal aspirate (NpA), secretions from the nasopharynx were aspirated by a catheter connected to an electronic suction device. About 1 ml NaCl (0.85%) was then aspirated to collect the secretions situated within the catheter. Culture and mPCR of sputum, nasopharyngeal swabs (NpSs), and NpAs were performed as described previously (12). Multiplex PCR was performed blindly with samples previously frozen at Ϫ70°C.To establish the definite etiologies of CAP, we performed the following: blood culture for 235 patients, the NOW S. pneumoniae urinary antigen test (Binax) for 215 patients, the complement fixation test for M. pneumoniae (8) with paired serum specimens for 216 patients, the microimmunofluorescence test for C. pneumoniae (3) with paired serum specimens for 216 patients, and the indirect immunofluorescence test for H. influenzae (13) with paired serum specimens for 48 patients. Our criteria for definite CAP etiologies were as follows: for S. pneumoniae, a positive blood culture or a positive urinary antigen test (only visible result lines at l...

Evaluation of a multiplex PCR for bacterial pathogens applied to bronchoalveolar lavage

“…PCR has previously been shown to identify a high rate of colonisation of H. influenzae in patients with chronic obstructive pulmonary disease [33], but H. influenzae was also identified in 22% by BAL culture and 37% by BAL mPCR in 27 patients who had never smoked. As mPCR has shown a high analytical specificity for H. influenzae [12], and as sputum mPCR was positive for H. influenzae in only two (7.7%) out of 26 CAP patients with definite aetiologies other than H. influenzae in the present authors' previous study [13], the high rate of H. influenzae in the present study probably represents colonisation. Hence, it would be interesting to test mPCR on BAL samples from another LRTI population and another control population with a lower smoking frequency.…”

“…In the present authors' previous study of 235 patients with CAP [13], three patients were microimmunofluorescence-test positive for C. pneumoniae in paired sera; these three patients were also mPCR positive for C. pneumoniae in nasopharyngeal secretions. The fact that no control and only one LRTI patient was BAL mPCR positive in the present study indicates that BAL mPCR is probably specific for C. pneumoniae.…”

“…The method showed an analytical sensitivity of 100% (89 out of 89) and a specificity of 99% (167 out of 168) when 257 bacterial strains (37 different species) were tested [12]. mPCR has been evaluated on sputum samples and nasopharyngeal samples from adults with community-acquired pneumonia (CAP) and controls with promising results [13].…”

Detection of Streptococcus pneumoniae Strain Cocolonization in the Nasopharynx

“…The sensitivity of plyNCR PCR for the detection of pneumococcal colonization was significantly higher than that of conventional culture (50% versus 40%). This corresponds to the results for other PCR-based methods for the detection of pneumococcal colonization directly in nasopharyngeal swabs (15,28).…”

Diagnosis of Streptococcus pneumoniae Infections in Adults with Bacteremia and Community-Acquired Pneumonia: Clinical Comparison of Pneumococcal PCR and Urinary Antigen Detection

“…In general, when performed on sputum samples they have good sensitivity but poor specificity (12,17). In order to overcome the latter problem, PCR assays have been evaluated on blood samples (5,8,14,15), giving excellent specificity but relatively poor sensitivity.…”