Investigation of bacterial etiology with conventional and multiplex PCR methods in adult patients with community-acquired pneumonia

Community-acquired pneumonia (CAP) is still a serious life-threatening disease, in which the etiologic agent cannot be identified in more than 50% of patients despite advanced diagnostic methods. The most commonly used methods in the determination of CAP etiology are culture and serological tests. Since early and accurate therapy reduces the mortality in CAP cases, rapid and reliable diagnostic methods are needed. The aim of this study was to determine the bacterial etiology in adult patients with CAP by implementing multiplex polymerase chain reaction/reverse line blot hybridization (M-PCR/RLBH) assay combined with conventional methods. A total of 128 cases (94 were male; age range: 19-81 years, mean age: 58) who were admitted to our hospital and clinically diagnosed as CAP between November 2008 - November 2010, were included in the study. Respiratory samples (sputum and/or bronchoalveolar lavage) obtained from patients were searched by M-PCR/RLBH method (Gen ID®, Autoimmun Diagnostika GmbH, Germany) in terms of the presence of Streptococcus pneumoniae, Haemophilus influenzae, Moraxella catarrhalis, Mycoplasma pneumoniae, Chlamydia pneumoniae and Legionella pneumophila nucleic acids. The samples were simultaneously inoculated onto 5% sheep blood agar, chocolate agar, haemophilus isolation agar, buffered charcoal yeast extract-selective agar and EMB agar media for cultivation. Serum samples obtained from the cases were tested for IgM and IgG antibodies against C.pneumoniae by microimmunofluorescence (Focus Diagnostic, USA) and against L.pneumophila and M.pneumoniae by indirect immunofluorescence (Euroimmun, Germany) methods. The bacterial etiology was identified in 59 (46.1%) of 128 patients with CAP and a total of 73 pathogens were detected. The leading organism was S.pneumoniae (n= 32, 25%), followed by H.influenzae and M.pneumoniae (n= 9, 7%), gram-negative bacilli (n= 10, 7.8%), M.catarrhalis (n= 6, 4.7%), C.pneumoniae (n= 4, 3.2%), L.pneumophila (n= 2, 1.6%) and Staphylococcus aureus (n= 1, 1.4%). Infection with atypical pathogens were detected in 15 (11.7%), and mixed infections in 14 (10.9%) patients. The detection rate of microorganisms (S.pneumoniae, H.influenzae, M.catarrhalis, C.pneumoniae, L.pneumophilia, M.pneumoniae) searched by M-PCR/RLBH method was 41.4% (53/128), while those microorganisms were detected in 23.4% (30/128) of the patients by conventional methods, representing a significant difference (p< 0.05). It was concluded that M-PCR/RL