Atypical bacterial etiology of acute respiratory infections and clinical characterizations among Iranian children.

Acute respiratory infections (ARIs) in children younger than 5 years of age are one of the leading causes of morbidity and mortality, particularly in developing countries. Mycoplasma pneumoniae and Chlamydophila pneumoniae are prevalent causative agents of ARIs, worldwide. We sought M. pneumoniae and C. pneumoniae in respiratory samples from Iranian children with ARIs.  From November 2014 to April 2015, respiratory samples of 150 children aged 1 month to 15 years old were screened for presence of M. pneumoniae and C. pneumoniae. Polymerase chain reaction (PCR) and culture methods were used to detect these bacteria in respiratory samples in the form of throat swabs and nasopharyngeal aspirates. A questionnaire containing demographic and clinical information has been filled up for all participants in this study. Our obtained data showed that out of 150 tested samples, 7 (4.7%) were PCR positive for M. pneumoniae and only one (0.7%) positive sample for C. pneumoniae was detected. However, none of the tested samples was detected M. pneumoniae using the bacterial culture method. All patients with ARIs due to M. pneumoniae showed up with sore throat and flu like symptoms. According to our data, PCR method is more sensitive than culture for detection of M. pneumoniae. With regards to our results, it appears that M. pneumoniae and especially C. pneumoniae were infrequent causative agents in our studied population.

Molecular characterisation of Chlamydia pneumoniae associated to atherosclerosis.

Chlamydia pneumoniae is a respiratory pathogen associated with chronic inflammatory diseases such as asthma and atherosclerosis, and its detection in human carotid and coronary atheroma suggests some support for its involvement in atherogenesis. The main objective of our study was to evaluate the association between Chlamydia pneumoniae and atherosclerosis in Moroccan patients through a case-control approach and detected strain genotyping. A total of 137 cases and 124 controls were enrolled, nested PCR was performed for Chlamydia pneumoniae screening of the peripheral blood mononuclear cells (PBMCs) of both cases and controls as well as atheroma plaques from 37 cases, and positive samples were subjected to sequencing for genotyping and phylogenetic analysis. The results showed 54% and 18%, respectively, for positivity in cases and control PBMCs and 86.5% in atheroma plaques, the difference being significant between the two groups (P < 0.001, ORa = 8.580, CI, 95% [3.273-22.491]). Strain sequence analyses showed more than 98% similarity with human reference strains, and revealed various genotypes. This study supports the involvement of Chlamydia pneumoniae in atherosclerosis in the studied population and genotyping revealed that detected strains were identical to human strains circulating worldwide.

Chlamydia pneumoniae Clinical Isolate from Gingival Crevicular Fluid: A Potential Atherogenic Strain.

Chlamydia pneumoniae has been associated to atherosclerotic cardiovascular diseases. The aim of our study was to characterize, for the first time, a C. pneumoniae strain isolated from the gingival crevicular fluid of a patient with chronic periodontitis, described as a risk factor for cardiovascular diseases. C. pneumoniae isolate was characterized and compared to the respiratory AR-39 strain by VD4-ompA genotyping and by investigating the intracellular growth in epithelial and macrophage cell lines and its ability to induce macrophage-derived foam cells. Inflammatory cytokine levels were determined in the gingival crevicular fluid sample. C. pneumoniae isolate showed a 99% similarity with the AR-39 strain in the VD4-ompA gene sequence and shared a comparable growth kinetic in epithelial cells and macrophages, as evidenced by the infectious progeny and by the number of chlamydial genomic copies. C. pneumoniae isolate significantly increased the number of foam cells as compared to uninfected and LDL-treated macrophages (45 vs. 6%, P = 0.0065) and to the AR-39 strain (45 vs. 30%, P = 0.0065). Significantly increased levels of interleukin 1-ß (2.1 ± 0.3 pg/µL) and interleukin 6 (0.6 ± 0.08 pg/µL) were found. Our results suggest that C. pneumoniae may harbor inside oral cavity and potentially be atherogenic, even though further studies will be needed to clarify the involvement of C. pneumoniae in chronic periodontitis as a risk factor for cardiovascular diseases.

Phylogenetic analysis of human Chlamydia pneumoniae strains reveals a distinct Australian indigenous clade that predates European exploration of the continent.

BACKGROUND: The obligate intracellular bacterium Chlamydia pneumoniae is a common respiratory pathogen, which has been found in a range of hosts including humans, marsupials and amphibians. Whole genome comparisons of human C. pneumoniae have previously highlighted a highly conserved nucleotide sequence, with minor but key polymorphisms and additional coding capacity when human and animal strains are compared. RESULTS: In this study, we sequenced three Australian human C. pneumoniae strains, two of which were isolated from patients in remote indigenous communities, and compared them to all available C. pneumoniae genomes. Our study demonstrated a phylogenetically distinct human C. pneumoniae clade containing the two indigenous Australian strains, with estimates that the most recent common ancestor of these strains predates the arrival of European settlers to Australia. We describe several polymorphisms characteristic to these strains, some of which are similar in sequence to animal C. pneumoniae strains, as well as evidence to suggest that several recombination events have shaped these distinct strains. CONCLUSIONS: Our study reveals a greater sequence diversity amongst both human and animal C. pneumoniae strains, and suggests that a wider range of strains may be circulating in the human population than current sampling indicates.

Comparative analysis of the growth and biological activity of a respiratory and atheroma isolate of Chlamydia pneumoniae reveals strain-dependent differences in inflammatory activity and innate immune evasion.

BACKGROUND: Chlamydia pneumoniae is a common human pathogen that is associated with upper and lower respiratory tract infections. It has also been suggested that C. pneumoniae infection can trigger or promote a number of chronic inflammatory conditions, including asthma and atherosclerosis. Several strains of C. pneumoniae have been isolated from humans and animals, and sequence data demonstrates marked genetic conservation, leaving unanswered the question as to why chronic inflammatory conditions may occur following some respiratory-acquired infections. METHODS: C. pneumoniae strains AR39 and AO3 were used in vitro to infect murine bone marrow derived macrophages and L929 fibroblasts, or in vivo to infect C57BL/6 mice via the intranasal route. RESULTS: We undertook a comparative study of a respiratory isolate, AR39, and an atheroma isolate, AO3, to determine if bacterial growth and host responses to infection varied between these two strains. We observed differential growth depending on the host cell type and the growth temperature; however both strains were capable of forming plaques in vitro. The host response to the respiratory isolate was found to be more inflammatory both in vitro, in terms of inflammatory cytokine induction, and in vivo, as measured by clinical response and lung inflammatory markers using a mouse model of respiratory infection. CONCLUSIONS: Our data demonstrates that a subset of C. pneumoniae strains is capable of evading host innate immune defenses during the acute respiratory infection. Further studies on the genetic basis for these differences on both the host and pathogen side could enhance our understanding how C. pneumoniae contributes to the development chronic inflammation at local and distant sites.

Comparative genomic analysis of human Chlamydia pneumoniae isolates from respiratory, brain and cardiac tissues.

Chlamydia pneumoniae is an obligate intracellular bacterium implicated in a wide range of human diseases including atherosclerosis and Alzheimer's disease. Efforts to understand the relationships between C. pneumoniae detected in these diseases have been hindered by the availability of sequence data for non-respiratory strains. In this study, we sequenced the whole genomes for C. pneumoniae isolates from atherosclerosis and Alzheimer's disease, and compared these to previously published C. pneumoniae genomes. Phylogenetic analyses of these new C. pneumoniae strains indicate two sub-groups within human C. pneumoniae, and suggest that both recombination and mutation events have driven the evolution of human C. pneumoniae. Further fine-detailed analyses of these new C. pneumoniae sequences show several genetically variable loci. This suggests that similar strains of C. pneumoniae are found in the brain, lungs and cardiovascular system and that only minor genetic differences may contribute to the adaptation of particular strains in human disease.

Prevalence and characterization of Chlamydia DNA in zoo animals in Japan.

Because many people visit zoos, prevention of zoonoses is important from the standpoint of public health. This study examined the prevalence of Chlamydia among zoo animals in Japan by PCR and characterized these bacteria by performing phylogenetic analyses of the sequences of the variable domain (VD) 2 and VD4 regions of the ompA gene, which encodes the Chlamydia major outer membrane protein. Fecal samples were collected from 1150 zoo animals in five zoos and examined for Chlamydia DNA. Chlamydia psittaci DNA was found in 3.9% of mammals, 7.2% of birds and 8.1% of reptiles. The prevalence of Chlamydia pneumoniae DNA was significantly higher in reptiles (5.8%) than in mammals (0.3%) and birds (0.3%). Phylogenetic analysis of the ompA VD2 region from 18 samples showed that nine were in three different clusters containing C. psittaci strains, six were in a cluster containing C. pneumoniae strains and three each formed a distinct branch. Furthermore, phylogenetic analysis of the ompA VD4 region showed that C. pneumoniae DNAs from reptiles were close to those from human patients. The C. pneumoniae DNAs from the European glass lizard, Emerald tree boa, and Panther chameleon were classified in clusters that were distinct from other strains, suggesting that these reptiles had each been infected with a specific C. pneumoniae genotype. This study showed that diverse Chlamydia strains have been prevalent among a variety of zoo animals.

Pneumonia Outbreak Caused by Chlamydophila pneumoniae among US Air Force Academy Cadets, Colorado, USA.

During October 2013-May 2014, there were 102 cases of pneumonia diagnosed in US Air Force Academy cadets. A total of 73% of tested nasal washes contained Chlamydophila pneumoniae. This agent can be considered to be present on campus settings during outbreaks with numerous, seemingly disconnected cases of relatively mild pneumonia.

Evolution to a chronic disease niche correlates with increased sensitivity to tryptophan availability for the obligate intracellular bacterium Chlamydia pneumoniae.

The chlamydiae are obligate intracellular parasites that have evolved specific interactions with their various hosts and host cell types to ensure their successful survival and consequential pathogenesis. The species Chlamydia pneumoniae is ubiquitous, with serological studies showing that most humans are infected at some stage in their lifetime. While most human infections are asymptomatic, C. pneumoniae can cause more-severe respiratory disease and pneumonia and has been linked to chronic diseases such as asthma, atherosclerosis, and even Alzheimer's disease. The widely dispersed animal-adapted C. pneumoniae strains cause an equally wide range of diseases in their hosts. It is emerging that the ability of C. pneumoniae to survive inside its target cells, including evasion of the host's immune attack mechanisms, is linked to the acquisition of key metabolites. Tryptophan and arginine are key checkpoint compounds in this host-parasite battle. Interestingly, the animal strains of C. pneumoniae have a slightly larger genome, enabling them to cope better with metabolite restrictions. It therefore appears that as the evolutionarily more ancient animal strains have evolved to infect humans, they have selectively become more "susceptible" to the levels of key metabolites, such as tryptophan. While this might initially appear to be a weakness, it allows these human C. pneumoniae strains to exquisitely sense host immune attack and respond by rapidly reverting to a persistent phase. During persistence, they reduce their metabolic levels, halting progression of their developmental cycle, waiting until the hostile external conditions have passed before they reemerge.

Chlamydia pneumoniae: modern insights into an ancient pathogen.

Chlamydia pneumoniae is an enigmatic human and animal pathogen. Originally discovered in association with acute human respiratory disease, it is now associated with a remarkably wide range of chronic diseases as well as having a cosmopolitan distribution within the animal kingdom. Molecular typing studies suggest that animal strains are ancestral to human strains and that C. pneumoniae crossed from animals to humans as the result of at least one relatively recent zoonotic event. Whole genome analyses appear to support this concept - the human strains are highly conserved whereas the single animal strain that has been fully sequenced has a larger genome with several notable differences. When compared to the other, better known chlamydial species that is implicated in human infection, Chlamydia trachomatis, C. pneumoniae demonstrates pertinent differences in its cell biology, development, and genome structure. Here, we examine the characteristic facets of C. pneumoniae biology, offering insights into the diversity and evolution of this silent and ancient pathogen.

[The phylogenetic position of Chlamydia strains isolated from monkeys and humans with Chlamydial pathology in the family Chiamydiaceae. Genotypic and phenotypic properties of this pathogen].

Based on the results of the comparative analysis concerning relatedness and evolutional difference of the 16S - 23S nucleotide sequences of the middle ribosomal cluster and 23S rRNA I domain, and based on identification of phylogenetic position for Chlamydophila pneumoniae and Chlamydia trichomatis strains released from monkeys, relatedness of the above stated isolates with similar strains released from humans and with strains having nucleotide sequences presented in the GenBank electronic database has been detected for the first time ever. Position of these isolates in the Chlamydiaceae family phylogenetic tree has been identified. The evolutional position of the investigated original Chlamydia and Chlamydophila strains close to analogous strains from the GenBank electronic database has been demonstrated. Differences in the 16S - 23S nucleotide sequence of the middle ribosomal cluster and 23S rRNA I domain of plasmid and non-plasmid Chlamydia trachomatis strains released from humans and monkeys relative to different genotype groups (group B- B, Ba, D, Da, E, L1, L2, L2a; intermediate group - F, G, Ga) have been revealed for the first time ever. Abnormality in incA chromosomal gene expression resulting in Chlamydia life and development cycle disorder and decrease of Chlamydia virulence can be related to probable changes in the nucleotide sequence of the gene under consideration.

A graph-based algorithm for mining multi-level patterns in genomic data.

Comparative genomics is concerned with the study of genome structure and function of different species. It can provide useful information for the derivation of evolutionary and functional relationships between genomes. Previous work on genome comparison focuses mainly on comparing the entire genomes for visualization without further analysis. As many interesting patterns may exist between genomes and may lead to the discovering of functional gene segments (groups of genes), we propose an algorithm called Multi-Level Genome Comparison Algorithm (MGC) that can be used to facilitate the analysis of genomes at multi-levels during the comparison process to discover sequential and regional consistency in gene segments. Different genomes may have common sub-sequences that differ from each other due to mutations, lateral gene transfers, gene rearrangements, etc., and these sub-sequences are usually not easily identified. Not all the genes can have a perfect one-to-one matching with each other. It is quite possible for one-to-many or many-to-many ambiguous relationships to exist between them. To perform the tasks effectively, MGC takes such ambiguity into consideration during genome comparison by representing genomes in a graph and then make use of a graph mining algorithm called the Multi-Level Attributed Graph Mining Algorithm (MAGMA) to build a hierarchical multi-level graph structure to facilitate genome comparison. To determine the effectiveness of these proposed algorithms, experiments were performed using intra- and inter-species of Microbial genomes. The results show that the proposed algorithms are able to discover multiple level matching patterns that show the similarities and dissimilarities among different genomes, in addition to confirming the specific role of the genes in the genomes.

Isolation of Chlamydia pneumoniae from serum samples of the patients with acute coronary syndrome.

BACKGROUND: Limited body of evidence suggests that lipopolysaccharide of C. pneumoniae as well as C. pneumoniae-specific immune complexes can be detected and isolated from human serum. The aim of this study was to investigate the presence of viable elementary bodies of C.pneumoniae in serum samples of patients with acute coronary syndrome and healthy volunteers. MATERIAL AND METHODS: Serum specimens from 26 healthy volunteers and 56 patients with acute coronary syndrome were examined subsequently by serological (C.pneumoniae-specific IgA and IgG), PCR-based and bacteriological methods. Conventional, nested and TaqMan PCR were used to detect C.pneumoniae genetic markers (ompA and 16S rRNA) in DNA from serum specimens extracted with different methods. An alternative protocol which included culturing high-speed serum sediments in HL cells and further C.pneumoniae growth evaluation with immunofluorescence analysis and TaqMan PCR was established. Pellet fraction of PCR-positive serum specimens was also examined by immunoelectron microscopy. RESULTS: Best efficiency of final PCR product recovery from serum specimens has been shown with specific C. pneumoniae primers using phenol-chloroform DNA extraction protocol. TaqMan PCR analysis revealed that human serum of patients with acute coronary syndrome may contain genetic markers of C. pneumoniae with bacterial load range from 200 to 2000 copies/ml serum. However, reliability and reproducibility of TaqMan PCR were poor for serum specimens with low bacterial copy number (<200 /ml). Combination of bacteriological, immunofluorescence and PCR- based protocols applied for the evaluating HL cells infected with serum sediments revealed that 21.0 % of the patients with acute coronary syndrome have viable forms C.pneumoniae in serum. The detection rate of C.pneumoniae in healthy volunteers was much lower (7.7%). Immunological profile of the patients did not match accurately C.pneumoniae detection rate in serum specimens. Elementary bodies of C.pneumoniae with typical ultrastructural characteristics were also identified in serum sediments using immunoelectron microscopy. CONCLUSIONS: Viable forms C. pneumoniae with typical electron microscopic structure can be identified and isolated from serum specimens of the patients with acute coronary syndrome and some healthy volunteers. Increased detection rate of C. pneumoniae in serum among the patients with an acute coronary syndrome may contribute towards enhanced pro-inflammatory status in cardiovascular patients and development of secondary complications of atherosclerosis.

Identification of phylogenetic position in the Chlamydiaceae family for Chlamydia strains released from monkeys and humans with chlamydial pathology.

Based on the results of the comparative analysis concerning relatedness and evolutional difference of the 16S-23S nucleotide sequences of the middle ribosomal cluster and 23S rRNA I domain, and based on identification of phylogenetic position for Chlamydophila pneumoniae and Chlamydia trichomatis strains released from monkeys, relatedness of the above stated isolates with similar strains released from humans and with strains having nucleotide sequences presented in the GenBank electronic database has been detected for the first time ever. Position of these isolates in the Chlamydiaceae family phylogenetic tree has been identified. The evolutional position of the investigated original Chlamydia and Chlamydophila strains close to analogous strains from the Gen-Bank electronic database has been demonstrated. Differences in the 16S-23S nucleotide sequence of the middle ribosomal cluster and 23S rRNA I domain of plasmid and nonplasmid Chlamydia trachomatis strains released from humans and monkeys relative to different genotype groups (group B-B, Ba, D, Da, E, L1, L2, L2a; intermediate group-F, G, Ga) have been revealed for the first time ever. Abnormality in incA chromosomal gene expression resulting in Chlamydia life development cycle disorder, and decrease of Chlamydia virulence can be related to probable changes in the nucleotide sequence of the gene under consideration.

Evidence that human Chlamydia pneumoniae was zoonotically acquired.

Zoonotic infections are a growing threat to global health. Chlamydia pneumoniae is a major human pathogen that is widespread in human populations, causing acute respiratory disease, and has been associated with chronic disease. C. pneumoniae was first identified solely in human populations; however, its host range now includes other mammals, marsupials, amphibians, and reptiles. Australian koalas (Phascolarctos cinereus) are widely infected with two species of Chlamydia, C. pecorum and C. pneumoniae. Transmission of C. pneumoniae between animals and humans has not been reported; however, two other chlamydial species, C. psittaci and C. abortus, are known zoonotic pathogens. We have sequenced the 1,241,024-bp chromosome and a 7.5-kb cryptic chlamydial plasmid of the koala strain of C. pneumoniae (LPCoLN) using the whole-genome shotgun method. Comparative genomic analysis, including pseudogene and single-nucleotide polymorphism (SNP) distribution, and phylogenetic analysis of conserved genes and SNPs against the human isolates of C. pneumoniae show that the LPCoLN isolate is basal to human isolates. Thus, we propose based on compelling genomic and phylogenetic evidence that humans were originally infected zoonotically by an animal isolate(s) of C. pneumoniae which adapted to humans primarily through the processes of gene decay and plasmid loss, to the point where the animal reservoir is no longer required for transmission.

[Chlamydophila pneumoniae: from its proteomics to arteriosclerosis]. / Chlamydophila pneumoniae: desde su proteómica hasta la arteriosclerosis.

Chlamydophila pneumoniae is a highly prevalent intracellular human pathogen with a unique biphasic life cycle. It is a common cause of upper respiratory infection and pneumonia, and is currently being studied as a potential risk factor for the development of atherosclerotic cardiovascular disease. The outer membrane surface antigens of C. pneumoniae are highly complex: some, such as the major outer membrane protein, are specific, but poorly immunodominant, whereas others have stronger immunogenicity, but are cross-reactive among Chlamydia species. Therefore, new, highly immunodominant, species-specific antigens should be sought. In this regard, the polymorphic membrane proteins (PMPs) are a) unique to Chlamydiae, b) often exposed on the surface of the bacteria, and c) highly immunogenic; these factors make them potential candidates for application in laboratory assays. Other chlamydial antigens, such as heat shock protein (HSP) 60, have been associated with atherosclerotic lesions because of their ability to induce an immunological attack on the endothelial wall. Over the last decade, several studies have suggested a potential role of chronic C. pneumoniae infection in human atherosclerosis. Nevertheless, prospective studies with sufficiently large samples and a healthy comparison group, using a combination of direct and indirect microbiological techniques in the same subject and sample, are needed to establish a relationship between the infection and disease activity.

Multi locus sequence typing of Chlamydiales: clonal groupings within the obligate intracellular bacteria Chlamydia trachomatis.

BACKGROUND: The obligate intracellular growing bacterium Chlamydia trachomatis causes diseases like trachoma, urogenital infection and lymphogranuloma venereum with severe morbidity. Several serovars and genotypes have been identified, but these could not be linked to clinical disease or outcome. The related Chlamydophila pneumoniae, of which no subtypes are recognized, causes respiratory infections worldwide. We developed a multi locus sequence typing (MLST) scheme to understand the population genetic structure and diversity of these species and to evaluate the association between genotype and disease. RESULTS: A collection of 26 strains of C. trachomatis of different serovars and clinical presentation and 18 strains of C. pneumoniae were included in the study. For comparison, sequences of C. abortus, C. psittaci, C. caviae, C. felis, C. pecorum (Chlamydophila), C. muridarum (Chlamydia) and of Candidatus protochlamydia and Simkania negevensis were also included. Sequences of fragments (400 - 500 base pairs) from seven housekeeping genes (enoA, fumC, gatA, gidA, hemN, hlfX, oppA) were analysed. Analysis of allelic profiles by eBurst revealed three non-overlapping clonal complexes among the C. trachomatis strains, while the C. pneumoniae strains formed a single group. An UPGMA tree produced from the allelic profiles resulted in three groups of sequence types. The LGV strains grouped in a single cluster, while the urogenital strains were distributed over two separated groups, one consisted solely of strains with frequent occurring serovars (E, D and F). The distribution of the different serovars over the three groups was not consistent, suggesting exchange of serovar encoding ompA sequences. In one instance, exchange of fumC sequences between strains of different groups was observed. Cluster analyses of concatenated sequences of the Chlamydophila and Chlamydia species together with those of Candidatus Protochlamydia amoebophila and Simkania negevensis resulted in a tree identical to that obtained with 23S RNA gene sequences. CONCLUSION: These data show that C. trachomatis and C. pneumoniae are highly uniform. The difference in genetic diversity between C. trachomatis and C. pneumoniae is in concordance with a later assimilation to the human host of the latter. Our data supports the taxonomy of the order of Chlamydiales.

Genetic diversity of the obligate intracellular bacterium Chlamydophila pneumoniae by genome-wide analysis of single nucleotide polymorphisms: evidence for highly clonal population structure.

BACKGROUND: Chlamydophila pneumoniae is an obligate intracellular bacterium that replicates in a biphasic life cycle within eukaryotic host cells. Four published genomes revealed an identity of > 99 %. This remarkable finding raised questions about the existence of distinguishable genotypes in correlation with geographical and anatomical origin. RESULTS: We studied the genetic diversity of C. pneumoniae by analysing synonymous single nucleotide polymorphisms (sSNPs) that are under reduced selection pressure. We conducted an in silico analysis of the four sequenced genomes, chose 232 representative sSNPs and analysed the loci in 38 C. pneumoniae isolates. We identified 15 different genotypes that were separated in four major clusters. Clusters were not associated with anatomical or geographical origin. However, animal lineages are basal on the C. pneumomiae phylogeny, suggesting a recent transmission to humans through successive bottlenecks some 150,000 years ago. A lack of detectable variation in 17 isolates emphasizes the extraordinary genetic conservation of this species and the high clonality of the population. Moreover, the largest cluster, which encompasses 80% of all analysed strains, is an extremely young clade, that went through an important population expansion some 3,300 years ago. CONCLUSION: sSNPs have proven useful as a sensitive marker to gain new insights into genetic diversity, population structure and evolutionary history of C. pneumoniae.

PCR and serology were effective for identifying Chlamydophila pneumoniae in a lower respiratory infection outbreak among military recruits.

During endemic infections, the sensitivity of diagnostic tests and rapid diagnosis of the respiratory tract pathogens is particularly important. Utilization of just one diagnostic technique, such as serological tests or polymerase chain reaction (PCR)-based detection methods, during outbreaks of lower respiratory tract infections (LRI) can result in some of the patients being missed. In this study we aimed to investigate the etiology of LRI in military recruits in Izmir, Turkey, among whom several pneumonia cases have been reported and 47 patients have been hospitalized. Nasopharyngeal swabs were used for PCR analysis of Chlamydophila pneumoniae, Mycoplasma pneumoniae and Legionella spp. Serum samples were collected in the acute and convalescent phase of infection for C. pneumoniae and M. pneumoniae. Thirty-nine patients were diagnosed with C. pneumoniae infection by PCR and/or serology. Diagnoses were established by PCR in the acute phase of infection in 40.4% of the group. Based on the results of these studies, PCR is a useful method for early detection and identification of C. pneumoniae-related LRI outbreaks. However, this technique is not sufficient to detect all positive cases per se. After effective therapy and introduction of appropriate infection control measures, the outbreak ceased without mortality. This is the first closed-community C. pneumoniae outbreak report from Turkey.