Chlamydia pneumoniae : A link between Periodontitis and Cardiovascular Disease? Debra Holtham Department of Biology, York College of Pennsylvania preparation of control and experimental samples Nested PCR of Samples ElectrophoresisAnalyze Result same size products Confirms Hypothesis (1) diff size/ no products Hypothesis (1) Incorrect Literature Cited Haraszthy VI., Zambon, J.J., Trevisan, M., Zeid, M., Genco, R.J. “Identification of Periodontal Pathogens in Atheromaotus Plaques”. Journal of Periodontal October Mattila, Kimmo J., Pussinen, Pirkko J., Paju, Susanna. “Dental Infections and Cardiovascular Diseases: A review”. Journal of Periodontal November N. SINADA*, J.K. SPITZNAGEL, and P. BAVOIL. Detection of Chlamydia pneumoniae in Human Dental Plaque. Abstract #2133, J. Dent. Res. Annual Meeting Abstract Issue. March, 2006 IADR, Orlando, FL Reed, S. G., D. E. Lopatin, B. Foxman, and B. A. Burt Oral Chlamydia trachomatis in patients with established periodontitis. Clin. Oral Investig. 4: Hypothesis (1) confirmed nPCR samplesDNA Sequencing same gene sequence Confirms Hypothesis (2) diff gene sequence Hypothesis (2) Incorrect This study involves genetic methods to discover if one possible linking factor between Chlamydia pneumoniae infection and cardiovascular disease is colonization from oral infection. Chlamydia pneumoniae is a gram negative, obligate intracellular bacterium. It lacks a peptidoglycan layer and was recognized to cause infections in humans as early as Interestingly, this bacterium has been found in gingival infections of patients with gum disease as well as in arterial walls of patients with hardened and clogged arteries in the heart. Periodontal diseases (gum disease), including gingivitis and periodontitis, are serious conditions that, if left untreated, can lead to tooth loss, blood, and bone infections. Periodontitis is a chronic infection that affects the teeth by creating pockets between the tooth, supporting bone, and soft tissue. Cardiovascular disease (CVD) refers to the class of diseases that involve malfunction of the heart and/or blood vessels (arteries and veins) which supply oxygen to all areas of the body. While the term technically refers to any disease that affects the cardiovascular system,, it is usually used to refer to those related to atherosclerosis (hardening and clogging of the arteries), heart attack, and stroke. From this study of Chlamydia pneumoniae from bacterial film samples in the oral cavity and hardened and clogged arterial walls of the arteries, a discovery can be made whether or not these two areas contain the same genotypic Chlamydia pneumoniae. Research Design Part 1 Research Design Part 2 Introduction Project Summary Chlamydia pnuemoniae is a bacteria found in patients with periodontal and cardiovascular disease. Periodontal disease exists from bacterial infection within gums and cardiovascular diseases range from hardened arterial walls to build up of “plaque” within heart walls. The objective is to determine whether these two regions, oral and cardiovascular, in fact contain the same genotypic Chlamydia pneumoniae that is found since we already know it exists in each region. Methods involved are polymerase chain reaction (PCR) to amplify the sample from each designated area followed by electrophoresis to determine if any common base pair regions exist between the samples and the Chlamydia pneumoniae control. Gene typing will determine if the genes are in fact identical. I suspect to find both regions to contain Chlamydia pneumoniae. Acknowledgements A short and sweet “Thank you” to Dr. Spitznagel for the stepping stone and Dr. Thompson for the helpful hints during Senior Thesis 1. nPCR is successful with tissue samples and Chlamydia pneumoniae is detected 2. Chlamydia pneumoniae has the same genotype in the oral and cardiovascular samples 1. Determine if nested Polymerase Chain Reaction system (nPCR) is as reliable for testing Chlamydia pneumonie in tissue samples as for dental plaque 2. Verify that the genotype of Chlamydia pneumoniae in dental plaque and atherosclerotic lesion are the same Chlamydia pneumoniae has been identified in aortic, coronary, carotid, iliac and femoral atherosclerotic lesions by various methods including PCR, electron microscopy, microbial culture and immunohistochemistry. Antigen traces have been found specifically in atherosclerotic lesions using various immunological assays such as Enzyme-Linked Immunosorbent Assay (ELISA) and Direct Fluorescent Antibody Test (DFA) and DNA traces have been detected by means of Polymerase Chain Reaction (PCR). Seroepidemiological association of C. pneumoniae with atherosclerosis demonstrated that patients with coronary heart disease(CHD) and acute myocardial infarction(AMI) had C. pneumoniae antibodies more often than the control population. Since then, other seroepidemiological studies have confirmed these findings, with IgA antibody being used as the marker. Chlamydia pneumoniae, periodontal pathogens, cause arthrosclerosis in experimental animals and have been found in human atherosclerotic lesions. C. pneumoniae is capable of producing inflammatory atherosclerosis-like changing in the aortas of infected rabbits. The prevalence of cardiovascular disease (CVD) seems to be highest in those individuals in whom periodontitis coexists. Patients with periodontal disease are, on average, at twice the risk for coronary vascular disease including myocardial infarction. Indicates that periodontitis is a CVD risk factor for individuals who react to an infection with a systemic inflammatory and immune response. Nested PCR (nPCR) amplification with 2 different primers increases both sensitivity and specificity which is fundamental for detection of unique Chlamydia pneumoniae base pairs. Results from this study specifically state that 80% of the 50 endarterectomy specimens were + in one or more PCR assays with 18% + for C.pneumoniae Haraszthy VI., Zambon, J.J., Trevisan, M., Zeid, M., Genco, R.J. “Identification of Periodontal Pathogens in Atheromaotus Plaques”. Journal of Periodontal October Objectives Review of Literature Expected Results Photo taken from (