Absence of Bacteria on Coronary Angioplasty Balloons from Unselected Patients: Results with Use of a High Sensitivity Polymerase Chain Reaction Assay

Abstract

Periodontitis is a chronic, bacterially-induced inflammatory disease of the tooth-supporting tissues, which may result in transient bacteremia and a systemic inflammatory response. Periodontitis is associated with coronary artery disease independently of established cardiovascular risk factors, and translocation of bacteria from the oral cavity to the coronary arteries may play a role in the development of coronary artery disease. Very few studies have used angioplasty balloons for in vivo sampling from diseased coronary arteries, and with varying results. Therefore, the aim of this study was to assess if bacterial DNA from primarily oral bacteria could be detected on coronary angioplasty balloons by use of an optimized sampling process combined with an internally validated sensitive polymerase chain reaction (PCR) assay. Coronary angioplasty balloons and control samples from a total of 45 unselected patients with stable angina, unstable angina/non-ST elevation myocardial infarction, and ST-elevation myocardial infarction (n = 15 in each group) were collected and analyzed using a PCR assay with high sensitivity and specificity for 16S rRNA genes of the oral microbiome. Despite elimination of extraction and purification steps, and demonstration of sensitivity levels of 25-125 colony forming units (CFU), we did not detect bacterial DNA from any of the coronary angioplasty balloons. A subsequent questionnaire indicated that the prevalence of periodontitis in the study cohort was at least 39.5%. Although coronary angioplasty balloons are unlikely to be useful for detection of bacteria with current PCR techniques in unselected patients with coronary artery disease, more studies are warranted to determine the extent to which bacteria contribute to atherosclerosis and its clinical manifestations and whether the presence of bacteria in the arteries is a transient phenomenon.

Fig 1. Photograph of agarose gel from one of the polymerase chain reaction (PCR) assay validation experiments.

(A) Results from PCR performed on a 10-fold dilution row of 5 μL human blood from healthy volunteers spiked with P. aeruginosa in concentrations ranging from 1.25x10⁵-1.25x10 colony forming units (CFU)/5 μL. Detection limit was 125 CFU. (B) Results from PCR performed directly on angioplasty balloons dilated and deflated while dipped in the same 10-fold dilution row of blood with P. aeruginosa. Detection limit was 25–125 CFU depending on the amount of blood adhering to the balloon, which in repeated experiments varied from 1–5 μL. Blood samples without added bacteria served as negative controls.

Fig 2. Transverse section of a deflated angioplasty balloon after percutaneous coronary intervention.

Close-up photograph shows the folds and grooves that potentially may contain atherosclerotic tissue material from the site of coronary lesion dilatation.

Fig 3. Photograph of agarose gel with polymerase chain reaction (PCR) results from six patient samples and controls.

Patient id 1.10, 1.12, 1.13, and 1.14: stable angina; patient id 2.09 and 2.10: unstable angina/non ST-elevation myocardial infarction. Lane a) angioplasty balloon, lane b) catheter control segment, lane c) arterial blood. Positive controls were arterial blood spiked with 10³−10⁴ colony-forming units (CFU) P. aeruginosa. Negative controls were PCR master mix run without template.