Background Molecular microbial techniques have identified several DNA sequences from oral and gut bacteria in atherosclerotic plaques. The composition of the plaque microbiome has shown great heterogeneity due to varying DNA extraction and amplification methods, small size sample cohorts, and a low abundance of bacteria in plaques, leading to interference by contaminating bacterial sequences from laboratory chemicals. We used a novel next-generation sequencing (NGS) approach to analyze the entirety of bacterial DNAs found in coronary artery plaques in a unique prospective autopsy series of out-of-hospital deaths representing a cross-section of the general population.

Methods DNA was extracted from aseptically collected and frozen (−80 °C) atherosclerotic coronary plaque samples taken from the left anterior descending (LAD) coronary arteries of 202 victims of sudden out-of-hospital death included in the Tampere Sudden Death Study. Bacterial DNA was amplified using nested 16S rRNA PCR and sequenced. Amplicon sequence variants (ASVs) were inferred using the DADA2 approach. Taxonomy assignment of the sequences was accomplished using the Silva database (version 138).The presence of bacteria in coronary artery atherosclerotic plaques was confirmed by immunohistochemistry.

Results ASVs from 230 bacteria at the genus level occurring in humans were detected in the coronary plaques. The most common ASVs detected in almost all plaques belonged to the oral biofilm-producing bacteria Veillonella and Streptococcus, which, along with Prevotella, Lactobacillus, Clostridium, and Fusobacterium were the genera with the most ASV reads following nested PCR. The immunopositivity of oral streptococci and Veillonella occurred mainly as biofilm-like structures around calcific plaque areas and associated with the severity of atherosclerosis.

Conclusion Coronary plaques harbor DNA sequences from dozens of mainly oral bacteria. However, only a few bacteria are so common that they likely have a role in the buildup of the microbiome inside a coronary artery atheroma.

The authors have declared no competing interest.

The Tampere Sudden Death Study (TSDS) was financially supported by EU's 7th Framework Programme (grant no. 201668 for AtheroRemo); the Jane and Aatos Erkko Foundation; the Tampere University Foundation; the Tampere University Hospital Medical Funds; the Finnish Foundation of Cardiovascular Research (K.S., P.K.,T.L. and V.H.); the Pirkanmaa Regional Fund of the Finnish Cultural Foundation (K.S.); the Yrjö Jahnsson Foundation; the Juho Vainio Foundation; the Sigrid Juselius Foundation (V.H.) the Tampere Tuberculosis Foundation; EU Horizon 2020 (grant 755320 for TAXINOMISIS and grant 848146 for To Aition); the Academy of Finland (grants 322098 and 356405 for T.L., 331946 for V.H.); the Tampere University Hospital Supporting Foundation; and the Finnish Society of Clinical Chemistry. Pashupati P. Mishra was supported by the Academy of Finland (grant number 349708). We acknowledge Biocenter Finland for infrastructure support.

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The TSDS protocol was approved by the Ethics Committee of the Pirkanmaa Hospital District (permission number R09097) and the National Supervisory Authority for Welfare and Health.

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