Analysis of novel genotypes of Mfa1 fimbriae in Porphyromonas gingivalis

Abstract: Objectives: Porphyromonas gingivalis, a periodontopathogenic gram-negative obligate anaerobic bacterium, generally expresses two types of fimbriae, FimA and Mfa1. Whereas FimA fimbriae are known to have I–V genotypes, the genotypes of Mfa1 fimbriae have not been fully clarified. In recent years, we have analyzed the genotypes of mfa1 in 84 strains from our laboratory stock and have shown that mfa1 has at least two different genotypes, namely mfa⁷⁰ and mfa⁵³. Through a series of analyses, we identified eight strains that could not be classified as either mfa⁷⁰ or mfa⁵³. In the present study, we performed whole-genome sequencing to classify the genotypes of mfa1 in these eight strains. Methods: Genomic DNA was extracted from the P. gingivalis strains 222, 1436, 1439, B158, EM3, JKG9, JKG10, and Kyudai-3, and sequenced using the Illumina NovaSeq PE150 system. After the raw sequences were trimmed and filtered for quality, the remaining reads were assembled de novo using BioBam OmicsBox. Based on the similarity of nucleotide and amino acid sequences, the protein-coding sequences of the strains were compared between the draft genomes using ClustalW. Results: Mfa1 fimbriae were composed of five proteins expressed from the mfa cluster (mfa1–mfa5) in the type strain of ATCC 33277; however, it was found that the von Willebrand factor-containing protein (VWA) gene corresponding to mfa5 was present in tandem in the EM3 and JKG10 strains. Notably, VWA was presented alone without mfa1–mfa4 in the 222 strain. It was also found that only mfa1 exists independently in the Kyudai-3 strain. Conclusions: We found the diversity of the mfa cluster among P. gingivalis strains. The genotyping of Mfa1 fimbriae requires sequence analysis to determine the presence or absence of the whole mfa cluster, including mfa1–mfa4 and VWA combination.