Fim3-dependent autoagglutination of Bordetella pertussis

Autoagglutination (Agg) of Bordetella pertussis is often observed in clinical laboratory. However, its causal factors and frequency in circulating strains are unknown. Repeated single colony isolation enabled us to detect an Agg^- mutant in the supernatant of an Agg⁺ strain of B. pertussis. Whole-genome sequencing and immunoblot analysis disclosed that the Agg^- mutant had a single C-deletion in its fim3 promoter region (Pfim3) which abolished Fim3 fimbriae production. A B. pertussis fim3-knock out mutant also lacked the Agg⁺ phenotype. Agg⁺ clinical isolates were detected a higher production of Fim3 than Fim3-producing Agg^- isolates. B. pertussis is known to harbor multiple Pfim3 poly(C) lengths within a single strain culture and our newly developed PCR/LDR assay revealed that Agg⁺ isolates harbor the highest Pfim3 poly-14C abundance. We evaluated the frequency of autoagglutination in clinical B. pertussis isolates collected in Japan between 1994 and 2018 (n = 203). Fim3 production was confirmed for 190 isolates and 74.7% of them displayed the Agg⁺ phenotype. The Agg⁺ phenotype was strongly associated with Pfim3 poly-14C abundance. Taken together, our findings demonstrated that B. pertussis autoagglutination occurs in response to high Fim3 levels and the Agg⁺ strain has predominated in Japan over the past two decades.