Genetic diversity and virulence of Bacillus cereus group isolates from bloodstream infections

Abstract

Bacillus cereus catheter-related bloodstream infections (CRBSIs) are an increasing concern in Japanese hospitals. Although their clinical characteristics have been explored, the genetic relationships and virulence profiles of B. cereus isolates from CRBSIs remain understudied. Here, using advanced genomic techniques, we investigated the genetic diversity, phylogenetic relationships, and virulence profiles of B. cereus isolates from patients with bloodstream infections. We analyzed 28 B. cereus group strains isolated from blood samples at the University of Tokyo Hospital between 2005 and 2017 using whole-genome sequencing, core-genome single-nucleotide polymorphism (SNP) typing, and virulence gene profiling. Core-genome SNP analysis revealed significant genetic diversity among the isolates, suggesting multiple independent sources of infection. The isolates predominantly belonged to panC clades III and IV, with distinct virulence gene profiles. All panC clade III isolates contained hbl operon genes, whereas four isolates from clade IV harbored cereulide synthetase genes (cesABCD). One isolate possessed a capsule gene operon (capBCADE), a rare finding among clinical B. cereus strains. Biofilm formation ability was observed in 50% of catheter-related isolates, although this ability was not significantly different from that of the noncatheter-related isolates.IMPORTANCEThis study provides novel insights into the genetic diversity and virulence potential of B. cereus strains causing bloodstream infections in a Japanese hospital setting. These findings suggest diverse infection pathways and highlight the importance of continuous molecular epidemiological surveillance for effective infection control.

Keywords: Bacillus cereus group; bloodstream infection; epidemiology; whole-genome analysis.

Fig 1

(a) The distribution of biofilm formation among the catheter and noncatheter groups showing the OD_{590 nm} values. ODc = Average OD of the negative control + 3 × SD of the negative control. Each dot represents the average OD value of each strain from triplicate samples in three independent experiments. One-way analysis of variance followed by a Dunnett test revealed a significant difference (P = 0.0355). The average value and range are shown for both groups. (b) Distribution of biofilm-forming ability among B. cereus isolates from catheter-related and noncatheter-related infections. The bar graph shows the number of B. cereus isolates that were biofilm-positive (black) or biofilm-negative (white) for catheter infection (n = 18) and noncatheter infection (n = 10) cases. Among the catheter infection isolates, nine (50%) were biofilm-positive; among the noncatheter infection isolates, two (20%) were biofilm-positive. However, this difference between infection source and biofilm formation ability was not statistically significant (P = 0.22, Fisher’s exact test).

Fig 2

Phylogenetic tree of B. cereus isolates based on core-genome single-nucleotide polymorphism (SNP) analysis The circular phylogenetic tree represents the genetic relationships between B. cereus group isolates, which were classified into seven phylogenetic groups based on panC gene sequencing. The tree is color-coded to indicate panC clades (I–VI), as shown by the outer ring. Isolates are labeled and color-coded as follows: bold italic blue text with dark red highlighting: 28 isolates from the current study; italic blue text with light red highlighting: previously reported isolates from Japan; italic blue text without highlighting, isolates from other East Asian countries; and black text, isolates from other regions or reference strains. The tree scale of 0.1 indicates the genetic distance.

Fig 3

SNP profiles and virulence gene distribution of B. cereus isolates from bloodstream infections The upper panel shows a heatmap of 592 selected SNPs for the 28 B. cereus isolates based on whole-genome sequencing data. Each column represents an isolate, and each row represents a specific SNP position. The colors indicate different nucleotides at each SNP position: blue, red, white, and pink represent the four possible bases (A, T, G, and C, respectively). The lower panel displays the presence (gray boxes) or absence (white boxes) of virulence-related genes in each isolate. Virulence genes are grouped into categories including capsule genes (capA–E), cereulide synthetase genes (cesA–D), hemolysin BL genes (hblA–D), nonhemolytic enterotoxin genes (nheA–C), and other virulence-associated genes (hlyA, hlyII, hlyIII, and plcR).

Fig 4

Genomic organization of the capsule gene operon in B. cereus BCER1 compared with the reference strain ATCC14579. The upper panel shows a section of the B. cereus ATCC14579 reference genome containing the ywqC_2, ywqD_2, fabZ_2, and mbl genes. The lower panel displays the corresponding region in the B. cereus BCER1 genome, where the capsule gene operon is inserted. Locus tags of BCER1_53350 to 54440 are marked. Arrows represent genes and their orientation. Black arrows indicate the capsule genes (capB (pgsB), capC(pgsC), capA(pgsA), capD′, capD″, and capE). White arrows represent other genes in this region, and BLAST hit annotations are provided for three genes adjacent to the capsule operon: group II intron reverse transcriptase/maturase, also classified as an integration/excision element (ItrA) in the mobileOG-db, and poly-γ-glutamate hydrolase enzyme and γ-dl-glutamyl hydrolase associated with cell wall/membrane/envelope biogenesis.