Antimicrobial and Phylogenomic Characterization of Bacillus cereus Group Strains Isolated from Different Food Sources in Italy

Abstract

Background:Bacillus cereus is a widespread environmental Gram-positive bacterium which is especially common in soil and dust. It produces two types of toxins that cause vomiting and diarrhea. At present, foodborne outbreaks due to Bacillus cereus group bacteria (especially Bacillus cereus sensu stricto) are rising, representing a serious problem in the agri-food supply chain. Methods: In this work, we analyzed 118 strains belonging to the Bacillus cereus group, isolated from several food sources, for which in vitro and in silico antibiotic resistance assessments were performed. Results: Many strains showed intermediate susceptibility to clindamycin, erythromycin, and tetracycline, suggesting an evolving acquisition of resistance against these antibiotics. Moreover, one strain showed intermediate resistance to meropenem, an antibiotic currently used to treat infections caused by Bacillus cereus. In addition to the phenotypic antimicrobial resistance profile, all strains were screened for the presence/absence of antimicrobial genes via whole-genome sequencing. There was inconsistency between the in vitro and in silico analyses, such as in the case of vancomycin, for which different isolates harbored resistance genes but, phenotypically, the same strains were sensitive. Conclusions: This would suggest that antibiotic resistance is a complex phenomenon due to a variety of genetic, epigenetic, and biochemical mechanisms.

Keywords: Bacillus cereus group; WGS; antibiotic resistance; food poisoning; minimum inhibitory concentration.

Figure 1

Phylogenetic tree based on cgMLST allele profiles from 118 B. cereus group isolates. The phylogenetic tree was constructed using the neighbor-joining (NJ) method. The isolates were indicated with pubMLST ID (Table S1). The clade, species, and source were added and color-coded, as reported in the legend. B. cytotoxicus NVH 391-98 (pubMLST ID: 2720) was used as an outgroup. BC_ss: B. cereus s.s.; BM: B. mosaicus; BC_T: B. cereus s.s. biovar Thuringiensis; BM_C: B. mosaicus subsp. cereus; BM_T: B. mosaicus subsp. cereus biovar Thuringiensis; BM_E: B. mosaicus subsp. cereus biovar Emeticus; BT: B. thuringiensis; BC_C: B. cytotoxicus.

Figure 2

Heatmap of antibiotic resistance gene occurrence (clustered by antibiotic molecule class) among strains. Colors range from light yellow (n. of predicted genes for a drug class = 0) to dark red (n = 10); x-axis: antibiotic molecule class; y-axis (left side): strains hierarchical clustering by gene occurrence; y-axis (right side): strain ID; species nomenclature (“BC_ss”: B. cereus s.s.; “BM”: B. mosaicus; “BC_T”: B. cereus s.s. biovar Thuringiensis; “BM_C”: B. mosaicus subsp. cereus; “BM_T”: B. mosaicus subsp. cereus biovar Thuringiensis; “BM_E”: B. mosaicus subsp. cereus biovar Emeticus); source of isolation (“M”: milk and dairy products; “G”: gastronomic preparations; “Me”: meat; “V”: vegetable; “P”: pasta; “F”: fish; “PB”: pastry and bakery products); antibiotic drugs (“FLQ”: fluoroquinolone; “PHE”: phenicol; “TET”: tetracycline; “MCL”: macrolide; “CLI”: clindamycin; “GLY”: glycopeptide; “BLA”: β-lactam).