Search

The foodborne pathogen Listeria monocytogenes can persist in produce processing environments, which increases the risk for food contamination. Increased resistance to antimicrobials commonly used in cleaning and sanitizing procedures may contribute to L. monocytogenes' persistence in these environments. This study aimed to evaluate sanitizer resistance in L. monocytogenes isolates collected from three tree fruit packing facilities (F1, F2, and F3) during packing seasons 2020-2021 (Y1) and 2021-2022 (Y2), and to assess evidence of persistence based on the genomic similarity of isolates to historical isolates collected in previous years. L. monocytogenes isolates collected in 2020-2022 (n = 44) were tested for resistance to peroxyacetic acid (PAA) and a proprietary biofilm-removing agent using a broth microdilution assay. Further, L. monocytogenes isolates were whole genome sequenced and screened for the presence of antimicrobial resistance and virulence genes, as well as to assess the genomic similarity of isolates using the CFSAN SNP bioinformatic pipeline. Over half (57%) of the tested isolates had a PAA minimum inhibitory concentration (MIC) of 250 ppm, which was similar to the applied concentration of the PAA sanitizer in the three facilities (230 ppm). In contrast, 80% of tested isolates had a biofilm remover MIC of 0.13 ppm, which was substantially below the concentration applied in the facilities (137 ppm). Genomes of all tested isolates carried antimicrobial resistance (fosX, lin, mdrL, mprF, and norB) and virulence (inlA, inlB, plcA, plcB, prfA, hly, mpl, and iap) genes. L. monocytogenes isolates collected between 2020 and 2022 belonged to three distinct lineages, with 22 multilocus sequence types (MLSTs) belonging to 22 different clonal complexes. Genomic similarity analysis with historical isolates collected from the same facilities in 2016-2017 demonstrated a 5-year persistence of the genotypes ST 1003 and ST 554 in F2, which were no longer detected in 2022. Overall, our results highlight the need to re-evaluate sanitizer concentrations to effectively control persistent L. monocytogenes strains in tree fruit packing facilities., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

Gram-positive, spore-forming members of the Bacillus cereus group species complex are widespread in natural environments and display various degrees of pathogenicity. Recently, B. cereus group strain Bacillus mycoides Flugge ATCC 21929 was found to represent a novel lineage within the species complex, sharing a relatively low degree of genomic similarity with all B. cereus group genomes (average nucleotide identity [ANI] < 88). ATCC 21929 has been previously associated with the production of a patented antibiotic, antibiotic 60-6 (i.e., cerexin A); however, the virulence potential and growth characteristics of this lineage have never been assessed. Here, we provide an extensive genomic and phenotypic characterization of ATCC 21929, and we assess its pathogenic potential in vitro. ATCC 21929 most closely resembles Bacillus paramycoides NH24A2 T (ANI and in silico DNA-DNA hybridization values of 86.70 and 34.10%, respectively). Phenotypically, ATCC 21929 does not possess cytochrome c oxidase activity and is able to grow at a range of temperatures between 15 and 43°C and a range of pH between 6 and 9. At 32°C, ATCC 21929 shows weak production of diarrheal enterotoxin hemolysin BL (Hbl) but no production of nonhemolytic enterotoxin (Nhe); at 37°C, neither Hbl nor Nhe is produced. Additionally, at 37°C, ATCC 21929 does not exhibit cytotoxic effects toward HeLa cells. With regard to fatty acid composition, ATCC 21929 has iso-C17:0 present in highest abundance. Based on the characterization provided here, ATCC 21929 T (= PS00077A T = PS00077B T = PSU-0922 T = BHP T ) represents a novel effective B. cereus group species, which we propose as effective species " Bacillus clarus " IMPORTANCE The B. cereus group comprises numerous closely related lineages with various degrees of pathogenic potential and industrial relevance. Species-level taxonomic classification of B. cereus group strains is important for risk evaluation and communication but remains challenging. Biochemical and phenotypic assays are often used to assign B. cereus group strains to species but are insufficient for accurate taxonomic classification on a genomic scale. Here, we show that antibiotic-producing ATCC 21929 represents a novel lineage within the B. cereus group that, by all metrics used to delineate prokaryotic species, exemplifies a novel effective species. Furthermore, we show that ATCC 21929 is incapable of producing enterotoxins Hbl and Nhe or exhibiting cytotoxic effects on HeLa cells at human body temperature in vitro These results provide greater insight into the genomic and phenotypic diversity of the B. cereus group and may be leveraged to inform future public health and food safety efforts., (Copyright © 2020 Méndez Acevedo et al.)