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Sporulation is dispensable for the vegetable-associated life cycle of the human pathogen Bacillus cereus.

ABSTRACT: Bacillus cereus is a common food-borne pathogen that is responsible for important outbreaks of food poisoning in humans. Diseases caused by B. cereus usually exhibit two major symptoms, emetic or diarrheic, depending on the toxins produced. It is assumed that after the ingestion of contaminated vegetables or processed food, spores of enterotoxigenic B. cereus reach the intestine, where they germinate and produce the enterotoxins that are responsible for food poisoning. In our study, we observed that sporulation is required for the survival of B. cereus in leaves but is dispensable in ready-to-eat vegetables, such as endives. We demonstrate that vegetative cells of B. cereus that are originally impaired in sporulation but not biofilm formation are able to reach the intestine and cause severe disorders in a murine model. Furthermore, our findings emphasise that the number of food poisoning cases associated with B. cereus is underestimated and suggest the need to revise the detection protocols, which are based primarily on spores and toxins.

SUBMITTER: Antequera-Gomez ML

PROVIDER: S-EPMC8313275 | biostudies-literature | 2021 Jul

REPOSITORIES: biostudies-literature

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Sporulation is dispensable for the vegetable-associated life cycle of the human pathogen Bacillus cereus.

Antequera-Gómez María Luisa ML Díaz-Martínez Luis L Guadix Juan Antonio JA Sánchez-Tévar Ana María AM Sopeña-Torres Sara S Hierrezuelo Jesús J Doan Hung K HK Leveau Johan H J JHJ de Vicente Antonio A Romero Diego D

Microbial biotechnology 20210506 4

Bacillus cereus is a common food-borne pathogen that is responsible for important outbreaks of food poisoning in humans. Diseases caused by B. cereus usually exhibit two major symptoms, emetic or diarrheic, depending on the toxins produced. It is assumed that after the ingestion of contaminated vegetables or processed food, spores of enterotoxigenic B. cereus reach the intestine, where they germinate and produce the enterotoxins that are responsible for food poisoning. In our study, we observed ...[more]

PMID: 33955675

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Project description:BackgroundBacillus cereus is a Gram-positive, spore-forming bacterium that produces a spectrum of effectors integral to bacterial niche adaptation and the development of various infections. Among those is EsxA, whose secretion depends on the EssC component of the type VII secretion system (T7SS). EsxA's roles within the bacterial cell are poorly understood, although postulations indicate that it may be involved in sporulation. However, the T7SS repertoire in B. cereus has not been reported, and its functions are unestablished.MethodsWe used the type strain, B. cereus ATCC14579, to generate ΔessC mutant through homologous recombination using the homing endonuclease I-SceI mediated markerless gene replacement. Comparatively, we analyzed the culture supernatant of type strain and the ΔessC mutant through Liquid chromatography-tandem mass spectrometry (LC-MS/MS). We further generated T7SSb-specific gene mutations to explore the housekeeping roles of the T7SSb-dependent effectors. The sporulation process of B. cereus ATCC14579 and its mutants was observed microscopically through the classic Schaeffer-Fulton staining method. The spore viability of each strain in this study was established by enumerating the colony-forming units on LB agar.ResultsThrough LC-MS/MS, we identified a pair of nearly identical (94%) effector proteins named EsxA belonging to the sagEsxA-like subfamily of the WXG100 protein superfamily in the culture supernatant of the wild type and none in the ΔessC mutant. Homology analysis of the T7SSb gene cluster among B. cereus strains revealed diversity from the 3' end of essC, encoding additional substrates. Deletions in esxA1 and esxA2 neither altered cellular morphology nor growth rate, but the ΔesxA1ΔesxA2 deletion resulted in significantly fewer viable spores and an overall slower sporulation process. Within 24 h culture, more than 80% of wild-type cells formed endospores compared to less than 5% in the ΔesxA1ΔesxA2 mutant. The maximum spore ratios for the wild type and ΔesxA1ΔesxA2 were 0.96 and 0.72, respectively. Altogether, these results indicated that EsxA1 and EsxA2 work cooperatively and are required for sporulation in B. cereus ATCC14567.ConclusionB. cereus ATCC14579 possesses two nearly identical T7SSb-dependent effectors belonging to the sagEsxA-like proteins. Simultaneous deletion of genes encoding these effectors significantly delayed and reduced sporulation, a novel finding for EsxA.

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Sporulation is dispensable for the vegetable-associated life cycle of the human pathogen Bacillus cereus.

Publications

Sporulation is dispensable for the vegetable-associated life cycle of the human pathogen Bacillus cereus.

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