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The influence of sporulation medium on the spore coat protein composition of Bacillus subtilis spores.

ABSTRACT: Spores are of high interest to the food and health sectors because of their extreme resistance to harsh conditions especially against heat. Earlier research has shown that spores prepared on solid agar plates have a higher heat resistance than those prepared in a liquid medium. It has also been shown that the more mature a spore is the higher is its heat resistance contributed most likely by the progressive cross-linking of coat proteins. The current study for the first time enlightens the effect of sporulation medium on spore proteins. The 14N spores prepared on solid agar medium and 15N metabolically labelled spores prepared in liquid medium differ in their coat protein composition as revealed by LC-FT-MS/MS analyses. The 14N:15N isotopic ratio of the 1:1 mixture of liquid and solid medium cultured spores exposes that most of the identified inner coat and crust proteins are significantly upregulated while most of the outer coat proteins are significantly downregulated for the spores prepared on solid medium relative to the spores prepared in the liquid medium. Medium specific differences in isotopic ratios between the tryptic peptides of expected cross-linked proteins suggest that also the coat protein cross-linking may be medium specific. The spores prepared on solid medium show a higher thermal resistance than the spores prepared in liquid medium. Since the core DPA content is found to be similar in both the spore populations, it appears that the difference in wet heat resistance is connected to the differences in the coat protein composition and assembly. Supporting the proteomic analyses, the electron microscopy analyses show a significantly thinner outer coat layer of the spores cultures on the solid medium.

INSTRUMENT(S): apex ultra

ORGANISM(S): Bacillus Subtilis Subsp. Subtilis

SUBMITTER: Wishwas R. Abhyankar

LAB HEAD: Chris G. de Koster

PROVIDER: PXD004473 | Pride | 2017-09-20

REPOSITORIES: pride

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The Influence of Sporulation Conditions on the Spore Coat Protein Composition of <i>Bacillus subtilis</i> Spores.

Abhyankar Wishwas R WR Kamphorst Kiki K Swarge Bhagyashree N BN van Veen Henk H van der Wel Nicole N NN Brul Stanley S de Koster Chris G CG de Koning Leo J LJ

Frontiers in microbiology 20161013

Spores are of high interest to the food and health sectors because of their extreme resistance to harsh conditions, especially against heat. Earlier research has shown that spores prepared on solid agar plates have a higher heat resistance than those prepared under a liquid medium condition. It has also been shown that the more mature a spore is, the higher is its heat resistance most likely mediated, at least in part, by the progressive cross-linking of coat proteins. The current study for the ...[more]

PMID: 27790212

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Project description:Bacterial spores play an important role in disease initiation, transmission and persistence. The outermost spore layer, the exosporium, is important as it is the first point of contact between the spore and the environment and may be involved in spore adherence, protection and germination. Clostridium sordellii is a highly lethal, spore forming pathogen that causes soft-tissue infections, enteritis and toxic-shock syndrome. Despite the importance of C. sordellii spores in disease, spore proteins from this bacterium have not been defined or interrogated functionally. In this study, we identified the C. sordellii outer spore proteome and two of the identified proteins, CSA and CSB, were characterised using a genetic and phenotypic approach. Both proteins were essential for the correct formation and positioning of the C. sordellii spore coat and exosporium. The absence of CSA reduced sporulation levels and increased spore sensitivity to heat, sodium hydroxide and hydrochloric acid. By comparison, CSB was required for normal levels of spore adherence to cervical, but not vaginal, cells, with csb mutant spores having increased adherence properties. The establishment of a mouse infection model of the gastrointestinal tract for C. sordellii allowed the role of CSA and CSB to be interrogated in an infected host. Following the oral administration of spores to mice, the wild-type strain efficiently colonized the gastrointestinal tract, with the peak of bacterial numbers occurring at one day post-infection. Colonization was reduced by two logs at four days post-infection. By comparison, mice infected with the csb mutant did not show a reduction in bacterial numbers. The absence of CSB therefore allows the csb mutant to persist within the gastrointestinal tract. We conclude that C. sordellii outer spore proteins are important for the structural and functional integrity of spores, and for colonization and persistence during infection. Finally, Clostridium difficile, Bacillus cereus and Bacillus anthracis encode proteins with homology to CSA and CSB but these bacteria produce spores that are structurally dissimilar to those of C. sordellii, and the function of the proteins in these hosts is different to that in C. sordellii. These findings suggest that, despite their homology, spore proteins can have variable functions in different bacterial species which highlights the necessity of studying each spore protein in the cognate species from which it originates.

			Action	DRS
	CW2098Coat.xlsx	Xlsx
	CW2098_RE2_01_2089.mgf	Mgf
	CW2098_analysis.baf	Other
	CW2098_mzIdentML.mzid	Mzid
	CW2143Coat.xlsx	Xlsx