Stress response of Group I and II Clostridium botulinum
Year: 2015
Type: dissertation
Abstract: The capability of Clostridium botulinum to survive food processing and grow in foods is a substantial hazard to human health. Clostridium botulinum can thrive under stress conditions and the wide genetic variation among C. botulinum strains reflects differences in their growth properties. However, the strain variation in growth, molecular mechanisms, and genetic markers behind the stress tolerance of C. botulinum are poorly understood. This study focused on the variation in growth of Group II C. botulinum strains at temperatures out of the optimal range and characterized the role of the genes that are involved in the stress response of Group I ATCC 3502 and Group II Beluga strains. At extreme temperatures, Group II C. botulinum strains showed significant variation. Some strains showed better growth at 37 °C than their generally accepted optimum growth temperature of 30 °C. Although not absolute, the minimum growth temperatures of 24 Group II strains in our experimental setting were higher than expected. The AFLP typing indicated a weak association between genetic background and growth at extreme temperatures. In addition, clustering analysis showed that the type B and F strains are closely related, since they were clustered together, and the type E strains formed a separate cluster. Our data indicated that Group II C. botulinum strains show considerable differences in their growth properties, and this should be taken into account in designing safety measures against this foodborne pathogen. The study implied that csp genes play important roles in NaCl, pH, and ethanol stress and motility of C. botulinum ATCC 3502. The growth of the cspB mutant was impaired in relation to the wild-type strain under all stress conditions tested, suggesting a universal role in stress response. Additionally, functional cspC was required for efficient growth during NaCl, pH, and ethanol stress. The cspA did not take part in NaCl, pH, and ethanol stress responses, and inactivation of this gene enhanced growth, which suggested a possible growth repressor role. In addition, cspB was not involved in motility, but cspA and cspC were crucial for flagellation and motility. The two-component system CBO2306/CBO2307 of Group I C. botulinum ATCC 3502 and CLO3403/CLO3404 of the Group II Beluga strain were induced up to 4.4- and 3.4-fold, respectively, after cold shock, but not at optimum growth temperatures. This indicated that both systems were involved in rapid response to cold shock. This argument was further confirmed by inactivation of the CBO2306/CBO2307 or CLO3403/CLO3404 genes, which resulted in impaired ability of the mutant strains to grow at low temperatures compared with the wild-type strains. In addition, the histidine kinase encoding clo3403 was required for motility. The cbo2802 that encodes the DEAD-box RNA helicase in C. botulinum ATCC 3502 was cold-inducible. The RT-qPCR analysis showed a 7.6-fold rapid increase in the relative expression of cbo2802 after cold shock. The significance of cbo2802 in the growth at suboptimal temperature of C. botulinum ATCC 3502 was further characterized by disruption of this gene. The mutants without functional cbo2802 showed impaired growth and restricted motility at suboptimal temperature. This study concluded that strains of Group II C. botulinum pose considerable variation in growth properties, which may be of critical importance in predictive modeling of growth in foods. The major cold-shock protein-encoding gene cspB seems to play a universal role in stress response and cspC a role in NaCl, pH, ethanol stress, and motility. The cspA gene is not involved in the stress response of C. botulinum ATCC 3502, but is important for flagella formation. The CBO2306/CBO2307 in Group I C. botulinum ATCC 3502 and CLO3403/CLO3404 in the Group II Beluga strain are part of the cold stress response machinery. Moreover, clo3403 is required for motility. We report for the first time in C. botulinum that DEAD-box RNA helicase is crucial for growth at suboptimal temperature and motility.
Source: Työväentutkimus Vuosikirja
Author: Yağmur Derman
Language: English
Subfield: Clinical Neurology
Field: Medicine
Domain: Health Sciences
Open Access status: green
OpenAlex ID: W2243635615