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Growth characteristics, gene expression and toxin production of Bacillus cereus in the simulated small intestine

Siele Ceuppens UGent (2012)
abstract
The aim of this PhD study was to investigate the growth and enterotoxin production of diarrhoeal B. cereus strains in the gastrointestinal tract. Diarrhoeal food poisoning caused by B. cereus is usually relatively mild and short-lasting, but in some cases the outcome is lethal (Lund et al. 2000). The bacterial numbers required for disease are not exactly known: the infective dose is estimated between 5 and 8 log viable cells or spores (EFSA 2005). This dose may be strain-dependent, since large variability was reported among different strains in their capability to produce (entero)toxins (Guinebretiere et al. 2002;Moravek et al. 2006) and to grow at body temperature (Wijnands et al. 2006b). Moreover, the co-ingested food may play an important role in disease aggravation and/or mitigation (Clavel et al. 2004). During this thesis, an in vitro simulation experiment for gastrointestinal passage was developed. Simulations were performed with different food types and several strains in the form of spores and vegetative cells to determine the survival and toxin production of these strains during gastrointestinal transit in the presence of food particles. In this way, the most important factors for B. cereus diarrhoeal food poisoning were identified and their influence was further investigated. General information about B. cereus, the associated toxin-mediated food poisoning syndromes and the regulation of enterotoxin gene expression is reviewed in Chapter 1 ‘Research context of diarrhoeal B. cereus and the regulation of toxin production’. The selection and optimization of an appropriate real-time PCR protocol as molecular quantification method for B. cereus in gastrointestinal media with high numbers of intestinal bacteria is described in Chapter 2 ‘Quantification of B. cereus vegetative cells and spores in the gastrointestinal environment’. With the suitable methods at hand, the behaviour of B. cereus in gastrointestinal simulation media was assessed in batch experiments, which are presented in Chapter 3 ‘Impact of intestinal microbiota and gastrointestinal conditions on B. cereus’. Chapter 4 ‘Survival and germination of Bacillus cereus spores during in vitro simulation of gastrointestinal transit occurred without outgrowth and enterotoxin production’ describes the development of the dynamic in vitro experiment simulating the gastrointestinal passage and the results obtained from simulating the consumption of mashed potatoes highly contaminated with B. cereus spores. Similar simulations with vegetative cells in lasagne verde were performed in Chapter 5 ‘Inactivation of B. cereus vegetative cells by gastric acid and bile during in vitro gastrointestinal transit’, which show the elimination of vegetative cells by gastric acid and bile and the limitations of batch experiments to simulate gastric passage. Since virulence is closely associated with enterotoxin production under gastrointestinal conditions, this was investigated in more detail in Chapter 6 ‘Enterotoxin production by B. cereus under gastrointestinal conditions’. Chapter 7 ‘Small B. cereus subpopulations are responsible for enterotoxin expression’ presents the development and characterization of a reporter strain for Cytotoxin K expression. The results suggest that only a small fraction of the B. cereus population in culture are expressing enterotoxins during growth in a laboratory monoculture. Finally, Chapter 8 ‘General discussion’ contains the integration of the results obtained during this PhD, their general discussion and their implications.
Please use this url to cite or link to this publication:
author
promoter
UGent and UGent
organization
alternative title
Groeiknitiek, genexpressie en toxineproductie van Bacillus cereus in de gesimuleerde dunne darm
year
type
dissertation (monograph)
subject
keyword
fluorescent reporter strain., food poisoning, Bacillus cereus, toxin regulation, real-time PCR, RT-qPCR, SHIME, in vitro simulation of gastrointestinal passage, toxin production
pages
246 pages
publisher
Ghent University. Faculty of Bioscience Engineering
place of publication
Ghent, Belgium
defense location
Gent : Faculteit Bio-ingenieurswetenschappen (A0.030)
defense date
2012-09-25 16:00
ISBN
9789059895416
language
English
UGent publication?
yes
classification
D1
additional info
dissertation consists of copyrighted material
copyright statement
I have transferred the copyright for this publication to the publisher
id
2982965
handle
http://hdl.handle.net/1854/LU-2982965
date created
2012-09-06 15:53:27
date last changed
2012-09-07 09:28:54
@phdthesis{2982965,
  abstract     = {The aim of this PhD study was to investigate the growth and enterotoxin production of diarrhoeal B. cereus strains in the gastrointestinal tract. Diarrhoeal food poisoning caused by B. cereus is usually relatively mild and short-lasting, but in some cases the outcome is lethal (Lund et al. 2000). The bacterial numbers required for disease are not exactly known: the infective dose is estimated between 5 and 8 log viable cells or spores (EFSA 2005). This dose may be strain-dependent, since large variability was reported among different strains in their capability to produce (entero)toxins (Guinebretiere et al. 2002;Moravek et al. 2006) and to grow at body temperature (Wijnands et al. 2006b). Moreover, the co-ingested food may play an important role in disease aggravation and/or mitigation (Clavel et al. 2004). During this thesis, an in vitro simulation experiment for gastrointestinal passage was developed. Simulations were performed with different food types and several strains in the form of spores and vegetative cells to determine the survival and toxin production of these strains during gastrointestinal transit in the presence of food particles. In this way, the most important factors for B. cereus diarrhoeal food poisoning were identified and their influence was further investigated.
General information about B. cereus, the associated toxin-mediated food poisoning syndromes and the regulation of enterotoxin gene expression is reviewed in Chapter 1 {\textquoteleft}Research context of diarrhoeal B. cereus and the regulation of toxin production{\textquoteright}. The selection and optimization of an appropriate real-time PCR protocol as molecular quantification method for B. cereus in gastrointestinal media with high numbers of intestinal bacteria is described in Chapter 2 {\textquoteleft}Quantification of B. cereus vegetative cells and spores in the gastrointestinal environment{\textquoteright}. With the suitable methods at hand, the behaviour of B. cereus in gastrointestinal simulation media was assessed in batch experiments, which are presented in Chapter 3 {\textquoteleft}Impact of intestinal microbiota and gastrointestinal conditions on B. cereus{\textquoteright}. Chapter 4 {\textquoteleft}Survival and germination of Bacillus cereus spores during in vitro simulation of gastrointestinal transit occurred without outgrowth and enterotoxin production{\textquoteright} describes the development of the dynamic in vitro experiment simulating the gastrointestinal passage and the results obtained from simulating the consumption of mashed potatoes highly contaminated with B. cereus spores. Similar simulations with vegetative cells in lasagne verde were performed in Chapter 5 {\textquoteleft}Inactivation of B. cereus vegetative cells by gastric acid and bile during in vitro gastrointestinal transit{\textquoteright}, which show the elimination of vegetative cells by gastric acid and bile and the limitations of batch experiments to simulate gastric passage. Since virulence is closely associated with enterotoxin production under gastrointestinal conditions, this was investigated in more detail in Chapter 6 {\textquoteleft}Enterotoxin production by B. cereus under gastrointestinal conditions{\textquoteright}. Chapter 7 {\textquoteleft}Small B. cereus subpopulations are responsible for enterotoxin expression{\textquoteright} presents the development and characterization of a reporter strain for Cytotoxin K expression. The results suggest that only a small fraction of the  B. cereus population in culture are expressing enterotoxins during growth in a laboratory monoculture. Finally, Chapter 8 {\textquoteleft}General discussion{\textquoteright} contains the integration of the results obtained during this PhD, their general discussion and their implications.},
  author       = {Ceuppens, Siele},
  isbn         = {9789059895416},
  keyword      = {fluorescent reporter strain.,food poisoning,Bacillus cereus,toxin regulation,real-time PCR,RT-qPCR,SHIME,in vitro simulation of gastrointestinal passage,toxin production},
  language     = {eng},
  pages        = {246},
  publisher    = {Ghent University. Faculty of Bioscience Engineering},
  school       = {Ghent University},
  title        = {Growth characteristics, gene expression and toxin production of Bacillus cereus in the simulated small intestine},
  year         = {2012},
}

Chicago
Ceuppens, Siele. 2012. “Growth Characteristics, Gene Expression and Toxin Production of Bacillus Cereus in the Simulated Small Intestine”. Ghent, Belgium: Ghent University. Faculty of Bioscience Engineering.
APA
Ceuppens, S. (2012). Growth characteristics, gene expression and toxin production of Bacillus cereus in the simulated small intestine. Ghent University. Faculty of Bioscience Engineering, Ghent, Belgium.
Vancouver
1.
Ceuppens S. Growth characteristics, gene expression and toxin production of Bacillus cereus in the simulated small intestine. [Ghent, Belgium]: Ghent University. Faculty of Bioscience Engineering; 2012.
MLA
Ceuppens, Siele. “Growth Characteristics, Gene Expression and Toxin Production of Bacillus Cereus in the Simulated Small Intestine.” 2012 : n. pag. Print.