Download MSc and BSc thesis projects

BSc and MSc thesis projects Food Microbiology - Physiology of Food-related Micro-organisms
Stress-response and survival strategies of Bacillus cereus
Keywords:
Bacillus cereus, sigma factors, polymerase, molecular biology,
bioinformatics
Supervision:
Marcel Tempelaars
Tjakko Abee ([email protected])
Project duration:
MSc - 6 months
Project description:
Bacillus cereus is a food pathogen that is often involved in outbreaks of food-borne illness. Because B.
cereus is able to form spores, this organism can survive the production processes used in the food
industry. These spores can germinate in the final food product and after that outgrowth might occur.
Under these processing conditions the cells are exposed to several types of stress, such as a low or
high temperature, high concentrations of salt, a lack of energy sources and/or a low pH. The central
question in this research project is to elucidate how vegetative cells of B. cereus can withstand the
different stresses they can encounter with a special emphasis on the role of regulators of gene
expression such as sigma factors.
Sigma factors are subunits of the enzyme RNA polymerase, which takes care of the transcription of
DNA to RNA. Sigma factors recognize specific promoter elements and can therefore "switch on"
specific sets of genes.
The role of sigma factors in the stress-response of Bacillus cereus
In this research project the role of different regulators of of the stress response of B. cereus will be
characterized using wild type strains and selected deletion mutants. Using different techniques, such
as mutant construction, phenotypic analysis, micro arrays, and by promoter-reporter fusions, the
expression of genes and roles of corresponding enzymes/proteins will be determined. This molecular
biological characterization can be coupled to the analysis of different physiological parameters (such as
membrane integrity and energy status), with the aim to obtain insight in the signals that lead to the
activation of adpative stress responses. Next, characterisation of the cellular defense systems involved
and their impact on adaptation and survival can be determined.
BSc and MSc thesis projects Food Microbiology - Physiology of Food-related Micro-organisms
Identification of genes and molecular mechanisms involved in biofilm
formation
Keywords:
Biofilm, pathogens, molecular biology
Supervisors:
Marcel Tempelaars ([email protected])
Heidy den Besten ([email protected])
Tjakko Abee ([email protected])
Project duration:
BSc and MSc – 3-6 months
Project description:
Many bacteria are capable of colonizing surfaces and grow in structures called biofilms. Biofilms
consist of the bacteria, attached to a surface and embedded in an extrapolymeric matrix, which can
consist of polysaccharides, protein, DNA etc. Biofilm cells are relatively hard to remove, because
they are more resistant to antimicrobial compounds and disinfectants. When food pathogens and
spoilage organisms manage to establish biofilm growth in food processing equipment, they will
form a persistent source of recontamination. The research in this project focuses on Gram-positive
bacteria that can occur in a variety of food products such as Listeria monocytogenes, Bacillus
cereus and Lactobacillus plantarum.
Different stages of biofilm development
To be able to prevent or fight biofilm formation, more knowledge is required on the genes and
molecular mechanism behind this physiological process. The complete genome sequence of
representatives of the bacteria listed above are available. The project concerns the generation of a
knock-out mutant library in one of the sequenced strains and the subsequent screening for
mutants with changes in biofilm phenotype. Interesting mutants will be further analyzed using
techniques like RT-PCR and DNA array analysis.
BSc and MSc thesis projects Food Microbiology - Physiology of Food-related Micro-organisms
Mixed biofilm formation of L. monocytogenes and lactic acid bacteria
Keywords:
biofilms, Listeria monocytogenes, multi-species, molecular biology
Supervisors:
Marcel Tempelaars ([email protected])
Heidy den Besten ([email protected])
Tjakko Abee ([email protected])
Project duration:
BSc and MSc – 3-6 months
Project description:
Biofilm are surface associated structures of bacterial cells that are attached to a substratum,
embedded in a matrix of extracellular polymeric substances. Bacteria in a biofilm have altered
phenotypes compared to cells in liquids with respect to their growth rate and gene transcription,
and in general more resistant to environmental adverse conditions. In natural environments,
biofilms are composed of more than one species resulting in so-called mixed species biofilms.
Listeria monocytogenes, the causative agent of listeriosis, can form biofilms and it is encountered
in similar niches as biofilm forming L. plantarum. Therefore, they are good model organisms to
study formation of mixed biofilms.
To date, mechanisms involved in mixed species biofilm formation involving food-associated
microbes including pathogens have hardly been studied. Therefore, we will study mixed species
biofilm formation by L. plantarum in co-culture with L. monocytogenes model strains and targeted
mutants. These L monocytogenes strains have deletions in specific genes, which might play a role
in biofilm formation. Mixed species biofilm formation will be analysed by different techniques
including (fluorescence) microscopy analysis and counting of biofilm cells)
Microscopic images of mixed biofilms
The impact of environmental conditions on mixed species biofilm formation will be assessed
including the effect of medium composition Both bacteria are fermentative but the pH growth limit
for L. plantarum is lower than for L. monocytogenes. The presence of glucose allows L. plantarum
to acidify the medium that can reduce survival of L. monocytogenes in the mixed biofilm.
Using targeted L. monocytogenes mutants, factors contributing to performance in mixed species
biofilms may be identified providing further insight into L. monocytogenes behaviour.
BSc and MSc thesis projects Food Microbiology - Physiology of Food-related Micro-organisms
Spore resistance and germination capacity of mesophilic and
thermophilic Bacillus species.
Keywords:
emetic B. cereus, thermophiles, sporulation, germination,
High temperature
Supervision:
Heidy den Besten ([email protected])
Tjakko Abee ([email protected])
Project duration:
MSc - 6 months
Project description:
Spores produced by mesophilic and thermophilic sporeformers may be introduced and/or produced
in food processing environments for example in so-called biofilms. In the latter case spores may be
released into fluid streams in pipe lines and end up in the final product. Characterization of stress
resistance and germination capacity of the spores produced in different conditions, may lead to
development of more effective cleaning procedures and cleaning strategies.
A combination of selected mesophilic and thermophilic microorganisms including respectively
(enterotoxic and emetic) B. cereus and G. stearothermophilus isolated from foods and food
processing environments will be included in the project.
The following parameters/methods may be investigated/used:
Spore formation capacity, spore stress resistance, germination capacity, fluorescence microscopy,
flow cytometry.
References:
- van der Voort M, Abee T. (2013) Sporulation environment of emetic toxin-producing Bacillus cereus strains
determines spore size, heat resistance and germination capacity. J Appl Microbiol. 114(4):1201-1210.
- den Besten HM, van Melis CC, Sanders JW, Nierop Groot MN, Abee T. (2012) Impact of sorbic acid on
germination and outgrowth heterogeneity of Bacillus cereus ATCC 14579 spores. Appl Environ Microbiol.
78(23):8477-8480.
BSc and MSc thesis projects Food Microbiology - Physiology of Food-related Micro-organisms
Heterogeneity in germination and outgrowth of Bacillus cereus
endospores
Keywords:
B. cereus, sporulation, germination, outgrowth
Supervision:
Marcel Tempelaars ([email protected])
Heidy den Besten ([email protected])
Tjakko Abee ([email protected])
Project duration:
MSc - 6 months
Project description:
Bacillus cereus is a Gram-positive, spore-forming bacterium that can be found in nearly every
environment. This means that it is easily transmitted into the food chain. The spores formed by B.
cereus are highly resistant against stresses, and are thus able to survive several processing conditions
used in the food industry. After germination in the final food product, outgrowth of B. cereus can
occur, after which it is able to cause food spoilage or food poisoning due to production of toxins. In
order to improve control of sporeformers, extensive research efforts are aimed at the improvement of
preservation methods. This study focusses on the germination and outgrowth capacity of B. cereus
spores with specific emphasis on the heterogeneity of these processes including the role of so-called
superdormant spores that are not easily triggered by nutrient germinants.
Previous research has shown that a large diversity occurs in germination between individual
spores, even under similar conditions. While one spore is already germinated and growing, others
will still be dormant. Heterogeneity in spore germination and outgrowth capacity will be studied at
single cell level by flow cytometry (FCM) or by direct imaging on Anopore The impact of
preservation treatments such as (mild) heating on spore germination and outgrowth capacity will
be determined. An extension of the research may include the assessment of factors contributing to
recovery of the (heat)damaged spores using selected deletion mutants.
Planning of the project:
1. production of spores (different conditions)
2. stress exposure of spores
3. germination capacity (FCM) or outgrowth capacity (anapore method)
5. heterogeneity assessment
http://www.biochemsoctrans.org/bst/034/1173/bst0341173f03.htm?resolution=HIGH
Cells and spores in cells
BSc and MSc thesis projects Food Microbiology - Physiology of Food-related Micro-organisms
Microbial warfare; understanding how small metabolites and peptides
produced by one organism inhibit the growth of other microorganisms
Keywords:
Antimicrobial compounds, Bacillus cereus, valinomycin, cereulide,
flow cytometry
Supervision:
Marcel Tempelaars ([email protected])
Tjakko Abee ([email protected])
Project duration:
MSc - 6 months
Project description:
Bacteria and fungi are known to produce substances that are secreted into their habitat and that
may have antimicrobial properties against other microorganisms. These antimicrobial agents can
be of importance for their potential as food preservatives and antagonistic effect against important
pathogens.
Similar compounds may also be produced by food-borne pathogens, such as the emetic toxin
cereulide, that is produced by Bacillus cereus. This toxin is associated with the onset of the emetic
syndrome, that is characterized by vomiting and nausea. Antimicrobial compounds are produced in
a variety of environments and in many cases their relevance in survival and/or competitive
advantage of the producing organism is not clear. In this project the effects of cereulide produced
by B. cereus on intraspecies and interspecies interactions will be investigated.
A range of techniques will be used including analysis of growth inhibition, viable counts, spore
counts, live/dead analysis and cellular activity detection using fluorescence microscopy and flow
cytometry.
The results obtained in this project will contribute to our understanding of the impact of cereulide
on ecology and performance in foods of emetic B. cereus.
BSc and MSc thesis projects Food Microbiology - Physiology of Food-related Micro-organisms
Pulsed electric field pasteurisation: influence of food matrix composition
and strain diversity
Keywords:
spoilage micro-organisms, pathogens, pulsed electrical field
Supervisors:
Rian Timmermans ([email protected])
Masja Nierop Groot (FBR)
Tjakko Abee
BSc and MSc – 4-6 months
Project duration:
Project description:
Pulsed electrical field (PEF) treatment is a novel pasteurisation technique which can be used to
inactivate spoilage micro-organisms and food pathogens, with minimal changes in the sensorial and
nutritional properties of food.
Previous research at Food & Biobased Research towards the sensitivity of spoilage and pathogenic
micro-organisms using PEF has been carried out with three different types of fruit juices. The
variation in these fruit juices was mainly dominated by a difference in pH, where the PEF process
showed to be more effective in acidic matrixes than at neutral pH.
The aim of this project is to evaluate the influence of the food matrix composition and strain
diversity on the effectiveness of PEF to inactivate spoilage yeast strain Saccharomyces cerevisiae,
E.coli ATCC 35218 as surrogate for E.coli O157:H7 and pathogens Salmonella Panama, and Listeria
monocytogenes. We would like to evaluate the impact of food matrix composition on sensitivity of
food pathogens and spoilage microorganisms towards PEF. Food products of interest include liquid
dairy products, vegetable juices, and lemonades with variable composition (pH, sugar
composition, fat content and conductivity). Comparison of the variability of the matrix and its
corresponding sensitivity towards the PEF process will be made, to determine the impact of matrix
composition on microbial behaviour towards PEF.
BSc and MSc thesis projects Food Microbiology - Physiology of Food-related Micro-organisms
Pulsed electric field pasteurisation: effect of strain history and recovery
conditions on survival
Keywords:
Supervisors:
Project duration:
spoilage micro-organisms, pathogens, pulsed electrical field, cultivation,
stress adaptation, outgrowth, quantification
Rian Timmermans ([email protected])
Masja Nierop Groot (FBR)
Tjakko Abee
MSc – 6 months
Project description:
Pulsed electrical field (PEF) treatment is a novel pasteurisation technique with proven effectiveness
against spoilage micro-organisms and food pathogens, with minimal changes in the sensorial and
nutritional properties of food.
Previous research at Food & Biobased Research has demonstrated that the spoilage microorganism Saccharomyces cerevisiae, E.coli ATCC 35218 as surrogate for E.coli O157:H7 and food
pathogens Salmonella Panama and Listeria monocytogenes can be inactivated by PEF in fruit
juices. These experiments were carried out with a standardized protocols for cultivation of target
strains. However, strain history may have impact on the sensitivity of the micro-organisms towards
an electrical pulse. Moreover, the ability to grow out of cells damaged by PEF treatment may be
influenced by matrix composition.
The aim of this project is to evaluate the influence of strain history and recovery conditions on
inactivation of the food pathogens Salmonella Panama and Listeria monocytogenes, and E.coli
ATCC 35218 as PEF-resistant surrogate for E.coli O157 by PEF. Variation in history conditions
includes cultivation temperature, growth phase, and cultivation medium composition.