BSc and MSc thesis projects Food Microbiology - Quantitative Microbiology/Human Nutrition
Dietary microbial exposure: Relation with allergies?
Keywords: allergies, nutritional micro-organism load, food habits
Berber Vlieg-Boerstra (Academic Medical Center),
Jeanne de Vries (
[email protected])
Project duration: BSc and MSc - 3-6 months
Project description:
Background:
Declining microbial exposure is suggested as a major cause for the increase in allergic disease in
recent decennia. This concept has been based on the “hygiene hypothesis” in which it is
hypothesized that frequent infections in early childhood could be protective for the development of
allergic disease. Over the years, the focus on microbial exposures has widened from the influence
of pathogenic microbes causing infections towards non-pathogenic types of strains (commensals
and environmental strains) and components of microbes such as bacterial endotoxins by farm
living. In parallel with the potential association of microbial load and the increase in the prevalence
of allergic disease in the past decennia, food habits have changed dramatically. However, there are
no data available on the total microbial exposure of our diet. In earlier theses, we identified foods
contributing to 95% of the total microbial load and developed a food frequency questionnaire (FFQ)
for assessment of the dietary microbial content of the diet.
To follow-up these studies, we aim to:
1. Study factors influencing on minimum and maximum microbial contents and to incorporate
these factors into the FFQ;
2. Evaluate the FFQ on the individual level;
3. To qualitatively describe differences between our actual food habits and those of several
decennia ago, and the suspected effect on the dietary microbial exposure;
4. To quantitatively compare the microbial load of the Dutch National Food Consumption Survey
2003 with the diet used several decennia ago.
Activities:
Literature research, recruitment of participants, development and validation of an instrument by
sampling and analysis of duplicate foods, statistical analyses using SPSS, report and presentation.
BSc and MSc thesis projects Food Microbiology - Quantitative Microbiology/Environmental
Technology
The use of HACCP for pathogen control in new sanitation concepts
Keywords: Hazard Analysis, CCP’s, validation, verification, water teratment processes
Grietje Zeeman (Environmental Technology)
Project duration: BSc and MSc - 3-6 months
Project description:
Background:
Water re-use will become a more and more important challenge in the coming years. Both within
food industry as well as in agricultural production and in households, treatment of water for re-use
and valorisation of specific streams, will increase in importance. Boundary condition for these
feed-back streams are of course the control of pathogens.
Grey water
Black water
Rainwater
In these studies we aim to:
5. Study factors influencing pathogen levels;
6. Apply principles of HACCP on the water and other streams as safety management system;
7. Carry out validation experiments in treatment processes.
8. Carry out verification experiments in treatment processes.
Activities:
Literature research, inactivation experiments, process monitoring, HACCP-analysis
BSc and MSc thesis projects Food Microbiology - Quantitative Microbiology & Food Safety
Management
Meta-analysis on the thermal resistance of microorganisms
Keywords: meta-analysis, D- and z-value, variability
Marcel Zwietering (
[email protected])
Project duration: BSc and MSc – 4-6 months
Project description
Numerous thermal inactivation studies have been done for foodborne pathogens and spoilage
microorganisms to determine their thermal resistance. However, the results of the thermal
inactivation studies vary because of the differences between used inactivation method, strain,
inactivation medium, physiological state of the cells, etc. To determine the global thermal
inactivation parameters of a pathogen, such as the D-value and z-value, a meta-analysis study can
be initiated (1). In a meta-analysis study, a vast number of thermal inactivation data of a
foodborne pathogen are collected, and the global thermal inactivation parameters, as well as the
factors influencing thermal resistance can be extracted. In this way, the differences found in many
studies can be explained and the sources of variability can be listed and quantified. This
information is critical to realistically predict inactivation of microorganisms in foods.
After the first meta-analysis initiative in 2006 for pathogens, more and more thermal inactivation
studies of foodborne pathogens have been published. Thus, there is a need to incorporate those
recent publication data into the existing database, especially for the pathogens for which only few
studies were available at that time. The extension of this database is expected to result in more
reliable information on the thermal inactivation profile of the pathogen in question. Furthermore,
we want to extend the data base with information on spoilage organisms.
Also you will perform experiments with several strains from a same species (e.g Salmonella, spores
of B. subtilis) to compare strain variability to the variability found in literature (2). This will help us
to understand how strain variability affects thermal inactivation efficiency.
1. van Asselt, E. D., and M. H. Zwietering. 2006. A systematic approach to determine global thermal
inactivation parameters for various food pathogens. International Journal of Food Microbiology. 107:73-82.
2. Aryani D.C., den Besten H.M.W., Hazeleger W.C., Zwietering M.H. 2015. Quantifying variability and the
effect of growth history on thermal resistance of Listeria monocytogenes. International Journal of Food
Microbiology 193, 130-138
BSc and MSc thesis projects Food Microbiology - Quantitative Microbiology & Food Safety
Management
Sampling plans for spices and dried herbs
Key words: microbiological limits, sample size, low aw food commodities, microbial
distributions
Heidy den Besten (
[email protected])
Marcel Zwietering
Project duration: BSc, MSc 3-6 months
Project description:
Spices and dried herbs are produced from diverse plants from mostly tropical and subtropical soils
and climates. Depending on the product, different parts of the plant are used to produce the spice
or the herb. In case of mustard and nutmeg the seeds are used, for pepper the berries, in oregano
and bay the leaves and so on [1]. These products commonly carry large numbers of micro-
organisms. Spore-forming organisms capable of causing gastroenteritis and Salmonella spp. can be
found in spices. In fresh herbs pathogenic types E. coli have been reported [2].
For a producer to bring a safe product in the market he has to ensure that levels of microorganisms
in a product do not pose threat to human health and the way to achieve this is not only to prevent
and control contamination, but also to monitor the process. Sampling is a method to verify a
product’s safety throughout the food chain and the sampling strategy plays a significant role
whether the monitoring is effective.
In this project, batches of spices or dried herbs will be examined for presence of pathogenic
microorganisms, such as Salmonella spp. and also non-pathogenic microorganisms’ levels will be
assessed, such as total viable counts, Enterobacteriaceae and/or sporeformers. This will provide
information on the distribution of these organisms in batches of spices or dried herbs which are
important to evaluate efficient sampling strategies.
1. ICMFS, 1980, Microorganisms in Foods 1: Their Significance and Methods of Enumeratio. 2nd ed.
(1978); reprinted 1982, 1988 with revisions. Toronto: University of Toronto Press. ISBN: 0802022936,
pp 731-735
2. ICMFS, 1986, Microorganisms in Foods 2. Sampling for microbiological analysis: Principles and specific
applications. International Commission on Microbiological Specifications for Foods. 2nd ed. (1986).
Toronto: University of Toronto Press. ISBN: 0802056938, pp. 213-216
BSc and MSc thesis projects Food Microbiology - Quantitative Microbiology & Food Safety
Management
Risk assessment of Listeria monocytogenes
Key words: exposure assessment, risk assessment, maximum dose,
competitive flora, ready-to-eat products
Project duration: MSc only – 3-6 months
Project description:
Listeria monocytogenes has caused foodborne disease with major public health consequences,
mainly in the elderly. The organism is known to grow in a variety of ready-to-eat products at
refrigeration temperatures. Various authors have estimated the risk of listeriosis and the risk of
ingesting a certain number of Listeria has been assumed to depend on time and refrigeration
temperature; higher temperatures being associated with higher maximal doses and lower
temperatures with lower maximal doses. The maximum number of organisms that can be reached
forms an essential parameter in exposure assessment of L. monocytogenes.
Recent experimental research suggests that the maximal number of L. monocytogenes that can be
present in ready-to-eat food products is higher than previously assumed. The objective of this
research is to update existing an existing risk assessment model with recently obtained data on the
maximum concentration of L. monocytogenes. The work will include literature search, computer-
based modelling and a limited amount of laboratory work.
BSc and MSc thesis projects Food Microbiology - Quantitative Microbiology & Food Safety
Management
Sporeformers in infant formula
Key words: pathogens, infant formula, modelling, validation
Gerrieke van Middendorp
Project duration: MSc only – 3-6 months
Project description:
Reconstituted infant formulae are excellent growth media for numerous bacteria that may be
present in the dry infant formula or in the water used for reconstitution, or in the bottles in which
the formula is stored. Hot reconstitution at temperatures over 70°C and storage at low
temperature has therefore been recommended as control measures to prevent microbial growth.
Placing a container with reconstituted liquid formula in the refrigerator, however, does not mean
that the temperature of the liquid is directly the same temperature as the set-point of the
refrigerator. The temperatures may allow sporeformers, such as Bacillus cereus, to germinate and
grow in infant formula.
In this project temperature profiles of infant formula during the cooling process have been obtained
and a model was built that predicts germination and outgrowth of sporeformers. The aim of this
project is to further develop the model and to validate it by testing a range of strains.
BSc and MSc thesis projects Food Microbiology - Quantitative Microbiology & Food Safety
Management
The impact of inhomogenous distributions of micro-organisms on the
outcome of MPN based methods
Key words: sampling plans, distributions, estimation, fitting
Ida Jongenburger
Project duration: MSc only – 4-6 months
Project description:
Microbiological analysis of foods often aims at the enumeration of microorganisms in a sample to
verify compliance with microbiological criteria. Enumeration is mostly done with the “plate count
method” that is based on the traditional petri dish in combination with an agar medium. The
sensitivity of this method depends on the maximum amount of sample that can be added to a petri
dish (typically 0.1g). If a lower limit of detection is required the Most Probable Number (MPN)
approach can be used that is based on presence-absence testing of multiple test portions
containing up to 100g of product (and dilutions thereof) in combination with a statistical evaluation
of the results.
The official ISO MPN procedure requires that only one sample is taken from which only one primary
suspension is prepared; from this several test portions are taken that are examined separately.
However, in many labs a slightly different procedure is used that consists of taking multiple
samples and preparing multiple primary suspensions. Depending on the distribution of
microorganisms in foods and the possibility that clumps of microorganisms will disperse during the
preparation of the primary dilution the latter procedure may result in severe underestimation of the
MPN. This may consequently lead to erroneous management decisions concerning the acceptability
of a lot, but also to wrong estimation of level based on MPN methods.
The aim of this project is to compare both procedures and to gain more insight in the distribution
of microorganisms in different types of food products.
MSc project
• Experimental work: gather data on distributions of micro-organisms in a group of products
(Cronobacter spp. and/or Salmonella in infant formula, milk and cheese, artificially adding
different levels of organisms).
• Compare microbial counts expressed as plate counts to Most Probable Number with respect to
estimated numbers and variability.
• In silico: Find and fit statistical distributions that describe the CFU and MPN numbers best.
• In a later stage: review scientific literature about validation of microbiological methods with
respect to spiking procedure and repeat determination of the limit of detection LOD50 using
both MPN procedures.
BSc and MSc thesis projects Food Microbiology - Quantitative Microbiology & Food Safety
Management
Miniaturized MPN method to enumerate Salmonella: simulation and
validation
Key words: sampling plans, distributions, estimation, fitting
Martine Reij (
[email protected])
Project duration: MSc only – 4-6 months
Project description:
The presence of Salmonella spp. in broiler chickens is a human health risk. To reduce this risk,
measures are taken to prevent and control of Salmonella in the poultry production chain. Not only
a method to detect, but especially a rapid method to quantify low numbers of Salmonella will be
convenient to investigate the critical points in the production chain. One of the ways to estimate
concentrations of microorganisms is the most probable number technique (MPN), testing
absence/presence of microorganisms in a series of tubes of multiple dilutions.
Scheme for an MPN method for solid products, which is diluted (1:10) and the test portions are divided over
pre-enrichment media (10 mL of 10-1 ≈ 1 g, 1 mL of 10-1≈0.1 g, 0.1 mL of 10-1≈ 0.01 g
The figure illustrates an example of an MPN for a solid product. A principle of the MPN method is,
that the individual tubes of the sample are independent. Although the MPN method is useful to
estimate low numbers, it is time consuming and labour intensive. Therefore, a new method (ISO
6579-2; 2012 ) has been launched to enumerate Salmonella using a miniaturized MPN technique.
However, the dilutions are not independent of each other.
The aim of this project is to determine the importance of the dilutions being independent. The
method will be to simulate outcomes in silico. In addition, the method will be validated in the
laboratory.
MSc project
• Literature study: gather information on the traditional and miniaturized MPN methods.
• In silico: simulate MPN scenarios with various levels of Salmonella.
• Experimental work: validate the method
ISO/TS 6579-2:2012, Microbiology of food and animal feed - Horizontal method for the
detection, enumeration and serotyping of Salmonella - Part 2: Enumeration by a
miniaturized most probable number technique
BSc and MSc thesis projects Food Microbiology - Quantitative Microbiology & Food Safety
Management
Quantifying the transfer of Enterobacteriaceae under dry conditions
Key words: Enterobacteriaceae, dry products, transmission routes, transfer, tracing
Salmonella, detection
Gerrieke van Middendorp
Project duration: MSc only – 3-6 months
Project description:
Enterobacteriaceae are a large family of Gram-negative bacteria, some of which are pathogens like
Salmonella, pathogenic E. coli, and Cronobacter. In order to prevent foodborne illness,
transmission routes of these pathogens have been studied. Most of these transmission studies
were performed under wet conditions. Recently, however, low-moisture foods such are seeds, nuts
and peanut butter have been implicated in outbreaks of Salmonella.In this study we aim to
quantify the transfer of Enterobacteriaceae from air and dust to surfaces and ultimately to dry
food products.
BSc and MSc thesis projects Food Microbiology - Quantitative Microbiology & Food Safety
Management
Quantifying the effect on washing and handling on microflora of
vegetables
Key words: Enterobacteriaceae, vegetables, salads, transmission routes, transfer, tracing
Salmonella, detection
Gerrieke van Middendorp
James Noah Ssemanda
Project duration: MSc only – 3-6 months
Project description:
The importance of vegetables in nutritious, healthy diets is well recognized, and in recent years
consumers have been encouraged to eat more of these products. For many countries, particularly
developing countries, products such as lettuce, spinach, tomatoes, carrots, onions, cabbages,
beets, cucumber and coriander, have increasingly become valuable commodities. Whereas some
vegetables can be cooked and others eaten raw, the consumption of sliced ready-to-eat; shredded;
skinned raw vegetables or salads, has become popular. This popularity has led to a remarkable
growth in trade of fresh vegetables in globalized market settings. As a result, almost in all
countries, agricultural practices to increase the production levels of vegetables have been adopted.
However, the increased production, distribution and consumption of ready-to-eat vegetables has
also been associated with an increased number foodborne outbreaks for the last three decades. It
is against this background that a study is planned. This study is aimed at providing scientific
answers to the concerns of the general public that vegetables (salads) served in food outlets in
developing countries may be a potential source food microbiological contamination and hence a
threat to food safety and food security overall.
In the frame work of a larger study in collaboration with the Rwanda Standards Board, various
subtopics can be studied such as:
- Characterisation of foodborne pathogens originating from farm vegetables in Rwanda
- Quantifying the effect of washing, sanitizers and handling on microflora of vegetables.
BSc and MSc thesis projects Food Microbiology - Quantitative Microbiology & Food Safety
Management
Prediction of spoilage of fresh poultry meat under dynamic temperature
regimes
Key words: shelf life fresh poultry meat, Pseudomonas ssp., cold chain,
temperature abuse
Supervisors: Masja Nierop Groot (FBR) ([email protected])
Martijntje Vollebregt (FBR) (
[email protected])
Marcel Zwietering (WU) (
[email protected])
Project duration: MSc only – 3-6 months
Project description:
The microbial shelf life of fresh poultry meat is largely determined by the temperature in the chain
starting from production and distribution chain to storage at home of consumers. To control the
spoilage of fresh poultry meat, it is of importance to control the temperature. Predictive models
could help to understand the impact of temperature on shelf life of products. Moreover, it allows
prediction on the effect of interventions on the shelf life. Predictive information can be used is
decision processes on temperature management and cleaning and disinfection protocols. This
project aims to develop a predictive shelf life model for fresh poultry meat as a function of
temperature. The project will be executed in collaboration with a Dutch producer of fresh poultry
meat.
The following activities are foreseen in the project:
1) Construction of a predictive shelf life model for fresh poultry meat
Models that predict the growth of the spoilage microorganisms Pseudomonas sp. on fresh poultry
meat as a function of temperature will be constructed. Using these models, different temperature
regimes representative for the production chain of fresh poultry meat will be developed. As an
input to these models, growth parameters for Pseudomonas sp. previously determined for fresh
poultry meat (Bruckner et al. 2013) will be used.
2) Validation of the predictive models
The predictive shelf life model for fresh poultry meat will be validated in practice. To this end, fresh
poultry meat naturally contaminated with Pseudomonas sp. will be subjected to different
temperature scenario’s that mimic conditions realistic for the poultry meat production and
distribution chain. At regular time intervals, samples will be taken and Pseudomonas sp. will be
enumerated. To validate the model, the predicted shelf life of fresh poultry meat stored at different
temperature regimes will be compared to the observed shelf life.
3) Effect of interventions on the shelf life of fresh poultry meat
Using the model developed under 1) the effect of interventions aiming for reduction of inital on
shelf life can be predicted. Intervention than can be included are for example altered cleaning and
desinfection regimes, use of UV or pulsed light treatment. Depending on the availability of
literature data, relevant input that can be obtained from literature, historical data present of the
producing industries or should be experimentally determined.
References:
− Bruckner et al. (2013). A preductive shelf life model as a tool for the improvement of quality management
in pork and poultry chains. Food Control 29: 451-460.
− Sylvain Dabadé et al. (2015). Prediction of shelf life of tropical shrimp (Penaeus notialis) under dynamic
temperature regimes. Int. Journal of Food Microbiology 210: 121-130.
− Sylvain Dabadé et al. (2015). Spoilage evaluation, shelf life prediction and potential spoilage organisms of
tropical brackish water shrimp (Penaeus notialis) at different storage temperatures. Food Microbiology 48:
8-16.
BSc and MSc thesis projects Food Microbiology - Quantitative Microbiology & Food Safety
Management
Prediction of outgrowth of Bacillus cereus under dynamic temperature
regimes and MAP conditions
Key words: Microbial food safety, Bacillus cereus, hurdle technology, modified
atmosphere, chilled temperature
Supervisors: Hermien van Bokhorst-van de Veen (FBR) ([email protected])
Masja Nierop Groot (FBR) (
[email protected])
Heidy den Besten (WU) (
[email protected])
Project duration: MSc thesis project- 6 months or BSc project – 4 months
Project description:
Foodborne illness caused by bacteria associated with contaminated food is still of great concern.
Conventional heat treatments of food to inactivate food spoilers and pathogens are familiar to food
processes, but consumers demand higher product quality that requires milder preservation
strategies. This poses new challenges in assuring the stability and safety of a variety of foods. To
ensure food safety, additional hurdles need to be included in the product. For example, in
pasteurised products with pH above 4.6, storage at refrigeration temperatures or in combination
with a modified atmosphere is required to control outgrowth of spoilage and pathogenic microbes.
This category includes processed foods of extended durability including ready-to-eat meals.
Microbes of concern with respect to the safety of this type of products include psychrotrophic
pathogenic sporeformers such as certain B. cereus strains. Food and Biobased Research (FBR), part
of Wageningen UR, develops mild preservation concepts for food producers that enhance shelf life
and microbiological safety. However, knowledge about the growth kinetics of B. cereus, especially
under modified atmosphere conditions, like high oxygen, carbon dioxide, or nitrogen
concentrations, is lacking. This is of great value concerning food safety of for instance fresh
produce or ready-to-eat products with extended shelf life.
The following activities are foreseen in the project:
1) Construction of a predictive growth-no growth model for B. cereus
Models that predict the growth/no growth of B. cereus as a function of temperature, pH and water
activity are available in literature (Daelman et al., 2013). A literature study will be performed to
extend the growth parameters database.
2) Validation of the predictive models
The project continues with determination of growth parameters of psychrotrophic B. cereus strains
under modified atmosphere packaging conditions and different low temperatures. Additional
hurdles will be included like variation in salt concentration/water activity and in pH. The most
BSc and MSc thesis projects Food Microbiology - Quantitative Microbiology & Food Safety
Management
promising conditions will be taken in a next series of experiments and when time allows, a
pasteurization step combined with the use of B. cereus spores can be included. Furthermore, the
gained data will be combined to build a generic model to predict growth/no growth conditions of B.
cereus.
Skills:
Under supervision, the student will perform a small literature study to become familiar with the
subject and to retrieve relevant data from literature. He or she will prepare and perform the
experiments, analyse the results that will be presented in a written report and oral presentation. In
this project, the student will gain knowledge on:
− setting up and performing experiments, interpret results and report them in a clear and correct
way,
− working in a ML-II laboratory (pathogen lab),
− performing kinetic measurements and growth assays,
− working with a modified atmosphere packaging system, and
− application of the results in a growth/no growth predicting model.
References:
− Samapundo et al. (2014). Incidence, diversity and characteristics of spores of psychrotolerant spore formers
in various REPFEDS produced in Belgium. Food microbiology, 44:288-295
− Daelman et al. (2013). Growth/no growth models for heat-treated psychrotrophic Bacillus cereus spores
under cold storage. International journal of food microbiology, 161:7-15
− Bennik et al. (1995). Growth of psychrotrophic foodborne pathogens in a solid surface model system under
the influence of carbon dioxide and oxygen. Food microbiology, 12, 509-519
