Non-toxic Clostridium botulinum strains and their contribution to product and process innovations

Why this project?

Consumers' increasing demand for clean label products such as ready meals, means food companies are continuously investing in innovations regarding conservatives and the treatment of food products. For example, in the meat industry the focus is on replacing nitrate additives (e.g. E 250) and its precursor nitrate (e.g. ovv E 251) due to growing concerns regarding the health aspects of nitrite.  
These trends also include risks for food safety. Probably the most important spore-forming bacteria that must be kept under control in products is Clostridium botulinum. On the one hand, because nitrite has a performant inhibiting effect against this organism (EFSA 2003: 50-150 mg/kg nitrite essential) and, on the other hand, because there are psychrotrophic strains that are of great concern regarding foodstuffs that must be refrigerated. 

Suitable product and process adaptations are thus needed to keep C. botulinum under control. 'Label friendly’ alternatives must be identified that show anticlostridial activity when replacing nitrate and nitrite. Optimum process parameters in combination with supplementary product conditioning (pH, aw, temperature, organic acids, packaging atmosphere) must be examined for processing. 
Challenge tests are essential for assessing these adaptations for their effectiveness against C. botulinum. The classic challenge tests for C. botulinum require exceptional safety measures, specific (anaerobe) growth conditions and specialised personnel. There is also no selective culture for C. botulinum, which is not beneficial to plate counts because the target cells are difficult to differentiate from the background microbiota. All of this decreases the degree to which these classic tests can be applied. A solution to this challenge is using marked, non-toxic C. botulinum strains. For example, gene technology makes it possible to develop attenuated C. botulinum that are non-toxic but have other properties that are equivalent to the pathogenic variant.   

Research approach

BotulinSafe is a collective research project (type VIS-CO) that wants to use these non-toxic C. botulinum strains with the objective of: 

  • Developing easily accessible challenge tests for C. botulinum for every target product type model 
  • Validating ‘label friendly’ alternatives for nitrate/nitrite anticlostridial activity 
  • Determining process intensity parameters in combination with conditioning conditions and their impact on the control of C. botulinum per target product type model 
  • Translating the project results for implementation in the industrial products sector. 

We wish to obtain this objective by:  

  • Developing non-toxic strains that represent the diversity of C. botulinum
  • Developing markers in the various representative strains' genome to make a selective count and thus make it possible to determine a specific germ count for C. botulinum 
  • Demonstrating the proof of concept for using C. botulinum strain cocktails for challenge studies and application of anticlostridial strategies in model products 
  • Validating the anticlostridial activity of natural antimicrobial substances  
  • Validating the anticlostridial activity and technological implementation of coagulase-negative Staphylococcus (CNS) as a starter culture in fermented meat products 
  • Providing an innovation forum aimed at further exploring the applications of these non-toxic strains 

Target group and expected results

 We expect the following results from the BotulinSafe project:  

  • Product type-specific challenge tests that are also validated for their application by commercial analytical laboratories,
  • Candidate natural antimicrobial substances with validated anticlostridial activity (including but not limited to, applicable as a nitrate/nitrite replacement in meat products), possible starter cultures with validated, well-characterised anticlostridial activity and supplemented with knowledge about their technological functions
  • Food products with adapted contents (in particular regarding nitrate/nitrite) or prepared using adapted (milder) thermal processing will be validated for their food safety aspects regarding C. botulinum by using challenge tests 

The project is, in the first place, geared towards meat processers and ready-made meal producers who have a positive attitude toward or export to countries with strict regulations regarding C. botulinum-related food safety risks. In addition, the results may also have added value for ingredient suppliers and commercial analytical laboratories. 

Project partners 

Flanders’ FOOD manages and coordinates the project.  
Responsibility for the execution: 

  • The research group for Food and Microbial Technology from KU Leuven, under the supervision of Prof. Chris Michiels 
  • The research group for Industrial Microbiology and Food Biotechnology from VUB, under the supervision of Prof. Frédéric Leroy 
  • The research group for Technology and Quality of Animal Products from KU Leuven, under the supervision of Prof. Ilse Fraeye 
  • The research group for Food Microbiology and Preservation from Ghent University, under the supervision of Prof. Frank Devlieghere 
Flanders' FOOD logo
KU Leuven
Vrije Universiteit Brussel
Universiteit Gent