Growth and survival of pathogens in selected Nigerian fermented foods

My research focus is on the safety of indigenous fermented foods and starter culture development. The aim is to reduce the drudgery associated with local food processing operations and to develop Nigerian fermented foods to meet international safety standards. My gratitude goes to the Society for Applied Microbiology for the award of an International Capacity Building Fund (ICB) Grant in order to fast track the achievement of my professional philosophy of “Food processing for safety and wealth creation”. My sincere and special thanks also go to Professor Jane Sutherland of the Microbiology Research Unit at London Metropolitan University for supervising my research under the SfAM fund.

Fermented foods are very popular in Nigeria, yet the processing operations involved in their production remain indigenous and at the household level of technology (Edema & Omemu, 2007). Many of the processes involve uncontrolled fermentations by mixed cultures of microorganisms under largely unhygienic conditions, with a resultant effect of minimizing the potential benefits of fermentation viz: preservation/shelf stability, improved digestibility, reduced toxicity, increased nutrient availability and desirable sensory modifications. Hence, fermented foods which should normally be beneficial may harbour pathogens, thereby constituting microbiological hazards (Gadaga et al, 2004). Research on fermented foods has focused principally on isolation and identification of the microorganisms associated with the fermentations and determination of the biochemical roles of the microorganisms but relatively little on the improvement of processing conditions of these foods (Edema & Fawole, 2006; Edema & Anetor, 2009). In order to assure the safety of these food products, it is necessary to determine their status with respect to presence or otherwise of pathogens.

Recent advances in identification techniques indicate that classical procedures do not adequately describe microorganisms. Lack of funds, infrastructure and facilities for scientific research and training remain major constraints to the development and use of latest cutting-edge molecular techniques in the developing world, hence the continued dependence on the less definitive, conventional (phenotypic) methods for strain identification. As a result, lots of research work on microbial identification based on these traditional, culture-dependent techniques does not get published in high impact international journals. This limits the applications of indigenous fermented foods, despite their significant potential for food safety and security.

The research work conducted under the SfAM ICB fund therefore aimed at isolating Campylobacter and Listeria monocytogenes, two of the most common food-borne pathogens (at least in the Western world – there is dearth of statistics for African countries including Nigeria) from iru and kunu. Iru is a soup condiment made from African locust beans (Parkia biglobosa) by Bacillus species in an alkaline fermentation process. Kunu, on the other hand, is a cereal-based beverage produced through the acidic fermentation of millet, sorghum or maize. The work was divided into two parts: 1) A training component with focus on gaining knowledge of cultivation, isolation and molecular identification of Campylobacter and L. monocytogenes (Inadequate facilities in Nigeria prevent adequate diagnosis or recognition of these pathogens) 2) A research component involving survival of Salmonella and Staphylococcus aureus in the two food types.  St. aureus (a Gram-positive bacterial pathogen) and Salmonella (a Gram-negative bacterial pathogen) are two of the most commonly isolated pathogenic species from Nigerian foods, be they fermented or not. Some studies have indicated substantial reduction or elimination of these organisms when simple hygiene practices in food processing are applied (Edema, 2011). However, there remains a strong indication that there could be a food safety risk associated with survival of these pathogens in fermented foods.

Professor Jane Sutherland has collections of these pathogens from which we selected 3 species of Campylobacter and Listeria each as references for the training component. One strain of Staphylococcus aureus and 5 species of Salmonella were also selected and used as test pathogens for the research component. The two food types were inoculated with the test pathogens and incubated at 15oC and 30oC for up to two weeks. Selective media were used for the isolation of Listeria and Campylobacter from the food samples and the cultivation of reference strains used for comparison: Listeria selective agar base along with Listeria enrichment broth base and Campylobacter blood-free agar base for Campylobacter. All media and appropriate supplements for each medium were obtained from Oxoid.

Total bacterial counts were in the range of log 5 for kunu and log 7 for iru. Counts of inoculated pathogens were log 2 at the start of the experiments. Iru allowed survival of pathogens longer than kunu, wherein the pathogens were not recovered by the 3rd day at both temperatures. Typical colonies from the Listeria and Campylobacter selective media were sub-cultured by repeated streaking before DNA extraction (InstaGene Matrix), purification (QIAquick Purification kit) and precipitation (with sodium acetate). The samples have been sent for sequencing to confirm their identities and the results will be included in a manuscript being prepared for publication as a research paper or short communication. The findings of the research work also provide some information on the growth and survival of St. aureus and Salmonella in Iru and kunu. This will contribute to development of a programme of Good Manufacturing Practice (GMP) for the selected fermented foods, thus improving the reliability, consistency and safety of these foods.

The training through this ICB fund grant has promoted my knowledge and understanding of the procedures in microbiological food safety research. More important for me was the expertise gained from the experience in molecular characterisation of the microorganisms. This experience is vital for my ongoing research for which I have recently obtained a TWAS (Academy of Sciences for the Developing World) research grant. Special thanks go to Dr. Irene Ouoba who took time out of her busy schedule to assist with the molecular biology aspect. Her contribution was invaluable to the training component and pivotal to my preparations towards developing a molecular biology research laboratory in my University.

I am also grateful to Dr. Amara Anyogu, Dr. Brigitte Awamaria, Elyn Xu, Daniel Amund, Nasim Farahmand and Shahram Naghizadeh, all of the Microbiology Research Unit at London Metropolitan University for their kind support, which contributed immensely to the success of my work and without which I could not have achieved as much as I did.
Mojisola O. Edema

References
Edema M. O., Fawole O. (2006) Evaluation and optimization of critical control points in the production of iru. Research Journal of Microbiology 1 (6) 503 – 511.

Edema M. O., Omemu A. M. (2007) Safety of small-scale food fermentations in Nigeria.5th International Conference of the Nigerian Society for Experimental Biology (NISEB) Kogi State University, Anyigba, Kogi State. Feb 28 – Mar 3, 2007 Proceedings pp 60 – 61.

Edema M. O., Anetor O. H. (2009) Optimization of processing conditions for Kunun-zaki, a spontaneously fermented cereal-based beverage. Journal of Food Quality 32: 469–480.

Edema M. O. (2011) Application of HACCP Principles for improving safety of indigenous Nigerian foods. Journal of Applied and Environmental Science 6 (3): 28 – 33.

Gadaga T. H., Nyanga L. K., Mutukumira A. N. (2004). The occurrence, growth and control of pathogens in African fermented foods. African Journal of Food, Agriculture, Nutrition and Development. 4 (1): http://www.bioline.org.br/request?nd04009



Categories: Early Career Scientists

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