Microbial test organisms for sterilization processes in food industry

Abstract

The importance of sterilization processes has been a major topic in the food manufacturing industry since it preserves the quality and provides longer shelf-life by preventing microbiological recontamination of products. Aseptic packaging is an alternative technique to pack commercially sterile products into containers with a very low recontamination rate, thus, the entire production chain must be designed according to the aseptic condition, including the packaging material, the filling machines, and the surrounding area where the process takes place.

To evaluate a sterilization process, there is a validation test in which the process is challenged with certain test organisms. VDMA or Verband Deutscher Maschinen- und Anlagenbau (German Engineering Federation) has published a guideline (FS NuV No. 10/2005 Appendix A) containing the recommendation of the required outcomes for the validation test of the filling machines depending on the sterilization processes and test organisms.

Furthermore, the guideline has motivated us to obtain further confidence data on the performance of the test organisms. The present study aimed to investigate and compare the resistance of spore-producing organisms, A. brasiliensis, B. atrophaeus, and B. subtilis, against different disinfection/sterilization processes relevant to the food industry as indicated by VDMA, i.e. chemical (hydrogen peroxide and peracetic acid products of Oxonia), thermal (steam, hot water, and dry heat), as well as UV irradiation. A review of existing research was then compiled with the laboratory results to suggest suitable test organisms depending on the sterilization processes applied.

Based on the observations, B. subtilis spores are the test organism that showed highest resistance against H2O2 and Oxonia sterilization process with D60 °C = 6,17 s and Dambient = 13,89 s respectively. This result was in accordance with the guidelines of VDMA. For the thermal processes, VDMA suggested A. brasiliensis spores as the recommended test organisms. However, our findings resulted that for disinfection and sterilization processes that employ hot water and dry heat, B. atrophaeus spores were found as the ideal test organism exhibiting the desired resistance towards the given processes with a D90 °C and D121 °C of 8,1 min and 71,94 min. Our review study on steam sterilization suggested G. stearothermophilus as the suitable test organisms for steam sterilization, although VDMA suggested A. brasiliensis spores. Throughout the study, A. brasiliensis spores have demonstrated rapid reductions for all the sterilization process evaluated, indicating low resistance and therefore were not considered as an ideal test organism to use. However, when looking into the available literature on UV irradiation, it was found that A. brasiliensis could be a possible test organism to assess its effectiveness considering its black spore pigment, aspergillin, which is responsible to the high UV resistance as it absorbs high amounts of the UV radiation to protect the cell.

Popular Abstract

Food spoilage is the process leading to a product becoming either undesirable or unacceptable for human consumption, involving alterations in appearance, taste, texture or smell. This condition can be caused by the activity of specific groups of microorganisms, including spoilage bacteria, yeasts or mould. To make sure the food is easy to consume, the food is sterilized using a controlled heating process used to eliminate microorganisms that can reproduce inside the product under a nonrefrigerated state.

However, some microorganisms can form protective structures called spores when they are triggered by the stressful environment such as lack of nutrient. These spores are in a dormant state, which means their growth is temporarily stopped and helped them to conserve energy. Compared to the living cell, spores are more challenging to kill by heat or even by chemical agents and irradiation. Thus, spores are known to be resistant organisms.

Nowadays, aseptic technology is gaining popularity. The aseptic process allows food to be shelf stable for an extended period and eliminates the need for preservatives. The word ‘aseptic’ itself refers to a condition free of harmful microorganisms. For example, after UHT milk is sterilized for certain temperature and time, the milk is filled into a pre-sterilized container and the package is sealed under a sterilized environment. The aseptic technique is aimed to ensure that no recontamination occurs throughout the sterilization process.

To make sure the sterilization process is working correctly, the aseptic process can be monitored using a validation or challenge test. This method is performed using resistant organisms, such as spore-producing organisms, to measure the effectiveness of certain sterilization processes. The challenge test is carried by firstly applying a defined load of the test organisms on the filling machines, subjecting the organisms to the sterilization process, and finally counting the number of spores that are recovered after the treatment. These values are known as logarithmic cycle reduction (LCR), expressing the number of spores that are killed throughout the sterilization whereas reflecting the effectiveness of the sterilization. Generally, killing the spores up to 4-5 LCR is needed to achieve a trustworthy sterilization process.

In relation to that, VDMA or Verband Deutscher Maschinen- und Anlagenbau (German Engineering Federation) has published a guideline containing the required outcomes for the challenge test based on different sterilization processes and test organisms. In this guideline, VDMA suggests different test organisms for certain sterilization processes relevant to the food industry; chemical processes of hydrogen peroxide and peracetic acid product (e.g. Oxonia), heat processes of steam, dry heat, and hot water; as well as Ultraviolet (UV) irradiation process. Furthermore, the VDMA guidelines have motivated the present study to obtain further confidence data on the performance of the test organisms. Therefore, the aim of the study is to investigate and compare the level of resistance of three different test organisms of A. brasiliensis, B. atrophaeus, and B. subtilis against different sterilization process as listed by VDMA. After that, the results of the experiments will be compiled with a review of existing studies to finally propose suitable combinations of test organisms and sterilization processes.

Based on the research outcomes, both B. atrophaeus and B. subtilis spores are recommended as suitable test organisms to challenge the chemical sterilization processes. Meanwhile, for hot water and dry heat processes, B. atrophaeus is more suitable. Comparing with the guidelines from VDMA, B. subtilis is also recommended to test the chemical processes. However, for hot water and dry heat, VDMA guidelines reported that A. brasiliensis is more suitable. In spite of the recommendation from VDMA, our test results implied that A. brasiliensis spores are highly sensitive to the sterilization processes, meaning that the spores are easier to kill, thus, not reliable to be an indicator of a successful sterilization process. However, when looking into existing researches on UV irradiation, it was known that A. brasiliensis could be suitable to evaluate a UV irradiation process, as the spores have a specific mechanism of a spore pigment that could absorb the UV to protect themselves. Additionally, Geobacillus stearothermophilus are the test organisms that are recommended for the steam sterilization based on literature reviews since many studies have shown the spores’ resistance towards the steam process.

• author: Nabila, Yasmin
• supervisor: Jenny Schelin
• organization: Biotechnology; Applied Microbiology
• course: KMBM01 20181
• year: 2018
• type: H2 - Master's Degree (Two Years)
• subject: Technology and Engineering; Agriculture and Food Sciences
• keywords: Aseptic processing and packaging, validation, test organisms, Aspergillus brasiliensis, Bacillus atrophaeus, Bacillus subtilis, sterilization, disinfection, applied microbiology, teknisk mikrobiologi
• language: English
• id: 8958049
• date added to LUP: 2018-09-12 11:05:05
• date last changed: 2018-09-12 11:05:05