Research Article: Effect of cultivation pH on the surface hydrophobicity of Bacillus subtilis spores

Date Published: July 27, 2017

Publisher: Springer Berlin Heidelberg

Author(s): Elisabeth Eschlbeck, Simon A. W. Bauer, Ulrich Kulozik.


Bacillus subtilis spores are often used as biological indicators (BI) to monitor decontamination processes with gaseous hydrogen peroxide. Results in practical inactivation validation tests, however, vary considerably with no available explanation so far. This study reports on the effect of cultivation pH on spore surface hydrophobicity. Surface hydrophobicity is suspected to have an impact on the decontamination of technical surfaces such as packaging material when gaseous, condensing hydrogen peroxide is applied. It is the aim of this study to examine the impact of different cultivation pH levels on surface hydrophobicity and resistance of B. subtilis spores. Submersed cultivation of B. subtilis in bioreactors at controlled conditions with different static pH levels led to contact angles ranged between 50° and 80°, which was analyzed with water on a homogeneous layer of spores on a filter sheet. Resistance of spores was also affected by the cultivation pH. The results show that the culturing conditions during BI production should be controlled to obtain BI with specified characteristics in inactivation validation tests.

Partial Text

In food and pharma industries sterile surfaces for aseptic packaging and clean rooms are often indispensable. The processes to obtain surface sterility are either based on physical or on chemical action. Hydrogen peroxide (H2O2) is among the most often used disinfectants. H2O2, mostly a mixture of 35% H2O2 in water and low amounts of stabilizers, can be applied in its liquid form as immersion bath, as spray or as vapor (Engelhard and Kulozik 2006; Pruß et al. 2012).

The obtained amount of spores are in a range of 1.28 × 107–3.40 × 108 cfu per mL and thus slightly dependent on cultivation pH. For the applied cultivation conditions including aeration, media composition and cultivation time, a pH level of 8.00 resulted in a maximum amount of spores. pH-values from 7.00 to 9.00 resulted in similarily high spore yields and provided enough spores for all further experiments. Further optimization of the cultivation conditions will presumably result in higher spore yields, however this was not the aim of the study.




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