Movement and containment of microbial contamination in the nutrient mist bioreactor

Abstract

In vitro plant cultures tend to get contaminated easily with bacteria and fungi because they are grown for long times in sugar-rich media. Contamination of bioreactors is particularly problematic as larger volumes entail larger losses. To study the movement and develop subsequent control of contaminants in the mist bioreactor, the spore-forming microbes Penicillium chrysogenum and Bacillus subtilis were deliberately inoculated into three possible locations in the reactor: the growth chamber (GC), the medium reservoir (R), or the mist-generating chamber (MG). Compared to inoculation into either R or MG regions, the growth of P. chrysogenum inoculated into the GC required 3 more days (c. 60% more time) to move throughout the rest of the reactor. In contrast, regardless of where B. subtilis was inoculated (GC, R, or MG), it took 7d to contaminate the entire system. The movement of filamentous fungi and bacteria seems to follow the same route of contamination throughout this reactor. Once visibly present in the reactor, neither contaminant was controllable by addition of the biocide, Plant Preservative Mixture (PPM). Both microbes were completely inhibited if PPM was added to the MG at the time of inoculation and then again 2-d post-inoculation of plants. Reactors were fun for 3 wk. Plants remained free of contamination. These results will prove useful in the implementation of large-scale in vitro culture systems.