Multiple linear regression models to simulate spore yields of Bacillus amyloliquefaciens BS13 through optimization of medium composition.

Bacillus amyloliquefaciens is a food spoilage spore-forming bacterium. Its spores are useful for multiple biotechnological applications. Nevertheless, few reports are available regarding the achievement of a high cell density and good sporulation effectiveness under fermentation conditions. Therefore, the current study was designed to optimize a low-cost fermentation medium allowing the highest sporulation yield by B. amyloliquefaciens strain BS13. Our data revealed that tryptone and starch were the best carbon and energy sources. In addition, two nitrogen sources namely, corn steep liquor (CSL) and yeast extract (YE), allowed a significant enhancement of spore production and they were both retained for further optimization. A combination of CaCl2 , MgSO4 , and MnSO4 showed a positive impact on spores' production. The composition of the optimized medium was (in g/L); tryptone 3, starch 15, CSL 13.5, YE 1.5, CaCl2 0.1, MgSO4 ·7H2 O 0.012, and MnSO4 ·7H2 O 0.0012. Such medium was further validated in a 400-L fermentor. The spore yield by B. amyloliquefaciens strain BS13 was enhanced from 3.0 × 1010 spores/mL under flask culture conditions to 6.2 × 1010 spores/mL when cultures were performed on large scale. Therefore, strain BS13 spore preparation could be proposed as a promising probiotic and a biocontrol agent useful for plants, animals, and humans.

Morphological differences between aerial and submerged sporidia of bio-fongicide Pseudozyma flocculosa CBS 16788.

Pseudozyma flocculosa is a fungus very useful and highly efficient as a biocontrol agent against powdery mildew. The reproduction of this fungus occurs exclusively by asexual production of conidia or sporidia that are the most suitable form for agricultural use and seems to be the most resistant to storage conditions. Despite the advantages offered by P. flocculosa in biological control, the use of this fungus use remains largely limited compared to that of chemical fungicides, at least partly due to the difficulty to obtain sporidia resistant to adverse environmental stresses in submerged culture conditions. Under solid-state and submerged-state cultivation, P. flocculosa strain CBS 16788 produced different types of sporidia. The submerged sporidia (SS) appeared relatively uniform in size, which was 15,4 ± 1,6 µm µm long, and 2,8 ± 0.8 µm wide. The aerial sporidia (AS) varied in shape and size, with a mean length of 8,2 ± 3 µm and width of 2,3 ± 0.6 µm. Under scanning and transmission electron microscopy, the cell wall of submerged sporidia was thinner than that of aerial spores, and the surface was smooth in contrast to the aerial sporidia that had a tendency to have verrucous, brittle surface characteristics. The thickness of the aerial sporidia wall is due to the presence of an outer layer rich in melanin. The sporidia germination was compared on YMPD (yeast extract, malt extract, soy peptone, dextrose and agar) coated coverslips. The aerial sporidia did not show germ tubes until 5 h of incubation, while the submerged sporidia showed many germ tubes after the same time. The resistance against the adverse environmental conditions in relation to the type of sporidia of P. flocculosa is discussed.

An enhanced process for the production of a highly purified extracellular lipase in the non-conventional yeast Yarrowia lipolytica.

Yarrowia lipolytica LgX64.81 is a non-genetically modified mutant that was previously identified as a promising microorganism for extracellular lipase production. In this work, the development of a fed-batch process for the production of this enzyme in this strain was described. A lipolytic activity of 2,145 U/mL was obtained after 32 h of batch culture in a defined medium supplemented with 10 g/L of tryptone, an enhancer of lipase expression. To maximize the volumetric productivity, two different fed-batch strategies had been investigated. In comparison to batch process, the intermittent fed-batch strategy had not improved the volumetric lipase productivity. In contrast, the stepwise feeding strategy combined with uncoupled cell growth and lipase production phases resulted in a 2-fold increase in the volumetric lipase productivity, namely, the lipase activity reached 10,000 U/mL after 80 h of culture. Furthermore, this lipase was purified to homogeneity by anion exchange chromatography on MonoQ resin followed by gel filtration on Sephacryl S-100. This process resulted in an overall yield of 72% and a 3.5-fold increase of the specific lipase activity. The developed process offers a great potential for an economic production of Lip2 at large scale in Y. lipolytica LgX64.81.

Selection of a cold-adapted bacterium for bioremediation of wastewater at low temperatures.

Amongst more than 1000 isolates collected in various cold environments, the strain Arthrobacter psychrolactophilus Sp 31.3 has been selected for its ability to grow and to produce exoenzymes at low temperatures, its inability to grow at 37 degrees C, its non-halophilic character and its growth versatility on various media. This non-pathogenic strain displays a strong resistance to desiccation and storage at room temperature and is suitable for the production of freeze-dried bacterial starters. When grown in a synthetic wastewater at 10 degrees C, the strain induces a complete clarification of the turbid medium and efficiently hydrolyses proteins, starch and lipids in the broth. Furthermore, this strain has a remarkable capacity to improve the biodegradability of organic compounds in wastewater as indicated by a BOD(5)/COD ratio of 0.7.

Culture and spray-drying of Tsukamurella paurometabola C-924: stability of formulated powders.

The nematocidal agent, Tsukamurella paurometabola C-924, was cultured in a 300 l bioreactor. Spray-dried formulations of this microorganism were prepared using sucrose. At an outlet temperature 62 degrees C, survival rates between 12 and 85% were reached with sucrose up to 10% (w/w). The stability study of the powders showed that the best storage condition was at 4 degrees C under vacuum. A new method for the calculation of cell death order for bacteria stored at low temperatures was developed. Powders stored under vacuum showed an Arrhenius behavior in relation to cell death kinetics.

Utilization of methyloleate in production of microbial lipase.

In this article, we report the development and optimization of an industrial culture medium for the production of extracellular lipase in the yeast Yarrowia lipolytica. Until now olive oil in combination with glucose was used as the carbon source and inducer for the production of lipase. Our results demonstrate that methyloleate, a cheap hydrophobic compound, could efficiently substitute olive oil as the inducer and carbon source for lipase production. A new process of lipase production was developed yielding a twofold increase in the level of production compared with the levels in previous reports.