Biochemical limitations of Bacillus thuringiensis based biopesticides production in a wheat bran culture medium.

Bacillus thuringiensis, a gram-positive sporulating bacteria found in the environment, produces, during its sporulation phase, crystals responsible for its insecticidal activity, constituted of an assembly of pore-forming δ-endotoxins. This has led to its use as a biopesticide, an eco-friendly alternative to harmful chemical pesticides. To minimize production cost, one endemic Bacillus thuringiensis sv. kurstaki (Btk) strain Lip, isolated from Lebanese soil, was cultivated in a wheat bran (WB) based medium (IPM-4-Citrus project EC n° 734921). With the aim of studying the biochemical limitations of Btk biopesticide production in a wheat bran based medium, the WB was sieved into different granulometries, heat treated, inoculated with Btk Lip at flask scale, then filtered and separated into an insoluble and a permeate fractions. Several biochemical analyses, ie. bio performances, starch, elemental composition, total nitrogen and ashes, were then conducted on both fractions before and after culture. On a morphological level, two populations were distinguished, the fine starch granules and the coarse lignocellulosic particles. The biochemical analyses showed that both the raw and sieved WB have a similar proteins content (0.115 g/gdm WB), water content (0.116 g/gdm WB) and elemental composition (carbon: 45%, oxygen: 37%, nitrogen: 3%, hydrogen: 6%, ashes: 5%). The starch content was 17%, 14% and 34% and the fermentable fraction was estimated to 32.1%, 36.1% and 51.1% respectively for classes 2, 3 and 4. Both the elemental composition and Kjeldahl analyses showed that the nitrogen is the limiting nutrient of the culture.

The N-Glycan cluster from Xanthomonas campestris pv. campestris: a toolbox for sequential plant N-glycan processing.

N-Glycans are widely distributed in living organisms but represent only a small fraction of the carbohydrates found in plants. This probably explains why they have not previously been considered as substrates exploited by phytopathogenic bacteria during plant infection. Xanthomonas campestris pv. campestris, the causal agent of black rot disease of Brassica plants, possesses a specific system for GlcNAc utilization expressed during host plant infection. This system encompasses a cluster of eight genes (nixE to nixL) encoding glycoside hydrolases (GHs). In this paper, we have characterized the enzymatic activities of these GHs and demonstrated their involvement in sequential degradation of a plant N-glycan using a N-glycopeptide containing two GlcNAcs, three mannoses, one fucose, and one xylose (N2M3FX) as a substrate. The removal of the α-1,3-mannose by the α-mannosidase NixK (GH92) is a prerequisite for the subsequent action of the ß-xylosidase NixI (GH3), which is involved in the cleavage of the ß-1,2-xylose, followed by the α-mannosidase NixJ (GH125), which removes the α-1,6-mannose. These data, combined to the subcellular localization of the enzymes, allowed us to propose a model of N-glycopeptide processing by X. campestris pv. campestris. This study constitutes the first evidence suggesting N-glycan degradation by a plant pathogen, a feature shared with human pathogenic bacteria. Plant N-glycans should therefore be included in the repertoire of molecules putatively metabolized by phytopathogenic bacteria during their life cycle.