Members of the genus Bacillus are known to produce a wide arsenal of antimicrobial substances, including peptide and lipopeptide antibiotics, and bacteriocins. Many of the Bacillus bacteriocins belong to the lantibiotics, a category of post-translationally modified peptides widely disseminated among different bacterial clades. Lantibiotics are among the best-characterized antimicrobial peptides at the levels of peptide structure, genetic determinants and biosynthesis mechanisms. Members of the genus Bacillus also produce many other nonmodified bacteriocins, some of which resemble the pediocin-like bacteriocins of the lactic acid bacteria (LAB), while others show completely novel peptide sequences. Bacillus bacteriocins are increasingly becoming more important due to their sometimes broader spectra of inhibition (as compared with most LAB bacteriocins), which may include Gram-negative bacteria, yeasts or fungi, in addition to Gram-positive species, some of which are known to be pathogenic to humans and/or animals. The present review provides a general overview of Bacillus bacteriocins, including primary structure, biochemical and genetic characterization, classification and potential applications in food preservation as natural preservatives and in human and animal health as alternatives to conventional antibiotics. Furthermore, it addresses their environmental applications, such as bioprotection against the pre- and post-harvest decay of vegetables, or as plant growth promoters.

Diversity and applications of Bacillus bacteriocins

Chitin is among the most abundant biomass present on Earth. Chitinase plays an important role in the decomposition of chitin and potentially in the utilization of chitin as a renewable resource. During the previous decade, chitinases have received increased attention because of their wide range of applications. Chito-oligomers produced by enzymatic hydrolysis of chitin have been of interest in recent years due to their broad applications in medical, agricultural, and industrial applications, including antibacterial, antifungal, hypocholesterolemic, and antihypertensive activity, and as a food quality enhancer. Microorganisms, particularly bacteria, form one of the major sources of chitinase. In this article, we have reviewed some of the chitinases produced by bacterial systems that have gained worldwide research interest for their diverse properties and potential industrial uses.

Bacterial chitinases: properties and potential.

Bacillus thuringiensis (Bt) is known as the most successful microbial insecticide against different orders of insect pests in agriculture and medicine. Moreover, Bt toxin genes also have been efficiently used to enhance resistance to insect pests in genetically modified crops. In light of the scientific advantages of new molecular biology technologies, recently, some other new potentials of Bt have been explored. These new environmental features include the toxicity against nematodes, mites, and ticks, antagonistic effects against plant and animal pathogenic bacteria and fungi, plant growth-promoting activities (PGPR), bioremediation of different heavy metals and other pollutants, biosynthesis of metal nanoparticles, production of polyhydroxyalkanoate biopolymer, and anticancer activities (due to parasporins). This review comprehensively describes recent advances in the Bt whole-genome studies, the last updated known Bt toxins and their functions, and application of cry genes in plant genetic engineering. Moreover, the review thoroughly describes the new features of Bt which make it a suitable cell factory that might be used for production of different novel valuable bioproducts.

Bacillus thuringiensis: a successful insecticide with new environmental features and tidings

Chitin is the second most abundant polymer in nature after cellulose and plays a major role in fungal cell walls. As a producer of variety of chitinase enzymes Trichoderma has become an important means of biological control of fungal diseases. A simple and sensitive method based on the use of basal medium with colloidal chitin as sole carbon source supplemented with Bromo cresol purple (pH indicator dye) is proposed to evaluate large populations of Trichoderma for chitinase activity. The soluble substrate with pH indicator dye (Bromo cresol purple, BCP) for the assay of chitinase activity on solid media is sensitive, easy, reproducible semi-quantitative enzyme diffusion plate assay and economic option to determine chitinases. Colloidal chitin derived from Rhizoctonia cell wall and commercial chitin included as a carbon source in broth also allowed selection and comparison of chitinolytic and exochitinase activity in Trichoderma spectrophotometrically. Released N-acetyl-β--D-glucosamine (NAGA) ranged from 37.67 to 174.33 mg/ml and 37.67 to 327.67 mg/ml and p-nitrophenol (pNP) ranged from 0.17 to 35.78 X 10-3 U/ml and 0.62 to 32.6 X 10-3 U/ml) respectively with Rhizoctonia cell wall and commercial chitin derived colloidal chitin supplemented broth.

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Chitinolytic assay of indigenous Trichoderma isolates collected from different geographical locations of Chhattisgarh in Central India

Colloidal chitin stained with Remazol Brilliant Blue R®, a useful substrate to select chitinolytic microorganisms and to evaluate chitinases

Three selected strains of Bacillus thuringiensis native to Mexico produced endochitinases, chitobiosidases, and N-acetyl-β-glucosaminidases in a medium containing colloidal chitin as a main carbon source. Two types of chitinases were clearly identified: endochitinases and chitobiosidases. Chromosomal location of a chitinase gene in B. thuringiensis LBIT-82 was resolved.

Selection of chitinolytic strains of Bacillus thuringiensis

Recently, we reported the synthesis of five bacteriocin-like inhibitor substances (Bt-BLIS: morricin 269, kurstacin 287, kenyacin 404, entomocin 420, and tolworthcin 524) by Mexican strains of Bacillus thuringiensis. Here we show that, collectively, these Bt-BLIS have a moderate to broad spectrum of antibacterial activity, being toxic to clinically significant against Gram-positive and Gram-negative bacteria, including common etiological agents of human diseases, such as strep throat and scarlet fever, septicemia, pneumonia, urinary tract infection, and emetic and gastrointestinal syndromes. Although synthesis of the five Bt-BLIS was independent of the presence of a target inducing bacterium, we demonstrated for the first time that a proteinaceous component(s) secreted by, or liberated by proteolytic cleavage of Bacillus cereus 183 following treatment with proteinase K, enhanced Bt-BLIS synthesis.

Enhanced synthesis and antimicrobial activities of bacteriocins produced by Mexican strains of Bacillus thuringiensis.

Aims: To synthesize two heterologous endochitinases in Escherichia coli and demonstrate their potential for applied use in generating antibacterial chitin-derived oligosaccharides (OGS).



Methods and Results:  Heterologous endochitinase genes, chiA Nima and chiA74, were expressed in E. coli. Endochitinases were secreted by the E. coli export machinery and by ∼20 h maximal chitinolytic activity was observed. The highest chitinolytic activity was observed with ChiA Nima, which produced antibacterial OGS with activities against Enterobacter cloacae, Escherichia coli, Staphylococcus aureus and S. xylosus.



Conclusions:  It was shown that the export machinery of E. coli is well suited for the secretion of bioactive ChiA74 and ChiA Nima endochitinases, and that the latter can generate antibacterial OGS.



Significance and Impact of the Study:  Our study suggests that it is feasible to synthesize endochitinases ChiA Nima and ChiA74 codified by E. coli and mass-produce these enzymes in culture supernatants. As signal peptides in native ChiA Nima and ChiA74 were recognized by the protein export molecular apparatus in E. coli, these short peptides could be included as signal sequences for transport in E. coli of other proteins with applied value. This is the first report suggesting that ChiA Nima can be used to produce OGS to control food-borne pathogenic bacteria.

https://sfamjournals.onlinelibrary.wiley.com/doi/pdf/10.1111/j.1365-2672.2008.03904.x

M. Bautista-Justo

Papers

Selection of chitinolytic strains of Bacillus thuringiensis

Enhanced synthesis and antimicrobial activities of bacteriocins produced by Mexican strains of Bacillus thuringiensis.

Generation of antibacterial oligosaccharides derived from chitin using heterologous endochitinase synthesized in Escherichia coli.