Improving Beneficial Traits in Bacillus cabrialesii subsp. cabrialesii TE3T through UV-Induced Genomic Changes
Authors: Morales Sandoval Pamela Helué , Ortega Urquieta María Edith , Valenzuela Ruíz Valeria , Montañez Acosta Kevin , Campos Castro Kevin Alejandro , Parra Cota Fannie I. , Santoyo Gustavo , de los Santos Villalobos Sergio 
Published: September 2024

It is essential to hunt for new technologies that promote sustainable practices for agroecosystems; thus, the bioprospecting of beneficial microorganisms complementing with mutation induction techniques to improve their genomic, metabolic, and functional traits is a promising strategy for the development of sustainable microbial inoculants. Bacillus cabrialesii subsp. cabrialesii strain TE3T, a previously recognized plant growth-promoting and biological control agent, was subjected to UV mutation induction to improve these agro-biotechnological traits. Dilutions were made which were spread on Petri dishes and placed under a 20 W UV lamp at 10-min intervals for 60 min. After the UV-induced mutation of this strain, 27 bacterial colonies showed morphological differences compared to the wild-type strain; however, only a strain named TE3T-UV25 showed an improvement in 53.6% of the biocontrol against Bipolaris sorokiniana vs. the wild-type strain, by competition of nutrient and space (only detected in the mutant strain), as well as diffusible metabolites. Furthermore, the ability to promote wheat growth was evaluated by carrying out experiments under specific greenhouse conditions, considering un-inoculated, strain TE3T, and strain TE3T-UV25 treatments. Thus, after 120 days, biometric traits in seedlings were quantified and statistical analyses were performed, which showed that strain TE3T-UV25 maintained its ability to promote wheat growth in comparison with the wild-type strain. On the other hand, using bioinformatics tools such as ANI, GGDC, and TYGS, the Overall Genome Relatedness Index (OGRI) and phylogenomic relationship of mutant strain TE3T-UV25 were performed, confirming that it changed its taxonomic affiliation from B. cabrialesii subsp. cabrialesii to Bacillus subtilis. In addition, genome analysis showed that the mutant, wild-type, and B. subtilis strains shared 3654 orthologous genes; however, a higher number of shared genes (3954) was found between the TE3T-UV25 mutant strain and B. subtilis 168, while the mutant strain shared 3703 genes with the wild-type strain. Genome mining was carried out using the AntiSMASH v7.0 web server and showed that mutant and wild-type strains shared six biosynthetic gene clusters associated with biocontrol but additionally, pulcherriminic acid cluster only was detected in the genome of the mutant strain and Rhizocticin A was exclusively detected in the genome of the wild-type strain. Finally, using the PlaBase tool, differences in the number of genes (17) associated with beneficial functions in agroecosystems were detected in the genome of the mutant vs. wild-type strain, such as biofertilization, bioremediation, colonizing plant system, competitive exclusion, phytohormone, plant immune response stimulation, putative functions, stress control, and biocontrol. Thus, the UV-induced mutation was a successful strategy to improve the bioactivity of B. cabrialesii subsp. cabrialesii TE3T related to the agro-biotecnology applications. The obtained mutant strain, B. subtilis TE3T-UV25, is a promising strain to be further studied as an active ingredient for the bioformulation of bacterial inoculants to migrate sustainable agriculture.