Screening and genome-wide analysis of lignocellulose-degrading bacteria from humic soil

Abstract

Crop straw contains huge amounts of exploitable energy, and efficient biomass degradation measures have attracted worldwide attention. Mining strains with high yields of cellulose-degrading enzymes is of great significance for developing clean energy and industrial production of related enzymes. In this study, we reported a high-quality genome sequence of Bacillus velezensis SSF6 strain using high-throughput sequencing technology (Illumina PE150 and PacBio) and assessed its lignocellulose degradation potential. The results demonstrated that the genome of B. velezensis SSF6 was 3.89 Mb and contained 4,015 genes, of which 2,972, 3,831 and 158 genes were annotated in the COGs (Clusters of Orthologous Groups), KEGG (Kyoto Encyclopedia of Genes and Genomes) and CAZyme (Carbohydrate-Active enZymes) databases, respectively, and contained a large number of genes related to carbohydrate metabolism. Furthermore, B. velezensis SSF6 has a high cellulose degradation capacity, with a filter paper assay (FPA) and an exoglucanase activity of 64.48 ± 0.28 and 78.59 ± 0.42 U/mL, respectively. Comparative genomic analysis depicted that B. velezensis SSF6 was richer in carbohydrate hydrolase gene. In conclusion, the cellulose-degrading ability of B. velezensis SSF6 was revealed by genome sequencing and the determination of cellulase activity, which laid a foundation for further cellulose degradation and bioconversion.

Introduction of Cellulolytic Bacterium Bacillus velezensis Z2.6 and Its Cellulase Production Optimization

“…This list compromised two endoglucanases (EC 3.2.1.4) from the GH51 family and nine β-glucosidases (EC 3.2.1.21), among which seven and two were categorized in families GH1 and GH3, respectively. The co-existence and approximate quantities of these annotated related proteins indicated the potential of strain Z2.6 in cellulose degradation, consistent with other B. velezensis strains such as FZB42 T and SSF6 [14]. To validate the authenticity of this finding, five fragments of genes (V7S33_01155, V7S33_4525, V7S33_5150, V7S33_13785, V7S33_13965) were cloned and sequenced (Table 2).…”

“…Under light microscopy by Gram staining (Figure S1c), Z2.6 was Gram-positive and short and rod-shaped with endospores forming. This strain was initially selected based on the comparison of the cellulose hydrolysis zone (H) to the colony size (C), with reference to the superiority in H/C ratio and increasing speed along time [14]. Strain Z2.6 met both parameters with final average H/C being 5.45 ± 0.82 on day 5 (Table S5).…”

“…The BLAST results declared that V7S33_05150 could be regarded as an endoglucanase (EC 3.2.1.4) with 100% identity, while V7S33_13785 belonged to β-glucosidase (EC 3.2.1.21) in strain Z2.6 with 99.57% identities. Notably, V7S33_05150 was detected with a carbohydrate-binding site, which may enhance carbohydrate catalytic abilities [14,52], and V7S33_13965 also contained a signal peptide, suggesting a possible extracellular role in hydrolyzing (1->4)beta-D-glucosidic linkages. These genes all exhibited similarities above 99.5% against sequences in other B. velezensis strains.…”

“…Following the completion of the eight parameters confirmed using the one-factor-ata-time design, we were next inclined to figure out predominant factors exerting higher contributions to cellulase production. Recent investigations on B. velezensis mainly centered on the isolation, characterization, and optimization of single fermentation conditions at one time [5,14,18,19], while a limited number of studies extended their analyses to incorporate response surfaces aimed at more efficient cellulase production [3,53]. To find the three most key factors among the eight factors, of which the carbon and nitrogen source were already affirmed as CMC-Na and tryptone, a PB design was used for this duty, shown in Table 3, where the response results are shown.…”

“…The utilization of these cellulose-decomposing enzymes presents promising prospects in the biorefinery and biologics industries [12]. Among the numerous beneficial attributes of such organisms, B. velezensis displays a wide spectrum of abilities in biocontrol, hydrolase production, and probiotics, all of which have substantial applications in the agriculture industry and relative utilization in pharmaceutics [13][14][15][16]. Nonetheless, recent applications of B. velezensis from recycled compost for secondary usage following fermentation have not been explored much.…”