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Phenotypic and Genomic Characterization of Bacillus sp. DLB3, a Black Pepper Rhizosphere Isolate with Antagonistic, Abiotic Stress-Tolerant, and Antibiotic-Susceptible Traits

Year: 2026

Type: article

Abstract: Rhizosphere bacteria combining biocontrol activity with resilience to environmental stress are valuable candidates for sustainable agriculture. This study characterized strain DLB3, isolated from black pepper rhizosphere soil, through dual-culture and disk-diffusion antagonism assays, enzymatic and siderophore screening, physiological tolerance tests, antibiotic susceptibility testing, whole-genome sequencing, and genome mining. Phylogenomic analysis and average nucleotide identity (>97%) and digital DNA–DNA hybridization (>79%) values identified the isolate as Bacillus velezensis. The strain inhibited Fusarium falciforme, Fusarium oxysporum, and Colletotrichum gloeosporioides (47.6–71.2% radial growth inhibition) and produced an inhibition zone (8.4 mm) against Ralstonia solanacearum, alongside extracellular protease, amylase, CMCase, and catecholate-type siderophore production. DLB3 tolerated a broad range of temperatures (20–50 °C, with optimal growth at 25–40 °C, peaking at 35 °C), pH (5–9), and salinity (up to 10% NaCl), and exhibited minimum inhibitory concentrations (MICs) of 1800, 500, 450, and 250 mg L−1 for Pb, Zn, Cu, and Cr, respectively, while remaining susceptible to all nine antibiotics tested. The 3,913,361 bp draft genome was assembled into 44 contigs (N50 = 990,239 bp; L50 = 2), with 99.81% completeness and 0.12% contamination according to CheckM. It harbored 18 biosynthetic gene clusters, carbohydrate-active enzymes, peptidases, and genes associated with heavy metal resistance, broadly consistent with the observed phenotypes, as well as a homolog of the cfr methyltransferase gene (97.4% identity) that was not phenotypically expressed. These findings provide a comprehensive genomic and phenotypic profile of strain DLB3 and its biocontrol-related potential.

Source: International Journal of Molecular Sciences

DOI: 10.3390/ijms27219536