Bacillus krulwichiae sp. nov., a halotolerant obligate alkaliphile that utilizes benzoate and m-hydroxybenzoate

Researchers isolated two bacterial strains, AM31DT and AM11D, from Japanese soil that represent a novel species of obligate alkaliphile capable of degrading aromatic compounds. These Gram-positive, rod-shaped bacteria belonging to the genus Bacillus grow optimally at pH 8-10 but cannot survive at neutral pH, and tolerate salt concentrations up to 14% NaCl. Notably, both strains utilize benzoate and m-hydroxybenzoate as sole carbon sources—the first alkaliphilic Bacillus strains documented to degrade these aromatic compounds. Phylogenetic analysis based on 16S rRNA gene sequencing placed the isolates in the alkaliphile group within Bacillus, with highest similarity to Bacillus alcalophilus and Bacillus pseudalcaliphilus. However, DNA-DNA hybridization demonstrated low relatedness (less than 19.3%) to these phylogenetically close neighbors. The strains contained characteristic menaquinone quinones and branched-chain fatty acids typical of Bacillus species. Based on combined phenotypic characteristics, phylogenetic data, and genetic relatedness measurements, the authors established a new species, Bacillus krulwichiae, with strain AM31DT designated as the type strain. This discovery expands understanding of alkaliphilic bacteria capable of aromatic compound degradation and provides potential sources for enzymes functioning in alkaline conditions.

Key findings

• Two obligate alkaliphilic Bacillus strains capable of utilizing benzoate and m-hydroxybenzoate as carbon sources were isolated, representing the first documented alkaliphilic Bacillus strains with aromatic compound degradation ability.
• 16S rRNA gene sequencing and phylogenetic analysis placed the isolates in Bacillus group 6 (alkaliphiles), most closely related to B. alcalophilus and B. pseudalcaliphilus, but DNA-DNA hybridization showed low relatedness (<19.3%) confirming species distinction.
• The novel species Bacillus krulwichiae exhibits obligate alkaliphily (growth at pH 8-10 but not pH 7), halotolerance (up to 14% NaCl), and characteristic biochemical properties including menaquinone-5/-6/-7 as major quinones and branched-chain fatty acids.
• Both strains are facultatively anaerobic, Gram-positive, spore-forming rods with peritrichous flagella that produce starch-, DNA-, and Tween hydrolysis but differ slightly in protease activity.
• Type strain AM31DT has DNA G+C content of 41.5 mol%, placing it within the Bacillus genus range, and grows strongly on both aromatic compounds, while AM11D shows greater preference for benzoate.