Genomic and physiological characterization of Neobacillus ammoniipjilus sp. nov. reveals versatile inorganic nitrogen utilization and ammonia-mitigation potential

A novel bacterial strain (KR4-4T), was isolated from soil in the Republic of Korea and subjected to polyphasic taxonomic and functional characterization. Phylogenetic analysis based on the 16S rRNA gene sequence placed strain KR4-4T within the genus Neobacillus. Phylogenomic analysis based on 92 bacterial core genes further supported its distinct taxonomic position. Average nucleotide identity and digital DNA-DNA hybridization values between KR4-4T and related Neobacillus type strains were 69.76-82.31% and 18.3-25.9%, respectively, supporting its recognition as a distinct species. Cells were Gram-stain-positive, non-motile, facultatively anaerobic rods, with growth occurring at 15-45 {degrees}C, pH 6.0-8.0, and 0-2.0% (w/v) NaCl. The major cellular fatty acids were iso-C15:0 and anteiso-C15:0, while the major polar lipids included diphosphatidylglycerol, phosphatidylglycerol, and phosphatidylethanolamine. Genome analysis revealed genes associated with inorganic nitrogen transport, nitrate and nitrite reduction, and ammonium assimilation. In liquid culture, KR4-4T removed NH4+-N under aerobic conditions and was able to utilize ammonium, nitrate, and nitrite as individual nitrogen sources, with removal efficiencies of 86.5%, 69.35%, and 62.9%, respectively. The strain also contributed to the reduction of gaseous NH3 in a closed-flask assay. Together, these physiological and genomic characteristics indicate a versatile capacity for inorganic nitrogen utilization and transformation and suggest potential for further evaluation in biological ammonia-mitigation applications. Based on phylogenetic, genomic, phenotypic, and chemotaxonomic evidence, strain KR4-4T represents a novel species of the genus Neobacillus, for which the name Neobacillus ammoniiphilus sp. nov. is proposed. The type strain is KR4-4T (= KACC 24003T = CCTCC AB 2025077T).