Genomic and physiological characterization of Neobacillus ammoniipjilus sp. nov. reveals versatile inorganic nitrogen utilization and ammonia-mitigation potential
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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 °C, pH 6.0–8.0, and 0–2.0% (w/v) NaCl. The major cellular fatty acids were iso-C 15:0 and anteiso-C 15:0 , 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).
Importance
Neobacillus species occur in diverse environments, but their capacity to use and transform multiple inorganic nitrogen forms remains poorly characterized. This study describes Neobacillus ammoniiphilus sp. nov. strain KR4-4ᵀ and links its taxonomic and genomic features with physiological nitrogen-utilization phenotypes. The strain used ammonium, nitrate, and nitrite as individual nitrogen sources, showed substantial ammonium removal under aerobic conditions, and enhanced the reduction of gas-phase ammonia in a sealed-flask assay. Genome analysis identified systems associated with inorganic nitrogen transport, nitrate and nitrite reduction, ammonium assimilation, and nitrogen regulation. These findings expand knowledge of nitrogen metabolism within Neobacillus and identify KR4-4ᵀ as a candidate for further evaluation in biological ammonia-mitigation applications.
