Abstract

A novel acid-tolerant HNAD bacteria, Acinetobacter soli HY24, was isolated. • Strain HY24 achieved efficient nitrogen removal under acidic conditions. • Assimilation was the main nitrogen metabolism of strain HY24. • The nitrogen removal pathway and acid-tolerant mechanisms of strain HY24 was clarified. The biological treatment of acidic wastewater presents a notable environmental engineering challenge. This study investigated a novel acid-tolerant bacterium, Acinetobacter soli HY24, focusing on its assimilation-dominated nitrogen removal, potential heterotrophic nitrification-aerobic denitrification (HNAD) capabilities, underlying nitrogen removal mechanisms, and the potential molecular basis for the coupling between acid tolerance and nitrogen removal. Optimal nitrogen removal was achieved under the following conditions: C/N ratio of 15, temperature of 30 °C, shaking speed of 150–200 rpm, pH 4.5, and sodium citrate as the carbon source. The maximum NH 4 + − N <math><msubsup is="true"><mi is="true">NH</mi><mn is="true">4</mn><mo is="true">+</mo></msubsup><mo is="true">−</mo><mi mathvariant="normal" is="true">N</mi></math> and NO 3 − − N <math><msubsup is="true"><mi is="true">NO</mi><mn is="true">3</mn><mo is="true">−</mo></msubsup><mo is="true">−</mo><mi mathvariant="normal" is="true">N</mi></math> removal efficiencies of strain HY24 reached 100% and 75.15 ± 6.38%, and the maximum removal rates were 14.92 ± 1.87 and 8.82 ± 0.57 mg/L h −1 at pH 4.5, respectively. Genome, nitrogen balance, intracellular amino acid, and RT-qPCR analyses revealed that cellular assimilation and HNAD serve as potential nitrogen removal pathways, among which assimilation was identified as the core pathway of strain HY24, accounting for 80.13 ± 4.37% ( NH 4 + − N <math><msubsup is="true"><mi is="true">NH</mi><mn is="true">4</mn><mo is="true">+</mo></msubsup><mo is="true">−</mo><mi mathvariant="normal" is="true">N</mi></math> ), 44.92 ± 3.47% ( NO 3 − − N <math><msubsup is="true"