Metabolic and genetic basis of heterotrophic nitrification and aerobic denitrification coupled with phosphorus...

Eutrophication caused by excess nitrogen and phosphorus requires efficient biological removal strategies. This study investigated Comamonas testosteroni ZSJS, a heterotrophic nitrifying-aerobic denitrifying (HNAD) bacterium capable of simultaneous nitrogen and phosphorus removal under aerobic conditions. Under optimal parameters (P/N = 0.1, C/N = 20, 30 °C, pH 7.0, 160 rpm), the strain removed > 97% of NH₄⁺-N and 99.85% of PO₄^3--P and utilized NH₄⁺-N, NO₂^--N, or NO₃^--N as sole nitrogen sources. Removal kinetics of nitrogen, phosphorus, and COD were well described by the modified Gompertz model (R² > 0.9). Nitrogen was partly assimilated into biomass and partly converted to gaseous products. Functional genes associated with nitrogen metabolism (napAB, nirK, amoC) and phosphorus metabolism (ppk, ppx, pst) were expressed. Phosphorus removal occurred exclusively under aerobic conditions and strongly depended on dissolved oxygen, with most phosphorus incorporated into extracellular polymeric substances. Cross-regulation between nitrogen and phosphorus metabolism was observed: moderate phosphorus levels enhanced nitrogen metabolism, whereas excessive phosphorus suppressed ammonia assimilation; conversely, nitrogen availability influenced phosphorus-related gene expression, particularly ppk and pst. Overall, strain ZSJS demonstrates a robust aerobic pathway for integrated nitrogen and phosphorus removal, highlighting its potential for simplified and efficient wastewater treatment processes.