The strong biocidal effect of free nitrous acid on anaerobic sewer biofilms

Several recent studies showed that nitrite dosage to wastewater results in long-lasting reduction of the sulfate-reducing and methanogenic activities of anaerobic sewer biofilms. In this study, we revealed that the quick reduction in these activities is due to the biocidal effect of free nitrous acid (FNA), the protonated form of nitrite, on biofilm microorganisms. The microbial viability was assessed after sewer biofilms being exposed to wastewater containing nitrite at concentrations of 0-120 mg-N/L under pH levels of 5-7 for 6-24 h. The viable fraction of microorganisms was found to decrease substantially from approximately 80% prior to the treatment to 5-15% after 6-24 h treatment at FNA levels above 0.2 mg-N/L. The level of the biocidal effect has a much stronger correlation with the FNA concentration, which is well described by an exponential function, than with the nitrite concentration or with the pH level, suggesting that FNA is the actual biocidal agent. An increase of the treatment from 6 to 12 and 24 h resulted in only slight decreases in microbial viability. Physical disrupted biofilm was more susceptible to FNA in comparison with intact biofilms, indicating that the biocidal effect of FNA on biofilms was somewhat reduced by mass transfer limitations. The inability to achieve 2-log killing even in the case of disrupted biofilms suggests that some microorganisms may be more resistant to FNA than others. The recovery of biofilm activities in anaerobic reactors after being exposed to FNA at 0.18 and 0.36 mg-N/L, respectively, resembled the regrowth of residual sulfate-reducing bacteria and methanogens, further confirming the biocidal effects of FNA on microorganisms in biofilms.