Isolation, Identification And Immobilization Of Novel Denitrification Strains

Water eutrophication is one of major problems of water environment in our country, and the nitrogen is the main pollution factor. There are several ways for wastewater denitrification, and the biological denitrification is considered as one of most cost-effective methods. Traditional biological denitrification theory suggests that the wastewater denitrification is mainly completed by a group of autotrophic, aerobic nitrifying bacteria and a group of heterotrophic, anaerobic denitrifying bacteria. However, in practice, it is discovered that there are many defects for these two kinds of bacteria used in the denitrifying process. So, in recent years the research and application of heterotrophic nitrifier and aerobic denitrifier being a hot spot, and it is expected to overcome the shortcomings of the traditional nitrogen removal process and enrich the biological denitrification theory. During the applied study of biology denitrifying technology, immobilization technology has been attracted attention beacuse of the superiority of itself. As one of immobilization methods, microcapsule immobilized has been concerned because of its good stabiliy, biocompatibility and mass trasfer. But the synchronous microcapsule immobilization of the heterotrophic nitrifier and aerobic denitrifier has not been repoted.We obtained 26 activated sludged samples from sewage plants, chemical fertilizer plants and farmland. After performing a series of stemps, finally a high-efficient heterotrophic nitrifier and a high-efficient aerobic denitrifier were obtained. We identified the strains and studied their denitrification conditions and characteristics. Then we mixed the two kinds of strains, and optimized the mixed strains denitrification conditions, investgated the denitrification effects in real wasrewater, synchronous immobilized the mixed strains by microcapsules.The results of this study are as follows.1. After sludge domestication, gradient dilution of domestication liquid, isolation from streaking plate, utilization of color indicator as rapid denitrification detection method, finally a high-efficient heterotrophic nitrifier and a high-efficient aerobic denitrifier were obtained. The two strains were identified as Alcaligenes sp. and Acinetobacter lwoffii respectively, and named as Alcaligenes sp. HN-S and Acinetobacter lwoffii DN-S. 2. HN-S was a obligate heterotrophic nitrifying bacteria, no autotrophic nitrification capacity. In the artificial wastewater with 182.30 mg/L ammonia nitrogen as initial concentration, the removal efficiency by the strain was 99.8% after 30 h cultivation. The average nitrogen removal rate was 9.61 mg-N/L/h in its exponential phase, faster than the most heterotrophic nitrifiers that screened before. It produced almost no NO2--N and NO3--N in the entire nitrification process. The optimal carbon source is sodium citrate. Higher dissolved oxygen and C/N ratio favored its nitrification. When temperature is ranged from 30℃to 35℃and pH is ranged from 5.0 to 9.0, it can completely remove ammonia nitrogen. It was able to denitrify when nitrite nitrogen was used as nitrogen source, but it could not utilized nitrate nitrogen. The most removed ammonia nitrogen was converted to N2. This was different from most current coveraged heterotrophic nitrifier.3. In the artificial wastewater with 166.05 mg/L nitrate nitrogen as initial concentration, the removal efficiency by the DN-S was 99.8% after 72 h cultivation. The average nitrogen removal rate was 8.2 mg-N/L/h in its exponential phase. It produced almost no NO2--N in the entire denitrification process. Higher dissolved oxygen favored its denitrification. The optimal C/N ratio was ranged from 9 to 12. The most removed nitrate nitrogen was converted to N2. DN-S was s simultaneous nitrification and denitrification bacteria, it also could utilize ammonia nitrogen and nitrite nitrogen as its nitrogen source.4. We mixed the HN-S and DN-S, and optimized the culture conditions of the mixed strains by orthogonal test. The most optimal nitrogen,removal condition was as follows:the inoculum concentration proportion of HN-S and DN-S was 1:2, C/N=12.5, temperature was 35℃, pH=9.0, dissolved oxygen was 2.5mg/L, Nitrogen removal ability of mixed strains was evaluated in real wastewater. Compared with activated sludge, the mixed strains had a stronger ability to adapt to different wastewater. By adding sodium citrate can significantly promote their denitrification capacity.5. We immobilized the mixed strains by microcapsules and used they to treatment artificial wastewater. The results showed that the denitrification speed of immobilized bacteria was lower than the mobilized bacteria, but its denitrification lasting longer, the production of bacteria mud was smaller, and the nitrogen assimilation was lower, thus improved the total nitrogen removal rate. Immobilization also improved the capacity of strains to resist the bad environment.