牙鲆海水循环养殖系统生物膜上3种细菌的数量与代谢活性

Metabolic responses and quantitative counts of heterotrophic bacteria,ammonifiers and nitrifiers in moving bed bioflter bio?lms in a marine recirculating aquaculture system

从牙鲆(Paralichthysolivaceus)海水循环养殖系统中移动床生物滤器载体生物膜上获取异养细菌、氨化细菌和硝化细菌样本。异养细菌用平板涂布法计数,其代谢活性以培养8h的氧吸收速率表示。氨化细菌、氨氧化细菌(AOB)和亚硝酸氧化细菌(NOB)用MPN法计数,其代谢活性分别以培养24hNH4+-N浓度的增加、NH4+-N浓度的减少和NO2--N浓度的减少表示。结果表明,生物膜成熟之后,4种生理类群细菌的数量和代谢活性基本保持稳定。异养细菌数量为107～108CFU-·-m-2(载体),氨化细菌、AOB和NOB数量分别为107～108MPN-·-m-2(载体)、105～106MPN-·-m-2(载体)和105～106MPN-·-m-2(载体)。分批培养测得异养细菌的氧吸收速率、氨化细菌的氨化速率、AOB的氨氧化速率和NOB的亚硝酸氧化速率分别是0.591～0.738g(O2)-·-m-2-·-d-1、0.081～0.135g(NH4+-N)-·-m-2-·-d-1、0.017～0.031g(NH4+-N)-·-m-2-·-d-1和0.020～0.038g(NO2--N)-·-m-2-·-d-1。异养细菌的氧吸收速率、氨化细菌的氨化速率和AOB的氨氧化速率与异养细菌、氨化细菌和AOB的数量呈正相关。移动床生物滤器生物膜的平均氨和亚硝酸盐去除速率为(0.121±0.076)g(NH4+-N)-·-m-2-·-d-1和(0.139±0.181)g(NO2--N)·-m-2-·-d-1。同时测定了底物浓度和pH对氨化细菌、AOB和NOB代谢活性的影响。结果显示,氨化细菌的氨化速率、AOB的氨氧化速率和NOB的亚硝酸氧化速率与底物浓度(蛋白胨、NH4+-N和NO2--N)呈线性相关,pH为8.0时3种生理类群细菌的代谢活性较pH6.5或pH9.5时高。

In the present study, viable numbers and metabolic activities of aerobic heterotrophic bacteria, ammonifiers, ammonia oxidizing bacteria （AOB）, nitrite oxidizing bacteria （NOB） in marine biofilms were investigated. The studied biofilms were associated with media of a moving bed biofilter in a marine aquaculture recirculating system. The system was stocked with Japanese flounder （Paralichthys olivaceus）. Numbers of heterotrophic bacteria were determined by plate counts and metabolic activities were measured oxygen uptake rate （OUR） determination during eight hours incubation. Numbers of ammonifiers, AOB and NOB involved in N metabolism were evaluated by the most probable number-test （MPN） method. Their metabolic activities were estimated by calculating the increase in NH4^＋-N concentration, the decrease in NH4^＋-N and NO2^--N concentrations during 24 h incubation. Results showed that the numbers and activities of the four physiological groups of bacteria kept relatively stable after biofilm maturation. The numbers of heterotrophic bacteria expressed as colony-forming units per square meter of media and the MPN of ammonifiers, AOB and NOB per square meter of media were 10^7-10^8, 10^7-10^8, 10^5-10^6 and 10^5-10^6, respectively. In batch culture, the OURs of heterotrophic bacteria, the ammonification rates of ammonifiers, the ammonia oxidizing rates of AOB and the nitrite oxidizing rates of NOB ranged from 0.591-0.73 8 g（O2） · m^-2· d^-1, 0.081-0.135 g （NH4^＋-N） · m^-2· d^-1, 0.017-0.031 g （NH4^＋-N） · m^-2 · d^-1 and 0.020-0.038 g （NO2^-N）· m^-2· d^-2, respectively. The oxygen uptake rates of heterotrophic bacteria, thea mmonification rates of ammonifiers and the ammonia oxidizing rates of AOB were correlated with the numbers of heterotrophic bacteria, ammonifiers and AOB. The mean ammonia and nitrite removal rates of the moving bed biofilter were （0.121 ＋ 0.076） g （NH4^＋-N）· m^-2· d^-1 and （0.139 ＋ 0.181） g （NO2^-N）·m^-2· d^-1 calculated according to the difference between NH4^＋-N, NO2^--N inlet and outlet biofilter concentrations. Furthermore, influences of substrate concentration and pH on metabolic activities of ammonifiers and nitrifiers （AOB and NOB） were detected. Results showed that ammonification rate, ammonia- and nitrite-oxidizing rates were linearly correlated with substrate concentrations （peptone, NH4^＋-N or NO2^--N）, and at pH 8.0 higher ammonification and nitrogen conversion rates were Rot than those of pH 6.5 and pH 9.5.