Elucidation of Ammonia inhibition in anaerobic treatment process by using 16S rRNA /DNA based microbial identification techniques

Abstract

A high ammonia landfill leachate was anaerobically treated for more than 1000 days in laboratory scale two different anaerobic reactors configured as sludge blanket and hybrid bed. Effects of high ammonia concentrations on reactor performances were correlated to the variations in methanogenic diversity by using 16S rRNA/DNA based microbial identification techniques such as FISH (fluorescent in-situ hybridization), DGGE (denaturing gradient gel electrophoresis), cloning and DNA sequencing. However, sudden and unexpected fluctuations in the characteristics of leachate resulted in complications during evaluation of results. Therefore, five laboratory scale anaerobic reactors seeded with different sludges were operated for 450 days under stable COD loads and gradually increasing total ammonia levels from 1000 to 6000 mg/l. Methanosaeta-related species mainly prevailing in seed sludges were substituted for Methanosarcina when FAN (free ammonia nitrogen) exceeded 100 mg/l. Subsequently, as FAN level elevated, rather than any shifts in the methanogenic community, single coccus shaped Methanosarcina cells formed stringent multicellular packets. However, when the FAN level exceeded 600 mg/l, disintegration of Methanosarcina clusters was observed. In contrast, inorganic particles originated from seed sludge provided a good support for Methanosarcina clusters in one reactor and with this special feature it successfully resisted to FAN level as high as 750 mg/l.