Enrichment Of Denitrifying Anaerobic Methane Oxidizing Microorganisms In A Hollow Fiber Membrane Bioreactor

Anaerobic oxidation of methane (AOM) is a critical process for controlling the flux of methane from anoxic environments. Recently, it was found that the process of denitrifying anaerobic methane oxidation (DAMO) could happen in freshwater environment. The DAMO process is catalyzed by DAMO bacteria called NC10with or without the involvement of DAMO archaea (ANME). However, for microbes involved in which, strictly confortable conditions are necessary, their doubling times are long and pure culture is difficult to be obtained. The enrichment of DAMO microbes often takes a long time (5-16months) to show a faint denitrification function in reported DAMO process, and the denitrification rate is low. This may be caused by the low solubility of methane which limits its mass transfer in the DAMO process.This research aims to fast enrich DAMO process in a hollow fiber membrane bioreactor (HFMB), with the special structure of fiber in the HFMB to increase the mass transfer of methane. Experimental inoculum came from the Ronggui river in the Pearl River Delta, Lake Thai and Dushu lake in Yangtze River Delta, respectively. It was found that the HFMBs inoculated with the three different sediments were all successful to show DAMO activities in a very short period of time (<3months). The denitrification rate of Ronggui reactor was30mg N L-1d-1, the denitrification rate of Dushu reactor once reached60mg N L-1d-1before inhibition, and was40mg N L-1d-1eventually, Thai reactor got the highest denitrification rate of60mg N L-1d-1without any suppressed. These denitrification rates were higher than the reported ones. Compared with batch experiments in serum bottles, the HFMB could tolerate nitrate concentration up to200mg N L-1, while the former was less than30mg N L-1with the same biomass concentration, However, when the nitrate concentration was higher than240mg N L-1, the DAMO activity in the HFMB was obviously inhibited by the accumulated nitrite.The analysis of phylogenetic analysis of the microbes from Thai reactor found that two kinds of bacteria dominated in the reactor, one had a long distance relationship with the DAMO bacteria, another belonged to proteobacteria, while only one kind of archaea existed in the reactor, which were similar to AMO archaea (ANME-1and ANME-2). Microbial analysis of Ronggui and Dushu reactors illustrated that microbes in the two reactors were more complex, DAMO archaea were dominant with40%and37%, respectively. Proteobacteria were about12%, and other bacteria (45-50%) were anaerolineaceae, protebacteria and desulfurellaceae. Anaerolineaceae and protebacteria often appear in the sewage plant and anaerobic ammonia oxidation environment. The existence of desulfurellaceae might be due to the sulphate in the medium. There were less than4%of methylococcus (aerobic methane oxidation bacteria) in the two reactors. So, likely AMO coupled to sulphate reduction and aerobic methane oxidation might happen in the two reactors.