The doubling time of anaerobic ammonium-oxidizing (anammox) bacteria in an anaerobic biological filtrated (ABF) reactor was determined. Fluorescence in situ hybridization analysis was used to detect and count anammox bacteria cells in anammox sludge. As a result, the populations of anammox bacteria at 14th and 21st days were 1.1 x 10(6) and 1.7 x 10(7) cells/ml reactor, respectively. From these results, the doubling time of anammox bacteria was calculated as 1.8 days, and the specific growth rate (mu) was 0.39 day(-1). This result indicated that the anammox bacteria have higher growth rate than the reported value (doubling time, 11 days). Furthermore, it was clearly demonstrated that nitrogen conversion rate was proportional to the population of anammox bacteria. Maintaining the ideal environment for the growth of anammox bacteria in the ABF reactor might lead to faster growth. This is the first report of the growth rate of anammox bacteria based on the direct counting of anammox bacteria.
Anaerobic ammonium-oxidizing (anammox) bacteria were immobilized in polyethylene glycol gel carriers. A small amount of seed sludge [0.24% (w/v)] was entrapped in the carriers, and continuous feeding tests were performed. Nitrogen removal activity increased gradually, reaching 3.7 kg N/m(3) reactor per day on day 67. The average of nitrogen conversion rate was calculated as 3.4 kg N/m(3) reactor per day. Microscopic examination clearly showed that small red clusters formed in the gel carrier. Moreover, fluorescence in situ hybridization analysis revealed that these clusters consisted of anammox bacteria. From real-time polymerase chain reaction analysis, the growth of anammox bacteria in the gel carriers was clearly shown by increased concentration of 16S rRNA gene of planctomycete from 4.3 x 10(8) to 4.2 x 10(9) copies/ml between days 41 and 55. To determine the effects of inoculation on the start-up of the reactor, the amount of seed sludge in the gel carrier was varied and it was found that the start-up period could be reduced to as little as 25 days when a sludge concentration of 1.4% (w/v) was used. This is the first report of successful immobilization and cultivation of anammox bacteria in a gel carrier.
An anaerobic ammonium oxidation (anammox) process for ammonia-rich wastewater treatment has not been reported at temperatures below 15 degrees C. This study used a gel carrier with entrapped anammox bacteria to obtain a stable nitrogen removal performance at low temperatures. In a continuous feeding test, a high nitrogen conversion rate (6.2 kg N m(-3) day(-1)) was confirmed at 32 degrees C. Nitrogen removal activity decreased gradually with decreasing operation temperature; however, it still occurred at 6 degrees C. Nitrogen conversion rates at 22 and 6.3 degrees C were 2.8 and 0.36 kg N m(-3) day(-1), respectively. Moreover, the stability of anammox activity below 20 degrees C was confirmed for more than 130 days. In batch experiments, anammox gel carriers were characterized with respect to temperature. The optimum temperature for anammox bacteria was found to be 37 degrees C. Furthermore, it was clear that the temperature dependence changed at about 28 degrees C. The apparent activation energy in the temperature range from 22 to 28 degrees C was calculated as 93 kJ mol(-1), and that in the range from 28 to 37 degrees C was 33 kJ mol(-1). This value agrees with the result of a continuous feeding test (94 kJ mol(-1), between 6 and 22 degrees C). The nitrogen removal performance demonstrated at the low temperatures used in this study will open the door for the application of anammox processes to many types of industrial wastewater treatment.
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