ABSTRACT Aceticlastic and hydrogenotrophic methanogens were comparatively investigated by 16S rRNA‐based fluorescence in situ hybridization (FISH) technique combined with Confocal Scanning Laser Microscopy (CSLM) to evaluate temperature influence on two functional groups of methanogens. Microorganisms were grown in the form of anaerobic granular sludge in two lab‐scale Upflow Anaerobic Sludge Bed (UASB) reactors fed with synthetic wastewater and operated at psychrophilic and mesophilic temperature conditions. Since hydrogenotrophic and aceticlastic methanogenesis are known to play a crucial role, digestion may be effectively blocked if these processes are ceased due to unfavorable environmental temperature conditions. Mesophilic (35 ± 2C) and psychrophilic (10 ± 1C) reactors were inoculated by the same mesophilic granules taken from a full‐scale UASB reactor. According to FISH results, Archaea representing methanogens were found dominating at bottom sampling ports of reactors and dominant member of Archaea was the genus aceticlastic Methanosaeta (MX825). Other aceticlastic methanogen such as Methanosarcina‐like (SARCI645) species were also identified which were higher in psychrophilic granules. Members of order Methanobacteriales (MB310) constituted the major hydrogenothrophic methanogens in both reactors, whereas other hydrogenothrophic methanogens members of order Methanococcales (MC1109) and Methanogenium (MG1200) relatives were absent. Abundance of originally mesophilic Methanosaeta‐related Archaea under low temperature conditions demonstrated the adaptive capacity of microorganisms to psychrophilic conditions. This might be attributed to enzymatic alterations in Methanosaeta cells originating from seed sludge, which were exposed to sub‐mesophilic temperatures at start‐up and then to psychrophilic conditions during gradual temperature decreases. Both sludge granules remained undisturbed and kept their rigidity even after exposure to very low temperatures. Quantification study was done using Image Pro‐plus Version 4.0. Archaeal cells detected with probe ARC915 were found around 60% of the 4,6‐diamidino‐2‐phenylindole stained cells at both reactors. Methanosaeta spp./Archaea ratios for mesophilic and psychrophilic UASB reactors were calculated as 71% and 84%, respectively. FISH and CSLM results together with quantification study showed that the microbial community of anaerobic granular sludges was dominated by Methanosaeta spp. which was identified as the major methanogenic Archaea.PRACTICAL APPLICATIONSSince effects on bioreactor performances were correlated to variations in microbial diversity, investigation of methanogenic diversity in anaerobic reactors is important during operation. Identification of microorganisms should be done for formation, stability, and physiological properties of the biomass in bioreactors, which is a key factor during process performance evaluation. Monitoring the changes in dominant archea of seed added at start‐up, gives information on system performance. A detailed investigation on diversity, structure and function of mixed microbial communities in anaerobic reactors is necessary to improve treatment efficiency and stability against inhibitory compounds. Among 16S rRNA based microbial identification techniques, FISH is a well‐known technique that helps determination of dominant species by giving a detailed analysis of microbial populations while CSLM is the most appropriate method for monitoring the formation of 3‐D structure of microbial biomass. In this study, temperature influence on aceticlastic and hydrogenotrophic methanogen groups was evaluated using FISH coupled with CSLM.

Effect of low temperature on up-flow anaerobic sludge bed (UASB) reactor performance treating raw sewage was investigated in terms of the variations in methanogenic diversity using the 16S rRNA based Fluorescence In-Situ Hybridization (FISH) technique. The diversity of microorganisms present in the anaerobic granular sludge and the structure of the granules operated at 13 degrees C have been investigated using FISH combined with CSLM (Confocal Scanning Laser Microscopy). According to FISH results, archaeal cells representing methanogens were found intensively dominant in the bottom sampling port of the UASB reactor and acetoclastic Methanosaeta was the abundant methanogen. Other methanogens such as Methanosarcina and Methanobacterium like species were also observed. The abundance of originally mesophilic Methanosaeta-related Archaea under low temperature at all sampling days revealed the microbial adaptation to psychrophilic conditions. This might be attributed to the enzymatic alterations in Methanosaeta cells originating from seed sludge, which were exposed to sub-mesophilic temperatures at start-up and then to psychrophilic conditions during gradual decreases of temperature. According to CSLM observation, even though the sludge retained in the reactor kept its granular form as a whole, the majority of the granules had a tendency to be partly broken and they lost their rigidity when raw sewage was fed following synthetic sewage. Besides, Methanosaeta related species prevailing in seed sludge have noticeably lost their long filamentous forms and deteriorated during raw sewage feeding. Members of the order Methanobacteriales constituted the major hydrogenothrophic methanogens present in the psychrophilic UASB reactor, whereas the other hydrogenothrophic methanogens-members of the order Methanococcales and Methanogenium relatives-were absent.

Istanbul has experienced rapid increases in population to more than 12 million people, which has created infrastructure problems of water supply and wastewater treatment and disposal. In this article, the achievements and approaches of the Istanbul Water and Sewerage Administration (ISKI) to solve the water shortage problem and to improve services are summarized. Istanbul had a very severe water shortage problem in 1994 because of ignorance of the implementation of the needed projects. After reviewing the reasons and causes of the problem, new priority criteria adopted after 1994 are given. Following the implementation of the projects determined according to the aforementioned criteria, water supplied has exceeded the water demand. The added capacity is equal to one to three times of the capacity built up to 1994 for water treatment, service reservoirs, pumping stations, transmission lines, and the water distribution network; water quality has been improved the meet local and international potable water standards. Unaccounted for water has been reduced from 60% to 27%. The percentage of treated wastewater has been increased from 10% to 90% in 8 years, resulting in drastic improvements and rehabilitation of the Golden Horn and coastal water quality. Through improved customer services, complaints were reduced from 33% in 1994 to 0.3%. Some of the main criteria and the approaches behind this success are summarized.