Microbial ecology of activated sludge

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Microbial ecology of activated sludge Powered By Docstoc
     ecology of                            The activated sludge process relies on a                                                              an ideal model system for developing

      activated                            complex microbial community to clean
                                                                                                                                                 new methodology and for evaluating
                                                                                                                                                 hypotheses relating to the ecology of

                                                                                                                                                 Discovery of microbial key

        sludge                             up wastewaters and thus protect our
                                           environment. Michael Wagner explains
                                                                                                                                                 In developing a detailed theoretical
                                                                                                                                                 framework for activated sludge micro-
                                                                                                                                                 biology, it is important to find out about
                                                                                                                                                 the identity and ecophysiology of the
                                           how new molecular tools are shedding light                                                            micro-organisms responsible for key
                                                                                                                                                 processes or major problems in waste-
                                           on this ecosystem and how the information                                                             water treatment plants. Only if we can
                                                                                                                                                 measure the abundance of desirable
                                                                                                                                                 or undesirable populations will we be
                                           can be used to improve the treatment                                                                  able to decipher (and later exploit) con-
                                                                                                                                                 ditions that might selectively promote
                                           process.                                                                                              their success or failure, and thus ration-
                                                                                                                                                 ally influence community assembly.
                                                                                                                                                 Traditional cultivation methods are
                                                                                                                                                 inadequate for identifying the key play-
                                                       he activated sludge pro-               activated sludge research has been                 ers, since only 10–15 % of the activated

                                                       cess was invented about a              dominated for decades by engineers                 sludge micro-organisms form colonies
                                                       century ago and has found              and until quite recently little was                on standard nutrient agar plates.
                                                       worldwide application in               known about which micro-organisms                  This unintentional selectivity has
                                                       the treatment of wastewater            were important in these man-made                   led to an incomplete and sometimes
                                                       before it is discharged into           systems and how changes in environ-                even erroneous perception of which
                                           the environment. Modern wastewater                 mental parameters or process design                bacterial species catalyse such import-
                                           treatment plants use this process not              affect the structure and function of the           ant steps as nitrification, denitrification
                                           only to remove organic matter from                 microbial community. This situation                and enhanced biological phosphorus
                                           the sewage, but also to minimize the               has changed dramatically during the                removal, or what the causative agents
                                           concentrations of nitrogen and phos-               last few years thanks to exciting                  for activated sludge bulking and
                                           phorus compounds which stimulate                   technical advances in microbial ecol-              foaming are. For example, Nitrobacter
                                           eutrophication in lakes, rivers and the            ogy. Using cutting-edge molecular                  spp. and Acinetobacter spp. were con-
                                           ocean. It has long been known that                 tools, many key players in microbial               sidered for decades to be responsible
                                           activated sludge consists mainly of                activated sludge have been identified              for nitrite oxidation and enhanced biol-
                                           bacteria (in concentrations of up to               and partially characterized, and we                ogical phosphorus removal in activated
                                           1011 cells ml–1) which catalyse all the            have begun to understand the laws                  sludge, respectively, and are still listed
                                           important steps of nutrient removal,               of biology which underpin this process.            as model organisms for these processes
                                           but are also responsible for the major             In parallel, microbial ecologists have             in recent issues of several standard
                                           problems in these plants. Nevertheless,            discovered that activated sludge is                textbooks on wastewater treatment.

                                             Pool for treating wastewater in British Columbia, Canada. Kaj R. Svensson / Science Photo Library

28        microbiology today february 05   microbiology today february 05                                                                                                                29
   Using cultivation-independent, 16S      available which allows microbial              agricultural soil to marine sponges.        project to decipher the genome of Candidatus ‘Accumulibacter         process stability and performance. Such insights cannot be
rRNA-based censuses of microbial           ecologists to identify which bacteria            Almost at the same time, Alexander       phosphatis’. These genome sequences will significantly speed         obtained by traditional bulk parameter measurements, but
activated sludge communities we have       metabolize the added substrates               Zehnder’s group at the EWAG (Düben-         up the process of understanding these recently discovered            require high resolution analyses of the structure and function
learnt that a single reactor may harbour   and so assign specific functions to           dorf, Switzerland) reported on the          micro-organisms, which serve us in ameliorating anthro-              of activated sludge microbial communities, as well as a
several hundred different bacterial        populations.                                  enrichment of uncultured betaproteo-        pogenic damage to the environment.                                   commitment of researchers and funding agencies to this
species. This diversity can be easily         This toolbox was used to identify the      bacteria tentatively named Candidatus                                                                            interdisciplinary research.
traced by DNA microarrays which            actual key players in nitrification,          ‘Accumulibacter phosphatis’ which are       Ecology and engineers
could, for example, be used in the         denitrification and enhanced biological       responsible for enhanced biological         Some engineers tend to undervalue progress in activated                Professor Michael Wagner
future to monitor community dynam-         phosphorus removal, most of which are         phosphorus removal in wastewater            sludge microbial ecology by stating that these new findings            Head of the Department of Microbial Ecology,
ics in activated sludge tanks or the       still uncultured. For example, Stefan         treatment plants. In addition, several      simply lead to a change in the names of bacteria responsible           University of Vienna, Althanstr. 14, A-1090 Vienna,
biogeography of activated sludge           Juretschko, a former PhD student in           novel denitrifiers have also been identi-   for certain processes in wastewater treatment plants, with             Austria (t +43 1 4277 54390; f +43 1 4277 54389;
bacteria. The abundance of selected        my group, discovered in 1998 that             fied in activated sludge. It has become     no relevance to their functions. My response is that these             e wagner@microbial-ecology.net)
populations in activated sludge can be     Nitrospira-related nitrite-oxidizers and      clear that the morphology-based             different names represent genetically very different organisms
precisely measured by quantitative         not Nitrobacter were the numerically          identification key in widespread use for    which have diverged early in evolutionary history and
                                                                                                                                                                                                          Further reading
fluorescence in situ hybridization         dominant nitrite oxidizers in most            filamentous activated sludge bacteria       thus are most likely to have very different physiological
(FISH) using rRNA-targeted oligo-          nitrifying activated sludge systems.          dramatically underestimates their           properties. For example, in an rRNA tree Nitrospira is as                         .,
                                                                                                                                                                                                            Curtis, T.P Head, I.M. & Graham, D.W. (2003). Theoretical
nucleotide probes. In 1994 and 1995,       Andreas Schramm and colleagues (now           actual diversity by lumping together        ‘closely’ related to Nitrobacter as humans are to sunflowers           ecology for engineering biology. Environ Sci Technol 37, 64A–70A.
by using this approach, we demon-          at the University of Aarhus, Denmark)         genetically very different organisms        (Fig. 2). Now that we have identified the microbial key players                                              .H.,
                                                                                                                                                                                                            Daims, H., Nielsen, J.L., Nielsen, P Schleifer, K.H. &
strated that neither Nitrobacter nor       later raised the hypothesis that these        which share the same morphology.            in many important processes and problems in activated                  Wagner, M. (2001). In situ characterization of Nitrospira-like
Acinetobacter were of major importance     nitrospiras are K-strategists for nitrite        Recent technological advances have       sludge, and have tools at hand to measure their abundance              nitrite-oxidizing bacteria active in wastewater treatment plants.
for N or P removal in most waste-          and oxygen and thus outcompete the            encouraged several research groups          and activities, we can begin to design experiments to show             Appl Environ Microbiol 67, 5273–5284.
water treatment plants. While perfectly    putative r-strategist Nitrobacter in react-   to attempt to determine the whole-          how operating conditions in a treatment plant affect these                                .,
                                                                                                                                                                                                            Hesselmann, R.P Werlen, C., Hahn, D., van der Meer, J.R. &
suited to the analysis of the microbial    ors with low nitrite availability. Recent     genome sequences of important but           parameters. A central follow-up question is how the diversity          Zehnder, A.J. (1999). Enrichment, phylogenetic analysis and
community structure, FISH results          data from my and other labs support           uncultured activated sludge bacteria.       of functionally important micro-organisms affects the                  detection of a bacterium that performs enhanced biological
alone are insufficient for understand-     this theory. Furthermore, it is now appa-     Together with the French national           stability of the respective processes (Fig. 1). For example, it is     phosphate removal in activated sludge. Syst Appl Microbiol 22,
ing the ecophysiology of activated         rent that these novel nitrite oxidizers       sequencing institute (Genoscope) and        tempting to speculate that a greater diversity of nitrifying           454–465.
sludge bacteria. This limitation can be    can grow mixotrophically in activated         collaborators from the Netherlands          bacteria increases the resistance of the nitrification process in      Wagner, M. (2004). Deciphering the function of uncultured
over-come in microcosm experiments         sludge, that different Nitrospira popu-       and Germany we are currently ana-           a wastewater treatment plant against certain perturbations.            micro-organisms. ASM News 70, 63–70.
which offer the activated sludge           lations co-occur in nitrifying sludges        lysing the sequences of anaerobic           Thus, engineers should be interested in monitoring the                 Wagner, M. & Loy, A. (2002). Bacterial community composition
community isotope-labelled substrates      (Fig. 1) and that these peculiar nitrite      ammonium oxidizers and Nitrospira-          diversity of important bacterial groups in their treatment             and function in sewage treatment systems. Curr Opin Biotechnol 13,
under defined conditions. Today, a         oxidizers are also of importance in           like nitrite oxidizers. Furthermore, the    plants and ultimately learn how they can influence diversity           218–227.
whole battery of molecular tools is        many other systems ranging from               Joint Genome Institute has launched a       by changing the operating conditions in order to maximize

                                                                                                                                                                                                            An outlet pipe discharges sewage onto a North Sea beach. Simon
 1a                                                                 1b                                                                2                                    Uncultured Nitrospira sp.        Fraser / Science Photo Library
                                                                                                                                                 Nitrospina                 from activated sludge
                                                                                                                                                  gracilis                                                  Fig. 1. FISH of nitrifying bacteria in activated sludges from two
                                                                                                                                                                                                            different wastewater treatment plants. (a) Two different populations
                                                                                                                                                                                                            of Nitrospira-like nitrite oxidizers are stained red and green,
                                                                                                                                                                                                            respectively. Ammonia oxidizers of the genus Nitrosomonas are
                                                                                                                                                                                                            labelled in blue. (b) Nitrospira-like nitrite oxidizers are stained red
                                                                                                                                                                                                            and ammonia oxidizers of the genus Nitrosomonas are labelled in
                                                                                                                                                 Nitrococcus                   Nitrobacter                  green. Bars, 10 µm. Courtesy Kilian Stoecker
                                                                                                                                                   mobilis                    hamburgensis
                                                                                                                                                                                                            Fig. 2. A comparison of phylogenetic distances between
                                                                                                                                                                          Amoeba                            recognized nitrite-oxidizing bacteria and between selected
                                                                                                                                                                      Acanthamoeba sp.                      eukaryotes. Phylogenetic 16S rRNA and 18S rRNA trees were
                                                                                                                                                                                                            calculated using neighbour joining with the Jukes–Cantor correction
                                                                                                                                                                                                            and alignment masks which excluded highly variable positions in
                                                                                                                                                  Helianthus annuus                                         the respective data sets. Bar represents 10 % estimated sequence

                                                                                                                                                                           Homo sapiens

                                                                                                                                          10 %

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