Application of microautoradiography to the study of substrate uptake by filamentous microorganisms in activated sludge

Abstract

Excessive growth of filamentous microorganisms in activated-sludge treatment plants is a major operational problem which causes poor settlement of activated sludge. An enhanced understanding of the factors controlling growth of different filamentous microorganisms is necessary in order to establish more successful control strategies. In the present study, the in situ substrate uptake was investigated by means of microautoradiography. It was demonstrated that the uptake of labeled organic substrates by the filamentous microorganisms, during short-term incubation, could be detected by microautoradiography. Viability and respiratory activity of the filaments were also detected by reduction of CTC (5-cyano-2,3-ditolyl tetrazolium chloride) and by incorporation of [ 3 H]thymidine. Gram, Neisser, and fluorescence staining techniques were used for the localization and identification of the filaments. Activated-sludge samples from five wastewater treatment plants with bulking problems due to filamentous microorganisms were investigated. Microthrix parvicella, Nostocoida limicola, and Eikelboom's type 0041 and type 021N were investigated for their ability to take up organic substrates. A panel of six substrates, i.e., [ 14 C]acetate, [ 3 H]glucose, [ 14 C]ethanol, [ 3 H]glycine, [ 3 H]leucine, and [ 3 H]oleic acid, was tested. The uptake response was found to be very specific not only between the different filamentous types but also among filaments of the same type from different treatment plants. Interestingly, M. parvicella consistently took up only oleic acid among the tested substrates. It is concluded that microautoradiography is a useful method for investigation of in situ substrate uptake by filamentous microorganisms in activated sludge.

Studies on the in situ physiology of Thiothrix spp. present in activated sludge

“…Heterotrophic Thiothrix strains are known (Williams and Unz, 1985) and the preincubation step of adding non-labelled acetate for 1 h before the labelled acetate was added and taken up (during the following 2±3 h incubation) strongly suggests that it was not a transient uptake and storage, but actual growth. Thiothrix filaments were not able to take up glucose which agrees with both the pure culture studies and our earlier studies of the in situ physiology of Thiothrix in different water treatment plants (Andreasen and Nielsen, 1997;Nielsen et al, 1998).…”

In situ substrate conversion and assimilation by nitrifying bacteria in a model biofilm

“…Vertical cryosections of the model biofilm from each experiment were hybridized with the different probes (Table 1) as described above. The sections were subsequently covered with a photographic emulsion for microautoradiography and allowed to expose for approximately 7 days before development as described earlier (Andreasen and Nielsen, 1997). The developed slides were dried and immediately inspected under the microscope.…”

Ecophysiology of mycolic acid-containing Actinobacteria (Mycolata) in activated sludge foams

“…The MAR experiments were performed using several 3 H-labeled and 14 C-labeled organic compounds. The procedure, which includes incubation, fixation, and FISH with oligonucleotide probes, addition of a radiosensitive emulsion, exposure, processing, and microscopic investigations, is described in detail elsewhere (Andreasen & Nielsen, 1997;Lee et al, 1999;Nielsen et al, 2000). A range of potential electron donors was assessed for their uptake under aerobic conditions.…”