CHARACTERIZATION OF BACTERIAL COMMUNITIES FROM ACTIVATED-SLUDGE - CULTURE-DEPENDENT NUMERICAL IDENTIFICATION VERSUS IN-SITU IDENTIFICATION USING GROUP-SPECIFIC AND GENUS-SPECIFIC RIBOSOMAL-RNA-TARGETED OLIGONUCLEOTIDE PROBES

The structures of bacterial communities were studied in activated slud ge samples obtained from the aerobic and anaerobic zones of a wastewat er treatment plant showing enhanced phosphorous removal. Samples were analyzed by in situ hybridization with oligonucleotide probes compleme ntary to selected regions of the 16S and 23S ribosomal RNA (rRNA) char acteristic for defined phylogenetic entities (genera and larger groups ). The microbial community structures revealed by molecular techniques were compared with the compositions of culturable bacterial communiti es, obtained from the characterization of 255 isolates from tryptone-s oy (TS) agar and R2A agar. These isolates were characterized by 89 phy siological tests and their cellular fatty acid patterns, and identifie d. Culture-dependent techniques indicated the following distribution: different Aeromonas spp. (2.7-8.3% on R2A agar; 45.0-63.7% on TS agar) , Acinetobacter spp. (5.4-9.0% on R2A agar; 5.0-9.1% on TS agar), Pseu domonas spp. (up to 10% on R2A agar) and Shewanella putrefaciens (up t o 3.0% on R2A agar), all members of the gamma subclass of Proteobacter ia, were isolated most frequently. The relatively rare isolates of the beta subclass were identified as Acidovorax spp., Alcaligenes spp., a nd Comamonas spp.. The Gram-positive bacteria (high DNA G+C) were assi gned mainly to Arthrobacter spp., Microbacterium spp., and Mycobacteri um phlei. Zn order to assess the in situ abundance of the most frequen tly isolated genus, Aeromonas, two rRNA-targeted oligonucleotide probe s were developed. The two gamma proteobacterial genera Aeromonas and A cinetobacter constituted less than 5% of all bacteria. In situ, Proteo bacteria belonging to the beta subclass and high G+C Gram-positive bac teria were dominant. From filamentous bacteria, Sphaerotilus spp. and Leptothrix spp. could be detected occasionally. In addition, one sampl e contained a high proportion of the morphologically distinct filament s of Microthrix parvicella. As for the genus Acinetobacter the relativ e abundance of the most frequently gamma-proteobacterial genus Aeromon as was overestimated by the intrinsic selectivity of cultivation. Cult ivation on nutrient-rich medium (TS-agar) especially supported an enha nced isolation of bacteria belonging to these two genera.