ABSTRACT
AIM: Identification of the predominating cultivable bacteria in granular activated carbon (GAC) filters used in a variety of water treatment plants for selecting representative strains to study the role of bacteria in the removal of dissolved organic matter. METHODS AND RESULTS: Bacterial isolates were collected from 21 GAC filters in nine water treatment plants treating either ground water or surface water with or without oxidative pretreatment. Enrichment of samples in dilute liquid medium improved culturability of the bacteria by approximately log unit, to 9% up to 70% of the total cell counts. Genomic fingerprinting and 16S rDNA sequence analysis revealed that most (68%) of the isolates belonged to the Betaproteobacteria and 25% were identified as Alphaproteobacteria. The number of different genera within the Betaproteobacteria was higher in the GAC filters treating ozonated water than in the filters treating nonozonated water. Polaromonas was observed in nearly all of the GAC filters (86%), and the genera Hydrogenophaga, Sphingomonas and Afipia were observed in 43%, 33% and 29% of the filter beds, respectively. AFLP analysis revealed that the predominating genus Polaromonas included a total of 23 different genotypes. CONCLUSIONS: This study is the first to demonstrate that Polaromonas, which has mainly been observed in ultraoligotrophic freshwater environments, is a common component of the microbial community in GAC filters used in water treatment. SIGNIFICANCE AND IMPACT OF THE STUDY: The predominance of ultraoligotrophic bacteria in the GAC filters indicates that very low concentrations of substrates are available for microbial growth. Polaromonas species are suited for further studies on the nutritional versatility and growth kinetics enabling the modelling of biodegradation processes in GAC filters.
Subject(s)
Bacteria/isolation & purification , Carbon , Comamonadaceae/isolation & purification , Filtration/methods , Water Microbiology , Water Supply , Bacteria/genetics , Biofilms/growth & development , Colony Count, Microbial , Comamonadaceae/classification , Comamonadaceae/genetics , DNA Fingerprinting , Ozone , Phylogeny , Polymerase Chain Reaction/methods , RNA, Ribosomal, 16S , Sequence Analysis, DNA , Water Supply/standardsABSTRACT
In order to aid the monitoring of the overall quality of (surface) waters a new analytical approach has been developed, combining on-line solid-phase extraction, HPLC separation and effect-related detection. Compounds present in surface water or wastewater samples are extracted on-line with Oasis [poly(divinylbenzene-co-N-vinylpyrrolidone)] material and directly fractionated by reversed-phase HPLC. The eluent of the total chromatogram is collected on a microtitre plate in fractions of 1 min each. After evaporation and re-dissolvation in a suitable solvent, the (geno)toxicity of the individual fractions before and after enzymatic activation with S9, is determined with the umu test. In this way, harmful compounds can be detected and localized in the HPLC-diode array detection trace even without their identity and exact concentration being known at that moment. The method was developed using two test compounds, 4-nitroquinoline-N-oxide and 2-aminoanthracene. Compounds with mutagenic properties comparable to those of the test compounds can be detected from 0.1 microg/l, which is a concentration relevant for surface waters. The new analytical approach was successfully applied to various types of model samples, as well as real wastewater.
Subject(s)
Chromatography, High Pressure Liquid/methods , Mutagenicity Tests , 4-Nitroquinoline-1-oxide/toxicity , Anthracenes/toxicity , Biotransformation , Industrial Waste , Mutagens/toxicity , Water Pollutants, Chemical/toxicityABSTRACT
During the past few years, Ralstonia (Pseudomonas) solanacearum race 3, biovar 2, was repeatedly found in potatoes in Western Europe. To detect this bacterium in potato tissue samples, we developed a method based on fluorescent in situ hybridization (FISH). The nearly complete genes encoding 23S rRNA of five R. solanacearum strains and one Ralstonia pickettii strain were PCR amplified, sequenced, and analyzed by sequence alignment. This resulted in the construction of an unrooted tree and supported previous conclusions based on 16S rRNA sequence comparison in which R. solanacearum strains are subdivided into two clusters. Based on the alignments, two specific probes, RSOLA and RSOLB, were designed for R. solanacearum and the closely related Ralstonia syzygii and blood disease bacterium. The specificity of the probes was demonstrated by dot blot hybridization with RNA extracted from 88 bacterial strains. Probe RSOLB was successfully applied in FISH detection with pure cultures and potato tissue samples, showing a strong fluorescent signal. Unexpectedly, probe RSOLA gave a less intense signal with target cells. Potato samples are currently screened by indirect immunofluorescence (IIF). By simultaneously applying IIF and the developed specific FISH, two independent targets for identification of R. solanacearum are combined, resulting in a rapid (1-day), accurate identification of the undesired pathogen. The significance of the method was validated by detecting the pathogen in soil and water samples and root tissue of the weed host Solanum dulcamara (bittersweet) in contaminated areas.
Subject(s)
Nucleic Acid Conformation , Pseudomonas/isolation & purification , RNA, Ribosomal, 23S/chemistry , RNA, Ribosomal, 23S/genetics , Solanum tuberosum/microbiology , Base Sequence , In Situ Hybridization, Fluorescence , Molecular Sequence Data , Phylogeny , Plant Diseases , Polymerase Chain Reaction/methods , Pseudomonas/genetics , RNA, Bacterial/chemistry , RNA, Bacterial/genetics , Sequence Alignment , Sequence Homology, Nucleic AcidABSTRACT
A 16S rRNA sequence analysis of Syntrophobacter wolinii was done by using PCR amplification of the 16S rRNA-genes from DNA isolated from the S. wolinii-Desulfovibrio sp. coculture. Phylogenetic analysis using the obtained sequence revealed that S. wolinii was not related to bacteria growing syntrophically on other fatty acids than propionate, but was related to sulfate-reducing bacteria. The closest related bacteria are Desulfomonile tiedjei and Desulfoarculus baarsii.
Subject(s)
Phylogeny , Sulfur-Reducing Bacteria/classification , Sulfur-Reducing Bacteria/genetics , Base Sequence , Cloning, Molecular , DNA, Bacterial/genetics , Desulfovibrio/classification , Desulfovibrio/genetics , Molecular Sequence Data , Polymerase Chain Reaction , RNA, Bacterial/genetics , RNA, Ribosomal, 16S/genetics , Species SpecificityABSTRACT
Free-living N2-fixing Frankia strains isolated from Casuarina sp. were investigated for genomic polymorphism. We used six 10-mer oligonucleotides as single arbitrary primers (AP) for the polymerase chain reaction (PCR) in order to amplify random DNA fragments in the genome of free-living Frankia strains. Agarose-gels of the amplified genomic DNA revealed that two of the six arbitrary primers showed polymorphism in the eight different Frankia genomes. Analysis of the AP-PCR products showed 9 polymorphic bands ranging from 4.1-0.60 kb. We conclude that single arbitrary primers can be used to amplify genomic DNA, and that polymorphism can be detected between the amplification products of the different Frankia genomes.
