SummaryTechnical developments in molecular biology have found extensive applications in the field of microbial ecology. Among these techniques, fingerprinting methods such as denaturing gel electrophoresis (DGE, including the three options: DGGE, TGGE and TTGE) has been applied to environmental samples over this last decade. Microbial ecologists took advantage of this technique, originally developed for the detection of single mutations, for the analysis of whole bacterial communities. However, until recently, the results of these high quality fingerprinting patterns were restricted to a visual interpretation, neglecting the analytical potential of the method in terms of statistical significance and ecological interpretation. A brief recall is presented here about the principles and limitations of DGE fingerprinting analysis, with an emphasis on the need of standardization of the whole analytical process. The main content focuses on statistical strategies for analysing the gel patterns, from single band examination to the analysis of whole fingerprinting profiles. Applying statistical method make the DGE fingerprinting technique a promising tool. Numerous samples can be analysed simultaneously, permitting the monitoring of microbial communities or simply bacterial groups for which occurrence and relative frequency are affected by any environmental parameter. As previously applied in the fields of plant and animal ecology, the use of statistics provides a significant advantage for the nonambiguous interpretation of the spatial and temporal functioning of microbial communities.

The current knowledge of microbial biocenoses (communities) in pristine aquifers is presented in a review, which also discusses their relevance for questions of groundwater protection. Aquifers are heterogeneous on all scales and structured in a variety of habitats. The void spaces in many aquifers are small. The biocenoses are thus predominantly composed of microorganisms and, often, microinvertebrates. Larger voids and macroorganisms occur in karst cavities. Due to the absence of light, the biocenoses depend on chemical energy resources, which are, however, scarce in non-contaminated groundwater. The microorganisms thus show small cell sizes, low population densities and reduced activity; they developed specific strategies to survive oligotrophic conditions. The review also discusses the impact of contamination on the biocenoses, and the potential use of the biocenoses or specific organisms as indicators for groundwater quality, and the limits of this approach. Bacteria are either planktonic or attached to aquifer material, which requires both fluid and solid phase sampling. Most groundwater bacteria are viable but non-culturable. Consequently, cultivation techniques give an incomplete picture of the biocenoses, while methods from molecular microbiology provide genetic fingerprints of the entire community. Different analytical methods are available to count microorganisms, identify species, characterise microbial diversity, and measure activity.Résumé Cette revue expose l'état actuel des connaissances concernant les biocénoses microbiennes présentes dans les aquifères oligotrophes. L'impact d'une contamination sur les biocénoses est discuté, ainsi que le potentiel que Les bactéries sont planctoniques ou attachées aux matériaux de l'aquifère, ce qui demande unéchantillonnagè a la fois de l'eau et du substrat. De nombreuses méthodes sont aujourd'hui disponibles pour le comptage, l'identification et la caractérisation de la diversité, ainsi que la mesure des activités des organismes des aquifères. Comme la grande majorité des bactéries est viable mais non cultivable, les techniques de cultures actuelles ne donnent qu'une image incomplète des communautés, alors que les méthodes moléculaires développées récemment offrent la possibilité d'obtenir un profil de la communauté plus complet.Resumen Se presentauna reseña crítica del conocimiento actual de biocenosis microbiana (comunidades) en acuíferos prístinos la cual también discute su relevancia en términos de protección de aguas subterráneas. Los acuíferos son heterogéneos en todas las escalas y estructurados en una variedad de habitats. Los espacios vacíos en muchos acuíferos son pequeños. La biocenosis está por lo tanto compuesta predominantemente por microorganismos y, frecuentemente, microinvertebrados. Espacios más grandes y macroorganismos ocurren en cavidades kársticas. Debido a la ausencia de luz la biocenosis depende de recursos energéticos químicos los cuales, sin embargo, son escasos en agua subterránea no contaminada. Los microorganismos muestran...

Aerobic granular sludge is attractive for high-rate biological wastewater treatment. Biomass wash-out conditions stimulate the formation of aerobic granules. Deteriorated performances in biomass settling and nutrient removal during start-up have however often been reported. The effect of wash-out dynamics was investigated on bacterial selection, biomass settling behavior, and metabolic activities during the formation of early-stage granules from activated sludge of two wastewater treatment plants (WWTP) over start-up periods of maximum 60 days. Five bubble-column sequencing batch reactors were operated with feast-famine regimes consisting of rapid pulse or slow anaerobic feeding followed by aerobic starvation. Slow-settling fluffy granules were formed when an insufficient superficial air velocity (SAV; 1.8 cm s−1) was applied, when the inoculation sludge was taken from a WWTP removing organic matter only, or when reactors were operated at 30°C. Fast-settling dense granules were obtained with 4.0 cm s−1 SAV, or when the inoculation sludge was taken from a WWTP removing all nutrients biologically. However, only carbon was aerobically removed during start-up. Fluffy granules and dense granules were displaying distinct predominant phylotypes, namely filamentous Burkholderiales affiliates and Zoogloea relatives, respectively. The latter were predominant in dense granules independently from the feeding regime. A combination of insufficient solid retention time and of leakage of acetate into the aeration phase during intensive biomass wash-out was the cause for the proliferation of Zoogloea spp. in dense granules, and for the deterioration of BNR performances. It is however not certain that Zoogloea-like organisms are essential in granule formation. Optimal operation conditions should be elucidated for maintaining a balance between organisms with granulation propensity and nutrient removing organisms in order to form granules with BNR activities in short start-up periods.

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Two bubble column sequencing batch reactors fed with an artificial wastewater were operated at 20 degrees C, 30 degrees C, and 35 degrees C. In a first stage, stable granules were obtained at 20 degrees C, whereas fluffy structures were observed at 30 degrees C. Molecular analysis revealed high abundance of the operational taxonomic unit 208 (OTU 208) affiliating with filamentous bacteria Leptothrix spp. at 30 degrees C, an OTU much less abundant at 20 degrees C. The granular sludge obtained at 20 degrees C was used for the second stage during which one reactor was maintained at 20 degrees C and the second operated at 30 degrees C and 35 degrees C after prior gradual increase of temperature. Aerobic granular sludge with similar physical properties developed in both reactors but it had different nutrient elimination performances and microbial communities. At 20 degrees C, acetate was consumed during anaerobic feeding, and biological phosphorous removal was observed when Rhodocyclaceae-affiliating OTU 214 was present. At 30 degrees C and 35 degrees C, acetate was mainly consumed during aeration and phosphorous removal was insignificant. OTU 214 was almost absent but the Gammaproteobacteria-affiliating OTU 239 was more abundant than at 20 degrees C. Aerobic granular sludge at all temperatures contained abundantly the OTUs 224 and 289 affiliating with Sphingomonadaceae indicating that this bacterial family played an important role in maintaining stable granular structures.

Abstract. Bacteriophages are increasingly used as tracers for quantitative analysis in both hydrology and hydrogeology. The biological particles are neither toxic nor pathogenic for other living organisms as they penetrate only a specific bacterial host. They have many advantages over classical fluorescent tracers and offer the additional possibility of multi-point injection for tracer tests. Several years of research make them suitable for quantitative transport analysis and flow boundary delineation in both surface and ground waters, including karst, fractured and porous media aquifers. This article presents the effective application of bacteriophages based on their use in differing Swiss hydrological environments and compares their behaviour to conventional coloured dye or salt-type tracers. In surface water and karst aquifers, bacteriophages travel at about the same speed as the typically referenced fluorescent tracers (uranine, sulphurhodamine G extra). In aquifers of interstitial porosity, however, they appear to migrate more rapidly than fluorescent tracers, albeit with a significant reduction in their numbers within the porous media. This faster travel time implies that a modified rationale is needed for defining some ground water protection area boundaries. Further developments of other bacteriophages and their documentation as tracer methods should result in an accurate and efficient tracer tool that will be a proven alternative to conventional fluorescent dyes.

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The enrichment culture SL2 dechlorinating tetrachloroethene (PCE) to ethene with strong trichloroethene (TCE) accumulation prior to cis-1,2-dichloroethene (cis-DCE) formation was analyzed for the presence of organohalide respiring bacteria and reductive dehalogenase genes (rdhA). Sulfurospirillum-affiliated bacteria were identified to be involved in PCE dechlorination to cis-DCE whereas "Dehalococcoides"-affiliated bacteria mainly dechlorinated cis-DCE to ethene. Two rdhA genes highly similar to tetrachloroethene reductive dehalogenase genes (pceA) of S. multivorans and S. halorespirans were present as well as an rdhA gene very similar to the trichloroethene reductive dehalogenase gene (tceA) of "Dehalococcoides ethenogenes" strain 195. A single strand conformation polymorphism (SSCP) method was developed allowing the simultaneous detection of the three rdhA genes and the estimation of their abundance. SSCP analysis of different SL2 cultures showed that one pceA gene was expressed during PCE dechlorination whereas the second was expressed during TCE dechlorination. The tceA gene was involved in cis-DCE dechlorination to ethene. Analysis of the internal transcribed spacer region between the 16S and 23S rRNA genes revealed two distinct sequences originating from Sulfurospirillum suggesting that two Sulfurospirillum populations were present in SL2. Whether each Sulfurospirillum population was catalyzing a different dechlorination step could however not be elucidated.