Prokaryotic community analysis with CARD-FISH in comparison to FISH in ultra-oligotrophic ground- and drinking water

AIMS: We compared the applicability of catalyzed reporter deposition fluorescence in situ hybridization (CARD-FISH) and FISH to enumerate prokaryotic populations in ultra-oligotrophic alpine ground waters and bottled mineral water METHODS AND RESULTS: The fluorescent oligonucleotide probes EUB338 and the EUB338mix (EUB338/EUB338-II/EUB338-III) were used to enumerate Bacteria and probes EURY806 and CREN537 for Euryarchaea and Crenarchaea, respectively. Improved detection of Planctomycetales by probe EUB338-II was tested using a different permeabilization step (proteinase K instead of lysozyme). Total detection efficiency of cells in spring water (normalized to DAPI counts) of four different alpine karst aquifers was on average 83% for CARD-FISH and only 15% for FISH. Applying CARD-FISH on bottled natural mineral waters resulted in an average total hybridization efficiency of 89%, with 78% (range 77% - 96%) Bacteria and 11% (range 3% - 22%) of cells identified as Archaea. CONCLUSIONS: CARD-FISH appears to result in substantially higher recovery efficiency than the conventional FISH approach and hence, is more suitable for the enumeration of specific prokaryotic groups in ground- and drinking water. SIGNIFICANCE AND IMPACT OF THE STUDY: This study represents the first evaluation of CARD-FISH on ultra-oligotrophic ground- and drinking water. Results are relevant for basic research and drinking water distributors. Archaea can comprise a significant fraction of the prokaryotic community in bottled mineral water

Bacterial community composition and potential controlling mechanisms along a trophic gradient in a barrier reef system

Bacterial abundance and community composition were investigated along trophic gradients in the barrier reef lagoon of Noumea, New Caledonia. Bacterial abundance and the percentage of high nucleic acid (%HNA) bacteria (a potential indicator for bacterial production) increased from offshore waters towards the head of the bays. 16S rRNA gene PCR and denaturing gradient gel electrophoresis (DGGE) were used as genetic fingerprints for assessing differences in bacterial community composition. Sequences of DGGE bands were assigned to (1) the genera Rugeria and Roseobacter (Rhodobacteriaceae), (2) the SAR11 cluster, (3) other Alphaproteobacteria, and (4) the genus Alteromonas. Removal of the operationally defined attached bacteria by prefiltration did not affect community profiles in offshore waters but had a strong influence in the bays, probably clue to the much higher particle load and thus, attached bacteria in the bays. For the free-living community, the number of bands decreased linearly with increasing water residence time, chlorophyll a concentration, and viral abundance. Specific bands were found for offshore waters and the 2 investigated semi-enclosed bays, whereas the lagoon showed no specific bands. A similarity analysis showed specific clusters for offshore water, the lagoon, and the bays. A principle component analysis together with cluster and correlation analysis indicated that water residence time, viruses, and a complex top-down cascading effect of ciliate grazers on flagellates influenced community composition. Also, data from fingerprints of the total and free-living communities suggest that the free-living and the attached community are controlled by different mechanisms

Diversity of planktonic and attached bacterial communities in a phenol-contaminated sandstone aquifer.

Polluted aquifers contain indigenous microbial communities with the potential for in situ bioremediation. However, the effect of hydrogeochemical gradients on in situ microbial communities (especially at the plume fringe, where natural attenuation is higher) is still not clear. In this study, we used culture-independent techniques to investigate the diversity of in situ planktonic and attached bacterial communities in a phenol-contaminated sandstone aquifer. Within the upper and lower plume fringes, denaturing gradient gel electrophoresis profiles indicated that planktonic community structure was influenced by the steep hydrogeochemical gradient of the plume rather than the spatial location in the aquifer. Under the same hydrogeochemical conditions (in the lower plume fringe, 30 m below ground level), 16S rRNA gene cloning and sequencing showed that planktonic and attached bacterial communities differed markedly and that the attached community was more diverse. The 16S rRNA gene phylogeny also suggested that a phylogenetically diverse bacterial community operated at this depth (30 mbgl), with biodegradation of phenolic compounds by nitrate-reducing Azoarcus and Acidovorax strains potentially being an important process. The presence of acetogenic and sulphate-reducing bacteria only in the planktonic clone library indicates that some natural attenuation processes may occur preferentially in one of the two growth phases (attached or planktonic). Therefore, this study has provided a better understanding of the microbial ecology of this phenol-contaminated aquifer, and it highlights the need for investigating both planktonic and attached microbial communities when assessing the potential for natural attenuation in contaminated aquifers