The microbial characterization of granular biomass samples received from WP1 partners of INNOWATECH project was performed by IRSA-Rome. TU-Delf provided samples from Sequencing Batch Granular Reactors (SBGR) fed with different synthetic substrated and industrial wastewater while IRSA-BARI provided samples from a Sequencing Batch Biofilter Granular Reactor (SBBGR) fed with landfill leachate. The main objective of the characterization was to elucidate the microbial composition and distribution in granules. The abundance and composition of the EPS was also evaluated. The analysis was performed by adopting a combination of biomolecular methods and microscopic techniques. Fluorescence in Situ Hybridization (FISH) has been chosen as elective screening tool to identify, visualize and quantify bacterial species and to investigate the biomass composition and granule development. Originally introduced by DeLong et al. in 1989, the use of FISH expanded to the study of bacterial populations in environmental and medical applications (Wagner et al., 2003). During the last years, many rRNA targeted oligonucleotide probes for in situ detection of bacteria present in biological treatment systems have been designed. Some of these probe sequences and stringency conditions can be easily accessed on-line using probeBase [http://www.microbial-ecology.net/probebase]. By applying these probes under the recommended hybridization conditions, the respective target microorganisms can be specifically identified in appropriately-fixed microbial samples within about three hours. The technique consists basically of three main steps: proper fixation of the sample, hybridization with the probes and successive washing, and finally, sample examination by epifluorescence microscopy. Sample fixation assures rRNA content preservation and maintenance of cellular integrity even during exposure to high temperatures and high osmotic gradients during the hybridisation phase. The protocol of microscopic characterization includes also staining procedures for the identification of filamentous bacteria, the estimation of EPS composition and of intracellular polyidroxyalcanoate (PHA) inclusions. Totally 35 biomass samples from WP1 project partners were analysed.
Deliverable D1.8 Report on the results of microscopic characterisation of sectioned granules
Simona Rossetti;Marco De Sanctis
2010
Abstract
The microbial characterization of granular biomass samples received from WP1 partners of INNOWATECH project was performed by IRSA-Rome. TU-Delf provided samples from Sequencing Batch Granular Reactors (SBGR) fed with different synthetic substrated and industrial wastewater while IRSA-BARI provided samples from a Sequencing Batch Biofilter Granular Reactor (SBBGR) fed with landfill leachate. The main objective of the characterization was to elucidate the microbial composition and distribution in granules. The abundance and composition of the EPS was also evaluated. The analysis was performed by adopting a combination of biomolecular methods and microscopic techniques. Fluorescence in Situ Hybridization (FISH) has been chosen as elective screening tool to identify, visualize and quantify bacterial species and to investigate the biomass composition and granule development. Originally introduced by DeLong et al. in 1989, the use of FISH expanded to the study of bacterial populations in environmental and medical applications (Wagner et al., 2003). During the last years, many rRNA targeted oligonucleotide probes for in situ detection of bacteria present in biological treatment systems have been designed. Some of these probe sequences and stringency conditions can be easily accessed on-line using probeBase [http://www.microbial-ecology.net/probebase]. By applying these probes under the recommended hybridization conditions, the respective target microorganisms can be specifically identified in appropriately-fixed microbial samples within about three hours. The technique consists basically of three main steps: proper fixation of the sample, hybridization with the probes and successive washing, and finally, sample examination by epifluorescence microscopy. Sample fixation assures rRNA content preservation and maintenance of cellular integrity even during exposure to high temperatures and high osmotic gradients during the hybridisation phase. The protocol of microscopic characterization includes also staining procedures for the identification of filamentous bacteria, the estimation of EPS composition and of intracellular polyidroxyalcanoate (PHA) inclusions. Totally 35 biomass samples from WP1 project partners were analysed.I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.