Project description:Polyhydroxyalkanoates (PHAs) are bio-based, biodegradable polyesters that can be produced from organic-rich waste streams using mixed microbial cultures. To maximize PHA production, mixed microbial cultures may be enriched for PHA-producing bacteria with a high storage capacity through the imposition of cyclic, aerobic feast-famine conditions in a sequencing batch reactor (SBR). Though enrichment SBRs have been extensively investigated a bulk solutions-level, little evidence at the proteome level is available to describe the observed SBR behavior to guide future SBR optimization strategies. As such, the purpose of this investigation was to characterize proteome dynamics of a mixed microbial culture in an SBR operated under aerobic feast-famine conditions using fermented dairy manure as the feedstock for PHA production. At the beginning of the SBR cycle, excess PHA precursors were provided to the mixed microbial culture (i.e., feast), after which followed a long duration devoid of exogenous substrate (i.e., famine). Two-dimensional electrophoresis was used to separate protein mixtures during a complete SBR cycle, and proteins of interest were identified.
Project description:Metaproteomic analysis of an enriched anaerobic rumen consortium (ERAC) using sugarcane bagasse and rumen as unique carbon and microbial sources
Project description:A shotgun metagenome microarray was created and used to investigate gene transcription during vinyl chloride (VC) dechlorination by a microbial enrichment culture called KB1. The array was constructed by spotting genomic fragments amplified from short-insert libraries of KB1 metagenomic DNA. Subsequently, the microarrays were interrogated with RNA extracted from KB1 during VC dechlorination (VC+methanol), and in the absence of VC (methanol-only). The most differentially expressed spots, and spots with the highest intensities, were then chosen to be sequenced. Sequencing revealed that Dehalococcoides (Dhc) genes involved in transcription, translation and energy generation were up-regulated during VC degradation. Furthermore, the results indicated that the reductive dehalogenase homologous (RDH) gene KB1rdhA14 is the only RDH gene up-regulated upon VC degradation, and that multiple RDH genes were more highly transcribed in the absence of VC. Numerous hypothetical genes from Dehalococcoides were also more highly transcribed in methanol only treatments and indicate that many uncharacterized proteins are involved in cell maintenance in the absence of chlorinated substrates. Spots with genes from Spirochaetes, Chloroflexi, Geobacter, Methanogens and phage organisms were differentially expressed and sequencing provided information from these uncultivated organisms that can be used to design primers for more targeted studies. This array format is powerful, as it does not require a priori sequence knowledge. This study provides the first report of such arrays being used to investigate transcription in a mixed community, and shows that this array format can be used to screen metagenomic libraries for functionally important genes.
