Unveiling PHA-storing populations using molecular methods

Abstract

Enrichment of mixed microbial cultures (MMCs) in polyhydroxyalkanoate (PHA)-storing microorganisms must take place to develop a successful PHA production process. Moreover, throughout the operational period of a MMC system, the population needs to be checked in order to understand the changes in the performance that eventually occurred. For these reasons, it is necessary to monitor the population evolution, in order to identify the different groups of microorganisms and relate them with the storage capacity and kinetics of the MMC. Regarding this particular process, several culture-independent molecular techniques were already applied, with the use of hybridization techniques such fluorescence in situ hybridization and also PCR-based methods like denaturing gradient gel electrophoresis, terminal restriction fragment length polymorphism, pyrosequencing, and quantitative PCR standing out. This review intends, thus, to look at the molecular methods currently applied in monitoring the PHA-storing population evolution and how they can be combined with the evolutionary engineering step in order to optimize the overall process.

Impact of Organic Acids Supplementation to Hardwood Spent Sulfite Liquor as Substrate for the Selection of Polyhydroxyalkanoates-Producing Organisms

“…Such characterization can contribute towards the development of operational process strategies designed to optimize the structure of the microbial community [21]. Several works that address the microbial characterization of their enrichments already revealed diverse communities that probably resulted from the operational conditions applied and the type of substrate used [15].…”

“…Specific probes for Thauera and Azoarcus genera belonging to Betaproteobacteria and often detected in MMC processes as PHAs-accumulating organisms were also tested [15]. Positive results were obtained for both genera, but only in small amounts, Thauera corresponded to 0.72 ± 0.25%, and Azoarcus to 0.62 ± 0.02%, remaining most of Betaproteobacteria unidentified.…”

“…As reviewed by Queirós et al [15], in recent years, several works dedicated to study the composition of MMC populations selected in PHAs producing processes, revealed a broad phylogenetic diversity. However, a link between operational conditions and the selection of a specific taxon over others, as well as the impact of the amount and type of accumulating organisms on the overall PHAs production, remains unclear [20,21].…”

“…By alternating periods of excess with periods of lack of carbon source, a selective pressure favorable to PHAs-storing organisms is imposed. This procedure, also known as aerobic dynamic feeding (ADF), is the key to the success of the process [15]. ADF can be tuned by manipulating several parameters such as organic loading rate (OLR) and cycle length.…”

“…MMC were reported preferentially to store polysaccharides instead of PHAs from sugar-based compounds when submitted to feast and famine conditions [19]. Queirós et al [9,15] observed that notwithstanding being able to select a MCC with HSSL under ADF conditions, presenting feast-to-famine ratios indicative of PHAs storage, low accumulations values were obtained. Moreover, Queirós et al [9] observed the accumulation of a glucose polymer that was probably produced by a side population able to uptake sugars present in HSSL.…”