(660b) High Throughput Screening of Cyanobacteria Cultures for Maximized PHA Production

Dr. Michael Benton     benton@lsu.edu   Courtney Lane             clane6@tigers.lsu.edu

High Throughput Screening of Cyanobacteria Cultures for Maximized Bioplastic Production

Polyhydroxyalkanoates (PHAs), biodegradable and biocompatible biopolymers used for carbon storage by many microorganisms, are receiving much attention as possible replacements for petroleum-based polymers.  Typical PHA fermentation processes utilize heterotrophic organisms such as Escherichia coli or Cupriavidus necator and show high yields, approaching 80-90% dry cell weight in some cases. Still, such production remains economically uncompetitive with petroleum-based plastics due to high production costs, 50% of which can be attributed to the carbon feedstock.  Cyanobacteria are phototrophic prokaryotes and have the potential to make PHA bioproduction economically viable as they can fix atmospheric carbon through photosynthesis, lessening the cost of carbon feedstock.  Unfortunately, typical PHA yields in cyanobacteria are typically around 10% dry cell weight, far below the level needed for such viability.  However, some atypical PHA producing cyanobacteria have been found which can accumulate up to 60% dry cell weight of PHA.  Therefore, strain choice is paramount in the design of a cyanobacteria-based PHA production process and more high-yield strains of cyanobacteria must be identified if this bioprocess is to become viable.  Cyanobacteria are an extremely diverse phylum ranging from simple unicellular organisms to fully differentiating vegetative colonies and span over 10,000 taxonomic entries in the National Center for Biotechnology Information database.  With so many strains to categorize, a rapid, robust, and efficient screening process is needed to maximize the opportunity of identifying species capable of economically feasible PHA production.  In this work, a high-throughput method to determine the genotypic and phenotypic PHA accumulation characteristics in cyanobacteria via PCR amplification and fluorescent staining was developed and tested on a diverse range of cyanobacteria: Plectonema sp. UTEX 1541, Spirulina platensis UTEX LB 2340, Synechococcus sp. UTEX 2434, Synechocystis sp. PCC 6803, and Synechocystis sp. UTEX 2470.  Positive PHA accumulation was observed via staining and PCR amplification in PCC 6803, UTEX LB 2340, and UTEX 2470, while no signs of PHA accumulation were observed in UTEX 1541 or UTEX 2434.  This novel assay will rapidly identify those cyanobacteria which could (based on culture conditions) accumulate relatively high amounts of PHA.  The characterization of these outlier strains would lead to new insights and a better understanding of PHA accumulation in cyanobacteria.  The atypical producers detected utilizing this test could ultimately be implemented in a more economically and environmentally sustainable cyanobacteria-based PHA bioprocess, creating a sustainable production system more competitive with petroleum-based processes.