Optimization of Ralstonia solanacearum cell growth using a central composite rotational design for the P(3HB) production: Effect of agitation and aeration

Abstract

The intracellular accumulation of polyhydroxyalkanoates (PHAs) normally occurs after cell growth, during the second fermentation stage and under nutrient-limited conditions in the presence of a carbon excess. However, some microorganisms are able to accumulate PHAs as poly(3-hydroxybutyrate) [P(3HB)] during the first fermentation stage, the cell growth phase, without nutrient limitation, once they have been reported to utilize type II metabolism during the polymer accumulation phase. This study evaluated the effect of aeration and agitation on cell growth and P(3HB) accumulation in Ralstonia solanacearum RS, performed in a bioreactor for 24h at 32°C. A 22 central composite rotational design (CCRD) was used, with agitation (150 to 250 rpm) and aeration (0.3 to 1 vvm) as independent variables and optical density (OD600nm), dry cell weight (DCW), and P(3HB) yield as dependent variables. A significant polymer accumulation, until 70% of P(3HB), was observed, proving that R. solanacearum RS exhibited metabolism type II, regardless of the aeration process. The best results were obtained for 1 vvm and 250 rpm (+1, +1), with values of OD600nm (18.04) and DCW (4.82 g.L-1).

Advanced Killing Potential of Thymol against a Time and Temperature Optimized Attached Listeria monocytogenes Population in Lettuce Broth

“…Α central composite rotational design (CCRD) including 10 experiments, each one twice executed (i.e., 20 experiments in total) ( Table 1 ), was applied to determine the putative interactive effects of the two independent factors, that is the incubation time ( X 1 , varying between 14.1 and 81.9 h) and temperature ( X 2 , varying between 3 and 37 °C) on the concentration of the attached/biofilm cells (Log 10 CFU/cm 2 ) of the four-strain L. monocytogenes cocktail on the SS coupons, as previously described [ 29 ]. That design also allowed to determine those values of the two factors that upon concurrently applied would maximize the final population density (Log 10 CFU/cm 2 ) of the attached/biofilm bacteria.…”

“…Considering all the above, the main aim of the current study was to compare the effectiveness of THY to that of benzalkonium chloride (BAC), a well-known quaternary ammonium compound (QAC) widely used as biocide in many sanitizing formulations applied in industrial, health care, home, and cosmetics settings [ 28 ], against sessile L. monocytogenes bacteria under attachment conditions trying to simulate as much as possible those encountered in the salad industry. Those latter conditions were initially here optimized following response surface methodology (RSM) [ 29 ] to predict that incubation time and temperature combination favoring the attachment of a four-strain L. monocytogenes cocktail to stainless steel (SS) coupons placed fully immersed in diluted sterile lettuce broth (dLB). The involvement of each strain in the formation of the mixed sessile community and its antimicrobial recalcitrance was also monitored by a repetitive sequence-based polymerase chain reaction (rep-PCR) approach [ 30 ].…”

Insights into the Polyhydroxybutyrate Biosynthesis in Ralstonia solanacearum Using Parallel 13C Tracers and Comparative Genome Analysis

“…R. solanacearum , although a plant pathogen, has the ability to accumulate P­(3HB), and it is essential to understand its metabolic capabilities in order to make it promising industrially. R. solanacearum is a Gram-negative bacteria belonging to the class beta proteobacteria. , Most of the research is focused on the virulence factors of R. solanacearum , with recent studies showing it as a potential candidate for PHB bioprocessing. , The latest studies show that for large-scale P­(3HB) production, R. solanacearum can be considered as it can metabolize glucose efficiently and researchers have achieved up to 50% yield (w/w) of P­(3HB) with modified extraction parameters. , The studies on R. solanacearum as P­(3HB) producer is currently limited to bioprocessing and recovery methods, and there is need to map biosynthetic pathways to understand potential metabolic bottlenecks. With the aim to investigate the intactness of P­(3HB) biosynthesis pathways and potential sources of NADPH needed for PHB in R. solanacearum , we undertook pathway analysis using comparative genomics and 13 C tracer studies.…”

“…6,7 Most of the research is focused on the virulence factors of R. solanacearum, with recent studies showing it as a potential candidate for PHB bioprocessing. 8,9 The latest studies show that for large-scale P(3HB) production, R. solanacearum can be considered as it can metabolize glucose efficiently and researchers have achieved up to 50% yield (w/w) of P(3HB) with modified extraction parameters. 8,9 The studies on R. solanacearum as P(3HB) producer is currently limited to bioprocessing and recovery methods, and there is need to map biosynthetic pathways to understand potential metabolic bottlenecks.…”

“…8,9 The latest studies show that for large-scale P(3HB) production, R. solanacearum can be considered as it can metabolize glucose efficiently and researchers have achieved up to 50% yield (w/w) of P(3HB) with modified extraction parameters. 8,9 The studies on R. solanacearum as P(3HB) producer is currently limited to bioprocessing and recovery methods, and there is need to map biosynthetic pathways to understand potential metabolic bottlenecks. With the aim to investigate the intactness of P(3HB) biosynthesis pathways and potential sources of NADPH needed for PHB in R. solanacearum, we undertook pathway analysis using comparative genomics and 13 C tracer studies.…”