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Heterologeous production of rhamnolipids in Pseudomonas putida is characterized by advantages of a non-pathogenic host and avoidance of the native quorum sensing regulation in Pseudomonas aeruginosa. Yet, downstream processing is a major problem in rhamnolipid production and increases in complexity at low rhamnolipid titers and when using chemical foam control. This leaves the necessity of a simple concentrating and purification method. Foam fractionation is an elegant method for in situ product removal when producing microbial surfactants. However, up to now in situ foam fractionation is nearly exclusively reported for the production of surfactin with Bacillus subtilis. So far no cultivation integrated foam fractionation process for rhamnolipid production has been reported. This is probably due to excessive bacterial foam enrichment in that system. In this article a simple integrated foam fractionation process is reported for heterologous rhamnolipid production in a bioreactor with easily manageable bacterial foam enrichments. Rhamnolipids were highly concentrated in the foam during the cultivation process with enrichment factors up to 200. The described process was evaluated at different pH, media compositions and temperatures. Foam fractionation processes were characterized by calculating procedural parameter including rhamnolipid and bacterial enrichment, rhamnolipid recovery, YX/S, YP/X, and specific as well as volumetric productivities. Comparing foam fractionation parameters of the rhamnolipid process with the surfactin process a high effectiveness of the integrated foam fractionation for rhamnolipid production was demonstrated.Electronic supplementary materialThe online version of this article (doi:10.1186/s13568-016-0183-2) contains supplementary material, which is available to authorized users.

The human pathogenic bacterium Pseudomonas aeruginosa produces rhamnolipids, glycolipids with functions for bacterial motility, biofilm formation, and uptake of hydrophobic substrates. Rhamnolipids represent a chemically heterogeneous group of secondary metabolites composed of one or two rhamnose molecules linked to one or mostly two 3-hydroxyfatty acids of various chain lengths. The biosynthetic pathway involves rhamnosyltransferase I encoded by the rhlAB operon, which synthesizes 3-(3-hydroxyalkanoyloxy)alkanoic acids (HAAs) followed by their coupling to one rhamnose moiety. The resulting mono-rhamnolipids are converted to di-rhamnolipids in a third reaction catalyzed by the rhamnosyltransferase II RhlC. However, the mechanism behind the biosynthesis of rhamnolipids containing only a single fatty acid is still unknown. To understand the role of proteins involved in rhamnolipid biosynthesis the heterologous expression of rhl-genes in non-pathogenic Pseudomonas putida KT2440 strains was used in this study to circumvent the complex quorum sensing regulation in P. aeruginosa. Our results reveal that RhlA and RhlB are independently involved in rhamnolipid biosynthesis and not in the form of a RhlAB heterodimer complex as it has been previously postulated. Furthermore, we demonstrate that mono-rhamnolipids provided extracellularly as well as HAAs as their precursors are generally taken up into the cell and are subsequently converted to di-rhamnolipids by P. putida and the native host P. aeruginosa. Finally, our results throw light on the biosynthesis of rhamnolipids containing one fatty acid, which occurs by hydrolyzation of typical rhamnolipids containing two fatty acids, valuable for the production of designer rhamnolipids with desired physicochemical properties.

Heterologous mono-rhamnolipid production by Pseudomonas putida KT2440 pSynPro8oT_rhlAB using glucose as the single carbon source was characterized in fed-batch bioreactor cultivations. For the described experiments, a defined mineral salt medium was used, and a two phase glucose feeding profile was applied, which yielded a final rhamnolipid concentration of 14.9 g/L. Applying the feeding profile, glucose stayed almost constant until 28 h of cultivation and decreased afterwards to limiting levels. Until the end of cultivation 253.0 ± 0.1 g glucose was added to the bioreactor of which a total of 252.0 ± 0.6 g glucose was metabolized. By modeling the fed-batch bioreactor cultivations the time courses of generated biomass, rhamnolipid and consumed glucose were described. The model was furthermore used to derive key process parameters from the collected data. The obtained values for the specific product formation rates (qRL) reached 18 mg/(g h) and yield coefficients (YRL/S) 10 mg/g respectively.

Summary Bacillus subtilis GLB191 (hereafter GLB191) is an efficient biological control agent against the biotrophic oomycete Plasmopara viticola , the causal agent of grapevine downy mildew. In this study, we show that GLB191 supernatant is also highly active against downy mildew and that the activity results from both direct effect against the pathogen and stimulation of the plant defences (induction of defence gene expression and callose production). High‐performance thin‐layer chromatography analysis revealed the presence of the cyclic lipopeptides fengycin and surfactin in the supernatant. Mutants affected in the production of fengycin and/or surfactin were thus obtained and allowed us to show that both surfactin and fengycin contribute to the double activity of GLB191 supernatant against downy mildew. Altogether, this study suggests that GLB191 supernatant could be used as a new biocontrol product against grapevine downy mildew.

Surfactants are an important class of industrial chemicals. Nowadays oleochemical surfactants such as alkyl polyglycosides (APGs) become increasingly important. This trend towards the utilization of renewable resources continues and consumers increasingly demand for environmentally friendly products. Consequently, research in microbial surfactants has drastically increased in the last years. While for mannosylerythritol lipids and sophorolipids established industrial processes exist, an implementation of other microbially derived surfactants has not yet been achieved. Amongst these biosurfactants, rhamnolipids synthesized by Pseudomonas aeruginosa and surfactin produced by Bacillus subtilis are so far the most analyzed biosurfactants due to their exceptional properties and the concomitant possible applications. In this review, a general overview is given regarding the current status of biosurfactants and benefits attributed to these molecules. Furthermore, the most recent research approaches for both rhamnolipids and surfactin are presented with respect to possible methods for industrial processes and the occurring drawbacks and limitations researchers have to address and overcome.