Regulation of carbon flux and NADH/NAD+ supply to enhance 2,3-butanediol production in Enterobacter aerogenes.

As a valuable platform chemical, 2,3-Butanediol (2,3-BDO) has a variety of industrial applications, and its microbial production is particularly attractive as an alternative to petroleum-based production. In this study, the regulation of intracellular carbon flux and NADH/NAD+ was used to increase the 2,3-BDO production of Enterobacter aerogenes. The genes encoding lactate dehydrogenase (ldh) and pyruvate formate lyase (pfl) were disrupted using the λ-Red recombination method and CRISPR-Cas9 to reduce the production of several byproducts and the consumption of NADH. Knockout of ldh or pfl increased intracellular NADH/NAD+ by 111 % and 113 %, respectively. Moreover, two important genes in the 2,3-BDO biosynthesis pathway, acetolactate synthase (budB) and acetoin reductase (budC), were overexpressed in E. aerogenes to further amply the metabolic flux toward 2,3-BDO production. And the overexpression of budB or budC increased intracellular NADH/NAD+ by 46 % and 57 %, respectively. In shake-flask cultivation with sucrose as carbon source, the 2,3-BDO titer of the IAM1183-LPBC was 3.55 times that of the wild type. In the 5-L fermenter, the maximal 2,3-BDO production produced by the IAM1183-LPBC was 2.88 times that of the original strain. This work offers new ideas for promoting the biosynthesis of 2,3-BDO for industrial applications.

Volatile fatty acids as an added value from biowaste.

The aim of the present work was to provide proof of concept of employing a co-culture of K. mobilis and E. coli for producing short and medium chain volatile fatty acids (VFAs) from kitchen biowaste and potato peels. To this aim, experiments were carried out at pilot-scale installation with a bioreactor of 250L. Different feeding strategies were tested under microaerobic conditions, at pH 6.0-6.5 in order to enhance chain elongation. Acetic acid and ethanol were dominating products in the initial stages of the bioprocess, but in a relatively short time of approx. 20-22h from the process start accumulation of propionic acid took place followed by a chain elongation to butyric and valeric acids. The highest final products yield of 325mg/g TS was achieved for the substrate load of 99.1g TS/L (VS of 91.1g/L) and pH 6.5, with the productivity of 448mg/L/h. However, the highest average VFAs chain length (3.77C) was observed in the process run with the loading of 63.2g TS/L and pH 6.0. In this study, we demonstrated that the existing symbiosis of the co-culture of K. mobilis and E. coli favours formation and chain elongation of VFA, induced most likely by the enhanced ethanol formation. Our finding differs from the previous research which focus mostly on anaerobic conditions of VFAs production. The results provide good basis for further optimisation of VFAs production process.

Biohydrogen and polyhydroxyalkanoate co-production by Enterobacter aerogenes and Rhodobacter sphaeroides from Calophyllum inophyllum oil cake.

The feasibility of coupled biohydrogen and polyhydroxyalkanoate production by Enterobacter aerogenes and Rhodobacter sphaeroides using Calophyllum inophyllum oil cake was studied under dark and photo fermentation conditions. The utilization of a non-edible acidic oil cake (C. inophyllum), and exploitation of a modified minimal salt media led to reduction in the cost of media. Cost of fermentation is reduced by implementation of alternate dark-photo fermentative periods and through the use of a co-culture consisting of a dark fermentative (E. aerogenes) and a photo fermentative (R. sphaeroides) bacterium. The biohydrogen and polyhydroxyalkanoate produced were 7.95 L H2/L media and 10.73 g/L media, respectively, under alternate dark and photo fermentation and were 3.23 L H2/L media and 5.6g/L media, respectively under complete dark fermentation. The characteristics of the oil cake and alternate dark (16 h) and photo (8h) fermentative conditions were found to be supportive in producing high biohydrogen and polyhydroxyalkanoate (PHA) yield.

Essential oils from Moroccan plants as potential chemosensitisers restoring antibiotic activity in resistant Gram-negative bacteria.

Bacterial drug resistance is a worrying public health problem. Antibiotic efflux is a major non-specific resistance mechanism used by bacteria, and efflux pumps are involved in the low-level susceptibility of various important Gram-negative pathogens. Use of molecules that can block bacterial pumps is an attractive strategy, but several studies report only partial efficacy owing to limits of these molecules (stability, selectivity, bioavailability, toxicity, etc.). The objective of this study was to search for natural sources of molecules able to inhibit efflux pump systems of resistant Gram-negative bacteria (Escherichia coli, Enterobacter aerogenes, Klebsiella pneumoniae, Salmonella enterica serotype Typhimurium and Pseudomonas aeruginosa). The results indicate that the studied essential oils exhibit interesting activity against the tested bacteria. This activity was significantly enhanced in the presence of an efflux pump inhibitor such as phenylalanine arginyl ß-naphthylamide (PAßN). The role of lipopolysaccharide (LPS) structure in the effect of essential oils was also reported in Salmonella LPS deep-rough mutants. In addition, essential oils of Thymus maroccanus and Thymus broussonetii, used at a low concentration (a fraction of the minimum inhibitory concentration), are able to significantly increase chloramphenicol susceptibility of several resistant isolates. These results demonstrate that these essential oils can alter efflux pump activity and may be attractive candidates to develop new drugs for chemosensitising multidrug-resistant strains to clinically used antibiotics.

Antioxidative and antibacterial effects of seeds and fruit rind of nutraceutical plants belonging to the Fabaceae family.

In the present study, the seeds and fruit rind of six plants of the Fabaceae family were selected to evaluate their potential as antioxidant and antibacterial agents. The dried powders were individually extracted with various organic solvents by the cold percolation method, were evaluated for antibacterial activity and methanol extracts used for antioxidant activities. Total phenol, protein and sugar contents were also measured. Antioxidant activities were measured by DPPH free radical scavenging activity, superoxide anion radical scavenging activity and reducing capacity assessment. Antibacterial activity was measured by the agar well diffusion method against four Gram positive and four Gram negative bacteria. The methanol extract of the fruit rind of C. indica showed the maximum DPPH free radical scavenging activity, superoxide anion radical scavenging activity, a high reducing capacity assessment and also had the highest total phenol content. There was a direct correlation between the phenol content and the antioxidant activity. The antibacterial activity of all the extracts was more pronounced on Gram positive bacteria than on Gram negative bacteria. Thus, the fruit rind of C. indica showed the best antioxidant and antibacterial activities.