Employment of the CRISPR/Cas9 system to improve cellulase production in Trichoderma reesei.

Trichoderma reesei has been explored intensively in the laboratory and on an industrial scale for its highly potent cellulase secretion machinery since its characterization over 70 years ago. Emergence of new genetic tools over the past decade has strengthened the understanding of mechanism involved in transcription of cellulase genes in fungi and provided a boost to edit them at molecular level. Since several transcriptional factors work synergistically for cellulase expression in fungi; engineering of cellulase secretome for enhanced cellulase titer require combined manipulation of these factors. In the same context, CRISPR/Cas9 has emerged as a powerful, versatile genetic engineering tool for multiplex gene editing in fungi. It is true that considerable efforts with CRISPR technologies have largely developed fungal genetic engineering, but its application in fungi is still challenging and limited. The present review illustrates the precision, strengths and challenges of using CRISPR/Cas9 technology for cellulase engineering in T. reesei, highlighting key strategies that could be employed for strain improvement.

Integrated production of ethanol and xylitol from Brassica juncea using Candida sojae JCM 1644.

The present study demonstrates a novel strategy involving two-step fermentation of lignocellulosic hydrolysate for the integrated production of ethanol and xylitol using a newly isolated yeast strain, Candida sojae JCM 1644. The isolated strain was characterised by its carbohydrate assimilation efficiency and tolerance towards inhibitors generated during pretreatment and fermentation of lignocellulosic biomass. In brief, the study comprised alkali treatment of Brassica juncea followed by its saccharification with cellulase consortia. An isolated strain was used for the co-production of xylitol and ethanol from sugar hydrolysate, and several parameters were systematically optimised for maximum co-production of ethanol and xylitol. Out of total glucose (149.72 g/L) and xylose (84.21 g/L) present in biomass hydrolysate, a product yield of 0.45 g/g (ethanol) and 0.62 g/g (xylitol) was achieved for a two-step fermentation process, which was 15.57% and 11.78% higher than the yield achieved for ethanol and xylitol, respectively, in a one-step fermentation process.

Seed bio-priming with tri-species consortia of phosphate solubilizing rhizobacteria (PSR) and its effect on plant growth promotion.

Three potential rhizobacteria namely Burkholderia gladioli (MTCC 10216), Pseudomonas sp. (MTCC 9002) and Bacillus subtilis (MTCC 8528) procured from IMTECH, Chandigarh (India) were evaluated individually and as consortia for its phosphate (P) solubilizing ability and effect of growth of fenugreek (Trigonella foenum-graecum L.) and tomato (Lycopersicon esculentum L.). Phosphate solubilizing ability of these strains individually and as consortia was tested on Pikovskayas agar medium, Phosphate solubilizing agar medium and National Botanical Research Institute phosphate agar medium containing six different sources of insoluble inorganic phosphate such as tri-calcium phosphate (TCP), di-calcium phosphate (DCP), zinc phosphate (ZP), ferric phosphate (FP), sodium di-hydrogen phosphate (SP), and aluminum phosphate (AP), and two organic P such as calcium and sodium phytate. The maximum P solubilizing ability was recorded in consortium-4 having all three potential bacterial strains. Phosphate solubilization after 7th day of incubation was 37.9 mg/100 ml of TCP, 40.01 mg/100 ml of DCP, 15.79 mg/100 ml of FP, 43.02 mg/100 ml of SP, no solubilization of ZP and AP, 39.75 mg/100 ml of calcium phytate and 24.01mg/100 ml of sodium phytate. Seed germination and the other plant parameters such as plant height and weight significantly increased in fenugreek and tomato seeds, bio-primed with consortium-4 followed by consortium-3. After bio-priming of seeds in pot assay, the level of phosphorus in soil got increased by 54% in consortium-4 treated soil followed by consortium-3 (47%) over untreated control soil. Based on these findings, consoritium-4 could be recommended as a good bio-inoculant for fenugreek, tomato and other crops in comparison to individual strains and other consortia.

Potentiation of antimicrobial activity of ciprofloxacin by Pelargonium graveolens essential oil against selected uropathogens.

The recent approach of using herbs and antibiotics in combination constitutes a strategy to overcome the problems of resistance and side effects associated with conventional antibiotics. In the present study, the antimicrobial effect of Pelargonium graveolens L' Hér essential oil in combination with ciprofloxacin was evaluated on uropathogens, namely, Klebsiella pneumoniae KT2, Proteus mirabilis PRT3 and Staphylococcus aureus ST2. Minimum inhibitory concentrations of P. graveolens essential oil and ciprofloxacin were determined by the microbroth dilution method and further, the interaction between these two agents was studied by a checkerboard method. The fractional inhibitory concentration index (FICI) was calculated to be 0.375 for both K. pneumoniae KT2 and P. mirabilis PRT3, while for S. aureus ST2 it was found to be 0.5. The values of FICI for the tested microorganisms were found to be ≤0.5, which indicates synergism between P. graveolens essential oil and ciprofloxacin. The concave shaped curve in the isobolograms also depicted a synergistic effect of P. graveolens essential oil and ciprofloxacin against the tested microorganisms. Hence, the synergistic action of P. graveolens essential oil and ciprofloxacin may be applied for the treatment of UTIs, which have hitherto been treated by using only synthetic drugs.