Bacterial Cellulose-Based Blends and Composites: Versatile Biomaterials for Tissue Engineering Applications.

Cellulose of bacterial origin, known as bacterial cellulose (BC), is one of the most versatile biomaterials that has a huge potential in tissue engineering due to its favourable mechanical properties, high hydrophilicity, crystallinity, and purity. Additional properties such as porous nano-fibrillar 3D structure and a high degree of polymerisation of BC mimic the properties of the native extracellular matrix (ECM), making it an excellent material for the fabrication of composite scaffolds suitable for cell growth and tissue development. Recently, the fabrication of BC-based scaffolds, including composites and blends with nanomaterials, and other biocompatible polymers has received particular attention owing to their desirable properties for tissue engineering. These have proven to be promising advanced materials in hard and soft tissue engineering. This review presents the latest state-of-the-art modified/functionalised BC-based composites and blends as advanced materials in tissue engineering. Their applicability as an ideal biomaterial in targeted tissue repair including bone, cartilage, vascular, skin, nerve, and cardiac tissue has been discussed. Additionally, this review briefly summarises the latest updates on the production strategies and characterisation of BC and its composites and blends. Finally, the challenges in the future development and the direction of future research are also discussed.

Agr Quorum Sensing influences the Wood-Ljungdahl pathway in Clostridium autoethanogenum.

Acetogenic bacteria are capable of fermenting CO2 and carbon monoxide containing waste-gases into a range of platform chemicals and fuels. Despite major advances in genetic engineering and improving these biocatalysts, several important physiological functions remain elusive. Among these is quorum sensing, a bacterial communication mechanism known to coordinate gene expression in response to cell population density. Two putative agr systems have been identified in the genome of Clostridium autoethanogenum suggesting bacterial communication via autoinducing signal molecules. Signal molecule-encoding agrD1 and agrD2 genes were targeted for in-frame deletion. During heterotrophic growth on fructose as a carbon and energy source, single deletions of either gene did not produce an observable phenotype. However, when both genes were simultaneously inactivated, final product concentrations in the double mutant shifted to a 1.5:1 ratio of ethanol:acetate, compared to a 0.2:1 ratio observed in the wild type control, making ethanol the dominant fermentation product. Moreover, CO2 re-assimilation was also notably reduced in both hetero- and autotrophic growth conditions. These findings were supported through comparative proteomics, which showed lower expression of carbon monoxide dehydrogenase, formate dehydrogenase A and hydrogenases in the ∆agrD1∆agrD2 double mutant, but higher levels of putative alcohol and aldehyde dehydrogenases and bacterial micro-compartment proteins. These findings suggest that Agr quorum sensing, and by inference, cell density play a role in carbon resource management and use of the Wood-Ljungdahl pathway as an electron sink.

Effective pretreatment of lignocellulosic co-substrates using barley straw-adapted microbial consortia to enhanced biomethanation by anaerobic digestion.

Microbial pretreatments have been identified as a compatible and sustainable process with anaerobic digestion compared to energy-intensive physicochemical pretreatments. In this study, barley straw and hay co-substrate was pretreated with a microaerobic barley straw-adapted microbial (BSAM) consortium prior to anaerobic digestion. The improved digestibility was investigated through 16S rRNA gene sequencing, microbial counts and C:N ratios. BSAM pretreatment resulted in 15.2 L kg-1 TS of methane yield after 35 days, almost 40 times more than the control. The methane content in total biogas produced were 58% (v/v) and 10% (v/v) in BSAM and control, respectively. This research demonstrated that BSAM-based pretreatment significantly increased the digestibility and surface area of the lignocellulosic material and considerably enhanced biomethanation. This study generates new potential bio-research opportunities in the emerging field of lignocellulosic anaerobic digestion-biorefineries.

Influence of nutrient status on the biohydrogen and lipid productivity in Parachlorella kessleri: a biorefinery approach.

The commercial reality of microalgal biotechnology for the production of individual bioactives is constrained by the high cost of production and requires a biorefinery approach. In this investigation, we examined the influence of different nutrient deprivation (nitrogen (N), phosphorus (P), sulphur (S) and manganese (Mn)) on growth, chlorophyll a (Chl a), biohydrogen (H2) and fatty acid profiles in Parachlorella kessleri EMCCN 3073 under both aerobic and anaerobic conditions. Anaerobic conditions combined with the nutrient deprivation resulted in cell division blockage, reduction in Chl a and remarkable changes in pH, whereas a significant increase in the H2 production was observed after 24 h. The highest cumulative H2 productivity was observed in N-deficient medium (300 µL/L, day 9) followed by Mn-deficient medium (250 µL/L, day 7). The highest H2 production rate (3.37 µL/L/h) was achieved by Mn-deficient medium after 24 h. In terms of fatty acid composition, P. kessleri exhibited a differential response to different nutrient stresses. Under aerobic conditions, N-deficient media resulted in the highest lipid content (119% compared to control, day 7), whereas earlier lipid induction at (1-3 days) was observed with Mn- and S-deficient media with 18-91% and 25-34% increase, respectively, compared with the replete control. Meanwhile, higher lipid content was observed under anaerobic conditions combined with Mn-, N-, P- and S-deprived media (day 1) with 20%, 13%, 8% and 7% increases respectively compared with the control. This investigation, for the first time clearly, highlights the potential of P. kessleri as a sustainable biorefinery platform, for H2 and fatty acid bio-production under anaerobic conditions. KEY POINTS: • Parachlorella kessleri could provide a future sustainable biorefinery platform. • Nutrient-deprived anaerobic conditions blocked cell growth but differentially induced H2 production. • Nutrient status, under both aerobic/anaerobic conditions, alters lipids and fatty acids profile of P. kessleri. • Nutrient-deprived (N- and Mn-) anaerobic conditions: future biorefinery platform.

Publisher Correction: Deciphering the unique cellulose degradation mechanism of the ruminal bacterium Fibrobacter succinogenes S85.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Deciphering the unique cellulose degradation mechanism of the ruminal bacterium Fibrobacter succinogenes S85.

Fibrobacter succinogenes S85, isolated from the rumen of herbivores, is capable of robust lignocellulose degradation. However, the mechanism by which it achieves this is not fully elucidated. In this study, we have undertaken the most comprehensive quantitative proteomic analysis, to date, of the changes in the cell envelope protein profile of F. succinogenes S85 in response to growth on cellulose. Our results indicate that the cell envelope proteome undergoes extensive rearrangements to accommodate the cellulolytic degradation machinery, as well as associated proteins involved in adhesion to cellulose and transport and metabolism of cellulolytic products. Molecular features of the lignocellulolytic enzymes suggest that the Type IX secretion system is involved in the translocation of these enzymes to the cell envelope. Finally, we demonstrate, for the first time, that cyclic-di-GMP may play a role in mediating catabolite repression, thereby facilitating the expression of proteins involved in the adhesion to lignocellulose and subsequent lignocellulose degradation and utilisation. Understanding the fundamental aspects of lignocellulose degradation in F. succinogenes will aid the development of advanced lignocellulosic biofuels.

Quantitative proteomic analysis of the influence of lignin on biofuel production by Clostridium acetobutylicum ATCC 824.

BACKGROUND: Clostridium acetobutylicum has been a focus of research because of its ability to produce high-value compounds that can be used as biofuels. Lignocellulose is a promising feedstock, but the lignin-cellulose-hemicellulose biomass complex requires chemical pre-treatment to yield fermentable saccharides, including cellulose-derived cellobiose, prior to bioproduction of acetone-butanol-ethanol (ABE) and hydrogen. Fermentation capability is limited by lignin and thus process optimization requires knowledge of lignin inhibition. The effects of lignin on cellular metabolism were evaluated for C. acetobutylicum grown on medium containing either cellobiose only or cellobiose plus lignin. Microscopy, gas chromatography and 8-plex iTRAQ-based quantitative proteomic technologies were applied to interrogate the effect of lignin on cellular morphology, fermentation and the proteome. RESULTS: Our results demonstrate that C. acetobutylicum has reduced performance for solvent production when lignin is present in the medium. Medium supplemented with 1 g L(-1) of lignin led to delay and decreased solvents production (ethanol; 0.47 g L(-1) for cellobiose and 0.27 g L(-1) for cellobiose plus lignin and butanol; 0.13 g L(-1) for cellobiose and 0.04 g L(-1) for cellobiose plus lignin) at 20 and 48 h, respectively, resulting in the accumulation of acetic acid and butyric acid. Of 583 identified proteins (FDR < 1 %), 328 proteins were quantified with at least two unique peptides. Up- or down-regulation of protein expression was determined by comparison of exponential and stationary phases of cellobiose in the presence and absence of lignin. Of relevance, glycolysis and fermentative pathways were mostly down-regulated, during exponential and stationary growth phases in presence of lignin. Moreover, proteins involved in DNA repair, transcription/translation and GTP/ATP-dependent activities were also significantly affected and these changes were associated with altered cell morphology. CONCLUSIONS: This is the first comprehensive analysis of the cellular responses of C. acetobutylicum to lignin at metabolic and physiological levels. These data will enable targeted metabolic engineering strategies to optimize biofuel production from biomass by overcoming limitations imposed by the presence of lignin.

Influence of Substrates on the Surface Characteristics and Membrane Proteome of Fibrobacter succinogenes S85.

Although Fibrobacter succinogenes S85 is one of the most proficient cellulose degrading bacteria among all mesophilic organisms in the rumen of herbivores, the molecular mechanism behind cellulose degradation by this bacterium is not fully elucidated. Previous studies have indicated that cell surface proteins might play a role in adhesion to and subsequent degradation of cellulose in this bacterium. It has also been suggested that cellulose degradation machinery on the surface may be selectively expressed in response to the presence of cellulose. Based on the genome sequence, several models of cellulose degradation have been suggested. The aim of this study is to evaluate the role of the cell envelope proteins in adhesion to cellulose and to gain a better understanding of the subsequent cellulose degradation mechanism in this bacterium. Comparative analysis of the surface (exposed outer membrane) chemistry of the cells grown in glucose, acid-swollen cellulose and microcrystalline cellulose using physico-chemical characterisation techniques such as electrophoretic mobility analysis, microbial adhesion to hydrocarbons assay and Fourier transform infra-red spectroscopy, suggest that adhesion to cellulose is a consequence of an increase in protein display and a concomitant reduction in the cell surface polysaccharides in the presence of cellulose. In order to gain further understanding of the molecular mechanism of cellulose degradation in this bacterium, the cell envelope-associated proteins were enriched using affinity purification and identified by tandem mass spectrometry. In total, 185 cell envelope-associated proteins were confidently identified. Of these, 25 proteins are predicted to be involved in cellulose adhesion and degradation, and 43 proteins are involved in solute transport and energy generation. Our results supports the model that cellulose degradation in F. succinogenes occurs at the outer membrane with active transport of cellodextrins across for further metabolism of cellodextrins to glucose in the periplasmic space and inner cytoplasmic membrane.

Novel bacteriophage therapy for controlling metallo-beta-lactamase producing Pseudomonas aeruginosa infection in catfish.

BACKGROUND: The bacteriophage therapy is an effective antimicrobial approach with potentially important applications in medicine and biotechnology which can be seen as an additional string in the bow. Emerging drug resistant bacteria in aquaculture industry due to unrestricted use of antibiotics warrants more sustainable and environmental friendly strategies for controlling fish infections.The isolated bacteria from fish lesions was characterised based on isolation on selective and differential medium like Pseudomonas agar, gram staining, biochemical tests and 16SrRNA sequencing. The metallo-beta-lactamase (MBL) producing bacterial isolate was evaluated using Imipenem - Ethylenediaminetetraacetic acid (EDTA) disk method. The specific bacteriophage was isolated and concentrated using coal bed developed in our lab at CSIR-NEERI. The isolated and enriched bacteriophage was characterised by nucleotide sequencing and electron microscopy. The phage therapy was applied for treating ulcerative lesion in fish. RESULTS: The pathogenic bacterium responsible for causing ulcerative lesions in catfish species (Clarias gariepinus) was identified as Pseudomonas aeruginosa. One out of twenty P. aeruginosa isolate showing multi drug resistance (MDR) was incidentally found to be MBL producing as determined by Imipenem-EDTA disk method. The phage therapy effectively cured the ulcerative lesions of the infected fish in 8-10 days of treatment, with a sevenfold reduction of the lesion with untreated infection control. CONCLUSION: Bacteriophage therapy can have potential applications soon as an alternative or as a complement to antibiotic treatment in the aquaculture. We present bacteriophage therapy as a treatment method for controlling MDR P. aeruginosa infection in C. gariepinus. To the best of our knowledge this is a first report of application of phage therapy against MBL producing P. aeruginosa isolated from aquatic ecosystem.