In vitro and in vivo evaluation of probiotic potential and safety assessment of Bacillus coagulans SKB LAB-19 (MCC 0554) in humans and animal healthcare.

Bacillus coagulans is Gram positive, spore forming and high lactic acid producing bacteria; however, probiotic and safety assessment of the isolated strain need to be investigated for commercial applications. Current study aimed to screen SKB LAB-19 for potential probiotic characteristics viz. enzyme production, antimicrobial properties, pH/bile salt tolerance, temperature stability, antidiarrheal activity in Swiss albino mice and Wistar rats; and acute oral toxicity in mice. The results showed that, SKB LAB-19 produces eight potential enzymes, effective against E. coli and C. perfringensis, tolerant to bile salt (0.3%w/v)/gastric pH (2.5), stable at 40-90 °C and nontoxic to cells. SKB LAB-19 was found to be safe and displayed promising results to reverse E. coli and castor oil induced diarrhoea. Histopathological studies showed repair to damaged mucosal epithelium cells and improves integrity of the goblet cells of colon. SKB LAB-19 showed immunomodulatory effects with increased immunoglobulins in blood and augmented weight of spleen and thymus. In addition, SKB LAB-19 showed significant in-vitro antioxidant activity (82.93%), reducing capacity and ascorbate auto-oxidation inhibition effect (94.62%). These preliminary results suggested that, SKB LAB-19 was found to be safe and has the potential to be used as effective probiotic and anti-diarrhoeal agent in humans and animal healthcare.

Investigation on mycelial growth requirements of Cantharellus cibarius under laboratory conditions.

The golden chanterelle represents one of the commonly found, edible mushrooms that is highly valued in various cuisines. The present study focused on assessing the requirements of Cantharellus cibarius such as pH, temperature, as well as the carbon and nitrogen sources for mycelial growth. Optimization of the growth parameters was carried out by one-factor-at-a-time method. The optimal pH and temperature were determined to be 6.0 and 22.5 °C, respectively. Among the various carbon sources studied, sucrose at a concentration of 2% gave maximum mycelial growth and proved to be the most suitable one. Amongst the nitrogen sources studied, peptone, ammonium sulphate, and sodium nitrate, gave the maximum mycelial growth at an optimized concentration of 0.5%. In the presence of beef extract and yeast extract, a change in colony pigmentation from yellow to dark grey was observed. Finally, the carbon to nitrogen ratio of 2:0.5 proved to be optimal for mycelial growth. This study is the first report on the optimisation of in vitro growth requirements of C. cibarius.

Emergent Approaches to Efficient and Sustainable Polyhydroxyalkanoate Production.

Petroleum-derived plastics dominate currently used plastic materials. These plastics are derived from finite fossil carbon sources and were not designed for recycling or biodegradation. With the ever-increasing quantities of plastic wastes entering landfills and polluting our environment, there is an urgent need for fundamental change. One component to that change is developing cost-effective plastics derived from readily renewable resources that offer chemical or biological recycling and can be designed to have properties that not only allow the replacement of current plastics but also offer new application opportunities. Polyhydroxyalkanoates (PHAs) remain a promising candidate for commodity bioplastic production, despite the many decades of efforts by academicians and industrial scientists that have not yet achieved that goal. This article focuses on defining obstacles and solutions to overcome cost-performance metrics that are not sufficiently competitive with current commodity thermoplastics. To that end, this review describes various process innovations that build on fed-batch and semi-continuous modes of operation as well as methods that lead to high cell density cultivations. Also, we discuss work to move from costly to lower cost substrates such as lignocellulose-derived hydrolysates, metabolic engineering of organisms that provide higher substrate conversion rates, the potential of halophiles to provide low-cost platforms in non-sterile environments for PHA formation, and work that uses mixed culture strategies to overcome obstacles of using waste substrates. We also describe historical problems and potential solutions to downstream processing for PHA isolation that, along with feedstock costs, have been an Achilles heel towards the realization of cost-efficient processes. Finally, future directions for efficient PHA production and relevant structural variations are discussed.

Cross-linked enzyme aggregates of arylamidase from Cupriavidus oxalaticus ICTDB921: process optimization, characterization, and application for mitigation of acrylamide in industrial wastewater.

Acrylamidase produced by Cupriavidus oxalaticus ICTDB921 was recovered directly from the fermentation broth by ammonium sulfate (40-50%) precipitation and then stabilized by cross-linking with glutaraldehyde. The optimum conditions for the preparation of cross-linked enzyme aggregates of acrylamidase (acrylamidase-CLEAs) were using 60 mM glutaraldehyde for 10 min at 35 °C and initial broth pH of 7.0. Acrylamidase-CLEAs were characterized by SDS-PAGE, FTIR, particle size analyzer and SEM. Cross-linking shifted the optimal temperature and pH from 70 to 50 °C and 5-7 to 6-8, respectively. It also altered the secondary structure fractions, pH and thermal stability along with the kinetic constants, Km and Vmax, respectively. A complete degradation of acrylamide ~ 1.75 g/L in industrial wastewater was achieved after 60 min in a batch process under optimum operating conditions, and the kinetics was best represented by Edward model (R2 = 0.70). Acrylamidase-CLEAs retained ~ 40% of its initial activity after three cycles for both pure acrylamide and industrial wastewater, and were stable for 15 days at 4 °C, retaining ~ 25% of its original activity.

Fermentative production of extracellular amylase from novel amylase producer, Tuber maculatum mycelium, and its characterization.

Truffles are symbiotic hypogeous edible fungi (form of mushroom) that form filamentous mycelia in their initial phase of the growth cycle as well as a symbiotic association with host plant roots. In the present study, Tuber maculatum mycelia were isolated and tested for extracellular amylase production at different pH on solid agar medium. Furthermore, the mycelium was subjected to submerged fermentation for amylase production under different culture conditions such as variable carbon sources and their concentrations, initial medium pH, and incubation time. The optimized conditions after the experiments included soluble starch (0.5% w/v), initial medium pH of 7.0, and incubation time of 7 days, at room temperature (22 ± 2 °C) under static conditions which resulted in 1.41 U/mL of amylase. The amylase thus obtained was further characterized for its biocatalytic properties and found to have an optimum activity at pH 5.0 and a temperature of 50 °C. The enzyme showed good thermostability at 50 °C by retaining 98% of the maximal activity after 100 min of incubation. The amylase activity was marginally enhanced in presence of Cu2+ and Na+ and slightly reduced by K+, Ca2+, Fe2+, Mg2+, Co2+, Zn2+, and Mn2+ ions at 1 mM concentration.

Mixed Culture Cultivation in Microbial Bioprocesses.

Mixed culture cultivation is well renowned for industrial applications due to its technological and economic benefits in bioprocess, food processing, and pharmaceutical industries. A mixed consortium encompasses to achieve growth in unsterile conditions, robustness to environmental stresses, perform difficult functions, show better substrate utilization, and increase productivity. Hence, mixed cultures are being valorized currently and has also augmented our understanding of microbial activities in communities. This chapter covers a wide range of discussion on recent improvements in mixed culture cultivation for microbial bioprocessing and multifarious applications in different areas. The history of microbial culture, microbial metabolism in mixed culture, biosynthetic pathway studies, isolation and identification of strains, along with the types of microbial interactions involved during their production and propagation, are meticulously detailed in the current chapter. Besides, parameters for evaluating mixed culture performance, large-scale production, and challenges associated with it are also discussed vividly. Microbial community, characteristics of single and mixed culture fermentation, and microbe-microbe interactions in mixed cultures have been summarized comprehensively. Lastly, various challenges and opportunities in the area of microbial mixed culture that are obligatory to improve the current knowledge of microbial bioprocesses are projected.

Assessment of physiochemical properties and concentration of heavy metals in agricultural soils fertilized with chemical fertilizers.

The present study aim to assess the effect of phosphate and urea fertilizers on the physicochemical properties, pH and electrical conductivity (EC) of the soil. The effect of these fertilizers on cation exchange capacity (CEC), organic matter (OM), and the possibility of contamination with heavy metals (HM) (Cr, Cu, Cd, Mn, Zn, Ni, Fe, and Pb) were studied on the soils of Alshati agricultural project at different seasons after forty years of fertilization. Uncultivated soil samples were also analyzed for comparison and considered as reference. Mean values of soil pH, EC, CEC, and OM ranged between 6.88-7.32, 0.14-0.26 µS/cm, 2.95-4.19 Cmol/kg and 0.49-0.53%, respectively, in all seasons. Concentrations of HMs were 9.50-38.38, 0.0-2.05, 0.0-0.47, 0.0-29.81, 0.0-13.85, 2.83-25.95 and 104-512.20 mg/kg respectively, for Cr, Cu, Cd, Mn, Zn, Ni and Fe. The concentrations of the HMs in the soil were found to be vary significantly with the seasons (winter, spring, summer, and autumn). However, no traces of Pb was found in the soil samples. The result showed a significant correlation between, pH, EC, CEC, OM and HM content of the soil. The geochemical index of contamination shows that there was no contamination with Cu, Cd, Zn, Pb, Cr, Mn, Ni, and Fe.

Chitosan coated calcium alginate beads for covalent immobilization of acrylamidase: Process parameters and removal of acrylamide from coffee.

This study reports on removal of acrylamide from roasted coffee by acrylamidase from Cupriavidus oxalaticus ICTDB921. Chitosan coated calcium alginate beads were functionalized with citric acid as nontoxic cross linker and activated by 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC) and N-hydroxysuccinimide (NHS) (1.66:1 w/w) for covalent immobilization of acrylamidase. The optimum beads were obtained using 5% sodium alginate, 1.5% chitosan, and 0.6 mol/L citric acid. The beads prepared at each step were characterized by FTIR and SEM. Coating of chitosan matrix on calcium alginate beads enhanced the mechanical stability over that of calcium alginate and/or chitosan. The immobilized acrylamidase showed optimum pH/temperature of 8.5/65 °C, improved pH/thermal/shelf stability, and retained 80% activity after four cycles. Haldane model could describe the degradation kinetics of acrylamide in batch study. In packed bed column, a bed height, feed flow rate and inlet acrylamide concentration of 20 cm, 1 mL/min, and 100 mg/L gave best results.

Magnetic cross-linked enzyme aggregates of acrylamidase from Cupriavidus oxalaticus ICTDB921 for biodegradation of acrylamide from industrial waste water.

Acrylamidase from Cupriavidus oxalaticus ICTDB921 was immobilized on magnetic nanoparticles (MNPs) for degradation of acrylamide (a group 2A carcinogen and an environmental contaminant) from industrial waste water. Acrylamidase-MNPs were prepared (maximum recovery ∼94%) at optimized process parameters viz. 1.5:1 (v/v) of acetone: crude acrylamidase/5 mM of glutaraldehyde/90 min/1.5:1 of enzyme: MNP ratio. MNPs and acrylamidase-MNPs were characterized by particle size analysis, FTIR, XRD, SEM and vibrating sample magnetometer. Acrylamidase-MNPs showed a shift in optimum pH (8-8.5) and temperature (60-65 °C) with higher pH/thermal stability vis-à-vis free enzyme. A significant increase in kinetic constants, thermal inactivation constants and thermodynamic parameters were noted for acrylamidase-MNPs. A complete degradation of acrylamide ∼2100 mg/L was achieved in industrial waste water under optimized conditions for batch process and the kinetics was best represented by Haldane model. Acrylamidase-MNPs retained >80% of its initial activity after 4 cycles for both pure acrylamide and industrial waste water.

Biodegradation of acrylamide by a novel isolate, Cupriavidus oxalaticus ICTDB921: Identification and characterization of the acrylamidase produced.

Acrylamide is neurotoxic, genotoxic, teratogenic and carcinogenic. Its widespread use in various industrial processes leads to environmental contamination. Acrylamidase produced by certain bacteria degrade acrylamide to acrylic acid and ammonia. The present study details the isolation and identification of soil bacterium which could degrade acrylamide. Among the 18 acrylamide-degrading isolates tested, isolate ICTDB921 demonstrated superior acrylamide degradation which was confirmed by HPLC, FTIR and GC-MS. The partial 16S rRNA sequencing confirmed the isolate to be Cupriavidus oxalaticus ICTDB921, which showed highest growth at 60 mM acrylamide, neutral pH and 30 °C. The kinetic model predictions were consistent with experimental results. The acrylamidase from this isolate showed potency at pH (6-8) and temperatures (30-60 °C), with reasonable pH (6-8) and thermal stability (upto 60 °C). The enzyme was stable against most metal ions and amino acids, and also degraded other aliphatic amides, demonstrating its potential in remediation of acrylamide from the environment and food systems.

Isolation and Characterization of Acrylamidase from Arthrobacter sp. DBV1 and Its Ability to Biodegrade Acrylamide.

Although acrylamide finds diverse industrial applications, its presence in the environment is hazardous due to its carcinogenic, neurotoxic, and teratogenic properties. In spite of the general toxicity of acrylamide in the monomer form, some microorganisms are able to use it as a source of energy by catabolizing it to ammonia and acrylic acid by means of acrylamidase (EC 3.5.1.4). The present work reports on a novel soil isolate as an acrylamide-degrading bacteria. Based on biochemical characterization and 16S ribosomal RNA (rRNA) gene sequence, the bacterial strain was identified as Gram-positive Arthrobacter sp. DBV1. The optimum growth conditions were found to be temperature (30 °C) and pH 6.0 to 7.0. Evaluation of the effect of concentration of acrylamide (10-50 mM) incorporated into minimal medium showed maximum growth of Arthrobacter sp. DBV1 at 30 mM acrylamide. The biodegradation of acrylamide was confirmed by HPLC analysis. Acrylamidase was isolated and characterized for temperature and pH optima, substrate specificity by using different amides, and the effect of different activators/inhibitors such as metal ions and amino acids. These finding suggests that the strain could be attractive for biodegradation of acrylamide from the environment and also possibly from foods containing preformed acrylamide.

Biochemical Characterization of Extracellular Cellulase from Tuber maculatum Mycelium Produced Under Submerged Fermentation.

Interaction of truffle mycelium with the host plant involves the excretion of extracellular enzymes. The ability of Tuber maculatum mycelium to produce an extracellular cellulase during submerged fermentation was demonstrated for the first time. T. maculatum mycelia were isolated and tested for extracellular cellulase production at variable pH on solid agar medium, and the highest activity was observed at pH 7.0. Furthermore, T. maculatum was subjected to submerged fermentation in basal salt medium for cellulase production. Under optimized conditions using sodium carboxymethyl cellulose (0.5 % w/v) as carbon source and an initial pH of 7.0, the enzyme production yielded 1.70 U/mL of cellulase in the cell-free supernatant after 7 days of incubation time. The optimum of the obtained cellulase's activity was at pH 5.0 and a temperature of 50 °C. The enzyme showed good thermostability at 50 °C by retaining 99 % of its maximal activity over an incubation time of 100 min. The cellulase activity was inhibited by Fe2+ and slightly activated by Mn2+ and Cu2+ at 1 mM concentration. The results indicated that truffle mycelium is utilizing cellulosic energy source in the root system, and the optimal conditions are those existing in the acidic Finnish soil.

Characterization of natural habitats and diversity of Libyan desert truffles.

Desert truffles have traditionally been used as food in Libya. Desert truffle grows and gives fruit sporadically when adequate and properly distributed rainfall occurs with existence of suitable soil and mycorrhizal host plant. The present study aimed to identify and characterize two kinds of wild desert truffles from ecological and nutritional points that were collected from the studied area. The truffle samples were identified as Terfezia (known as red or black truffle) and Tirmania (known as white truffle). The nutritional values (protein, lipid and carbohydrate) of both Libyan wild truffle (Terfezia and Tirmania) were determined on a dry weight basis and result showed that Tirmania and Terfezia contained 16.3 and 18.5% protein, 6.2 and 5.9% lipid, 67.2 and 65% carbohydrate, respectively, in ascocarp biomass. The soil pH of the upper and lower regions of the Hamada Al-Hamra ranged between 8.2 and 8.5 giving suitable conditions for fructification. The plants, Helianthemum kahiricum and Helianthemum lippii were the dominant plants in Hamada Al-Hamra region found to form a mycorrhiza with desert truffles. The phylogenetic analysis of the genomic rDNA ITS region showed that, out of five collections three represented Tirmania pinoyi (Maire) Malencon, one Tirmania nivea (Desf.) Trappe, and one Terfezia boudieri Chatin.