Vegan grade medium component screening and concentration optimization for the fermentation of the probiotic strain Lactobacillus paracasei IMC 502® using Design of Experiments.

Lactobacillus paracasei IMC502® is a commercially successful probiotic strain. However, there are no reports that investigate growth medium composition in relation to improved biomass production for this strain. The major outcome of the present study is the design and optimization of a growth medium based on vegan components to be used in the cultivation of Lactobacillus paracasei IMC502®, by using Design of Experiments. Besides comparing different carbon sources, the use of plant-based peptones as nitrogen sources was considered. In particular, the use of guar peptone as the main nitrogen source, in the optimization of fermentation media for the production of probiotics, could replace other plant peptones (e.g. potato, rice, wheat, and soy) which are part of the human diet, thereby avoiding an increase in product and process prices. A model with R2 and adjusted R2 values higher than 95% was obtained. Model accuracy was equal to 94.11%. The vegan-optimized culture medium described in this study increased biomass production by about 65% compared to growth on De Man-Rogosa-Sharpe (MRS) medium. Moreover, this approach showed that most of the salts and trace elements generally present in MRS are not affecting biomass production, thus a simplified medium preparation can be proposed with higher probiotic biomass yield and titer. The possibility to obtain viable lactic acid bacteria at high density from vegetable derived nutrients will be of great interest to specific consumer communities, opening the way to follow this approach with other probiotics of impact for human health.

Characterization of Salmonella enterica Biofilms and Antibiofilm Effect of Carvacrol and 2-Aminobenzimidazole.

Biofilm-associated foodborne Salmonella infections in poultry have become increasingly challenging for veterinarians, particularly in developing countries, and warrant thorough investigation. We assessed the biofilm-forming tendency of poultry isolates of Salmonella enterica, namely Salmonella Typhimurium (n = 23), Salmonella Infantis (n = 28), and Salmonella Heidelberg (n = 18), in nutrient-rich Rappaport-Vassiliadis Soya (RVS) peptone broth and nutrient-deficient diluted Tryptone Soya Broth (TSB). Seven of the tested isolates exhibited moderate biofilm formation in diluted TSB, whereas two showed such formation in RVS. In addition, the Congo red agar assay revealed curli and cellulose production in seven isolates. Fourteen specific biofilm-associated genes were analyzed identifying sdiA and seqA to be the most prevalent (100%), and glyA the least prevalent (69.5%). The prevalence of the genes bcsA and csgA was significantly lower in moderate and weak biofilm formers, respectively, as compared with nonbiofilm formers in RVS peptone broth. Furthermore, the compounds carvacrol and 2-aminobenzimidazole (2-ABI) effectively inhibited biofilm formation by Salmonella serovars in RVS peptone and TSB media, respectively. Whereas the antibiofilm activity of 2-ABI against Salmonella has not been reported previously, we determined its most effective concentration at 1.5 mM among tested antibiofilm treatments. These findings indicate that Salmonella strains prevalent in poultry farms have the potential to form biofilms, and the tested compounds should be further explored as supportive or alternative antimicrobials.

Xylitol biosynthesis enhancement by Candida tropicalis via medium, process parameter optimization, and co-substrate supplementation.

The present study examines the impact of nitrogen sources (yeast extract, ammonium sulfate peptone, ammonium nitrate, urea, and sodium nitrate), salt solution (0.5 g/L MgSO4, 0.5 g/L KH2PO4, 0.3 g/L CaCl2), trace elements solution (0.1 g/L CuSO4, 0.1 g/L FeSO4, 0.02 g/L MnCl2, 0.02 g/L ZnSO4), operational parameters (temperature, aeration, agitation, initial pH and xylose concentration) and co- substrate supplementation (glucose, fructose, maltose, sucrose, and glycerol) on xylitol biosynthesis by Candida tropicalis ATCC 13803 using synthetic xylose. The significant medium components were identified using the Plackett Burman design followed by central composite designs to obtain the optimal concentration for the critical medium components in shaker flasks. Subsequently, the effect of operational parameters was examined using the One Factor At a Time method, followed by the impact of five co-substrates on xylitol biosynthesis in a 1 L bioreactor. The optimal media components and process parameters are as follows: peptone: 12.68 g/L, yeast extract: 6.62 g/L, salt solution (0.5 g/L MgSO4, 0.5 g/L KH2PO4, and 0.3 g/L CaCl2): 1.23 X (0.62 g/L, 0.62 g/L, and 0.37 g/L respectively), temperature: 30 °C, pH: 6, agitation: 400 rpm, aeration: 1 vvm, and xylose: 50 g/L. Optimization studies resulted in xylitol yield and productivity of 0.71 ± 0.004 g/g and 1.48 ± 0.018 g/L/h, respectively. Glycerol supplementation (2 g/L) further improved xylitol yield (0.83 ± 0.009 g/g) and productivity (1.87 ± 0.020 g/L/h) by 1.66 and 3.12 folds, respectively, higher than the unoptimized conditions thus exhibiting the potential of C. tropicalis ATCC 13803 being used for commercial xylitol production.

Bidirectional fermentation of Monascus and Mulberry leaves enhances GABA and pigment contents: establishment of strategy, studies of bioactivity and mechanistic.

Bidirectional fermentation is a technology that utilizes fungi to ferment medicinal edible substrates, with synergistic and complementary advantages. In this work, a fermentation strategy was established to produce a high yield of γ-aminobutyric acid (GABA) and Monascus pigments (MPs) using Monascus and mulberry leaves (MLs). Firstly, the basic fermentation parameters were determined using single-factor experiments, followed by Plackett-Burman (PB) experimental design to identify MLs, glucose, peptone, and temperature as significant influencing factors. The fermentation parameters were optimized using an artificial neural network (ANN). Finally, the effects of bidirectional fermentation of MLs and Monascus were investigated by bioactivity analysis, microstructure observation, and RT-qPCR. The outcomes showed that the bidirectional fermentation significantly increased the bioactive content and promoted the secondary metabolism of Monascus. The established fermentation conditions were 44.2 g/L of MLs, 57 g/L of glucose, 15 g/L of peptone, 1 g/L of MgSO4, 2 g/L of KH2PO4, 8% (v/v) of inoculum, 180 rpm, initial pH 6, 32 °C and 8 days. The content of GABA reached 13.95 g/L and the color value of MPs reached 408.07 U/mL. This study demonstrated the feasibility of bidirectional fermentation of MLs and Monascus, providing a new idea for the application of MLs and Monascus.

Whey proteins as multifunctional food materials: Recent advancements in hydrolysis, separation, and peptidomimetic approaches.

Whey protein derived bioactives, including α-lactalbumin, ß-lactoglobulin, bovine serum albumin, lactoferrin, transferrin, and proteose-peptones, have exhibited wide ranges of functional, biological and therapeutic properties varying from anticancer, antihypertensive, and antimicrobial effects. In addition, their functional properties involve gelling, emulsifying, and foaming abilities. For these reasons, this review article is framed to understand the relationship existed in between those compound levels and structures with their main functional, biological, and therapeutic properties exhibited either in vitro or in vivo. The impacts of hydrolysis mechanism and separation techniques in enhancing those properties are likewise discussed. Furthermore, special emphasize is given to multifunctional effects of whey derived bioactives and their future trends in ameliorating further food, pharmaceutical, and nutraceutical products. The underlying mechanism effects of those properties are still remained unclear in terms of activity levels, efficacy, and targeted effectiveness. For these reasons, some important models linking to functional properties, thermal properties and cell circumstances are established. Moreover, the coexistence of radical trapping groups, chelating groups, sulfhydryl groups, inhibitory groups, and peptide bonds seemed to be the key elements in triggering those functions and properties. Practical Application: Whey proteins are the byproducts of cheese processing and usually the exploitation of these food waste products has increasingly getting acceptance in many countries, especially European countries. Whey proteins share comparable nutritive values to milk products, particularly on their richness on important proteins that can serve immune protection, structural, and energetic roles. The nutritive profile of whey proteins shows diverse type of bioactive molecules like α-lactalbumin, ß-lactoglobulin, lactoferrin, transferrin, immunoglobulin, and proteose peptones with wide biological importance to the living system, such as in maintaining immunological, neuronal, and signaling roles. The diversification of proteins of whey products prompted scientists to exploit the real mechanisms behind of their biological and therapeutic effects, especially in declining the risk of cancer, tumor, and further complications like diabetes type 2 and hypertension risk effects. For these reasons, profiling these types of proteins using different proteomic and peptidomic approaches helps in determining their biological and therapeutic targets along with their release into gastrointestinal tract conditions and their bioavailabilities into portal circulation, tissue, and organs. The wide applicability of those protein fractions and their derivative bioactive products showed significant impacts in the field of emulsion and double emulsion stabilization by playing roles as emulsifying, surfactant, stabilizing, and foaming agents. Their amphoteric properties helped them to act as excellent encapsulating agents, particularly as vehicle for delivering important vitamins and bioactive compounds. The presence of ferric elements increased their transportation to several metal-ions in the same time increased their scavenging effects to metal-transition and peroxidation of lipids. Their richness with almost essential and nonessential amino acids makes them as selective microbial starters, in addition their richness in sulfhydryl amino acids allowed them to act a cross-linker in conjugating further biomolecules. For instance, conjugating gold-nanoparticles and fluorescent materials in targeting diseases like cancer and tumors in vivo is considered the cutting-edges strategies for these versatile molecules due to their active diffusion across-cell membrane and the presence of specific transporters to these therapeutic molecules.

Transcriptomic profiling reveals differences in the adaptation of two Tetragenococcus halophilus strains to a lupine moromi model medium.

BACKGROUND: Tetragenococcus (T.) halophilus is a common member of the microbial consortia of food fermented under high salt conditions. These comprises salty condiments based on soy or lupine beans, fish sauce, shrimp paste and brined anchovies. Within these fermentations this lactic acid bacterium (LAB) is responsible for the formation of lactic and other short chain acids that contribute to the flavor and lower the pH of the product. In this study, we investigated the transcriptomic profile of the two T. halophilus strains TMW 2.2254 and TMW 2.2256 in a lupine moromi model medium supplied with galactose. To get further insights into which genomic trait is important, we used a setup with two strains. That way we can determine if strain dependent pathways contribute to the overall fitness. These strains differ in the ability to utilize L-arginine, L-aspartate, L-arabinose, D-sorbitol, glycerol, D-lactose or D-melibiose. The lupine moromi model medium is an adapted version of the regular MRS medium supplied with lupine peptone instead of casein peptone and meat extract, to simulate the amino acid availabilities in lupine moromi. RESULTS: The transcriptomic profiles of the T. halophilus strains TMW 2.2254 and TMW 2.2256 in a lupine peptone-based model media supplied with galactose, used as simulation media for a lupine seasoning sauce fermentation, were compared to the determine potentially important traits. Both strains, have a great overlap in their response to the culture conditions but some strain specific features such as the utilization of glycerol, sorbitol and arginine contribute to the overall fitness of the strain TMW 2.2256. Interestingly, although both strains have two non-identical copies of the tagatose-6P pathway and the Leloir pathway increased under the same conditions, TMW 2.2256 prefers the degradation via the tagatose-6P pathway while TMW 2.2254 does not. Furthermore, TMW 2.2256 shows an increase in pathways required for balancing out the intracellular NADH/NADH+ ratios. CONCLUSIONS: Our study reveals for the first time, that both versions of tagatose-6P pathways encoded in both strains are simultaneously active together with the Leloir pathway and contribute to the degradation of galactose. These findings will help to understand the strain dependent features that might be required for a starter strain in lupine moromi.

The effects of sodium sulfite on Helicobacter pylori by establishing a hypoxic environment.

Helicobacter pylori (H. pylori) is an obligate microaerobion and does not survive in low oxygen. Sodium sulfite (SS) reacts and consume oxygen in solutions. The present study aimed to investigate the effects of SS on H. pylori. The effects of SS on oxygen concentrations in solutions and on H. pylori in vivo and in vitro were examined, and the mechanisms involved were explored. The results showed that SS decreased the oxygen concentration in water and artificial gastric juice. In Columbia blood agar and special peptone broth, SS concentration-dependently inhibited the proliferation of H. pylori ATCC43504 and Sydney strain-1 in Columbia blood agar or special peptone broth, and dose-dependently decreased the number of H. pylori in Mongolian gerbils and Kunming mouse infection models. The H. pylori was relapsed in 2 weeks withdrawal and the recurrence in the SS group was lower than that in the positive triple drug group. These effects were superior to positive triple drugs. After SS treatments, the cell membrane and cytoplasm structure of H. pylori were disrupted. SS-induced oxygen-free environment initially blocked aerobic respiration, triggered oxidative stress, disturbed energy production. In conclusion, SS consumes oxygen and creates an oxygen-free environment in which H. pylori does not survive. The present study provides a new strategy and perspective for the clinical treatment of H. pylori infectious disease.

Isolation, identification, and optimization of the fibrinolytic protease-producing strains.

Cardiovascular complications due to thrombosis have become one of the main causes of death worldwide. The high cost and undesirable side effects of existing thrombolytic agents have led researchers to isolate potential strains that produce fibrinolytic enzymes for therapeutic applications. Fibrinolytic enzymes, especially of microbial origin, are recognized as potential therapeutic candidates for thrombosis. In this study, isolation, identification, and optimization of fibrinolytic protease enzyme-producing strains were performed using fermentative protein sources. Fibrinolytic protease-producing strains were selected by analyzing the isolated strains on skim milk agar medium. The selected strains were examined on blood agar and fibrin plate medium, and the ones showing high enzymatic activity were determined. The strain determined to have the highest activity was identified as Acinetobacter johnsonii TR01 by 16S rRNA analysis. The maximum fibrinolytic protease production of the strain occurred at 60 °C and pH 7.0. Under different medium conditions used for enzyme production, fructose was found to be the best carbon source, while yeast extract and peptone were the best nitrogen sources. It was observed that CaCl2, KH2PO4, and MgSO4 components had a negative effect, while MnCl2 and ZnC4H6O4 components had a positive effect on enzyme production. The medium composition for maximum enzyme activity (8.30 IU/ml) determined by Response Surface Methodology was 14.22 g/L fructose, 11.190 g/L yeast extract, 14.22 g/L peptone, 0.5 g/L MnCl2, and 0.5 g/L ZnC4H6O4.

Holtiella tumoricola gen. nov. sp. nov., isolated from a human clinical sample.

The diversity of bacteria associated with biopsy material obtained from patients with colorectal cancer was investigated using culture techniques. A novel bacterium, strain CC70AT, was isolated by diluting a sample of homogenized tissue in anaerobic medium, and then plating to yield a pure culture. Strain CC70AT was a Gram-positive, strictly anaerobic, motile, rod-shaped bacterium. Formate, but not acetate, was a fermentative end-product from growth in peptone-yeast extract and peptone-yeast-glucose broth. The G+C content of DNA from strain CC70AT was 34.9 mol%. 16S rRNA gene sequence analysis revealed that the isolate was part of the phylum Bacillota. The closest described relatives of strain CC70AT were Cellulosilyticum lentocellum (93.3 %) and Cellulosilyticum ruminicola (93.3 and 91.9% sequence similarity across 16S rRNA gene, respectively). According to the data obtained in this work, strain CC70AT represents a novel bacterium belonging to a new genus for which the name Holtiella tumoricola gen. nov., sp. nov. is proposed. The type strain for our described novel species is CC70AT (=DSM 27931T= JCM 30568T).

Modeling and optimization of antibacterial effect of lichen-associated bacteria, Bacillus subtilis KSRLAB3 strain against marine fouling bacteria, Vibrio alginolyticus.

One of the most commonly occurring bacteria, Bacillus subtilis, can produce a wide variety of secondary metabolites. In this study, the antimicrobial effect of B. subtilis KSRLAB3 against Vibrio alginolyticus was optimized using the Plackett-Burman design (PBD) method, response surface methodology (RSM), and genetic algorithm (GA). Initially, the effects of carbon source, nitrogen source, NaCl concentration, pH, temperature, and incubation time on antimicrobial effects were studied. Among the carbon and nitrogen sources investigated, mannose and peptone elicited maximum antimicrobial effect. Using PBD, the most significant variables that influence the antimicrobial effect were identified, including incubation time, peptone concentration, and temperature. The optimum conditions required for attaining maximum antimicrobial effect was identified using the RSM-GA hybrid method, and the optimum condition includes 49.999 h of incubation time, 4.39 g/L of peptone concentration, and 27.629°C of incubation temperature. The confirmatory experiments performed around the optimum condition showed a zone of inhibition of 35 ± 0.52 mm. Methanolic extract also proved the presence of antibacterial lipopeptide surfactin. Therefore, the RSM-GA hybrid method was successfully used in this study to model the antimicrobial effect of B. subtilis KSRLAB3 against V. alginolyticus. The effective inhibition of V. alginolyticus can be investigated further for the development of antifouling coatings.

Development of an animal-free nitrogen source for the liquid surface culture of Cordyceps militaris.

Cordyceps militaris is a medicinal mushroom in Asia in the 21st century, which cordycepin is a significant bioactive compound. This study, investigated the effect of culture conditions and vegetable seed extract powder as a supplementary source of animal-free nitrogen on the production of cordycepin by C. militaris in liquid surface culture. The highest cordycepin production was observed under soybean extract powder (SBEP) conditions, and 80 g L-1 of SBEP supplementation increased cordycepin production to 2.52 g L-1, which was greater than the control (peptone). Quantitative polymerase chain reaction was used to examine the transcription levels, and the results showed that supplementing with SBEP 80 g L-1 significantly increased the expression of genes associated with the carbon metabolic pathway, amino acid metabolism, and two key genes involved in the cordycepin biosynthesis (cns1 and NT5E) compared to peptone-supplemented culture. Under optimal culture conditions, the model predicted a maximum response of cordycepin production of 2.64 g L-1 at a working volume of 147.5 ml, an inoculum size of 8.8% v/v, and a cultivation time of 40.0 days. This optimized culture condition could be used to increase cordycepin production in large-scale bioreactors. Additional research can be conducted to assess the economic viability of this process.

Transfer of Enterococcus faecium and Salmonella enterica during simulated postharvest handling of yellow onions (Allium cepa).

Bacterial transfer during postharvest handling of fresh produce provides a mechanism for spreading pathogens, but risk factors in dry environments are poorly understood. The aim of the study was to investigate factors influencing bacterial transfer between yellow onions (Allium cepa) and polyurethane (PU) or stainless steel (SS) under dry conditions. Rifampin-resistant Enterococcus faecium NRRL B-2354 or a five-strain cocktail of Salmonella was inoculated onto onion skin or PU surfaces at high or moderate levels using peptone, onion extract, or soil water as inoculum carriers. Transfer from inoculated to uninoculated surfaces was conducted using a texture analyzer to control force, time, and number of contacts. Transfer rates (ratio of recipient surface to donor surface populations) of E. faecium (4-5%) were significantly higher than those of Salmonella (0.5-0.6%) at the high (7 log CFU/cm2) but not moderate (5 log CFU/cm2) inoculum levels. Significantly higher populations of E. faecium transferred from onion to PU than from PU to onion. The transfer rates of E. faecium were impacted by inoculum carrier (61% [onion extract], 1.6% [peptone], and 0.31% [soil]) but not by inoculation level or recipient surface (PU versus SS). Bacterial transfer during dry onion handling is significantly dependent on bacterial species, inoculation levels, inoculum carrier, and transfer direction.

Evaluation of the Different Nutritional and Environmental Parameters on Microbial Pyrene Degradation by Mangrove Culturable Bacteria.

Mangrove ecosystems play curial roles in providing many ecological services and alleviating global climate change. However, they are in decline globally, mainly threatened by human activities and global warming, and organic pollutants, especially PAHs, are among the crucial reasons. Microbial remediation is a cost-effective and environmentally friendly way of alleviating PAH contamination. Therefore, understanding the effects of environmental and nutritional parameters on the biodegradation of polycyclic aromatic hydrocarbons (PAHs) is significant for the bioremediation of PAH contamination. In the present study, five bacterial strains, designated as Bp1 (Genus Rhodococcus), Sp8 (Genus Nitratireductor), Sp13 (Genus Marinobacter), Sp23 (Genus Pseudonocardia), and Sp24 (Genus Mycolicibacterium), have been isolated from mangrove sediment and their ring hydroxylating dioxygenase (RHD) genes have been successfully amplified. Afterward, their degradation abilities were comprehensively evaluated under normal cultural (monoculture and co-culture) and different nutritional (tryptone, yeast extract, peptone, glucose, sucrose, and NPK fertilizer) and environmental (cetyl trimethyl ammonium bromide (CTAB), sodium dodecyl sulfate (SDS)) parameters, as well with different co-contaminants (phenanthrene and naphthalene) and heavy metals (Cd2+, Cu2+, Fe3+, Ni2+, Mg2+, Mn2+, and Co2+). The results showed that strain Sp24 had the highest pyrene degradation rate (85%) in the monoculture experiment after being cultured for 15 days. Adding nitrogen- and carbon-rich sources, including tryptone, peptone, and yeast extract, generally endorsed pyrene degradation. In contrast, the effects of carbon sources (glucose and sucrose) on pyrene degradation were distinct for different bacterial strains. Furthermore, the addition of NPK fertilizer, SDS, Tween-80, phenanthrene, and naphthalene enhanced the bacterial abilities of pyrene removal significantly (p < 0.05). Heavy metals significantly reduced all bacterial isolates' degradation potentials (p < 0.05). The bacterial consortia containing high bio-surfactant-producing strains showed substantially higher pyrene degradation. Moreover, the consortia of three and five bacterial strains showed more degradation efficiency than those of two bacterial strains. These results provide helpful microbial resources for mangrove ecological remediation and insight into optimized culture strategies for the microbial degradation of PAHs.

Artificial neural network modeling and statistical optimization of medium components to enhance production of exopolysaccharide by Bacillus sp. EPS003.

Microbial Exopolysaccharides (EPS) have a wide range of applications in food, cosmetics, agriculture, pharmaceutical industries, and environmental bioremediation. The present study aims at enhancing the production of EPS from a soil-isolate Bacillus sp. EPS003. Effects of carbon and nitrogen sources and process conditions were evaluated one factor at a time. Box-Behnken design has been used and a 2.5-fold increase in yield is reported after optimizing the most influential parameters sucrose, yeast extract, and peptone as identified by the Plackett-Burman method. An artificial neural network (ANN) with two different topologies (EPS-NN1 and EPS-NN2) was developed. On comparing prediction accuracy, EPS-NN2 formulated as one input layer with four input variables (sucrose, yeast extract, peptone, biomass), a single hidden layer with seven neurons and EPS yield in the output layer showed a high coefficient of determination (R2-0.98) and low error (NRMSE-0.024). This study concludes that the consideration of biomass value has increased the prediction accuracy of the model.

Study on the use of bovine blood protein hydrolysate as a peptone in microbial culture media.

Livestock blood is a protein-rich waste byproduct produced during meat production processes in slaughterhouses. Its utilization through conversion into value-added products is an intriguing management strategy. In this study, bovine blood was used to obtain the protein hydrolysate for use as a peptone for microbial growth medium. Lyophilized bovine blood was heat treated to make it susceptible to enzymic hydrolysis, and then enzymatically treated with trypsin (bovine pancreas protease) to produce protein hydrolysate. Physico-chemical features were determined for protein hydrolysate and compared to commercial Merck peptone from meat. Amino acid compositions of bovine blood and commercial peptones were subjected to multivariate analysis based on Euclidean similarity matrix using software PAST. Strains of Staphylococcus aureus 25,923, Pseudomonas aeruginosa 27,853, Staphylococcus aureus 6538 P, Enterococcus faecalis 11,700, Escherichia coli 8739, Klebsiella pneumoniae 13,883, Salmonella typhimurium 14,028 and Listeria monocytogenes 13,932 were used as test microbial strains. Growth of bacteria in culture media based on the peptone from bovine protein hydrolysate was compared to that in corresponding reference media based on commercial peptone. The results of these growth tests were comparable. Growth data were depicted and statistically analyzed using R packages ggplot2 and growthcurver, respectively, providing data fitting a standard form of logistic equation.

Modified Drug-Susceptibility Testing and Screening Culture Agar for Colistin-Susceptible Enterobacteriaceae Isolates Harboring a Mobilized Colistin Resistance Gene mcr-9.

Three isolates of the Enterobacter cloacae complex harboring mcr-9, a member of the colistin resistance mcr gene family encoded on plasmids, were susceptible to colistin, with MICs of 0.125 to 0.5 µg/mL in standard broth microdilution (BMD) tests using cation-adjusted Mueller-Hinton broth (CA-MHB) in accordance with European Committee on Antimicrobial Susceptibility Testing guidelines. In contrast, their MICs for colistin were significantly higher (4 to 128 µg/mL) when BMD tests were performed using brain-heart infusion (BHI) medium, Luria-Bertani (LB) broth, tryptic soy broth (TSB), or CA-MHB supplemented with casein, tryptonen or peptone. Colistin significantly induced mcr-9 expression in a dose-dependent manner when these mcr-9-positive isolates were cultured in BHI or CA-MHB supplemented with peptone/casein. Pretreatment of mcr-9-positive isolates and Escherichia coli DH5α harboring mcr-9 with colistin significantly increased their survival rates against LL-37, a human antimicrobial peptide. Electrospray ionization time-of-flight mass spectrometry analysis showed that a lipid A moiety of lipopolysaccharide was partially modified by phosphoethanolamine in E. coli DH5α harboring mcr-9 when treated with colistin. Of 93 clinical isolates of Enterobacteriaceae, only the mcr-9-positive isolates showed MICs to colistin that were at least 32 times higher in BHI than in CA-MHB. These mcr-9-positive isolates grew on a modified BHI agar, MCR9-JU, containing 3 µg/mL colistin. These results suggest that the BMD method using BHI is useful when performed together with the BMD method using CA-MHB to detect mcr-9-positive isolates and that MCR9-JU agar is useful in screening for Enterobacteriaceae isolates harboring mcr-9 and other colistin-resistant isolates.

Production and optimization of bioplastic (Polyhydroxybutyrate) from Bacillus cereus strain SH-02 using response surface methodology.

BACKGROUND: Polyhydroxybutyrate (PHB) is a biopolymer formed by some microbes in response to excess carbon sources or essential nutrient depletion. PHBs are entirely biodegradable into CO2 and H2O under aerobic and anaerobic conditions. It has several applications in various fields such as medicine, pharmacy, agriculture, and food packaging due to its biocompatibility and nontoxicity nature. RESULT: In the present study, PHB-producing bacterium was isolated from the Dirout channel at Assiut Governorate. This isolate was characterized phenotypically and genetically as Bacillus cereus SH-02 (OM992297). According to one-way ANOVA test, the maximum PHB content was observed after 72 h of incubation at 35 °C using glucose and peptone as carbon and nitrogen source. Response surface methodology (RSM) was used to study the interactive effects of glucose concentration, peptone concentration, and pH on PHB production. This result proved that all variables have a significant effect on PHB production either independently or in the interaction with each other. The optimized medium conditions with the constraint to maximize PHB content and concentration were 22.315 g/L glucose, and 15.625 g/L peptone at pH 7.048. The maximum PHB content and concentration were 3100.799 mg/L and 28.799% which was close to the actual value (3051 mg/l and 28.7%). The polymer was identified as PHB using FTIR, NMR, and mass spectrometry. FT-IR analysis showed a strong band at 1724 cm- 1 which attributed to the ester group's carbonyl while NMR analysis has different peaks at 169.15, 67.6, 40.77, and 19.75 ppm that were corresponding to carbonyl, methine, methylene, and methyl resonance. Mass spectroscopy exhibited molecular weight for methyl 3- hydroxybutyric acid. CONCLUSION: PHB-producing strain was identified as Bacillus cereus SH-02 (OM992297). Under optimum conditions from RSM analysis, the maximum PHB content and concentration of this strain can reach (3100.799 mg/L and 28.799%); respectively. FTIR, NMR, and Mass spectrometry were used to confirm the polymer as PHB. Our results demonstrated that optimization using RSM is one of the strategies used for reducing the production cost. RSM can determine the optimal factors to produce the polymer in a better way and in a larger quantity without consuming time.

Optimization and physicochemical characterization of bacterial cellulose by Komagataeibacter nataicola and Komagataeibacter maltaceti strains isolated from grape, thorn apple and apple vinegars.

Bacterial cellulose (BC) is a valuable biopolymer that is increasingly used in medical, pharmaceutical and food industries with its excellent physicochemical properties as high water-holding capacity, nanofibrillar structure, large surface area, porosity, mechanical strength and biocompatibility. Accordingly, the isolation, identification and characterization of potent BC producers from grape, thorn apple and apple vinegars were performed in this study. The strains isolated from grape and apple vinegars were identified as Komagataeibacter maltaceti and the strain isolated from thorn apple vinegar was identified as Komagataeibacter nataicola with 16S rRNA analysis. Optimized conditions were found as 8% dextrin, 1.5% (peptone + yeast extract) and 10% inoculation amount at pH 6.0 with a productivity rate of 1.15 g/d/L, a yield of 8.06% and a dry weight of 6.45 g/L for K. maltaceti, and 10% maltose, 1% (peptone + yeast extract) and 10% inoculation amount at pH 6.0 with a productivity rate of 0.96 g/L/d, a yield of 5.35% and a dry weight of 5.35 g/L for K. nataicola. Obtained BC from K. maltaceti and K. nataicola strains was more than 2.56- and 1.86-fold when compared with BC obtained from HS media and exhibited 95.1% and 92.5% WHC, respectively. Based on the characterization results, BC pellicles show characteristic FT-IR bands and have ultrafine 3D structures with high thermal stability. By means of having ability to assimilate monosaccharides, disaccharides and polysaccharide used in this study, it is predicted that both isolated Komagataeibacter species can be used in the production of biopolymers from wastes containing complex carbon sources in the future.

Degradation and decolourization potential of ligninolytic enzyme producing Bacillus paramycoides BL2 and Micrococcus luteus BL3 for pulp paper industrial effluent and its toxicity evaluation.

Aim of this study was to optimize the production of Ligninolytic enzyme for the degradation of complex pollutants present in pulp paper industrial effluent (PPIE). Two ligninolytic enzyme-producing bacterial strains were isolated from PPIE and identified as Bacillus paramycoides strain BL2 (MZ676667) and Micrococcus luteus strains BL3 (MZ676668). The identified bacterial strain Bacillus paramycoides strain BL2 showed optimum production of LiP (4.30 U/ml), MnP (3.38 U/ml) at 72 h of incubation, while laccase (4.43 U/ml) at 96 h of incubation. While, Micrococcus luteus strains BL3 produced maximum LiP (3.98) and MnP (3.85 U/ml) at 96 h of incubation and maximum laccase (3.85 U/ml) at 72 h of incubation, pH 7-8, and temperatures of 30-35 °C. Furthermore, in the presence of glucose (1.0%) and peptone (0.5%) as nutrient sources, the enzyme activity of consortium leads to reduction of lignin (70%), colour (63%) along with COD (71%) and BOD (58%). The pollutants detected in control i.e. 3.6-Dioxa-2,7-disilaoctane, 2-Heptnoic acid,trimethylsilyl ester, 7-Methyldinaphtho [2,1-b,1',2'-d] silole, Hexadeconoic acid, trimethylysilyl ester, Methyl1(Z)-3,3-dipheny.1-4-hexenoale, 2,6,10,14,18,22-Tetracosahexane,2,2-dimethylpropyl(2Z,6E)-10,11epoxy5,6 Dihyrostigmasterol, acetate were completely diminished. The toxicity of PPIE was reduced up to 75%. Hence, knowledge of this study will be very useful for industrial sector for treatment of complex wastewater.

Granulimonas faecalis gen. nov., sp. nov., and Leptogranulimonas caecicola gen. nov., sp. nov., novel lactate-producing Atopobiaceae bacteria isolated from mouse intestines, and an emended description of the family Atopobiaceae.

Two strictly anaerobic, Gram-stain-positive, non-motile bacteria (strains OPF53T and TOC12T) were isolated from mouse intestines. Strains OPF53T and TOC12T grew at pH 5.5-9.0 and 5.0-9.0, respectively, and at temperatures of 30-45 °C. The cell morphologies of these strains were short rods and rods, respectively, and the cells possessed intracellular granules. The major cellular fatty acids of OPF53T were C18  :  1 cis 9 and C18  :  1 cis 9 dimethyl acetal, whereas those of TOC12T were C18  :  0 and C18  :  1 cis 9. In OPF53T, the main end-products of modified peptone-yeast extract-glucose (PYG) fermentation were lactate, formate and butyrate, whereas, in addition to these acids, TOC12T also produced hydrogen. The genomes of OPF53T and TOC12T were respectively 2.2 and 2.0 Mbp in size with a DNA G+C contents of 69.1 and 58.7 %. The 16S rRNA gene sequences of OPF53T and TOC12T showed the highest similarity to members of the family Atopobiaceae, namely, Olsenella phocaeensis Marseille-P2936T (94.3 %) and Olsenella umbonata KCTC 15140T (93.2 %), respectively. Phylogenetic analyses revealed that both isolates formed distinct lineages from other genera of the family Atopobiaceae. In addition, the two strains were characterized by relatively low 16S rRNA gene sequence similarity (93.4 %) and can be distinguished by their distinctive traits (including cell shape, DNA G+C content, and major fatty acids profiles). On the basis of their polyphasic taxonomic properties, these isolates represent two noel species of two novel genera within the family Atopobiaceae, for which the names Granulimonas faecalis gen. nov., sp. nov. (OPF53T=JCM 35015T=KCTC 25474T) and Leptogranulimonas caecicola gen. nov., sp. nov. (TOC12T=JCM 35017T=KCTC 25472T) are proposed.