Bioprocess development for extraction and purification of cellulases from Aspergillus niger 3ASZ using statistical experimental design techniques.

The amount of cellulosic materials is large and may lead to environmental pollution, so they can be converted into useful materials for use in food or energy. Statistical design (Plackett-Burman and Box-Behnken) was the main topic of this study and was used to optimize the effect of environmental factors on cellulase production by Aspergillus niger. Cellulase production using Plackett-Burman was 6.86-fold higher than the production of cellulase using the basal medium. B0X-Benken showed an increase in the cellulase production equal to 18 times compared to the basal medium, where the cellulase produced had an activity equal to 79.4 U/mL/min. Ammonium sulfate precipitation was applied to the crude enzyme, followed by sequential fractionation with an Amicon system. The Amicon was used to demonstrate the final volume, total enzyme activity, specific activity, purification fold, and yield of cellulase (partially purified enzyme). Numerous cellulolytic enzymes are abundant in Aspergillus species. All of the data showed that Aspergillus sp. might be a reliable source of industrially and economically useful cellulases. By statistically calculating the relevance of a large number of elements in one experiment using a multifactorial statistical design, time may be saved while still maintaining the validity of each component.

2,4-Dichlorophenol biotransformation using immobilized marine halophilic Bacillus subtilis culture and laccase enzyme: application in wastewater treatment.

BACKGROUND: 2,4-Dichlorophenol (2,4-DCP) is a very toxic aromatic compound for humans and the environment and is highly resistant to degradation. Therefore, it is necessary to develop efficient remediation and cost-effective approaches to this pollutant. Microbial enzymes such as laccases can degrade phenols, but limited information is known about immobilized bacterial laccase and their reuse. METHODS: Immobilization of marine halophilic Bacillus subtilis AAK cultures via entrapment and adsorption techniques and degradation of different phenolic compounds by immobilized cells were estimated. Partial purification and immobilization of laccase enzymes were carried out. In addition, the biodegradation of 2,4-DCP and others contaminated by wastewater was investigated. RESULTS: Immobilization of cells and partially purified laccase enzymes by adsorption into 3% alginate increased 2,4-DCP biotransformation compared with free cells and free enzymes. In addition, the reuse of both the immobilized culture and laccase enzymes was evaluated. The highest removal of 2,4-DCP from pulp and paper wastewater samples inoculated by immobilized cells and the immobilized enzyme was 90% and 95%, respectively, at 50 h and 52 h of incubation, compared to free cells and free enzyme. CONCLUSION: The results of this study have revealed the immobilization of a biocatalyst and its laccase enzyme as a promising technique for enhancing the degradation of 2,4-DCP and other toxic phenolic and aromatic compounds. The reuse of the biocatalyst and its laccase enzyme enabled the application of this cost-effective bioremediation strategy.

Lead adsorption and antibacterial activity using modified magnetic biochar/sodium alginate nanocomposite.

Biochar is one of the most promising wastewater treatment materials. As shown in the Scanning Electron Micrograph, the magnetic biochar (BC) cross-linked glutaraldehyde (G) with sodium alginate (SA) (BC-G-SA) nanocomposite formed with uniform particle size without aggregation, and an X-Ray Diffraction study revealed that the BC-G-SA nanocomposite has an amorphous structure. The BC-G-SA nanocomposite enhanced the microwave adsorption process for Pb (II). The maximum metal capacity value was obtained using the microwave adsorption technique at pH 5.0 and contact time 20 s for Pb (II) at medium and low microwave power (940 and 1400 µmol g-1, respectively). Pb (II) adsorption isotherm follows a pseudo-second-order model. Also, the BC-G-SA nanocomposite effectively inhibited bacterial growth throughout the growth kinetics experiment. BC-G-SA inhibited the growth of S. aureus at a MIC of 200 g mL-1, whereas L. monocytogenes had a MIC of 200 g mL-1. The MIC values for E. faecalis and E. faecium were significantly lower (50 and 100 g mL-1, respectively).

Incidence of virulence determinants and antibiotic resistance in lactic acid bacteria isolated from food products.

Background: Lactic acid bacteria (LAB) confer beneficial health effects in humans. However, the safety of these bacteria and their potential to spread resistance in the environment must be evaluated. Materials & methods: Fifty-three LAB were isolated from different food samples and assessed for the prevalence of virulence determinants and antibiotic resistance profile. Results: Multiple resistance was reported for Lactobacillus brevis MIM04, having revealed phenotypic resistance to vancomycin (MIC >128 µg/ml), ampicillin, cefotaxime, oxacillin and gentamicin. Virulence traits (cylA, gelE, esp and agg) were detected using specific primers. Enterococcus faecium CHE32, Lactobacillus plantarum CHE37 and E. faecium MLK68 lack virulence genes, possess antimicrobial activity and survive in low pH and bile salt conditions. Conclusion: Isolated LAB revealed probiotic properties.

Novel Siwa propolis and colistin-integrated chitosan nanoparticles: elaboration; in vitro and in vivo appraisal.

Aim: The present study aimed to formulate novel cremophore-decorated chitosan nanoparticles of colistin, integrated with Siwa propolis extract, to solve bacterial resistance to colistin. Materials & methods: The novel nanoformula was prepared using an incorporation method. Physicochemical assessment and in vivo studies of the selected nanoformulations were performed. Results: The nanoformulation exhibited a nanosize of 48.3 nm, high ζ potential (43.6 mV), high entrapment efficiency (75%) and complete bacterial growth eradication within 2 h (minimum inhibitory concentration = 6.25 µg/ml). Histological examination showed that incorporation of colistin into the nanoformulation could successfully prevent its nephrotoxicity. Conclusion: Tailoring of proper nanocarrier could successfully revert bacteria from being colistin-resistant to colistin-sensitive. The developed nanoformulation can be considered as a potential antibacterial agent in pneumonia treatment.

Assessment of the heavy metal bioremediation efficiency of the novel marine lactic acid bacterium, Lactobacillus plantarum MF042018.

Heavy metal pollution is one of the most serious environmental and human health risk problem associated with industrial progress. The present study was conducted with the goal of isolation and characterization of metal-resistant lactic acid bacteria (LAB) from the Alexandrian Mediterranean Seacoast, Egypt, with their possible exploitation in metal remediation. Lactobacillus plantarum MF042018 exhibited high degree of resistance, up to 500 and 100 ppm, to both nickel and chromium, respectively, with multiple antibiotic resistance (MAR) index above 0.5. In an attempt to improve chromium removal by L. plantarum MF042018, Plackett-Burman followed by Box-Behnken statistical designs were applied. An initial Cr2+ concentration of 100 ppm and inoculum size of 3% presented the best conditions for the accumulation of chromium by L. plantarum MF042018. The study was also navigated to assess the biosorption capacity of L. plantarum MF042018, the maximum uptake capacity (q) of both Cd2+ and Pb2+ was recorded at pH 2.0 and a temperature of 22 °C after 1 hr. The biosorption process of Cd2+ and Pb2+ was well explained by the Langmuir isotherm model better than the Freundlich isotherm. Furthermore, the results revealed that the use of L. plantarum MF042018 is an effective tool for the treatment of hazardous metal-polluted battery-manufacturing effluent. Therefore, the present study implies that L. plantarum MF042018 can be applied as a promising biosorbent for the removal of heavy metals from industrial wasterwaters.

Draft Genome Sequence of an Enterococcus faecalis Strain (24FS) That Was Isolated from Healthy Infant Feces and Exhibits High Antibacterial Activity, Multiple-Antibiotic Resistance, and Multiple Virulence Factors.

Enterococcus faecalis 24FS is a bacteriocin-producing, multiply antibiotic-resistant, and potentially virulent bacterium isolated from healthy infant feces. The draft 2.9-Mb genome sequence revealed 2,968 protein-encoding genes; 11 antibiotic resistance, 8 virulence, and 3 bacteriocin genes; and 2 plasmids, 4 prophages, 30 insertion sequence (IS) elements, 1 transposon, and 1 integron.

Statistically optimized ceftriaxone sodium biotransformation through Achromobacter xylosoxidans strain Cef6: an unusual insight for bioremediation.

The present study underlines a unique promising approach toward efficient biotransformation of ceftriaxone sodium (Ceftx), a highly frequent prescribed cephalosporin antibiotic, by a newly bacterium namely Achromobacter xylosoxidans strain Cef6 isolated from Ceftx contaminated raw materials in pharmaceutical industries. A three step sequential statistical-mathematical approach (Plackett-Burman design [PBD], Central Composite Design [CCD], and ridge-canonical analyses) was anticipated to optimize the biotransformation process. Ceftx concentration and medium volume: bottle volume ratio, two key determinants, significantly (p < 0.05) affected the process outcome deduced by regression analysis of PBD' data. CCD and ridge-canonical analyses localized the optimal levels of Ceftx concentration and medium volume: 250 ml bottle volume ratio to be 0.39 and 7.973 g Ceftx/L modified tryptic soy broth achieving Ceftx biotransformation (100%) after 39 h under aerobic static conditions at 30 °C, irrespectively deduced via HPLC analysis. Impressively, only one of five Ceftx byproducts was detected by the end of the biotransformation process. To the best of authors' knowledge, this is the first report addressing a detailed study regarding efficient biotransformation of Ceftx by single bacterium not bacterial consortium under aerobic conditions. Present data would greatly encourage applying this approach for decontamination of some Ceftx contaminated environmental sites.

Complete Genome Sequence of Lactobacillus plantarum 10CH, a Potential Probiotic Lactic Acid Bacterium with Potent Antimicrobial Activity.

Lactobacillus plantarum 10CH is a bacteriocin-producing potential probiotic lactic acid bacterium (LAB) strain isolated from cheese. Its complete nucleotide sequence shows a single circular chromosome of 3.3 Mb, with a G+C content of 44.51%, a 25-gene plantaricin bacteriocin gene cluster, and the absence of recognized virulence factors.

Hydrogel Dressing with a Nano-Formula against Methicillin-Resistant Staphylococcus aureus and Pseudomonas aeruginosa Diabetic Foot Bacteria.

This study proposes an alternative approach for the use of chitosan silver-based dressing for the control of foot infection with multidrug-resistant bacteria. Sixty-five bacterial isolates were isolated from 40 diabetic patients. Staphylococcus aureus (37%) and Pseudomonas aeruginosa (18.5%) were the predominant isolates in the ulcer samples. Ten antibiotics were in vitro tested against diabetic foot clinical bacterial isolates. The most resistant S. aureus and P. aeruginosa isolates were then selected for further study. Three chitosan sources were tested individually for chelating silver nanoparticles. Squilla chitosan silver nanoparticles (Sq. Cs-Ag(0)) showed the maximum activity against the resistant bacteria when mixed with amikacin that showed the maximum synergetic index. This, in turn, resulted in the reduction of the amikacin MIC value by 95%. For evaluation of the effectiveness of the prepared dressing using Artemia salina as the toxicity biomarker, the LC50 was found to be 549.5, 18,000, and 10,000 µg/ml for amikacin, Sq. Cs-Ag(0), and dressing matrix, respectively. Loading the formula onto chitosan hydrogel dressing showed promising antibacterial activities, with responsive healing properties for the wounds in normal rats of those diabetic rats (polymicrobial infection). It is quite interesting to note that no emergence of any side effect on either kidney or liver biomedical functions was noticed.

In vitro antagonistic activity, plant growth promoting traits and phylogenetic affiliation of rhizobacteria associated with wild plants grown in arid soil.

The role of plant growth-promoting rhizobacteria (PGPR) in adaptation of plants in extreme environments is not yet completely understood. For this study native bacteria were isolated from rhizospeheric arid soils and evaluated for both growth-promoting abilities and antagonistic potential against phytopathogenic fungi and nematodes. The phylogentic affiliation of these representative isolates was also characterized. Rhizobacteria associated with 11 wild plant species from the arid soil of Almadinah Almunawarah, Kingdom of Saudi Arabia (KSA) were investigated. From a total of 531 isolates, only 66 bacterial isolates were selected based on their ability to inhibit Fusarium oxysporum, and Sclerotinia sclerotiorum. The selected isolates were screened in vitro for activities related to plant nutrition and plant growth regulation as well as for antifungal and nematicidal traits. Isolated bacteria were found to exhibit capabilities in fix atmospheric nitrogen, produce ammonia, indoleacetic acid (IAA), siderophores, solubilize phosphate and zinc, and showed an antagonistic potential against some phytopathogenic fungi and one nematode species (Meloidogyne incognita) to various extent. Isolates were ranked by their potential ability to function as PGPR. The 66 isolates were genotyped using amplified rDNA restriction analysis (ARDRA) and 16S rRNA gene sequence analysis. The taxonomic composition of the representative genotypes from both rhizosphere and rhizoplane comprised Bacillus, Enterobacter and Pseudomonas. Out of the 10 genotypes, three strains designated as PHP03, CCP05, and TAP02 might be regarded as novel strains based on their low similarity percentages and high bootstrap values. The present study clearly identified specific traits in the isolated rhizobacteria, which make them good candidates as PGPR and might contribute to plant adaption to arid environments. Application of such results in agricultural fields may improve and enhance plant growth in arid soils.

Synthesis, characterization and the antimicrobial activity of new eco-friendly ionic liquids.

A green microwave-assisted procedure for the preparation of a series of 24 new 1-alkyl-3-ethylimidazolium ionic liquids with different functional groups in the alkyl chain is described. Moreover, the synthesis of a variety of ten new geminal dicationic ionic liquids is reported. Their structures were characterized by FT-IR, (1)H NMR, (13)C NMR, (11)B, (19)F, (31)P, and mass spectrometry. Several ionic liquids were selected for antimicrobial activity studies, yielding very interesting and promising results.

Solid-state fermentation for the production of meroparamycin by streptomyces sp. strain MAR01.

The antibiotic meroparamycin was produced in the free culture system of Streptomyces sp. strain MAR01. Five solid substrates (rice, wheat bran, Quaker, bread, and ground corn) were screened for their ability to support meroparamycin production in solid-state fermentation. In batch culture, wheat bran recorded the highest antibacterial activity with the lowest residual substrate values. The highest residual substrate values were recorded for both ground corn and Quaker. On the other hand, no antibacterial activity was detected for rice as a solid substrate. The use of the original strength of starch-nitrate medium in the solid-state fermentation gave a lower antibacterial activity compared with the free culture system. Doubling the strength of this medium resulted in the increase in the activity to be equivalent to the free culture. The initial pH (7.0) of the culture medium and 2 ml of spore suspension (1 ml contains 5x10(9) spores/ml) were the optima for antibiotic production. The water was the best eluent for the extraction of the antibiotic from the solid-state culture. Ten min was enough time to extract the antibiotic using a mixer, whereas, 60 min was required when shaking was applied. Semicontinuous production of meroparamycin using a percolation method demonstrated a more or less constant antibacterial activity over 4 runs (450-480 microg/ml). The semicontinuous production of the antibiotic was monitored in a fixed-bed bioreactor and the maximum activity was attained after the fourth run (510 microg/ml) and the overall process continued for 85 days.

Kosinostatin, a major secondary metabolite isolated from the culture filtrate of Streptomyces violaceusniger strain HAL64.

During a screening program, an actinomycete strain isolated from the Egyptian soil was investigated for its potential to show antimicrobial activity. The identification of this isolate was performed according to spore morphology and cell wall chemo-type, which suggested that this strain is a streptomycete. Further cultural, physiological characteristics and the analysis of the nucleotide sequence of the 16S rRNA gene (1480 bp) of this isolate indicated that this strain is identical to Streptomyces violaceusniger (accession number EF063682) and then designated S. violaceusniger strain HAL64. In its culture supernatant, this organism could produce one major compound strongly inhibits the growth of Gram-positive but the inhibition of Gram-negative indicator bacteria was lower. The antibiotic was separated by silica gel column chromatography and then purified on a sephadex LH-20 column and finally the purity was checked by HPLC. The chemical structure of the purified compound was determined using spectroscopic analyses (molecular formula of C33H32N2O10 and molecular weight of 617.21) and found to be identical to the kosinostatin, a quinocycline antibiotic which is known to be produced by Micromonspora sp. TP-A0468 (Igarashi et al., 2002) and to quinocycline B isolated from Streptomyces aureofaciens (Celmer et al., 1958). Although the antibiotic is known, the newly isolated strain was able to produce the antibiotic as a major product providing an important biotechnological downstream advantage.

Meroparamycin production by newly isolated Streptomyces sp. strain MAR01: taxonomy, fermentation, purification and structural elucidation.

Twelve actinomycete strains were isolated from Egyptian soil. The isolated actinomycete strains were then screened with regard to their potential to generate antibiotics. The most potent of the producer strains was selected and identified. The cultural and physiological characteristics of the strain identified the strain as a member of the genus Streptomyces. The nucleotide sequence of the 16S rRNA gene (1.5 kb) of the most potent strain evidenced a 99% similarity with Streptomyces spp. and S. aureofaciens 16S rRNA genes, and the isolated strain was ultimately identified as Streptomyces sp. MAR01. The extraction of the fermentation broth of this strain resulted in the isolation of one major compound, which was active in vitro against gram-positive, gram-negative representatives and Candida albicans. The chemical structure of this bioactive compound was elucidated based on the spectroscopic data obtained from the application of MS, IR, UV, 1H NMR, 13C NMR, and elemental analysis techniques. Via comparison to the reference data in the relevant literature and in the database search, this antibiotic, which had a molecular formula of C19H29NO2 and a molecular weight of 303.44, was determined to differ from those produced by this genus as well as the available known antibiotics. Therefore, this antibiotic was designated Meroparamycin.

Evaluation of in vitro antibacterial activity of some disinfectants on Escherichia coli serotypes.

Three disinfectants commonly used in poultry farms (formalin, TH4+, and Virkon-S) were chosen for the present study. The effect of disinfectant concentration and the duration of exposure to these disinfectants on the survival of Escherichia coli serotypes (O114:K-, O86, O55:K39, and O86:K60) were investigated. Formalin (0.6%), TH4+ (0.06%), and Virkon (0.5%) all killed the four serotypes within 5 min of exposure. As the disinfectant concentration decreases, the length of exposure time to kill serotype increases. At 0.03%, 0.007%, and 0.03% of formalin, TH4+ and Virkon-S concentrations failed to kill the four E. coli serotypes within 360 min, respectively. An improvement of the inhibitory effect of these disinfectants occurred when added together with the inoculum instead of an established population. The influence of formalin, TH4+, and Virkon-S on the cell morphology of E. coli O55:K39 was investigated by using transmission electron microscopy. Formalin-treated cells exhibited normal cell morphology, with the exception that the treated cell was less fimbriated, and more destruction of pili increased when formalin concentrations were doubled. Cells treated with TH4+ (0.03%) showed destruction of the cell wall and cell surface membrane after 5 min. Cell filamentation occurred at 0.015% and increased with the increase of exposure time to this drug. Spheroplasts were observed only when cells were treated with 0.125% Virkon-S for 60 min, and cell lysis started to occur when 0.25% Virkon-S was applied for 15 min. Scanning electron microscope study revealed that Virkon-S at 0.03% and TH4+ at 0.007% completely prevented the adherence of E. coli O55:K39 serotype to chicken tracheal organ, whereas formalin (0.03%) disinfection minimized the adherence of E. coli cells to tracheal explants after 360 min of incubation.