Molecular basis for the increased activity of ZMS-2 serine protease in the presence of metal ions and hydrogen peroxide.

Serine proteases are important enzymes widely used in commercial products and industry. Recently, we identified a new serine protease from the desert bacterium Bacillus subtilis ZMS-2 that showed enhanced activity in the presence of Zn2+, Ag+, or H2O2. However, the molecular basis underlying this interesting property is unknown. Here, we report comparative studies between the ZMS-2 protease and its homolog, subtilisin E (SubE), from B. subtilis ATCC 6051. In the absence of Zn2+, Ag+, or H2O2, both enzymes showed the same level of proteolytic activity, but in the presence of Zn2+, Ag+, or H2O2, ZMS-2 displayed increased activity by 22%, 8%, and 14%, whereas SubE showed decreased activity by 16%, 12%, and 9%, respectively. In silico studies showed that both proteins have almost identical amino acid sequences and folding structures, except for two amino acids located in the protruding loops of the proteins. ZMS-2 contains Ser236 and Ser268, whereas SubE contains Thr236 and Thr268. Replacing Ser236 or Ser268 in ZMS-2 with threonine resulted in variants whose activities were not enhanced by Zn2+ or Ag+. However, this single mutation did not affect the enhancement by H2O2. This finding may be used as a basis for engineering better proteases for industrial uses.

Characterization and Pilot-Scale Application of the ZMS-2 Serine Protease with Novel Properties for the Eco-friendly Leather Processing.

Microbial alkaline proteases are dominating the global enzyme market with a share of over 65% due to their multifarious catalytic potentials. Hence, production of proteases with novel properties of commercial significance is highly desirable to meet the global enzyme demand. Here, we report the purification, characterization, and pilot-scale application of a serine protease from the desert soil bacterium Bacillus subtilis ZMS-2 with novel properties as dehairing agent in leather processing. The enzyme was purified 16.5-fold with a specific activity of 1543.5 U mg-1 and recovery percentage of 33.6% using ammonium sulfate precipitation, ion exchange, and gel filtration chromatography. The purified enzyme was characterized as a metal ion-, surfactant-, and denaturant-compatible alkaline serine protease having a molecular weight of 36.1 kDa with an optimum activity at pH 8.5 and 60 °C. The catalytic activity of the enzyme was enhanced by Zn+2 (204%), Ag+ (110%), H2O2 (123%), Triton X-100 (110%), iso-octane (109%), chloroform (110%), ethanol (105%), ethyl acetate (110%), and acetonitrile (128%). During pilot-scale applications, the optimum condition was found to be a combination of enzyme (1.5%, 460 U mL-1), sodium sulfide (2%), and calcium hydroxide (lime) (3%). Under this condition, the time required for complete dehairing was 90 min. Chemoenzymatically processed skins exhibited better physical properties than chemically processed skin, including tensile strength (16.35 ± 6.68 N/mm), ball burst (452.88 ± 6.06 N/mm), percent elongation (38.85 ± 1.06 N), tear strength (50.16 ± 4.42 N/mm), and softness (6.5 mm). Electron microscopy analysis of the treated skin showed complete removal of hairs with roots, confirming the keratin specificity of the enzyme. Moreover, the enzyme-assisted dehairing process reduced chemical oxygen demand (COD), biochemical oxygen demand (BOD), total dissolved solids (TDS), and total suspended solids (TSS) by 68, 77, 34, and 39%, respectively. Thus, the alkaline serine protease from B. subtilis ZMS-2 is a potential dehairing agent for the eco-friendly processing of animal skins on industrial scales.

In-vitro evaluation of virulence markers and antifungal resistance of clinical Candida albicans strains isolated from Karachi, Pakistan.

Candidiasis is a significant fungal infection with high mortality and morbidity rates worldwide. Candida albicans is the most dominant species responsible for causing different manifestations of candidiasis. Certain virulence traits as well as its resistance to antifungal drugs contribute to the pathogenesis of this yeast. This study was designed to determine the production of some virulence factors, such as biofilm formation and extracellular hydrolytic enzymes (esterase, coagulase, gelatinase, and catalase) by this fungus, as well as its antifungal resistance profile. A total of 304 clinical C. albicans isolates obtained from different clinical specimens were identified by a conventional diagnostic protocol. The antifungal susceptibility of C. albicans strains was determined by disk diffusion technique against commercially available antifungal disks, such as nystatin 50 µg, amphotericin B 100 unit, fluconazole 25 µg, itraconazole 10 µg, ketoconazole 10 µg, and voriconazole 1 µg. The assessment of biofilm formation was determined by the tube staining assay and spectrophotometry. Gelatinase, coagulase, catalase, and esterase enzyme production was also detected using standard techniques. A total of 66.1% (201/304) and 28.9% (88/304) of C. albicans strains were susceptible-dose dependent (SDD) to nystatin and itraconazole, respectively. Among the antifungal drugs, C. albicans strains showed high resistance to ketoconazole 24.7% (75/304); however, no statistically significant relationship between the clinical origin of C. albicans isolates and antifungal drug resistance pattern was detected. For virulence factors, the majority of the C. albicans strains actively produced biofilm and all hydrolytic enzymes. Biofilm formation was demonstrated by 88% (267/304) of the strains with a quantitative mean value 0.1762 (SD ± 0.08293). However, 100% (304/304) of isolates produced catalase enzyme, 69% (211/304) produced coagulase, 66% (197/304) produced gelatinase, and 52% (157/304) produced esterase enzyme. A significant relationship between the source of specimens and biofilm formation by C. albicans was observed; nevertheless, there was no significant relationship between different sources of C. albicans strains and the production of different enzymatic virulence factors. The study found that C. albicans strains have excellent potential to produce virulence markers and resistance to antifungals, which necessitates surveillance of these opportunistic pathogens to minimize the chances of severe invasive infections.

Biological pretreatment of sugarcane bagasse for the production of fungal laccase and bacterial cellulase.

Sugarcane bagasse (SB) is a promising source of appreciable quantities of fermentable sugars. However, the presence of lignin hinders utilization of these carbohydrates and hence pretreatment to remove lignin is necessarily carried out. Here, a biological pretreatment method was synchronized with the production of a thermostable cellulase using SB as a raw material. Initially, bagasse was fermented by a laccase producing fungus, Trametes pubescens MB 89 under solid state fermentation (SSF) and a titer of 1758 IU mL-1 of laccase was obtained. Investigations of nine factors affecting laccase production through Plackett Burman design improved the titers to 6539 IU mL-1 . Five factors (incubation period, concentration of CuSO4 , temperature, moisture content, and particle size) were found significant which were optimized through Central Composite design leading to an improvement in the titers by ~5 folds (8841 IU mL-1 ). Biologically pretreated SB was fermented by a thermophilic bacterium, Neobacillus sedimentimangrovi UE25, that yielded 8.64 IU mL-1 of cellulase. Delignification and cellulose utilization were affirmed by structural analysis through FTIR and SEM. The synchronized process yielded higher titers of laccase and cellulase under SSF of SB with the minimum use of corrosive chemicals.

Aspergillus violaceofuscus alleviates cadmium and chromium stress in Okra through biochemical modulation.

Endophytic fungi from the Chilli were used to help okra plants exposed to cadmium (Cd) or chromium (Cr) stress. Initially, the strain Ch06 produced higher amounts of indole acetic acid (IAA) (230.5 µg/mL), sugar (130.7 µg/mL), proteins (128.2 µg/mL), phenolics (525.6 µg/mL) and flavonoids (98.4 µg/mL) in Czapek broth supplemented with Cd or Cr. The production of IAA and other metabolites in such a higher concentration suggested that Ch06 might improve plant growth under heavy metal stress. For this reason, an experiment was designed, in which biomass of Ch06 (at 2g/100g of sand) were applied to the okra plants exposed to Cd or Cr stress (at 100 or 500 µg/g). The results exhibited that Ch06 improved the total chlorophyll (36.4±0.2 SPAD), shoot length (22.6±0.2 cm), root length (9.1±0.6 cm), fresh weight (5±0.6 g), dry weight (1.25±0.01 g), sugars (151.6 µg/g), proteins (114.8 µg/g), proline (6.7 µg/g), flavonoids (37.9 µg/g), phenolics (70.7 µg/g), IAA (106.7 µg/g), catalase (0.75 enzyme units/g tissue) and ascorbic acid oxidaze (2.2 enzyme units/g tissue) of the associated okra plants. Similar observations have been recorded in Ch06 associated okra plants under Cd and Cr stress. Also, Ch06 association reduced translocation of Cd (35% and 45%) and Cr (47% and 53%) to the upper parts of the okra plants and thus reduced their toxicity. The internal transcribed spacer (ITS) region amplification of 18S rDNA (ribosomal deoxyribo nucleic acid) exhibited that the potent strain Ch06 was Aspergillus violaceofuscus. The results implied that A. violaceofuscus has the ability to promote host species growth exposed to Cd and Cr. Moreover, it helped the host plants to recover in Cd and Cr polluted soils, hence can be used as biofertilizer.

Porostereum spadiceum-AGH786 Regulates the Growth and Metabolites Production in Triticum aestivum L. Under Salt Stress.

The role of the most fungal endophytes in the host plant growth and production of metabolites under stress conditions is still unknown. Fungal endophytes occur in almost all plants to benefit the host plants exposed to biotic and abiotic stress. In the present work, we investigated salt (NaCl) stress alleviation capability of a fungal endophyte (Porostereum spadiceum-AGH786). The culture filtrate (CF: 1.5 mL.) of P. spadiceum-AGH786 contained IAA (158 µg/ml), SA (29.3 µg/ml), proline (114.6 µg/ml), phenols (167.4 µg/ml), lipids (71.4 µg/ml), sugar (133.2 µg/ml), flavonoids (105.04 µg/ml). Smaller amounts of organic acids, such as butyric acid (5.8 µg/ml), formic acid (2.34 µg/ml), succinic acid (2.02 µg/ml), and quinic acid (2.25 µg/ml) were also found in CF of P. spadiceum-AGH786. Similarly, the CF displayed antioxidant activity in 2,2-Diphenyl-1-picrylhydrazyl (DPPH) and 2,2'-Azino-bis-3-ethylbenzothiazoline-6-sulfonic acid (ABTS) assays. Moreover, wheat plants colonized by P. spadiceum-AGH786 showed significantly (P = 0.05) higher polyphenol oxidases activity (2.2 mg/g DW) under normal conditions as compared to the NaCl-treated plants. We also observed that P. spadiceum-AGH786 improved biomass (0.30 g) of wheat plants subjected to 140 mM NaCl stress. The results conclude that the wheat plant colonization by P. spadiceum-AGH786 greatly improved the plant growth under 70 mM and 140 mM NaCl stress. Thus, the biomass of the P. Spadiceum-AGH786 can be used in saline soil to help the host plants.

Characterization of the genome and serine protease of a novel Bacillus subtilis isolate.

Current study was undertaken to carry out the genome-wide analysis of a multipotent isolate from desert soil which was previously identified as Bacillus tequilensis based on 16S rDNA analysis. This study also aims to characterize the serine protease and its biocatalytic potentials implying a combination of empirical and in-silico approaches. Next generation sequencing and short read de novo assembly generated the 4,235,084 bp draft genome of Bacillus sp. ZMS-2. Genome sequence analysis by digital DNA:DNA hybridization (dDDH) and average nucleotide identity classified the isolate as Bacillus subtilis ZMS-2 (Bioproject ID: PRJNA691551). Genome annotation revealed 10 antibiotic resistance genes, 8 antibiotic/antifungal gene clusters and 25 genes encoding proteases including subtilisin E, an extracellular alkaline protease. This extracellular protease (ZMS-2 protease) was produced using a statistically optimized medium, purified partially and characterized as alkaline serine protease. The partially purified ZMS-2 protease (780 U/mL) showed a 21 mm zone of casein hydrolysis and dehaired goat skin by pulling out hair with roots. These catalytic potentials of ZMS-2 protease were further confirmed using scanning electron microscopy of casein beads and dehaired skin. The study concludes B. subtilis ZMS-2 as a potent producer of a protease with promising potentials of commercial importance.

In vivo hepatoprotective and in vitro antimicrobial potential of Ceasalpinia bonduc (Linn): Pharmacological correlation with identified phytochemicals.

The in vivo hepatoprotective potential of methanolic extract of Ceasalpinia bonduc (CBLM) has been explored against carbon tetrachloride (CCl4) induced acute liver injury in rats. Treatment of plant extract on CCl4 intoxicated liver significantly reduced the hepatoxicity, along with serum enzymes GPT and GOT. To explore the chemical constituents from CBLM extract, it was fractionated into non-polar to moderately polar fractions (CBLM-H, CBLM-HEt, CBLM-Et, CBLM-EtM, CBLM-M) and subjected to GC/GC-MS analysis. Altogether twenty seven (~71%) phytochemicals were identified from different fractions by using Electronic Mass Spectral Library GC-MS (NIST 20). Out of which twenty one are first time reported from Ceasalpinia bonduc, fourteen from genus Caesalpinia and ten from family Fabaceae. The identified phytochemicals 2-ethyl-2-hydroxy-1,3-dimethylcyclopentanecarboxylic acid, ethyl ester (21) and 1,3,5-triazine-2,4-diamine,6-hydroxy-N,N-dicyclohexyl (23) are first time identified as plant metabolites. To explore the antimicrobial potential four strains of Gram-positive and eight strains of Gram-negative bacteria were used along with pure cultures of five saprophytic fungus (molds) and two strains of yeast were utilized. CBLM-H and CBLM-HEt were exhibited praiseworthy antimicrobial potential. CBLM-H showed complete growth inhibition of P. mirabilis and V. cholerae at the concentration of 0.1g/mL while CBLM-HEt at 0.05g/mL halted the growth of S. aureus.

Bio-functional hydrogel membranes loaded with chitosan nanoparticles for accelerated wound healing.

Wounds are often recalcitrant to traditional wound dressings and a bioactive and biodegradable wound dressing using hydrogel membranes can be a promising approach for wound healing applications. The present research aimed to design hydrogel membranes based on hyaluronic acid, pullulan and polyvinyl alcohol and loaded with chitosan based cefepime nanoparticles for potential use in cutaneous wound healing. The developed membranes were evaluated using dynamic light scattering, proton nuclear magnetic resonance, Fourier transform infrared spectroscopy, thermogravimetric analysis, and scanning electron microscopy. The results indicated the novel crosslinking and thermal stability of the fabricated hydrogel membrane. The in vitro analysis demonstrates that the developed membrane has water vapors transmission rate (WVTR) between 2000 and 2500 g/m2/day and oxygen permeability between 7 and 14 mg/L, which lies in the range of an ideal dressing. The swelling capacity and surface porosity to liberate encapsulated drug (cefepime) in a sustained manner and 88% of drug release was observed. The cefepime loaded hydrogel membrane demonstrated a higher zone of inhibition against Staphylococcus aureus, Pseudomonas aeruginosa, and Escherichia coli and excisional rat model exhibit expeditious recovery rate. The developed hydrogel membrane loaded with cefepime nanoparticles is a promising approach for topical application and has greater potential for an accelerated wound healing process.

Luffa cylindrica Immobilized with Aspergillus terreus QMS-1: an Efficient and Cost-Effective Strategy for the Removal of Congo Red using Stirred Tank Reactor.

Microbial populations within the rhizosphere have been considered as prosperous repositories with respect to bioremediation aptitude. Among various environmental contaminants, effluent from textile industries holds a huge amount of noxious colored materials having high chemical oxygen demand concentrations causing ecological disturbances. The study was aimed to explore the promising mycobiome of rhizospheric soil for the degradation of azo dyes to develop an efficient system for the exclusion of toxic recalcitrants. An effluent sample from the textile industry and soil samples from the rhizospheric region of Musa acuminata and Azadirachta indica were screened for indigenous fungi to decolorize Congo red, a carcinogenic diazo dye, particularly known for its health hazards to the community. To develop a bio-treatment process, Aspergillus terreus QMS-1 was immobilized on pieces of Luffa cylindrica and exploited in stirred tank bioreactor under aerobic and optimized environment. Quantitative estimation of Congo red decolorization was carried out using UV-Visible spectrophotometer. The effects of fungal immobilization and biosorption on the native structure of Luffa cylindrica were evaluated using a scanning electron microscope. A. terreus QMS-1 can remove (92%) of the dye at 100 ppm within 24 h in the presence of 1% glucose and 1% ammonium sulphate at pH 5.0. The operation of the bioreactor in a continuous flow for 12 h with 100 ppm of Congo red dye in simulated textile effluent resulted in 97% decolorization. The stirred tank bioreactor was found to be a dynamic, well maintained, no sludge producing approach for the treatment of textile effluents by A. terreus QMS-1 of the significant potential for decolorization of Congo red.Microbial populations within the rhizosphere have been considered as prosperous repositories with respect to bioremediation aptitude. Among various environmental contaminants, effluent from textile industries holds a huge amount of noxious colored materials having high chemical oxygen demand concentrations causing ecological disturbances. The study was aimed to explore the promising mycobiome of rhizospheric soil for the degradation of azo dyes to develop an efficient system for the exclusion of toxic recalcitrants. An effluent sample from the textile industry and soil samples from the rhizospheric region of Musa acuminata and Azadirachta indica were screened for indigenous fungi to decolorize Congo red, a carcinogenic diazo dye, particularly known for its health hazards to the community. To develop a bio-treatment process, Aspergillus terreus QMS-1 was immobilized on pieces of Luffa cylindrica and exploited in stirred tank bioreactor under aerobic and optimized environment. Quantitative estimation of Congo red decolorization was carried out using UV-Visible spectrophotometer. The effects of fungal immobilization and biosorption on the native structure of Luffa cylindrica were evaluated using a scanning electron microscope. A. terreus QMS-1 can remove (92%) of the dye at 100 ppm within 24 h in the presence of 1% glucose and 1% ammonium sulphate at pH 5.0. The operation of the bioreactor in a continuous flow for 12 h with 100 ppm of Congo red dye in simulated textile effluent resulted in 97% decolorization. The stirred tank bioreactor was found to be a dynamic, well maintained, no sludge producing approach for the treatment of textile effluents by A. terreus QMS-1 of the significant potential for decolorization of Congo red.

Bacillus tequilensis ZMS-2: A novel source of alkaline protease with antimicrobial, anti-coagulant, fibrinolytic and dehairing potentials.

Emerging resistance to existing antimicrobial agents is one of the growing concerns and a serious problem for public health globally. Currently available antimicrobial agents are potent and effective but surfacing resistance to these drugs has not been ruled out so far. Therefore, it is utmost important to explore new bioactive compounds from natural sources to meet future needs. The present study was designed to produce, optimize, characterize and evaluate antimicrobial, fibrinolytic and anti-coagulant potential of a new alkaline protease. Proteolytic strain from desert soils of Tharparkar, Pakistan was subjected to 16S rDNA sequencing and identified as Bacillus tequilensis ZMS-2(Genbank Accession No. MK101013). During submerged fermentation at 37ºC, maximum enzyme production (454 U/ml) was observed with 24h old inoculum. The best incubation time was 72h (544 U/ml), optimum inoculum size and pH was 10% at pH 8 with 494 and 506 U/ml, respectively. The best carbon source was starch (571 U/ml), while ideal substrate was wheat bran (536 U/ml). Optimal temperature and pH for proteolytic activity was 60ºC (420 U/ml) and 8 (332 U/ml). Alkaline protease showed antibacterial activity against Staphylococcus aureus (27mm), Bacillus licheniformis (20mm), Klebsiella pneumoniae (17mm) and Escherichia coli (15mm). The strain B. tequilensis ZMS-2 also exhibited anticoagulant, fibrinolytic and dehairing potential suggesting application of its protease in various industries.

Non-albicans Candida species: Emergence of neglected pathogens among population of Karachi.

Candida albicans was considered as the principal cause of opportunistic candidiasis but nowadays, neglected non-albicans Candida (NAC) species are evolving as more virulent and drug resistant strains. This research was intended to assess pervasiveness of candidiasis mainly caused by NAC species in Karachi city. A total of 562 clinical isolates of Candida spp. collected during the period of one year were identified by microscopic as well as morphological (germ tube formation, characteristics on CHROM agar and Corn meal agar) and Biochemical (sugar assimilation and fermentation) characteristics. Doubtful species were further identified by using Remel RapIDTM yeast plus kit. The results were statistically analyzed by SPSS 16.0 version software. Isolated strains of candida revealed slight predominance of C. albicans (54.5%) over non- albicans Candida species (45.5%). Among NAC species, C. tropicalis and C. glabrata were isolated as the predominant species. These clinical species were procured mainly from urine samples of females (73.7%) of age group 20-30 years. No significant correlations exist between Candida species and their months of isolation as well as their isolation from different districts of Karachi. Emergence of NAC species may predict an upcoming threat in health care facilities and hence, require prompt management and accurate identification to suggest empirical antifungal therapy.

Antimicrobial potential of newly isolated Aspergillusterreus MK-1: An approach towards new antibiotics.

OBJECTIVE: To attempt discovering new bioactive metabolites from fungal sources. METHODS: The exploratory study was conducted at the Department of Microbiology, Federal Urdu University for Arts, Science and Technology, Karachi from January 2016 to November 2017and comprised of soil samples collected from rhizosphere region of different garden plants from the city. Fungi were screened for production of antibiotics by testing cell-free culture filtrates obtained by Shake-flask fermentation technique. Agar-Well diffusion assay method was used to evaluate antagonistic activity against pathogenic microorganisms. RESULTS: Bioactive compounds extracted by ethyl acetate and thin layer chromatography revealed mixture of compounds in the crude extract. AspergillusterreusMK-1 showed significant inhibition of medically important test pathogens namely Staphylococcus aureus, Pseudomonas aeruginosa, Escherichiacoli, Salmonella typhi, Micrococcus luteus, Streptococcus epidermidis, Bacillus subtilis, Candida albicans and Aspergillusniger. The best biological activity of crude ethyl acetate extract was observed against Pseudomonas aeruginosa (63mm). CONCLUSIONS: Newly isolated AspergillusterreusMK-1 emerged as a potent candidate for the production of antimicrobial compounds.

Fungal contamination in smokeless tobacco products traditionally consumed in Pakistan.

OBJECTIVE: To isolate potential pathogenic fungi from smokeless tobacco products. METHODS: The study was conducted from January 2015 to February 2017 during which samples of smokeless tobacco products such as Mainpuri, Tambako, Khiwam, Gutkha, Naswar and Mawa etc. were collected from different cities of Pakistan. The samples were tested for fungal contamination by spread plate method. Different strains of fungi were isolated and identified on the basis of their macroscopic as well as microscopic characteristics. The fungal strains isolated were also screened for their susceptibility to commonly used antifungal drugs by disc diffusion method. RESULTS: Of the 600 samples collected, 300(50%) were from Sindh, 70(11.7%) Balochistan, 74(12.3%) from Khyber Pakhtunkhwa, 105(17.5%) from Punjab and 51(8.5%) from Azad Kashmir. In terms of products, there were 404(67.3%) samples of Naswar, 69(11.5%) Patti, 40(6.6%) Khiwam, 35(5.8%) Mawa, 32(5.3%) Gutkha, and 20(3.3%) Mainpuri samples. Different species of Aspergillus were predominantly isolated followed by Penicillium, Mucor, Sepedonium and Trichophyton. The isolated strains of Aspergillus also revealed resistance against many commonly-used anti-fungals such as Amphotericin B and Itraconazole.. CONCLUSIONS: There was high prevalence of opportunistic fungi in study samples, posing a threat for human health which requires prompt notice and management.

Mother's Work Status on Children's Bruxism in a Subset of Saudi Population.

AIMS AND OBJECTIVES: The aims and objectives of this study were to determine if an association exists between mothers work status and her children's incidence of bruxism and habits related to bruxism. MATERIALS AND METHODS: A cross-sectional study was conducted through data collection of a questionnaire answered by 561 mothers' about their working status and their child's habits and behaviors. The survey consisted of 5 parts with a total of 34 questions: mother's information, child's behavior, child's sleeping pattern, mother's knowledge about bruxism, and child's medical history. Odds ratios, Chi-square, and their corresponding 95% confidence intervals are reported. Statistical significance was set at P ≤ 0.05. RESULTS: The work status of the mother was not statistically significant in increasing the incidence of a child to have bruxism. However, this study clearly elucidates that 7 of the 15 habits correlate significantly with a status of bruxism. According to this sample, a child, that is, reported to be aggressive is more than twice as likely to have nocturnal bruxism. Likewise, any child that bites their nails, complains of headaches, drools in their sleep, snores, complains of muscle cramps, and colic is more than twice as likely to be a nocturnal bruxer than a child that does not have these habits. CONCLUSION: The prevalence of children's bruxism in this convenient sample was 34.5% (n = 141). The concerning habits related to bruxism can serve the pediatric dentist, general dentist, general practitioner, and primary care provider of children having these red flags as indicators of bruxism. It is imperative that parents of these children be made aware these habits that may occur together, alone or even simultaneously with bruxism.

Phytochemical and Biological Activities of Pseudocalymma elegans: A False Garlic.

Evaluation of phytochemical constituents and antioxidant and antimicrobial activities of hexane (PELH), dichloromethane (PELDCM), ethyl acetate (PELEA), and MeOH (PELM) extracts of young leaves of Pseudocalymma elegans have been carried out. Moreover, extracts have also been explored for the presence of sulphur containing compounds, 1,2-dithiolane (33), diallyl disulfide (35), 3-vinyl-1,2-dithiacyclohex-5-ene (37), and diallyl trisulfide (38) responsible for the garlic like smell of P. elegans. All the extracts were found to be antioxidant and showed potent inhibition with IC50 values of 0.168 ± 0.001, 0.128 ± 0.002, 0.221 ± 0.011, and 0.054 ± 0.001, respectively, as compared to standard drugs ascorbic acid (AA) and butylated hydroxytoluene (BHT). The ethyl acetate extract (PELE) showed excellent activities against few Gram-positive and Gram-negative bacteria and some fungi as compared with standard drug ceftriaxone (3rd generation cephalosporin) and nystatin, respectively. Chemical constituents of hexane, dichloromethane, and ethyl acetate extracts were identified by gas chromatography-mass spectrometry and mass spectral library search. Over all 55 chemical constituents were first time identified from the leaves which included branched and n-hydrocarbons, fatty acids, fatty acid methyl esters, fatty alcohols, terpenes, alkaloid, vitamins, glycosides, aromatic compounds, and sulfur containing compounds. Two known chemical constituents, ursolic acid (1) and ß-amyrin (2), were also purified for the first time from the MeOH extract. To elucidate the structures of these compounds, UV, IR, EI-MS, 1 H- and 13 C-NMR spectroscopy were used.

Isolation of potentially pathogenic fungi from selected pigeons' feeding sites in Karachi: A new dimension to health hazard.

OBJECTIVE: To determine the presence of pathogenic fungal strains in areas where pigeons are present in a large number. METHODS: This study was conducted at the Federal Urdu University of Arts, Science and Technology, Karachi, from February 2015 to March2016, and comprised samples of soil contaminated with pigeons' excreta. The samples were collected from 20 different pigeon-feeding places in the city. These samples were processed for the isolation and identification of fungi by using standard conventional methods. The fungal strains isolated were also tested for their susceptibility to commonly used antifungal agents by disc diffusion technique. RESULTS: There were 105 samples. A wide variety of fungal strains belonging to different genera of Aspergillus, Rhizopus, Penicillium, Fusarium and Candida were isolated and identified by using conventional methods. The antifungal resistance pattern of these strains also depicts emergence of resistance against commonly used antifungal agents such as amphotericin B and fluconazole. CONCLUSIONS: The soil and air of places densely populated with pigeons were found to be loaded with fungal spores and many of them were potential pathogens.

Synthesis, spectroscopic characterization and antimicrobial activities of benzoxazolone derivatives.

: Background: Pathogenic microbial diseases are now the key virulence in our daily life. Significant research has been carried out in order to trigger the bacterial infections. Amongst the organic molecules, oxazolone and derivatives were found to have excellent bioactivities including antimicrobial activities. METHODS: By keeping in mind the considerable antimicrobial activities of class benzoxazolones, a series of benzoxazolone derivatives 3-16 have been synthesized. Out of which five compounds 10, 11, 14, 15, and 16 were new synthetic derivatives whereas compounds 9, 12, and 13 were already known compounds. These compounds have been synthesized by refluxing of amino phenol and 1,1-carbonyldiimidazole1 (C3H3N2)2CO) (CDI) in a dry THF and then treated with commercially available acid chloride. The structures of the compounds were elucidated on the basis of 1H-NMR, EIMS and elemental analysis. All the compounds were screened for their antibacterial activities and tested by agar well diffusion method. RESULTS: Compounds 14 and 16 showed good activity against S. aureus. Compound 5 showed good while 14 and 16 were found to be most active against E. coli using cefuroxime as a standard. Antifungal activities were carried out by using standard drug nystatin and compounds 4, 5, 9, 11 and compound 12 were found to be active against C. albicans. Compounds 4, 5, 9 and compound 10 showed good while 7, 11, and compound 13 showed excellent activities against Chrysosporium sp. Compounds 6, 7 and compound 12 were found to be most active against A niger and A. flavus, respectively. CONCLUSION: A number of derivatives were identified to have potent antimicrobial activities and may serve as lead compounds for future research.

Radiative Peristaltic Flow of Jeffrey Nanofluid with Slip Conditions and Joule Heating.

Mixed convection peristaltic flow of Jeffrey nanofluid in a channel with compliant walls is addressed here. The present investigation includes the viscous dissipation, thermal radiation and Joule heating. Whole analysis is performed for velocity, thermal and concentration slip conditions. Related problems through long wavelength and low Reynolds number are examined for stream function, temperature and concentration. Impacts of thermal radiation, Hartman number, Brownian motion parameter, thermophoresis, Joule heating and slip parameters are explored in detail. Clearly temperature is a decreasing function of Hartman number and radiation parameter.