Showing papers on "Agar plate published in 2011"

Use of Blue Agar CAS Assay for Siderophore Detection.

Abstract: Schwyn and Neiland developed a universal siderophore assay using chrome azurol S(CAS) and hexadecyltrimethylammonium bromide (HDTMA) as indicators. In Schwyn and Neiland’s original paper, the procedure given for making CAS agar is written in general terms and can be difficult to follow, especially for an individual who has limited experience making more complex media. Here, we give a step-by-step protocol for making the CAS agar plates, and we discuss how this media can be incorporated in a comprehensive project in a microbiology lab course for biology majors.

Malassezia cuniculi sp. nov., a novel yeast species isolated from rabbit skin

Abstract: Members of the genus Malassezia have rarely been associated with lagomorphs. During the course of an investigation of the lipophilic mycobiota of rabbit skin, two lipid-dependent isolates which could not be identified were recovered on Leeming and Notman agar medium from different animals. No growth of Malassezia yeasts was obtained either on Sabouraud's glucose agar or modified Dixon agar media. In this study, we describe a new taxon, Malassezia cuniculi sp. nov., including its morphological and physiological characteristics. The validation of this new species was supported by analysis of the D1/D2 regions of the 26S rRNA gene and the ITS-5.8S rRNA gene sequences. The results of these studies confirm the separation of this new species from the other species of the genus Malassezia, as well as the presence of Malassezia yeasts on lagomorphs.

Clinical use of photodynamic antimicrobial chemotherapy for the treatment of deep carious lesions.

Abstract: The purpose of this study was to assess photodynamic antimicrobial chemotherapy (PACT) via irradiation, using a low power laser associated with a photosensitization dye, as an alternative to remove cariogenic microorganisms by drilling. Remaining dentinal samples in deep carious lesions on permanent molars (n = 26) were treated with 0.01% methylene blue dye and irradiated with a low power laser (InGaAIP - indium gallium aluminum phosphide; λ = 660 nm; 100 mW; 320 Jcm(-2); 90 s; 9J). Samples of dentin from the pulpal wall region were collected with a micropunch before and immediately after PACT and kept in a transport medium for microbiological analysis. Samples were cultured in plates of Brucella blood agar, Mitis Salivarius Bacitracin agar and Rogosa SL agar to determine the total viable bacteria, mutans streptococci and Lactobacillus spp. counts, respectively. After incubation, colony-forming units were counted and microbial reduction was calculated for each group of bacteria. PACT led to statistically significant reductions in mutans streptococci (1.38 log), Lactobacillus spp. (0.93 log), and total viable bacteria (0.91 log). This therapy may be an appropriate approach for the treatment of deep carious lesions using minimally invasive procedures.

Optimization, production and partial purification of extracellular α-amylase from Bacillus sp. marini

Abstract: The aim of the current study was to isolate amylase producing bacteria from the soil samples collected from marine environment of Andaman and Nicobar Islands, India. The isolation was performed by serial dilution and plating method. Total 35 bacteria were isolated from the collected soil samples. All isolates were screened for amylolytic activity by starch agar plate method, among 35 bacterial isolates only 4 isolates showed the amylolytic activity, one isolate was selected for further study. The isolate (potential strain) was identified as Bacillus sp. marini by microscopic, biochemical and molecular experiments. The best enzyme activity was observed at pH 7 and temperature 40°C, 5.5% NaCl concentration, starch as carbon source and yeast extract as nitrogen source. At the optimum conditions B. sp. marini produced 8000 U of amylase per litre of culture broth, which is 3 fold higher than before optimization.

Detection of Salmonella and Vibrio species in some seafood in Alexandria

Abstract: Sea foods are prone to bacterial contamination and could cause health risk to consumers. The present work aimed to study the occurrence of Salmonella and Vibrio in some seafood from different markets in Alexandria. The study was carried out on 150 seafood samples (shrimp, oyster (Gandofli) and mussel (Om El Kholoul)). For detection of Salmonella; samples were cultured on CHROM agar Salmonella Plus medium and Xylose lysine desoxycholate (XLD) agar plates. Thiosulphate Citrate Bile Salt Sucrose (TCBS) agar was used for Vibrio isolation. Salmonella was isolated from 10.0% of samples, distributed as 7 (14.0%), 4 (8%) and 4 (8%) from shrimp, oyster and mussel respectively. Vibrios were isolated from 52.0% of tested seafood with the highest percentage (88.0%) from oyster. The most frequently isolated Vibrio spp. were V. alginolyticus (52.5%), V. parahaemolyticus (14.1%) and V. mimcus (11.5%). Seven different Salmonella serotypes were identified (Typhimurium, Derby, Typhi, Paratyphi A, Paratyphi B, Infantis, and Abortus equi). Our results confirm that bacterial contamination in seafood products is common, and suggest that routine examination of such products for pathogenic agents would be advisable. (Wafaa MK Bakr, Walaa A Hazzah, Amani F Abaza. Detection of Salmonella and Vibrio species in some seafood

Responses of Escherichia coli, Listeria monocytogenes, and Staphylococcus aureus to Simulated Food Processing Treatments, Determined Using Fluorescence-Activated Cell Sorting and Plate Counting

Abstract: Three common food pathogenic microorganisms were exposed to treatments simulating those used in food processing. Treated cell suspensions were then analyzed for reduction in growth by plate counting. Flow cytometry (FCM) and fluorescence-activated cell sorting (FACS) were carried out on treated cells stained for membrane integrity (Syto 9/propidium iodide) or the presence of membrane potential [DiOC2(3)]. For each microbial species, representative cells from various subpopulations detected by FCM were sorted onto selective and nonselective agar and evaluated for growth and recovery rates. In general, treatments giving rise to the highest reductions in counts also had the greatest effects on cell membrane integrity and membrane potential. Overall, treatments that impacted cell membrane permeability did not necessarily have a comparable effect on membrane potential. In addition, some bacterial species with extensively damaged membranes, as detected by FCM, appeared to be able to replicate and grow after sorting. Growth of sorted cells from various subpopulations was not always reflected in plate counts, and in some cases the staining protocol may have rendered cells unculturable. Optimized FCM protocols generated a greater insight into the extent of the heterogeneous bacterial population responses to food control measures than did plate counts. This study underlined the requirement to use FACS to relate various cytometric profiles generated by various staining protocols with the ability of cells to grow on microbial agar plates. Such information is a prerequisite for more-widespread adoption of FCM as a routine microbiological analytical technique.

Frequency of Detection of Escherichia coli, Salmonella spp., and Campylobacter spp. in the Faeces of Wild Rats (Rattus spp.) in Trinidad and Tobago

Abstract: The study was conducted to determine the frequency of isolation of Salmonella, Campylobacter and E. coli O157 in the faecal samples of rats trapped across the regional corporations in Trinidad and to assess their resistance to antimicrobial agents. A total of 204 rats were trapped for the detection of selected bacteria. Standard methods were used to isolate Salmonella, Campylobacter and E. coli O157. Characterization of E. coli was done on sorbitol MacConkey agar to determine non-sorbitol fermentation, blood agar to determine haemolytic and mucoid colonies and by using E. coli O157 antiserum to determine O157 strain. The disc diffusion method was used to determine resistance to nine antimicrobial agents. Of the 204 rats, 4 (2.0%), 7 (3.4%) and 171 (83.8%) were positive for Salmonella spp., Campylobacter spp. and E. coli, respectively. Of the 171 isolates of E. coli tested 0 (0.0%), 25 (14.6%) and 19 (11.1%) were haemolytic, mucoid and non-sorbitol fermenters, respectively. All isolates were negative for the O157 strain. The frequency of resistance to the 9 antimicrobial agents tested was 75% (3 of 4) for Salmonella, 85.7% (6 of 7) of Campylobacter spp. and 36.3% (62 of 171) for E. coli (P < .05; χ2).

Antimicrobial activity of Calendula officinalis, Camellia sinensis and chlorhexidine against the adherence of microorganisms to sutures after extraction of unerupted third molars

Abstract: OBJECTIVE: The objective of this study was to compare the antimicrobial effect of mouthwashes containing Calendula officinalis L., Camellia sinensis (L.) Kuntze and 0.12% chlorhexidine digluconate on the adherence of microorganisms to suture materials after extraction of unerupted third molars. MATERIAL AND METHODS: Eighteen patients with unerupted maxillary third molars indicated for extraction were selected (n=6 per mouthwash). First, the patients were subjected to extraction of the left tooth and instructed not to use any type of antiseptic solution at the site of surgery (control group). After 15 days, the right tooth was extracted and the patients were instructed to use the Calendula officinalis, Camellia sinensis or chlorhexidine mouthwash during 1 week (experimental group). For each surgery, the sutures were removed on postoperative day 7 and placed in sterile phosphate-buffered saline. Next, serial dilutions were prepared and seeded onto different culture media for the growth of the following microorganisms: blood agar for total microorganism growth; Mitis Salivarius bacitracin sucrose agar for mutans group streptococci; mannitol agar for Staphylococcus spp.; MacConkey agar for enterobacteria and Pseudomonas spp., and Sabouraud dextrose agar containing chloramphenicol for Candida spp. The plates were incubated during 24-48 h at 37oC for microorganism count (CFU/mL). RESULTS: The three mouthwashes tested reduced the number of microorganisms adhered to the sutures compared to the control group. However, significant differences between the control and experimental groups were only observed for the mouthwash containing 0.12% chlorhexidine digluconate. CONCLUSIONS: Calendula officinalis L. and Camellia sinensis (L.) Kuntze presented antimicrobial activity against the adherence of microorganisms to sutures but were not as efficient as chlorhexidine digluconate.

EXTRACELLULAR ENZYME PRODUCTION BY INDIGENOUS THERMOPHILIC BACTERIA: PARTIAL PURIFICATION AND CHARACTERIZATION OF α-AMYLASE BY BACILLUS SP. WA21

Abstract: Thermostability is a characteristic of most of the enzymes available for bulk industrial usage. Thermophilic microorganisms are of special interest as a source of novel thermostable enzymes. A total of 50 bacterial strains, isolated from local hot springs and ash samples were screened for the extracellular enzyme production including amylase, lipase, esterase, cellulase and β-galactosidase. As a follow up, studies on α-amylase were carried out with a bacterial strain identified as Bacillus sp. WA21 (from hot spring) on the basis of maximum zone of starch hydrolysis in agar plate medium. Bacillus WA21 showed growth over a wide range of temperature (35-55oC) and pH (3-11) with optimum being 45oC and pH 6. Maximum enzyme production was observed after 144 hours. The enzyme was found optimally active at 55oC and pH 6. Temperature stability profile revealed that enzyme α-amylase retained more than half of its activity at 85oC and between pH 5-9. Thus, Bacillus WA21 may be regarded as a promising source of α-amylase for biotechnological and industrial applications.

Resveratrol Inhibits Periodontal Pathogens In Vitro

Abstract: The gram-negative anaerobic bacteria A. actinomycetemcomitans (Aa) and P. gingivalis (Pg) are key components in the aetiology of periodontal disease, and associated hard-tissue destruction. Resveratrol is a phytoalexin, produced naturally by several plants when under attack by bacterial or fungal pathogens. It is found in many foods including mulberries, peanuts and the skin of labrusca and muscadine grapes. The objective of this study was to evaluate the effect of resveratrol on the in vitro growth of periodontal pathogens Aa and Pg. For comparison, resveratrol's effect on a variety of other oral microorganisms was also evaluated. Resveratrol demonstrates a poor solubility in water, thus different concentrations of resveratrol in the solvent dimethyl sulphoxide (DMSO) were added to calibrated suspensions of Aa and Pg. As a control, a parallel series of dilutions containing the vehicle DMSO alone was made to measure the effect of the solvent. Minimum inhibitory concentrations of the periodontal pathogens were calculated. All suspensions were incubated for 1, 3, 6 and 24 h in an anaerobic chamber at 37 °C. At each time interval, selected dilutions from each culture broth were plated on blood agar plates. Colonies appearing on blood agar plates were visually counted at 3 days for Aa, and at 5 days for Pg. The periodontal bacteria showed a significant decrease (p < 0.05) in viable counts after 1 h, whilst no colony forming units could be observed after 24 h. The results suggest that resveratrol possesses significant antimicrobial properties on periodontal pathogens in vitro. Copyright © 2011 John Wiley & Sons, Ltd.

Laboratory Detection of Extended-Spectrum-Beta-Lactamase-Producing Enterobacteriaceae: Evaluation of Two Screening Agar Plates and Two Confirmation Techniques

Abstract: The worldwide prevalence of extended-spectrum-beta-lactamase-producing ESBL-producing Enterobacteriaceae (ESBL-E) is increasing, making the need for optimized detection techniques more urgent. In this study we investigated the performance of two ESBL-E screening and two ESBL-E confirmation techniques. In accordance with the Dutch national guidelines (www.wip.nl), a collection of 642 highly resistant Enterobacteriaceae strains, as identified by Vitek2, was used to test the performances of two screening techniques (EbSA ESBL agar plate and ChromID ESBL agar plate) and of two confirmation techniques (MIC-strip ESBL and Vitek2 ESBL test panel). The individual test results were compared by using Etest, followed by a combination disk test if Etest results were inconclusive. Among group 1 isolates (Escherichia coli, Klebsiella spp., Proteus spp., Salmonella spp., and Shigella spp.) 291 (57.6%) were ESBL-E, versus 65 (47.4%) in group 2 (Enterobacter spp., Citrobacter spp., Morganella morganii, Serratia spp., and Providencia spp.). The sensitivities of all four tests for group 1 were comparable (EbSA, 96.6%; ChromID, 97.3%; MIC-strip, 99.6%; and Vitek2, 95.1%). The specificities of the EbSA and ChromID were the same (93.9%). However, the confirmation techniques produced many inconclusive test results, which reduces the applicability in routine laboratories. Only the two screening agar plates were validated for ESBL testing of group 2 microorganisms. They showed comparable sensitivities; however, the EbSA screening agar plate had a significantly higher specificity (78.6% versus 44.3%). In conclusion the screening agar plates performed better than the two confirmation techniques. The EbSA agar plate had the best overall performance.

Inhibitory activity of Pseudomonas sp. on Flavobacterium psychrophilum, in vitro

Abstract: A Pseudomonas sp. isolate MSB1 efficiently inhibited the growth of Flavobacterium psychrophilum of different serotypes on agar medium. A significant difference in the inhibition was observed between isolates of the less virulent Fp(T) serotype compared to the Fd and Th serotypes. In broth coculture experiments, a low number of cells of MSB1 inhibited and outcompeted the F. psychrophilum cells. Also cell-free culture supernatant of MSB1 clearly repressed the growth of F. psychrophilum. A chromoazurol S assay suggested that MSB1 produced efficient siderophores, which most probably were responsible for the iron deficiency in the supernatant. The limited growth of F. psychrophilum in the supernatant was found to be partly because of the lack of available iron, but the results also indicated that some other mechanisms were probably involved in the observed inhibition. A potential use of MSB1 as a probiotic in rainbow trout aquaculture, especially in early life stages of the fish, is suggested, but future in vivo experiments needs to be carried out to verify this suggestion. This study also indicates a low iron acquisition efficiency of F. psychrophilum, compared to other examined bacterial fish pathogens.

Antimicrobial Activity of Pyocyanin Produced by Pseudomonas aeruginosa Isolated from Surgical Wound-Infections

Abstract: 2 Abstract: Five different isolates of Pseudomonas aeruginosa were obtained from surgical specimens and minced meat. The isolates were tested for the production of the blue pigment; pyocyanin. Considerable amounts of blue pigment were produced by P. aeruginosa isolates when grown on the four tested media. Muller-Hinton agar was further used for growth, pigment production and sensitivity tests. Pigment production began during the first 24 hrs of growth and maximal pigment production was achieved after 48 hrs by isolates No. 1, 2 and 3. At this time, isolate No. 5 produced only moderate level of pyocyanin. While isolate No. 4 reached the highest yield after 72 hrs. Pyocyanin pigments obtained from culture supernatants of the tested P.aeruginosa isolates "grown in peptone water liquid medium" by serial chloroform extractions reached about 62.8µg/ml. The growth of all Gram-positive bacteria and Candida species was completely inhibited when cultivated on the agar plates containing the blue pigment. Whereas, Gram negative bacteria; Klebsiella pneumoniae and P. aeruginosa were resistant to pyocyanin. Salmonella typhi and Proteus mirabilis were intermediately affected. Thus, the five tested P. aeruginosa isolates were resistant to their own produced pyocyanin pigments. In a Conclusion, the Gram-positive bacteria were more susceptible as a group to the antibiotic action of pyocyanin than were the Gram negative bacteria. The antibacterial and antifungal nature of the pigment is attractive for the topical treatment of wound infections.

Comparative evaluation of a chromogenic agar medium-PCR protocol with a conventional method for isolation of Vibrio parahaemolyticus strains from environmental and clinical samples.

Abstract: Screening for pathogenic Vibrio parahaemolyticus has become routine in certain areas associated with food-borne outbreaks. This study is an evaluation of the CHROMagar Vibrio (CV) medium – PCR prot...

Production and optimization of acid protease by Aspergillus spp under submerged fermentation

Abstract: Proteases are the most important industrial enzymes and comprise about 25% of commercial enzymes in the world. Two third of the industrially produced proteases are from microbial sources. The proteolytic fungi Aspergillus spp was isolated from soil contaminated with abattoir waste and screened on skim milk agar medium for proteolytic activity. The enzyme was optimized supplementation of cheese whey and other favorable conditions like pH, temperature and incubation time to the medium. The optimized conditions for enzyme production were 5.0, 32 ± 20C and 5 days intervals respectively. The fermentation medium was supplemented with different nitrogen and carbon sources to improve the enzyme production. Among the sources tested in the present study, soybean meal (1%) was proved the best nitrogen source whereas glucose and rice flour were shown at 1.5% and 1% for enzyme production by Aspergillus spp respectively.

A simple and rapid plate assay for screening of inulindegrading microorganisms using Lugol’s iodine solution

Abstract: In this report, a simple and rapid agar plate assay was established for screening of halophilic, inulindegrading microorganisms. Two strains considered inulinolytic with this method were chosen and the inulinolytic activities in their culture supernatant were measured with the Somogyi-Nelson method, while their hydrolysis products of inulin were detected with TLC chromatogram. Key words: Screening, halophilic microorganism, inulinase, Lugol’s iodine solution.

Molecular identification of antagonistic bacteria from Tehran soils and evaluation of their inhibitory activities toward pathogenic fungi

Abstract: Background and Objectives: To find antagonistic bacteria with potential antifungal activity against some pathogenic fungi, including Aspergillus niger, A. flavus, Fusarium moniliforme and Penicillium marneffei, a total of 148 agricultural soil samples from different sites of Tehran were examined. Materials and Methods: Antagonistic soils were selected by screening against A. niger on glucose-yeast extract (GY) agar using a visual agar plate assay method. All growing bacteria were examined for antifungal activity, and antagonistic bacteria identified based on 16S rRNA sequence analysis. Among a total number of 97 bacteria isolated form inhibitory soils (36 samples), 16 bacteria were reported as strong growth inhibitors in co-cultures on GY agar with all tested fungi at variable degrees. Fungal growth inhibitory bacteria were cultured against all fungi and growth inhibition was measured and analyzed between test and control groups by statistical analysis (ANOVA). Results: Molecular identification of antagonistic bacteria indicated that most bacterial isolates belonged to the genus Bacillus (81.25%), including B. subtilis (5 isolates), B. amyloliquefaciens (6 isolates) and B. valismortis (2 isolates), followed by one isolate (6.25%) from each Streptomyces sp., Pseudomonas chlororaphis and Acinetobacter baumannii. Based on the visual plate assay results, total fungal growth inhibition of all bacteria was reported in the range of 13.2 to 68.3%. P. chlororaphis S105 was reported as the most potent antagonistic bacterium which inhibited the growth of A. niger by 68.3%, followed by F. moniliforme (66.4%), A. flavus (64.7%) and P. marneffei (57.1%). Conclusion: P. chlororaphis and some other inhibitory bacteria reported in the present study, they may be considered not only as a rich source of useful metabolites with potential application in antifungal drug discovery, but also as potential candidates for biological control programs.

Antibacterial Activity of Extracts of Ajuga Iva, and Teucrium Polium

Abstract: Antibiotics provide the main basis for the therapy of microbial (bacterial and fungal) infections. Since the discovery of these antibiotics and their use as chemotherapeutic agents there was a belief in the medical fraternity that this would lead to the eventual eradication of infectious diseases. However, overuse of antibiotics has become the major factor in the emergence and dissemination of multi-drug resistant strains of several groups of microorganisms. Many plants which are used in traditional medicine contain antimicrobial compounds. In this study, the antibacterial activity of methanolic extracts of Teucrium polium and Ajuga iva were tested against five bacteria, E. coli MC 4100, Bacillus subtilis, Pseudomonas diminutus, Paracoccus paratrophus and Micrococcus luteus. T. polium and A. iva plants were collected and allowed to dry in the dark at room temperature. Dried plant material (100g) was added to 1 L of methanol and incubated at room temperature for three days. The crude solution was filtered through muslin cloth, and the filtrate evaporated to dryness. The dried material was dissolved in 2 ml of methanol. Bacterial suspensions (100 µl) were spread on tryptone soya agar (TSA) medium. Plant extracts (10 µl) were applied to discs of filter paper and placed on agar plates containing the microorganisms. The plates were incubated at 37°C for 48h. After incubation the zones of inhibition around the discs was measured. Extracts of T. polium gave zones of inhibition against Bacillus subtilis, Micrococcus luteus and Paracoccus paratrophus of 3.7, 2.0 and 2.0 mm, respectively. A. iva extract only inhibited the growth of Paracoccus paratrophus, giving a zone of inhibition of 3.0.mm. The present results showed that extracts of T. polium inhibited the growth of three bacterial species. Extracts of A. iva, on the other hand, inhibited only one bacterium. Key word: Ajuga iva, antibacterial activity, plant extracts, Teucrium polium.

SOLUBILIZACIÓN in vitro DE FOSFATOS POR UNA CEPA DE Paecilomyces lilacinus (Thom) Samson In vitro PHOSPHATE SOLUBILIZATION BY A STRAIN OF Paecilomyces lilacinus (Thom) Samson

Abstract: Phosphorus is an element with limited availability in the soil, which can be improved by solubilizing fungi of calcium and iron phosphates. Therefore, the objective of this study was to evaluate in vitro the solubilization of phosphate compounds by Paecilomyces lilacinus in solid and liquid culture media. In solid media agar plates were used with each phosphate compound, and a control without phosphates, which were inoculated with spores of the fungus. The halos of solubilization were evaluated and the relative efficiency of solubilization (ERS) was calculated. In tests with liquid media, P. lilacinus was inoculated in culture media with calcium and iron phosphate without agar, in which the soluble phosphorus (P) and pH were quantified. In agar plates with calcium phosphate solubilization halos were observed and ESR was 305 % at 8 d, but in media with iron phosphate halos were not formed. In the liquid medium with calcium phosphate a greater amount of soluble P (71.287.88 mg L 1 ) was obtained compared with the medium with iron phosphate (1.750.32 mg L 1 ). In the presence of iron phosphate, P. lilacinus immobilized most of the solubilized phosphorus and left a small proportion available. The pH of both culture media decreased in the presence of fungus, which is likely that the main mechanism of solubilization is the production of organic acids.

Cellulase production by pink pigmented facultative methylotrophic strains (PPFMs).

Abstract: Pink pigmented facultative methylotrophs (PPFM) isolated from water samples of Cooum and Adyar rivers in Chennai and soil samples of forests located in various districts of Tamil Nadu, India were screened for cellulase production using carboxymethylcellulose agar (CMC agar) medium. The strains showed wide variations in the production of clearing zones around the colonies on CMC agar medium flooded with Congo red. CMCase and filter paper assays were used to quantitatively measure the cellulase activity of 13 PPFM strains. Among the strains, Methylobacterium gregans, MNW 60, MHW 109, MSF 34, and MSF 40 showed cellulolytic activity ranging from 0.73 to 1.16 U mL−1 with wide temperature (35–65°C) and pH (5 to 8) tolerance. SDS-PAGE analysis of the crude enzyme of PPFM strain MNW 60 exhibited several protein bands, and zymogram analysis revealed two dimeric cellulase bands with molecular mass of ~92 and 42 kDa. Scanning electron microscopic studies revealed significant morphological differences between the cells grown in normal and CMC amended medium. The strain MNW 60 was identified as Methylobacterium sp. based on biochemical, physiological, and morphological analyses, and the methylotrophic nature was authenticated by the presence of mxaF gene, encoding methanol dehydrogenase as a key indicator enzyme of methylotrophs, with 99% similarity to Methylobacterium lusitanum. With the 16S ribosomal RNA sequence showing 97% similarity to M. lusitanum strain MP2, this can be proposed as a novel taxon of the genus Methylobacterium. The study forms the first detailed report on the extracellular cellulase production by pink pigmented Methylobacterium sp., and it is expected that this might be the basis for further studies on cellulase production by PPFMs to explore the molecular mechanism, strain improvement, and large-scale cellulase production for its application.

Antimicrobial activity of different extracts of roots of Rumex nepalensis Spreng

Abstract: Six crude extracts from the roots of Rumex nepalensis Spreng. were evaluated for in vitro antimicrobial activity against two Gram positive (Staphylococcus aureus, Streptococcus mutans) and two Gram negative bacteria (Escherichia coli, Pseudomonas aeruginosa) and one fungus, Candida albicans. The agar plate well diffusion method was used for different samples. Benzene and ethyl acetate extracts demonstrated significant inhibitory effect against majority of microorganisms under test.

Antibacterial effect of calcium carbonate nanoparticles on Agrobacterium tumefaciens

Abstract: Aims: Improvements in nanotechnology in the past decayed has created various opportunities for evaluation of biologic effects such as anti-bacterial effects of nanoparticles. The purpose of this study was to evaluate the antibacterial activity of calcium carbonate nanoparticles on two different bacteria including A grobacterium tumefaciens and Staphylococcus aureu s. Methods: The antibacterial effect of calcium carbonate nanoparticles against mentioned bacteria was evaluated by dilution in agar containing medium and dilution in Broth medium. Each of prepared Broth Media (10 ml) was inoculated with 1 ml of bacterial suspension (10 6 CFU/ml) and incubated. Sampling of culture Media was performed in specific intervals and diluted as 10 -1 , to 10 -6 . Then 100 µl of each sample was transferred to agar plates and was spread carefully and then incubated. Grown colonies were counted and MIC and MBC was determined. Results: Calcium carbonate nanoparticles showed very good antibacterial effect and after 16 hours the bacteria were totally diminished. The lowest and highest MIC concentration of these nanoparticles in solid medium was 31.2 and 125 µg/ml respectively. The MIC of calcium carbonate nanoparticles in Broth Medium was two times more than the MIC concentration in solid medium, while different concentrations of ordinary calcium carbonate not only revealed antibacterial effects but also supported the bacterial growth. Conclusion: Use of calcium carbonate nanoparticles as an anti-microbial agent is recommended in different fields of medicine, food industry and agriculture and can be of importance considering health and economic issues.

Accurate assessment of antibiotic susceptibility and screening resistant strains of a bacterial population by linear gradient plate.

Abstract: The dynamics of a bacterial population exposed to the minimum inhibitory concentration (MIC) of an antibiotic is an important issue in pharmacological research. Therefore, a novel antibiotic susceptibility test is urgently needed that can both precisely determine the MIC and accurately select antibiotic-resistant strains from clinical bacterial populations. For this purpose, we developed a method based on Fick’s laws of diffusion using agar plates containing a linear gradient of antibiotic. The gradient plate contained two layers. The bottom layer consisted of 15 mL agar containing the appropriate concentration of enrofloxacin and allowed to harden in the form of a wedge with the plate slanted such that the entire bottom was just covered. The upper layer consisted of 15 mL plain nutrient agar added with the plate held in the horizontal position. After allowing vertical diffusion of the drug from the bottom agar layer for 12 h, the enrofloxacin concentration was diluted in proportion to the ratio of the agar layer thicknesses. The uniform linear concentration gradient was verified by measuring the enrofloxacin concentration on the agar surface. When heavy bacterial suspensions were spread on the agar surface and incubated for more than 12 h, only resistant cells were able to form colonies beyond the boundary of confluent growth of susceptible cells. In this way, the true MIC of enrofloxacin was determined. The MICs obtained using this linear gradient plate were consistent with those obtained using conventional antibiotic susceptibility tests. Discrete colonies were then spread onto a gradient plate with higher antibiotic concentrations; the boundary line increased significantly, and gene mutations conferring resistance were identified. This new method enables the rapid identification of resistant strains in the bacterial population. Use of the linear gradient plate can easily identify the precise MIC and reveal the dynamic differentiation of bacteria near the MIC. This method allows the study of genetic and physiological characteristics of individual strains, and may be useful for early warning of antibiotic resistance that may occur after use of certain antimicrobial agents, and guide clinical treatment.

Incidence of Aeromonas species isolated from water and fish sources of Lake Manzala in Egypt

Abstract: Lake Manzala situated in the east of Nile Delta, between the Damietta branch of Nile River and Suez Canal. The lake receives pollution from different sources, which make the lake polluted and eutrophicated. Samples were taken from five sites of the lake. Faecal coliforms showed the highest counts 2.2× 103 cfu ml–1 and 4× 103 cfu ml–1 in water in El-Kapoty and El-Mataryia areas, respectively. Aeromonas sppwere counted on Endo Agar, and Aeromonason selective Agar plates which reached the highest counts in water samples collected from Bahr El-Bakar drain (1.2× 103 cfu ml–1) and in intestine of fishes collected from El-Mataryia area (4.1 × 102 cfu g–1). There was a correlation between faecal coliforms and Aeromonas sppin water samples. A total of 80 isolates of water and fish samples, were obtained from Aeromonasselective agar and other different selective media and identified with API 20 E system. Strains of Acrimonies sobriawere found highly β-hemolytic on fresh human blood agar, as well as the extra cellular product of ECP and appeared to be destructive to the mammalian cells.

Antibacterial efficacy of lytic bacteriophages against antibiotic-resistant Klebsiella species.

Abstract: Bacterial resistance to antibiotics is a leading and highly prevalent problem in the treatment of infectious diseases. Bacteriophages (phages) appear to be effective and safe alternatives for the treatment of resistant infections because of their specificity for bacterial species and lack of infectivity in eukaryotic cells. The present study aimed to isolate bacteriophages against Klebsiella spp. and evaluate their efficacy against antibiotic-resistant species. Seventy-two antibiotic-resistant Klebsiella spp. were isolated from samples of patients who referred to the Ghaem Hospital (Mashhad, Iran). Lytic bacteriophages against Klebsiella spp. were isolated from wastewater of the septic tank of the same hospital. Bactericidal activity of phages against resistant Klebsiella spp. was tested in both liquid (tube method; after 1 and 24 h of incubation) and solid (double-layer agar plate method; after 24 h of incubation) phases. In each method, three different concentrations of bacteriophages (low: 107 PFU/mL) were used. Bacteriophages showed promising bactericidal activity at all assessed concentrations, regardless of the test method and duration of incubation. Overall, bactericidal effects were augmented at higher concentrations. In the tube method, higher activity was observed after 24 h of incubation compared to the 1-h incubation. The bactericidal effects were also higher in the tube method compared to the double-layer agar plate method after 24 h of incubation. The findings of the present study suggest that bacteriophages possess effective bactericidal activity against resistant Klebsiella spp. These bactericidal activities are influenced by phage concentration, duration of incubation, and test method.

Improvement of Xylanase Production from Streptomyces Pseudogriseolus via UV Mutagenesis

Abstract: 3 Abstract: Streptomyces pseudogriseolus isolated from terrestrial environment was subjected to UV irradiation for different periods producing 139 mutants. Twenty-nine mutants were considered as morphological mutants as they showed abnormality in growth, color and/or shape. Qualitative screening has been done to those mutants using agar plate medium stained with Congo red after growth for 3 days at 30 C. Ninety-four mutant exhibited clear zones more than the wild strain, 48 o mutants were selected to be tested quantitatively using submerged culture medium containing xylan as sole carbon source. Quantitatively, 16 mutants showed xylanase activity more than the wild strain. Mutant no. 121 was the promizing potent xylanase-producing mutant. It had an activity of 161% as compared to the wild strain.

Improved method to induce sporulation of Colletotrichum gloeosporioides, causal fungus of grape ripe rot

Abstract: Colletotrichumgloeosporioides and C. acutatum are causal agents of grape ripe rot, but with available methods, sporulation of C.gloeosporioides on plate media has been unstable and inferior to that of C. acutatum. To facilitate studies on C. gloeosporioides, I developed an improved method to induce conidiation of this fungus. Isolates of C. gloeosporioides were pre-cultured in potato dextrose broth for 1 week, then pulverized in whole broth. The homogenate was then spread on diluted oatmeal agar (15–20% commercial oatmeal agar medium, 1.5% agar) plates. After the plates were cultured at 25°C under continuous light for another week, the C. gloeosporioides isolates sporulated stably on the plate medium.

Extracellular Amylase Activity of Amylolytic Bacteria Isolated from Quail's (Coturnix japonica) Intestinal Tract in Corn Flour Medium

Abstract: 2 Abstract: Amylolitic microorganisms have capability of producing amylase which is an important enzyme in industries such as paper, textile and food industries. These microorganisms are found as a normal intestinal microflora in poultry's digestive tracts including quail (Coturnix japonica), especially with carbohydrates as a most nutrient in their ration. Corn becomes most important feed for quail because it has high energy related to its high amylum content. Amilolytic microorganisms in the digestive tract digest amylum by producing an extracellular enzyme (amylase) which breakdown amylum into simpler molecules facilitating for its absorbtion in the digestive tract. This research aimed to isolate amylolytic bacteria from quail's digestive tract, to determine their growth and amylolytic activity in corn flour medium. Isolation was done by using Starch Agar medium, then amylolytic activity was indicated based on intensity of clear zone formation in the media and by amylase assay using DNS method in Starch Broth medium. Growth curve and amylase assay was carried out in 2% Corn Flour Broth medium. Data were subjected to analysis of Pearson correlation. The results showed that six isolates called as BAP1, BAP2, BAP3, BAP4, BAP5 and BAP6 were found. BAP6 isolate showed the widest clear zone formation in Starch Agar medium, highest amylase activity and non pathogenic characters. Thus, it was selected and used in further analysis. BAP6 isolate grew well in 2% Corn Flour Broth medium with the highest cell number, 2.2 x 10 CFU/ml at 24 h incubation and the 8 highest amylase activity, 0.0201 Unit at 12 h incubation.