In vivo and in vitro control activity of plant essential oils against three strains of Aspergillus niger.

PubMed

Kumar, Peeyush; Mishra, Sapna; Kumar, Atul; Kumar, Sanjeev; Prasad, Chandra Shekhar

2017-09-01

Contamination of environment and food from the prevalent spores and mycotoxins of Aspergillus niger has led to several diseases in humans and other animals. The present study investigated the control activity of plant essential oils against three strains of A. niger. In the elaborate assays done through microdilution plate assay and agar disk diffusion assay in the lab condition and in vivo assay on the stored wheat grains, the essential oil of Thymus vulgaris depicted overall superior efficacy. In microdilution plate assay, the oil of Anethum graveolens showed best fungistatic activity, while best fungicidal activity was depicted by Syzygium aromaticum oil. The oil of T. vulgaris showed moderate control efficacy against A. niger strains with its antifungal activity resulting mainly due to killing of microorganism rather than growth inhibition. In agar disk diffusion assay, T. vulgaris oil with a zone of inhibition (ZOI) of 23.3-61.1% was the most effective fungicide. The in vivo assay to evaluate the protection efficacy of oils for stored wheat grains against A. niger (AN1) revealed T. vulgaris (90.5-100%) to be the best control agent, followed by the oil of S. aromaticum (61.9-100%). The GC-MS analysis of T. vulgaris oil indicated the presence of thymol (39.11%), γ-terpinene (19.73%), o-cymene (17.21%), and β-pinene (5.38%) as major oil components. Phytotoxic effects of the oils on wheat seeds showed no significant phytotoxic effect of oils in terms of seed germination or seedling growth. The results of the study demonstrated control potentiality of essential oils for the protection of stored wheat against A. niger with prospect for development of eco-friendly antifungal products.

Antifungal susceptibility testing of Malassezia yeast: comparison of two different methodologies.

PubMed

Rojas, Florencia D; Córdoba, Susana B; de Los Ángeles Sosa, María; Zalazar, Laura C; Fernández, Mariana S; Cattana, María E; Alegre, Liliana R; Carrillo-Muñoz, Alfonso J; Giusiano, Gustavo E

2017-02-01

All Malassezia species are lipophilic; thus, modifications are required in susceptibility testing methods to ensure their growth. Antifungal susceptibility of Malassezia species using agar and broth dilution methods has been studied. Currently, few tests using disc diffusion methods are being performed. The aim was to evaluate the in vitro susceptibility of Malassezia yeast against antifungal agents using broth microdilution and disc diffusion methods, then to compare both methodologies. Fifty Malassezia isolates were studied. Microdilution method was performed as described in reference document and agar diffusion test was performed using antifungal tablets and discs. To support growth, culture media were supplemented. To correlate methods, linear regression analysis and categorical agreement was determined. The strongest linear association was observed for fluconazole and miconazole. The highest agreement between both methods was observed for itraconazole and voriconazole and the lowest for amphotericin B and fluconazole. Although modifications made to disc diffusion method allowed to obtain susceptibility data for Malassezia yeast, variables cannot be associated through a linear correlation model, indicating that inhibition zone values cannot predict MIC value. According to the results, disc diffusion assay may not represent an alternative to determine antifungal susceptibility of Malassezia yeast. © 2016 Blackwell Verlag GmbH.

Antimicrobial activity of honey from the stingless bee Trigona carbonaria determined by agar diffusion, agar dilution, broth microdilution and time-kill methodology.

PubMed

Boorn, K L; Khor, Y-Y; Sweetman, E; Tan, F; Heard, T A; Hammer, K A

2010-05-01

The aim of this study was to determine the spectrum of antimicrobial activity of 11 samples of stingless bee honey compared to medicinal, table and artificial honeys. Activity was assessed by agar diffusion, agar dilution, broth microdilution and time-kill viability assays. By agar dilution, minimum inhibitory concentration (MIC) ranges were 4% to >10% (w/v) for Gram-positive bacteria, 6% to >16% (w/v) for Gram-negative bacteria and 6% to >10% (w/v) for Candida spp. By broth microdilution, all organisms with the exception of Candida albicans and Candida glabrata were inhibited at 3 log for Pseudomonas aeruginosa and <1 log for C. albicans. Similar treatment with each control honey resulted in decreases of <1 log for all organisms. Stingless bee honey has broad-spectrum antibacterial activity although activity against Candida was limited. Stingless bee honey samples varied in activity and the basis for this remains to be determined. Stingless bee honey had similar activity to medicinal honey and may therefore have a role as a medicinal agent.

Antibacterial properties of Ag-exchanged Philippine natural zeolite-chitosan composites

NASA Astrophysics Data System (ADS)

Taaca, Kathrina Lois M.; Olegario, Eleanor M.; Vasquez, Magdaleno R.

2017-12-01

Zeolites are microporous minerals composed of silicon, aluminum and oxygen. These aluminosilicates consist of tetrahedral units which produce open framework structures to generate a system of pores and cavities of molecular dimensions. Zeolites are naturally abundant and can be mined in most parts of the world. In this study, natural zeolites (NaZ) which are locally-sourced here in the Philippines were investigated to determine its properties. An ion-exchange process was utilized, using the zeolite to silver (Ag) solution ratio of 1:20 (w/v), to incorporate Ag into the zeolite framework. Characterizations such as XRD, AAS, and Agar diffusion assay were used to evaluate the properties of the synthesized Ag-exchanged zeolites (AgZ). X-ray diffraction revealed that both NaZ and AgZ have peaks mostly corresponding to the clinoptilolite structure, with some trace peaks of the mordenite and quartz. Absorption spectroscopy revealed that the ion exchange process added about 0.61188g of silver into the zeolite structure. This Ag content was seen to be enough to make the AgZ sample exhibit an antibacterial effect where clearing zones against E. coli and S. aureus were observed in the agar diffusion assay, respectively. The AgZ sample was also tested as ceramic filler to a polymer matrix-chitosan. The diffusion assay revealed presence of antibacterial activity to the polymer composite with AgZ fillers. These results indicate that the Philippine natural zeolite, incorporated with metals such as Ag, can be used as an antibacterial agent and can be developed as a ceramic filler to improve the antibacterial property of composite materials for biomedical application.

Effect of pH alkaline salts of fatty acids on the inhibition of bacteria associated with poultry processing

USDA-ARS?s Scientific Manuscript database

The agar diffusion assay was used to examine the effect of pH on the ability of alkaline salts of three fatty acids (FA) to inhibit growth of bacteria associated with poultry processing. FA solutions were prepared by dissolving 0.5 M concentrations of caprylic, capric, or lauric acid in separate ali...

Anti-Quorum Sensing Potential of Crude Kigelia africana Fruit Extracts

PubMed Central

Chenia, Hafizah Y.

2013-01-01

The increasing incidence of multidrug-resistant pathogens has stimulated the search for novel anti-virulence compounds. Although many phytochemicals show promising antimicrobial activity, their power lies in their anti-virulence properties. Thus the quorum sensing (QS) inhibitory activity of four crude Kigelia africana fruit extracts was assessed qualitatively and quantitatively using the Chromobacterium violaceum and Agrobacterium tumefaciens biosensor systems. Inhibition of QS-controlled violacein production in C. violaceum was assayed using the qualitative agar diffusion assay as well as by quantifying violacein inhibition using K. africana extracts ranging from 0.31–8.2 mg/mL. Qualitative modulation of QS activity was investigated using the agar diffusion double ring assay. All four extracts showed varying levels of anti-QS activity with zones of violacein inhibition ranging from 9–10 mm. The effect on violacein inhibition was significant in the following order: hexane > dichloromethane > ethyl acetate > methanol. Inhibition was concentration-dependent, with the ≥90% inhibition being obtained with ≥1.3 mg/mL of the hexane extract. Both LuxI and LuxR activity were affected by crude extracts suggesting that the phytochemicals target both QS signal and receptor. K. africana extracts with their anti-QS activity, have the potential to be novel therapeutic agents, which might be important in reducing virulence and pathogenicity of drug-resistant bacteria in vivo. PMID:23447012

A novel bicomponent hemolysin from Bacillus cereus.

PubMed Central

Beecher, D J; MacMillan, J D

1990-01-01

A procedure combining isoelectric focusing (Sephadex IEF) and fast protein liquid chromatography (Superose 12; Mono-Q) removed hemolytic activity (presumably a contaminant) from partially purified preparations of the multicomponent diarrheal enterotoxin produced by Bacillus cereus. However, when the separated fractions were recombined, hemolytic activity was restored, suggesting that hemolysis is a property of the enterotoxin components. Combined fractions exhibited a unique ring pattern in gel diffusion assays in blood agar. During diffusion of the hemolysin from an agar well, the erythrocytes closest to the well were not lysed initially. After diffusion, hemolysis was observed as a sharp ring beginning several millimeters away from the edge of the well. With time the cells closer to the well were also lysed. This novel hemolysin consists of a protein (component B) which binds to or alters cells, allowing subsequent lysis by a second protein (component L). Sodium dodecyl sulfate-polyacrylamide gel electrophoresis, isoelectric focusing, and Western blot analysis showed that hemolysin BL has properties similar to those described previously for the enterotoxin and that both components are distinct from cereolysin and cereolysin AB. Images PMID:2114359

A new effective assay to detect antimicrobial activity of filamentous fungi.

PubMed

Pereira, Eric; Santos, Ana; Reis, Francisca; Tavares, Rui M; Baptista, Paula; Lino-Neto, Teresa; Almeida-Aguiar, Cristina

2013-01-15

The search for new antimicrobial compounds and the optimization of production methods turn the use of antimicrobial susceptibility tests a routine. The most frequently used methods are based on agar diffusion assays or on dilution in agar or broth. For filamentous fungi, the most common antimicrobial activity detection methods comprise the co-culture of two filamentous fungal strains or the use of fungal extracts to test against single-cell microorganisms. Here we report a rapid, effective and reproducible assay to detect fungal antimicrobial activity against single-cell microorganisms. This method allows an easy way of performing a fast antimicrobial screening of actively growing fungi directly against yeast. Because it makes use of an actively growing mycelium, this bioassay also provides a way for studying the production dynamics of antimicrobial compounds by filamentous fungi. The proposed assay is less time consuming and introduces the innovation of allowing the direct detection of fungal antimicrobial properties against single cell microorganisms without the prior isolation of the active substance(s). This is particularly useful when performing large screenings for fungal antimicrobial activity. With this bioassay, antimicrobial activity of Hypholoma fasciculare against yeast species was observed for the first time. Copyright © 2012 Elsevier GmbH. All rights reserved.

The in-vitro antimicrobial activities of some medicinal plants from Cameroon.

PubMed

Gangoué-Piéboji, J; Pegnyemb, D E; Niyitegeka, D; Nsangou, A; Eze, N; Minyem, C; Mbing, J Ngo; Ngassam, P; Tih, R Ghogomu; Sodengam, B L; Bodo, B

2006-04-01

The antimicrobial activities of 10 plant species (Voacanga africana, Crepis cameroonica, Plagiostyles africana, Crotalaria retusa, Mammea africana, Lophira lanceolata, Ochna afzelii, Ouratea elongata, Ou. flava and Ou. sulcata), each of which is currently used in the traditional medicine of Cameroon, were investigated in vitro. The activities of a methanol extract of each plant were tested, in disc-diffusion assays, against 37 reference or laboratory strains of seven species of microorganism (Staphylococcus aureus, S. epidermidis, Enterococcus hirae, Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa and Candida albicans). The minimal inhibitory concentrations of each extract were then estimated, against each of the more susceptible microorganisms (i.e. those giving an inhibition zone measuring at least 9 mm in diameter in the disc-diffusion assays), by agar dilution. Although, in the disc-diffusion assays, each of the 10 methanol extracts investigated displayed some degree of antimicrobial activity against at least one species of microorganism, no activity against the Gram-negative bacteria (Es. coli, K. pneumoniae and Ps. aeruginosa) was observed. The extract with the greatest antimicrobial activity was that of Pl. africana (Euphorbiaceae).

A fixed-time diffusion analysis method determines that the three cheV genes of Helicobacter pylori differentially affect motility

PubMed Central

Lowenthal, Andrew C.; Simon, Christopher; Fair, Amber S.; Mehmood, Khalid; Terry, Karianne; Anastasia, Stephanie; Ottemann, Karen M.

2009-01-01

Helicobacter pylori is a chemotactic bacterium that has three CheV proteins in its predicted chemotaxis signal transduction system. CheV proteins contain both CheW- and response-regulator-like domains. To determine the function of these proteins, we developed a fixed-time diffusion method that would quantify bacterial direction change without needing to define particular behaviours, to deal with the many behaviours that swimming H. pylori exhibit. We then analysed mutants that had each cheV gene deleted individually and found that the behaviour of each mutant differed substantially from wild-type and the other mutants. cheV1 and cheV2 mutants displayed smooth swimming behaviour, consistent with decreased cellular CheY-P, similar to a cheW mutant. In contrast, the cheV3 mutation had the opposite effect and the mutant cells appeared to change direction frequently. Additional analysis showed that the cheV mutants displayed aberrant behaviour as compared to the wild-type in the soft-agar chemotaxis assay. The soft-agar assay phenotype was less extreme compared to that seen in the fixed-time diffusion model, suggesting that the cheV mutants are able to partially compensate for their defects under some conditions. Each cheV mutant furthermore had defects in mouse colonization that ranged from severe to modest, consistent with a role in chemotaxis. These studies thus show that the H. pylori CheV proteins each differently affect swimming behaviour. PMID:19332820

The effect of lactic acid bacteria isolates on the urinary tract pathogens to infants in vitro.

PubMed

Lim, In Seok; Lee, Ho Seok; Kim, Won Yong

2009-01-01

Urinary tract infections are common clinical problems in children, even though lots of treatment strategies have been tried. Many studies of the application of probiotics for urinary tract infection in female adults exist, but there is a lack of studies in children. The aims of this study were to screen probiotic strains for inhibiting the uropathogens in vitro, to find candidates for in vivo study. Nine strains of E. coli were isolated from children with urinary tract infection and six uropathogens were obtained from Korean Collection for Type Cultures and American Type Culture Collection. Also 135 lactic acid bacteria (LAB) strains were isolated from healthy children, and were identified through physiologic, biochemical methods, 16S rDNA PCR, and data analysis. And with agar disk diffusion assay technique the antimicrobial activities of these LAB strains against those uropathogens were examined. Three strains of separated LAB strains demonstrated major antimicrobial activity against all the uropathogens. In the agar disk diffusion assay technique, antimicrobial activities increased most in the 4th day culture broth with separated Lactobacillus. In summary, some LAB can be used as candidates to develop the probiotic microorganisms that inhibit uropathogens in children, and are expected to be applied to treatment and prevention of pediatric urinary tract infection.

Prunus mume extract exhibits antimicrobial activity against pathogenic oral bacteria.

PubMed

Seneviratne, Chamida J; Wong, Ricky W K; Hägg, Urban; Chen, Yong; Herath, Thanuja D K; Samaranayake, P Lakshman; Kao, Richard

2011-07-01

Prunus mume is a common fruit in Asia, which has been used in traditional Chinese medicine. In this study, we focused on the antimicrobial properties of Prunus mume extract against oral pathogens related to dental caries and periodontal diseases. A total of 15 oral pathogens including Streptococcus mutans, S. sobrinus, S. mitis, S. sanguinis, Lactobacillus acidophilus, P. gingivalis, Aggregatibacter actinomycetemcomitans, and Candida species were included in the study. Initially, agar diffusion assay was performed to screen the antimicrobial activities of Prunus mume extract. Minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) were then determined for sensitive species. Effect of Prunus mume extract on human oral keratinocytes (HOK) viability was also tested. In the agar diffusion assay, drug suspension of 2 g/mL was able to inhibit all the bacterial species tested, but not the fungal species. MIC and MBC range of Prunus mume extract against the oral bacteria was 0.15625-0.0003 g/mL and P. gingivalis being the most susceptible species. Prune extract did not cause any detrimental effect on HOK. Prunus mume extract may be a potential candidate for developing an oral antimicrobial agent to control or prevent dental diseases associated with oral pathogenic bacteria. © 2011 The Authors. International Journal of Paediatric Dentistry © 2011 BSPD, IAPD and Blackwell Publishing Ltd.

Antimicrobial Activity of Rhoeo discolor Phenolic Rich Extracts Determined by Flow Cytometry.

PubMed

García-Varela, Rebeca; García-García, Rebeca M; Barba-Dávila, Bertha A; Fajardo-Ramírez, Oscar R; Serna-Saldívar, Sergio O; Cardineau, Guy A

2015-10-14

Traditional medicine has led to the discovery of important active substances used in several health-related areas. Phytochemicals in Rhoeo discolor extracts have proven to have important antimicrobial activity. In the present study, our group determined the antimicrobial effects of extracts of Rhoeo discolor, a plant commonly used in Mexico for both medicinal and ornamental purposes. We evaluated the in vitro activity of phenolic rich extracts against specifically chosen microorganisms of human health importance by measuring their susceptibility via agar-disc diffusion assay and flow cytometry: Gram-positive Listeria innocua and Streptococcus mutans, Gram-negative Escherichia coli and Pseudomonas aeruginosa, and lastly a fungal pathogen Candida albicans. Ten different extracts were tested in eight different doses on all the microorganisms. Analytical data revealed a high content of phenolic compounds. Both agar-disc diffusion assay and flow cytometry results demonstrated that Pseudomonas aeruginosa was the least affected by extract exposure. However, low doses of these extracts (predominantly polar), in a range from 1 to 4 μg/mL, did produce a statistically significant bacteriostatic and bactericidal effect on the rest of the microorganisms. These results suggest the addition of certain natural extracts from Rhoeo discolor could act as antibacterial and antimycotic drugs or additives for foods and cosmetics.

Combination Studies of Oreganum Vulgare Extract Fractions and Volatile Oil along with Ciprofloxacin and Fluconazole against Common Fish Pathogens

PubMed Central

Bharti, Veni; Vasudeva, Neeru; Dhuhan, Joginder Singh

2013-01-01

Purpose: The study is aimed at finding new antibiotic therapy for aquaculture due to potential of bacteria to develop resistance to the existing therapies. Use of large quantities of synthetic antibiotics in aquaculture thus has the potential to be detrimental to fish health, to the environment and wildlife and to human health. Methods: Antimicrobial potential of volatile oil and fractions of chloroform extract of Oreganum vulgare was evaluated alone and in the presence of standard antimicrobials against common fish pathogens by disc-diffusion, agar well assay and two fold microdilution method by nanodrop spectrophotometric method. Results: The best results were represented by volatile oil followed by phenolic fraction by disc-diffusion, agar well and microdilution assays (Minimum inhibitory concentration). By the interaction studies, it was observed that the volatile oil and phenolic fraction were able to inhibit the pathogens at very low concentration compared to standard drugs. The fractional inhibitory concentration index (FICI) was calculated and volatile oil and phenolic fractions were found to be synergistic against Pseudomonas fluorescens and Candida albicans. Conclusion: The experimental data suggests the use of volatile oil and phenolic fraction in combination with standard antimicrobials to maintain healthy aquaculture with lesser adverse effects as compared to synthetic antibiotic therapy. PMID:24312842

Evaluation of the effectiveness of peracetic acid in the sterilization of dental equipment.

PubMed

Ceretta, R; Paula, M M S; Angioletto, Ev; Méier, M M; Mitellstädt, F G; Pich, C T; Junior, S A; Angioletto, E

2008-01-01

To evaluate the effectiveness of peracetic acid in the microbiological sterilisation of dental materials. Peracetic acid solution was evaluated at concentrations of 800, 1500 and 2500 ppm. At these concentrations, it was determined whether peracetic acid caused corrosion to dental instruments and induced cellular mutagenicity and cytotoxicity. In addition, the minimum inhibitory concentration (MIC), the minimum bactericidal concentration (MBC), agar diffusion and diffusion by well method, were also verified. The corrosion rate, calculated from potentiodynamic assays was 10(-6) cm/year, indicating that the product does not damage equipment. The sterilisation capacity of peracetic acid at 2500 ppm was the best. The comet assay indicated genotoxic activity at 2500 ppm. This study demonstrated the effectiveness of peracetic acid for sterilizing dental equipment, providing another alternative for the prevention of infections in clinics.

Cytotoxicity of Light-Cured Dental Materials according to Different Sample Preparation Methods

PubMed Central

Lee, Myung-Jin; Kim, Mi-Joo; Kwon, Jae-Sung; Lee, Sang-Bae; Kim, Kwang-Mahn

2017-01-01

Dental light-cured resins can undergo different degrees of polymerization when applied in vivo. When polymerization is incomplete, toxic monomers may be released into the oral cavity. The present study assessed the cytotoxicity of different materials, using sample preparation methods that mirror clinical conditions. Composite and bonding resins were used and divided into four groups according to sample preparation method: uncured; directly cured samples, which were cured after being placed on solidified agar; post-cured samples were polymerized before being placed on agar; and “removed unreacted layer” samples had their oxygen-inhibition layer removed after polymerization. Cytotoxicity was evaluated using an agar diffusion test, MTT assay, and confocal microscopy. Uncured samples were the most cytotoxic, while removed unreacted layer samples were the least cytotoxic (p < 0.05). In the MTT assay, cell viability increased significantly in every group as the concentration of the extracts decreased (p < 0.05). Extracts from post-cured and removed unreacted layer samples of bonding resin were less toxic than post-cured and removed unreacted layer samples of composite resin. Removal of the oxygen-inhibition layer resulted in the lowest cytotoxicity. Clinicians should remove unreacted monomers on the resin surface immediately after restoring teeth with light-curing resin to improve the restoration biocompatibility. PMID:28772647

Antioxidant and antibacterial activities on foodborne pathogens of Artocarpus heterophyllus Lam. (Moraceae) leaves extracts.

PubMed

Loizzo, M R; Tundis, R; Chandrika, U G; Abeysekera, A M; Menichini, F; Frega, N G

2010-06-01

Total water extract, ethyl acetate, and aqueous fractions from the leaves of Artocarpus heterophyllus were evaluated for phenolic content, antioxidant, and antibacterial activities against some foodborne pathogens such as E. coli, Listeria monocytogenes, Salmonella typhimurium, Salmonella enterica, Bacillus cereus, Enterococcus faecalis, and Staphylococcus aureus. The minimum inhibitory concentration (MICs) of extract and fractions determined by the agar dilution method were ranged from 221.9 microg/mL for ethyl acetate fraction to 488.1 microg/mL for total extract. In the agar diffusion method the diameters of inhibition were 12.2 for the total extract, 10.7 and 11.5 for ethyl acetate and aqueous fractions, respectively. A. heterophyllus showed significant antioxidant activity tested in different in vitro systems (DPPH, ABTS, FRAP, and Fe(2+) chelating activity assay). In particular, in DPPH assay A. heterophyllus total extract exhibited a strong antiradical activity with an IC(50) value of 73.5 microg/mL while aqueous fraction exerted the highest activity in FRAP assay (IC(50) value of 72.0 microg/mL). The total phenols content by Folin-Ciocalteau method was determined with the purpose of testing its relationship with the antioxidant and antibacterial activities.

One-week 96-well soft agar growth assay for cancer target validation.

PubMed

Ke, Ning; Albers, Aaron; Claassen, Gisela; Yu, De-hua; Chatterton, Jon E; Hu, Xiuyuan; Meyhack, Bernd; Wong-Staal, Flossie; Li, Qi-Xiang

2004-05-01

Soft agar growth, used to measure cell anchorage-independent proliferation potential, is one of the most important and most commonly used assays to detect cell transformation. However, the traditional soft agar assay is time-consuming, labor-intensive, and plagued with inconsistencies due to individual subjectivity. It does not, therefore, meet the increasing demands of today's oncology drug target screening or validation processes. This report describes an alternative 96-well soft agar growth assay that can function as a replacement for the traditional method and overcomes the aforementioned limitations. It offers the following advantages: a shortened assay duration (1 week instead of 4 weeks) that makes transient transfection or treatment possible; plate reader quantification of soft agar growth (measuring cloning efficiency and colony size); and a significant reduction in required labor. Higher throughput also makes it possible to process large numbers of samples and treatments simultaneously and in a much more efficient manner, while saving precious workspace and overall cost.

The radiosensitivity of a murine fibrosarcoma as measured by three cell survival assays.

PubMed Central

Rice, L.; Urano, M.; Suit, H. D.

1980-01-01

The radiation sensitivity of a weakly immunogenic spontaneous fibrosarcoma of the C3Hf/Sed mouse (designated FSa-II) was assessed by three in vivo cell survival methods: end-point dilution (TD50) assay, lung colony (LC) assay, and agar diffusion chamber (ADC) assay. The hypoxic fraction of this tumour was also determined by the ADC method. Although there was a good agreement of the cell survival data between the ADC and LC methods, the TD50 method yielded a considerably less steep cell survival curve. Beneficial aspects and limitations of each assay are discussed. In addition, the use of the ADC method for the growth of xenogeneic cell lines and a preliminary experiment with human tumour cells in non-immunosuppressed hosts suggest that this method may be a valuable adjunct for studying the growth and therapeutic responses of human tumour cells. PMID:6932931

Improved soft-agar colony assay in a fluid processing apparatus.

PubMed

Forsman, A D; Herpich, A R; Chapes, S K

1999-01-01

The standard method for quantitating bone marrow precursor cells has been to count the number of colony-forming units that form in semisolid (0.3%) agar. Recently we adapted this assay for use in hardware, the Fluid Processing Apparatus, that is flown in standard payload lockers of the space shuttle. When mouse or rat macrophage colony-forming units were measured with this hardware in ground-based assays, we found significantly more colony growth than that seen in standard plate assays. The improved growth correlates with increased agar thickness but also appears to be due to properties inherent to the Fluid Processing Apparatus. This paper describes an improved method for determining bone marrow macrophage precursor numbers in semisolid agar.

Comparison between Disk Diffusion and Microdilution Methods for Determining Susceptibility of Clinical Fungal Isolates to Caspofungin▿

PubMed Central

Milici, Maria Eleonora; Maida, Carmelo Massimo; Spreghini, Elisabetta; Ravazzolo, Barbara; Oliveri, Salvatore; Scalise, Giorgio; Barchiesi, Francesco

2007-01-01

We compared the caspofungin (CAS) susceptibility testing results generated by the disk diffusion (DD) assay with the results of the Clinical and Laboratory Standards Institute (CLSI) broth microdilution (BD) reference method for 106 yeast isolates. The isolates represented 11 different fungal species, including Candida albicans (n = 50), C. parapsilosis (n = 10), C. glabrata (n = 10), C. tropicalis (n = 10), C. guillermondii (n = 6), C. rugosa (n = 5), C. krusei (n = 5), C. kefyr (n = 2), C. pelliculosa (n = 2), Saccharomyces cerevisiae (n = 3), and Geotrichum candidum (n = 3). The DD assay was performed in supplemented Mueller-Hinton agar with CAS, which was tested at concentrations of 2, 10, and 25 μg per disk. MICs and inhibition zone diameters were evaluated at 24 and 48 h. In general, the results obtained by the DD assay correlated well with those obtained by the BD method. In particular, a significant correlation between methods was observed when CAS was used at concentration of 2 μg/disk at a reading time of either 24 or 48 h. PMID:17728477

Significant antibacterial activity and synergistic effects of camel lactoferrin with antibiotics against methicillin-resistant Staphylococcus aureus (MRSA).

PubMed

Redwan, Elrashdy M; El-Baky, Nawal Abd; Al-Hejin, Ahmed M; Baeshen, Mohammed N; Almehdar, Hussein A; Elsaway, Abdulrahman; Gomaa, Abu-Bakr M; Al-Masaudi, Saad Berki; Al-Fassi, Fahad A; AbuZeid, Isam Eldin; Uversky, Vladimir N

2016-01-01

Methicillin-resistant Staphylococcus aureus (MRSA) causes major healthcare problems in many countries, as it is present as several hospital- and community-associated strains. Hospital-associated MRSA is one of the most prevalent nosocomial pathogens throughout the world and infections caused by community-acquired MRSA are rising. This emphasizes the need for new and efficient anti-MRSA agents. We evaluated the antibacterial effects of camel lactoferrin (cLf) and human lactoferrin (hLf) alone and in combination with several antibiotics against MRSA. Antimicrobials were tested against MRSA and an S. aureus control strain by the agar disc diffusion method. The minimum inhibitory concentration (MIC) was determined for antimicrobials by the broth microdilution method. Synergy between cLf or hLf and antibiotics was examined by checkerboard and time-kill assays. The agar disc diffusion assay showed that MRSA growth was inhibited by cLf at 0.25-3 mg/ml and hLf at 1-3 mg/ml. cLf demonstrated 3 times higher inhibitory activity against MRSA than hLf in terms of MIC values (250 vs. 750 μg/ml, respectively). Biotinylated cLf was recognized by two membrane proteins of MRSA, 66-67 KDa. Combinations of cLf or hLf and oxacillin or vancomycin at sub-MIC levels enhanced in vitro antibacterial activity against MRSA compared with each agent alone. Copyright © 2016 Institut Pasteur. Published by Elsevier Masson SAS. All rights reserved.

Antibacterial properties of aged dental cements evaluated by direct-contact and agar diffusion tests.

PubMed

Lewinstein, Israel; Matalon, Shlomo; Slutzkey, Shimshon; Weiss, Ervin I

2005-04-01

Since failure of fixed partial dentures is most frequently caused by caries, it would be advantageous if cements possessed antibacterial properties. The purpose of this study was to evaluate the antibacterial properties of 3 dental cements using the direct-contact test and agar diffusion test. For the direct-contact test, wells (n = 4) of microtiter plates were coated with the tested cements (Harvard cement, Duralon, and Ketac-Cem) while Streptococcus mutans suspension was placed directly on the cements. Bacterial growth was evaluated by a temperature-controlled microplate spectrophotometer. Eight wells of bacteria without the tested cements served as the positive control. Six wells of the tested cement without bacteria served as the negative control. For the agar diffusion test, triplicate specimens of freshly mixed cements were poured into uniform wells (5 mm in diameter) punched in the agar plates inoculated with Streptococcus mutans . After incubation at 37 degrees C for 24 hours, the agar plates were examined for bacterial growth and the diameter of the halo formed in the bacterial lawn was measured. In both tests, each cement was mixed in 2 different powder/liquid ratios. For the direct-contact test, data were initially recorded after 1 hour of incubation. Additional experiments were performed on specimens that were aged for 24 hours, 1 week, 1 month, and 3 months before assessment by either direct-contact test or agar diffusion test. The data were subjected to 1-way ANOVA with the Tukey post hoc test (alpha=.05). Compared with the control group, Duralon and Harvard cements demonstrated antibacterial properties even after 3 months with the direct-contact test (P <.002), while Ketac-Cem exhibited no antibacterial properties. In the agar diffusion test, no antibacterial activity was observed for any of the tested cements. The different powder/liquid ratios had a negligible effect on the antibacterial properties of the tested cements. Within the limitations of this study, Duralon and Harvard cements possessed prolonged antibacterial properties, while Ketac-Cem exhibited no antibacterial activity. The direct-contact test may be a more suitable test than the agar diffusion test to evaluate antibacterial properties of definitive cements.

[Evaluation of four methods for detecting methicillin-resistant Staphylococcus aureus isolates from clinical specimens at a regional hospital in Mexico].

PubMed

Acosta-Pérez, Gabriel; Rodríguez-Ábrego, Gabriela; Longoria-Revilla, Ernesto; Castro-Mussot, María Eugenia

2012-01-01

To estimate the prevalence of methicillin-resistant Staphylococcus aureus (MRSA) in clinical isolates and to compare different methods for detection of MRSA in a lab with limited available personnel and resources. 140 Staphylococcus aureus strains isolated from patients in several departments were assayed for β-lactamase production, MIC-Vitek 2 oxacillin, ChromID MRSA, disk diffusion in agar for cefoxitin 30 μg and PBP2a detection. The results of conventional tests were compared with the "gold standard" PCR test for mecA gene. Cohen´s kappa index was also calculated in order to evaluate the intra assay agreement between the used methods. The found prevalence was 90.7%. Sensitivity and specificity were: disk diffusion for cefoxitin 97 and 92% respectively, MIC Vitek 2-XL 97 and 69%, ChromoID MRSA 97 and 85%, and PBP2a detection 98 and 100%. All methods are very good for detecting MRSA, choosing a method to use will depend on each laboratory infrastructure.

Naphtho[1,2-b]furan-4,5-dione is a potent anti-MRSA agent against planktonic, biofilm and intracellular bacteria.

PubMed

Yang, Shih-Chun; Yen, Feng-Lin; Wang, Pei-Wen; Aljuffali, Ibrahim A; Weng, Yi-Han; Tseng, Chih-Hua; Fang, Jia-You

2017-09-01

Naphtho[1,2-b]furan-4,5-dione (N12D) and naphtho[2,3-b]furan-4,9-dione (N23D) are furanonaphthoquinone derivatives from natural resources. We examined the antimicrobial activity of N12D and N23D against drug-resistant Staphylococcus aureus. Minimum inhibitory concentration, minimum bactericidal concentration, bacterial viability and agar diffusion assay were conducted against methicillin-resistant S. aureus (MRSA) and clinical isolates of vancomycin-resistant S. aureus. The minimum inhibitory concentration of N12D and N23D against MRSA was 4.9-9.8 and 39 μM, respectively. With regard to the agar diffusion test, the inhibition zone of the quinone compounds was threefold larger than that of oxacillin. N12D was found to inhibit MRSA biofilm thickness from 24 to 16 μm as observed by confocal microscopy. N12D showed a significant reduction of the intracellular MRSA burden without decreasing the macrophage viability. The antibacterial mechanisms of N12D may be bacterial wall/membrane damage and disturbance of gluconeogenesis and the tricarboxylic acid cycle.

Antimicrobial Activity of Lippia Species from the Brazilian Semiarid Region Traditionally Used as Antiseptic and Anti-Infective Agents

PubMed Central

Pinto, Cristiana da Purificação; Rodrigues, Velize Dias; Pinto, Fernanda da Purificação; Pinto, Renata da Purificação; Uetanabaro, Ana Paula Trovatti; Pinheiro, Carla Santos Ribeiro; Gadea, Suzana Ferreira Magalhães; Silva, Tânia Regina dos Santos; Lucchese, Angélica Maria

2013-01-01

Lippia origanoides Kunth, Lippia alnifolia Schauer, and Lippia thymoides Martius and Schauer are shrubs used in the traditional Brazilian medicine as antiseptics, as well as in the treatment of infectious diseases. This study was designed to investigate the antibacterial and antifungal activities of the methanolic extracts of these species, as new potential sources of antimicrobial drugs. The antimicrobial activity of methanolic extracts was investigated against resistant yeasts and bacteria by agar disk diffusion. Then, the MIC determination of the most active species and its fractions in hexane, dichloromethane, ethyl acetate, and water was performed. By the agar diffusion assay, all species were active against at least two microorganisms, giving evidence to support their use in the popular medicine. L. origanoides leaves exhibited the widest antimicrobial action, inhibiting the growth of two Gram-positive bacteria and two yeasts; this activity was also confirmed by the MIC evaluation. The fractionation of L. origanoides crude extracts improved the activity in spectrum and intensity. The results obtained in this study indicate that L. origanoides may be a promising alternative in the treatment of bacterial and fungal infections and in the seeking of new antimicrobial drugs. PMID:24109492

The Mediterranean red alga Asparagopsis taxiformis has antifungal activity against Aspergillus species.

PubMed

Genovese, Giuseppa; Leitner, Sandra; Minicante, Simona A; Lass-Flörl, Cornelia

2013-09-01

The red algae Asparagopsis taxiformis collected from the Straits of Messina (Italy) were screened for antifungal activity against Aspergillus species. EUCAST methodology was applied and extracts showed antifungal activity against A. fumigatus, A. terreus and A. flavus. The lowest minimum inhibitory concentrations observed were <0.15 mg ml(-1) and the highest were >5 mg ml(-1) for Aspergillus spp. tested. Agar diffusion assays confirmed antifungal activity of A. taxiformis extracts in Aspergillus species. © 2013 Blackwell Verlag GmbH.

Antibacterial activity of selected plant essential oils against Escherichia coli O157:H7.

PubMed

Burt, S A; Reinders, R D

2003-01-01

To quantify the antibacterial properties of five essential oils (EO) on a non-toxigenic strain of Escherichia coli O157:H7 in the presence and absence of a stabilizer and an emulsifier and at three different temperatures. Five EOs known to exhibit antibacterial properties were screened by disc diffusion assay and the most active were selected for further study in microdilution colorimetric assays. Oregano (Origanum vulgare) and thyme (Thymus vulgaris; light and red varieties) EO had the strongest bacteriostatic and bactericidal properties, followed by bay (Pimenta racemosa) and clove bud (Eugenia caryophyllata synonym: Syzygium aromaticum) EO. Oregano oil was colicidal at 625 microl l(-1) at 10, 20 and 37 degrees C. The addition of 0.05% (w/v) agar as stabilizer reinforced the antibacterial properties, particularly at 10 degrees C, whereas 0.25% (w/v) lecithin reduced antibacterial activity. Scanning electron micrographs showed extensive morphological changes to treated cells. Oregano and thyme EO possess significant in vitro colicidal and colistatic properties, which are exhibited in a broad temperature range and substantially improved by the addition of agar as stabilizer. Bay and clove bud EO are less active. Lecithin diminished antibacterial properties. The bactericidal concentration of oregano EO irreversibly damaged E. coli O157:H7 cells within 1 min. Oregano and light thyme EO, particularly when enhanced by agar stabilizer, may be effective in reducing the number or preventing the growth of E. coli O157:H7 in foods.

Quantitative Assessment of Antimicrobial Activity of PLGA Films Loaded with 4-Hexylresorcinol

PubMed Central

Kemme, Michael; Heinzel-Wieland, Regina

2018-01-01

Profound screening and evaluation methods for biocide-releasing polymer films are crucial for predicting applicability and therapeutic outcome of these drug delivery systems. For this purpose, we developed an agar overlay assay embedding biopolymer composite films in a seeded microbial lawn. By combining this approach with model-dependent analysis for agar diffusion, antimicrobial potency of the entrapped drug can be calculated in terms of minimum inhibitory concentrations (MICs). Thus, the topical antiseptic 4-hexylresorcinol (4-HR) was incorporated into poly(lactic-co-glycolic acid) (PLGA) films at different loadings up to 3.7 mg/cm2 surface area through a solvent casting technique. The antimicrobial activity of 4-HR released from these composite films was assessed against a panel of Gram-negative and Gram–positive bacteria, yeasts and filamentous fungi by the proposed assay. All the microbial strains tested were susceptible to PLGA-4-HR films with MIC values down to 0.4% (w/w). The presented approach serves as a reliable method in screening and quantifying the antimicrobial activity of polymer composite films. Moreover, 4-HR-loaded PLGA films are a promising biomaterial that may find future application in the biomedical and packaging sector. PMID:29324696

Simulation of Bacillus subtilis biofilm growth on agar plate by diffusion-reaction based continuum model

NASA Astrophysics Data System (ADS)

Zhang, Xianlong; Wang, Xiaoling; Nie, Kai; Li, Mingpeng; Sun, Qingping

2016-08-01

Various species of bacteria form highly organized spatially-structured aggregates known as biofilms. To understand how microenvironments impact biofilm growth dynamics, we propose a diffusion-reaction continuum model to simulate the formation of Bacillus subtilis biofilm on an agar plate. The extended finite element method combined with level set method are employed to perform the simulation, numerical results show the quantitative relationship between colony morphologies and nutrient depletion over time. Considering that the production of polysaccharide in wild-type cells may enhance biofilm spreading on the agar plate, we inoculate mutant colony incapable of producing polysaccharide to verify our results. Predictions of the glutamate source biofilm’s shape parameters agree with the experimental mutant colony better than that of glycerol source biofilm, suggesting that glutamate is rate limiting nutrient for Bacillus subtilis biofilm growth on agar plate, and the diffusion-limited is a better description to the experiment. In addition, we find that the diffusion time scale is of the same magnitude as growth process, and the common-employed quasi-steady approximation is not applicable here.

Simulation of Bacillus subtilis biofilm growth on agar plate by diffusion-reaction based continuum model.

PubMed

Zhang, Xianlong; Wang, Xiaoling; Nie, Kai; Li, Mingpeng; Sun, Qingping

2016-07-19

Various species of bacteria form highly organized spatially-structured aggregates known as biofilms. To understand how microenvironments impact biofilm growth dynamics, we propose a diffusion-reaction continuum model to simulate the formation of Bacillus subtilis biofilm on an agar plate. The extended finite element method combined with level set method are employed to perform the simulation, numerical results show the quantitative relationship between colony morphologies and nutrient depletion over time. Considering that the production of polysaccharide in wild-type cells may enhance biofilm spreading on the agar plate, we inoculate mutant colony incapable of producing polysaccharide to verify our results. Predictions of the glutamate source biofilm's shape parameters agree with the experimental mutant colony better than that of glycerol source biofilm, suggesting that glutamate is rate limiting nutrient for Bacillus subtilis biofilm growth on agar plate, and the diffusion-limited is a better description to the experiment. In addition, we find that the diffusion time scale is of the same magnitude as growth process, and the common-employed quasi-steady approximation is not applicable here.

Inhibition of norsolorinic acid accumulation to Aspergillus parasiticus by marine actinomycetes

NASA Astrophysics Data System (ADS)

Yan, Peisheng; Shi, Cuijuan; Shen, Jihong; Wang, Kai; Gao, Xiujun; Li, Ping

2014-11-01

Thirty-six strains of marine actinomycetes were isolated from a sample of marine sediment collected from the Yellow Sea and evaluated in terms of their inhibitory activity on the growth of Aspergillus parasiticus and the production of norsolorinic acid using dual culture plate assay and agar diffusion methods. Among them, three strains showed strong antifungal activity and were subsequently identified as Streptomyces sp. by 16S rRNA gene sequencing analysis. The supernatant from the fermentation of the MA01 strain was extracted sequentially with chloroform and ethyl acetate, and the activities of the extracts were determined by tip culture assay. The assay results show that both extracts inhibited mycelium growth and toxin production, and the inhibitory activities of the extracts increased as their concentrations increased. The results of this study suggest that marine actinomycetes are biologically important for the control of mycotoxins, and that these bacteria could be used as novel biopesticides against mycotoxins.

Assay for adhesion and agar invasion in S. cerevisiae.

PubMed

Guldal, Cemile G; Broach, James

2006-11-08

Yeasts are found in natural biofilms, where many microorganisms colonize surfaces. In artificial environments, such as surfaces of man-made objects, biofilms can reduce industrial productivity, destroy structures, and threaten human life. 1-3 On the other hand, harnessing the power of biofilms can help clean the environment and generate sustainable energy. 4-8 The ability of S. cerevisiae to colonize surfaces and participate in complex biofilms was mostly ignored until the rediscovery of the differentiation programs triggered by various signaling pathways and environmental cues in this organism. 9, 10 The continuing interest in using S. cerevisiae as a model organism to understand the interaction and convergence of signaling pathways, such as the Ras-PKA, Kss1 MAPK, and Hog1 osmolarity pathways, quickly placed S. cerevisiae in the junction of biofilm biology and signal transduction research. 11-20 To this end, differentiation of yeast cells into long, adhesive, pseudohyphal filaments became a convenient readout for the activation of signal transduction pathways upon various environmental changes. However, filamentation is a complex collection of phenotypes, which makes assaying for it as if it were a simple phenotype misleading. In the past decade, several assays were successfully adopted from bacterial biofilm studies to yeast research, such as MAT formation assays to measure colony spread on soft agar and crystal violet staining to quantitatively measure cell-surface adherence. 12, 21 However, there has been some confusion in assays developed to qualitatively assess the adhesive and invasive phenotypes of yeast in agar. Here, we present a simple and reliable method for assessing the adhesive and invasive quality of yeast strains with easy-to-understand steps to isolate the adhesion assessment from invasion assessment. Our method, adopted from previous studies, 10, 16 involves growing cells in liquid media and plating on differential nutrient conditions for growth of large spots, which we then wash with water to assess adhesion and rub cells completely off the agar surface to assess invasion into the agar. We eliminate the need for streaking cells onto agar, which affects the invasion of cells into the agar. In general, we observed that haploid strains that invade agar are always adhesive, yet not all adhesive strains can invade agar medium. Our approach can be used in conjunction with other assays to carefully dissect the differentiation steps and requirements of yeast signal transduction, differentiation, quorum sensing, and biofilm formation.

Assay for Adhesion and Agar Invasion in S. cerevisiae

PubMed Central

Guldal, Cemile G; Broach, James

2006-01-01

Yeasts are found in natural biofilms, where many microorganisms colonize surfaces. In artificial environments, such as surfaces of man-made objects, biofilms can reduce industrial productivity, destroy structures, and threaten human life. 1-3 On the other hand, harnessing the power of biofilms can help clean the environment and generate sustainable energy. 4-8 The ability of S. cerevisiae to colonize surfaces and participate in complex biofilms was mostly ignored until the rediscovery of the differentiation programs triggered by various signaling pathways and environmental cues in this organism. 9, 10 The continuing interest in using S. cerevisiae as a model organism to understand the interaction and convergence of signaling pathways, such as the Ras-PKA, Kss1 MAPK, and Hog1 osmolarity pathways, quickly placed S. cerevisiae in the junction of biofilm biology and signal transduction research. 11-20 To this end, differentiation of yeast cells into long, adhesive, pseudohyphal filaments became a convenient readout for the activation of signal transduction pathways upon various environmental changes. However, filamentation is a complex collection of phenotypes, which makes assaying for it as if it were a simple phenotype misleading. In the past decade, several assays were successfully adopted from bacterial biofilm studies to yeast research, such as MAT formation assays to measure colony spread on soft agar and crystal violet staining to quantitatively measure cell-surface adherence. 12, 21 However, there has been some confusion in assays developed to qualitatively assess the adhesive and invasive phenotypes of yeast in agar. Here, we present a simple and reliable method for assessing the adhesive and invasive quality of yeast strains with easy-to-understand steps to isolate the adhesion assessment from invasion assessment. Our method, adopted from previous studies, 10, 16 involves growing cells in liquid media and plating on differential nutrient conditions for growth of large spots, which we then wash with water to assess adhesion and rub cells completely off the agar surface to assess invasion into the agar. We eliminate the need for streaking cells onto agar, which affects the invasion of cells into the agar. In general, we observed that haploid strains that invade agar are always adhesive, yet not all adhesive strains can invade agar medium. Our approach can be used in conjunction with other assays to carefully dissect the differentiation steps and requirements of yeast signal transduction, differentiation, quorum sensing, and biofilm formation. PMID:18704175

Biologically active traditional medicinal herbs from Balochistan, Pakistan.

PubMed

Zaidi, Mudassir A; Crow, Sidney A

2005-01-04

The biological activities of the following four important medicinal plants of Balochistan, Pakistan were checked; Grewia erythraea Schwein f. (Tiliaceae), Hymenocrater sessilifolius Fisch. and C.A. Mey (Lamiaceae), Vincetoxicum stocksii Ali and Khatoon (Asclepiadaceae) and Zygophyllum fabago L. (Zygophyllaceae). The methanolic extracts were fractionated into hexane, ethyl acetate, chloroform, butanol and water. The antifungal and antibacterial activities of these plants were determined against 12 fungal and 12 bacterial strains by agar well diffusion and disk diffusion assays. The extract of Zygophyllum fabago was found to be highly effective against Candida albicans and Escherichia coli. The extract of Vincetoxicum stocksii was also found to be significantly active against Candida albicans, Bacillus subtilis and Bacillus cereus. Extracts of Hymenocrater sessilifolius and Grewia erythraea showed good activity only against Pseudomonas aeruginosa.

Application of firefly luciferase assay for adenosine triphosphate (ATP) to antimicrobial drug sensitivity testing

NASA Technical Reports Server (NTRS)

Picciolo, G. L.; Tuttle, S. A.; Schrock, C. G.; Deming, J. W.; Barza, M. J.; Wienstein, L.; Chappelle, E. W.

1977-01-01

The development of a rapid method for determining microbial susceptibilities to antibiotics using the firefly luciferase assay for adenosine triphosphate (ATP) is documented. The reduction of bacterial ATP by an antimicrobial agent was determined to be a valid measure of drug effect in most cases. The effect of 12 antibiotics on 8 different bacterial species gave a 94 percent correlation with the standard Kirby-Buer-Agar disc diffusion method. A 93 percent correlation was obtained when the ATP assay method was applied directly to 50 urine specimens from patients with urinary tract infections. Urine samples were centrifuged first to that bacterial pellets could be suspended in broth. No primary isolation or subculturing was required. Mixed cultures in which one species was predominant gave accurate results for the most abundant organism. Since the method is based on an increase in bacterial ATP with time, the presence of leukocytes did not interfere with the interpretation of results. Both the incubation procedure and the ATP assays are compatible with automation.

Biodegradation and bioconversion of coals by fungi: Technical report No. 6 for the period December 1, 1986-February 28, 1987. [Candida sp. , Cunninghamella sp. , Acremonium sp. , and Trametes versicolor

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ward, B.

We continued our tests for biosolubilization activity on lignite, subbituminous, and bituminous coals, using both surface solubilization and agar diffusion assays for selected lignite-solubilizing strains challenged with untreated and treated coals shown in earlier tests to be resistant to biosolubilization. These latter were subjected to treatments with heat or pressurized oxygen atmosphere, or to combinations of the same. We have observed biosolubilization of some weathered high-rank coals by some of the fungal isolates tested. 1 tab.

Standard Nutrient Agar 1 as a substitute for blood-supplemented Müller-Hinton agar for antibiograms in developing countries.

PubMed

Niederstebruch, N; Sixt, D

2013-02-01

In the industrial world, the agar diffusion test is a standard procedure for the susceptibility testing of bacteria isolates. Beta-hemolytic Streptococcus spp. are tested with Müller-Hinton agar supplemented with 5 % blood, a so-called blood agar. The results are interpreted using standardized tables, which only exist for this type of nutrient matrix. Because of a number difficulties, both with respect to technical issues and to manual skills, blood agar is not a feasible option in many developing countries. Beta-hemolytic Streptococcus spp. also grow on Standard Nutrient Agar 1 (StNA1). This suggests using that type of nutrient medium for running agar diffusion tests. However, there are no standardized tables that can be used for interpreting the diameters of the zones of inhibition on StNA1 1. Using the existing standardized tables for blood agar to interpret cultures on StNA1 1 would be of great benefit under such circumstances where blood agar is not available. With this in mind, we conducted comparative tests to evaluate the growth characteristics of beta-hemolytic Streptococcus spp. on StNA1 1 compared to Müller-Hinton agar supplemented with 5 % sheep blood. In this study, we were able to show that beta-hemolytic Streptococcus spp. develop similar zones of inhibition on blood agar and on StNA1 1. Therefore, it is suggested that, for the interpretation of antibiograms of beta-hemolytic Streptococcus spp. performed on StNA1 1, the standard tables for blood agar can be used.

Potential of Zimbabwean commercial probiotic products and strains of Lactobacillus plantarum as prophylaxis and therapy against diarrhoea caused by Escherichia coli in children.

PubMed

Chingwaru, Walter; Vidmar, Jerneja

2017-01-01

To evaluate the potential of commercial fermented products sold in the country, and strains of Lactobacillus plantarum (L. plantarum) as prophylaxis and therapy against diarrhoea in children. The antimicrobial potential of cultures of lactobacilli enriched from 4 Zimbabwean commercial food/beverage products: Dairibord Lacto sour milk (DLSM), Probrand sour milk (PSM), Kefalos Vuka cheese (KVC) and Chibuku opaque beer (COB); and four strains of L. plantarum obtained from Balkan traditional cheeses against clinical strains of Escherichia coli (E. coli) was assayed using the well diffusion method. Three commercial paediatric antidiarrhoeal drug products: Biogaia (BG), Prolife (PL) and Probio Junior (PJ) and a mutant strain of E. coli [strain 11105 (ATCC) - a vitamin B-12 auxotroph and penicillin G acylase-producing strain] were used as controls. An agar diffusion assay and a competitive exclusion assay were carried out on Mueller Hinton agar. Crude cultures of putative lactobacillus strains obtained from Zimbabwean dairy products (Probrand sour milk, Kefalos Vuka vuka cheese and Chibuku opaque beer) had significantly higher antimicrobial activities against clinical strains of E. coli than strains of L. plantarum isolated from Balkan cheeses (CLP1, CLP2 or CLP3) and crude microbial cultures from commercial paediatric probiotic products (BG, PJ and PL) of a culture of Lactobacillus rhamnosus LGG (P < 0.05). The putative Lactobacilli from four commercial Zimbabwean dairy products (Probrand sour milk, Kefalos Vuka vuka cheese and Chibuku opaque beer), and three strains of L. plantarum from Balkan cheeses (CLP1, CLP2 or CLP3) exhibited high antibacterial activities that can be harnessed to control paediatric diarrhoea that is caused by pathogenic strains of E. coli. Studies to characterise the probiotic potential of the live cultures in the products and the new strains of L. plantarum are underway. Copyright © 2017 Hainan Medical University. Production and hosting by Elsevier B.V. All rights reserved.

Can Ceftazidime-Avibactam and Aztreonam Overcome β-Lactam Resistance Conferred by Metallo-β-Lactamases in Enterobacteriaceae?

PubMed

Marshall, Steven; Hujer, Andrea M; Rojas, Laura J; Papp-Wallace, Krisztina M; Humphries, Romney M; Spellberg, Brad; Hujer, Kristine M; Marshall, Emma K; Rudin, Susan D; Perez, Federico; Wilson, Brigid M; Wasserman, Ronald B; Chikowski, Linda; Paterson, David L; Vila, Alejandro J; van Duin, David; Kreiswirth, Barry N; Chambers, Henry F; Fowler, Vance G; Jacobs, Michael R; Pulse, Mark E; Weiss, William J; Bonomo, Robert A

2017-04-01

Based upon knowledge of the hydrolytic profile of major β-lactamases found in Gram-negative bacteria, we tested the efficacy of the combination of ceftazidime-avibactam (CAZ-AVI) with aztreonam (ATM) against carbapenem-resistant enteric bacteria possessing metallo-β-lactamases (MBLs). Disk diffusion and agar-based antimicrobial susceptibility testing were initially performed to determine the in vitro efficacy of a unique combination of CAZ-AVI and ATM against 21 representative Enterobacteriaceae isolates with a complex molecular background that included bla IMP , bla NDM , bla OXA-48 , bla CTX-M , bla AmpC , and combinations thereof. Time-kill assays were conducted, and the in vivo efficacy of this combination was assessed in a murine neutropenic thigh infection model. By disk diffusion assay, all 21 isolates were resistant to CAZ-AVI alone, and 19/21 were resistant to ATM. The in vitro activity of CAZ-AVI in combination with ATM against diverse Enterobacteriaceae possessing MBLs was demonstrated in 17/21 isolates, where the zone of inhibition was ≥21 mm. All isolates demonstrated a reduction in CAZ-AVI agar dilution MICs with the addition of ATM. At 2 h, time-kill assays demonstrated a ≥4-log 10 -CFU decrease for all groups that had CAZ-AVI with ATM (8 μg/ml) added, compared to the group treated with CAZ-AVI alone. In the murine neutropenic thigh infection model, an almost 4-log 10 -CFU reduction was noted at 24 h for CAZ-AVI (32 mg/kg every 8 h [q8h]) plus ATM (32 mg/kg q8h) versus CAZ-AVI (32 mg/kg q8h) alone. The data presented herein require us to carefully consider this new therapeutic combination to treat infections caused by MBL-producing Enterobacteriaceae . Copyright © 2017 American Society for Microbiology.

Comparative evaluation of three chromogenic media combined with broth enrichment and the real-time PCR-based Xpert MRSA assay for screening of methicillin-resistant Staphylococcus aureus in nasal swabs.

PubMed

Lee, Seungok; Park, Yeon-Joon; Park, Kang-Gyun; Jekarl, Dong Wook; Chae, Hyojin; Yoo, Jin-Kyung; Seo, Sin Won; Choi, Jung Eun; Lim, Jung Hye; Heo, Seon Mi; Seo, Ju Hee

2013-07-01

We evaluated the performance of three chromogenic media (Brilliance agar I [Oxoid, UK], Brilliance agar II [Oxoid], and ChromID MRSA [Biomérieux, France]) combined with broth enrichment and the Xpert MRSA assay for screening of methicillin-resistant Staphylococcus aureus (MRSA). We obtained 401 pairs of duplicate nasal swabs from 321 patients. One swab was suspended overnight in tryptic soy broth; 50-µL aliquots of suspension were inoculated on the three chromogenic media. Brilliance agar I and II were examined after 24 hr, and ChromID MRSA, after 24 and 48 hr. The paired swab was processed directly using real-time PCR-based Xpert MRSA assay. True positives, designated as MRSA growth in any of the culture media, were detected with the prevalence of 17% in our institution. We report the sensitivity, specificity, positive predictive value, and negative predictive value of MRSA growth as follows: 92.3%, 94.0%, 75.9%, and 98.4% in Brilliance agar I (24 hr); 92.7%, 97.9%, 90.0%, and 98.5% in Brilliance agar II (24 hr); 95.6%, 95.8%, 82.3%, and 99.1% in ChromID MRSA (24 hr); 100%, 92.5%, 73.1%, and 100% in ChromID MRSA (48 hr); 92.6%, 96.7%, 85.1%, and 98.5% in Xpert MRSA assay. The agreement between the enriched culture and Xpert MRSA assay was 96.0%. Three chromogenic culture media combined with enrichment and Xpert MRSA assay demonstrated similar capabilities in MRSA detection. The Xpert MRSA assay yielded results comparable to those of culture methods, saving 48-72 hr, thus facilitating earlier detection of MRSA in healthcare settings.

A supplemented soft agar chemotaxis assay demonstrates the Helicobacter pylori chemotactic response to zinc and nickel

PubMed Central

Sanders, Lisa; Andermann, Tessa M.

2013-01-01

Directed motility, or chemotaxis, is required for Helicobacter pylori to establish infection in the stomach, although the full repertoire of this bacterium’s chemotactic responses is not yet known. Here we report that H. pylori responds to zinc as an attractant and nickel as a repellent. To reach this conclusion, we employed both a temporal chemotaxis assay based on bacterial reversals and a supplemented soft agar spatial assay. We refined the temporal assay using a previously described chemorepellent, acid, and found that H. pylori requires rich media with serum to maintain optimal swimming motility. Surprisingly, we found that some strains respond to acid as an attractant, and that the TlpC chemoreceptor correlated with whether acid was sensed as an attractant or repellent. Using this same assay, we detected weak repellent responses to nickel and copper, and a varied response to zinc. We thus developed an alternative spatial chemotactic assay called the supplemented soft agar assay, which utilizes soft agar medium supplemented with the test compound. With Escherichia coli, the attractant serine slowed overall bacterial migration, while the repellent nickel increased the speed of overall migration. In H. pylori we detected slowed migration with doubled tryptone media, as well as zinc, consistent with an attractant response. In contrast, nickel increased migration, consistent with repulsion. PMID:23139399

Relative diffusion of paramagnetic metal complexes of MRI contrast agents in an isotropic hydrogel medium.

PubMed

Weerakoon, Bimali Sanjeevani; Osuga, Toshiaki

2017-03-01

The observation of molecular diffusion by means of magnetic resonance imaging (MRI) is significant in the evaluation of the metabolic activity of living tissues. Series of MRI examinations were conducted on a diffusion model to study the behaviour of the diffusion process of different-molecular-weight (MW) paramagnetic MRI contrast agents in an isotropic agar hydrogel medium. The model consisted of a solidified 1 % agar gel with an initial concentration of 0.5 mmol/L contrast solution layered on top of the gel. The diffusion process was monitored at pre-determined time intervals of immediately, 1, 6, 9, 23, and 48 h after introduction of the contrast agents onto the agar gel with a T1-weighted spin-echo (SE) pulse sequence. Three types of paramagnetic contrast agents, Gd-DTPA with a MW of 547.57 g/mol, Prohance with a MW of 558.69 g/mol and MnCl 2 with a MW of 125.84 g/mol, resulted in an approximate average diffusional displacement ratio of 1:1:2 per hour, respectively, within 48 h of the experiment. Therefore, the results of this study supported the hypothesis that the rate of the diffusion process of MRI contrast agents in the agar hydrogel medium is inversely related to their MWs. However, more repetitions are necessary under various types of experimental conditions and also with various types of contrast media of different MWs for further confirmation and validation of these results.

Antibacterial activities of extracts from Ugandan medicinal plants used for oral care.

PubMed

Ocheng, Francis; Bwanga, Freddie; Joloba, Moses; Borg-Karlson, Ann-Karin; Gustafsson, Anders; Obua, Celestino

2014-08-08

Medicinal plants are widely used for treatment of oral/dental diseases in Uganda. To investigate antibacterial activities of 16 commonly used medicinal plants on microorganisms associated with periodontal diseases (PD) and dental caries (DC). Pulp juice and solvent extracts (hexane, methanol and water) from the plants were tested against Aggregatibacter actinomycetemcomitans, Porphyromonas gingivalis, Tannerella forsythia associated with PD and Streptococcus mutans, Streptococcus sobrinus, Lactobacillus acidophilus associated with DC. Tests were done using agar well-diffusion (pulp juice) and agar-dilution (Solvent extracts) assays. Pulp juice from Zanthoxylum chalybeum and Euclea latidens showed activity against all the bacteria, Zanthoxylum chalybeum being most active. Hexane extract from aerial part of Helichrysum odoratissimum was most active (MIC: 0.125-0.5 mg/ml). Methanol extract from leaves of Lantana trifolia showed activity against all bacteria (MIC: 0.25-1 mg/ml). Several of the tested plants showed antibacterial activities against bacteria associated with PD and DC, meriting further investigations. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

Methicillin-Resistant Staphylococcus aureus (MRSA) Detection: Comparison of Two Molecular Methods (IDI-MRSA PCR Assay and GenoType MRSA Direct PCR Assay) with Three Selective MRSA Agars (MRSA ID, MRSASelect, and CHROMagar MRSA) for Use with Infection-Control Swabs▿

PubMed Central

van Hal, S. J.; Stark, D.; Lockwood, B.; Marriott, D.; Harkness, J.

2007-01-01

Methicillin-resistant Staphylococcus aureus (MRSA) is an increasing problem. Rapid detection of MRSA-colonized patients has the potential to limit spread of the organism. We evaluated the sensitivities and specificities of MRSA detection by two molecular methods (IDI-MRSA PCR assay and GenoType MRSA Direct PCR assay) and three selective MRSA agars (MRSA ID, MRSASelect, and CHROMagar MRSA), using 205 (101 nasal, 52 groin, and 52 axillary samples) samples from consecutive known MRSA-infected and/or -colonized patients. All detection methods had higher MRSA detection rates for nasal swabs than for axillary and groin swabs. Detection of MRSA by IDI-MRSA was the most sensitive method, independent of the site (94% for nasal samples, 80% for nonnasal samples, and 90% overall). The sensitivities of the GenoType MRSA Direct assay and the MRSA ID, MRSASelect, and CHROMagar MRSA agars with nasal swabs were 70%, 72%, 68%, and 75%, respectively. All detection methods had high specificities (95 to 99%), independent of the swab site. Extended incubation for a further 24 h with selective MRSA agars increased the detection of MRSA, with a corresponding decline in specificity secondary to a significant increase in false-positive results. There was a noticeable difference in test performance of the GenoType MRSA Direct assay in detection of MRSA (28/38 samples [74%]) compared with detection of nonmultiresistant MRSA (17/31 samples [55%]) (susceptible to two or more non-β-lactam antibiotics). This was not observed with selective MRSA agar plates or IDI-MRSA. Although it is more expensive, in addition to rapid turnaround times of 2 to 4 h, IDI-MRSA offers greater detection of MRSA colonization, independent of the swab site, than do conventional selective agars and GenoType MRSA Direct. PMID:17537949

Antibacterial activity and effects of Colla corii asini on Salmonella typhimurium invasion in vitro and in vivo.

PubMed

Park, Kwang-Il; Lee, Mi-Ra; Oh, Tae-Woo; Kim, Kwang-Youn; Ma, Jin-Yeul

2017-12-04

Salmonella enterica serovar Typhimurium is a foodborne pathogen that triggers inflammatory responses in the intestines of humans and livestock. Colla corii asini is a traditional medicine used to treat gynecologic and chronic diseases in Korea and China. However, the antibacterial activity of Colla corii asini has been unknown. In this study, we investigated the antibacterial activity and effects of Colla corii asini extract on Salmonella typhimurium invasion. To tested for antibacterial effects of Colla corii asini extracts, we confirmed the agar diffusion using Luria solid broth medium. Also, we determined the MIC (minimum inhibitory concentration) and the MBC (minimum bactericidal concentration) value of the Colla corii asini ethanol extract (CEE) by using two-fold serial dilution methods. We evaluated the expression of salmonella invasion proteins including SipA, SipB and SipC by using Western blot and qPCR at the concentration of CEE without inhibition of bacterial growth. In vitro and vivo, we determined the inhibitory effect of invasion of S. typhimurium on CEE by using gentamicin assay and S. typhimurium-infected mice. CEE significantly inhibited the growth of Salmonella typhimurium in an agar diffuse assay and had an MIC of 0.78 mg/ml and an MBC of 1.56 mg/ml. Additionally, CEE reduced Salmonella typhimurium cell invasion via the inhibition of Salmonella typhimurium invasion proteins, such as SipA, SipB and SipC. Furthermore, CEE significantly suppressed invasion in the small intestines (ilea) of mice injected with Salmonella typhimurium. These findings show that Colla corii asini exerts antibacterial activity and suppresses Salmonella typhimurium invasion in vitro and in vivo. Together, these findings demonstrate that Colla corii asini is a potentially useful therapeutic herbal medicine for treating salmonella-mediated diseases.

Direct and transdentinal (indirect) antibacterial activity of commercially available dental gel formulations against Streptococcus mutans.

PubMed

Tüzüner, Tamer; Ulusoy, Ayça Tuba; Baygin, Ozgul; Yahyaoglu, Gorkem; Yalcin, Ilkay; Buruk, Kurtulus; Nicholson, John

2013-01-01

To evaluate the direct and transdentinal (indirect) agar diffusion antibacterial activity of different commercially available antibacterial dental gel formulations against Streptococcus mutans. The commercially available dental gel formulations were Corsodyl® (COG, 1% chlorhexidine), Cervitec® (CEG, 0.2% chlorhexidine + 0.2% sodium fluoride), Forever Bright® (FOB, aloe vera), Gengigel® (GEG, 0.2% hyaluronic acid), 35% phosphoric acid gel and distilled water (control). Direct agar diffusion was performed by isolating three wells from brain-heart infusion agar plates using sterile glass pipettes attached to a vacuum pump and adding 0.1 ml of the gels to each well. Transdentinal (indirect) agar diffusion was performed by applying gel to 0.2- and 0.5-mm-thick human dentin discs previously etched with phosphoric acid and rinsed with distilled water. Zones formed around the wells and the dentin discs were measured and analyzed using Kruskal-Wallis and Mann-Whitney U tests with Bonferroni correction (p < 0.01). Direct agar diffusion tests showed significant differences among all gel formulations (p < 0.01) except for COG and CEG (p > 0.01). COG and CEG exhibited higher antibacterial effects compared to FOB and GEG (p < 0.01) in both direct and transdentinal (indirect) testing procedures. GEG did not show any antimicrobial activity in transdentinal (indirect) testing. Commercially available dental gels inhibited S. mutans, which may indicate their potential as cavity disinfectants. Copyright © 2013 S. Karger AG, Basel.

Production and Characterization of Antifungal Compounds Produced by Lactobacillus plantarum IMAU10014

PubMed Central

Wang, HaiKuan; Yan, YanHua; Wang, JiaMing; Zhang, HePing; Qi, Wei

2012-01-01

Lactobacillus plantarum IMAU10014 was isolated from koumiss that produces a broad spectrum of antifungal compounds, all of which were active against plant pathogenic fungi in an agar plate assay. Two major antifungal compounds were extracted from the cell-free supernatant broth of L. plantarum IMAU10014. 3-phenyllactic acid and Benzeneacetic acid, 2-propenyl ester were carried out by HPLC, LC-MS, GC-MS, NMR analysis. It is the first report that lactic acid bacteria produce antifungal Benzeneacetic acid, 2-propenyl ester. Of these, the antifungal products also have a broad spectrum of antifungal activity, namely against Botrytis cinerea, Glomerella cingulate, Phytophthora drechsleri Tucker, Penicillium citrinum, Penicillium digitatum and Fusarium oxysporum, which was identified by the overlay and well-diffusion assay. F. oxysporum, P. citrinum and P. drechsleri Tucker were the most sensitive among molds. PMID:22276116

Fabrication, Characterization, and Evaluation of Bionanocomposites Based on Natural Polymers and Antibiotics for Wound Healing Applications.

PubMed

Rădulescu, Marius; Holban, Alina Maria; Mogoantă, Laurențiu; Bălşeanu, Tudor-Adrian; Mogoșanu, George Dan; Savu, Diana; Popescu, Roxana Cristina; Fufă, Oana; Grumezescu, Alexandru Mihai; Bezirtzoglou, Eugenia; Lazar, Veronica; Chifiriuc, Mariana Carmen

2016-06-10

The aim of our research activity was to obtain a biocompatible nanostructured composite based on naturally derived biopolymers (chitin and sodium alginate) loaded with commercial antibiotics (either Cefuroxime or Cefepime) with dual functions, namely promoting wound healing and assuring the local delivery of the loaded antibiotic. Compositional, structural, and morphological evaluations were performed by using the thermogravimetric analysis (TGA), scanning electron microscopy (SEM), and fourier transform infrared spectroscopy (FTIR) analytical techniques. In order to quantitatively and qualitatively evaluate the biocompatibility of the obtained composites, we performed the tetrazolium-salt (MTT) and agar diffusion in vitro assays on the L929 cell line. The evaluation of antimicrobial potential was evaluated by the viable cell count assay on strains belonging to two clinically relevant bacterial species (i.e., Escherichia coli and Staphylococcus aureus).

Phytochemical Screening and Antimicrobial Activity of Some Medicinal Plants Against Multi-drug Resistant Bacteria from Clinical Isolates

PubMed Central

Dahiya, Praveen; Purkayastha, Sharmishtha

2012-01-01

The in vitro antibacterial activity of various solvents and water extracts of aloe vera, neem, bryophyllum, lemongrass, tulsi, oregano, rosemary and thyme was assessed on 10 multi-drug resistant clinical isolates from both Gram-positive and Gram-negative bacteria and two standard strains including Staphylococcus aureus ATCC 25923 and Escherichia coli ATCC 25922. The zone of inhibition as determined by agar well diffusion method varied with the plant extract, the solvent used for extraction, and the organism tested. Klebsiella pneumoniae 2, Escherichia coli 3 and Staphylococcus aureus 3 were resistant to the plant extracts tested. Moreover, water extracts did not restrain the growth of any tested bacteria. Ethanol and methanol extracts were found to be more potent being capable of exerting significant inhibitory activities against majority of the bacteria investigated. Staphylococcus aureus 1 was the most inhibited bacterial isolate with 24 extracts (60%) inhibiting its growth whereas Escherichia coli 2 exhibited strong resistance being inhibited by only 11 extracts (28%). The results obtained in the agar diffusion plates were in fair correlation with that obtained in the minimum inhibitory concentration tests. The minimum inhibitory concentration of tulsi, oregano, rosemary and aloe vera extracts was found in the range of 1.56-6.25 mg/ml for the multi-drug resistant Staphylococcus aureus isolates tested whereas higher values (6.25-25 mg/ml) were obtained against the multi-drug resistant isolates Klebsiella pneumoniae 1 and Escherichia coli 1 and 2. Qualitative phytochemical analysis demonstrated the presence of tannins and saponins in all plants tested. Thin layer chromatography and bioautography agar overlay assay of ethanol extracts of neem, tulsi and aloe vera indicated flavonoids and tannins as major active compounds against methicillin-resistant Staphylococcus aureus. PMID:23716873

Alternative to the soft-agar assay that permits high-throughput drug and genetic screens for cellular transformation

PubMed Central

Rotem, Asaf; Janzer, Andreas; Izar, Benjamin; Ji, Zhe; Doench, John G.; Garraway, Levi A.; Struhl, Kevin

2015-01-01

Colony formation in soft agar is the gold-standard assay for cellular transformation in vitro, but it is unsuited for high-throughput screening. Here, we describe an assay for cellular transformation that involves growth in low attachment (GILA) conditions and is strongly correlated with the soft-agar assay. Using GILA, we describe high-throughput screens for drugs and genes that selectively inhibit or increase transformation, but not proliferation. Such molecules are unlikely to be found through conventional drug screening, and they include kinase inhibitors and drugs for noncancer diseases. In addition to known oncogenes, the genetic screen identifies genes that contribute to cellular transformation. Lastly, we demonstrate the ability of Food and Drug Administration-approved noncancer drugs to selectively kill ovarian cancer cells derived from patients with chemotherapy-resistant disease, suggesting this approach may provide useful information for personalized cancer treatment. PMID:25902495

Alternative to the soft-agar assay that permits high-throughput drug and genetic screens for cellular transformation.

PubMed

Rotem, Asaf; Janzer, Andreas; Izar, Benjamin; Ji, Zhe; Doench, John G; Garraway, Levi A; Struhl, Kevin

2015-05-05

Colony formation in soft agar is the gold-standard assay for cellular transformation in vitro, but it is unsuited for high-throughput screening. Here, we describe an assay for cellular transformation that involves growth in low attachment (GILA) conditions and is strongly correlated with the soft-agar assay. Using GILA, we describe high-throughput screens for drugs and genes that selectively inhibit or increase transformation, but not proliferation. Such molecules are unlikely to be found through conventional drug screening, and they include kinase inhibitors and drugs for noncancer diseases. In addition to known oncogenes, the genetic screen identifies genes that contribute to cellular transformation. Lastly, we demonstrate the ability of Food and Drug Administration-approved noncancer drugs to selectively kill ovarian cancer cells derived from patients with chemotherapy-resistant disease, suggesting this approach may provide useful information for personalized cancer treatment.

Comparison of Agar Dilution, Disk Diffusion, MicroScan, and Vitek Antimicrobial Susceptibility Testing Methods to Broth Microdilution for Detection of Fluoroquinolone-Resistant Isolates of the Family Enterobacteriaceae

PubMed Central

Steward, Christine D.; Stocker, Sheila A.; Swenson, Jana M.; O’Hara, Caroline M.; Edwards, Jonathan R.; Gaynes, Robert P.; McGowan, John E.; Tenover, Fred C.

1999-01-01

Fluoroquinolone resistance appears to be increasing in many species of bacteria, particularly in those causing nosocomial infections. However, the accuracy of some antimicrobial susceptibility testing methods for detecting fluoroquinolone resistance remains uncertain. Therefore, we compared the accuracy of the results of agar dilution, disk diffusion, MicroScan Walk Away Neg Combo 15 conventional panels, and Vitek GNS-F7 cards to the accuracy of the results of the broth microdilution reference method for detection of ciprofloxacin and ofloxacin resistance in 195 clinical isolates of the family Enterobacteriaceae collected from six U.S. hospitals for a national surveillance project (Project ICARE [Intensive Care Antimicrobial Resistance Epidemiology]). For ciprofloxacin, very major error rates were 0% (disk diffusion and MicroScan), 0.9% (agar dilution), and 2.7% (Vitek), while major error rates ranged from 0% (agar dilution) to 3.7% (MicroScan and Vitek). Minor error rates ranged from 12.3% (agar dilution) to 20.5% (MicroScan). For ofloxacin, no very major errors were observed, and major errors were noted only with MicroScan (3.7% major error rate). Minor error rates ranged from 8.2% (agar dilution) to 18.5% (Vitek). Minor errors for all methods were substantially reduced when results with MICs within ±1 dilution of the broth microdilution reference MIC were excluded from analysis. However, the high number of minor errors by all test systems remains a concern. PMID:9986809

Evaluating the Effect of Peptoid Lipophilicity on Antimicrobial Potency, Cytotoxicity, and Combinatorial Library Design.

PubMed

Turkett, Jeremy A; Bicker, Kevin L

2017-04-10

Growing prevalence of antibiotic resistant bacterial infections necessitates novel antimicrobials, which could be rapidly identified from combinatorial libraries. We report the use of the peptoid library agar diffusion (PLAD) assay to screen peptoid libraries against the ESKAPE pathogens, including the optimization of assay conditions for each pathogen. Work presented here focuses on the tailoring of combinatorial peptoid library design through a detailed study of how peptoid lipophilicity relates to antibacterial potency and mammalian cell toxicity. The information gleaned from this optimization was then applied using the aforementioned screening method to examine the relative potency of peptoid libraries against Staphylococcus aureus, Acinetobacter baumannii, and Enterococcus faecalis prior to and following functionalization with long alkyl tails. The data indicate that overall peptoid hydrophobicity and not simply alkyl tail length is strongly correlated with mammalian cell toxicity. Furthermore, this work demonstrates the utility of the PLAD assay in rapidly evaluating the effect of molecular property changes in similar libraries.

Assessment of antimicrobial usage and residues in commercial chicken eggs from smallholder poultry keepers in Morogoro municipality, Tanzania.

PubMed

Nonga, H E; Simon, C; Karimuribo, E D; Mdegela, R H

2010-08-01

Occurrence of antimicrobial residues in commercial chicken eggs was determined in Morogoro municipality between January and February 2007. Twenty smallholder farmers were interviewed on the types of antimicrobials, reasons of use and their awareness on antimicrobial withdrawal period. Seventy egg samples were collected for qualitative antimicrobial drug residues analysis by use of agar well diffusion and Delvotest SP assays. It was found that farmers use antimicrobial drugs as prophylaxis and treatment of common chicken diseases namely fowl typhoid (85%), infectious bursa disease (Gumboro) (65%) infectious coryza (65%), collibacilosis (55%), coccidiosis (54%), Newcastle disease (50%), helminthosis (20%) and fowl pox (15%). Antimicrobials accounted for 85% of the drugs commonly used. It was also found that 65% of the farmers treat their chicken themselves. The common drugs were oxytetracycline (75%), egg booster (50%), amprolium (35%), sulphamethoxypyridazine (35%), sulphanilamide (25%), chlortetracyclines (10%), chloramphenicol (10%), sulphadiazine-trimethoprim (20%), duoxycycline (20%), sulphadiazine (25%) and flumequine (10%). Eighty per cent of the farmers had knowledge on antimicrobial withdrawal period sold eggs before withdrawal period and almost 85% were unaware of possible effects of antimicrobial residues in humans. All 70 eggs were positive to antimicrobial residues by Delvotest kit, but 21.4% positive with agar well diffusion test. It was concluded that the presence of antimicrobial residues in table eggs could be of public health significance to the egg consumers in Morogoro municipality.

Comparison of media for detection of fungi on spacecraft

NASA Technical Reports Server (NTRS)

Herring, C. M.; Brandsberg, J. W.; Oxborrow, G. S.; Puleo, J. R.

1974-01-01

Five media, including Trypticase soy agar (TSA; BBL) pour plates, spread plates of TSA, Mycophil agar with chloromycetin, Mycophil agar with chloromycetin and Actidione, and cornmeal agar with chloromycetin were quantitatively and qualitatively compared for the detection of fungi on spacecraft. Cornmeal agar with chloromycetin yielded the highest number of fungal colonies, although not always significantly higher than Mycophil agar with chloromycetin or TSA spread plates. Cornmeal agar with chloromycetin also gave the best qualitative representation of fungi on the spacecraft, recovering 68% of the genera found from all media. This medium yielded 10 times the number of fungal colonies and 3 times the number of genera found on TSA pour plates as currently used for spacecraft assay.

Comparison of media for detection of fungi on spacecraft.

PubMed

Herring, C M; Brandsberg, J W; Oxborrow, G S; Puleo, J R

1974-03-01

Five media, including Trypticase soy agar (TSA; BBL) pour plates, spread plates of TSA, Mycophil agar with chloromycetin, Mycophil agar with chloromycetin and Actidione, and cornmeal agar with chloromycetin were quantitatively and qualitatively compared for the detection of fungi on spacecraft. Cornmeal agar with chloromycetin yielded the highest number of fungal colonies, although not always significantly higher than Mycophil agar with chloromycetin or TSA spread plates. Cornmeal agar with chloromycetin also gave the best qualitative representation of fungi on the spacecraft, recovering 68% of the genera found from all media. This medium yielded 10 times the number of fungal colonies and 3 times the number of genera found on TSA pour plates as currently used for spacecraft assay.

Antifungal efficacy of plant essential oils against stored grain fungi of Fusarium spp.

PubMed

Kumar, Peeyush; Mishra, Sapna; Kumar, Atul; Sharma, Amit Kumar

2016-10-01

The control potential of seven plant essential oils was evaluated against Fusarium proliferatum (Matsushima) Nirenberg and Fusarium verticillioides Sheldon. The fungicidal activity was assessed through microtiter plate assay to determine the minimum inhibitory and fungicidal concentration of essential oils. The essential oil of Mentha arvensis was adjudged as best for inhibiting the fungal growth, while oil of Thymus vulgaris and Anethum graveolens showed high efficacy in terms of fungicidal activity. The oil of M. arvensis and T. vulgaris also showed good inhibition activity in agar disc diffusion assay. M. arvensis essential oil was analysed for its composition using gas chromatography/mass spectrometry revealing menthol (63.18 %), menthone (15.08 %), isomenthyl acetate (5.50 %) and limonene (4.31 %) as major components. Significant activity of M. arvensis essential oil against F. proliferatum and F. verticillioides isolates obtained, pave the way for its use as antifungal control agents.

Multiparametric comparison of chromogenic-based culture methods used to assess the microbiological quality of drinking water and the mFC method combined with a molecular confirmation procedure.

PubMed

Maheux, Andrée F; Dion-Dupont, Vanessa; Bisson, Marc-Antoine; Bouchard, Sébastien; Jubinville, Éric; Nkuranga, Martine; Rodrigue, Lynda; Bergeron, Michel G; Rodriguez, Manuel J

2015-03-01

MI agar and Colilert(®), as well as mFC agar combined with an Escherichia coli-specific molecular assay (mFC + E. coli rtPCR), were compared in terms of their sensitivity, ease of use, time to result and affordability. The three methods yielded a positive E. coli signal for 11.5, 10.8, and 11.5% of the 968 well water samples tested, respectively. One hundred and thirty-six (136) samples gave blue colonies on mFC agar and required confirmation. E. coli-specific rtPCR showed false-positive results in 23.5% (32/136) of cases. In terms of ease of use, Colilert was the simplest method to use while the MI method provided ease of use comparable to all membrane filtration methods. However, the mFC + E. coli rtPCR assay required highly trained employees for confirmation purposes. In terms of affordability, and considering contamination rate of well water samples tested, the Colilert method and the mFC + E. coli rtPCR assay were at least five times more costly than the MI agar method. Overall, compared with the other two methods tested, the MI agar method offers the most advantages to assess drinking water quality.

Inhibition of Listeria monocytogenes by Food-Borne Yeasts†

PubMed Central

Goerges, Stefanie; Aigner, Ulrike; Silakowski, Barbara; Scherer, Siegfried

2006-01-01

Many bacteria are known to inhibit food pathogens, such as Listeria monocytogenes, by secreting a variety of bactericidal and bacteriostatic substances. In sharp contrast, it is unknown whether yeast has an inhibitory potential for the growth of pathogenic bacteria in food. A total of 404 yeasts were screened for inhibitory activity against five Listeria monocytogenes strains. Three hundred and four of these yeasts were isolated from smear-ripened cheeses. Most of the yeasts were identified by Fourier transform infrared spectroscopy. Using an agar-membrane screening assay, a fraction of approximately 4% of the 304 red smear cheese isolates clearly inhibited growth of L. monocytogenes. Furthermore, 14 out of these 304 cheese yeasts were cocultivated with L. monocytogenes WSLC 1364 on solid medium to test the antilisterial activity of yeast in direct cell contact with Listeria. All yeasts inhibited L. monocytogenes to a low degree, which is most probably due to competition for nutrients. However, one Candida intermedia strain was able to reduce the listerial cell count by 4 log units. Another four yeasts, assigned to C. intermedia (three strains) and Kluyveromyces marxianus (one strain), repressed growth of L. monocytogenes by 3 log units. Inhibition of L. monocytogenes was clearly pronounced in the cocultivation assay, which simulates the conditions and contamination rates present on smear cheese surfaces. We found no evidence that the unknown inhibitory molecule is able to diffuse through soft agar. PMID:16391059

Study on E. coli and Salmonella biofilms from fresh fruits and vegetables.

PubMed

Amrutha, Balagopal; Sundar, Kothandapani; Shetty, Prathapkumar Halady

2017-04-01

Foodborne outbreaks associated with fresh fruits and vegetables are on the rise worldwide. Biofilm formation is one of the important traits of pathogens making them strongly attached to substrates as well as express virulence phenotypes. Present study investigates the biofilm forming ability of E. coli and Salmonella sp. isolated from fresh fruits and vegetables. A total of 53 strains, including 35 E. coli and 18 Salmonella sp. isolated from different fruit and vegetable samples were taken into account for the study. Initial screening for biofilm formation was done using Congo Red agar plate test. Results revealed that 22.8% E. coli and 22.2% Salmonella sp. were potential biofilm formers. However, the MTP (Micro-Titre Plate) assay suggested more isolates of both E. coli and Salmonella sp. were moderate to strong biofilm producers. Agar plate diffusion assay with Agrobacterium tumefaciens NTL-4 showed the production of quorum signaling molecules (AHLs) by three isolates of E. coli and one Salmonella sp. Two E. coli isolates showed a significant amount of EPS production indicating higher biofilm forming potential. The Presence of LUX R homologue gene ( sdi A) in two of the Salmonella isolates were confirmed by PCR which demonstrated their potential pathogenicity. Results of the work underline the biofilm forming and potentially virulent capacities of isolates from the surface of fruits and vegetables.

Detection of group B streptococci in Lim broth by use of group B streptococcus peptide nucleic acid fluorescent in situ hybridization and selective and nonselective agars.

PubMed

Montague, Naomi S; Cleary, Timothy J; Martinez, Octavio V; Procop, Gary W

2008-10-01

The sensitivity, specificity, and positive and negative predictive values for the detection of group B streptococci from Lim enrichment broth with sheep blood agar (SBA), with selective Streptococcus agar (SSA), and by a peptide nucleic acid fluorescent in situ hybridization (PNA FISH) assay were as follows: for culture on SBA, 68.4%, 100%, 100%, and 87.9%, respectively; for culture on SSA, 85.5%, 100%, 100%, and 94.1%, respectively; and for the PNA FISH assay, 97.4%, 98.3%, 96.1%, and 98.9%, respectively.

Practical direct plaque assay for coliphages in 100-ml samples of drinking water.

PubMed Central

Grabow, W O; Coubrough, P

1986-01-01

A practical single-agar-layer plaque assay for the direct detection of coliphages in 100-ml samples of water was designed and evaluated. With this assay a 100-ml sample of water, an agar medium containing divalent cations, and the host Escherichia coli C (ATCC 13706) were mixed in a single container, and the mixture was plated on 10 14-cm-diameter petri dishes. It was more sensitive, reliable, and accurate than various other methods and proved rapid, simple, and economic. PMID:3532952

Ultra-sensitive detection of tumorigenic cellular impurities in human cell-processed therapeutic products by digital analysis of soft agar colony formation.

PubMed

Kusakawa, Shinji; Yasuda, Satoshi; Kuroda, Takuya; Kawamata, Shin; Sato, Yoji

2015-12-08

Contamination with tumorigenic cellular impurities is one of the most pressing concerns for human cell-processed therapeutic products (hCTPs). The soft agar colony formation (SACF) assay, which is a well-known in vitro assay for the detection of malignant transformed cells, is applicable for the quality assessment of hCTPs. Here we established an image-based screening system for the SACF assay using a high-content cell analyzer termed the digital SACF assay. Dual fluorescence staining of formed colonies and the dissolution of soft agar led to accurate detection of transformed cells with the imaging cytometer. Partitioning a cell sample into multiple wells of culture plates enabled digital readout of the presence of colonies and elevated the sensitivity for their detection. In practice, the digital SACF assay detected impurity levels as low as 0.00001% of the hCTPs, i.e. only one HeLa cell contained in 10,000,000 human mesenchymal stem cells, within 30 days. The digital SACF assay saves time, is more sensitive than in vivo tumorigenicity tests, and would be useful for the quality control of hCTPs in the manufacturing process.

Ultra-sensitive detection of tumorigenic cellular impurities in human cell-processed therapeutic products by digital analysis of soft agar colony formation

PubMed Central

Kusakawa, Shinji; Yasuda, Satoshi; Kuroda, Takuya; Kawamata, Shin; Sato, Yoji

2015-01-01

Contamination with tumorigenic cellular impurities is one of the most pressing concerns for human cell-processed therapeutic products (hCTPs). The soft agar colony formation (SACF) assay, which is a well-known in vitro assay for the detection of malignant transformed cells, is applicable for the quality assessment of hCTPs. Here we established an image-based screening system for the SACF assay using a high-content cell analyzer termed the digital SACF assay. Dual fluorescence staining of formed colonies and the dissolution of soft agar led to accurate detection of transformed cells with the imaging cytometer. Partitioning a cell sample into multiple wells of culture plates enabled digital readout of the presence of colonies and elevated the sensitivity for their detection. In practice, the digital SACF assay detected impurity levels as low as 0.00001% of the hCTPs, i.e. only one HeLa cell contained in 10,000,000 human mesenchymal stem cells, within 30 days. The digital SACF assay saves time, is more sensitive than in vivo tumorigenicity tests, and would be useful for the quality control of hCTPs in the manufacturing process. PMID:26644244

Assessment of formulas for calculating critical concentration by the agar diffusion method.

PubMed Central

Drugeon, H B; Juvin, M E; Caillon, J; Courtieu, A L

1987-01-01

The critical concentration of antibiotic was calculated by using the agar diffusion method with disks containing different charges of antibiotic. It is currently possible to use different calculation formulas (based on Fick's law) devised by Cooper and Woodman (the best known) and by Vesterdal. The results obtained with the formulas were compared with the MIC results (obtained by the agar dilution method). A total of 91 strains and two cephalosporins (cefotaxime and ceftriaxone) were studied. The formula of Cooper and Woodman led to critical concentrations that were higher than the MIC, but concentrations obtained with the Vesterdal formula were closer to the MIC. The critical concentration was independent of method parameters (dilution, for example). PMID:3619419

Estimating the Diffusion Coefficients of Sugars Using Diffusion Experiments in Agar-Gel and Computer Simulations.

PubMed

Miyamoto, Shuichi; Atsuyama, Kenji; Ekino, Keisuke; Shin, Takashi

2018-01-01

The isolation of useful microbes is one of the traditional approaches for the lead generation in drug discovery. As an effective technique for microbe isolation, we recently developed a multidimensional diffusion-based gradient culture system of microbes. In order to enhance the utility of the system, it is favorable to have diffusion coefficients of nutrients such as sugars in the culture medium beforehand. We have, therefore, built a simple and convenient experimental system that uses agar-gel to observe diffusion. Next, we performed computer simulations-based on random-walk concepts-of the experimental diffusion system and derived correlation formulas that relate observable diffusion data to diffusion coefficients. Finally, we applied these correlation formulas to our experimentally-determined diffusion data to estimate the diffusion coefficients of sugars. Our values for these coefficients agree reasonably well with values published in the literature. The effectiveness of our simple technique, which has elucidated the diffusion coefficients of some molecules which are rarely reported (e.g., galactose, trehalose, and glycerol) is demonstrated by the strong correspondence between the literature values and those obtained in our experiments.

Impact of pectin esterification on the antimicrobial activity of nisin-loaded pectin particles.

PubMed

Krivorotova, Tatjana; Staneviciene, Ramune; Luksa, Juliana; Serviene, Elena; Sereikaite, Jolanta

2017-01-01

The relationship between pectin structure and the antimicrobial activity of nisin-loaded pectin particles was examined. The antimicrobial activity of five different nisin-loaded pectin particles, i.e., nisin-loaded high methoxyl pectin, low methoxyl pectin, pectic acid, dodecyl pectin with 5.4 and 25% degree of substitution were tested in the pH range of 4.0-7.0 by agar-diffusion assay and agar plate count methods. It was found that the degree of esterification of carboxyl group of galacturonic acid in pectin molecule is important for the antimicrobial activity of nisin-loaded pectin particles. Nisin-loaded particles prepared using pectic acid or the pectin with low degree of esterification exhibit higher antimicrobial activity than nisin-loaded high methoxyl pectin particles. Pectins with free carboxyl groups or of low degree of esterification are the most suitable for particles preparation. Moreover, nisin-loaded pectin particles were active at close to neutral or neutral pH values. Therefore, they could be effectively applied for food preservation. © 2016 American Institute of Chemical Engineers Biotechnol. Prog., 33:245-251, 2017. © 2016 American Institute of Chemical Engineers.

Comparison of Media for Detection of Fungi on Spacecraft

PubMed Central

Herring, C. M.; Brandsberg, J. W.; Oxborrow, G. S.; Puleo, J. R.

1974-01-01

Five media, including Trypticase soy agar (TSA; BBL) pour plates, spread plates of TSA, Mycophil agar with chloromycetin, Mycophil agar with chloromycetin and Actidione, and cornmeal agar with chloromycetin were quantitatively and qualitatively compared for the detection of fungi on spacecraft. Cornmeal agar with chloromycetin yielded the highest number of fungal colonies, although not always significantly higher than Mycophil agar with chloromycetin or TSA spread plates. Cornmeal agar with chloromycetin also gave the best qualitative representation of fungi on the spacecraft, recovering 68% of the genera found from all media. This medium yielded 10 times the number of fungal colonies and 3 times the number of genera found on TSA pour plates as currently used for spacecraft assay. PMID:4151044

Differentiating non-0157:H7 STEC serogroups from ground beef plated on agar media by hyperspetral imaging

USDA-ARS?s Scientific Manuscript database

Introduction: The development of an assay to detect and confirm a positive non-O157:H7 isolate is challenging when mixed morphologically results are obtained from the serogroups growing on Rainbow agar. Rainbow agar is only claimed by the manufacturer to be very specific for E.coli O157:H7 strain...

Immunochemical and biological properties of a mouse monoclonal antibody reactive to prunus necrotic ringspot ilarvirus.

PubMed

Aebig, J A; Jordan, R L; Lawson, R H; Hsu, H T

1987-01-01

A monoclonal antibody reacting with prunus necrotic ringspot ilarvirus was tested in immunochemical studies, neutralization of infectivity assays, and by immuno-electron microscopy. The antibody was able to detect the 27,000 Mr coat protein of prunus necrotic ringspot ilarvirus in western blots and also detected all polypeptide fragments generated after incubation of whole virus with proteolytic enzymes. In neutralization of infectivity studies, the antibody blocked virus infectivity, although it did not precipitate the antigen in agar gel Ouchterlony double diffusion tests. Immuno-electron microscopy confirmed that the antibody coats virions but does not cause clumping. The antibody may be a useful tool for investigating coat protein-dependent initiation of ilarvirus infection.

Inhibitory Effects of Pterodon emarginatus Bean Oil and Extract on Staphylococcus aureus

PubMed Central

Mendes, V. S.; Sant'Anna, J. B.; Oliveira, S. C. C.; Maldonade, Iriani Rodrigues; Machado, Eleuza Rodrigues

2017-01-01

Background: Pterodon emarginatus is a tree of the Brazilian Savannah. The beans of this tree are used in folk medicine as anti-inflammatory preparations, especially for infections caused by Staphylococcus aureus. These bacteria can cause simple infections or serious illnesses such as pneumonia, meningitis, endocarditis, toxic shock syndrome, septicemia, and others. Objective: This study had the goal of verifying the effect of the essential oil (OE) from P. emarginatus on the inhibition of S. aureus in culture medium, i.e., “ in vitro” tests. Materials and Methods: The vegetable material was cut and crushed with a press. The OE was obtained by extraction using hexane, alcohol, and water. The P. emarginatus extracts obtained were used to evaluate the antimicrobial effect on S. aureus (ATCC 25923) by tests of well diffusion, disc diffusion, and microdilution. The strain used in the assays was maintained in brain heart infusion broth and nutrient agar until testing. Afterward, the bacteria were spread on agar plates with Mueller-Hinton agar medium. In the wells and on the paper discs, the OE suspensions were placed in the following volumes: 10, 15, 20, 25, 30, 40, and 80 μL and subsequently they were incubated at 35°C ± 2°C. After 24 h, the number of colony-forming unit was determined. Results: Pure OE and hydroalcoholic extract inhibited the growth of S. aureus, while aqueous extract had no effect on bacterial growth in all microbial methods used. Conclusion: Thus, the present study showed the potential of sucupira-based extracts against S. aureus growth, opening new perspectives for the evaluation of these bioactive compounds as phytopharmaceutical products. SUMMARY Plant extract act as antimicrobials to prevent and reduce bacterial contaminationBeans of Pterodon emarginatus has antibacterial propertiesExtraction with different solvents might implicate on the rate of bacterial deathThe effect of different microbiological methods (well diffusion, disc diffusion and microdilution) was evaluated on reducing CFUThe results showed by MBC that concentrations superior to 10% (v/v) using AC and 7.5% (v/v) using OE were necessary to eliminate colonies formedAccording to data of MIC, at 2.5% of AC and OE was enough to kill S. aureusThe well diffusion technique demonstrated better performance than disc diffusion test for OE and AC extractsHydroalcoholic and oil extracts of sucupira beans had highest effect against Staphylococcus aureusAqueous extract had no effect on bacterial growth in all microbial methods testedThe sucupira-based extracts is a promising source as herbal drug due to therapeutic value Abbreviations Used: OE: Essencial oil; AC: Hydroalcoholic oil extract; AQ: Aqueous extracts; MIC: Minimum inhibitory concentration; MBC: Minimum bactericidal concentration; CFU: Colony formed unit. PMID:29263627

Inhibitory Effects of Pterodon emarginatus Bean Oil and Extract on Staphylococcus aureus.

PubMed

Mendes, V S; Sant'Anna, J B; Oliveira, S C C; Maldonade, Iriani Rodrigues; Machado, Eleuza Rodrigues

2017-01-01

Pterodon emarginatus is a tree of the Brazilian Savannah. The beans of this tree are used in folk medicine as anti-inflammatory preparations, especially for infections caused by Staphylococcus aureus . These bacteria can cause simple infections or serious illnesses such as pneumonia, meningitis, endocarditis, toxic shock syndrome, septicemia, and others. This study had the goal of verifying the effect of the essential oil (OE) from P. emarginatus on the inhibition of S. aureus in culture medium, i.e., " in vitro " tests. The vegetable material was cut and crushed with a press. The OE was obtained by extraction using hexane, alcohol, and water. The P. emarginatus extracts obtained were used to evaluate the antimicrobial effect on S. aureus (ATCC 25923) by tests of well diffusion, disc diffusion, and microdilution. The strain used in the assays was maintained in brain heart infusion broth and nutrient agar until testing. Afterward, the bacteria were spread on agar plates with Mueller-Hinton agar medium. In the wells and on the paper discs, the OE suspensions were placed in the following volumes: 10, 15, 20, 25, 30, 40, and 80 μL and subsequently they were incubated at 35°C ± 2°C. After 24 h, the number of colony-forming unit was determined. Pure OE and hydroalcoholic extract inhibited the growth of S. aureus , while aqueous extract had no effect on bacterial growth in all microbial methods used. Thus, the present study showed the potential of sucupira-based extracts against S. aureus growth, opening new perspectives for the evaluation of these bioactive compounds as phytopharmaceutical products. Plant extract act as antimicrobials to prevent and reduce bacterial contaminationBeans of Pterodon emarginatus has antibacterial propertiesExtraction with different solvents might implicate on the rate of bacterial deathThe effect of different microbiological methods (well diffusion, disc diffusion and microdilution) was evaluated on reducing CFUThe results showed by MBC that concentrations superior to 10% (v/v) using AC and 7.5% (v/v) using OE were necessary to eliminate colonies formedAccording to data of MIC, at 2.5% of AC and OE was enough to kill S. aureus The well diffusion technique demonstrated better performance than disc diffusion test for OE and AC extractsHydroalcoholic and oil extracts of sucupira beans had highest effect against Staphylococcus aureus Aqueous extract had no effect on bacterial growth in all microbial methods testedThe sucupira-based extracts is a promising source as herbal drug due to therapeutic value Abbreviations Used: OE: Essencial oil; AC: Hydroalcoholic oil extract; AQ: Aqueous extracts; MIC: Minimum inhibitory concentration; MBC: Minimum bactericidal concentration; CFU: Colony formed unit.

Improved quantitative recovery of Listeria monocytogenes from stainless steel surfaces using a one-ply composite tissue.

PubMed

Vorst, Keith L; Todd, Ewen C D; Rysert, Elliot T

2004-10-01

Four sampling devices, a sterile environmental sponge (ES), a sterile cotton-tipped swab (CS), a sterile calcium alginate fiber-tipped swab (CAS), and a one-ply composite tissue (CT), were evaluated for quantitative recovery of Listeria monocytogenes from a food-grade stainless steel surface. Sterile 304-grade stainless steel plates (6 by 6 cm) were inoculated with approximately 106 CFU/cm2 L. monocytogenes strain Scott A and dried for 1 h. The ES and CT sampling devices were rehydrated in phosphate buffer solution. After plate swabbing, ES and CT were placed in 40 ml of phosphate buffer solution, stomached for 1 min and hand massaged for 30 s. Each CS and CAS device was rehydrated in 0.1% peptone before swabbing. After swabbing, CS and CAS were vortexed in 0.1% peptone for 1 min. Samples were spiral plated on modified Oxford agar with modified Oxford agar Rodac Contact plates used to recover any remaining cells from the stainless steel surface. Potential inhibition from CT was examined in both phosphate buffer solution and in a modified disc-diffusion assay. Recovery was 2.70, 1.34, and 0.62 log greater using CT compared with ES, CS, and CAS, respectively, with these differences statistically significant (P < 0.001) for ES and CT and for CAS, CS, and CT (P < 0.05). Rodac plates were typically overgrown following ES, positive after CS and CAS, and negative after CT sampling. CT was noninhibitory in both phosphate buffer solution and the modified disc-diffusion assay. Using scanning electron microscopy, Listeria cells were observed on stainless steel plates sampled with each sampling device except CT. The CT device, which is inexpensive and easy to use, represents a major improvement over other methods in quantifying L. monocytogenes on stainless steel surfaces and is likely applicable to enrichment of environmental samples.

Secondary metabolites extracted from marine sponge associated Comamonas testosteroni and Citrobacter freundii as potential antimicrobials against MDR pathogens and hypothetical leads for VP40 matrix protein of Ebola virus: an in vitro and in silico investigation.

PubMed

Skariyachan, Sinosh; Acharya, Archana B; Subramaniyan, Saumya; Babu, Sumangala; Kulkarni, Shruthi; Narayanappa, Rajeswari

2016-09-01

The current study explores therapeutic potential of metabolites extracted from marine sponge (Cliona sp.)-associated bacteria against MDR pathogens and predicts the binding prospective of probable lead molecules against VP40 target of Ebola virus. The metabolite-producing bacteria were characterized by agar overlay assay and as per the protocols in Bergey's manual of determinative bacteriology. The antibacterial activities of extracted metabolites were tested against clinical pathogens by well-diffusion assay. The selected metabolite producers were characterized by 16S rDNA sequencing. Chemical screening and Fourier Transform Infrared (FTIR) analysis for selected compounds were performed. The probable lead molecules present in the metabolites were hypothesized based on proximate analysis, FTIR data, and literature survey. The drug-like properties and binding potential of lead molecules against VP40 target of Ebola virus were hypothesized by computational virtual screening and molecular docking. The current study demonstrated that clear zones around bacterial colonies in agar overlay assay. Antibiotic sensitivity profiling demonstrated that the clinical isolates were multi-drug resistant, however; most of them showed sensitivity to secondary metabolites (MIC-15 μl/well). The proximate and FTIR analysis suggested that probable metabolites belonged to alkaloids with O-H, C-H, C=O, and N-H groups. 16S rDNA characterization of selected metabolite producers demonstrated that 96% and 99% sequence identity to Comamonas testosteroni and Citrobacter freundii, respectively. The docking studies suggested that molecules such as Gymnastatin, Sorbicillactone, Marizomib, and Daryamide can designed as probable lead candidates against VP40 target of Ebola virus.

Clinical test to detect mecA and antibiotic resistance in Staphylococcus aureus, based on novel biotechnological methods.

PubMed

Shahmohammadi, Mohammad Reza; Nahaei, Mohammad Reza; Akbarzadeh, Abolfazl; Milani, Morteza

2016-09-01

Methicillin-resistant Staphylococcus aureus (MRSA) is one of the most common organisms isolated from clinical samples, and has been associated with morbidity and mortality among hospitalized patients. The aim of this study was to evaluate the prevalence and antibiotic susceptibility patterns among MRSA and methicillin-sensitive S. aureus (MSSA) isolates collected from four hospitals in Iran. A total of 183 isolates of S. aureus were collected from various clinical specimens of four hospitals in Iran. The isolates were identified by using the conventional biochemical tests. Three methods-oxacillin agar disk diffusion, oxacillin agar screening, and PCR- were applied to determine susceptibility to oxacillin. The conventional disk agar diffusion test was used to evaluate the antibiotic sensitivity of our isolates against 15 antibiotics, according to the guidelines of the Clinical and Laboratory Standards Institute (CLSI). Of 183 isolates, 77 isolates (42.1%) were found to be MRSA, by the PCR method. The highest antibiotic resistance was found to be against penicillin, co-trimoxazole, erythromycin, and tetracycline respectively. All isolates were susceptible to vancomycin, according to the results of disk agar diffusion. Among other antibiotics, teicoplanin (84%) and fusidic acid (80.5%) were more active against MRSA isolates. For the different methods evaluated, the sensitivities and specificities were as follows: for disk agar diffusion (84.9% and 95.9%) and for agar screening test with oxacillin concentrations of 0.6 μg/ml (70.8% and 97.4%), 4 μg/ml (96.1%and 97.2%) and 6 μg/ml (96% and 96.3%), respectively. The results of our study showed that 47% of S. aureus isolates were MRSA. Overall, in this research study, resistance to all test antimicrobial agents in MRSA isolates were higher than that of MSSA isolates. Our results also revealed that 85% of mecA-positive isolates and 15% of mecA-negative isolates were resistant to methicillin; while 96% of mecA-negative isolates were sensitive to methicillin. Meanwhile 4% of mecA-positive isolates were also sensitive to methicillin.

Immobilization of pectin degrading enzyme from Bacillus licheniformis KIBGE IB-21 using agar-agar as a support.

PubMed

Rehman, Haneef Ur; Aman, Afsheen; Zohra, Raheela Rahmat; Qader, Shah Ali Ul

2014-02-15

Pectinase from Bacillus licheniformis KIBGE IB-21 was immobilized in agar-agar matrix using entrapment technique. Effect of different concentrations of agar-agar on pectinase immobilization was investigated and it was found that maximum immobilization was achieved at 3.0% agar-agar with 80% enzyme activity. After immobilization, the optimum temperature of enzyme increased from 45 to 50 °C and reaction time from 5 to 10 minutes as compared to free enzyme. Due to the limited diffusion of high molecular weight substrate, K(m) of immobilized enzyme slightly increased from 1.017 to 1.055 mg ml(-1), while Vmax decreased from 23,800 to 19,392 μM min(-1) as compared to free enzyme. After 120 h entrapped pectinase retained their activity up to 82% and 71% at 30 °C and 40 °C, respectively. The entrapped pectinase showed activity until 10th cycle and maintain 69.21% activity even after third cycle. Copyright © 2013 Elsevier Ltd. All rights reserved.

Control of the pattern of perithecium development in Sordaria fimicola on agar medium.

PubMed

Pollock, R T

1975-06-01

In a Sordaria fimicola (Rob.) Ces. and de Not. colony grown on agar medium in a petri plate, perithecia developed in a narrow band around the plate edge after the colony margin reached the edge. Physical wounding of the colony carried out shortly before or during the time perithecia were developing around the plate edge stimulated perithecium development in the wound area. Diffusion barriers were created by cutting small trenches in the agar parallel to the plate edge. The trenches were made at several different positions between the plate center and edge using cultures of several different ages, and the resultant distribution of perithecia along the trench edges suggested that the colony center and periphery produce diffusible inhibitors of perithecium development. These inhibitors may be responsible, in part, for the observed pattern of perithecium development in the colony.

Molecular Diffusion Coefficients: Experimental Determination and Demonstration.

ERIC Educational Resources Information Center

Fate, Gwendolyn; Lynn, David G.

1990-01-01

Presented are laboratory methods which allow the demonstration and determination of the diffusion coefficients of compounds ranging in size from water to small proteins. Included are the procedures involving the use of a spectrometer, UV cell, triterated agar, and oxygen diffusion. Results including quantification are described. (CW)

Individual based simulations of bacterial growth on agar plates

NASA Astrophysics Data System (ADS)

Ginovart, M.; López, D.; Valls, J.; Silbert, M.

2002-03-01

The individual based simulator, INDividual DIScrete SIMulations (INDISIM) has been used to study the behaviour of the growth of bacterial colonies on a finite dish. The simulations reproduce the qualitative trends of pattern formation that appear during the growth of Bacillus subtilis on an agar plate under different initial conditions of nutrient peptone concentration, the amount of agar on the plate, and the temperature. The simulations are carried out by imposing closed boundary conditions on a square lattice divided into square spatial cells. The simulator studies the temporal evolution of the bacterial population possible by setting rules of behaviour for each bacterium, such as its uptake, metabolism and reproduction, as well as rules for the medium in which the bacterial cells grow, such as concentration of nutrient particles and their diffusion. The determining factors that characterize the structure of the bacterial colony patterns in the presents simulations, are the initial concentrations of nutrient particles, that mimic the amount of peptone in the experiments, and the set of values for the microscopic diffusion parameter related, in the experiments, to the amount of the agar medium.

Ultrafiltration and thermal processing effects on Maillard reaction products and biological properties of date palm sap syrups (Phoenix dactylifera L.).

PubMed

Makhlouf-Gafsi, Ines; Krichen, Fatma; Mansour, Riadh Ben; Mokni, Abir; Sila, Assad; Bougatef, Ali; Blecker, Christophe; Attia, Hamadi; Besbes, Souhail

2018-08-01

The effect of ultrafiltration process and temperature concentration on MRPs content and antioxidant, antimicrobial and cytotoxic properties of date palm sap syrups were investigated. MRPs were analyzed by HPLC. Antioxidant activity was evaluated by reducing power and DPPH free radical and H 2 O 2 scavenging activities. Antimicrobial activity was evaluated by the agar disk diffusion method. In vitro cytotoxic activity was examined by cell proliferation assay. Date sap syrups displayed strong antioxidant activities which are correlated 5HMF and 2F contents. In addition, concentration at 100 °C, unlike ultrafiltration process, enhanced significantly the antioxidant activities sap syrups and total phenolic contents. The antimicrobial activities showed marked activity against S. enterica, P. aeruginosa, S. aureus, L. monocytogenes with an inhibition zone of 21, 34, 27 and 34 mm respectively. Cytotoxicity assays showed that sap syrups can inhibit the proliferation of HeLa cell lines at high concentration. Published by Elsevier Ltd.

Inhibitory effects of terpene alcohols and aldehydes on growth of green alga Chlorella pyrenoidosa

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ikawa, Miyoshi; Mosley, S.P.; Barbero, L.J.

1992-10-01

The growth of the green alga Chlorella pyrenoidosa was inhibited by terpene alcohols and the terpene aldehyde citral. The strongest activity was shown by citral. Nerol, geraniol, and citronellol also showed pronounced activity. Strong inhibition was linked to acyclic terpenes containing a primary alcohol or aldehyde function. Inhibition appeared to be taking place through the vapor phase rather than by diffusion through the agar medium from the terpene-treated paper disks used in the system. Inhibition through agar diffusion was shown by certain aged samples of terpene hydrocarbons but not by recently purchased samples.

In vitro anti-biofilm and anti-bacterial activity of Junceella juncea for its biomedical application

PubMed Central

Kumar, P; Selvi, S Senthamil; Govindaraju, M

2012-01-01

Objective To investigate the anti-biofilm and anti-bacterial activity of Junceella juncea (J. juncea) against biofilm forming pathogenic strains. Methods Gorgonians were extracted with methanol and analysed with fourier transform infrared spectroscopy. Biofilm forming pathogens were identified by Congo red agar supplemented with sucrose. A quantitative spectrophotometric method was used to monitor in vitro biofilm reduction by microtitre plate assay. Anti-bacterial activity of methanolic gorgonian extract (MGE) was carried out by disc diffusion method followed by calculating the percentage of increase with crude methanol (CM). Results The presence of active functional group was exemplified by FT-IR spectroscopy. Dry, black, crystalline colonies confirm the production of extracellular polymeric substances responsible for biofilm formation in Congo red agar. MGE exhibited potential anti-biofilm activity against all tested bacterial strains. The anti-bacterial activity of methanolic extract was comparably higher in Salmonella typhii followed by Escherichia coli, Vibrio cholerae and Shigella flexneri. The overall percentage of increase was higher by 50.2% to CM. Conclusions To conclude, anti-biofilm and anti-bacterial efficacy of J. juncea is impressive over biofilm producing pathogens and are good source for novel anti-bacterial compounds. PMID:23593571

In vitro antibacterial effects of glass-ionomer cement containing ethanolic extract of propolis on Streptococcus mutans

PubMed Central

Topcuoglu, Nursen; Ozan, Fatih; Ozyurt, Mustafa; Kulekci, Guven

2012-01-01

Objective: The aim of this study was to evaluate the antibacterial property of glass-ionomer cement (GIC) containing propolis against Streptococcus mutans and its effect on the in vitro S. mutans biofilm formation. Methods: Ethanolic extract of propolis (EEP) was prepared at two concentrations as 25 and 50%. Three different experimental GIC disks were prepared using pure liquid and liquid solutions diluted with 25 and 50 percent of EEP concentrations. Minimum inhibitory concentration (MIC) of EEP on the growth of S. mutans ATCC 25175 was determined by using agar dilution method. Agar diffusion test and an in vitro S. mutans biofilm assay for GIC disks with and without EEP were performed. Results: MIC values of Turkish propolis for S. mutans ATCC 25175 was found as 25 μg/mL. Experimental GICs containing propolis exhibited inhibition zones and their dry biofilm weights were less than the pure GIC. The bacterial density was lower in the GIC containing 50% EEP. Conclusions: A distinct antibacterial and antibiofilm efficacy of propolis containing GIC on S. mutans has been observed. Although further research is needed to show clinical results, antibacterial GIC containing propolis would be a promising material for restoration. PMID:23077424

Antibacterial activity of Zuccagnia punctata Cav. ethanolic extracts.

PubMed

Zampini, Iris C; Vattuone, Marta A; Isla, Maria I

2005-12-01

The present study was conducted to investigate antibacterial activity of Zuccagnia punctata ethanolic extract against 47 strains of antibiotic-resistant Gram-negative bacteria and to identify bioactive compounds. Inhibition of bacterial growth was investigated using agar diffusion, agar macrodilution, broth microdilution and bioautographic methods. Zuccagnia punctata extract was active against all assayed bacteria (Escherichia coli, Klebsiella pneumoniae, Proteus mirabilis, Enterobacter cloacae, Serratia marcescens, Morganella morganii, Acinetobacter baumannii, Pseudomonas aeruginosa, Stenotrophomonas maltophilia) with minimal inhibitory concentration (MIC) values ranging from 25 to 200 microg/mL. Minimal bactericidal concentration (MBC) values were identical or two-fold higher than the corresponding MIC values. Contact bioautography, indicated that Zuccagnia punctata extracts possess one major antibacterial component against Pseudomonas aeruginosa and at least three components against. Klebsiella pneumoniae and Escherichia coli. Activity-guided fractionation of 1he ethanol extract on a silica gel column yielded a compound (2',4'-dihydroxychalcone), which exhibited strong antibacterial activity with MIC values between 0.10 and 1.00 microg/mL for Proteus mirabilis, Enterobacter cloacae, Serratia marcescens, Morganella morganii, Acinetobacter baumannii, Pseudomonas aeruginosa, Stenotrophomonas maltophilia. These values are lower than imipenem (0.25-16 microg/mL). Zuccagnia punctata might provide promising therapeutic agents against infections with multi-resistant Gram-negative bacteria.

Study of methods for the improvement of bacterial transport media

NASA Technical Reports Server (NTRS)

Gardner, R. L.; Beakley, J. W.

1973-01-01

A series of 500 transport media recipes was tested for ability to hold pure cultures of Streptococcus equisimilus, Corynebacterium equi, Neisseria perflava, and Haemophilus parainfluenzae for 21 days. Stuart Medium Base with 0.4% agar was used as the control medium for this and the other experiments in the investigation. At the end of the holding period inoculated transport media were quantitatively assayed, and the control media were assayed immediately after inoculation. Three vials of each medium were inoculated with an organism, and each vial's medium was diluted and spread on duplicate plates. Assay media for this experiment included Brain Heart Infusion,(BHIA) Tryptic Soy Agar, and BHIA with 1% Isovitalex enrichment.

The effects of nutrient chemotaxis on bacterial aggregation patterns with non-linear degenerate cross diffusion

NASA Astrophysics Data System (ADS)

Leyva, J. Francisco; Málaga, Carlos; Plaza, Ramón G.

2013-11-01

This paper studies a reaction-diffusion-chemotaxis model for bacterial aggregation patterns on the surface of thin agar plates. It is based on the non-linear degenerate cross diffusion model proposed by Kawasaki et al. (1997) [5] and it includes a suitable nutrient chemotactic term compatible with such type of diffusion, as suggested by Ben-Jacob et al. (2000) [20]. An asymptotic estimation predicts the growth velocity of the colony envelope as a function of both the nutrient concentration and the chemotactic sensitivity. It is shown that the growth velocity is an increasing function of the chemotactic sensitivity. High resolution numerical simulations using Graphic Processing Units (GPUs), which include noise in the diffusion coefficient for the bacteria, are presented. The numerical results verify that the chemotactic term enhances the velocity of propagation of the colony envelope. In addition, the chemotaxis seems to stabilize the formation of branches in the soft-agar, low-nutrient regime.

In-vitro Antimicrobial Activities of Some Iranian Conifers

PubMed Central

Afsharzadeh, Maryam; Naderinasab, Mahboobe; Tayarani Najaran, Zahra; Barzin, Mohammad; Emami, Seyed Ahmad

2013-01-01

Male and female leaves and fruits of eleven different taxons of Iranian conifers (Cupressus sempervirens var. horizontalis, C. sempervirens var. sempervirens, C. sempervirens cv. Cereifeormis, Juniperus communis subsp. hemisphaerica, J. excelsa subsp. excelsa, J. excelsa subsp. polycarpos, J. foetidissima, J. oblonga, J. sabina, Platycladus orientalis and Taxus baccata) were collected from different localities of Iran, dried and extracted with methanol. The extracts were tested for their antimicrobial activity against Pseudomonas aeruginosa, Staphylococcus aureus, Escherichia coli and Candida albicans. The extracts were screened qualitatively using four different methods, the disc diffusion, hole plate, cylinder agar diffusion and agar dilution methods, whereas the minimum inhibitory concentrations (MIC) of each extract were determined by the agar dilution method. The best result was obtained by means of hole plate method in qualitative determination of antimicrobial activities of extracts and the greatest activity was found against S. aureus in all tested methods. PMID:24250573

Influence of Structural Properties and Kinetic Constraints on Bacillus cereus Growth

PubMed Central

Stecchini, Mara Lucia; Del Torre, Manuela; Sarais, Ileana; Saro, Onorio; Messina, Mariella; Maltini, Enrico

1998-01-01

The influence of structural properties and kinetic constraints on the behavior of Bacillus cereus was investigated on agar media. Dimensional criteria were used to study the growth in bacterial colonies. The architecture of the agar gel as modified by the agar content was found to influence the colony size, and smaller colonies were observed on media containing 50 to 70 g of agar liter−1. Except at low nutrient levels, colonies responded to nutrient gradients by decreasing in size the farther away they were from the nutrient source, and the decrease in colony size was influenced by the agar content. The diffusivities of glucose and a protein (insulin-like growth factor) were not affected by the gel architecture, suggesting that other factors, such as mechanical factors, could influence microbial growth in the agar systems used. Increasing the viscosity of the liquid phase of the agar media by adding polyvinylpyrrolidone resulted in a reduction in colony size. When the agar concentration was increased, the colony areas were not influenced by the viscosity of the system. PMID:9501447

Application of Paramagnetically Tagged Molecules for Magnetic Resonance Imaging of Biofilm Mass Transport Processes▿

PubMed Central

Ramanan, B.; Holmes, W. M.; Sloan, W. T.; Phoenix, V. R.

2010-01-01

Molecules become readily visible by magnetic resonance imaging (MRI) when labeled with a paramagnetic tag. Consequently, MRI can be used to image their transport through porous media. In this study, we demonstrated that this method could be applied to image mass transport processes in biofilms. The transport of a complex of gadolinium and diethylenetriamine pentaacetic acid (Gd-DTPA), a commercially available paramagnetic molecule, was imaged both in agar (as a homogeneous test system) and in a phototrophic biofilm. The images collected were T1 weighted, where T1 is an MRI property of the biofilm and is dependent on Gd-DTPA concentration. A calibration protocol was applied to convert T1 parameter maps into concentration maps, thus revealing the spatially resolved concentrations of this tracer at different time intervals. Comparing the data obtained from the agar experiment with data from a one-dimensional diffusion model revealed that transport of Gd-DTPA in agar was purely via diffusion, with a diffusion coefficient of 7.2 × 10−10 m2 s−1. In contrast, comparison of data from the phototrophic biofilm experiment with data from a two-dimensional diffusion model revealed that transport of Gd-DTPA inside the biofilm was by both diffusion and advection, equivalent to a diffusion coefficient of 1.04 × 10−9 m2 s−1. This technology can be used to further explore mass transport processes in biofilms, either by using the wide range of commercially available paramagnetically tagged molecules and nanoparticles or by using bespoke tagged molecules. PMID:20435773

Non-invasive Measurement of Thermal Diffusivity Using High-Intensity Focused Ultrasound and Through-Transmission Ultrasonic Imaging.

PubMed

Yeshurun, Lilach; Azhari, Haim

2016-01-01

Thermal diffusivity at the site ablated by high-intensity focused ultrasound (HIFU) plays an important role in the final therapeutic outcome, as it influences the temperature's spatial and temporal distribution. Moreover, as tissue thermal diffusivity is different in tumors as compared with normal tissue, it could also potentially be used as a new source of imaging contrast. The aim of this study was to examine the feasibility of combining through-transmission ultrasonic imaging and HIFU to estimate thermal diffusivity non-invasively. The concept was initially evaluated using a computer simulation. Then it was experimentally tested on phantoms made of agar and ex vivo porcine fat. A computerized imaging system combined with a HIFU system was used to heat the phantoms to temperatures below 42°C to avoid irreversible damage. Through-transmission scanning provided the time-of-flight values in a region of interest during its cooling process. The time-of-flight values were consequently converted into mean values of speed of sound. Using the speed-of-sound profiles along with the developed model, we estimated the changes in temperature profiles over time. These changes in temperature profiles were then used to calculate the corresponding thermal diffusivity of the studied specimen. Thermal diffusivity for porcine fat was found to be lower by one order of magnitude than that obtained for agar (0.313×10(-7)m(2)/s vs. 4.83×10(-7)m(2)/s, respectively, p < 0.041). The fact that there is a substantial difference between agar and fat implies that non-invasive all-ultrasound thermal diffusivity mapping is feasible. The suggested method may particularly be suitable for breast scanning. Copyright © 2016 World Federation for Ultrasound in Medicine & Biology. Published by Elsevier Inc. All rights reserved.

Electron microscopy of antigen precipitates extracted from gel diffusion plates

PubMed Central

Watson, D. H.; Le Bouvier, G. L.; Tomlinson, J. A.; Walkey, D. G. A.

1966-01-01

A method is described whereby material from virus precipitin lines from agar gel diffusion plates may be examined in the electron microscope by a negative staining technique. ImagesFIGS. 1-2FIGS. 3-4 PMID:4286708

Modeling of the Bacillus subtilis Bacterial Biofilm Growing on an Agar Substrate

PubMed Central

Wang, Xiaoling; Wang, Guoqing; Hao, Mudong

2015-01-01

Bacterial biofilms are organized communities composed of millions of microorganisms that accumulate on almost any kinds of surfaces. In this paper, a biofilm growth model on an agar substrate is developed based on mass conservation principles, Fick's first law, and Monod's kinetic reaction, by considering nutrient diffusion between biofilm and agar substrate. Our results show biofilm growth evolution characteristics such as biofilm thickness, active biomass, and nutrient concentration in the agar substrate. We quantitatively obtain biofilm growth dependence on different parameters. We provide an alternative mathematical method to describe other kinds of biofilm growth such as multiple bacterial species biofilm and also biofilm growth on various complex substrates. PMID:26355542

Modeling of the Bacillus subtilis Bacterial Biofilm Growing on an Agar Substrate.

PubMed

Wang, Xiaoling; Wang, Guoqing; Hao, Mudong

2015-01-01

Bacterial biofilms are organized communities composed of millions of microorganisms that accumulate on almost any kinds of surfaces. In this paper, a biofilm growth model on an agar substrate is developed based on mass conservation principles, Fick's first law, and Monod's kinetic reaction, by considering nutrient diffusion between biofilm and agar substrate. Our results show biofilm growth evolution characteristics such as biofilm thickness, active biomass, and nutrient concentration in the agar substrate. We quantitatively obtain biofilm growth dependence on different parameters. We provide an alternative mathematical method to describe other kinds of biofilm growth such as multiple bacterial species biofilm and also biofilm growth on various complex substrates.

Quorum sensing inhibitory potential and in silico molecular docking of flavonoids and novel terpenoids from Senegalia nigrescens.

PubMed

Bodede, Olusola; Shaik, Shakira; Chenia, Hafizah; Singh, Parvesh; Moodley, Roshila

2018-04-24

Senegalia nigrescens is used in traditional medicine for the treatment of dysentery and convulsions. This study was aimed at identifying bioactive compounds from S. nigrescens and carrying out in vitro and in silico anti-quorum sensing studies on the compounds. Extracts of S. nigrescens were chromatographed repeatedly. The isolated compounds were characterised using NMR spectroscopy and mass spectrometry. The anti-quorum sensing potential of S. nigrescens crude extracts and selected phytochemicals was quantified using Chromobacterium violaceum quorum sensing-controlled violacein inhibition assays. Qualitative modulation of quorum sensing activity and signal synthesis was investigated using agar diffusion double ring assays and C. violaceum. Molecular docking was conducted to explore the binding conformations of ent-kaurene diterpenes and flavonoids into the binding sites of quorum sensing regulator proteins, CviR and CviR'. Phytochemical investigation of S. nigrescens resulted in the isolation of a new ent-kaurene diterpenoid (ent-kaur-15-en-18,20-diol) alongside ent-kaur-15-en-18-ol, being isolated for the first time from a plant species. Other compounds isolated included 30-hydroxylup-20(29)-en-3β-ol, 3β-hydroxy-20(29)-en-lupan-30-al, lupeol, stigmasterol, a long chain alcohol (tetracosan-1-ol) and three flavonoids (melanoxetin, quercetin and quercetin-3-O-methyl ether). Structures of isolated compounds were elucidated using different spectroscopic techniques including 1D and 2D NMR. Inhibition of violacein production was concentration-dependent, with 56.52% inhibition being obtained with 200 µg of quercetin-3-O-methyl ether, while 53.38% inhibition was obtained with 600 µg of quercetin. Agar diffusion double ring assays indicated CviI synthase/CviR receptor modulation by S. nigrescens phytochemicals, suggesting that quorum signal synthesis was down-regulated and/or targeting binding of signal to the receptor. The computed binding energy data suggested that the flavonoids had a stronger tendency to inhibit both CviR and CviR' with varying binding affinities. S. nigrescens crude extracts together with the novel ent-kaurenoids and flavonoids demonstrated potential anti-quorum sensing activity. S. nigrescens may thus represent a source of anti-quorum sensing therapeutic candidates for the control of existing and emerging infectious diseases. Copyright © 2018 Elsevier B.V. All rights reserved.

Evaluation of different methods to detect methicillin resistance in Staphylococcus aureus (MRSA).

PubMed

Alipour, Farzad; Ahmadi, Malahat; Javadi, Shahram

2014-01-01

The studies suggest that dogs living with human are potential risk of becoming MRSA carrier and increased risk of infections caused by MRSA. Phenotypic methods to detect methicillin resistance in Staphylococcus aureus (MRSA) are inadequate. The objective of the present study was to determine methicillin resistance in S. aureus by phenotypic susceptibility test (oxacillin disk diffusion, cefoxitin disk diffusion, oxacillin screen agar) and molecular methods (PCR as a gold standard) and the latex agglutination test for the detection of PBP2a and to evaluate the results of these tests for its sensitivity and specificity. A total of 100 swab samples were taken from muzzle site, in more contact with human, of dogs and MRSA were isolated. Oxacillin (1 μg), cefoxitin (30 μg) disk diffusion and oxacillin screen agar method were used. The isolates were also subjected to latex agglutination test for detection of PBP2a and PCR to detect mecA gene. By PCR 37% of isolates show the presence of mecA. Latex agglutination was found to be the most sensitive (97.29%) and cefoxitin disk diffusion to be the most specific (96.82%) tests for detection of MRSA. Our finding showed that combining oxacillin screen agar or cefoxitin disk diffusion with latex agglutination improves sensitivity and specificity to detect methicillin resistance S. aureus (MRSA) isolates. Copyright © 2014 King Saud Bin Abdulaziz University for Health Sciences. Published by Elsevier Ltd. All rights reserved.

Antimicrobial activity and rutin identification of honey produced by the stingless bee Melipona compressipes manaosensis and commercial honey.

PubMed

Pimentel, Renah Boanerges de Queiroz; da Costa, Cristovão Alves; Albuquerque, Patrícia Melchionna; Junior, Sergio Duvoisin

2013-07-01

Honey has been identified as a potential alternative to the widespread use of antibiotics, which are of significant concern considering the emergence of resistant bacteria. In this context, this study aimed to evaluate the antimicrobial activity of honey samples produced by a stingless bee species and by Apis sp. against pathogenic bacteria, as well as to identify the presence of phenolic compounds. Honey samples from the stingless bee M. compressipes manaosensis were collected twice, during the dry and rainy seasons. Three commercial honey samples from Apis sp. were also included in this study. Two different assays were performed to evaluate the antibacterial potential of the honey samples: agar-well diffusion and broth macrodilution. Liquid-liquid extraction was used to assess phenolic compounds from honey. HPLC analysis was performed in order to identify rutin and apigenin on honey samples. Chromatograms were recorded at 340 and 290 nm. Two honey samples were identified as having the highest antimicrobial activity using the agar diffusion method. Honey produced by Melipona compressipes manaosensis inhibited the growth of Staphylococcus aureus, Escherichia coli (0157: H7), Proteus vulgaris, Shigella sonnei and Klebsiella sp. A sample of honey produced by Apis sp. also inhibited the growth of Salmonella paratyphi. The macrodilution technique presented greater sensitivity for the antibacterial testing, since all honey samples showed activity. Flavonoid rutin was identified in the honey sample produced by the stingless bee. Honey samples tested in this work showed antibacterial activity against Gram-positive and Gram-negative bacteria. The results reported herein highlight the potential of using honey to control bacterial growth.

A simple and convenient microtiter plate assay for the detection of bactericidal antibodies to Vibrio cholerae O1 and Vibrio cholerae O139.

PubMed

Boutonnier, Alain; Dassy, Bruno; Duménil, Rémy; Guénolé, Alain; Ratsitorahina, Maherisoa; Migliani, René; Fournier, Jean-Michel

2003-12-01

It is believed that the correlate of protection for cholera can be determined by the serum vibriocidal assay. The currently available vibriocidal assays, based on the conventional agar plating technique, are labor intensive. We developed a simple and convenient microtiter plate assay for the detection of vibriocidal antibodies that is equally as efficient for Vibrio cholerae O1 and for V. cholerae O139. The addition of succinate and neotetrazolium made it possible to measure the growth of surviving bacterial target cells by monitoring a color change. We evaluated assay parameters (target strains, growth of target cells, complement source and concentration) that may affect the reproducibility of the method for V. cholerae O139. The results obtained with the microtiter plate assay were uniformly similar to those obtained with the conventional agar plating assay, when testing both the Inaba and Ogawa serotypes of V. cholerae O1. The microtiter plate assay was also convenient for measuring the activity of animal sera and mouse monoclonal antibodies.

A quantitative in vitro assay for the evaluation of phototoxic potential of topically applied materials.

PubMed

Tenenbaum, S; DiNardo, J; Morris, W E; Wolf, B A; Schnetzinger, R W

1984-10-01

A quantitative in vitro method for phototoxic evaluation of chemicals has been developed and validated. The assay uses Saccharomyces cerevisiae, seeded in an agar overlay on top of a plate count agar base. 8-Methoxy psoralen is used as a reference standard against which materials are measured. Activity is quantified by cytotoxicity measured as zones of inhibition. Several known phototoxins (heliotropine, lyral, phantolid, and bergamot oil) and photoallergens (6-methyl coumarin and musk ambrette) are used to validate the assay. An excellent correlation is observed between in vivo studies employing Hartley albino guinea pigs and the in vitro assay for several fragrance raw materials and other chemicals. The in vitro assay exhibits a greater sensitivity from 2-500 fold. For three fragrance oils, the in vitro assay detects low levels of photobiological activity while the in vivo assay is negative. Although the in vitro assay does not discriminate between phototoxins and photoallergens, it can be used for screening of raw materials so that reduction in animal usage can be achieved while maintaining the protection of the consumer.

Novel Selective Detection Method of Tumor Angiogenesis Factors Using Living Nano-Robots.

PubMed

Al-Fandi, Mohamed; Alshraiedeh, Nida; Owies, Rami; Alshdaifat, Hala; Al-Mahaseneh, Omamah; Al-Tall, Khadijah; Alawneh, Rawan

2017-07-14

This paper reports a novel self-detection method for tumor cells using living nano-robots. These living robots are a nonpathogenic strain of E. coli bacteria equipped with naturally synthesized bio-nano-sensory systems that have an affinity to VEGF, an angiogenic factor overly-expressed by cancer cells. The VEGF-affinity/chemotaxis was assessed using several assays including the capillary chemotaxis assay, chemotaxis assay on soft agar, and chemotaxis assay on solid agar. In addition, a microfluidic device was developed to possibly discover tumor cells through the overexpressed vascular endothelial growth factor (VEGF). Various experiments to study the sensing characteristic of the nano-robots presented a strong response toward the VEGF. Thus, a new paradigm of selective targeting therapies for cancer can be advanced using swimming E. coli as self-navigator miniaturized robots as well as drug-delivery vehicles.

Development, optimization and validation of a rapid colorimetric microplate bioassay for neomycin sulfate in pharmaceutical drug products.

PubMed

Francisco, Fabiane Lacerda; Saviano, Alessandro Morais; Pinto, Terezinha de Jesus Andreoli; Lourenço, Felipe Rebello

2014-08-01

Microbiological assays have been used to evaluate antimicrobial activity since the discovery of the first antibiotics. Despite their limitations, microbiological assays are widely employed to determine antibiotic potency of pharmaceutical dosage forms, since they provide a measure of biological activity. The aim of this work is to develop, optimize and validate a rapid colorimetric microplate bioassay for the potency of neomycin in pharmaceutical drug products. Factorial and response surface methodologies were used in the development and optimization of the choice of microorganism, culture medium composition, amount of inoculum, triphenyltetrazolium chloride (TTC) concentration and neomycin concentration. The optimized bioassay method was validated by the assessment of linearity (range 3.0 to 5.0μg/mL, r=0.998 and 0.994 for standard and sample curves, respectively), precision (relative standard deviation (RSD) of 2.8% and 4.0 for repeatability and intermediate precision, respectively), accuracy (mean recovery=100.2%) and robustness. Statistical analysis showed equivalency between agar diffusion microbiological assay and rapid colorimetric microplate bioassay. In addition, microplate bioassay had advantages concerning the sensitivity of response, time of incubation, and amount of culture medium and solutions required. Copyright © 2014 Elsevier B.V. All rights reserved.

Biological studies on Brazilian plants used in wound healing.

PubMed

Schmidt, C; Fronza, M; Goettert, M; Geller, F; Luik, S; Flores, E M M; Bittencourt, C F; Zanetti, G D; Heinzmann, B M; Laufer, S; Merfort, I

2009-04-21

n-Hexanic and ethanolic extracts from twelve plants (Brugmansia suaveolens Brecht. et Presl., Eupatorium laevigatum Lam., Galinsoga parviflora Cav., Iresine herbstii Hook., Kalanchöe tubiflora Hamet-Ahti, Petiveria alliacea L., Pluchea sagittalis (Lam.) Cabrera, Piper regnellii DC., Schinus molle L., Sedum dendroideum Moç et Sessé ex DC., Waltheria douradinha St. Hill., Xanthium cavanillesii Schouw.) used in traditional South Brazilian medicine as wound healing agents were investigated in various biological assays, targeting different aspects in this complex process. The extracts were investigated on NF-kappaB DNA binding, p38alpha MAPK, TNF-alpha release, direct elastase inhibition and its release as well as on caspase-3. Fibroblasts migration to and proliferation into the wounded monolayers were evaluated in the scratch assay, the agar diffusion test for antibacterial and the MTT assay for cytotoxic effects. The hydrophilic extracts from Galinsoga parviflora, Petiveria alliacea, Schinus molle, Waltheria douradinha and Xanthium cavanillesii as well as the lipophilic extract of Waltheria douradinha turned out to be the most active ones. These results increase our knowledge on the wound healing effects of the investigated medicinal plants. Further studies are necessary to find out the effective secondary metabolites responsible for the observed effects.

Susceptibilities of Norwegian Candida albicans strains to fluconazole: emergence of resistance. The Norwegian Yeast Study Group.

PubMed Central

Sandven, P; Bjørneklett, A; Maeland, A

1993-01-01

All Candida albicans isolates in Norwegian microbiological laboratories in 1991 judged clinically important (except vaginal isolates) were collected. The isolates were tested for susceptibility to fluconazole with an agar dilution test and a commercially available agar diffusion test. A total of 212 strains (95%) were susceptible to fluconazole, and MICs for most of the strains (92%) were < or = 1.56 micrograms/ml. The agar diffusion test using a 15-micrograms tablet and a 48-h incubation period separated resistant from susceptible strains with a wide margin. The only exception was a strain for which the MIC was 6.25 micrograms/ml. The difference in zone size between the resistant and the susceptible populations of strains was 11 mm. Accordingly, it appears that the agar diffusion test is an appropriate method for detecting fluconazole resistance. The 12 fluconazole-resistant isolates originated from eight AIDS patients with oral or esophageal Candida infections. Seven of the patients had been given fluconazole for 1 month or more, often as self medication. Four had infections that were clinically resistant to fluconazole; one additional patient responded only when the dose was increased. All isolates recovered from these patients were analyzed by multilocus enzyme electrophoresis. The 12 C. albicans isolates belonged to five electrophoretic types, but three of four patients attending one hospital had isolates belonging to one electrophoretic type. One possible explanation for this finding could be that a nosocomial spread of resistant strains has occurred. PMID:8285631

Demonstrating Diffusion

ERIC Educational Resources Information Center

Foy, Barry G.

1977-01-01

Two demonstrations are described. Materials and instructions for demonstrating movement of molecules into cytoplasm using agar blocks, phenolphthalein, and sodium hydroxide are given. A simple method for demonstrating that the rate of diffusion of a gas is inversely proportional to its molecular weight is also presented. (AJ)

Preparation of an agar-silver nanoparticles (A-AgNp) film for increasing the shelf-life of fruits.

PubMed

Gudadhe, Janhavi A; Yadav, Alka; Gade, Aniket; Marcato, Priscyla D; Durán, Nelson; Rai, Mahendra

2014-12-01

Preparation of protective coating possessing antimicrobial properties is present day need as they increase the shelf life of fruits and vegetables. In the present study, preparation of agar-silver nanoparticle film for increasing the shelf life of fruits is reported. Silver nanoparticles (Ag-NPs) biosynthesised using an extract of Ocimum sanctum leaves, were mixed with agar-agar to prepare an agar-silver nanoparticles (A-AgNp) film. This film was surface-coated over the fruits, Citrus aurantifolium (Thornless lime) and Pyrus malus (Apple), and evaluated for the determination of antimicrobial activity of A-AgNp films using disc diffusion method, weight loss and shelf life of fruits. This study demonstrates that these A-AgNp films possess antimicrobial activity and also increase the shelf life of fruits.

Spreading of nonmotile bacteria on a hard agar plate: Comparison between agent-based and stochastic simulations

NASA Astrophysics Data System (ADS)

Rana, Navdeep; Ghosh, Pushpita; Perlekar, Prasad

2017-11-01

We study spreading of a nonmotile bacteria colony on a hard agar plate by using agent-based and continuum models. We show that the spreading dynamics depends on the initial nutrient concentration, the motility, and the inherent demographic noise. Population fluctuations are inherent in an agent-based model, whereas for the continuum model we model them by using a stochastic Langevin equation. We show that the intrinsic population fluctuations coupled with nonlinear diffusivity lead to a transition from a diffusion limited aggregation type of morphology to an Eden-like morphology on decreasing the initial nutrient concentration.

Agar Underlay Method for Recovery of Sublethally Heat-Injured Bacteria

PubMed Central

Kang, D. H.; Siragusa, G. R.

1999-01-01

A method of recovering sublethally heat-injured bacteria was developed. The procedure (termed the agar underlay method) uses a nonselective agar underlaid with a selective medium. In a two-chambered petri dish, the Lutri plate (LP), a nonselective agar is inoculated with a population of sublethally heat-injured bacteria. After a 2-h repair incubation period, selective agar is added to the bottom chamber of the LP and incubated. By diffusing through the nonselective top agar, selective agents from the underlay medium impart selectivity to the system. By the agar underlay method, recovery rates of the heat-injured food-borne pathogens Escherichia coli O157:H7 and Salmonella typhimurium were not different (P > 0.05) from recovery rates determined with nonselective media. Sublethally heat-injured cells (60°C for 1.5 min in buffer or 80°C for 30 s on meat surfaces) grew and produced a typical colony morphology and color reaction when the agar underlay procedure was used with the appropriate respective selective agars. Unlike agar overlay methods for injury repair, the agar underlay procedure allows the typical selective-medium colony morphology to develop and allows colonies to be more easily picked for further characterization. Higher recovery rates of heat-injured fecal enterococci from bovine fecal samples and total coliforms from animal waste lagoons were obtained by the agar underlay method with selective agars than by direct plating on the respective selective media. PMID:10583985

Compliance of clinical microbiology laboratories in the United States with current recommendations for processing respiratory tract specimens from patients with cystic fibrosis.

PubMed

Zhou, Juyan; Garber, Elizabeth; Desai, Manisha; Saiman, Lisa

2006-04-01

Respiratory tract specimens from patients with cystic fibrosis (CF) require unique processing by clinical microbiology laboratories to ensure detection of all potential pathogens. The present study sought to determine the compliance of microbiology laboratories in the United States with recently published recommendations for CF respiratory specimens. Microbiology laboratory protocols from 150 of 190 (79%) CF care sites were reviewed. Most described the use of selective media for Burkholderia cepacia complex (99%), Staphylococcus aureus (82%), and Haemophilus influenzae (89%) and identified the species of all gram-negative bacilli (87%). Only 52% delineated the use of agar diffusion assays for susceptibility testing of Pseudomonas aeruginosa. Standardizing laboratory practices will improve treatment, infection control, and our understanding of the changing epidemiology of CF microbiology.

Evaluation of antibacterial, antioxidant and GC-MS analysis of ethanolic seed extract of Myristica dactyloides

NASA Astrophysics Data System (ADS)

Bhavani, R.; Vijayalakshmi, R.; Venkat Kumar, S.; Rajeshkumar, S.

2017-11-01

In this present investigation we analysed the antimicrobial and antioxidant activities of ethanol extract of Myristica dactyloides. The antimicrobial activity of the ethanol extract was evaluated by the agar well diffusion method against of E. coli, Klebsiella pneumonia, Streptococcus sp, and Staphylococcus aureus at different concentrations. The antibacterial activity showed the result in a dose-dependent manner. The free radical scavenging was evaluated against DPPH, hydroxyl, and nitric oxide radicals. In DPPH, hydroxyl and nitric oxide scavenging assay showed the IC 50 value of the extract was found to be 20 μg/ml, 48.25 and 30 μg/ml, respectively. The plant can be considered as promising antioxidant properties with high potential value for drug development for various diseases.

Polyphenolic extracts of edible flowers incorporated onto atelocollagen matrices and their effect on cell viability.

PubMed

López-García, Jorge; Kuceková, Zdenka; Humpolíček, Petr; Mlček, Jiři; Sáha, Petr

2013-10-30

The phenolic extract of chives flowers (Allium schoenoprasum, Liliaceae), introduced Sage (Salvia pratensis, Lamiaceae), European elderberry (Sambucus nigra, Caprifoliaceae) and common dandelion (Taraxacum officinale, Asteraceae) were characterised by High Performance Liquid Chromatography and incorporated in different concentrations onto atelocollagen thin films. In order to assess the biological impact of these phenolic compounds on cell viability, human immortalised non-tumorigenic keratinocyte cell line was seeded on the thin films and cell proliferation was determined by using an MTT assay. In addition, their antimicrobial activity was estimated by using an agar diffusion test. Data indicated the concomitance between cell viability and concentration of polyphenols. These findings suggest that these phenolic-endowed atelocollagen films might be suitable for tissue engineering applications, on account of the combined activity of polyphenols and collagen.

Comparison of commercial enzyme-linked immunosorbent assay kits with agar gel precipitation and hemagglutination-inhibition tests for detecting antibodies to avian influenza viruses.

PubMed

Shiraishi, Rikiya; Nishiguchi, Akiko; Tsukamoto, Kenji; Muramatsu, Masatake

2012-09-01

We evaluated the utility of 5 commercial enzyme-linked immunosorbent assay (ELISA) kits for detecting antibodies to avian influenza viruses. The sensitivities and specificities of the ELISA kits were compared with those of the agar gel precipitation (AGP) and hemagglutination-inhibition (HI) tests. The results suggest that some ELISA kits might not be suitable for monitoring during the early stages of avian influenza virus infections. Therefore, ELISA kits should only be used in conjunction with a profound knowledge about monitoring of avian influenza.

Phototoxicity assessment of drugs and cosmetic products using E. coli.

PubMed

Verma, K; Agrawal, N; Misra, R B; Farooq, M; Hans, R K

2008-02-01

A gram negative bacteria Escherichia coli (Dh5alpha strain) was developed as an alternate test system of phototoxicity. Eight drugs (antibiotics) and cosmetic products (eight face creams) were examined for their phototoxicity using this test system. Five known phototoxic compounds were used to validate the test system. UVA-radiation induced phototoxicity of these compounds was tested by agar gel diffusion assay. Decrease in colony forming units (CFU) was taken as an end point of phototoxicity. The phototoxic compounds and antibiotics produced significant reduction in CFU (p<0.001) at 80 microg/ml concentrations under exposure to UVA-radiation (5.4-10.8 J/cm(2)). One face cream was found phototoxic and produced significant decrease in CFU of E. coli at 1.0mg/ml concentration under UVA exposure (10.8 J/cm(2)). The minimum effective concentration of tetracycline and dose of UVA-radiation were also determined by observing growth inhibition of E. coli through disc diffusion assay. The observations suggested that E. coli can be used as an alternative test system for phototoxicity evaluation of chemicals. A battery of test systems is required to conclude the toxic/phototoxic potential of a chemical agent. In view of the speed, easiness, sensitivity and low cost, E. coli is introduced as one of the alternate test system for phototoxicity studies in safety evaluation of various chemical ingredients or formulations used in cosmetics and drugs.

Nitrate analogs as attractants for soybean cyst nematode.

PubMed

Hosoi, Akito; Katsuyama, Tsutomu; Sasaki, Yasuyuki; Kondo, Tatsuhiko; Yajima, Shunsuke; Ito, Shinsaku

2017-08-01

Soybean cyst nematode (SCN) Heterodera glycines Ichinohe, a plant parasite, is one of the most serious pests of soybean. In this paper, we report that SCN is attracted to nitrate and its analogs. We performed attraction assays to screen for novel attractants for SCN and found that nitrates were attractants for SCN and SCN recognized nitrate gradients. However, attraction of SCN to nitrates was not observed on agar containing nitrate. To further elucidate the attraction mechanism in SCN, we performed attraction assays using nitrate analogs ([Formula: see text], [Formula: see text], [Formula: see text]). SCN was attracted to all nitrate analogs; however, attraction of SCN to nitrate analogs was not observed on agar containing nitrate. In contrast, SCN was attracted to azuki root, irrespective of presence or absence of nitrate in agar media. Our results suggest that the attraction mechanisms differ between plant-derived attractant and nitrate.

Novel Selective Detection Method of Tumor Angiogenesis Factors Using Living Nano-Robots

PubMed Central

Alshraiedeh, Nida; Owies, Rami; Alshdaifat, Hala; Al-Mahaseneh, Omamah; Al-Tall, Khadijah; Alawneh, Rawan

2017-01-01

This paper reports a novel self-detection method for tumor cells using living nano-robots. These living robots are a nonpathogenic strain of E. coli bacteria equipped with naturally synthesized bio-nano-sensory systems that have an affinity to VEGF, an angiogenic factor overly-expressed by cancer cells. The VEGF-affinity/chemotaxis was assessed using several assays including the capillary chemotaxis assay, chemotaxis assay on soft agar, and chemotaxis assay on solid agar. In addition, a microfluidic device was developed to possibly discover tumor cells through the overexpressed vascular endothelial growth factor (VEGF). Various experiments to study the sensing characteristic of the nano-robots presented a strong response toward the VEGF. Thus, a new paradigm of selective targeting therapies for cancer can be advanced using swimming E. coli as self-navigator miniaturized robots as well as drug-delivery vehicles. PMID:28708066

On the Enhanced Antibacterial Activity of Antibiotics Mixed with Gold Nanoparticles

NASA Astrophysics Data System (ADS)

Burygin, G. L.; Khlebtsov, B. N.; Shantrokha, A. N.; Dykman, L. A.; Bogatyrev, V. A.; Khlebtsov, N. G.

2009-08-01

The bacterial action of gentamicin and that of a mixture of gentamicin and 15-nm colloidal-gold particles on Escherichia coli K12 was examined by the agar-well-diffusion method, enumeration of colony-forming units, and turbidimetry. Addition of gentamicin to colloidal gold changed the gold color and extinction spectrum. Within the experimental errors, there were no significant differences in antibacterial activity between pure gentamicin and its mixture with gold nanoparticles (NPs). Atomic absorption spectroscopy showed that upon application of the gentamicin-particle mixture, there were no gold NPs in the zone of bacterial-growth suppression in agar. Yet, free NPs diffused into the agar. These facts are in conflict with the earlier findings indicating an enhancement of the bacterial activity of similar gentamicin-gold nanoparticle mixtures. The possible causes for these discrepancies are discussed, and the suggestion is made that a necessary condition for enhancement of antibacterial activity is the preparation of stable conjugates of NPs coated with the antibiotic molecules.

Structural, morphological, optical and biological properties of pure ZnO and agar/zinc oxide nanocomposites.

PubMed

Magesh, G; Bhoopathi, G; Nithya, N; Arun, A P; Ranjith Kumar, E

2018-05-26

In this work, ZnO nanoparticles were prepared by in situ chemical precipitation method in the presence of Agar biopolymer. The influence of Agar concentrations on the structural, morphological and optical properties of ZnO have been investigated. The XRD pattern of Pure ZnO and Agar/ZnO nanocomposites indicates the hexagonal wurtzite phase of ZnO. The crystallite size of pure ZnO and Agar/ZnO nanocomposites was found to be in the range of 35.5 to 19.73 nm. Pure ZnO and Agar/ZnO nanocomposites showed nanospheroid and nanopaddy shaped morphology from FESEM studies. The interplanar distance observed from the HRTEM image confirms the plane of the prepared material. The elemental composition of the samples were characterized by EDX. The optical properties of Pure ZnO and Agar/ZnO nanocomposites were characterized by UV, FTIR and PL. The band gap of Agar/ZnO nanocomposites were varied with the Agar concentration. Oxygen vacancy induced photoluminescence of ZnO are observed and its intensity is found to be increased linearly with the Agar concentration. The antibacterial activity of ZnO and Agar/ZnO nanocomposites was evaluated by disc diffusion method against Gram-positive (B.subtilis) and Gram-negative (P. aeruginosa) bacteria. The cytotoxicity of Agar/ZnO nanocomposites was studied against Normal (L929) and Breast cancer cell line (MB231). The result of this investigation reveals that the Agar/ZnO nanocomposites deliver a dose dependent toxicity in normal and cancer cell line. Copyright © 2018. Published by Elsevier B.V.

Inhibitory effect of sodium hypochlorite and chlorhexidine digluconate in clinical isolates of Sporothrix schenckii.

PubMed

Madrid, Isabel Martins; Mattei, Antonella Souza; Santin, Rosema; dos Reis Gomes, Angelita; Cleff, Marlete Brum; Meireles, Mário Carlos Araújo

2012-05-01

The susceptibility of Sporothrix schenckii isolates from clinical cases of canine, feline and human sporotrichosis, and from the environment, was evaluated with 4% sodium hypochlorite and 6.6% chlorhexidine digluconate using the broth microdilution, agar diffusion and direct exposure techniques. The minimal inhibitory concentration was smaller than 0.8% for chlorhexidine digluconate and between 8% and 4% for sodium hypochlorite. Inhibition zones were not found in agar diffusion for sodium hypochlorite, and zones averaging 1.9 mm were found for chlorhexidine digluconate. In the direct exposure test, sodium hypochlorite demonstrated best performance at 20 min of contact, as chlorhexidine digluconate presented little antimicrobial activity. © 2011 Blackwell Verlag GmbH.

In-vitro and in-vivo anti-Trichophyton activity of essential oils by vapour contact.

PubMed

Inouye, S; Uchida, K; Yamaguchi, H

2001-05-01

The minimum inhibitory doses (MIDs) of essential oils by vapour contact to inhibit the growth of Trichophyton mentagrophytes and Trichophyton rubrum on agar medium were determined using airtight boxes. Among seven essential oils examined, cinnamon bark oil showed the least MID, followed by lemongrass, thyme and perilla oils. Lavender and tea tree oils showed moderate MID, and citron oil showed the highest MID, being 320 times higher than that of cinnamon bark oil. The MID values were less than the minimum inhibitory concentration (MIC) values determined by agar dilution assay. Furthermore, the minimum agar concentration (MAC) of essential oils absorbed from vapour was determined at the time of MID determination as the second antifungal measure. The MAC value by vapour contact was 1.4 to 4.7 times less than the MAC remaining in the agar at the time of MIC determination by agar dilution assay. Using selected essential oils, the anti-Trichophyton activity by vapour contact was examined in more detail. Lemongrass, thyme and perilla oils killed the conidia, inhibited germination and hyphal elongation at 1-4 micrograms ml-1 air, whereas lavender oil was effective at 40-160 micrograms ml-1 air. The in-vivo efficacy of thyme and perilla oils by vapour contact was shown against an experimental tinea pedis in guinea pigs infected with T. mentagrophytes. These results indicated potent anti-Trichophyton action of essential oils by vapour contact.

The upper surface of an Escherichia coli swarm is stationary.

PubMed

Zhang, Rongjing; Turner, Linda; Berg, Howard C

2010-01-05

When grown in a rich medium on agar, many bacteria elongate, produce more flagella, and swim in a thin film of fluid over the agar surface in swirling packs. Cells that spread in this way are said to swarm. The agar is a solid gel, with pores smaller than the bacteria, so the swarm/agar interface is fixed. Here we show, in experiments with Escherichia coli, that the swarm/air interface also is fixed. We deposited MgO smoke particles on the top surface of an E. coli swarm near its advancing edge, where cells move in a single layer, and then followed the motion of the particles by dark-field microscopy and the motion of the underlying cells by phase-contrast microscopy. Remarkably, the smoke particles remained fixed (diffusing only a few micrometers) while the swarming cells streamed past underneath. The diffusion coefficients of the smoke particles were smaller over the virgin agar ahead of the swarm than over the swarm itself. Changes between these two modes of behavior were evident within 10-20 microm of the swarm edge, indicating an increase in depth of the fluid in advance of the swarm. The only plausible way that the swarm/air interface can be fixed is that it is covered by a surfactant monolayer pinned at its edges. When a swarm is exposed to air, such a monolayer can markedly reduce water loss. When cells invade tissue, the ability to move rapidly between closely opposed fixed surfaces is a useful trait.

Synthesis and Bioactivity Evaluation of Novel 2-Salicyloylbenzofurans as Antibacterial Agents.

PubMed

Phan, Phuong-Thuy T; Nguyen, Thu-Trang T; Nguyen, Hong-Nhung T; Le, Bao-Khanh N; Vu, Thao T; Tran, Dong C; Pham, Tuan-Anh N

2017-04-25

In order to discover new antibacterial agents, series of 2-salicyloylbenzofuran derivatives were designed, synthesized and evaluated for their antibacterial activities against three Gram-(+) strains (methicillin-sensitive Staphylococcus aureus (MSSA) ATCC 29213, methicillin-resistant Staphylococcus aureus (MRSA) ATCC 43300, and Streptococcus faecalis ( S. faecalis ) ATCC 29212) and one Gram-(-) strain ( Escherichia coli (E. coli) ATCC 25922). The 2-salicyloylbenzofuran heterocycles were generated by Rap-Stoermer condensation of salicylaldehydes with phenacyl bromides and then converted to diverse O -ether derivatives by Williamson synthesis. The targeted products were screened for in vitro qualitative (zone of inhibition) and quantitative (MIC) antibacterial activities by agar well diffusion assay and agar dilution method. Amongst the compounds, those bearing carboxylic acid functional group were found to exhibit reasonable activity against Gram-(+) bacterial strains including S. faecalis , MSSA and MRSA with the most potent antibacterial agent 8h (MICs = 0.06-0.12 mM). Besides, the 2-salicyloylbenzofurans partly displayed inhibitory activity against MRSA with the best MICs = 0.14 mM ( 8f ) and 0.12 mM ( 8h ). Finally, the antibacterial results preliminarily suggested that the substituent bearing carboxylic acid group at salicyloyl-C2 and the bromine atoms on the benzofuran moiety seem to be the functionality necessary for antibacterial activities.

Antimicrobial activity of commercially available essential oils against Streptococcus mutans.

PubMed

Chaudhari, Lalit Kumar D; Jawale, Bhushan Arun; Sharma, Sheeba; Sharma, Hemant; Kumar, C D Mounesh; Kulkarni, Pooja Adwait

2012-01-01

Many essential oils have been advocated for use in complementary medicine for bacterial and fungal infections. However, few of the many claims of therapeutic efficacy have been validated adequately by either in vitro testing or in vivo clinical trials. To study the antibacterial activity of nine commercially available essential oils against Streptococcus mutans in vitro and to compare the antibacterial activity between each material. Nine pure essential oils; wintergreen oil, lime oil, cinnamon oil, spearmint oil, peppermint oil, lemongrass oil, cedarwood oil, clove oil and eucalyptus oil were selected for the study. Streptococcus mutans was inoculated at 37ºC and seeded on blood agar medium. Agar well diffusion assay was used to measure antibacterial activity. Zone of inhibition was measured around the filter paper in millimeters with vernier caliper. Cinnamon oil showed highest activity against Streptococcus mutans followed by lemongrass oil and cedarwood oil. Wintergreen oil, lime oil, peppermint oil and spearmint oil showed no antibacterial activity. Cinnamon oil, lemongrass oil, cedarwood oil, clove oil and eucalyptus oil exhibit antibacterial property against S. mutans. The use of these essential oils against S. mutans can be a viable alternative to other antibacterial agents as these are an effective module used in the control of both bacteria and yeasts responsible for oral infections.

Antibacterial efficacy of Mangifera indica L. kernel and Ocimum sanctum L. leaves against Enterococcus faecalis dentinal biofilm

PubMed Central

Subbiya, Arunajatesan; Mahalakshmi, Krishnan; Pushpangadan, Sivan; Padmavathy, Kesavaram; Vivekanandan, Paramasivam; Sukumaran, Vridhachalam Ganapathy

2013-01-01

Introduction: The Enterococcus faecalis biofilm in the root canal makes it difficult to be eradicated by the conventional irrigants with no toxicity to the tissues. Hence, plant products with least side effects are explored for their use as irrigants in the root canal therapy. Aim: To evaluate and compare the antibacterial efficacy of Mangifera indica L. kernel (mango kernel) and Ocimum sanctum L. leaves (tulsi) extracts with conventional irrigants (5% sodium hypochlorite (NaOCl) and 2% chlorhexidine) against E. faecalis dentinal biofilm. Materials and Methods: Agar diffusion and broth microdilution assay was performed with the herbal extracts and conventional irrigants (2% chlorhexidine and 5% NaOCl) against E. faecalis planktonic cells. The assay was extended onto 3 week E. faecalis dentinal biofilm. Results: Significant reduction of colony forming units (CFU)/mL was observed for the herbal groups and the antibacterial activity of the herbal groups was at par with 5% NaOCl. Conclusions: The antibacterial activity of these herbal extracts is found to be comparable with that of conventional irrigants both on the biofilm and planktonic counterparts. PMID:24082577

Volatile oil from striped African pepper (Xylopia parviflora, Annonaceae) possesses notable chemopreventive, anti-inflammatory and antimicrobial potential.

PubMed

Woguem, Verlaine; Fogang, Hervet P D; Maggi, Filippo; Tapondjou, Léon A; Womeni, Hilaire M; Quassinti, Luana; Bramucci, Massimo; Vitali, Luca A; Petrelli, Dezemona; Lupidi, Giulio; Papa, Fabrizio; Vittori, Sauro; Barboni, Luciano

2014-04-15

Fruits of Xylopia parviflora, well known as striped African pepper, are sold in the Cameroonian markets as a flavouring ingredient to make traditional soups. The essential oil hydrodistilled from fruits was analysed for in vitro biological activities, namely cytotoxic, anti-inflammatory, antimicrobial and antioxidant, by MTT, nitric oxide inhibitory assay, agar disc diffusion method, and DPPH and ABTS assays. The essential oil composition, analysed by GC and GC-MS, was dominated by monoterpene hydrocarbons (50.0%) responsible for the pepper odour, such as β-pinene (34.0%) and α-pinene (10.3%). The oil induced a strong inhibitory effect on tumour cells MDA-MB 231 and HCT116, with inhibition values close to those of cisplatin. A dose-dependent decrease in NO production was noted in RAW 264.7 macrophages treated with the oil, revealing a promising anti-inflammatory potential. The essential oil showed a measurable antimicrobial activity against all the species tested, while the radical scavenging activity was low. Copyright © 2013 Elsevier Ltd. All rights reserved.

Detection of Group B Streptococci in Lim Broth by Use of Group B Streptococcus Peptide Nucleic Acid Fluorescent In Situ Hybridization and Selective and Nonselective Agars▿

PubMed Central

Montague, Naomi S.; Cleary, Timothy J.; Martinez, Octavio V.; Procop, Gary W.

2008-01-01

The sensitivity, specificity, and positive and negative predictive values for the detection of group B streptococci from Lim enrichment broth with sheep blood agar (SBA), with selective Streptococcus agar (SSA), and by a peptide nucleic acid fluorescent in situ hybridization (PNA FISH) assay were as follows: for culture on SBA, 68.4%, 100%, 100%, and 87.9%, respectively; for culture on SSA, 85.5%, 100%, 100%, and 94.1%, respectively; and for the PNA FISH assay, 97.4%, 98.3%, 96.1%, and 98.9%, respectively. PMID:18667597

New closed tube loop mediated isothermal amplification assay for prevention of product cross-contamination

PubMed Central

Karthik, K.; Rathore, Rajesh; Thomas, Prasad; Arun, T.R.; Viswas, K.N.; Dhama, Kuldeep; Agarwal, R.K.

2014-01-01

Loop mediated isothermal amplification (LAMP) assay, a promising diagnostic test, has been developed for detection of different pathogens of human as well as animals. Various positive points support its use as a field level test but the major problem is product cross contamination leading to false positive results. Different methods were adopted by various researchers to control this false positive amplification due to cross contamination but all have their own advantages and disadvantages. A new closed tube LAMP assay based on agar dye capsule was developed in the present study and this technique has some advantages over the other closed tube technique.•Agar at the concentration of 1.5% was used to sandwich SYBR green dye I with the aid of intradermal syringe. This agar dye capsule was placed over the LAMP reaction mixture before it was amplified.•To eliminate the hazardous nature of Ultra Violet (UV) light during result visualization of LAMP products, the present study demonstrates the use of Light Emitting Diode (LED) lights for result visualization.•LAMP was carried out for Brucella species detection using this modified techniques yielding good results without any cross contamination and LED showed similar fluorescence compared to UV. PMID:26150945

New closed tube loop mediated isothermal amplification assay for prevention of product cross-contamination.

PubMed

Karthik, K; Rathore, Rajesh; Thomas, Prasad; Arun, T R; Viswas, K N; Dhama, Kuldeep; Agarwal, R K

2014-01-01

Loop mediated isothermal amplification (LAMP) assay, a promising diagnostic test, has been developed for detection of different pathogens of human as well as animals. Various positive points support its use as a field level test but the major problem is product cross contamination leading to false positive results. Different methods were adopted by various researchers to control this false positive amplification due to cross contamination but all have their own advantages and disadvantages. A new closed tube LAMP assay based on agar dye capsule was developed in the present study and this technique has some advantages over the other closed tube technique.•Agar at the concentration of 1.5% was used to sandwich SYBR green dye I with the aid of intradermal syringe. This agar dye capsule was placed over the LAMP reaction mixture before it was amplified.•To eliminate the hazardous nature of Ultra Violet (UV) light during result visualization of LAMP products, the present study demonstrates the use of Light Emitting Diode (LED) lights for result visualization.•LAMP was carried out for Brucella species detection using this modified techniques yielding good results without any cross contamination and LED showed similar fluorescence compared to UV.

Antibiotic susceptibility of methicillin-resistant staphylococci (MRS) of food origin: A comparison of agar disc diffusion method and a commercially available miniaturized test.

PubMed

Buzón-Durán, Laura; Capita, Rosa; Alonso-Calleja, Carlos

2018-06-01

Methicillin-resistant staphylococci (MRS) are a major concern to public and animal health. Thirty MRS (Staphylococcus aureus, S. cohnii, S. epidermidis, S. haemolyticus, S. hominis, S. lentus, S. lugdunensis, S. sciuri, and S. xylosus) isolates from meat and poultry preparations were tested for antimicrobial susceptibility to 11 antimicrobials (belonging to seven different categories) of clinical significance using both the standard agar disc diffusion method and a commercially available miniaturized system (Sensi Test Gram-positive). It is worth stressing that 16 isolates (53.33%) exhibited an extensively drug-resistant phenotype (XDR). The average number of resistances per strain was 4.67. These results suggest that retail meat and poultry preparations are a likely vehicle for the transmission of multi-drug resistant MRS. Resistance to erythromycin was the commonest finding (76.67% of strains), followed by tobramycin, ceftazidime (66.67%), ciprofloxacin (56.67%) and fosfomycin (53.33%). An agreement (kappa coefficient) of 0.64 was found between the two testing methods. Using the agar disc diffusion as the reference method, the sensitivity, specificity and accuracy of the miniaturized test were 98.44%, 69.44% and 83.33%, respectively. Most discrepancies between the two methods were due to isolates that were susceptible according to the disc diffusion method but resistant according to the miniaturized test (false positives). Copyright © 2017. Published by Elsevier Ltd.

Colony patterning and collective hyphal growth of filamentous fungi

NASA Astrophysics Data System (ADS)

Matsuura, Shu

2002-11-01

Colony morphology of wild and mutant strains of Aspergillus nidulans at various nutrient and agar levels was investigated. Two types of colony patterning were found for these strains. One type produced uniform colonies at all nutrient and agar levels tested, and the other exhibited morphological change into disordered ramified colonies at low nutrient levels. Both types showed highly condensed compact colonies at high nutrient levels on low agar media that was highly diffusive. Disordered colonies were found to develop with low hyphal extension rates at low nutrient levels. To understand basic pattern selection rules, a colony model with three parameters, i.e., the initial nutrient level and the step length of nutrient random walk as the external parameters, and the frequency of nutrient uptake as an internal parameter, was constructed. At low nutrient levels, with decreasing nutrient uptake frequency under diffusive conditions, the model colony exhibited onsets of disordered ramification. Further, in the growth process of A. nidulans, reduction of hyphal extension rate due to a population effect of hyphae was found when hyphae form three-dimensional dense colonies, as compared to the case in which hyphal growth was restricted into two-dimensional space. A hyphal population effect was introduced in the colony model. Thickening of colony periphery due to the population effect became distinctive as the nutrient diffusion effect was raised at high nutrient levels with low hyphal growth rate. It was considered that colony patterning and onset of disorder were strongly governed by the combination of nutrient diffusion and hyphal growth rate.

Initial cytotoxicity assays of media for sulfate-reducing bacteria: An endodontic biopharmaceutical product under development.

PubMed

Heggendorn, Fabiano Luiz; Silva, Gabriela Cristina de Carvalho; Cardoso, Elisama Azevedo; Castro, Helena Carla; Gonçalves, Lúcio Souza; Dias, Eliane Pedra; Lione, Viviane de Oliveira Freitas; Lutterbach, Márcia Teresa Soares

2016-01-01

This study assessed the cell viability of the inoculation vehicle of BACCOR (a combination of sulfate-reducing bacteria plus a culture media for bacteria), a biopharmaceutical product under development for dental use as aid in fractured endodontic file removal from the root canal. Different culture media for bacteria were evaluated: modified Postgate E (MCP-E mod), Modified Postgate E without Agar-agar (MCP-E w/Ag), Postgate C with Agar-agar (MCP-C Ag) and Postgate C without Agar-agar (MCP-C w/Ag). Cytotoxicity was quantified by the MTT test, exposing L929 and Vero cell lines to the vehicles over 24 h. The exposure of L929 cell line to MCP-E w/Ag resulted in biocompatibility (52% cell viability), while the exposure of the Vero kidney line revealed only MCP-E mod as cytotoxic. When diluted, all the vehicles showed biocompatibility with both cell lines. MCP-E w/Ag was the vehicle chosen for BACCOR, because of its biocompatibility with the cells used.

Antimicrobial activity and rutin identification of honey produced by the stingless bee Melipona compressipes manaosensis and commercial honey

PubMed Central

2013-01-01

Background Honey has been identified as a potential alternative to the widespread use of antibiotics, which are of significant concern considering the emergence of resistant bacteria. In this context, this study aimed to evaluate the antimicrobial activity of honey samples produced by a stingless bee species and by Apis sp. against pathogenic bacteria, as well as to identify the presence of phenolic compounds. Methods Honey samples from the stingless bee M. compressipes manaosensis were collected twice, during the dry and rainy seasons. Three commercial honey samples from Apis sp. were also included in this study. Two different assays were performed to evaluate the antibacterial potential of the honey samples: agar-well diffusion and broth macrodilution. Liquid-liquid extraction was used to assess phenolic compounds from honey. HPLC analysis was performed in order to identify rutin and apigenin on honey samples. Chromatograms were recorded at 340 and 290 nm. Results Two honey samples were identified as having the highest antimicrobial activity using the agar diffusion method. Honey produced by Melipona compressipes manaosensis inhibited the growth of Staphylococcus aureus, Escherichia coli (0157: H7), Proteus vulgaris, Shigella sonnei and Klebsiella sp. A sample of honey produced by Apis sp. also inhibited the growth of Salmonella paratyphi. The macrodilution technique presented greater sensitivity for the antibacterial testing, since all honey samples showed activity. Flavonoid rutin was identified in the honey sample produced by the stingless bee. Conclusions Honey samples tested in this work showed antibacterial activity against Gram-positive and Gram-negative bacteria. The results reported herein highlight the potential of using honey to control bacterial growth. PMID:23815879

An interlaboratory study on efficient detection of Shiga toxin-producing Escherichia coli O26, O103, O111, O121, O145, and O157 in food using real-time PCR assay and chromogenic agar.

PubMed

Hara-Kudo, Yukiko; Konishi, Noriko; Ohtsuka, Kayoko; Iwabuchi, Kaori; Kikuchi, Rie; Isobe, Junko; Yamazaki, Takumiko; Suzuki, Fumie; Nagai, Yuhki; Yamada, Hiroko; Tanouchi, Atsuko; Mori, Tetsuya; Nakagawa, Hiroshi; Ueda, Yasufumi; Terajima, Jun

2016-08-02

To establish an efficient detection method for Shiga toxin (Stx)-producing Escherichia coli (STEC) O26, O103, O111, O121, O145, and O157 in food, an interlaboratory study using all the serogroups of detection targets was firstly conducted. We employed a series of tests including enrichment, real-time PCR assays, and concentration by immunomagnetic separation, followed by plating onto selective agar media (IMS-plating methods). This study was particularly focused on the efficiencies of real-time PCR assays in detecting stx and O-antigen genes of the six serogroups and of IMS-plating methods onto selective agar media including chromogenic agar. Ground beef and radish sprouts samples were inoculated with the six STEC serogroups either at 4-6CFU/25g (low levels) or at 22-29CFU/25g (high levels). The sensitivity of stx detection in ground beef at both levels of inoculation with all six STEC serogroups was 100%. The sensitivity of stx detection was also 100% in radish sprouts at high levels of inoculation with all six STEC serogroups, and 66.7%-91.7% at low levels of inoculation. The sensitivity of detection of O-antigen genes was 100% in both ground beef and radish sprouts at high inoculation levels, while at low inoculation levels, it was 95.8%-100% in ground beef and 66.7%-91.7% in radish sprouts. The sensitivity of detection with IMS-plating was either the same or lower than those of the real-time PCR assays targeting stx and O-antigen genes. The relationship between the results of IMS-plating methods and Ct values of real-time PCR assays were firstly analyzed in detail. Ct values in most samples that tested negative in the IMS-plating method were higher than the maximum Ct values in samples that tested positive in the IMS-plating method. This study indicates that all six STEC serogroups in food contaminated with more than 29CFU/25g were detected by real-time PCR assays targeting stx and O-antigen genes and IMS-plating onto selective agar media. Therefore, screening of stx and O-antigen genes followed by isolation of STECs by IMS-plating methods may be an efficient method to detect the six STEC serogroups. Copyright © 2016 Elsevier B.V. All rights reserved.

Nisin depletes ATP and proton motive force in mycobacteria.

PubMed

Chung, H J; Montville, T J; Chikindas, M L

2000-12-01

This study examined the inhibitory effect of nisin and its mode of action against Mycobacterium smegmatis, a non-pathogenic species of mycobacteria, and M. bovis-Bacill Carmette Guerin (BCG), a vaccine strain of pathogenic M. bovis. In agar diffusion assays, 2.5 mg ml(-1) nisin was required to inhibit M. bovis-BCG. Nisin caused a slow, gradual, time- and concentration-dependent decrease in internal ATP levels in M. bovis-BCG, but no ATP efflux was detected. In mycobacteria, nisin decreased both components of proton motive force (membrane potential, Delta Psi and Delta pH) in a time- and concentration-dependent manner. However, mycobacteria maintained their intracellular ATP levels during the initial time period of Delta Psi and Delta pH dissipation. These data suggest that the mechanism of nisin in mycobacteria is similar to that in food-borne pathogens.

Antimicrobial activity of Arctium lappa constituents against microorganisms commonly found in endodontic infections.

PubMed

Pereira, Juliana Vianna; Bergamo, Débora Cristina Baldoqui; Pereira, José Odair; França, Suzelei de Castro; Pietro, Rosemeire Cristina Linhares Rodrigues; Silva-Sousa, Yara T Corrêa

2005-01-01

This study evaluated in vitro the antimicrobial activity of rough extracts from leaves of Arctium lappa and their phases. The following microorganisms, commonly found in the oral cavity, specifically in endodontic infections, were used: Enterococcus faecalis, Staphylococcus aureus, Pseudomonas aeruginosa, Bacillus subtilis and Candida albicans. The agar-diffusion method allowed detection of the hexanic phase as an inhibitor of microbial growth. Bioautographic assays identified antimicrobial substances in the extract. The results showed the existence, in the rough hexanic phase and in its fractions, of constituents that have retention factors (Rf) in three distinct zones, thereby suggesting the presence of active constituents with chemical structures of different polarities that exhibited specificity against the target microorganisms. It may be concluded that the Arctium lappa constituents exhibited a great microbial inhibition potential against the tested endodontic pathogens.

Thermal characterization of magnetically aligned carbonyl iron/agar composites.

PubMed

Diaz-Bleis, D; Vales-Pinzón, C; Freile-Pelegrín, Y; Alvarado-Gil, J J

2014-01-01

Composites of magnetic particles into polymeric matrices have received increasing research interest due to their capacity to respond to external magnetic or electromagnetic fields. In this study, agar from Gelidium robustum has been chosen as natural biocompatible polymer to build the matrix of the magnetic carbonyl iron particles (CIP) for their uses in biomedical fields. Heat transfer behavior of the CIP-agar composites containing different concentrations (5, 10, 15, 20, 25 and 30% w/w) of magnetically aligned and non-aligned CIP in the agar matrix was studied using photothermal radiometry (PTR) in the back-propagation emission configuration. The morphology of the CIP-agar composites with aligned and non-aligned CIP under magnetic field was also evaluated by scanning electron microscopy (SEM). The results revealed a dominant effect of CIP concentration over the alignment patterns induced by the magnetic field, which agrees with the behavior of the thermal diffusivity and thermal conductivity. Agar served as a perfect matrix to be used with CIP, and CIP-agar composites magnetically aligned at 20% CIP concentration can be considered as promising 'smart' material for hyperthermia treatments in the biomedical field. Copyright © 2013 Elsevier Ltd. All rights reserved.

Evaluation of phytochemical content, antimicrobial, cytotoxic and antitumor activities of extract from Rumex hastatus D. Don roots.

PubMed

Sahreen, Sumaira; Khan, Muhammad Rashid; Khan, Rahmat Ali; Hadda, Taibi Ben

2015-07-03

Being a part of Chinese as well as ayurdic herbal system, roots of Rumex hastatus D. Don (RH) is highly medicinal, used to regulated blood pressure. It is also reported that the plant is diuretic, laxative, tonic, used against microbial skin diseases, bilious complaints and jaundice. The present study is conducted to evaluate phytochemical, antimicrobial, antitumor and cytotoxic activities of extract obtained from R. hastatus roots. RH roots were powdered and extracted with methanol to get crude extract. Crude extract was further fractioned on the basis of increasing polarity, with n-hexane (HRR), chloroform (CRR), ethyl acetate (ERR), n-butanol (BRR) and residual aqueous fraction (ARR). Methanol extract and its derived fractions were subjected to phytochemical screening and assayed for antibacterial activities via agar well diffusion method. Antifungal activities were checked through agar tube dilution method whereas potato disc assay was employed for the determination of antitumor activity. On the other hand cytotoxic activities were conducted using brine shrimps procedures. The results obtained from phytochemical analysis indicate the presence of alkaloids, anthraquinones, flavonoids and saponins in all the fractions. Most of the plant fractions showed substantial antimicrobial activities, which is in accordance with the spacious use of tested plant samples in primary healthcare center. Fractions of R. hastatus roots for cytotoxicity were tested as an effective cytotoxic was found as BRR > MRR > CRR > ARR > ERR > HRR. Ranking order of fractions of R. hastatus roots for effective antitumor screening was found as MRR > BRR > ARR > CRR > ERR > HRR. These results showed that R. hastatus appeared as an important source for the discovery of new antimicrobial drugs and antitumor agents; verify its traditional uses and its exploitation as therapeutic agent.

Bioactive Secondary Metabolites from Schizogyne sericea (Asteraceae) Endemic to Canary Islands.

PubMed

Venditti, Alessandro; Bianco, Armandodoriano; Muscolo, Camilla; Zorzetto, Christian; Sánchez-Mateo, Candelaria C; Rabanal, Rosa M; Quassinti, Luana; Bramucci, Massimo; Damiano, Silvia; Iannarelli, Romilde; Lupidi, Giulio; Papa, Fabrizio; Petrelli, Dezemona; Vitali, Luca A; Vittori, Sauro; Maggi, Filippo

2016-07-01

Schizogyne sericea (Asteraceae) is a halophytic shrub endemic to the Canary Islands and traditionally employed as analgesic, astringent, anti-inflammatory, and vulnerary. A comprehensive phytochemical investigation was conducted on the flowering aerial parts by analyzing both essential oil constituents and polar compounds. The essential oil was dominated by p-cymene, with the noteworthy occurrence of β-pinene and thymol esters. From the EtOH extract, eight compounds were isolated and structurally elucidated. Essential oil, polar fractions, and isolates (2), (4), and (5) were separately in vitro assayed for antiproliferative activity on human tumor cell lines (A375, MDA-MB 231, and HCT116) by MTT assay, for antioxidant potential by DPPH, ABTS, and FRAP assays, and for antimicrobial activity by the agar disk diffusion method. Results revealed that essential oil and compounds 1 and 2 exert a strong inhibition on tumor cells, and in some cases, higher than that of cisplatin. Fractions containing thymol derivatives (1 and 2) and caffeoylquinic acid derivatives 4 and 5 displayed antioxidant activity comparable to that of Trolox, making S. sericea extract an interesting natural product with potential applications as preservative or in the treatment of diseases in which oxidative stress plays an important role. © 2016 Wiley-VHCA AG, Zürich.

In vitro synergism between berberine and miconazole against planktonic and biofilm Candida cultures.

PubMed

Wei, Guo-Xian; Xu, Xin; Wu, Christine D

2011-06-01

To investigate the antimycotic activity of the plant alkaloid berberine (BBR), alone and in combination with antifungal azoles, against planktonic and biofilm Candida cultures. The minimum inhibitory concentrations (MICs) of BBR, miconazole (MCZ), and fluconazole (FLC) towards Candida albicans, Candida glabrata, Candida kefyr, Candida krusei, Candida parapsilosis, and Candida tropicalis were determined by a microdilution method. For C. albicans, the synergistic effects of BBR combined with MCZ or FLC were examined in a paper disc agar diffusion assay and checkerboard microdilution assay. The effect of the BBR/MCZ combination was further investigated in a C. albicans biofilm formation model with a dual-chamber flow cell. The effect on metabolic activity of biofilm cells was established using 2,3-bis(2-methoxy-4-nitro-5-sulfophenyl)-2H-tetrazolium-5-carboxanilide (XTT)/menadione. Berberine inhibited the growth of various Candida species (MICs 0.98-31.25mg/L) in the following order of susceptibility: C. krusei > C. kefyr > C. glabrata > C. tropicalis > C. parapsilosis and C. albicans. Synergism between BBR and MCZ or FLC was observed in the disc diffusion assay as well as in suspension showing an FIC index <0.5 (∑FIC=0.19). Whilst neither BBR (16 mg/L) nor MCZ (0.8 mg/L) alone significantly inhibited biofilm formation of C. albicans, their combination reduced biofilm formation by >91% after 24 h, as established from the reduction in surface area coverage (P<0.01). The BBR/MCZ combination also exhibited synergy against the metabolic activity of pre-formed C. albicans biofilms in polystyrene microtiter plates (∑FIC=0.25). Berberine exhibits synergistic effects with commonly used antimycotic drugs against C. albicans, either in planktonic or in biofilm growth phases. Published by Elsevier Ltd.

Proposed quality control guidelines for antimicrobial susceptibility tests using tilmicosin.

PubMed Central

Shryock, T R; White, D W; Werner, C S; Staples, J M

1995-01-01

Quality control guidelines for tilmicosin, a novel veterinary-use-only macrolide, were developed in a multi-laboratory study according to established National Committee for Clinical Laboratory Standards (NCCLS) procedures (M23-T2). Tilmicosin was incorporated into Sensititre plates for broth microdilution endpoint testing and into two lots of 15-micrograms disks for Kirby-Bauer agar disk diffusion testing. One common lot and five unique lots of Mueller-Hinton media were used. (Broth was cation adjusted, and agar was supplemented with 5% defibrinated sheep blood.) Bacteria used for reference strains included Pasteurella haemolytica 128K, Pasteurella multocida ATCC 43137, and Staphylococcus aureus ATCC 29213 (microdilution) and ATCC 25923 (disk). Replicate tests were conducted. Disk diffusion and broth microdilution quality control ranges are proposed. PMID:7714188

Evaluation of Five Chromogenic Agar Media and the Rosco Rapid Carb Screen Kit for Detection and Confirmation of Carbapenemase Production in Gram-Negative Bacilli

PubMed Central

Gilmour, Matthew W.; DeGagne, Pat; Nichol, Kim; Karlowsky, James A.

2014-01-01

An efficient workflow to screen for and confirm the presence of carbapenemase-producing Gram-negative bacilli was developed by evaluating five chromogenic screening agar media and two confirmatory assays, the Rapid Carb screen test (Rosco Diagnostica A/S, Taastrup, Denmark) and the modified Hodge test. A panel of 150 isolates was used, including 49 carbapenemase-producing isolates representing a variety of β-lactamase enzyme classes. An evaluation of analytical performance, assay cost, and turnaround time indicated that the preferred workflow (screening test followed by confirmatory testing) was the chromID Carba agar medium (bioMérieux, Marcy l'Étoile, France), followed by the Rapid Carb screen test, yielding a combined sensitivity of 89.8% and a specificity of 100%. As an optional component of the workflow, a determination of carbapenemase gene class via molecular means could be performed subsequent to confirmatory testing. PMID:25355764

Comparison of disk diffusion and agar dilution methods for gentamicin susceptibility testing of Neisseria gonorrhoeae.

PubMed

Gianecini, Ricardo; Oviedo, Claudia; Irazu, Lucia; Rodríguez, Marcelo; Galarza, Patricia

2018-03-29

Gentamicin is a promising antibiotic for the treatment of multidrug-resistant gonorrhea. The aim of this study was to analyze the suitability and reliably of disk diffusion to monitor the susceptibility to gentamicin. We studied 237 Neisseria gonorrhoeae isolates obtained in 2013 and 2015. Reference MICs were correlated with inhibition zone diameters (in millimeters) of gentamicin 10 µg disks manufactured by BBL and Oxoid. The Pearson correlation between disk diffusion and agar dilution was r = -.68 (P < 0.001) for BBL disk and r = -.71 (P < 0.001) for Oxoid disk. No very major or major discrepancies were detected. However, a high percentage of minor discrepancies was observed (44.7%, BBL disk) and (21.9%, Oxoid disk). By adjusting the susceptible breakpoint to S ≥ 17 mm, the minor discrepancies rate was reduced to 19.4% (BBL disk) and 10.1% (Oxoid disk). The disk diffusion may be a screening method in clinical laboratories to detect the gentamicin susceptibility of N. gonorrhoeae. Copyright © 2018 Elsevier Inc. All rights reserved.

Antibacterial and phytochemical screening of Anethum graveolens, Foeniculum vulgare and Trachyspermum ammi

PubMed Central

Kaur, Gurinder J; Arora, Daljit S

2009-01-01

Background Anethum graveolens Linn., Foeniculum vulgare Mill. and Trachyspermum ammi L. are widely used traditional medicinal plants to treat various ailments. To provide a scientific basis to traditional uses of these plants, their aqueous and organic seed extracts, as well as isolated phytoconstituents were evaluated for their antibacterial potential. Methods Antibacterial activity of aqueous and organic seed extracts was assessed using agar diffusion assay, minimum inhibitory concentration and viable cell count studies; and their antibacterial effect was compared with some standard antibiotics. The presence of major phytoconstituents was detected qualitatively and quantitatively. The isolated phytoconstituents were subjected to disc diffusion assay to ascertain their antibacterial effect. Results Hot water and acetone seed extracts showed considerably good antibacterial activity against all the bacteria except Klebsiella pneumoniae and one strain of Pseudomonas aeruginosa. Minimum inhibitory concentration for aqueous and acetone seed extracts ranged from 20–80 mg/ml and 5–15 mg/ml respectively. Viable cell count studies revealed the bactericidal nature of the seed extracts. Statistical analysis proved the better/equal efficacy of some of these seed extracts as compared to standard antibiotics. Phytochemical analysis showed the presence of 2.80 – 4.23% alkaloids, 8.58 – 15.06% flavonoids, 19.71 – 27.77% tannins, 0.55–0.70% saponins and cardiac glycosides. Conclusion Antibacterial efficacy shown by these plants provides a scientific basis and thus, validates their traditional uses as homemade remedies. Isolation and purification of different phytochemicals may further yield significant antibacterial agents. PMID:19656417

Abilities of the mCP Agar Method and CRENAME Alpha Toxin-Specific Real-Time PCR Assay To Detect Clostridium perfringens Spores in Drinking Water

PubMed Central

Maheux, Andrée F.; Bérubé, Ève; Boudreau, Dominique K.; Villéger, Romain; Cantin, Philippe; Boissinot, Maurice; Bissonnette, Luc

2013-01-01

We first determined the analytical specificity and ubiquity (i.e., the ability to detect all or most strains) of a Clostridium perfringens-specific real-time PCR (rtPCR) assay based on the cpa gene (cpa rtPCR) by using a bacterial strain panel composed of C. perfringens and non-C. perfringens Clostridium strains. All non-C. perfringens Clostridium strains tested negative, whereas all C. perfringens strains tested positive with the cpa rtPCR, for an analytical specificity and ubiquity of 100%. The cpa rtPCR assay was then used to confirm the identity of 116 putative C. perfringens isolates recovered after filtration of water samples and culture on mCP agar. Colonies presenting discordant results between the phenotype on mCP agar and cpa rtPCR were identified by sequencing the 16S rRNA and cpa genes. Four mCP−/rtPCR+ colonies were identified as C. perfringens, whereas 3 mCP+/rtPCR− colonies were identified as non-C. perfringens. The cpa rtPCR was negative with all 51 non-C. perfringens strains and positive with 64 of 65 C. perfringens strains. Finally, we compared mCP agar and a CRENAME (concentration and recovery of microbial particles, extraction of nucleic acids, and molecular enrichment) procedure plus cpa rtPCR (CRENAME + cpa rtPCR) for their abilities to detect C. perfringens spores in drinking water. CRENAME + cpa rtPCR detected as few as one C. perfringens CFU per 100 ml of drinking water sample in less than 5 h, whereas mCP agar took at least 25 h to deliver results. CRENAME + cpa rtPCR also allows the simultaneous and sensitive detection of Escherichia coli and C. perfringens from the same potable water sample. In itself, it could be used to assess the public health risk posed by drinking water potentially contaminated with pathogens more resistant to disinfection. PMID:24077714

Screening of bacterial antagonists for biological control of Phytophthora blight of pepper.

PubMed

Rajkumar, M; Lee, Wang Hyu; Lee, Kui Jae

2005-01-01

The aim of this study was to assess the potential of bacterial antagonists to control Phytophthora blight of pepper caused by P. capsici using different screening methods. Among a collection of fluorescent pseudomonas isolated from the rhizosphere of pepper, twelve isolates were initially selected based on dual culture assay on potato dextrose agar and corn meal agar. Further, these twelve isolates were screened for the reduction of disease severity caused by P. capsici using detached leaves and seedling assay. Most of the antagonists showed varying levels of antagonism against P. capsici in both detached leaves and seedlings assay. In addition, few isolates increased shoot and root length of pepper in seedling assays. Among them, isolate PS119 showing highest ability to reduce the disease severity in the in vitro seedling assay was found to be the most efficient antagonists against P. capsici in the in vivo biological control tests. These results indicate that the in vitro seedling assay can be used as a rapid and more accurate technique for the selection of promising biocontrol agents against P. capsici. ((c) 2005 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim).

Agar Disk Diffusion and Automated Microbroth Dilution Produce Similar Antimicrobial Susceptibility Testing Results for Salmonella Serotypes Newport, Typhimurium, and 4,5,12:i-, But Differ in Economic Cost

PubMed Central

Cummings, Kevin J.; Warnick, Lorin D.; Schukken, Ynte H.; Siler, Julie D.; Gröhn, Yrjo T.; Davis, Margaret A.; Besser, Tom E.; Wiedmann, Martin

2011-01-01

Abstract Data generated using different antimicrobial testing methods often have to be combined, but the equivalence of such results is difficult to assess. Here we compared two commonly used antimicrobial susceptibility testing methods, automated microbroth dilution and agar disk diffusion, for 8 common drugs, using 222 Salmonella isolates of serotypes Newport, Typhimurium, and 4,5,12:i-, which had been isolated from clinical salmonellosis cases among cattle and humans. Isolate classification corresponded well between tests, with 95% overall category agreement. Test results were significantly negatively correlated, and Spearman's correlation coefficients ranged from −0.98 to −0.38. Using Cox's proportional hazards model we determined that for most drugs, a 1 mm increase in zone diameter resulted in an estimated 20%–40% increase in the hazard of growth inhibition. However, additional parameters such as isolation year or serotype often impacted the hazard of growth inhibition as well. Comparison of economical feasibility showed that agar disk diffusion is clearly more cost-effective if the average sample throughput is small but that both methods are comparable at high sample throughput. In conclusion, for the Salmonella serotypes and antimicrobial drugs analyzed here, antimicrobial susceptibility data generated based on either test are qualitatively very comparable, and the current published break points for both methods are in excellent agreement. Economic feasibility clearly depends on the specific laboratory settings, and disk diffusion might be an attractive alternative for certain applications such as surveillance studies. PMID:21877930

Development of a nematode offspring counting assay for rapid and simple soil toxicity assessment.

PubMed

Kim, Shin Woong; Moon, Jongmin; Jeong, Seung-Woo; An, Youn-Joo

2018-05-01

Since the introduction of standardized nematode toxicity assays by the American Society for Testing and Materials (ASTM) and International Organization for Standardization (ISO), many studies have reported their use. Given that the currently used standardized nematode toxicity assays have certain limitations, in this study, we examined the use of a novel nematode offspring counting assay for evaluating soil ecotoxicity based on a previous soil-agar isolation method used to recover live adult nematodes. In this new assay, adult Caenorhabditis elegans were exposed to soil using a standardized toxicity assay procedure, and the resulting offspring in test soils attracted by a microbial food source in agar plates were counted. This method differs from previously used assays in terms of its endpoint, namely, the number of nematode offspring. The applicability of the bioassay was demonstrated using metal-spiked soils, which revealed metal concentration-dependent responses, and with 36 field soil samples characterized by different physicochemical properties and containing various metals. Principal component analysis revealed that texture fraction (clay, sand, and silt) and electrical conductivity values were the main factors influencing the nematode offspring counting assay, and these findings warrant further investigation. The nematode offspring counting assay is a rapid and simple process that can provide multi-directional toxicity assessment when used in conjunction with other standard methods. Copyright © 2018 Elsevier Ltd. All rights reserved.

Biocatalytic and antibacterial visualization of green synthesized silver nanoparticles using Hemidesmus indicus.

PubMed

Latha, M; Sumathi, M; Manikandan, R; Arumugam, A; Prabhu, N M

2015-05-01

In the present investigation, we described the green synthesis of silver nanoparticles using plant leaf extract of Hemidesmus indicus. The synthesized silver nanoparticles were characterized by UV-visible spectroscopy, fourier transform infra-red spectroscopy (FTIR), X-ray diffraction (XRD), transmission electron microscopy (TEM) and energy dispersive X-ray spectroscopy (EDX). TEM images proved that the synthesized silver nanoparticles were spherical in shape with an average particle size of 25.24 nm. To evaluate antibacterial efficacy, bacteria was isolated from poultry gut and subjected to 16S rRNA characterization and confirmed as Shigella sonnei. The in vitro antibacterial efficacy of synthesized silver nanoparticles was studied by agar bioassay, well diffusion and confocal laser scanning microscopy (CLSM) assay. The H. indicus mediated synthesis of silver nanoparticles shows rapid synthesis and higher inhibitory activity (34 ± 0.2 mm) against isolated bacteria S. sonnei at 40 μg/ml. Copyright © 2015 Elsevier Ltd. All rights reserved.

Rosmarinus eriocalyx: An alternative to Rosmarinus officinalis as a source of antioxidant compounds.

PubMed

Bendif, Hamdi; Boudjeniba, Messaoud; Djamel Miara, Mohamed; Biqiku, Loreta; Bramucci, Massimo; Caprioli, Giovanni; Lupidi, Giulio; Quassinti, Luana; Sagratini, Gianni; Vitali, Luca A; Vittori, Sauro; Maggi, Filippo

2017-03-01

Rosmarinus eriocalyx is an aromatic evergreen bush endemic to Algeria where it is used as a condiment to flavour soups and meat and as a traditional remedy. In the present work we have analyzed for the first time the phenolic composition of polar extracts obtained from stems, leaves and flowers of R. eriocalyx by HPLC, and determined the antioxidant and antimicrobial effects by DPPH, ABTS, FRAP, ORAC and agar disc diffusion methods, respectively. Results showed that ethanolic extracts of leaves and flowers are a rich source of phenolic compounds, mainly rosmarinic acid, carnosic acid and carnosol that are the main responsible for the noteworthy antioxidant activity observed in the assays. This study showed that R. eriocalyx might be a spice to be included in the European food additive list and used as a preservative agent besides R. officinalis. Copyright © 2016 Elsevier Ltd. All rights reserved.

A study of tablet dissolution by magnetic resonance electric current density imaging.

PubMed

Mikac, Ursa; Demsar, Alojz; Demsar, Franci; Sersa, Igor

2007-03-01

The electric current density imaging technique (CDI) was used to monitor the dissolution of ion releasing tablets (made of various carboxylic acids and of sodium chloride) by following conductivity changes in an agar-agar gel surrounding the tablet. Conductivity changes in the sample were used to calculate spatial and temporal changes of ionic concentrations in the sample. The experimental data for ion migration were compared to a mathematical model based on a solution of the diffusion equation with moving boundary conditions for the tablet geometry. Diffusion constants for different acids were determined by fitting the model to the experimental data. The experiments with dissolving tablets were used to demonstrate the potential of the CDI technique for measurement of ion concentration in the vicinity of ion releasing samples.

Serotyping reanalysis of unserotypable Actinobacillus pleuropneumoniae isolates by agar gel diffusion test.

PubMed

Morioka, Ayako; Shimazaki, Yoko; Uchiyama, Mariko; Suzuki, Shoko

2016-05-03

We observed increasing unserotypable (UT) Actinobacillus pleuropneumoniae isolates using agar gel diffusion (AGD) test. To reanalyze their serovar, we performed rapid slide agglutination (RSA) test and multiplex PCR for 47 UT isolates. Of these, 25 were serovar 1 (UT-serovar 1), 20 were serovar 2 (UT-serovar 2) and 2 were serovar 15 (UT-serovar 15). We examined serotyping antigen extraction temperature to determine heat influence. UT-serovar 1 and 15 were influenced by heat, because their precipitation lines were observed in the case of low antigen extraction temperature. To investigate the relationship between antigenicity and genotype, we performed pulsed-field gel electrophoresis (PFGE) analysis using UT-serovar 2 and 15. The predominant PFGE pattern of UT-serovar 2 was identical to that of serovar 2.

Serotyping reanalysis of unserotypable Actinobacillus pleuropneumoniae isolates by agar gel diffusion test

PubMed Central

MORIOKA, Ayako; SHIMAZAKI, Yoko; UCHIYAMA, Mariko; SUZUKI, Shoko

2016-01-01

We observed increasing unserotypable (UT) Actinobacillus pleuropneumoniae isolates using agar gel diffusion (AGD) test. To reanalyze their serovar, we performed rapid slide agglutination (RSA) test and multiplex PCR for 47 UT isolates. Of these, 25 were serovar 1 (UT-serovar 1), 20 were serovar 2 (UT-serovar 2) and 2 were serovar 15 (UT-serovar 15). We examined serotyping antigen extraction temperature to determine heat influence. UT-serovar 1 and 15 were influenced by heat, because their precipitation lines were observed in the case of low antigen extraction temperature. To investigate the relationship between antigenicity and genotype, we performed pulsed-field gel electrophoresis (PFGE) analysis using UT-serovar 2 and 15. The predominant PFGE pattern of UT-serovar 2 was identical to that of serovar 2. PMID:26726101

Formulation and Evaluation of Tramadol hydrochloride Rectal Suppositories.

PubMed

Saleem, M A; Taher, M; Sanaullah, S; Najmuddin, M; Ali, Javed; Humaira, S; Roshan, S

2008-09-01

Rectal suppositories of tramadol hydrochloride were prepared using different bases and polymers like PEG, cocoa butter, agar and the effect of different additives on in vitro release of tramadol hydrochloride was studied. The agar-based suppositories were non-disintegrating/non-dissolving, whereas PEGs were disintegrating/dissolving and cocoa butter were melting suppositories. All the prepared suppositories were evaluated for various physical parameters like weight variation, drug content and hardness. The PEG and cocoa butter suppositories were evaluated for macromelting range, disintegration and liquefaction time. In vitro release study was performed by USP type I apparatus. The prepared suppositories were within the permissible range of all physical parameters. In vitro drug release was in the order of PEG>Agar>cocoa butter. Addition of PVP, HPMC in agar suppositories retards the release. The mechanism of drug release was diffusion controlled and follows first order kinetics. The results suggested that blends of PEG of low molecular weight (1000) with high molecular weight (4000 and 6000) in different percentage and agar in 10% w/w as base used to formulate rapid release suppositories. The sustained release suppositories can be prepared by addition of PVP, HPMC in agar-based suppositories and by use of cocoa butter as base.

The molecular bacterial load assay replaces solid culture for measuring early bactericidal response to antituberculosis treatment.

PubMed

Honeyborne, Isobella; Mtafya, Bariki; Phillips, Patrick P J; Hoelscher, Michael; Ntinginya, Elias N; Kohlenberg, Anke; Rachow, Andrea; Rojas-Ponce, Gabriel; McHugh, Timothy D; Heinrich, Norbert

2014-08-01

We evaluated the use of the molecular bacterial load (MBL) assay, for measuring viable Mycobacterium tuberculosis in sputum, in comparison with solid agar and liquid culture. The MBL assay provides early information on the rate of decline in bacterial load and has technical advantages over culture in either form. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

An (almost) solvable model for bacterial pattern formation

NASA Astrophysics Data System (ADS)

Grammaticos, B.; Badoual, M.; Aubert, M.

2007-10-01

We present a simple model for the description of ring-like concentric structures in bacterial colonies. We model the differences between Bacillus subtilis and Proteus mirabilis colonies by using a different dependence of the duration of the consolidation phase on the concentration of agar. We compare our results to experimental data from these two bacterial species colonies and obtain a good agreement. Based on this analysis, we formulate a hypothesis on the connection of the diffusion constant that appears in the model to the experimental agar concentration.

Antioxidant and antibacterial activity of Thai medicinal plant (Capparis micracantha)

NASA Astrophysics Data System (ADS)

Laoprom, Nonglak; Sangprom, Araya; Chaisri, Patcharaporn

2018-04-01

This work aims to study the antioxidants capacity, Total phenolic content and antibacterial activity of Thai medicinal plant for the treatment of dermatitis-related inflammations, Capparis micracantha. Crude extract from stem of Thai medicinal plant was extracted with hexane, ethyl acetate, methanol and water. The antioxidant activities (IC50) was evaluated with 1,1-diphenyl-1-princylhydrazyl (DPPH) radical scavenging assay. Total phenolic content (TPC) was determined by using Folin-Ciocalteu method. Bacterial activities was tested with four human pathogenic bacteria; Escherichia coli, Listeria monocytogenes, Staphylococcus aureus and Stapylococcus epidermidis by using agar diffusion assay. Minimum Inhibition Concentration (MIC) and Minimum Bactericidal Concentration (MBC) were also determined by broth dilution method. For antioxidant activity, the methanol fraction from stem extract showed the highest activity with an IC50 of 2.4 mg/ml. Water extraction was the high TPC with 10,136.9 mg GAE/g dry weight. Methanol and water extraction showed the remarkable inhibition of bacterial growth was shown against L. monocytogenes and S. aureus. In addition, ethyl acetate, methanol and water fraction from stem extract against S. epidermidis. The present finding suggests that the extract of C. micracantha could be used to discover bioactive natural products that may serve as pharmaceutical products.

Phytochemical Constituents and Antimicrobial Activity of the Ethanol and Chloroform Crude Leaf Extracts of Spathiphyllum cannifolium (Dryand. ex Sims) Schott.

PubMed

Dhayalan, Arunachalam; Gracilla, Daniel E; Dela Peña, Renato A; Malison, Marilyn T; Pangilinan, Christian R

2018-01-01

The study investigated the medicinal properties of Spathiphyllum cannifolium (Dryand. ex Sims) Schott as a possible source of antimicrobial compounds. The phytochemical constituents were screened using qualitative methods and the antibacterial and antifungal activities were determined using agar well diffusion method. One-way analysis of variance and Fisher's least significant difference test were used. The phytochemical screening showed the presence of sterols, flavonoids, alkaloids, saponins, glycosides, and tannins in both ethanol and chloroform leaf extracts, but triterpenes were detected only in the ethanol leaf extract. The antimicrobial assay revealed that the chloroform leaf extract inhibited Candida albicans, Escherichia coli, Staphylococcus aureus, Bacillus subtilis, and Pseudomonas aeruginosa , whereas the ethanol leaf extract inhibited E. coli , S. aureus , and B. subtilis only. The ethanol and chloroform leaf extracts exhibited the highest zone of inhibition against B. subtilis . The antifungal assay showed that both the leaf extracts have no bioactivity against Aspergillus niger and C. albicans . Results suggest that chloroform is the better solvent for the extraction of antimicrobial compounds against the test organisms used in this study. Findings of this research will add new knowledge in advancing drug discovery and development in the Philippines.

Evidence for Anti-Pseudogymnoascus destructans (Pd) Activity of Propolis.

PubMed

Ghosh, Soumya; McArthur, Robyn; Guo, Zhi Chao; McKerchar, Rory; Donkor, Kingsley; Xu, Jianping; Cheeptham, Naowarat

2017-12-21

White-nose syndrome (WNS) in bats, caused by Pseudogymnoascus destructans ( Pd ), is a cutaneous infection that has devastated North American bat populations since 2007. At present, there is no effective method for controlling this disease. Here, we evaluated the effect of propolis against Pd in vitro. Using Sabouraud dextrose agar (SDA) medium, approximately 1.7 × 10⁷ conidia spores of the Pd strain M3906-2/mL were spread on each plate and grown to form a consistent lawn. A Kirby-Bauer disk diffusion assay was employed using different concentrations of propolis (1%, 2%, 3%, 4%, 5%, 10%, 15%, 20%, 25%), in plates incubated at 8 °C and 15 °C. At 8 °C and 15 °C, as the concentration of propolis increased, there was an increasing zone of inhibition (ZOI), reaching the highest degree at 10% and 25% concentrations, respectively. A germule suppression assay showed a similar effect on Pd conidia germination. A MALDI-TOF-MS analysis of propolis revealed multiple constituents with a potential anti- Pd activity, including cinnamic acid, p-coumaric acid, and dihydrochalcones, which could be further tested for their individual effects. Our study suggests that propolis or its individual constituents might be suitable products against Pd .

Effect of propolis in gastric disorders: inhibition studies on the growth of Helicobacter pylori and production of its urease.

PubMed

Baltas, Nimet; Karaoglu, Sengul Alpay; Tarakci, Cemre; Kolayli, Sevgi

2016-01-01

There is considerable interest in alternative approaches to inhibit Helicobacter pylori (H. pylori) and thus treat many stomach diseases. Propolis is a pharmaceutical mixture containing many natural bioactive substances. The aim of this study was to use propolis samples to treat H. pylori. The anti-H. pylori and anti-urease activities of 15 different ethanolic propolis extracts (EPEs) were tested. The total phenolic contents and total flavonoid contents of the EPE were also measured. The agar-well diffusion assay was carried out on H. pylori strain J99 and the inhibition zones were measured and compared with standards. All propolis extracts showed high inhibition of H. pylori J99, with inhibition diameters ranging from 31.0 to 47.0 mm. Helicobacter pylori urease inhibitory activity was measured using the phenol-hypochlorite assay; all EPEs showed significant inhibition against the enzyme, with inhibition concentrations (IC 50 ; mg/mL) ranging from 0.260 to 1.525 mg/mL. The degree of inhibition was related to the phenolic content of the EPE. In conclusion, propolis extract was found to be a good inhibitor that can be used in H. pylori treatment to improve human health.

Inhibition of Fusarium solani Infection in Murine Keratocytes by Lactobacillus salivarius ssp. salivarius JCM1231 Culture Filtrate In Vitro.

PubMed

Hu, Jianzhang; Chen, Fang; Kan, Tong; Zhuang, Hua; Zhang, Jingjin; Han, Xiaoli

2017-10-01

To explore the inhibitory activity of Lactobacillus salivarius ssp. salivarius JCM1231 (L. salivarius JCM1231) culture filtrate against Fusarium solani (F. solani) and its effects on murine keratocytes (MKs) infected with F. solani. L. salivarius JCM1231 was cultured in an anaerobic incubator for 24 h, and the L. salivarius culture filtrate (LSCF) was prepared .The antifungal activity of L. salivarius JCM1231 against F. solani was determined with a plate overlay assay, agar diffusion assay, and conidial germination inhibition test. The effects of temperature, pH, and proteolytic enzymes on the antifungal activity of LSCF were detected with microtiter plate-well assay and conidial germination inhibition assay. Furthermore, the effects of LSCF on MKs infected with F. solani were detected. Cell activity and apoptosis were measured using methylthiazoletetrazolium assays and flow cytometry analysis, respectively. The levels of interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α) cytokines were measured using real-time polymerase chain reactions and enzyme-linked immunosorbent assays (ELISA), and mycotoxin production was detected with high-performance liquid chromatography tandem mass spectrometry. Conidial germination and mycelia growth of F. solani were significantly inhibited by LSCF. The antifungal substances produced by L. salivarius JCM1231 were heat unstable, proteinaceous, and sensitive to proteolytic enzymes and were active within a narrow acidic pH range between 2.0 and 4.0. In the presence of 15 µg/ml of LSCF, cell activity was significantly increased, and cell apoptosis, the level of IL-6 and TNF-α expressions, and mycotoxin (zearalenone and fumonisin B1) productions were decreased significantly in MKs infected with F. solani. L. salivarius JCM1231 culture filtrate can effectively inhibit F. solani growth and protect MKs against F. solani infection.

Screening of Microbial Extracts for Anticancer Compounds Using Streptomyces Kinase Inhibitor Assay.

PubMed

Shanbhag, Prashant; Bhave, Sarita; Vartak, Ashwini; Kulkarni-Almeida, Asha; Mahajan, Girish; Villanueva, Ivan; Davies, Julian

2015-07-01

Eukaryotic kinases are known to play an important role in signal transduction pathways by phosphorylating their respective substrates. Abnormal phosphorylations by these kinases have resulted in diseases. Hence inhibitors of kinases are of considerable pharmaceutical interest for a wide variety of disease targets, especially cancers. A number of reports have been published which indicate that eukaryotic-like kinases may complement two-component kinase systems in several bacteria. In Streptomyces sp. such kinases have been found to have a role in formation of aerial hyphae, spores, pigmentation & even in antibiotic production in some strains. Eukaryotic kinase inhibitors are seen to inhibit formation of aerial mycelia in Streptomyces without inhibiting vegetative mycelia. This property has been used to design an assay to screen for eukaryotic kinase inhibitors. The assay involves testing of compounds against Streptomyces 85E ATCC 55824 using agar well diffusion method. Inhibitors of kinases give rise to "bald" colonies where aerial mycelia and sporulation inhibition is seen. The assay has been standardized using known eukaryotic protein kinase inhibiting anticancer agents like AG-490, AG-1295, AG-1478, Flavopiridol and Imatinib as positive controls, at a concentration ranging from 10 μg/well to 100 μg/well. Anti-infective compounds which are not reported to inhibit eukaryotic protein kinases were used as negative controls. A number of microbial cultures have been screened for novel eukaryotic protein kinase inhibitors. Further these microbial extracts were tested in various cancer cell lines like Panel, HCT116, Calul, ACHN and H460 at a concentration of 10 μg/mL/ well. The anticancer data was seen correlating well with the Streptomyces kinase assay thus validating the assay.

A Simple Experiment for Visualizing Diffusion

ERIC Educational Resources Information Center

Helseth, L. E.

2011-01-01

We propose a simple and fascinating experiment for studying diffusion in gels using a pH-sensitive dye. By doping agar with methyl red, we obtain a gel which rapidly reacts to changes in pH by changing its absorption spectrum. The pH gradients can be followed using a digital camera, and we demonstrate here that the pH-sensitive colour changes can…

Fluorogenic membrane overlays to enumerate total coliforms, Escherichia coli, and total Vibrionaceae in shellfish and seawater

USDA-ARS?s Scientific Manuscript database

Three assays were developed to enumerate total coliforms, Escherichia coli, and total Vibrionaceae in shellfish and other foods and in seawater and other environmental samples. Assays involve membrane overlays of overnight colonies on non-selective agar plates to detect ß-glucuronidase and lysyl am...

DETECTION OF BACTERIAL CYTOTOXIC ACTIVITIES FROM WATER-DAMAGED CEILING TILE MATERIAL FOLLOWING INCUBATION ON BLOOD AGAR

EPA Science Inventory

Samples of ceiling tiles with high levels of bacteria exhibited cytotoxic activities on a HEP-2 tissue culture assay. Ceiling tiles containing low levels of bacterial colonization did not show cytotoxic activities on the HEP-2 tissue culture assay. Using a spread plate procedure ...

High-Throughput Biosensor Discriminates Between Different Algal H 2-Photoproducing Strains

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wecker, Matt S. A.; Maria L. Ghirardi

2014-02-27

A number of species of microalgae and cyanobacteria photosynthetically produce H 2 gas by coupling water oxidation with the reduction of protons to molecular hydrogen, generating renewable energy from sunlight and water. Photosynthetic H 2 production, however, is transitory, and there is considerable interest in increasing and extending it for commercial applications. Here we report a Petri-plate version of our previous, microplate-based assay that detects photosynthetic H 2 production by algae. The assay consists of an agar overlay of H 2-sensing Rhodobacter capsulatus bacteria carrying a green fluorescent protein that responds to H 2 produced by single algal colonies inmore » the bottom agar layer. The assay distinguishes between algal strains that photoproduce H 2 at different levels under high light intensities, and it does so in a simple, inexpensive, and high-throughput manner. The assay will be useful for screening both natural populations and mutant libraries for strains having increased H 2 production, and useful for identifying various genetic factors that physiologically or genetically alter algal hydrogen production.« less

Comparison of susceptibility test methods to detect penicillin susceptibility in Streptococcus pneumoniae isolates.

PubMed

Mohd Nasir, Mohd Desa; Parasakthi, Navaratnam

2004-06-01

The increasing prevalence of penicillin-resistant Streptococuus pneumoniae urges for fast and accurate susceptibility testing methods. This study evaluated the comparability of three commonly used techniques; disk diffusion, E-test and agar dilution, to detect penicillin susceptibility in clinical isolates of S. pneumoniae. Fifty pneumococcal isolates, obtained from patients at the University of Malaya Medical Centre, were selected to include both penicillin-susceptible strains and those that had decreased susceptibility (resistant and intermediate) to penicillin. The minimum inhibitory concentration (MIC) values of penicillin to serve as the reference was determined by the agar dilution method in which, based on the MIC breakpoints recommended by the National Committee for Clinical Laboratory Standards (NCCLS), 27 strains had decreased susceptibility to penicillin with 17 strains resistant and 10 intermediate. Comparing to the agar dilution method, oxacillin disk diffusion test detected all strains with decreased penicillin susceptibility as such while E-test showed a close agreement of susceptibility (92%) of the isolates to penicillin. This confirmed that oxacillin is a good screening test for S. pneumoniae isolates with decreased susceptibility to penicillin while E-test is very reliable for rapid and accurate detection of penicillin susceptibility.

Invasive growth of Saccharomyces cerevisiae depends on environmental triggers: a quantitative model.

PubMed

Zupan, Jure; Raspor, Peter

2010-04-01

In this contribution, the influence of various physicochemical factors on Saccharomyces cerevisiae invasive growth is examined quantitatively. Agar-invasion assays are generally applied for in vitro studies on S. cerevisiae invasiveness, the phenomenon observed as a putative virulence trait in this clinically more and more concerning yeast. However, qualitative agar-invasion assays, used until now, strongly limit the feasibility and interpretation of analyses and therefore needed to be improved. Besides, knowledge in this field concerning the physiology of invasive growth, influenced by stress conditions related to the human alimentary tract and food, is poor and should be expanded. For this purpose, a quantitative agar-invasion assay, presented in our previous work, was applied in this contribution to clarify the significance of the stress factors controlling the adhesion and invasion of the yeast in greater detail. Ten virulent and non-virulent S. cerevisiae strains were assayed at various temperatures, pH values, nutrient starvation, modified atmosphere, and different concentrations of NaCl, CaCl2 and preservatives. With the use of specific parameters, like a relative invasion, eight invasive growth models were hypothesized, which enabled intelligible interpretation of the results. A strong preference for invasive growth (meaning high relative invasion) was observed when the strains were grown on nitrogen- and glucose-depleted media. A significant increase in the invasion of the strains was also determined at temperatures typical for human fever (37-39 degrees C). On the other hand, a strong repressive effect on invasion was found in the presence of salts, anoxia and some preservatives. Copyright 2010 John Wiley & Sons, Ltd.

Immunomodulatory Effects of Lactobacillus salivarius LS01 and Bifidobacterium breve BR03, Alone and in Combination, on Peripheral Blood Mononuclear Cells of Allergic Asthmatics.

PubMed

Drago, Lorenzo; De Vecchi, Elena; Gabrieli, Arianna; De Grandi, Roberta; Toscano, Marco

2015-07-01

The aim of this study was to evaluate probiotic characteristics of Lactobacillus salivarius LS01 and Bifidobacterium breve BR03 alone and in combination and their immunomodulatory activity in asthmatic subjects. Subjects affected by allergic asthma were recruited. Initially, LS01 and BR03 were analyzed for their growth compatibility by a broth compatibility assay. To study the antimicrobial activity of probiotic strains, an agar diffusion assay was performed. Finally, cytokine production by peripheral blood mononuclear cells (PBMCs) stimulated with LS01 and BR03 was determined by means of specific quantitative enzyme-linked immunosorbent assay (ELISA). The growth of some clinical pathogens were slightly inhibited by LS01 and LS01-BR03 co-culture supernatant not neutralized to pH 6.5, while only the growth of E. coli and S. aureus was inhibited by the supernatant of LS01 and LS01-BR03 neutralized to pH 6.5. Furthermore, LS01 and BR03 combination was able to decrease the secretion of proinflammatory cytokines by PBMCs, leading to an intense increase in IL-10 production. L. salivarius LS01 and B. breve BR03 showed promising probiotic properties and beneficial immunomodulatory activity that are increased when the 2 strains are used in combination in the same formulation.

Visualization of the Charcoal Agar Resazurin Assay for Semi-quantitative, Medium-throughput Enumeration of Mycobacteria.

PubMed

Gold, Ben; Roberts, Julia; Ling, Yan; Lopez Quezada, Landys; Glasheen, Jou; Ballinger, Elaine; Somersan-Karakaya, Selin; Warrier, Thulasi; Nathan, Carl

2016-12-14

There is an urgent need to discover and progress anti-infectives that shorten the duration of tuberculosis (TB) treatment. Mycobacterium tuberculosis, the etiological agent of TB, is refractory to rapid and lasting chemotherapy due to the presence of bacilli exhibiting phenotypic drug resistance. The charcoal agar resazurin assay (CARA) was developed as a tool to characterize active molecules discovered by high-throughput screening campaigns against replicating and non-replicating M. tuberculosis. Inclusion of activated charcoal in bacteriologic agar medium helps mitigate the impact of compound carry-over, and eliminates the requirement to pre-dilute cells prior to spotting on CARA microplates. After a 7-10 day incubation period at 37 °C, the reduction of resazurin by mycobacterial microcolonies growing on the surface of CARA microplate wells permits semi-quantitative assessment of bacterial numbers via fluorometry. The CARA detects approximately a 2-3 log10 difference in bacterial numbers and predicts a minimal bactericidal concentration leading to ≥99% bacterial kill (MBC≥99). The CARA helps determine whether a molecule is active on bacilli that are replicating, non-replicating, or both. Pilot experiments using the CARA facilitate the identification of which concentration of test agent and time of compound exposure require further evaluation by colony forming unit (CFU) assays. In addition, the CARA can predict if replicating actives are bactericidal or bacteriostatic.

Disk Diffusion Testing Using Candida sp. Colonies Taken Directly from CHROMagar Candida Medium May Decrease Time Required To Obtain Results

PubMed Central

Klevay, Michael; Ebinger, Alex; Diekema, Daniel; Messer, Shawn; Hollis, Richard; Pfaller, Michael

2005-01-01

We compared results of disk diffusion antifungal susceptibility testing from Candida sp. strains passaged on CHROMagar and on potato dextrose agar. The overall categorical agreements for fluconazole and voriconazole disk testing were 95% and 98% with 0% and 0.5% very major errors, respectively. Disk diffusion testing by the CLSI (formerly NCCLS) M44-A method can be performed accurately by taking inocula directly from CHROMagar. PMID:16000489

Performance of transport and selective media for swine Bordetella bronchiseptica recovery and it comparison to polymerase chain reaction detection

PubMed Central

Coutinho, Tania Alen; Bernardi, Mari Lourdes; de Itapema Cardoso, Marisa Ribeiro; Borowski, Sandra Maria; Moreno, Andrea Micke; de Barcellos, David Emilio Santos Neves

2009-01-01

Three comparative assays were performed seeking to improve the sensitivity of the diagnosis of Bordetella bronchiseptica infection analyzing swine nasal swabs. An initial assay compared the recovery of B. bronchiseptica from swabs simultaneously inoculated with B. bronchiseptica and some interfering bacteria, immersed into three transport formulations (Amies with charcoal, trypticase soy broth and phosphate buffer according to Soerensen supplemented with 5% of bovine fetal serum) and submitted to different temperatures (10°C and 27°C) and periods of incubation (24, 72 and 120 hours). A subsequent assay compared three selective media (MacConkey agar, modified selective medium G20G and a ceftiofur medium) for their recovery capabilities from clinical specimens. One last assay compared the polymerase chain reaction to the three selective media. In the first assay, the recovery of B. bronchiseptica from transport systems was better at 27°C and the three formulations had good performances at this temperature, but the collection of qualitative and quantitative analysis indicated the advantage of Amies medium for nasal swabs transportation. The second assay indicated that MacConkey agar and modified G20G had similar results and were superior to the ceftiofur medium. In the final assay, polymerase chain reaction presented superior capability of B. bronchiseptica detection to culture procedures. PMID:24031390

Escherichia coli Overexpressing a Baeyer-Villiger Monooxygenase from Acinetobacter radioresistens Becomes Resistant to Imipenem

PubMed Central

Minerdi, Daniela; Zgrablic, Ivan; Castrignanò, Silvia; Catucci, Gianluca; Medana, Claudio; Terlizzi, Maria Elena; Gribaudo, Giorgio; Gilardi, Gianfranco

2015-01-01

Antimicrobial resistance is a global issue currently resulting in the deaths of hundreds of thousands of people a year worldwide. Data present in the literature illustrate the emergence of many bacterial species that display resistance to known antibiotics; Acinetobacter spp. are a good example of this. We report here that Acinetobacter radioresistens has a Baeyer-Villiger monooxygenase (Ar-BVMO) with 100% amino acid sequence identity to the ethionamide monooxygenase of multidrug-resistant (MDR) Acinetobacter baumannii. Both enzymes are only distantly phylogenetically related to other canonical bacterial BVMO proteins. Ar-BVMO not only is capable of oxidizing two anticancer drugs metabolized by human FMO3, danusertib and tozasertib, but also can oxidize other synthetic drugs, such as imipenem. The latter is a member of the carbapenems, a clinically important antibiotic family used in the treatment of MDR bacterial infections. Susceptibility tests performed by the Kirby-Bauer disk diffusion method demonstrate that imipenem-sensitive Escherichia coli BL21 cells overexpressing Ar-BVMO become resistant to this antibiotic. An agar disk diffusion assay proved that when imipenem reacts with Ar-BVMO, it loses its antibiotic property. Moreover, an NADPH consumption assay with the purified Ar-BVMO demonstrates that this antibiotic is indeed a substrate, and its product is identified by liquid chromatography-mass spectrometry to be a Baeyer-Villiger (BV) oxidation product of the carbonyl moiety of the β-lactam ring. This is the first report of an antibiotic-inactivating BVMO enzyme that, while mediating its usual BV oxidation, also operates by an unprecedented mechanism of carbapenem resistance. PMID:26459905

Escherichia coli Overexpressing a Baeyer-Villiger Monooxygenase from Acinetobacter radioresistens Becomes Resistant to Imipenem.

PubMed

Minerdi, Daniela; Zgrablic, Ivan; Castrignanò, Silvia; Catucci, Gianluca; Medana, Claudio; Terlizzi, Maria Elena; Gribaudo, Giorgio; Gilardi, Gianfranco; Sadeghi, Sheila J

2016-01-01

Antimicrobial resistance is a global issue currently resulting in the deaths of hundreds of thousands of people a year worldwide. Data present in the literature illustrate the emergence of many bacterial species that display resistance to known antibiotics; Acinetobacter spp. are a good example of this. We report here that Acinetobacter radioresistens has a Baeyer-Villiger monooxygenase (Ar-BVMO) with 100% amino acid sequence identity to the ethionamide monooxygenase of multidrug-resistant (MDR) Acinetobacter baumannii. Both enzymes are only distantly phylogenetically related to other canonical bacterial BVMO proteins. Ar-BVMO not only is capable of oxidizing two anticancer drugs metabolized by human FMO3, danusertib and tozasertib, but also can oxidize other synthetic drugs, such as imipenem. The latter is a member of the carbapenems, a clinically important antibiotic family used in the treatment of MDR bacterial infections. Susceptibility tests performed by the Kirby-Bauer disk diffusion method demonstrate that imipenem-sensitive Escherichia coli BL21 cells overexpressing Ar-BVMO become resistant to this antibiotic. An agar disk diffusion assay proved that when imipenem reacts with Ar-BVMO, it loses its antibiotic property. Moreover, an NADPH consumption assay with the purified Ar-BVMO demonstrates that this antibiotic is indeed a substrate, and its product is identified by liquid chromatography-mass spectrometry to be a Baeyer-Villiger (BV) oxidation product of the carbonyl moiety of the β-lactam ring. This is the first report of an antibiotic-inactivating BVMO enzyme that, while mediating its usual BV oxidation, also operates by an unprecedented mechanism of carbapenem resistance. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Mechanism of Enhancement of Virus Plaques by Cationic Polymers

PubMed Central

Wallis, Craig; Melnick, Joseph L.

1968-01-01

It has been assumed that plaque enhancement by cationic polymers is due to their binding of sulfated polysaccharides in agar. However, viruses that are enhanced by cationic polymers, diethylaminoethyl-dextran, and protamine were found not to be inhibited by polyanions in agar under the usual overlay conditions. In the case of adenovirus, enhancement by protamine seems to be due to the protamine serving as a source of arginine; enzymes released from the cultured cells digest the protamine and provide a reservoir of arginine for the cells. Other viruses (herpes and echovirus types 3, 4, 5, and 6) known to be susceptible to agar inhibitors were found to be enhanced by cationic polymers even under starch gel and methylcellulose overlays, which are free of polyanions. Since cationic polymers enhance the diffusion of virus through agar or starch gel, plaque enhancement seems to be the result of the gel becoming positively charged so that viruses can move effectively through them. The observation that starch gel and methylcellulose enhance plaque formation with viruses known to be inhibited under agar was also reinvestigated. When the consistency of the agar gel was reduced to the same viscosity of starch gel and methylcellulose overlays, the same plaque counts and sizes were observed under all three overlays. PMID:4986901

Preparation, Imaging, and Quantification of Bacterial Surface Motility Assays

PubMed Central

Morales-Soto, Nydia; Anyan, Morgen E.; Mattingly, Anne E.; Madukoma, Chinedu S.; Harvey, Cameron W.; Alber, Mark; Déziel, Eric; Kearns, Daniel B.; Shrout, Joshua D.

2015-01-01

Bacterial surface motility, such as swarming, is commonly examined in the laboratory using plate assays that necessitate specific concentrations of agar and sometimes inclusion of specific nutrients in the growth medium. The preparation of such explicit media and surface growth conditions serves to provide the favorable conditions that allow not just bacterial growth but coordinated motility of bacteria over these surfaces within thin liquid films. Reproducibility of swarm plate and other surface motility plate assays can be a major challenge. Especially for more “temperate swarmers” that exhibit motility only within agar ranges of 0.4%-0.8% (wt/vol), minor changes in protocol or laboratory environment can greatly influence swarm assay results. “Wettability”, or water content at the liquid-solid-air interface of these plate assays, is often a key variable to be controlled. An additional challenge in assessing swarming is how to quantify observed differences between any two (or more) experiments. Here we detail a versatile two-phase protocol to prepare and image swarm assays. We include guidelines to circumvent the challenges commonly associated with swarm assay media preparation and quantification of data from these assays. We specifically demonstrate our method using bacteria that express fluorescent or bioluminescent genetic reporters like green fluorescent protein (GFP), luciferase (lux operon), or cellular stains to enable time-lapse optical imaging. We further demonstrate the ability of our method to track competing swarming species in the same experiment. PMID:25938934

Comparative Evaluation of Antibacterial Effect of Dental Luting Cements on Streptococcus mutans and Lactobacillus acidophilus: An In vitro Study.

PubMed

Feroz, Sma; Bhoyar, A; Khan, S

2016-12-01

This study aimed at evaluating and comparing the antibacterial activity of six types of dental luting cements on Streptococcus mutans and Lactobacillus acidophilus using the agar diffusion test (ADT) and the direct-contact test (DCT). The antibacterial activity in ADT was measured based on the diameter of the zone of inhibition formed, whereas in DCT the density of the bacterial suspension was measured. The lower the density of the suspension, the more antibacterial activity the cement possesses. Agar diffusion test was carried out on the bacteria. After an incubation period of 24 hours, the plates were checked for the presence of zone of inhibition. In DCT the cement was mixed and applied. Once the cement was set, bacterial suspension and brain-heart infusion medium was poured and incubated for 24 hours. After 24 hours, the plate was placed in the enzyme-linked immunosorbent assay plate reader, which measured the optical density of the fluid. The first set of data was recorded approximately 1 hour after incubation. Overall, three sets of data were recorded. Additional experiments were performed on set test materials that were allowed to age for 24 hours, 1 week, 1, 3, and 6 months. When using ADT only two cements zinc oxide eugenol (ZOE) and zinc polycarboxylate (ZPC) cement showed antibacterial activity against the test organisms. When using DCT, all cements showed some amount of antibacterial activity. Zinc oxide eugenol and ZPC cement showed highest amount of antibacterial activity against S. mutans and L. acidophilus respectively. Within the limitations of study, ZOE cement and ZPC cement were most effective against the tested microorganisms followed by the newer resin cement. The glass ionomer cement was the weakest of all. Patients with high caries index can be treated more effectively using the abovementioned cements.

Impact of Cu(II)-doping on the vulnerability of Escherichia coli ATCC 10536 revealed by Atomic Force Microscopy.

PubMed

Rasheed, Wasia; Perveen, Samina; Mustafa, Ghulam; Shah, Muhammad Raza; Ahmed, Shakil; Uzzaman, Sami

2018-05-08

E. coli strain is a gram-negative bacterium known to induce both extra-intestinal infections and intestinal infections. For survival of microbes, metal intake and accessibility should be according to their physiological requirements. Peculiarly, copper homeostasis is critical for E. coli survival and growth. Therefore in this study, an extensive work is conducted to investigate the impact of Cu(II)-doping on the susceptibility of Escherichia coli ATCC 10536 against Cu(II)-selective Cefaclor-silver nanoconjugates (i.e., Cf-AgNPs) and its organic precursor (i.e. Cefaclor). At first, the maximal non-cytotoxic dose of Cu(II) that was sub-lethal for Escherichia coli was determined by MTT assay and was found to be 100 μg/L. Afterwards, MICs of Cf-AgNPs and Cefaclor against controlled and Cu(II)-doped E. coli cells were determined by using Agar well diffusion method. The susceptibility of E. coli cells against Cf-AgNPs was increased upon Cu(II) doping, whereas the bactericidal activity of Cefaclor against Cu(II)-doped E. coli cells was retarded due to hydrolysis. In addition, morphological changes induced in controlled and Cu(II)-doped samples of E. coli after treatment with Cefaclor and Cf-AgNPs were also monitored by Atomic force microscopy (AFM). The obtained results from both Agar well diffusion method and AFM confirmed that Cf-AgNPs are more effective against Cu(II)-doped Escherichia coli. Moreover, thermal profile of Cu(II)-selective Cf-AgNPs was also demonstrated by TGA and DSC. This study can be an important part of the relevant state-of-the-art. Indeed, further clinical studies are necessary to determine the relevant role of Cf-AgNPs compared with that of the Cefaclor now available. Copyright © 2018 Elsevier Ltd. All rights reserved.

Growth pattern of the surface of fungus Aspergillus colony

NASA Astrophysics Data System (ADS)

Matsuura, Shu; Miyazima, Sasuke

1992-05-01

Aspergillus oryzae colonies were grown under various glucose concentrations, temperatures, and agar concentrations, and the effects on the pattern were investigated. Patterns of colony were found to vary from uniform to diffusion-limited aggregation type.

Evaluation of antimicrobial efficacy of Aloe vera and its effectiveness in decontaminating gutta percha cones.

PubMed

Athiban, Prakash P; Borthakur, Bikash Jyoti; Ganesan, S; Swathika, B

2012-07-01

The aim of this study was to evaluate the antimicrobial efficacy of Aloe vera and to determine its effectiveness in decontaminating gutta percha cones. A concentrated extract of Aloe vera was used to check for the antimicrobial efficacy using the agar well diffusion method. Presence of zones' of diffusion was identified against three common GP contaminants namely, E.coli, E.faecalis and Staph. aureus. New GP Cones, freshly taken out of the packet were then decontaminated for 1minute using Aloe vera gel and then placed in thioglycolate broth to check for the presence of turbidity. The zones of inhibition on the agar plate were measured as 24mm,21mm and 24mm respectively. The broth remained clear even after 48 hours of incubation. We conclude that Aloe vera is indeed effective as a GP decontaminant and it holds a promising future as a medium for storage of GP cones.

Evaluation of the antimicrobial activities of chlorhexidine gluconate, sodium hypochlorite and octenidine hydrochloride in vitro.

PubMed

Tirali, Resmiye E; Bodur, Haluk; Sipahi, Bilge; Sungurtekin, Elif

2013-04-01

The objective of this study was to compare the antimicrobial activity of sodium hypochlorite (NaOCl), chlorhexidine gluconate (CHX) and octenidine hydrochloride (OCT) in different concentrations against endodontic pathogens in vitro. Agar diffusion procedure was used to determine the antimicrobial activity of the tested materials. Enterococcus faecalis, Candida albicans and the mixture of these were used for this study. In the agar diffusion test, 5.25% NaOCl exhibited better antimicrobial effect than the other concentrations of NaOCl for all strains. All concentrations of OCT were effective against C. albicans and E. faecalis. Some 0.2% CHX was ineffective on all microorganisms. Antibacterial effectiveness of all experimental solutions decreased on the mixture of all strains. Decreasing concentrations of NaOCl resulted in significantly reduced antimicrobial effect. © 2010 The Authors. Australian Endodontic Journal © 2010 Australian Society of Endodontology.

Isolation and Characterization of Isogenic Pairs of Domed Hemolytic and Flat Nonhemolytic Colony Types of Bordetella pertussis

PubMed Central

Peppler, Mark S.

1982-01-01

Four different serotype strains of Bordetella pertussis, 3779BL2S4, Tohama I, 353/Z, and 2753, were plated on Bordet-Gengou agar, where they grew as domed, hemolytic (D+H+) wild-type colonies. Cloned D+H+ colony types of all four strains were passed onto modified Stainer-Scholte medium solidified with 1% Noble Agar. Colonies were selected from Stainer-Scholte agar, and these subsequently grew as flat, nonhemolytic (D−H−) colonies when transferred back onto Bordet-Gengou agar. The frequency of D−H− organisms within a population of cloned D+H+ was determined to be between 5 × 10−5 and 5 × 10−6. The D−H− colony types maintained their flat, nonhemolytic characteristics for over 80 single-colony passages on Bordet-Gengou agar. The isogenic pairs of D+H+ and D−H− colony types from the four strains were compared for hemagglutination titer, lymphocytosis-promoting activity, adenylate cyclase activity, and presence of agglutinogens by agglutination. In all cases the D−H− colony types showed reduced activities or amounts of antigen compared with their D+H+ parents. Freely diffusible antigens were markedly different between the two phenotypes as noted by double diffusion of antisera added to plates on which colonies of the variants were growing. Antigens solubilized from the two colony types by Triton X-100 were also markedly different as judged by radial immunodiffusion with antifimbrial hemagglutinin, antilymphocytosis-promoting factor, and anti-353/Z adsorbed with autoclaved 353/Z. In addition, autoradiographs of 125I-surface-labeled whole cells separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed unique banding patterns for each colony type. Since all organisms, regardless of colony type, were grown on Bordet-Gengou agar, the differences reported could not be due to medium composition. Differences between phenotypes were also independent of passage number on Bordet-Gengou agar. By analogy to previous studies, the D−H− organisms appear to fulfill the criteria for phase III or phase IV in the system of Leslie and Gardner (P. H. Leslie and A. D. Gardner, J. Hyg. 31:423-434, 1931) or phase III in the system of Kasuga et al. (T. Kasuga, Y. Nakase, K. Ukishima, and K. Takatsu, Kitasato Arch. Exp. Med. 26:121-134, 1954). Images PMID:6279517

An HTS-compatible 3D colony formation assay to identify tumor-specific chemotherapeutics.

PubMed

Horman, Shane R; To, Jeremy; Orth, Anthony P

2013-12-01

There has been increasing interest in the development of cellular behavior models that take advantage of three-dimensional (3D) cell culture. To enable assessment of differential perturbagen impacts on cell growth in 2D and 3D, we have miniaturized and adapted for high-throughput screening (HTS) the soft agar colony formation assay, employing a laser-scanning cytometer to image and quantify multiple cell types simultaneously. The assay is HTS compatible, providing high-quality, image-based, replicable data for multiple, co-cultured cell types. As proof of concept, we subjected colorectal carcinoma colonies in 3D soft agar to a mini screen of 1528 natural product compounds. Hit compounds from the primary screen were rescreened in an HTS 3D co-culture matrix containing colon stromal cells and cancer cells. By combining tumor cells and normal, nontransformed colon epithelial cells in one primary screening assay, we were able to obtain differential IC50 data, thereby distinguishing tumor-specific compounds from general cytotoxic compounds. Moreover, we were able to identify compounds that antagonized tumor colony formation in 3D only, highlighting the importance of this assay in identifying agents that interfere with 3D tumor structural growth. This screening platform provides a fast, simple, and robust method for identification of tumor-specific agents in a biologically relevant microenvironment.

Antibacterial activity of selected Malaysian honey

PubMed Central

2013-01-01

Background Antibacterial activity of honey is mainly dependent on a combination of its peroxide activity and non-peroxide components. This study aims to investigate antibacterial activity of five varieties of Malaysian honey (three monofloral; acacia, gelam and pineapple, and two polyfloral; kelulut and tualang) against Staphylococcus aureus, Bacillus cereus, Escherichia coli, and Pseudomonas aeruginosa. Methods Minimum Inhibitory Concentration (MIC) and Minimum Bactericidal Concentration (MBC) were performed for semi-quantitative evaluation. Agar well diffusion assay was used to investigate peroxide and non-peroxide activities of honey. Results The results showed that gelam honey possessed lowest MIC value against S. aureus with 5% (w/v) MIC and MBC of 6.25% (w/v). Highest MIC values were shown by pineapple honey against E. coli and P. aeruginosa as well as acacia honey against E. coli with 25% (w/v) MIC and 50% (w/v) MBC values. Agar inhibition assay showed kelulut honey to possess highest total antibacterial activity against S. aureus with 26.49 equivalent phenol concentrations (EPC) and non-peroxide activity of 25.74 EPC. Lowest antibacterial activity was observed in acacia honey against E. coli with total activity of 7.85 EPC and non-peroxide activity of 7.59 EPC. There were no significant differences (p > 0.05) between the total antibacterial activities and non-peroxide activities of Malaysian honey. The intraspecific correlation between MIC and EPC of E. coli (r = -0.8559) was high while that between MIC and EPC of P. aeruginosa was observed to be moderate (r = -0.6469). S. aureus recorded a smaller correlation towards the opposite direction (r = 0.5045). In contrast, B.cereus showed a very low intraspecific correlation between MIC and EPC (r = -0.1482). Conclusions Malaysian honey, namely gelam, kelulut and tualang, have high antibacterial potency derived from total and non-peroxide activities, which implies that both peroxide and other constituents are mutually important as contributing factors to the antibacterial property of honey. PMID:23758747

Antibacterial activity of selected Malaysian honey.

PubMed

Zainol, Mohd Izwan; Mohd Yusoff, Kamaruddin; Mohd Yusof, Mohd Yasim

2013-06-10

Antibacterial activity of honey is mainly dependent on a combination of its peroxide activity and non-peroxide components. This study aims to investigate antibacterial activity of five varieties of Malaysian honey (three monofloral; acacia, gelam and pineapple, and two polyfloral; kelulut and tualang) against Staphylococcus aureus, Bacillus cereus, Escherichia coli, and Pseudomonas aeruginosa. Minimum Inhibitory Concentration (MIC) and Minimum Bactericidal Concentration (MBC) were performed for semi-quantitative evaluation. Agar well diffusion assay was used to investigate peroxide and non-peroxide activities of honey. The results showed that gelam honey possessed lowest MIC value against S. aureus with 5% (w/v) MIC and MBC of 6.25% (w/v). Highest MIC values were shown by pineapple honey against E. coli and P. aeruginosa as well as acacia honey against E. coli with 25% (w/v) MIC and 50% (w/v) MBC values. Agar inhibition assay showed kelulut honey to possess highest total antibacterial activity against S. aureus with 26.49 equivalent phenol concentrations (EPC) and non-peroxide activity of 25.74 EPC. Lowest antibacterial activity was observed in acacia honey against E. coli with total activity of 7.85 EPC and non-peroxide activity of 7.59 EPC. There were no significant differences (p > 0.05) between the total antibacterial activities and non-peroxide activities of Malaysian honey. The intraspecific correlation between MIC and EPC of E. coli (r = -0.8559) was high while that between MIC and EPC of P. aeruginosa was observed to be moderate (r = -0.6469). S. aureus recorded a smaller correlation towards the opposite direction (r = 0.5045). In contrast, B.cereus showed a very low intraspecific correlation between MIC and EPC (r = -0.1482). Malaysian honey, namely gelam, kelulut and tualang, have high antibacterial potency derived from total and non-peroxide activities, which implies that both peroxide and other constituents are mutually important as contributing factors to the antibacterial property of honey.

Characterization and bioactivities of a novel polysaccharide obtained from Gracilariopsis lemaneiformis.

PubMed

Shi, Chen-Shan; Sang, Ya-Xin; Sun, Gui-Qing; Li, Tian-Ye; Gong, Zheng-Si; Wang, Xiang-Hong

2017-01-01

Gracilariopsis lemaneiformis is a type of red alga that contains seaweed polysaccharide agar. In this study, a novel non-agar seaweed polysaccharide fraction named GCP (short of crude polysaccharide obtained from Gracilariopsis lemaneiformis) was isolated from Gracilariopsis lemaneiformis. Structural analysis showed that GCP shows triple helical chain conformation when dissolved in water and has many branches and long side chains. Also, 1→3 linkage is the major linkage and the sugar structures are galactopyranose configurations linked by β-type glycosidic linkages. Two macromolecular substance fractions (GCP-1 and GCP-2) were purified by DEAE Sepharose Fast Flow column chromatography. Moreover, a splenocyte damage assay and splenocyte proliferation assay were used to analyse the bioactivities of GCP, GCP-1 and GCP-2. It was demonstrated that polysaccharides could protect splenocyte damaged by H2O2; GCP-2 shows a greatest protection rate, that is, 92.8%, which significantly enhanced the splenocyte proliferation, and GCP showed the highest proliferation rate, 9.30%. The results suggested that this type of novel non-agar polysaccharide displayed remarkable antioxidant and immunomodulatory activities and early alkali treatment could decrease the activities. It may represent a potential material for health food and clinical medicines.

Screening of some essential oils against Trichosporon species.

PubMed

Uniyal, Veena; Saxena, Seema; Bhatt, R P

2013-01-01

White Piedra is a superficial mycoses characterized by nodules on the hair shaft, caused by the basidiomycetous yeast Trichosporon species. In this study 25 essential oils were extracted and screened against two Trichosporon species i.e. Trichosporon asahii and Trichosporon cutaneum. Both these fungi procured from MTCC Chandigarh were maintained on yeast malt agar plates and tubes at 25 degrees C. Two screening methods viz., agar well diffusion assay and minimum inhibitory concentration were adopted for the study. The results showed that the maximum anti-yeast activity against T. asahii and T. cutaneum was demonstrated by oil of Mentha piperita showing full inhibition of both the fungi, Melaleuca alternifolia with an inhibition zone of 45 and 40 mm, Cymbopogon winterians with inhibition zone of 45 and 45 mm and Cymbopogon flexuosus with 35 and 30 mm inhibition zones. The oil of Trachyspermum ammi exhibited 10 and 20 mm, Abelmoschus moschatus exhibited 30 and 20 mm, Salvia sclarea showed 20 and 18 mm and Jasminum officinale exhibited 25 and 15 mm inhibition zones showing moderate activity. The oil of Cyperus scariosus, Pogostemon patchouli and Rosa damascene showed no inhibition zone against both the fungi while Vetiveria zizanoides exhibited no inhibition in case of T. asahii and inhibition zone of 10 mm in case of T. cutaneum demonstrating comparatively low activity against both the fungi. These results support that the essential oils can be used to cure superficial mycoses and these oils may have significant role as pharmaceuticals and preservatives.

Native Brazilian plants against nosocomial infections: a critical review on their potential and the antimicrobial methodology.

PubMed

H Moreno, Paulo Roberto; da Costa-Issa, Fabiana Inácio; Rajca-Ferreira, Agnieszka K; Pereira, Marcos A A; Kaneko, Telma M

2013-01-01

The growing incidences of drug-resistant pathogens have increased the attention on several medicinal plants and their metabolites for antimicrobial properties. These pathogens are the main cause of nosocomial infections which led to an increasing mortality among hospitalized patients. Taking into consideration those factors, this paper reviews the state-of-the-art of the research on antibacterial agents from native Brazilian plant species related to nosocomial infections as well as the current methods used in the investigations of the antimicrobial activity and points out the differences in techniques employed by the authors. The antimicrobial assays most frequently used were broth microdilution, agar diffusion, agar dilution and bioautography. The broth microdilution method should be the method of choice for testing new antimicrobial agents from plant extracts or isolated compounds due to its advantages. At the moment, only a small part of the rich Brazilian flora has been investigated for antimicrobial activity, mostly with unfractionated extracts presenting a weak or moderate antibacterial activity. The combination of crude extract with conventional antibiotics represents a largely unexploited new form of chemotherapy with novel and multiple mechanisms of action that can overcome microbial resistance that needs to be further investigated. The antibacterial activity of essential oil vapours might also be an interesting alternative treatment of hospital environment due to their ability in preventing biofilm formation. However, in both alternatives more studies should be done on their mode of action and toxicological effects in order to optimize their use.

Molecular method for detection of total coliforms in drinking water samples.

PubMed

Maheux, Andrée F; Boudreau, Dominique K; Bisson, Marc-Antoine; Dion-Dupont, Vanessa; Bouchard, Sébastien; Nkuranga, Martine; Bergeron, Michel G; Rodriguez, Manuel J

2014-07-01

This work demonstrates the ability of a bacterial concentration and recovery procedure combined with three different PCR assays targeting the lacZ, wecG, and 16S rRNA genes, respectively, to detect the presence of total coliforms in 100-ml samples of potable water (presence/absence test). PCR assays were first compared to the culture-based Colilert and MI agar methods to determine their ability to detect 147 coliform strains representing 76 species of Enterobacteriaceae encountered in fecal and environmental settings. Results showed that 86 (58.5%) and 109 (74.1%) strains yielded a positive signal with Colilert and MI agar methods, respectively, whereas the lacZ, wecG, and 16S rRNA PCR assays detected 133 (90.5%), 111 (75.5%), and 146 (99.3%) of the 147 total coliform strains tested. These assays were then assessed by testing 122 well water samples collected in the Québec City region of Canada. Results showed that 97 (79.5%) of the samples tested by culture-based methods and 95 (77.9%), 82 (67.2%), and 98 (80.3%) of samples tested using PCR-based methods contained total coliforms, respectively. Consequently, despite the high genetic variability of the total coliform group, this study demonstrated that it is possible to use molecular assays to detect total coliforms in potable water: the 16S rRNA molecular assay was shown to be as efficient as recommended culture-based methods. This assay might be used in combination with an Escherichia coli molecular assay to assess drinking water quality. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

Molecular Method for Detection of Total Coliforms in Drinking Water Samples

PubMed Central

Boudreau, Dominique K.; Bisson, Marc-Antoine; Dion-Dupont, Vanessa; Bouchard, Sébastien; Nkuranga, Martine; Bergeron, Michel G.; Rodriguez, Manuel J.

2014-01-01

This work demonstrates the ability of a bacterial concentration and recovery procedure combined with three different PCR assays targeting the lacZ, wecG, and 16S rRNA genes, respectively, to detect the presence of total coliforms in 100-ml samples of potable water (presence/absence test). PCR assays were first compared to the culture-based Colilert and MI agar methods to determine their ability to detect 147 coliform strains representing 76 species of Enterobacteriaceae encountered in fecal and environmental settings. Results showed that 86 (58.5%) and 109 (74.1%) strains yielded a positive signal with Colilert and MI agar methods, respectively, whereas the lacZ, wecG, and 16S rRNA PCR assays detected 133 (90.5%), 111 (75.5%), and 146 (99.3%) of the 147 total coliform strains tested. These assays were then assessed by testing 122 well water samples collected in the Québec City region of Canada. Results showed that 97 (79.5%) of the samples tested by culture-based methods and 95 (77.9%), 82 (67.2%), and 98 (80.3%) of samples tested using PCR-based methods contained total coliforms, respectively. Consequently, despite the high genetic variability of the total coliform group, this study demonstrated that it is possible to use molecular assays to detect total coliforms in potable water: the 16S rRNA molecular assay was shown to be as efficient as recommended culture-based methods. This assay might be used in combination with an Escherichia coli molecular assay to assess drinking water quality. PMID:24771030

Cytotoxic and antifungal activities of melleolide antibiotics follow dissimilar structure-activity relationships.

PubMed

Bohnert, Markus; Nützmann, Hans-Wilhelm; Schroeckh, Volker; Horn, Fabian; Dahse, Hans-Martin; Brakhage, Axel A; Hoffmeister, Dirk

2014-09-01

The fungal genus Armillaria is unique in that it is the only natural source of melleolide antibiotics, i.e., protoilludene alcohols esterified with orsellinic acid or its derivatives. This class of natural products is known to exert antimicrobial and cytotoxic effects. Here, we present a refined relationship between the structure and the antimicrobial activity of the melleolides. Using both agar diffusion and broth dilution assays, we identified the Δ(2,4)-double bond of the protoilludene moiety as a key structural feature for antifungal activity against Aspergillus nidulans, Aspergillus flavus, and Penicillium notatum. These findings contrast former reports on cytotoxic activities and may indicate a different mode of action towards susceptible fungi. We also report the isolation and structure elucidation of five melleolides (6'-dechloroarnamial, 6'-chloromelleolide F, 10-hydroxy-5'-methoxy-6'-chloroarmillane, and 13-deoxyarmellides A and B), along with the finding that treatment with an antifungal melleolide impacts transcription of A. nidulans natural product genes. Copyright © 2014 Elsevier Ltd. All rights reserved.

Antitumor and antifungal activities of organic extracts of seacucumber Holothuria atra from the southeast coast of India

NASA Astrophysics Data System (ADS)

Dhinakaran, Devaraj Isaac; Lipton, Aaron Premnath

2015-02-01

In phylum Echinodermata, the family Holothuridae is distinguished by its capacity of bioactive compounds. Sea cucumber Holothuria atra is commonly known as the lollyfish. The antifungal activity was detected using agar well diffusion method against the various fungal strains such as Trichoderma viride, Aspergillus niger, Aspergillus flavis, Candida albicans and Penicillium chrysogenum. Relatively high antifungal activity was seen against Candida albicans at 100 μL-1 concentration of extracts. Zone of inhibition was measured at 18 mm of diameter. The anti-tumor activities were detected against the Vero and Hep2 cell lines using MTT assay. The cells were treated with H. atra extract at concentrations 0.078-10mg mL-1. The extract showed high proliferative activity against the Hep2 cells. The body wall extracts of sea cucumber ( H. atra) showed effective antifungal and antitumor activities. All these findings suggest that the extracts could be used for the development of drugs.

Assessment of antinociceptive, antipyretic and antimicrobial activity of Piper cubeba L. essential oil in animal models.

PubMed

Mothana, Ramzi; Alsaid, Mansour; Khaled, Jamal M; Alharbi, Naiyf S; Alatar, Abdulrahman; Raish, Mohammad; Al-Yahya, Mohammed; Rafatullah, Syed; Parvez, Mohammad Khalid; Ahamad, Syed Rizwan

2016-03-01

This study was designed to investigate the possible antiniciceptive, antipyretic and antimicrobial activities of the essential oil obtained from the fruits of Piper Cubeba (L.). To assess the antinociceptive and antipyretic activities, three doses (150, 300 and 600 mg/kg, i.p.) were tested in acetic acid-induced abdominal writhing, tail flick reaction and hot-plate and Brewer's yeast-induced hyperpyrexia test models in animals. Moreover, the antimicrobial activity was examined using agar diffusion method and broth micro-dilution assay for minimum inhibitory concentrations (MIC). The Piper Cubeba essential oil (PCEO) showed a marked antinociception (17, 30 and 54%) and an increase in reaction time in mice in the flick tailed and hot-plate tests. The brewer's yeast induced hyperpyrexia was decreased in a dose dependent manner. PCEO also exhibited a strong antimicrobial potential. These findings confirm the traditional analgesic indications of P. cubeba oil and provide persuasive evidence and support its use in Arab traditional medicine.

In Vitro Structural and Functional Evaluation of Gold Nanoparticles Conjugated Antibiotics

NASA Astrophysics Data System (ADS)

Saha, Biswarup; Bhattacharya, Jaydeep; Mukherjee, Ananda; Ghosh, Anup Kumar; Santra, Chitta Ranjan; Dasgupta, Anjan K.; Karmakar, Parimal

2007-12-01

Bactericidal efficacy of gold nanoparticles conjugated with ampicillin, streptomycin and kanamycin were evaluated. Gold nanoparticles (Gnps) were conjugated with the antibiotics during the synthesis of nanoparticles utilizing the combined reducing property of antibiotics and sodium borohydride. The conjugation of nanoparticles was confirmed by dynamic light scattering (DLS) and electron microscopic (EM) studies. Such Gnps conjugated antibiotics showed greater bactericidal activity in standard agar well diffusion assay. The minimal inhibitory concentration (MIC) values of all the three antibiotics along with their Gnps conjugated forms were determined in three bacterial strains, Escherichia coli DH5α, Micrococcus luteus and Staphylococcus aureus. Among them, streptomycin and kanamycin showed significant reduction in MIC values in their Gnps conjugated form whereas; Gnps conjugated ampicillin showed slight decrement in the MIC value compared to its free form. On the other hand, all of them showed more heat stability in their Gnps conjugated forms. Thus, our findings indicated that Gnps conjugated antibiotics are more efficient and might have significant therapeutic implications.

Development and characterization of bioactive edible films from spider crab (Maja crispata) chitosan incorporated with Spirulina extract.

PubMed

Balti, Rafik; Mansour, Mohamed Ben; Sayari, Nadhem; Yacoubi, Lamia; Rabaoui, Lotfi; Brodu, Nicolas; Massé, Anthony

2017-12-01

Active food packaging films based on crab chitosan and Spirulina extract (SE) were developed. The effects of the SE incorporation at different levels on physical (color, opacity water vapor and oxygen permeability) and mechanical (tensile strength and elongation at break) properties of chitosan films were investigated. FTIR was carried out to observe the potential modifications of the chitosan films when incorporated with SE. The obtained results suggested that incorporation of SE into chitosan films improved mechanical and barrier properties. The antioxidant activity of the chitosan/SE films was characterized by means of three different analytical assays (DPPH, FRAP and FIC). Crab chitosan edible films containing SE showed higher antioxidant activity, regardless concentrations and methods assayed. Furthermore, the antioxidant activity occurred in a concentration-dependent manner. The agar disc diffusion method was used to determine the antibacterial activities of chitosan edible films against Escherichia coli, Staphylococcus aureus, Pseudomonas aeruginosa, Listeria monocytogenes, Salmonella typhimurium, Bacillus subtilis and Bacillus cereus. The chitosan/SE films were more effective (p<0.05) against five of the seven tested bacteria. The obtained crab chitosan edible films incorporated with SE showed great potential to be used for active food packaging due to its excellent antioxidant and antibacterial activities. Copyright © 2017 Elsevier B.V. All rights reserved.

Anti-listerial activity of a polymeric film coated with hybrid coatings doped with Enterocin 416K1 for use as bioactive food packaging.

PubMed

Iseppi, Ramona; Pilati, Francesco; Marini, Michele; Toselli, Maurizio; de Niederhäusern, Simona; Guerrieri, Elisa; Messi, Patrizia; Sabia, Carla; Manicardi, Giuliano; Anacarso, Immacolata; Bondi, Moreno

2008-04-30

In this study, Enterocin 416K1, a bacteriocin produced by Enterococcus casseliflavus IM 416K1, was entrapped in an organic-inorganic hybrid coating applied to a LDPE (low-density polyethylene) film for its potential use in the active food packaging field. The antibacterial activity of the coated film was evaluated against Listeria monocytogenes NCTC 10888 by qualitative modified agar diffusion assay, quantitative determination in listeria saline solution suspension and direct contact with artificially contaminated food samples (frankfurters and fresh cheeses) stored at room and refrigeration temperatures. All investigations demonstrated that enterocin-activated coatings have a good anti-listeria activity. Qualitative tests showed a clear zone of inhibition in the indicator lawn in contact with and around the coated film. During the quantitative antibacterial evaluation the L. monocytogenes viable counts decreased to 1.5 log units compared to the control. The inhibitory capability was confirmed also in food-contact assays. In all food samples packed with coated films we observed a significant decrease in L. monocytogenes viable counts in the first 24 h compared to the control. This difference was generally maintained up to the seventh day and then decreased, with the exception of the cheese samples stored at refrigeration temperature.

Screening and Scoring of Antimicrobial and Biological Activities of Italian Vulnerary Plants against Major Oral Pathogenic Bacteria

PubMed Central

Ferrazzano, Gianmaria F.; Roberto, Lia; Catania, Maria Rosaria; Chiaviello, Angela; De Natale, Antonino; Roscetto, Emanuela; Pinto, Gabriele; Pollio, Antonino; Ingenito, Aniello; Palumbo, Giuseppe

2013-01-01

This study aims to evaluate the activity of Italian vulnerary plants against the most important oral pathogenic bacteria. This estimate was accomplished through a fivefold process: (a) a review of ethnobotanical and microbiological data concerning the Italian vulnerary plants; (b) the development of a scoring system to rank the plants; (c) the comparative assessment of microbiological properties; (d) the assessment of potential cytotoxic effects on keratinocyte-like cells and gingival fibroblasts in culture by XTT cell viability assay; (e) clinical evaluation of the most suitable plant extract as antibacterial agent in a home-made mouthwash. The study assays hexane (H), ethanol (E), and water (W) extracts from 72 plants. The agar diffusion method was used to evaluate the activity against Streptococcus mutans, Streptococcus sobrinus, Lactobacillus casei, and Actinomyces viscosus. Twenty-two plants showed appreciable activity. The extracts showing the strongest antibacterial power were those from Cotinus coggygria Scop., Equisetum hyemale L., Helichrysum litoreum Guss, Juniperus communis L., and Phyllitis scolopendrium (L.) Newman subsp. scolopendrium. The potential cytotoxic effect of these extracts was assessed. On the basis of these observations, a mouth-rinse containing the ethanolic extract of H. litoreum has been tested in vivo, resulting in reduction of the salivary concentration of S. mutans. PMID:24302963

A comparison of in vitro cytotoxicity assays in medical device regulatory studies.

PubMed

Liu, Xuemei; Rodeheaver, Denise P; White, Jeffrey C; Wright, Ann M; Walker, Lisa M; Zhang, Fan; Shannon, Stephen

2018-06-06

Medical device biocompatibility testing is used to evaluate the risk of adverse effects on tissues from exposure to leachates/extracts. A battery of tests is typically recommended in accordance with regulatory standards to determine if the device is biocompatible. In vitro cytotoxicity, a key element of the standards, is a required endpoint for all types of medical devices. Each validated cytotoxicity method has different methodology and acceptance criteria that could influence the selection of a specific test. In addition, some guidances are more specific than others as to the recommended test methods. For example, the International Organization for Standardization (ISO 1 ) cites preference for quantitative methods (e.g., tetrazolium (MTT/XTT), neutral red (NR), or colony formation assays (CFA)) over qualitative methods (e.g., elution, agar overlay/diffusion, or direct), while a recent ISO standard for contact lens/lens care solutions specifically requires a qualitative direct test. Qualitative methods are described in United States Pharmacopeia (USP) while quantitative CFAs are listed in Japan guidance. The aim of this review is to compare the methodologies such as test article preparation, test conditions, and criteria for six cytotoxicity methods recommended in regulatory standards in order to inform decisions on which method(s) to select during the medical device safety evaluation. Copyright © 2018. Published by Elsevier Inc.

Evidence for Anti-Pseudogymnoascus destructans (Pd) Activity of Propolis

PubMed Central

Ghosh, Soumya; McArthur, Robyn; Guo, Zhi Chao; McKerchar, Rory; Donkor, Kingsley

2017-01-01

White-nose syndrome (WNS) in bats, caused by Pseudogymnoascus destructans (Pd), is a cutaneous infection that has devastated North American bat populations since 2007. At present, there is no effective method for controlling this disease. Here, we evaluated the effect of propolis against Pd in vitro. Using Sabouraud dextrose agar (SDA) medium, approximately 1.7 × 107 conidia spores of the Pd strain M3906-2/mL were spread on each plate and grown to form a consistent lawn. A Kirby–Bauer disk diffusion assay was employed using different concentrations of propolis (1%, 2%, 3%, 4%, 5%, 10%, 15%, 20%, 25%), in plates incubated at 8 °C and 15 °C. At 8 °C and 15 °C, as the concentration of propolis increased, there was an increasing zone of inhibition (ZOI), reaching the highest degree at 10% and 25% concentrations, respectively. A germule suppression assay showed a similar effect on Pd conidia germination. A MALDI-TOF-MS analysis of propolis revealed multiple constituents with a potential anti-Pd activity, including cinnamic acid, p-coumaric acid, and dihydrochalcones, which could be further tested for their individual effects. Our study suggests that propolis or its individual constituents might be suitable products against Pd. PMID:29267199

Anti-infective properties of Lactobacillus fermentum against Staphylococcus aureus and Pseudomonas aeruginosa.

PubMed

Varma, Parvathi; Nisha, N; Dinesh, Kavitha R; Kumar, Anil V; Biswas, Raja

2011-01-01

Surgical wounds and implant-associated Staphylococcus aureus and Pseudomonas aeruginosa infections are often difficult to treat because of limited susceptibility of several of these strains to conventional antibiotics. As a result, there is a constant need for new alternative drugs. The aim of this study was to investigate the antimicrobial properties of Lactobacillus fermentum, a probiotic bacterium, which we have isolated from colonic biopsies. The inhibition of S. aureus and P. aeruginosa growth was evaluated by coincubating with L. fermentum strains. Growth inhibition was tested for several of their clinical isolates using agar well diffusion assays. For biofilm assay S. aureus and P. aeruginosa were grown on the glass slides and in 96-well plates in presence of 2.5 μg/ml culture filtrate of L. fermentum. Biofilms were photographed using confocal microscope or stained with 0.1% crystal violet. Reduction in the cytotoxicity of S. aureus and P. aeruginosa was observed in presence of 2.5 μg/ml L. fermentum-spent media. Using in vitroexperiments, we showed that L. fermentum-secreted compound(s) inhibits the growth, cytotoxicity and biofilm formation of several S. aureus and P. aeruginosa strains. Compound(s) present in the culture supernatant of L. fermentum may have promising applications in treating hospital-acquired infections. Copyright © 2011 S. Karger AG, Basel.

Antibacterial synergic effect of honey from two stingless bees: Scaptotrigona bipunctata Lepeletier, 1836, and S. postica Latreille, 1807

NASA Astrophysics Data System (ADS)

Nishio, E. K.; Ribeiro, J. M.; Oliveira, A. G.; Andrade, C. G. T. J.; Proni, E. A.; Kobayashi, R. K. T.; Nakazato, G.

2016-02-01

Several studies have tested antimicrobial activity of combinations of honey and various substances. In this study, we tested a combination of two stingless bee honeys against various bacterial strains. In particular: the antibacterial activity of honeys produced by Scaptotrigona bipunctata (SB) and Scaptotrigona postica (SP) was evaluated against Gram-positive and Gram-negative bacterial strains by agar well diffusion assays, minimum inhibitory concentration (MIC) assessment, construction of growth and viability curves and scanning electron microscopy (SEM). The interaction of the two honeys was also evaluated by the checkerboard assay. Inhibition zones ranged from 8 to 22 mm. The MIC values of the individual honeys ranged from 0.62 to 10% (v v-1) and decreased to 1/4 to 1/32 when the honeys were combined. SEM images showed division inhibition and cell wall disruption for the SB and SP honeys, respectively, and these alterations were observed in same field when the SB and SP honeys were combined. This study demonstrated that the natural honeys possess in vitro antimicrobial activity against Gram-positive and Gram-negative bacteria, including multidrug-resistant strains. Combination of the SB and SP honeys could lead to the development of new broad-spectrum antimicrobials that have the potential to prevent the emergence of resistant bacterial strains.

Antibacterial synergic effect of honey from two stingless bees: Scaptotrigona bipunctata Lepeletier, 1836, and S. postica Latreille, 1807

PubMed Central

Nishio, E. K.; Ribeiro, J. M.; Oliveira, A. G.; Andrade, C. G. T. J.; Proni, E. A.; Kobayashi, R. K. T.; Nakazato, G.

2016-01-01

Several studies have tested antimicrobial activity of combinations of honey and various substances. In this study, we tested a combination of two stingless bee honeys against various bacterial strains. In particular: the antibacterial activity of honeys produced by Scaptotrigona bipunctata (SB) and Scaptotrigona postica (SP) was evaluated against Gram-positive and Gram-negative bacterial strains by agar well diffusion assays, minimum inhibitory concentration (MIC) assessment, construction of growth and viability curves and scanning electron microscopy (SEM). The interaction of the two honeys was also evaluated by the checkerboard assay. Inhibition zones ranged from 8 to 22 mm. The MIC values of the individual honeys ranged from 0.62 to 10% (v v−1) and decreased to 1/4 to 1/32 when the honeys were combined. SEM images showed division inhibition and cell wall disruption for the SB and SP honeys, respectively, and these alterations were observed in same field when the SB and SP honeys were combined. This study demonstrated that the natural honeys possess in vitro antimicrobial activity against Gram-positive and Gram-negative bacteria, including multidrug-resistant strains. Combination of the SB and SP honeys could lead to the development of new broad-spectrum antimicrobials that have the potential to prevent the emergence of resistant bacterial strains. PMID:26869239

Anti-microbial activity and composition of manuka and portobello honey.

PubMed

Schneider, Monika; Coyle, Shirley; Warnock, Mary; Gow, Iain; Fyfe, Lorna

2013-08-01

Recently renewed interest in the therapeutic properties of honey has led to the search for new antimicrobial honeys. This study was undertaken to assess the antimicrobial activity and composition of a locally produced Portobello honey (PBH) on three bacteria known to infect wounds. Manuka honey (MH) was used for comparative purposes. Broth culture and agar disc diffusion assays were used to investigate the antimicrobial properties of honey. The honeys were tested at four concentrations: 75%, 50%, 10% and 1% (v/v) and compared with an untreated control. The composition of honey was determined by measuring: polyphenol content by Folin Ciocalteau method, antioxidant capacity by ferric ion reducing power assay, hydrogen peroxide (H2 O2 ) by catalase test, pH and sugar content by pH strips and refractometer, respectively. Both honeys at 75% and 50% inhibited the majority of the three bacteria tested. 10% PBH exhibited antimicrobial activity to the lesser extent than 10% MH. The difference was very significant (p ≤ 0.001). Both honeys were acidic with pH 4, and both produced H2 O2 . The sugar content of PBH was higher than MH, but the difference was not significant. The MH had significantly higher levels of the polyphenols and antioxidant activity than PBH. Copyright © 2012 John Wiley & Sons, Ltd.

validated microbiological and HPLC methods for the determination of moxifloxacin in pharmaceutical preparations and human plasma.

PubMed

Abdelaziz, Ahmed A; Elbanna, Tarek E; Gamaleldeen, Noha M

2012-10-01

The article presents a comparison between microbiological and high performance liquid chromatographic (HPLC) assays for quantification of moxifloxacin in tablets, ophthalmic solutions and human plasma. The microbiological method employed a cylinder-plate agar diffusion assay using a strain of Esherichia coli ATCC 25922 as the test organism and phosphate buffer (pH8) as the diluent. The calibration curves were linear (R(2) > 0.98) over a concentration range of 0.125 to 16 µgml(-1). The within day and between days precisions were ≤ 4.47% and ≤ 6.39% respectively. Recovery values were between 89.4 and 110.2%. The HPLC assay used Hypersil(®) BDS C18 reversed phase column (250×4.6 mm, 5µm) with a mobile phase comprising 20 mM ammonium dihydrogen orthophosphate (pH3) and acetonitrile (75:25) and flowing at 1.5 ml/min. The detection was at 295nm. The calibration curves were linear (R(2) > 0.999) over the range of 0.125 to 16 µg ml(-1). The within day and between days precisions were ≤ 4.07% and ≤ 5.09% respectively. Recovery values were between 97.7 and 107.6%. Similar potencies were obtained after the analysis of moxifloxacin tablets and ophthalmic solutions by both methods. Also pharmacokinetic parameters were calculated after the analysis of plasma samples of six male healthhy volunteers by both validated methods.

Comparison of methods for determining the effectiveness of antibacterial functionalized textiles.

PubMed

Haase, Hajo; Jordan, Lisa; Keitel, Laura; Keil, Claudia; Mahltig, Boris

2017-01-01

Antimicrobial functionalization of textiles is important for various applications, such as protection of textile materials from decomposition, generation of more effective wound dressings, and the prevention of infections or malodors resulting from bacterial growth. In order to test the efficacy of new products, their antibacterial activity needs to be evaluated. At present, several different procedures are being used for this purpose, hindering comparisons among different studies. The present paper compares five of these assays using a sample panel of different textiles functionalized with copper (Cu) and silver (Ag) as antibacterial agents, and discusses the suitability of these methods for different analytical requirements. Bacterial viability was determined by measuring the optical density at 600 nm, a colorimetric assay based on MTT (3-[4, 5-dimethylthiazol-2-yl]-2, 5 diphenyl tetrazolium bromide) conversion, an agar diffusion assay, and colony formation, either after culturing in media containing textile samples, or after recovery from textiles soaked with bacterial suspension. All experiments were performed with a Gram-negative (Escherichia coli) and a Gram-positive (Staphylococcus warneri) model organism. In general, the results yielded by the different methods were of good comparability. To identify the most suitable test system for the particular type of antibacterial coating, several factors need to be taken into account, such as choosing appropriate endpoints for analyzing passive or active antibacterial effects, selection of relevant microorganisms, correcting for potential interference by leaching of colored textile coatings, required hands on time, and the necessary sensitivity.

Comparison of methods for determining the effectiveness of antibacterial functionalized textiles

PubMed Central

Jordan, Lisa; Keitel, Laura; Keil, Claudia; Mahltig, Boris

2017-01-01

Antimicrobial functionalization of textiles is important for various applications, such as protection of textile materials from decomposition, generation of more effective wound dressings, and the prevention of infections or malodors resulting from bacterial growth. In order to test the efficacy of new products, their antibacterial activity needs to be evaluated. At present, several different procedures are being used for this purpose, hindering comparisons among different studies. The present paper compares five of these assays using a sample panel of different textiles functionalized with copper (Cu) and silver (Ag) as antibacterial agents, and discusses the suitability of these methods for different analytical requirements. Bacterial viability was determined by measuring the optical density at 600 nm, a colorimetric assay based on MTT (3-[4, 5-dimethylthiazol-2-yl]-2, 5 diphenyl tetrazolium bromide) conversion, an agar diffusion assay, and colony formation, either after culturing in media containing textile samples, or after recovery from textiles soaked with bacterial suspension. All experiments were performed with a Gram-negative (Escherichia coli) and a Gram-positive (Staphylococcus warneri) model organism. In general, the results yielded by the different methods were of good comparability. To identify the most suitable test system for the particular type of antibacterial coating, several factors need to be taken into account, such as choosing appropriate endpoints for analyzing passive or active antibacterial effects, selection of relevant microorganisms, correcting for potential interference by leaching of colored textile coatings, required hands on time, and the necessary sensitivity. PMID:29161306

Antimicrobial activity of the imipenem/rifampicin combination against clinical isolates of Acinetobacter baumannii grown in planktonic and biofilm cultures.

PubMed

Wang, Yang; Bao, Wanguo; Guo, Na; Chen, Haiying; Cheng, Wei; Jin, Kunqi; Shen, Fengge; Xu, Jiancheng; Zhang, Qiaoli; Wang, Chao; An, Yanan; Zhang, Kaiyu; Wang, Feng; Yu, Lu

2014-12-01

To investigate the antimicrobial activity of imipenem and rifampicin alone and in combination against clinical isolates of Acinetobacter baumannii grown in planktonic and biofilm cultures. Minimum inhibitory concentrations were determined for each isolate grown in suspension and in biofilm using a microbroth dilution method. Chequerboard assays and the agar disk diffusion assay were used to determine synergistic, indifferent or antagonistic interactions between imipenem and rifampicin. We used the tissue culture plate method for A. baumannii biofilm formation to measure the percentage of biofilm inhibition and the amount of extracellular DNA after the treatment. To understand the synergistic mechanisms, we conducted hydroxyl radical formation assays. The results were verified by confocal laser scanning microscopy. Imipenem and rifampicin showed effective antimicrobial activity against suspensions and biofilm cultures of A. baumannii, respectively. Synergistic antimicrobial effects between imipenem and rifampicin were observed in 13 and 17 of the 20 clinical isolates when in suspension and in biofilms, respectively. Imipenem and rifampicin alone and in combination generated hydroxyl radicals, which are highly reactive oxygen forms and the major components of bactericidal agents. Furthermore, treatment with imipenem and rifampicin individually or in combination has obvious antibiofilm effects. The synergistic activity of imipenem and rifampicin against clinical isolates of A. baumannii (in suspension and in biofilms) was observed in vitro. Therefore, we conclude that imipenem combined with rifampicin has the potential to be used as a combinatorial therapy for the treatment of infectious diseases caused by A. baumannii.

A vapour phase assay for evaluating the antimicrobial activities of essential oils against bovine respiratory bacterial pathogens.

PubMed

Amat, S; Baines, D; Alexander, T W

2017-12-01

The objectives of this study were to develop a new assay for the evaluation of the antimicrobial activities of essential oils (EOs) in vapour phase and to demonstrate the antimicrobial activities of commercial EOs against BRPs. To achieve the first objective, a microtube cap containing 100 μl of EO was embedded in an agar plate. An agar plug (diameter 13 mm) inoculated with a bacterial suspension containing10 8  CFU per ml was then placed over the cap and incubated at 37°C for 24 h. Subsequently, bacteria were recovered from the agar plug by immersion in 5 ml of broth for 10 min, followed by vortexing for 30 s, and the broths were then plated for enumeration. To demonstrate the usefulness of the assay, nine commercial EOs derived from the following specific plants: ajowan, carrot seed, cinnamon leaf, citronella, fennel, ginger grass, lavender, rosemary and thyme were first evaluated for their vapour phase antimicrobial activities against Mannheimia haemolytica serotype 1. Selected EOs were further tested against Pasteurella multocida and Histophilus somni. The EOs of ajowan, thyme and cinnamon leaf completely or partially inhibited BRPs growth. This new assay provided reproducible results on the vapour phase antimicrobial activities of EOs against BRPs. These results support further study of EOs as a potential mitigation strategy against BRPs. In this study, we present a new vapour phase assay for evaluating the antimicrobial activities of essential oils (EO) against bovine respiratory pathogens (BRPs). Using this assay, we identified EOs, such as ajowan, thyme and cinnamon leaf, that can effectively inhibit growth of the BRPs Mannheimia haemolytica serotype 1, Pasteurella multocida and Histophilus somni. This is the first study to demonstrate the vapour phase antimicrobial activity of EOs against BRPs. © 2017 Her Majesty the Queen in Right of Canada. © 2017 The Society for Applied Microbiology Reproduced with the permission of the Minister of the Department of Agriculture and Agri-Food Canada.

Essential oil of the leaves of Ricinus communis L.: in vitro cytotoxicity and antimicrobial properties.

PubMed

Zarai, Zied; Ben Chobba, Ines; Ben Mansour, Riadh; Békir, Ahmed; Gharsallah, Néji; Kadri, Adel

2012-08-13

The aim of the present study was to appraise the antimicrobial activity of Ricinus communis L. essential oil against different pathogenic microorganisms and the cytotoxic activity against HeLa cell lines. The agar disk diffusion method was used to study the antibacterial activity of Ricinus communis L. essential oil against 12 bacterial and 4 fungi strains. The disc diameters of zone of inhibition (DD), the minimum inhibitory concentrations (MIC) and the concentration inhibiting 50% (IC50) were investigated to characterize the antimicrobial activities of this essential oil. The in vitro cytotoxicity of Ricinus communis L. essential oil was examined using a modified MTT assay; the viability and the IC50 were used to evaluate this test. The essential oil from the leaves of Ricinus communis L. was analyzed by GC-MS and bioassays were carried out. Five constituents of the oil were identified by GC-MS. The antimicrobial activity of the oil was investigated in order to evaluate its efficacy against twelve bacteria and four fungi species, using disc diffusion and minimum inhibitory concentration methods. The essential oil showed strong antimicrobial activity against all microorganisms tested with higher sensitivity for Bacillus subtilis, Staphylococcus aureus and Enterobacter cloacae. The cytotoxic and apoptotic effects of the essential oil on HeLa cell lines were examined by MTT assay. The cytotoxicity of the oil was quite strong with IC50 values less than 2.63 mg/ml for both cell lines. The present study showed the potential antimicrobial and anticarcinogenic properties of the essential oil of Ricinus communis L., indicating the possibilities of its potential use in the formula of natural remedies for the topical treatment of infections.

Polymerase chain reaction for screening clinical isolates of corynebacteria for the production of diphtheria toxin.

PubMed

Pallen, M J; Hay, A J; Puckey, L H; Efstratiou, A

1994-04-01

To assess the performance of the polymerase chain reaction (PCR) when used to screen rapidly large numbers of corynebacteria for toxin production; and to determine the incidence of false positive PCR results with non-toxigenic Corynebacterium diphtheriae isolates. Eighty seven recent British isolates of corynebacteria were assayed by PCR. All isolates were assayed from both blood and tellurite agar within a five day period. Thirty three non-toxigenic isolates of C diphtheriae from six countries were also tested by PCR and by the Elek immunodiffusion assay. There was complete concordance between the results of PCR and traditional methods on the recent British isolates, with one exception: an Elek positive "C ulcerans" isolate, which was PCR positive from tellurite but not from blood agar. One of the thirty three (3%) non-toxigenic isolates of C diphtheriae was PCR positive. These results suggest that PCR compares favourably with traditional methods for the detection of toxigenic corynebacteria and that it represents a powerful new tool in the diagnosis of an old disease.

Polymerase chain reaction for screening clinical isolates of corynebacteria for the production of diphtheria toxin.

PubMed Central

Pallen, M J; Hay, A J; Puckey, L H; Efstratiou, A

1994-01-01

AIMS--To assess the performance of the polymerase chain reaction (PCR) when used to screen rapidly large numbers of corynebacteria for toxin production; and to determine the incidence of false positive PCR results with non-toxigenic Corynebacterium diphtheriae isolates. METHODS--Eighty seven recent British isolates of corynebacteria were assayed by PCR. All isolates were assayed from both blood and tellurite agar within a five day period. Thirty three non-toxigenic isolates of C diphtheriae from six countries were also tested by PCR and by the Elek immunodiffusion assay. RESULTS--There was complete concordance between the results of PCR and traditional methods on the recent British isolates, with one exception: an Elek positive "C ulcerans" isolate, which was PCR positive from tellurite but not from blood agar. One of the thirty three (3%) non-toxigenic isolates of C diphtheriae was PCR positive. CONCLUSIONS--These results suggest that PCR compares favourably with traditional methods for the detection of toxigenic corynebacteria and that it represents a powerful new tool in the diagnosis of an old disease. Images PMID:8027375

Validation of an automated colony counting system for group A Streptococcus.

PubMed

Frost, H R; Tsoi, S K; Baker, C A; Laho, D; Sanderson-Smith, M L; Steer, A C; Smeesters, P R

2016-02-08

The practice of counting bacterial colony forming units on agar plates has long been used as a method to estimate the concentration of live bacteria in culture. However, due to the laborious and potentially error prone nature of this measurement technique, an alternative method is desirable. Recent technologic advancements have facilitated the development of automated colony counting systems, which reduce errors introduced during the manual counting process and recording of information. An additional benefit is the significant reduction in time taken to analyse colony counting data. Whilst automated counting procedures have been validated for a number of microorganisms, the process has not been successful for all bacteria due to the requirement for a relatively high contrast between bacterial colonies and growth medium. The purpose of this study was to validate an automated counting system for use with group A Streptococcus (GAS). Twenty-one different GAS strains, representative of major emm-types, were selected for assessment. In order to introduce the required contrast for automated counting, 2,3,5-triphenyl-2H-tetrazolium chloride (TTC) dye was added to Todd-Hewitt broth with yeast extract (THY) agar. Growth on THY agar with TTC was compared with growth on blood agar and THY agar to ensure the dye was not detrimental to bacterial growth. Automated colony counts using a ProtoCOL 3 instrument were compared with manual counting to confirm accuracy over the stages of the growth cycle (latent, mid-log and stationary phases) and in a number of different assays. The average percentage differences between plating and counting methods were analysed using the Bland-Altman method. A percentage difference of ±10 % was determined as the cut-off for a critical difference between plating and counting methods. All strains measured had an average difference of less than 10 % when plated on THY agar with TTC. This consistency was also observed over all phases of the growth cycle and when plated in blood following bactericidal assays. Agreement between these methods suggest the use of an automated colony counting technique for GAS will significantly reduce time spent counting bacteria to enable a more efficient and accurate measurement of bacteria concentration in culture.

Screening of antagonistic bacteria for biological control of nursery wilt of black pepper (Piper nigrum).

PubMed

Anith, K N; Radhakrishnan, N V; Manomohandas, T P

2003-01-01

Bacterial antagonists of Phytophthora capsici were isolated from underground shoot portions of rooted cuttings of black pepper. Initially isolates were screened by dual culture on potato dextrose agar and carrot agar. Further, a screening was done on black pepper shoots for supression of lesion caused by the pathogen. Most of the antagonists showed varying levels of antagonism in the dual culture and the shoot assay. Isolate PN-026, showing the highest suppression of lesion development in the shoot assay was found to be the most efficient antagonist in reducing Phytophthora capsici induced nursery wilt of black pepper. This screening involving the host, pathogen, and the antagonist, performed on black pepper shoot (the planting material for this vegetatively propagated crop), could be used as a rapid and reliable method for the isolation of efficient bacterial antagonists of P. capsici.

Infection Assay of Cyst Nematodes on Arabidopsis Roots.

PubMed

Bohlmann, Holger; Wieczorek, Krzysztof

2015-09-20

Plant parasitic nematodes are devastating pests on many crops. Juveniles (J2) of cyst nematodes invade the roots to induce a syncytium. This feeding site is their only source of nutrients. Male nematodes leave the roots after the fourth molt to mate with females. The females stay attached to their syncytia throughout their life and produce hundreds of eggs, which are contained in their bodies. When the females die their bodies form the cysts, which protect the eggs. Cysts can survive for many years in the soil until favorable conditions induce hatching of the juveniles. The beet cyst nematode Heterodera schachtii ( H. schachtii )is a pathogen of sugar beet ( Beta vulgaris ) but can also complete its life cycle on Arabidopsis roots growing on agar plates under sterile conditions. We present here protocols for a stock culture of H. schachtii and an infection assay on agar plates.

Antagonism of Bacillus spp. isolated from marine biofilms against terrestrial phytopathogenic fungi.

PubMed

Ortega-Morales, B O; Ortega-Morales, F N; Lara-Reyna, J; De la Rosa-García, S C; Martínez-Hernández, A; Montero-M, Jorge

2009-01-01

We aimed at determining the antagonistic behavior of bacteria derived from marine biofilms against terrestrial phytopathogenic fungi. Some bacteria closely related to Bacillus mojavensis (three isolates) and Bacillus firmus (one isolate) displayed antagonistic activity against Colletotrichum gloeosporioides ATCC 42374, selected as first screen organism. The four isolates were further quantitatively tested against C. gloeosporioides, Colletotrichum fragariae, and Fusarium oxysporum on two culture media, potato dextrose agar (PDA) and a marine medium-based agar [yeast extract agar (YEA)] at different times of growth of the antagonists (early, co-inoculation with the pathogen and late). Overall antagonistic assays showed differential susceptibility among the pathogens as a function of the type of culture media and time of colonization (P < 0.05). In general, higher suppressive activities were recorded for assays performed on YEA than on PDA; and also when the antagonists were allowed to grow 24 h earlier than the pathogen. F. oxysporum was the most resistant fungus while the most sensitive was C. gloeosporioides ATCC 42374. Significant differences in antagonistic activity (P < 0.05) were found between the different isolates. In general, Bacillus sp. MC3B-22 displayed a greater antagonistic effect than the commercial biocontrol strain Bacillus subtilis G03 (Kodiak). Further incubation studies and scanning electronic microscopy revealed that Bacillus sp. MC3B-22 was able to colonize, multiply, and inhibit C. gloeosporioides ATCC 42374 when tested in a mango leaf assay, showing its potential for fungal biocontrol. Additional studies are required to definitively identify the active isolates and to determine their mode of antifungal action, safety, and biocompatibility.

Films based on soy protein-agar blends for wound dressing: Effect of different biopolymer proportions on the drug release rate and the physical and antibacterial properties of the films.

PubMed

Rivadeneira, Josefina; Audisio, M C; Gorustovich, Alejandro

2018-04-01

No single material can provide all requirements for wound dressings. Here, we evaluated the influence of different soy protein isolate and agar proportions (3:1, 1:1, and 1:3) in blend films on some of their physical-chemical and antibacterial properties to elucidate their potential as wound dressings. The films were synthesized by the gel casting method and ciprofloxacin hydrochloride was incorporated into the films. Films were characterized based on their surface morphology, water uptake ability, and weight loss profile. Also, the ciprofloxacin hydrochloride release kinetics was quantified spectrophotometrically. The antibacterial effect was evaluated against Staphylococcus aureus and Pseudomonas aeruginosa strains. The soy protein isolate-agar ratio affected the water uptake of the films and the release profile of ciprofloxacin hydrochloride but not the weight loss profile. The amount of drug released decreased near 80% because of the decrease in agar content in the films. The release kinetics of ciprofloxacin hydrochloride data best fitted to the Korsmeyer-Peppas model, suggesting that the mechanism of drug release was mainly of the diffusion type. All ciprofloxacin hydrochloride-releasing soy protein isolate-agar films strongly inhibited the cell viability of the bacterial strains studied. We concluded that water uptake and ciprofloxacin hydrochloride release can be controlled by changing the soy protein isolate-agar proportion. The proportions did not lead to changes in the antibacterial strength of the films.

Evaluation of a Method for Rapid Detection of Listeria monocytogenes in Dry-Cured Ham Based on Impedanciometry Combined with Chromogenic Agar.

PubMed

Labrador, Mirian; Rota, María C; Pérez, Consuelo; Herrera, Antonio; Bayarri, Susana

2018-05-01

The food industry is in need of rapid, reliable methodologies for the detection of Listeria monocytogenes in ready-to-eat products, as an alternative to the International Organization of Standardization (ISO) 11290-1 reference method. The aim of this study was to evaluate impedanciometry combined with chromogenic agar culture for the detection of L. monocytogenes in dry-cured ham. The experimental setup consisted in assaying four strains of L. monocytogenes and two strains of Listeria innocua in pure culture. The method was evaluated according to the ISO 16140:2003 standard through a comparative study with the ISO reference method with 119 samples of dry-cured ham. Significant determination coefficients ( R 2 of up to 0.99) for all strains assayed in pure culture were obtained. The comparative study results had 100% accuracy, 100% specificity, and 100% sensitivity. Impedanciometry followed by chromogenic agar culture was capable of detecting 1 CFU/25 g of food. L. monocytogenes was not detected in the 65 commercial samples tested. The method evaluated herein represents a promising alternative for the food industry in its efforts to control L. monocytogenes. Overall analysis time is shorter and the method permits a straightforward analysis of a large number of samples with reliable results.

The Resazurin-Agar Method - a Quick Test to Determine Water Quality

NASA Astrophysics Data System (ADS)

Huckfeldt, J.; Westphal, B.; Claußen, L.

2015-12-01

Rezasurin has been used as a smart tracer in stream ecosystems to indicate metabolic activity, specifically aerobic respiration by heterotrophic bacteria. Resazurin is a blue compound which is irreversibly reduced to the pink resorufin in the presence of aerobic bacteria. The degree and speed of colour change from blue to pink is a measure of the degree of oxygen consumption and thus an indirect indication of the concentration of aerobic bacteria in a given medium. A high concentration of bacteria in water indicates a bad water quality. In our work a method was developed using resazurin agar plates to find a quick and easy way for testing water quality and comparing concentrations of bacteria in freshwater and seawater samples. The theory was to concentrate bacteria from a defined volume of water sample onto polycarbonate filters (0.2 μm), which are then placed onto the resazurin agar plate. The presence of aerobic bacteria on the filter will reduce the resazurin in the agar and the compound changes its colour. First tests conducted with different dilutions of a pure culture of yoghurt bacteria showed promising results and confirmed the feasibility of the method. In a further assay, we used water samples from different water layers and different temperatures and were also able to observe differences in the concentration of bacteria, depending on these different environmental conditions.The assay was also successfully used with seawater samples, collected from 2 different stations at 3 different depths in the Baltic Sea (salinity=15). The discolouration of the plates showed good correlation with the oxygen concentrations in the water. The resazurin-agar plate method is economical and fast. Several samples could be investigated at the same time without sacrificing the reliability of the results. Thus it is a good pre-screening test for a quantitative evaluation of bacteria in a water sample.

A Simple Assay to Screen Antimicrobial Compounds Potentiating the Activity of Current Antibiotics

PubMed Central

Iqbal, Junaid; Kazmi, Shahana Urooj; Khan, Naveed Ahmed

2013-01-01

Antibiotic resistance continues to pose a significant problem in the management of bacterial infections, despite advances in antimicrobial chemotherapy and supportive care. Here, we suggest a simple, inexpensive, and easy-to-perform assay to screen antimicrobial compounds from natural products or synthetic chemical libraries for their potential to work in tandem with the available antibiotics against multiple drug-resistant bacteria. The aqueous extract of Juglans regia tree bark was tested against representative multiple drug-resistant bacteria in the aforementioned assay to determine whether it potentiates the activity of selected antibiotics. The aqueous extract of J. regia bark was added to Mueller-Hinton agar, followed by a lawn of multiple drug-resistant bacteria, Salmonella typhi or enteropathogenic E. coli. Next, filter paper discs impregnated with different classes of antibiotics were placed on the agar surface. Bacteria incubated with extract or antibiotics alone were used as controls. The results showed a significant increase (>30%) in the zone of inhibition around the aztreonam, cefuroxime, and ampicillin discs compared with bacteria incubated with the antibiotics/extract alone. In conclusion, our assay is able to detect either synergistic or additive action of J. regia extract against multiple drug-resistant bacteria when tested with a range of antibiotics. PMID:23865073

Alternative method for quantification of alfa-amylase activity.

PubMed

Farias, D F; Carvalho, A F U; Oliveira, C C; Sousa, N M; Rocha-Bezerrra, L C B; Ferreira, P M P; Lima, G P G; Hissa, D C

2010-05-01

A modification of the sensitive agar diffusion method was developed for macro-scale determination of alfa-amylase. The proposed modifications lower costs with the utilisation of starch as substrate and agar as supporting medium. Thus, a standard curve was built using alfa-amylase solution from Aspergillus oryzae, with concentrations ranging from 2.4 to 7,500 U.mL-1. Clear radial diffusion zones were measured after 4 hours of incubation at 20 A degrees C. A linear relationship between the logarithm of enzyme activities and the area of clear zones was obtained. The method was validated by testing alpha-amylase from barley at the concentrations of 2.4; 60; 300 and 1,500 U.mL-1. The proposed method turned out to be simpler, faster, less expensive and able to determine on a macro-scale alpha-amylase over a wide range (2.4 to 7,500 U.mL-1) in scientific investigation as well as in teaching laboratory activities.

Antigenic analyses of tissues and excretory and secretory products from Strongylus vulgaris.

PubMed

Wynne, E; Slocombe, J O; Wilkie, B N

1981-07-01

Rabbit antisera were prepared against veronal buffered saline extracts of L4 and L5 Strongylus vulgaris, adult S. vulgaris and adult Strongylus equinus retrieved from naturally infected horses. In agar gel diffusion with these antisera, adult S vulgaris and S. equinus each appeared to have at least one unique antigen; larval S. vulgaris appeared to have two species-specific and two stage-specific antigens. There were several common antigens. Excretory and secretory products were collected also from L4 and L5 an maintained over several days in tissue culture fluid. In agar gel diffusion against the above rabbit antisera, a stage-specific antigen was found also in excretory and secretory products. In addition, excretory and secretory products had three antigens in common with adult and larval S. vulgaris, but only one of these was common to adult S. equinus. The excretory and secretory products appear, therefore, to have two species-specific and one stage-specific antigens.

Evaluation of antimicrobial efficacy of Aloe vera and its effectiveness in decontaminating gutta percha cones

PubMed Central

Athiban, Prakash P; Borthakur, Bikash Jyoti; Ganesan, S; Swathika, B

2012-01-01

Aim: The aim of this study was to evaluate the antimicrobial efficacy of Aloe vera and to determine its effectiveness in decontaminating gutta percha cones. Materials and Methods: A concentrated extract of Aloe vera was used to check for the antimicrobial efficacy using the agar well diffusion method. Presence of zones’ of diffusion was identified against three common GP contaminants namely, E.coli, E.faecalis and Staph. aureus. New GP Cones, freshly taken out of the packet were then decontaminated for 1minute using Aloe vera gel and then placed in thioglycolate broth to check for the presence of turbidity. Results: The zones of inhibition on the agar plate were measured as 24mm,21mm and 24mm respectively. The broth remained clear even after 48 hours of incubation. Conclusion: We conclude that Aloe vera is indeed effective as a GP decontaminant and it holds a promising future as a medium for storage of GP cones. PMID:22876011

INFLUENCE OF IODOFORM ON ANTIMICROBIAL POTENTIAL OF CALCIUM HYDROXIDE

PubMed Central

Estrela, Carlos; Estrela, Cyntia Rodrigues de Araújo; Hollanda, Augusto César Braz; Decurcio, Daniel de Almeida; Pécora, Jesus Djalma

2006-01-01

The purpose of this research was to verify the influence of Iodoform on antimicrobial potential of calcium hydroxide. S. aureus, E. faecalis, P. aeruginosa, B. subtilis, C. albicans were the biological indicators. The substances tested were: calcium hydroxide + saline; calcium hydroxide + Iodoform + saline; Iodoform + saline. For the agar diffusion test, 18 Petri plates with 20 ml of BHI agar were inoculated with the microbial suspensions. Fifty-four cavities were made and filled with the substances tested. The diameters of microbial inhibition were then measured. In direct exposure test, 162 #50 sterile absorbent paper points were immersed in the experimental suspensions for 5 min, and covered with the pastes. At intervals of 24, 48 and 72 hours, the paper points were immersed in 10 ml of Letheen Broth, followed by incubation at 37°°C for 48h. Microbial growth was evaluated by turbidity of the culture medium. A 0.1 ml inoculum obtained from the Letheen Broth was transferred to 7 ml of BHI, and incubated at 37°°C for 48h. Bacterial growth was again evaluated by turbidity of the culture medium. The calcium hydroxide associated with the saline or the iodoform plus saline showed antimicrobial effectiveness in both experimental methods. The iodoform paste presented antimicrobial ineffectiveness for the agar diffusion test on all biological microorganisms and for the direct exposure test on B. subtilis and on the mixture. PMID:19089027

Assessment of Antioxidant and Antibacterial Properties on Meat Homogenates of Essential Oils Obtained from Four Thymus Species Achieved from Organic Growth

PubMed Central

Ballester-Costa, Carmen; Viuda-Martos, Manuel

2017-01-01

In the organic food industry, no chemical additives can be used to prevent microbial spoilage. As a consequence, the essential oils (EOs) obtained from organic aromatic herbs and spices are gaining interest for their potential as preservatives. The organic Thymus zygis, Thymus mastichina, Thymus capitatus and Thymus vulgaris EOs, which are common in Spain and widely used in the meat industry, could be used as antibacterial agents in food preservation. The aims of this study were to determine (i) the antibacterial activity using, as culture medium, extracts from meat homogenates (minced beef, cooked ham or dry-cured sausage); and (ii) the antioxidant properties of organic EOs obtained from T. zygis, T. mastichina, T. capitatus and T. vulgaris. The antioxidant activity was determined using different methodologies, such as Ferrous ion-chelating ability assay, Ferric reducing antioxidant power, ABTS radical cation (ABTS•+) scavenging activity assay and 2,2′-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging method; while the antibacterial activity was determined against 10 bacteria using the agar diffusion method in different meat model media. All EOs analyzed, at all concentrations, showed antioxidant activity. T. capitatus and T. zygis EOs were the most active. The IC50 values, for DPPH, ABTS and FIC assays were 0.60, 1.41 and 4.44 mg/mL, respectively, for T. capitatus whilst for T. zygis were 0.90, 2.07 and 4.95 mg/mL, respectively. Regarding antibacterial activity, T. zygis and T. capitatus EOs, in all culture media, had the highest inhibition halos against all tested bacteria. In general terms, the antibacterial activity of all EOs assayed was higher in the medium made with minced beef than with the medium elaborated with cooked ham or dry-cured sausage. PMID:28788051

Assessment of Antioxidant and Antibacterial Properties on Meat Homogenates of Essential Oils Obtained from Four Thymus Species Achieved from Organic Growth.

PubMed

Ballester-Costa, Carmen; Sendra, Esther; Fernández-López, Juana; Pérez-Álvarez, Jose A; Viuda-Martos, Manuel

2017-07-28

In the organic food industry, no chemical additives can be used to prevent microbial spoilage. As a consequence, the essential oils (EOs) obtained from organic aromatic herbs and spices are gaining interest for their potential as preservatives. The organic Thymus zygis , Thymus mastichina , Thymus capitatus and Thymus vulgaris EOs, which are common in Spain and widely used in the meat industry, could be used as antibacterial agents in food preservation. The aims of this study were to determine (i) the antibacterial activity using, as culture medium, extracts from meat homogenates (minced beef, cooked ham or dry-cured sausage); and (ii) the antioxidant properties of organic EOs obtained from T. zygis , T. mastichina , T. capitatus and T. vulgaris . The antioxidant activity was determined using different methodologies, such as Ferrous ion-chelating ability assay, Ferric reducing antioxidant power, ABTS radical cation (ABTS • +) scavenging activity assay and 2,2'-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging method; while the antibacterial activity was determined against 10 bacteria using the agar diffusion method in different meat model media. All EOs analyzed, at all concentrations, showed antioxidant activity. T. capitatus and T. zygis EOs were the most active. The IC 50 values, for DPPH, ABTS and FIC assays were 0.60, 1.41 and 4.44 mg/mL, respectively, for T. capitatus whilst for T. zygis were 0.90, 2.07 and 4.95 mg/mL, respectively. Regarding antibacterial activity, T. zygis and T. capitatus EOs, in all culture media, had the highest inhibition halos against all tested bacteria. In general terms, the antibacterial activity of all EOs assayed was higher in the medium made with minced beef than with the medium elaborated with cooked ham or dry-cured sausage.

Mechanism of oil-pulling therapy - in vitro study.

PubMed

Asokan, Sharath; Rathinasamy, T K; Inbamani, N; Menon, Thangam; Kumar, S Senthil; Emmadi, Pamela; Raghuraman, R

2011-01-01

Oil pulling has been used extensively as a traditional Indian folk remedy without scientific proof for many years for strengthening teeth, gums and jaws and to prevent decay, oral malodor, bleeding gums and dryness of throat and cracked lips. The aim of this study was to evaluate the antibacterial activity of sesame oil and lignans isolated from sesame oil on oral microorganisms and to check whether saponification or emulsification occurs during oil-pulling therapy. The in vitro study was carried out in three different phases: (1) Antibacterial activity of the lignans and sesame oil were tested by minimum inhibitory concentration assay by agar dilution method and agar well diffusion method, respectively. (2) Increase in free fatty acid level of oil and the quantity of sodium hydroxide (NaOH) used up in the titration are good indicators of saponification process. This was assessed using analytical tests for vegetable oils. (3) Swished oil was observed under light microscope to assess the status of the oil, presence of microorganisms, oral debris and foreign bodies. Sesamin and sesamolin isolated from sesame oil did not have any antibacterial effect against oral microorganisms like Streptococcus mutans, Streptococcus mitis and Streptococcus viridans. Emulsification of sesame oil occurs during oil-pulling therapy. Increased consumption of NaOH in titration is a definite indication of a possible saponification process. The myth that the effect of oil-pulling therapy on oral health was just a placebo effect has been broken and there are clear indications of possible saponification and emulsification process, which enhances its mechanical cleaning action.

Antagonistic and Quantitative Assessment of Indigenous Lactic acid Bacteria in Different Varieties of Ogi against Gastrointestinal Pathogens

PubMed Central

Afolayan, Ayorinde Oluwatobiloba; Ayeni, Funmilola Abidemi; Ruppitsch, Werner

2017-01-01

Introduction Ogi is a popular fermented cereal gruel consumed mainly in the western part of Nigeria. Traditionally, uncooked Ogi is normally administered to diarrhoea patients to reduce the frequency of stooling. This study was therefore undertaken to identify, quantify and determine the antimicrobial properties of lactic acid bacteria (LAB) isolated from Ogi. Methods The Ogi samples (Yellow, white, sorghum) were obtained from different market in Ibadan, Nigeria and Ogi control (cooked, uncooked and Omidun) were prepared with the viable counts of bacteria monitored over 5 days period. LAB were isolated from the varieties and identified by partial sequencing of 16S rRNA gene. The antimicrobial activities of the cell free supernatant (CFS) and the viable cells of the isolated LAB against Escherichia coli EC004, Salmonella sp. SS11, Shigella sp. SS10 were investigated by agar diffusion assay, agar overlay method, and coculture growth studies. Results Omidun had the highest LAB count while cooked ogi has the lowest LAB count. Weissella paramesenteroides , L. brevis, L. rossiae, L. fermentum, L. plantarum, Acetobacter pasteurianus, Paenibacillus sp. and Bacillus sp. were isolated from Ogi in this study. Large zone of inhibition (11≤x≤20) was observed with CFS against Salmonella sp. SS11 and Shigella sp. SS10 and also the overlay method. Coculture studies of Weissella paramesenteroides, Lactobacillus fermentum, and L. plantarum with Salmonella sp. SS11 showed a 5-8 log reduction of the pathogens' growth after 24 hours as compared with the control. Conclusion Ogi and its contents have antimicrobial properties against pathogenic organisms. PMID:28748023

Methicillin-Resistant Staphylococcus aureus: Comparison of Susceptibility Testing Methods and Analysis of mecA-Positive Susceptible Strains

PubMed Central

Sakoulas, George; Gold, Howard S.; Venkataraman, Lata; DeGirolami, Paola C.; Eliopoulos, George M.; Qian, Qinfang

2001-01-01

Methicillin-resistant Staphylococcus aureus (MRSA) is responsible for an increasing number of serious nosocomial and community-acquired infections. Phenotypic heterogeneous drug resistance (heteroresistance) to antistaphylococcal beta-lactams affects the results of susceptibility testing. The present study compared the MRSA-Screen latex agglutination test (Denka Seiken Co., Ltd., Tokyo, Japan) for detection of PBP 2a with agar dilution, the VITEK-1 and VITEK-2 systems (bioMérieux, St. Louis, Mo.), and the oxacillin agar screen test for detection of MRSA, with PCR for the mecA gene used as the “gold standard” assay. Analysis of 107 methicillin-susceptible S. aureus (MSSA) isolates and 203 MRSA isolates revealed that the MRSA-Screen latex agglutination test is superior to any single phenotype-based susceptibility testing method, with a sensitivity of 100% and a specificity of 99.1%. Only one isolate that lacked mecA was weakly positive by the MRSA-Screen latex agglutination test. This isolate was phenotypically susceptible to oxacillin and did not contain the mecA gene by Southern blot hybridization. The oxacillin agar screen test, the VITEK-1 system, the VITEK-2 system, and agar dilution showed sensitivities of 99.0, 99.0, 99.5, and 99%, respectively, and specificities of 98.1, 100, 97.2, and 100%, respectively. The differences in sensitivity or specificity were not statistically significant. Oxacillin bactericidal assays showed that mecA- and PBP 2a-positive S. aureus isolates that are susceptible to antistaphylococcal beta-lactams by conventional methods are functionally resistant to oxacillin. We conclude that the accuracy of the MRSA-Screen latex agglutination method for detection of PBP 2a approaches the accuracy of PCR and is more accurate than any susceptibility testing method used alone for the detection of MRSA. PMID:11682512

An evaluation of false-positive rifampicin resistance on the Xpert MTB/RIF.

PubMed

Cayci, Yeliz Tanriverdi; Bilgin, Kemal; Coban, Ahmet Yilmaz; Birinci, Asuman; Durupınar, Belma

2017-11-01

Mycobacterium tuberculosis (MTB) is one of the most significant causes of mortality and morbidity. Early diagnose is important especially in multiple drug resistant tuberculosis to avoid transmission. Traditional techniques requires at least one to three weeks for diagnosis of tuberculosis. Diagnostic delays with multiple drug resistant tuberculosis are associated with worse clinical outcomes and increased transmission The Xpert MTB/RIF assay is one of the new diagnostic device for the diagnosis of tuberculosis and rapid detection of rifampicin resistance. We assessed the performance of Xpert MTB/RIF assay for detecting rifampicin resistance using phenotypic drug susceptibility tests as automated BD MGIT 960. Total of 2136 specimens were included in the study. Xpert MTB/RIF testing was performed on samples, using version 4 cartridges, according to the manufacturer's recommendations. The MTBC culture and first-line phenotypic DST were performed in automated BD MGIT 960 (Becton & Dickinson, USA) according to the recommendations of the manufacturer. Agar proportion was used in the case of inconsistency for rifampicin resistance. Thirty-four samples (19 respiratory and 15 nonrespiratory samples) were determined as positive for M. tuberculosis complex by Xpert MTB/RIF (Cepheid GeneXpert® System, USA). Xpert MTB/RIF assay detected 4/34 (11.7%) specimens as rifampicin resistant. One of the rifampicin resistant isolates was determined susceptible in MGIT 960 automated system. This isolate was also tested with agar proportion method and found susceptible to rifampicin. The Xpert MTB/RIF assay can be used as first-line assay for the detection of M. tuberculosis. However, microbiologists must be aware of the limitations of the assay.

Effect of the fungus Pochonia chlamydosporia on Echinostoma paraensei (Trematoda: Echinostomatidae).

PubMed

Lelis, Rosane Teixeira; Braga, Fabio Ribeiro; de Carvalho, Lorendane Millena; de Paula, Alessandra Teixeira; Araujo, Juliana Milani; Fausto, Mariana Costa; Junior, Arnaldo Maldonado; Rodrigues, João Victor Facchini; de Freitas Soares, Filippe Elias; Garcia, Juberlan Silva; de Araújo, Jackson Victor

2014-11-01

Echinostoma paraensei is a trematode of the genus Echinostoma that causes echinostomiasis in humans. The objectives of this study were to: evaluate the ovicidal activity of the nematophagous fungus Pochonia chlamydosporia (VC1 and VC4) on a solid medium 2% water-agar (2% WA) against E. paraensei eggs (assay A); evaluate ovicidal effect (destruction of eggs) of the isolate VC4 in supplemented culture media (assay B); and evaluate the ovicidal ability of the crude extract (VC4) on E. paraensei eggs (assay C). Eggs of E. paraensei (assay A) were placed in Petri dishes containing 2% WA with an isolate of the fungus P. chlamydosporia (VC1 and VC4) grown for 10 days, and without fungus as a control and evaluated regarding their destruction. In assay B, eggs of E. paraensei were placed in Petri dishes with different supplemented culture media and with VC4 isolate and the destruction of eggs was examined at the end of 25 days of interaction. In assay C, effects of the crude extract of P. chlamydosporia (VC4) on eggs were evaluated at the end of 7 days. In assay A, there was no difference (p>0.05) in ovicidal activity among the tested isolates (VC1 and VC4); however, the highest percentage for ovicidal activity (type 3 effect) was demonstrated by the isolate VC4. In assay B, the culture medium starch-agar showed the best results for the destruction of the eggs, with a percentage of 46.6% at the end of the assay. In assay C, the crude extract of VC4 was effective in the destruction of E. paraensei eggs, with a percentage reduction of 53%. The results of this study demonstrate that a rich culture medium with a greater availability of carbon and nitrogen may interfere directly in the predatory characteristics of ovicidal fungi. Copyright © 2014 Elsevier B.V. All rights reserved.

Antimicrobial olive leaf gelatin films for enhancing the quality of cold smoked salmon

USDA-ARS?s Scientific Manuscript database

Olive leaf products were evaluated as antimicrobial/antioxidant ingredients in edible films for smoked fish preservation. Olive leaf powder (OLP) and its water/ethanol extract (OLE) were tested against Listeria monocytogenes, Escherichia coli O157:H7 and Salmonella enterica using agar diffusion test...

In vitro sensitivity of Hungarian Actinobaculum suis strains to selected antimicrobials.

PubMed

Biksi, I; Major, Andrea; Fodor, L; Szenci, O; Vetési, F

2003-01-01

In vitro antimicrobial sensitivity of 12 Hungarian isolates and the type strain ATCC 33144 of Actinobaculum suis to different antimicrobial compounds was determined both by the agar dilution and by the disc diffusion method. By agar dilution, MIC50 values in the range of 0.05-3.125 micrograms/ml were determined for penicillin, ampicillin, ceftiofur, doxycycline, tylosin, pleuromutilins, chloramphenicol, florfenicol, enrofloxacin and lincomycin. The MIC50 value of oxytetracycline and spectinomycin was 6.25 and 12.5 micrograms/ml, respectively. For ofloxacin, flumequine, neomycin, streptomycin, gentamicin, nalidixic acid, nitrofurantoin and sulphamethoxazole + trimethoprim MIC50 values were in the range of 25-100 micrograms/ml. With the disc diffusion method, all strains were sensitive to penicillin, cephalosporins examined, chloramphenicol and florfenicol, tetracyclines examined, pleuromutilins, lincomycin and tylosin. Variable sensitivity was observed for fluoroquinolones (flumequine, enrofloxacin, ofloxacin), most of the strains were susceptible to marbofloxacin. Almost all strains were resistant to aminoglycosides but most of them were sensitive to spectinomycin. A strong correlation was determined for disc diffusion and MIC results (Spearman's rho 0.789, p < 0001). MIC values of the type strain and MIC50 values of other tested strains did not differ significantly. Few strains showed a partially distinct resistance pattern for erythromycin, lincomycin and ampicillin in both methods.

Antifungal Activity of Wickerhamomyces anomalus and Lactobacillus plantarum during Sourdough Fermentation: Identification of Novel Compounds and Long-Term Effect during Storage of Wheat Bread ▿

PubMed Central

Coda, Rossana; Cassone, Angela; Rizzello, Carlo G.; Nionelli, Luana; Cardinali, Gianluigi; Gobbetti, Marco

2011-01-01

This study aimed at investigating the antifungal activity of Wickerhamomyces anomalus and sourdough lactic acid bacteria to extend the shelf life of wheat flour bread. The antifungal activity was assayed by agar diffusion, growth rate inhibition, and conidial germination assays, using Penicillium roqueforti DPPMAF1 as the indicator fungus. Sourdough fermented by Lactobacillus plantarum 1A7 (S1A7) and dough fermented by W. anomalus LCF1695 (D1695) were selected and characterized. The water/salt-soluble extract of S1A7 was partially purified, and several novel antifungal peptides, encrypted into sequences of Oryza sativa proteins, were identified. The water/salt-soluble extract of D1695 contained ethanol and, especially, ethyl acetate as inhibitory compounds. As shown by growth inhibition assays, both water/salt-soluble extracts had a large inhibitory spectrum, with some differences, toward the most common fungi isolated from bakeries. Bread making at a pilot plant was carried out with S1A7, D1695, or a sourdough started with a combination of both strains (S1A7-1695). Slices of the bread manufactured with S1A7-1695 did not show contamination by fungi until 28 days of storage in polyethylene bags at room temperature, a level of protection comparable to that afforded by 0.3% (wt/wt) calcium propionate. The effect of sourdough fermentation with W. anomalus LCF1695 was also assessed based on rheology and sensory properties. PMID:21441340

Differential quantification of SIgA and SC by two-directional rocket method.

PubMed Central

Kosaka, T; Asahina, T; Kobayashi, N

1980-01-01

The two-directional rocket method, a newly modified method for quantitative immunoelectrophoresis, was used as the assay for separating SC and SIgA, which have identical antigenicity but differ in mobility. This method proved to be sufficiently simple and sensitive to enable simultaneous assay of SC and SIgA in saliva. The method employs electrophoresis into antibody-containing agarose/agar gel in the presence of heparin-Ca EDTA. The height of the precipitation peaks formed in two directions is proportional to the concentration of the antigens. Concomitant use of agarose which has little electroendosmosis and agar which has high electroendosmosis facilitated cathodic migration of SIgA. Transfer of SC from beta-region to alpha 1-region without influencing the mobilities of SIgA, albumin or IgG was obtained by addition of heparin-Ca EDTA to agarose/agar gel. This effect of heparin-Ca EDTA is vulnerable to changes of pH of the gel, but is almost completely independent of change in composition or concentration of the gel. The function of heparin as a polyanion may be resonsible for it. Carbamylation of antibody was used to accelerate a clear-cut resolution of the cathodic rockets. This technique was found to be a method of choice for analysis of SIgA and SC in large numbers. In using this method as a screening assay for detection of primary immunodeficiency, by studying saliva samples collected from 3 month old infants on the occasion of regular check-up over a 2 year period, two cases of isolated IgA immunodeficiency and two cases of hypoglobulinaemia were discovered in 12,000 infants. Images Figure 1 Figure 7 PMID:6776036

An in vitro study of the antimicrobial effects of indigo naturalis prepared from Strobilanthes formosanus Moore.

PubMed

Chiang, Yin-Ru; Li, Ann; Leu, Yann-Lii; Fang, Jia-You; Lin, Yin-Ku

2013-11-21

Indigo naturalis is effective in treating nail psoriasis coexisting with microorganism infections. This study examines the antimicrobial effects of indigo naturalis prepared from Strobilanthes formosanus Moore. Eight bacterial and seven fungal strains were assayed using the agar diffusion method to examine the effects of indigo naturalis and its bioactive compounds. The bioactive compounds of indigo naturalis were purified sequentially using GFC, TLC, and HPLC. Their structures were identified using mass spectrometry and NMR spectroscopy. UPLC-MS/MS was applied to compare the metabolome profiles of indigo naturalis ethyl-acetate (EA) extract and its source plant, Strobilanthes formosanus Moore. The results of in vitro antimicrobial assays showed that indigo naturalis EA-extract significantly (≥1 mg/disc) inhibits Gram-positive bacteria (Staphylococcus aureus, S. epidermis and methicillin-resistant S. aureus (MRSA)) and mildly inhibits non-dermatophytic onychomycosis pathogens (Aspergillus fumigates and Candida albicans), but has little effect on dermatophyes. Isatin and tryptanthrin were identified as the bioactive compounds of indigo naturalis using S. aureus and S. epidermis as the bioassay model. Both bioactive ingredients had no effect on all tested fungi. In summary, indigo naturalis prepared from Strobilanthes formosanus Moore exhibits antimicrobial effects on Staphylococcus and non-dermatophytic onychomycosis pathogens. Tryptanthrin and isatin may be its major bioactive ingredients against Staphylococcus and the inhibitory effect on MRSA may be due to other unidentified ingredients.

Chemical composition and biological evaluation of the volatile constituents from the aerial parts of Nephrolepis exaltata (L.) and Nephrolepis cordifolia (L.) C. Presl grown in Egypt.

PubMed

El-Tantawy, Mona E; Shams, Manal M; Afifi, Manal S

2016-01-01

The essential oil from the aerial parts of Nephrolepis exaltata and Nephrolepis cordifolia obtained by hydro-distillation were analyzed by gas chromatography/ mass spectrometry. The essential oils exhibited potential antibacterial and antifungal activities against a majority of the selected microorganisms. NEA oil showed promising cytotoxicity in breast, colon and lung carcinoma cells. The results presented indicate that NEA oil could be useful alternative for the treatment of dermatophytosis. Comparative investigation of hydro-distilled volatile constituents from aerial parts (A) of Nephrolepis exaltata (NE) and Nephrolepis cordifolia (NC) (Family Nephrolepidaceae) was carried out. Gas chromatography/mass spectrometry revealed that oils differ in composition and percentages of components. Oxygenated compounds were dominant in NEA and NCA. 2,4-Hexadien-1-ol (16.1%), nonanal (14.4%), β-Ionone (6.7%) and thymol (2.7%) were predominant in NEA. β-Ionone (8.0%), eugenol (7.2%) and anethol (4.6%) were the main constituents in NCA. Volatile samples were screened for their antibacterial and antifungal activities using agar diffusion method and minimum inhibitory concentrations. The cytotoxic activity was evaluated using viability assay in breast (MCF-7), colon (HCT-116) and lung carcinoma (A-549) cells by the MTT assay. The results revealed that NEA oil exhibited potential antimicrobial activity against most of the tested organisms and showed promising cytotoxicity.

Pharmacological Assessment of the Medicinal Potential of Acacia mearnsii De Wild.: Antimicrobial and Toxicity Activities

PubMed Central

Olajuyigbe, Olufunmiso O.; Afolayan, Anthony J.

2012-01-01

Acacia mearnsii De Wild. (Fabaceae) is a medicinal plant used in the treatment of microbial infections in South Africa without scientific validation of its bioactivity and toxicity. The antimicrobial activity of the crude acetone extract was evaluated by both agar diffusion and macrobroth dilution methods while its cytotoxicity effect was assessed with brine shrimp lethality assay. The study showed that both bacterial and fungal isolates were highly inhibited by the crude extract. The MIC values for the gram-positive bacteria (78.1–312.5) μg/mL, gram-negative bacteria (39.1–625) μg/mL and fungal isolates (625–5000) μg/mL differ significantly. The bacteria were more susceptible than the fungal strains tested. The antibiosis determination showed that the extract was more (75%) bactericidal than bacteriostatic (25%) and more fungicidal (66.67%) than fungistatic (33.33%). The cytotoxic activity of the extract was observed between 31.25 μg/mL and 500 μg/mL and the LC50 value (112.36 μg/mL) indicates that the extract was nontoxic in the brine shrimp lethality assay (LC50 > 100 μg/mL). These results support the use of A. mearnsii in traditional medicine for treatment of microbial infections. The extract exhibiting significant broad spectrum antimicrobial activity and nontoxic effects has potential to yield active antimicrobial compounds. PMID:22605976

Microbioassay of Antimicrobial Agents

PubMed Central

Simon, Harold J.; Yin, E. Jong

1970-01-01

A previously described agar-diffusion technique for microbioassay of antimicrobial agents has been modified to increase sensitivity of the technique and to extend the range of antimicrobial agents to which it is applicable. This microtechnique requires only 0.02 ml of an unknown test sample for assay, and is capable of measuring minute concentrations of antibiotics in buffer, serum, and urine. In some cases, up to a 20-fold increase in sensitivity is gained relative to other published standardized methods and the error of this method is less than ±5%. Buffer standard curves have been established for this technique, concurrently with serum standard curves, yielding information on antimicrobial serum-binding and demonstrating linearity of the data points compared to the estimated regression line for the microconcentration ranges covered by this technique. This microassay technique is particularly well suited for pediatric research and for other investigations where sample volumes are small and quantitative accuracy is desired. Dilution of clinical samples to attain concentrations falling with the range of this assay makes the technique readily adaptable and suitable for general clinical pharmacological studies. The microassay technique has been standardized in buffer solutions and in normal human serum pools for the following antimicrobials: ampicillin, methicillin, penicillin G, oxacillin, cloxacillin, dicloxacillin, cephaloglycin, cephalexin, cephaloridine, cephalothin, erythromycin, rifamycin amino methyl piperazine, kanamycin, neomycin, streptomycin, colistin, polymyxin B, doxycycline, minocycline, oxytetracycline, tetracycline, and chloramphenicol. PMID:4986725

Antimicrobial efficacy of 3 oral antiseptics containing octenidine, polyhexamethylene biguanide, or Citroxx: can chlorhexidine be replaced?

PubMed

Rohrer, Nadine; Widmer, Andreas F; Waltimo, Tuomas; Kulik, Eva M; Weiger, Roland; Filipuzzi-Jenny, Elisabeth; Walter, Clemens

2010-07-01

Use of oral antiseptics decreases the bacterial load in the oral cavity. To compare the antimicrobial activity of 3 novel oral antiseptics with that of chlorhexidine, which is considered the "gold standard" of oral hygiene. Comparative in vitro study. Four common oral microorganisms (Streptococcus sanguinis, Streptococcus mutans, Candida albicans, and Fusobacterium nucleatum) were tested under standard conditions and at different concentrations, by use of a broth dilution assay and an agar diffusion assay and by calculating the log10 reduction factor (RF). The antimicrobial activity of each antiseptic was assessed by counting the difference in bacterial densities (ie, the log10 number of colony-forming units of bacteria) before and after the disinfection process. The oral antiseptics containing octenidine (with an RF in the range of 7.1-8.24 CFU/mL) and polyhexamethylene biguanide (with an RF in the range of 7.1-8.24 CFU/mL) demonstrated antimicrobial activity comparable to that of chlorhexidine (with an RF in the range of 1.03-8.24 CFU/mL), whereas the mouth rinse containing Citroxx (Citroxx Biosciences; with an RF in the range of 0.22-1.36 CFU/mL) showed significantly weaker antimicrobial efficacy. Overall, octenidine and polyhexamethylene biguanide were more active at lower concentrations.conclusion. Oral antiseptics containing the antimicrobial agent octenidine or polyhexamethylene biguanide may be considered as potent alternatives to chlorhexidine-based preparations.

Chemical Composition, Antioxidant and Antimicrobial Activities of Thymus capitata Essential Oil with Its Preservative Effect against Listeria monocytogenes Inoculated in Minced Beef Meat

PubMed Central

El Abed, Nariman; Kaabi, Belhassen; Smaali, Mohamed Issam; Chabbouh, Meriem; Habibi, Kamel; Mejri, Mondher; Marzouki, Mohamed Nejib; Ben Hadj Ahmed, Sami

2014-01-01

The chemical composition, antioxidant and antimicrobial activities, and the preservative effect of Thymus capitata essential oil against Listeria monocytogenes inoculated in minced beef meat were evaluated. The essential oil extracted was chemically analyzed by gas chromatography-mass spectrometry. Nineteen components were identified, of which carvacrol represented (88.89%) of the oil. The antioxidant activity was assessed in vitro by using both the DPPH and the ABTS assays. The findings showed that the essential oil exhibited high antioxidant activity, which was comparable to the reference standards (BHT and ascorbic acid) with IC50 values of 44.16 and 0.463 μg/mL determined by the free-radical scavenging DPPH and ABTS assays, respectively. Furthermore, the essential oil was evaluated for its antimicrobial activity using disc agar diffusion and microdilution methods. The results demonstrated that the zone of inhibition varied from moderate to strong (15–80 mm) and the minimum inhibition concentration values ranged from 0.32 to 20 mg/mL. In addition, essential oil evaluated in vivo against Listeria monocytogenes showed clear and strong inhibitory effect. The application of 0.25 or 1% (v/w) essential oil of T. capitata to minced beef significantly reduced the L. monocytogenes population when compared to those of control samples (P-value  <0.01). PMID:24719640

Development of a Flow Cytometry-Based Method for Rapid Detection of Escherichia coli and Shigella Spp. Using an Oligonucleotide Probe

PubMed Central

Xue, Yong; Wilkes, Jon G.; Moskal, Ted J.; Williams, Anna J.; Cooper, Willie M.; Nayak, Rajesh; Rafii, Fatemeh; Buzatu, Dan A.

2016-01-01

Standard methods to detect Escherichia coli contamination in food use the polymerase chain reaction (PCR) and agar culture plates. These methods require multiple incubation steps and take a long time to results. An improved rapid flow-cytometry based detection method was developed, using a fluorescence-labeled oligonucleotide probe specifically binding a16S rRNA sequence. The method positively detected 51 E. coli isolates as well as 4 Shigella species. All 27 non-E. coli strains tested gave negative results. Comparison of the new genetic assay with a total plate count (TPC) assay and agar plate counting indicated similar sensitivity, agreement between cytometry cell and colony counts. This method can detect a small number of E.coli cells in the presence of large numbers of other bacteria. This method can be used for rapid, economical, and stable detection of E. coli and Shigella contamination in the food industry and other contexts. PMID:26913737

Development of a Flow Cytometry-Based Method for Rapid Detection of Escherichia coli and Shigella Spp. Using an Oligonucleotide Probe.

PubMed

Xue, Yong; Wilkes, Jon G; Moskal, Ted J; Williams, Anna J; Cooper, Willie M; Nayak, Rajesh; Rafii, Fatemeh; Buzatu, Dan A

2016-01-01

Standard methods to detect Escherichia coli contamination in food use the polymerase chain reaction (PCR) and agar culture plates. These methods require multiple incubation steps and take a long time to results. An improved rapid flow-cytometry based detection method was developed, using a fluorescence-labeled oligonucleotide probe specifically binding a16S rRNA sequence. The method positively detected 51 E. coli isolates as well as 4 Shigella species. All 27 non-E. coli strains tested gave negative results. Comparison of the new genetic assay with a total plate count (TPC) assay and agar plate counting indicated similar sensitivity, agreement between cytometry cell and colony counts. This method can detect a small number of E.coli cells in the presence of large numbers of other bacteria. This method can be used for rapid, economical, and stable detection of E. coli and Shigella contamination in the food industry and other contexts.

Potential use of gallium-doped phosphate-based glass material for periodontitis treatment.

PubMed

Sahdev, Rohan; Ansari, Tahera I; Higham, Susan M; Valappil, Sabeel P

2015-07-01

This study aimed at evaluating the potential effect of gallium-incorporated phosphate-based glasses towards periodontitis-associated bacteria, Porphyromonas gingivalis, and matrix metalloproteinase-13. Periodontitis describes a group of inflammatory diseases of the gingiva and supporting structures of the periodontium. They are initiated by the accumulation of plaque bacteria, such as the putative periodontal pathogen Porphyromonas gingivalis, but the host immune response such as elevated matrix metalloproteinases are the major contributing factor for destruction of periodontal tissues. Antibacterial assays of gallium-incorporated phosphate-based glasses were conducted on Porphyromonas gingivalis ATCC 33277 using disc diffusion assay on fastidious anaerobe agar and liquid broth assay in a modified tryptic soy broth. In vitro study investigated the effect of gallium on purified recombinant human matrix metalloproteinase-13 activity using matrix metalloproteinase assay kit. In vivo biocompatibility of gallium-incorporated phosphate-based glass was evaluated in rats as subcutaneous implants. Antibacterial assay of gallium displayed activity against Porphyromonas gingivalis (inhibition zone of 22 ± 0.5 mm compared with 0 mm for control glass, c-PBG). Gallium in the glass contributed to growth inhibitory effect on Porphyromonas gingivalis (up to 1.30 reductions in log 10 values of the viable counts compared with control) in a modified tryptic soy broth. In vitro study showed gallium-incorporated phosphate-based glasses inhibited matrix metalloproteinase activity significantly (p ≤ 0.01) compared with c-PBG. Evaluation of in vivo biocompatibility of gallium-incorporated phosphate-based glasses in rats showed a non-toxic and foreign body response after 2 weeks of implantation. The results indicate that gallium ions might act on multiple targets of biological mechanisms underlying periodontal disease. Moreover, gallium-incorporated phosphate-based glasses are biocompatible in a rat model. The findings warrant further investigation and will have important clinical implications in the future treatment and management of periodontitis. © The Author(s) 2015 Reprints and permissions: sagepub.co.uk/journalsPermissions.nav.

Antimicrobial activities of three species of family mimosaceae.

PubMed

Mahmood, Adeel; Mahmood, Aqeel; Qureshi, Rizwana Aleem

2012-01-01

The antimicrobial activities of crude methanolic extract of leaves of Acacia nilotica L., Albizia lebbeck L. and Mimosa himalayana Gamble belonging to family mimosaceae were investigated in this research work. Antibacterial activity was studied by agar well diffusion method against one gram-positive Bacillus subtilis and three gram-negative Pseudomonas aeruginosa, Escherichia coli and Klebsiella pneumonia. Crude extract of all plants showed best activity against gram-negative bacterial strains while minor inhibition zones were found against gram positive bacterial strains. Antifungal activity of crude plant extract was screened by agar tube dilution method against Aspergillus nigar and Aspergillus flavus. These results showed that these plants extracts have potential against bacterias, while against fungi their activity is not much effective.

[Lipase and phospholipase C from Staphylococcus aureus of different origin. I. Determination and occurrence (author's transl)].

PubMed

Berete, Y J; Schaeg, W; Brückler, J; Blobel, H

1980-11-01

Lipase and phospholipase C from Staphylococcus aureus of different origin were demonstrated qualitatively by agar diffusion on tributyrin- and lecithin agar. On test media with either 0,3% Na-azide or 0,3% KCN lipase-activity was not inhibited, phospholipase C, on the other hand, completely blocked (Table 1, Fig. 2). In this manner a tentative differentiation was possible between lipase and phospholipase C. For the quantitative determination of lipase the hydrolysis of p-nitrophenyl palmitate proved to be most useful (Fig. 1). S. aureus-cultures of human origin produced more often and more actively lipase and phospholipase C than those from cattle (Table 2).

Antimicrobial Activity and Probable Mechanisms of Action of Medicinal Plants of Kenya: Withania somnifera, Warbugia ugandensis, Prunus africana and Plectrunthus barbatus

PubMed Central

Mwitari, Peter G.; Ayeka, Peter A.; Ondicho, Joyce; Matu, Esther N.; Bii, Christine C.

2013-01-01

Withania somnifera, Warbugia ugandensis, Prunus africana and Plectrunthus barbatus are used traditionally in Kenya for treatment of microbial infections and cancer. Information on their use is available, but scientific data on their bioactivity, safety and mechanisms of action is still scanty. A study was conducted on the effect of organic extracts of these plants on both bacterial and fungal strains, and their mechanisms of action. Extracts were evaluated through the disc diffusion assay. Bacteria and yeast test strains were cultured on Mueller-Hinton agar and on Sabouraud dextrose agar for the filamentous fungi. A 0.5 McFarland standard suspension was prepared. Sterile paper discs 6 mm in diameter impregnated with 10 µl of the test extract (100 mg/ml) were aseptically placed onto the surface of the inoculated media. Chloramphenicol (30 µg) and fluconazole (25 µg) were used as standards. Discs impregnated with dissolution medium were used as controls. Activity of the extracts was expressed according to zone of inhibition diameter. MIC was determined at 0.78–100 mg/ml. Safety studies were carried using Cell Counting Kit 8 cell proliferation assay protocol. To evaluate extracts mechanisms of action, IEC-6 cells and RT-PCR technique was employed in vitro to evaluate Interleukin 7 cytokine. Investigated plants extracts have both bactericidal and fungicidal activity. W. ugandensis is cytotoxic at IC50<50 µg/ml with MIC values of less than 0.78 mg/ml. Prunus africana shuts down expression of IL 7 mRNA at 50 µg/ml. W. somnifera has the best antimicrobial (1.5625 mg/ml), immunopotentiation (2 times IL 7 mRNA expression) and safety level (IC50>200 µg/ml). Fractions from W. ugandensis and W. somnifera too demonstrated antimicrobial activity. Mechanisms of action can largely be attributed to cytotoxicity, Gene silencing and immunopotentiation. Use of medicinal plants in traditional medicine has been justified and possible mechanisms of action demonstrated. Studies to isolate and characterize the bioactive constituents continue. PMID:23785437

Antimicrobial resistance in Campylobacter spp isolated from broiler flocks

PubMed Central

Kuana, Suzete Lora; dos Santos, Luciana Ruschel; Rodrigues, Laura Beatriz; Borsoi, Anderlise; Moraes, Hamilton Luis do Souza; Salle, Carlos Tadeu Pippi; do Nascimento, Vladimir Pinheiro

2008-01-01

The aim of this study was to assess the antimicrobial susceptibility of 62 Campylobacter spp. strains obtained from broiler flocks using the agar diffusion method. The Campylobacter spp strains were isolated from 22 flocks aged between 3 and 5 weeks of life, isolated from cloacae swabs, stools and cecal droppings in the farm and from the carcass rinsing in the slaughterhouse. Campylobacter spp strains were tested on Mueller-Hilton (MH) agar (27 samples) and MH plus TTC agar (35 samples). The antimicrobial susceptibility test revealed a 62.5% resistance to at least one drug, especially to enrofloxacin (71%), neomycin (50%), lincomycin (50%), tetracycline (43%), penicillin (42%), ceftiofur (33%) amoxicillin (27%), spiramycin (20%), ampicillin (18%) and norfloxacin (14%), whereas a lower percentage of strains was resistant to erythromycin (10%) and doxycycline (10%). All strains were sensitive to gentamicin and lincomycin-spectinomycin and 80% of them to colistin. These results indicate that it is necessary to reduce the use of antimicrobials in veterinary and human medicine. PMID:24031299

Light chain typing of immunoglobulins in small samples of biological material

PubMed Central

Rádl, J.

1970-01-01

A method is described for the typing of the light chains of immunoglobulins in small samples of sera or external secretions and without their previous isolation. It consists of immunoelectrophoresis in agar plates which contain specific antisera against one of the light chain types. All immunoglobulins of this type are thus selected by precipitation in the central area during the electrophoretic phase. Immunoglobulins of the opposite light chain type diffuse through the agar and react with the class specific antisera from the troughs. This results in the precipitin lines as in conventional immunoelectrophoresis. This technique has proved most useful for typing heterogenous or homogeneous immunoglobulins in normal and low concentration. The antisera used for incorporation in the agar should fulfil special requirements. They should contain a high level of antibodies against common surface determinants of the immunoglobulin light chains. The further possibilities of this immunoselection technique for typing different protein mixtures is discussed. ImagesFIG. 1FIG. 2FIG. 3FIG. 4FIG. 5FIG. 6 PMID:4098592

Growth, nisA Gene Expression, and In Situ Activity of Novel Lactococcus lactis subsp. cremoris Costarter Culture in Commercial Hard Cheese Production.

PubMed

Noutsopoulos, Dimitrios; Kakouri, Athanasia; Kartezini, Eleftheria; Pappas, Dimitrios; Hatziloukas, Efstathios; Samelis, John

2017-12-01

This study evaluated in situ expression of the nisA gene by an indigenous, nisin A-producing (NisA+) Lactococcus lactis subsp. cremoris raw milk genotype, represented by strain M78, in traditional Greek Graviera cheeses under real factory-scale manufacturing and ripening conditions. Cheeses were produced with added a mixed thermophilic and mesophilic commercial starter culture (CSC) or with the CSC plus strain M78 (CSC+M78). Cheeses were sampled after curd cooking (day 0), fermentation of the unsalted molds for 24 h (day 1), brining (day 7), and ripening of the brined molds (14 to 15 kg each) for 30 days in a fully controlled industrial room (16.5°C; 91% relative humidity; day 37). Total RNA was directly extracted from the cheese samples, and the expression of nisA gene was evaluated by real-time reverse transcription PCR (qRT-PCR). Agar overlay and well diffusion bioassays were correspondingly used for in situ detection of the M78 NisA+ colonies in the cheese agar plates and antilisterial activity in whole-cheese slurry samples, respectively. Agar overlay assays showed good growth (>8 log CFU/g of cheese) of the NisA+ strain M78 in coculture with the CSC and vice versa. The nisA expression was detected in CSC+M78 cheese samples only, with its expression levels being the highest (16-fold increase compared with those of the control gene) on day 1, followed by significant reduction on day 7 and almost negligible expression on day 37. Based on the results, certain intrinsic and mainly implicit hurdle factors appeared to reduce growth prevalence rates and decrease nisA gene expression, as well as the nisin A-mediated antilisterial activities of the NisA+ strain M78 postfermentation. To our knowledge, this is the first report on quantitative expression of the nisA gene in a Greek cooked hard cheese during commercial manufacturing and ripening conditions by using a novel, rarely isolated, indigenous NisA+ L. lactis subsp. cremoris genotype as costarter culture.

Evidence that bacteriophage λ lysogens may induce in response to the proton motive force uncoupler CCCP.

PubMed

Thomason, Lynn C; Court, Donald L

2016-02-01

We describe a genetic β-galactoside reporter system using a disk diffusion assay on MacConkey Lactose agar petri plates to monitor maintenance of the bacteriophage λ prophage state and viral induction in Escherichia coli K-12. Evidence is presented that the phage λ major lytic promoters, pL and pR, are activated when cells containing the reporters are exposed to the energy poison carbonyl cyanide m-chlorophenyl hydrazine, CCCP. This uncoupler of oxidative phosphorylation inhibits ATP synthesis by collapsing the proton motive force. Expression of the λ lytic promoters in response to CCCP requires host RecA function and an autocleavable CI repressor, as does SOS induction of the λ prophage that occurs by a DNA damage-dependent pathway. λ Cro function is required for CCCP-mediated activation of the λ lytic promoters. CCCP does not induce an sfi-lacZ SOS reporter. Published by Oxford University Press on behalf of FEMS 2015. This work is written by (a) US Government employee(s) and is in the public domain in the US.

Effect of glycerol and zinc oxide addition on antibacterial activity of biodegradable bioplastics from chitosan-kepok banana peel starch

NASA Astrophysics Data System (ADS)

Agustin, Y. E.; Padmawijaya, K. S.

2017-07-01

Bioplastic is a biopolymer plastic that can be degraded easily by microorganisms so it can be used as alternative replaced commercial plastic. This research aims to study the effects of additive (glycerol and zinc oxide) addition in the characteristic of antimicrobial activity and biodegradability bioplastic from chitosan and Kepok banana peel starch. In this research, bioplastics were synthesized by chitosan as the backbone and antimicrobial, Kepok banana peel starch as filler, glycerol as plasticizer, also ZnO as an amplifier. Bioplastics were characterized their antimicrobial activity using agar diffusion method (zone inhibition assay) and biodegradability test using microbe (EM4). The result showed the optimum composition of bioplastic is kitosan 4 - 30% starch - 5 mL glycerol - 5% ZnO gives the good antimicrobial activity towards gram positive and gram negative bacteria, and this bioplastic will be degraded within an hour and 12 min. Thus, this bioplastics may have potential to be use for food packaging by having biodegradable properties and also inhibit bacterial growth.

Influence of geosmin-producing Streptomyces on the growth and volatile metabolites of yeasts during chinese liquor fermentation.

PubMed

Du, Hai; Lu, Hu; Xu, Yan

2015-01-14

Diverse Streptomyces species act as geosmin producers in the Chinese liquor-making process, causing an earthy, off-odor containment. Through microbiological and metabolite analyses, this paper investigates the influence of several geosmin-producing Streptomyces on the microbial community of a brewing system. The antifungal activity against functional liquor-brewing microbes was assayed by an agar diffusion method. Several Streptomyces, most notably Streptomyces sampsonii QC-2, inhibited the growth of the brewing functional yeasts and molds in pure culture. In a simulated coculture, Streptomyces spp. reduced the flavor compounds (alcohols and esters) contributed by yeasts. Nine components in Streptomyces sampsonii QC-2 broth were detected by ultraperformance liquid chromatography coupled with photo diode array (UPLC–PDA), with characteristic ultraviolet absorptions at 360, 380, and 400 nm. The main products of Streptomyces sampsonii QC-2 were identified by ultraperformance liquid chromatography–quadrupole time-of-flight mass spectrometry (UPLC/Q-TOF–MS/MS), and confirmed by standard mass spectrometry. The antifungal active components were revealed as a series of heptaene macrolide antibiotics.

Evaluation of the Premi Test and comparison with the One-Plate Test for the detection of antimicrobials in kidney.

PubMed

Cantwell, H; O'Keeffe, M

2006-02-01

The Premi Test, a test kit designed for the rapid screening of antimicrobial residues in meat, fish and eggs, was evaluated and compared with the (modified) One-Plate Test, an agar diffusion assay. The performance characteristics described for qualitative, screening methods in Commission Decision 2002/657/EC were used for the evaluation. The Premi Test was found to detect a range of antimicrobials to MRL levels in kidney fluid but to have poorer sensitivity for some antimicrobials such as tetracyclines, sulphonamides, flumequine and streptomycin. The test was found not to be sensitive for the banned antimicrobial chloramphenicol. The One-Plate Test was found to detect most tetracyclines and flumequine to MRL levels but to be less sensitive than the Premi Test for most of the other classes of antimicrobials. Neither test alone provides a comprehensive screening test for antimicrobial residues in kidney at MRL levels. However, the Premi Test is fast, easy to use and rugged and, in combination with other antimicrobial tests, may be used to provide a comprehensive screening system for antimicrobials in tissues.

2,3-Dideoxyglucosides of selected terpene phenols and alcohols as potent antifungal compounds.

PubMed

James Bound, D; Murthy, Pushpa S; Srinivas, P

2016-11-01

The antifungal activities of novel 2,3-unsaturated and 2,3-dideoxy 1-O-glucosides of carvacrol, thymol, and perillyl alcohol were tested against Aspergillus flavus, Aspergillus ochraceus, Fusarium oxysporum, Saccharomyces cerevisiae and Candida albicans. In the agar well diffusion tests, zones of inhibition for the derivatives of carvacrol, thymol and perillyl alcohol were higher (15-30mm) in the case of filamentous fungi than those for the parent compounds. Their MIC and MFC values indicated that the 2,3-unsaturated and 2,3-dideoxy 1-O-glucosides of carvacrol and thymol exhibited more fungicidal activity than the other compounds. Further, the 2,3-dideoxyglucosides of carvacrol and thymol, exhibited antitoxigenic effects against A. ochraceus and A. flavus and inhibited the production of ochratoxin and aflatoxin-B2. Propidium iodide influx assay demonstrated the lysis of C. albicans cells by carvacrol and its 2,3-unsaturated 1-O-glucoside and the loss of the membrane integrity. These new 2,3-dideoxyglucosides can be useful as antifungal agents and condiments in foods. Copyright © 2016 Elsevier Ltd. All rights reserved.

Morphological changes of Ganoderma boninense mycelia after challenged by Trichoderma and Bacillus

DOE Office of Scientific and Technical Information (OSTI.GOV)

Alexander, Arnnyitte; Chong, Khim-Phin, E-mail: chongkp@ums.edu.my; Dayou, Jedol

Ganoderma boninense is a fungal pathogen that causes Basal Stem Rot (BSR) disease in oil palm. This deadly disease has caused major losses in the oil palm industry and no remedy is reported to date. The more promising control on G. boninense is the use of biological control agents (BCAs). Despite many attempts in using BCAs as a control agent but evidence on the colonization of BCAs and morphological changes of the pathogen is not well documented. We have investigated the effect of antagonist activity on the combination of Trichoderma spp. and Bacillus spp. on the morphology of G. boninense.more » The antagonist activity was evaluated using agar well diffusion assay. BCAs suppressed the mycelia growth of G. boninense up to 70%. Observation under Scanning Electron Microscopy (SEM) shows these BCAs induced stripping of G. boninense hyphal structure by destroying the cellular structure. Highly disrupted, disaggerated, shrivelled and lysis of G. boninense hyphal were also observed. The antifungal activity of Trichoderma spp. and Bacillus spp. observed could be associated with the production of Cell Wall Degrading Enzymes (CWDE)« less

Morphological changes of Ganoderma boninense mycelia after challenged by Trichoderma and Bacillus

NASA Astrophysics Data System (ADS)

Alexander, Arnnyitte; Dayou, Jedol; Chong, Khim-Phin

2015-07-01

Ganoderma boninense is a fungal pathogen that causes Basal Stem Rot (BSR) disease in oil palm. This deadly disease has caused major losses in the oil palm industry and no remedy is reported to date. The more promising control on G. boninense is the use of biological control agents (BCAs). Despite many attempts in using BCAs as a control agent but evidence on the colonization of BCAs and morphological changes of the pathogen is not well documented. We have investigated the effect of antagonist activity on the combination of Trichoderma spp. and Bacillus spp. on the morphology of G. boninense. The antagonist activity was evaluated using agar well diffusion assay. BCAs suppressed the mycelia growth of G. boninense up to 70%. Observation under Scanning Electron Microscopy (SEM) shows these BCAs induced stripping of G. boninense hyphal structure by destroying the cellular structure. Highly disrupted, disaggerated, shrivelled and lysis of G. boninense hyphal were also observed. The antifungal activity of Trichoderma spp. and Bacillus spp. observed could be associated with the production of Cell Wall Degrading Enzymes (CWDE).

Antimicrobial and Anti-Inflammatory Activities of Pterygota macrocarpa and Cola gigantea (Sterculiaceae)

PubMed Central

Agyare, Christian; Koffuor, George Asumeng; Boamah, Vivian Etsiapa; Adu, Francis; Mensah, Kwesi Boadu; Adu-Amoah, Louis

2012-01-01

Pterygota macrocarpa and Cola gigantea are African medicinal plants used in traditional medicine for the treatment of sores, skin infections, and other inflammatory conditions including pains. This study therefore aims at investigating the antimicrobial properties of ethanol leaf and stem bark extracts of P. macrocarpa and C. gigantea using the agar diffusion and the micro-dilution techniques and also determining the anti-inflammatory properties of the extracts of these plants in carrageenan-induced foot edema in seven-day old chicks. The minimum inhibitory concentration of both ethanol leaf and bark extracts of P. macrocarpa against the test organisms was from 0.125 to 2.55 mg/mL and that of C. gigantea extracts was 0.125 to 2.75 mg/mL. Extracts with concentration of 50 mg/mL were most active against the test organisms according to the agar diffusion method. All the extracts of P. macrocarpa and C. gigantea at 30, 100, and 300 mg/kg body weight except ethanol leaf extract of C. gigantea exhibited significant anti-inflammatory effects (P ≤ 0.001). PMID:22690251

Enhance the anti-microorganism activity of cinnamon oil by xanthan gum as emulsifying agent

NASA Astrophysics Data System (ADS)

Lieu, Dong M.; Dang, Thuy T. K.; Nguyen, Huong T.

2018-04-01

The aim of this study was to evaluate the effect of emulsifying agents (tween 20, DMSO (Dimethyl Sulfoxide) and xanthan gum) to inhibit Escherichia coli; Staphylococcus aureus; Saccharomyces cerevisiae and Aspergillus niger by cinnamon oil (Cinnamomum Cassia). Cinnamon oil was added in the emulsifying agents independently: tween 20 (0.3% v/v). DMSO (0.3% v/v) and xanthan gum (0.3% w/v) at different concentrations and evaluated their anti-microorganism activity by agar disk diffusion, mycelial growth inhibition and growth inhibition in liquid phase. The result indicated that, cinnamon oil diluted in different emulsifying agents showed the difference of the anti-microorganism activity, in which DMSO showed the lowest result. Xanthan gum and tween 20 show good stable emulsion. The anti-microorganism effect of cinnamon oil in tween 20 and xanthan gum was not significant difference. However, cinnamon oil in xanthan gum showed anti-microorganism activity better than tween 20 at low concentration in agar disk diffusion. This suggests that, cinnamon oil could be encapsulated by xanthan gum to enhance the anti-microorganism activity.

Evaluation of VIDAS UP Listeria assay (LPT) for the detection of Listeria in a variety of foods and environmental surfaces: First Action 2013.10.

PubMed

Crowley, Erin; Bird, Patrick; Flannery, Jonathan; Benzinger, M Joseph; Fisher, Kiel; Boyle, Megan; Huffman, Travis; Bastin, Ben; Bedinghaus, Paige; Judd, William; Hoang, Thao; Agin, James; Goins, David; Johnson, Ronald L

2014-01-01

The VIDAS UP Listeria (LPT) is an automated rapid screening enzyme phage-ligand based assay for the detection of Listeria species in human food products and environmental samples. The VIDAS LPT method was compared in a multi-laboratory collaborative study to AOAC Official Method 993.12 Listeria monocytogenes in Milk and Dairy Products reference method following current AOAC guidelines. A total of 14 laboratories participated, representing government and industry, throughout the United States. One matrix, queso fresco (soft Mexican cheese), was analyzed using two different test portion sizes, 25 and 125 g. Samples representing each test portion size were artificially contaminated with Listeria species at three levels, an uninoculated control level [0 colony-forming units (CFU)/test portion], a low-inoculum level (0.2-2 CFU/test portion), and a high-inoculum level (2-5 CFU/test portion). For this evaluation, 1800 unpaired replicate test portions were analyzed by either the VIDAS LPT or AOAC 993.12. Each inoculation level was analyzed using the Probability of Detection (POD) statistical model. For the low-level inoculated test portions, difference in collaborator POD (dLPOD) values of 0.01, (-0.10, 0.13), with 95% confidence intervals, were obtained for both 25 and 125 g test portions. The range of the confidence intervals for dLPOD values for both the 25 and 125 g test portions contains the point 0.0 indicating no statistically significant difference in the number of positive samples detected between the VIDAS LPT and the AOAC methods. In addition to Oxford agar, VIDAS LPT test portions were confirmed using Agar Listeria Ottavani and Agosti (ALOA), a proprietary chromogenic agar for the identification and differentiation of L. monocytogenes and Listeria species. No differences were observed between the two selective agars. The VIDAS LPT method, with the optional ALOA agar confirmation method, was adopted as Official First Action status for the detection of Listeria species in a variety of foods and environmental samples.

Multi-chamber electroosmosis using textile reinforced agar membranes--A promising concept for the future of hemodialysis.

PubMed

Kofler, Markus; Lenninger, Margit; Mayer, Gert; Neuwirt, Hannes; Grimm, Michael; Bechtold, Thomas

2016-01-20

Renal replacement therapy options are limited to hemodialysis and peritoneal dialysis (70% of US patients) or renal transplantation. Diffusion processes are the main physico-chemical principle behind hemodialysis. An alternative way to achieve liquid flow through membranes bases on the electroosmotic flow which is observed as electrokinetic phenomenon in porous membranes which bear surface charges. Agar consists of the non-ionic agarose and the negatively charged agaropectine thus an electroosmotic flux is observed in analytical electrophoresis. In this study the potential electroosmosis on textile reinforced agar membranes as separation method was investigated. Using a five-chamber electrolysis cell and an agar membrane/cellulose fabric composite an intensive electroosmotic flow of 1-2 ml cm(2) h(-1) at 100 mA cell current could be observed. The movement of cations in the negatively charged agar structure led to an intensive electroosmotic flux, which also transported uncharged molecules such as urea, glucose through the membrane. Separation of uncharged low molecular weight molecules is determined by the membrane characteristic. The transport of ions (K(+), PO4(3-), creatinine) and uncharged molecules (urea, glucose) in electroosmotic separation experiments was monitored using a pH 5.5 phosphate electrolyte with the aim to assess the overall transport processes in the electrochemical cell. The results demonstrate the potential of the method for filtration of biological fluids in the absence of external pressure or high shear rates. Copyright © 2015 Elsevier Ltd. All rights reserved.

Echinocandin Susceptibility Testing of Candida Species: Comparison of EUCAST EDef 7.1, CLSI M27-A3, Etest, Disk Diffusion, and Agar Dilution Methods with RPMI and IsoSensitest Media▿

PubMed Central

Arendrup, Maiken Cavling; Garcia-Effron, Guillermo; Lass-Flörl, Cornelia; Lopez, Alicia Gomez; Rodriguez-Tudela, Juan-Luis; Cuenca-Estrella, Manuel; Perlin, David S.

2010-01-01

This study compared nine susceptibility testing methods and 12 endpoints for anidulafungin, caspofungin, and micafungin with the same collection of blinded FKS hot spot mutant (n = 29) and wild-type isolates (n = 94). The susceptibility tests included EUCAST Edef 7.1, agar dilution, Etest, and disk diffusion with RPMI-1640 plus 2% glucose (2G) and IsoSensitest-2G media and CLSI M27A-3. Microdilution plates were read after 24 and 48 h. The following test parameters were evaluated: fks hot spot mutants overlapping the wild-type distribution, distance between the two populations, number of very major errors (VMEs; fks mutants misclassified as susceptible), and major errors (MEs; wild-type isolates classified as resistant) using a wild-type-upper-limit value (WT-UL) (two twofold-dilutions higher than the MIC50) as the susceptibility breakpoint. The methods with the lowest number of errors (given as VMEs/MEs) across the three echinocandins were CLSI (12%/1%), agar dilution with RPMI-2G medium (14%/0%), and Etest with RPMI-2G medium (8%/3%). The fewest errors overall were observed for anidulafungin (4%/1% for EUCAST, 4%/3% for CLSI, and 3%/9% for Etest with RPMI-2G). For micafungin, VME rates of 10 to 71% were observed. For caspofungin, agar dilution with either medium was superior (VMEs/MEs of 0%/1%), while CLSI, EUCAST with IsoSensitest-2G medium, and Etest were less optimal (VMEs of 7%, 10%, and 10%, respectively). Applying the CLSI breakpoint (S ≤ 2 μg/ml) for CLSI results, 89.2% fks hot spot mutants were classified as anidulafungin susceptible, 60.7% as caspofungin susceptible, and 92.9% as micafungin susceptible. In conclusion, no test was perfect, but anidulafungin susceptibility testing using the WT-UL to define susceptibility reliably identified fks hot spot mutants. PMID:19884370

In vitro antibacterial activity of adhesive systems on Streptococcus mutans.

PubMed

Paradella, Thaís Cachuté; Koga-Ito, Cristiane Yumi; Jorge, Antonio Olavo Cardoso

2009-04-01

To evaluate the antibacterial activity of three adhesive systems -- Prime & Bond 2.1 (PB), Clearfil SE Bond (CS) and One Up Bond F (OU) -- on Streptococcus mutans in vitro. Adherence and agar disk-diffusion tests were performed. For the adherence testing, 40 human enamel specimens (4 mm2) were sterilized and the adhesive sytems were applied (n = 10). The control group did not receive the application of any adhesive system. Specimens were immersed in brain heart infusion broth (BHI) inoculated with S. mutans standardized suspension (10(6) cells/ml) for 48 h at 37 degrees C and 5% CO2. The number of S. mutans cells adhered to each specimen was evaluated by the plating method on BHI agar. For agar disk-diffusion testing, adhesive disks and disks soaked in distilled water (negative control) or 0.2% chlorexidine (positive control) were incubated with S. mutans for 48 h. The diameters of the zones of bacterial inhibition were measured. Adherence data were transformed in logarithms of base 10 (log10). Data were submitted to Kruskal-Wallis and Student-Neuman-Keuls tests at the 5% level of significance. The results of the adherence test showed that One Up Bond F (OU) and Clearfil SE Bond (CS) did not differ significantly from one another, but allowed significantly less adherence than Prime & Bond 2.1 (PB) and control [mean log10 (standard deviation) values: PB 6.10 (0.19); CS primer 4.55 (0.98); OU 4.65 (0.54); control group 6.34 (0.27)]. The disk-diffusion test showed no significant difference between OU (diameter in mm: 3.02 +/- 0.13) and CS (3.0 +/- 0.12), but both were significantly more effective in inhibiting bacterial growth than PB (1.0 +/- 0.10). The self-etching systems Clearfil SE Bond and One Up Bond F presented a greater inhibitory effect against S. mutans, also in terms of adherence, than did the conventional system, Prime & Bond 2.1.

Detection of Salmonella spp. with the BACTEC 9240 Automated Blood Culture System in 2008 - 2014 in Southern Iran (Shiraz): Biogrouping, MIC, and Antimicrobial Susceptibility Profiles of Isolates.

PubMed

Anvarinejad, Mojtaba; Pouladfar, Gholam Reza; Pourabbas, Bahman; Amin Shahidi, Maneli; Rafaatpour, Noroddin; Dehyadegari, Mohammad Ali; Abbasi, Pejman; Mardaneh, Jalal

2016-04-01

Human salmonellosis continues to be a major international problem, in terms of both morbidity and economic losses. The antibiotic resistance of Salmonella is an increasing public health emergency, since infections from resistant bacteria are more difficult and costly to treat. The aims of the present study were to investigate the isolation of Salmonella spp. with the BACTEC automated system from blood samples during 2008 - 2014 in southern Iran (Shiraz). Detection of subspecies, biogrouping, and antimicrobial susceptibility testing by the disc diffusion and agar dilution methods were performed. A total of 19 Salmonella spp. were consecutively isolated using BACTEC from blood samples of patients between 2008 and 2014 in Shiraz, Iran. The isolates were identified as Salmonella, based on biochemical tests embedded in the API-20E system. In order to characterize the biogroups and subspecies, biochemical testing was performed. Susceptibility testing (disc diffusion and agar dilution) and extended-spectrum β-lactamase (ESBL) detection were performed according to the clinical and laboratory standards institute (CLSI) guidelines. Of the total 19 Salmonella spp. isolates recovered by the BACTEC automated system, all belonged to the Salmonella enterica subsp. houtenae. Five isolates (26.5%) were resistant to azithromycin. Six (31.5%) isolates with the disc diffusion method and five (26.3%) with the agar dilution method displayed resistance to nalidixic acid (minimum inhibitory concentration [MIC] > 32 μg/mL). All nalidixic acid-resistant isolates were also ciprofloxacin-sensitive. All isolates were ESBL-negative. Twenty-one percent of isolates were found to be resistant to chloramphenicol (MIC ≥ 32 μg/mL), and 16% were resistant to ampicillin (MIC ≥ 32 μg/mL). The results indicate that multidrug-resistant (MDR) strains of Salmonella are increasing in number, and fewer antibiotics may be useful for treating S. enterica infections. Routine investigation and reporting of antibiotic MICs in patients presenting with Salmonella infections is suggested.

Hair sheep blood, citrated or defibrinated, fulfills all requirements of blood agar for diagnostic microbiology laboratory tests.

PubMed

Yeh, Ellen; Pinsky, Benjamin A; Banaei, Niaz; Baron, Ellen Jo

2009-07-03

Blood agar is used for the identification and antibiotic susceptibility testing of many bacterial pathogens. In the developing world, microbiologists use human blood agar because of the high cost and inhospitable conditions for raising wool sheep or horses to supply blood. Many pathogens either fail to grow entirely or exhibit morphologies and hemolytic patterns on human blood agar that confound colony recognition. Furthermore, human blood can be hazardous to handle due to HIV and hepatitis. This study investigated whether blood from hair sheep, a hardy, low-maintenance variety of sheep adapted for hot climates, was suitable for routine clinical microbiology studies. Hair sheep blood obtained by jugular venipuncture was anticoagulated by either manual defibrination or collection in human blood bank bags containing citrate-phosphate-dextrose. Trypticase soy 5% blood agar was made from both forms of hair sheep blood and commercial defibrinated wool sheep blood. Growth characteristics, colony morphologies, and hemolytic patterns of selected human pathogens, including several streptococcal species, were evaluated. Specialized identification tests, including CAMP test, reverse CAMP test, and satellite colony formation with Haemophilus influenzae and Abiotrophia defectiva were also performed. Mueller-Hinton blood agar plates prepared from the three blood types were compared in antibiotic susceptibility tests by disk diffusion and E-test. The results of all studies showed that blood agar prepared from citrated hair sheep blood is suitable for microbiological tests used in routine identification and susceptibility profiling of human pathogens. The validation of citrated hair sheep blood eliminates the labor-intensive and equipment-requiring process of manual defibrination. Use of hair sheep blood, in lieu of human blood currently used by many developing world laboratories and as an alternative to cost-prohibitive commercial sheep blood, offers the opportunity to dramatically improve the safety and accuracy of laboratory diagnosis of pathogenic bacteria in resource-poor countries.

In vitro activity of natural honey alone and in combination with curcuma starch against Rhodotorula mucilaginosa in correlation with bioactive compounds and diastase activity

PubMed Central

Ahmed, Moussa; Djebli, Noureddine; Aissat, Saad; Khiati, Baghdad; Meslem, Abdelmalek; Bacha, Salima

2013-01-01

Objective To evaluate the in vitro activity and synergism of the combinations of natural honey and curcuma starch against Rhodotorula mucilaginosa in correlation with total phenolic, flavonoid contents, and diastase activity. Methods The Folin-Ciocalteu test was used to determine the total polyphenols content and the flavonoid content was analyzed using by the aluminum chloride method. The antifungal activity of the natural honey, determined by an agar well diffusion assay and agar incorporation method. Results Total phenolic content varied from (63.93±0.11) to (95.36±6.08) mg GAE/100 g honey as gallic acid equivalent. Total flavonoids content varied from (5.41±0.04) to (9.94±0.54) mg CE/100 g. Diastase activity values were between (7.3±2.8) and (26±2.8). The zone inhibition diameter for the six honey samples without starch ranged between 6 and 20 mm. When starch was mixed with honey and then added to well, a zone inhibition increase diameter 7 and 21 mm. The percentage increase was noticed with each variety and it ranged between 5% and 62.5%. The minimal inhibitory concentrations for the six varieties of honey without starch against Rhodotorula mucilaginosa ranged between 28% and 36% (v/v). When starch was incubated with honey and then added to media, a minimal inhibitory concentration drop has been noticed with each variety. It ranged between 6.66 % and 20% (w/v). No significant correlation was established between diastase activity and bioactive compounds. Conclusions The mixture of curcuma starch and honey could lead to the development of new combination antibiotics against Rhodotorula infections PMID:24075348

Propolis, A Hope for the Future in Treating Resistant Periodontal Pathogens.

PubMed

Shabbir, Ambreen; Rashid, Maryam; Tipu, Hamid N

2016-07-12

Periodontitis is one of the most common causes of tooth loss worldwide. Recently, special attention has been paid to natural medication for its treatment. For this purpose, propolis (bee glue) activity has also been investigated. Its antibacterial properties are mainly attributed to flavonones pinocembrin, flavonols galangin and to the caffeic acid phenethyl ester. This study is aimed at evaluating the antimicrobial effects of propolis from Pakistan on 35 clinical isolates of pigmented anaerobic periodontal pathogens. This study was conducted in the Microbiology department, University of Health Sciences, Lahore, Pakistan. Pathogens included were Porphyromonas asaccharolytica (n=9), Porphyromonas gingivalis (n=13), Prevotella intermedia (n=9), Prevotella melaninogenica (n=4). Minimum inhibitory concentration (MIC) to three antibiotics was obtained by E-test method. All strains were sensitive to amoxicillin plus clavulanic acid and metronidazole, but 100% of P asaccharolytica and P melaninogenica strains displayed intermediate resistance to tetracycline while 69.2% P gingivalis and 100% P intermedia strains exhibited complete resistance to tetracycline. Screening for antibacterial activity of propolis extract was done by agar well diffusion assay, and all strains were found sensitive to ethanolic extract of propolis. MIC was obtained by agar incorporation technique with values ranging from 0.064 to 0.512 mg/ml. It was also noticed that percentage yield of ethanolic extract of propolis prepared from ultrasonic extraction method was higher compared to extract obtained with maceration. These results indicate that propolis from this region has potent antimicrobial activity against pigmented anaerobic periodontal pathogens. Taking into consideration the increasing resistance in anaerobic bacteria, this effective antimicrobial activity of propolis gives hope in the treatment of oral cavity diseases.

Propolis, A Hope for the Future in Treating Resistant Periodontal Pathogens

PubMed Central

Rashid, Maryam; Tipu, Hamid N

2016-01-01

Introduction: Periodontitis is one of the most common causes of tooth loss worldwide. Recently, special attention has been paid to natural medication for its treatment. For this purpose, propolis (bee glue) activity has also been investigated. Its antibacterial properties are mainly attributed to flavonones pinocembrin, flavonols galangin and to the caffeic acid phenethyl ester. This study is aimed at evaluating the antimicrobial effects of propolis from Pakistan on 35 clinical isolates of pigmented anaerobic periodontal pathogens. Methods: This study was conducted in the Microbiology department, University of Health Sciences, Lahore, Pakistan. Pathogens included were Porphyromonas asaccharolytica (n=9), Porphyromonas gingivalis (n=13), Prevotella intermedia (n=9), Prevotella melaninogenica (n=4). Minimum inhibitory concentration (MIC) to three antibiotics was obtained by E-test method. All strains were sensitive to amoxicillin plus clavulanic acid and metronidazole, but 100% of P asaccharolytica and P melaninogenica strains displayed intermediate resistance to tetracycline while 69.2% P gingivalis and 100% P intermedia strains exhibited complete resistance to tetracycline. Screening for antibacterial activity of propolis extract was done by agar well diffusion assay, and all strains were found sensitive to ethanolic extract of propolis. Results: MIC was obtained by agar incorporation technique with values ranging from 0.064 to 0.512 mg/ml. It was also noticed that percentage yield of ethanolic extract of propolis prepared from ultrasonic extraction method was higher compared to extract obtained with maceration. Conclusion: These results indicate that propolis from this region has potent antimicrobial activity against pigmented anaerobic periodontal pathogens. Taking into consideration the increasing resistance in anaerobic bacteria, this effective antimicrobial activity of propolis gives hope in the treatment of oral cavity diseases. PMID:27563508

In vitro activity of natural honey alone and in combination with curcuma starch against Rhodotorula mucilaginosa in correlation with bioactive compounds and diastase activity.

PubMed

Ahmed, Moussa; Djebli, Noureddine; Aissat, Saad; Khiati, Baghdad; Meslem, Abdelmalek; Bacha, Salima

2013-10-01

To evaluate the in vitro activity and synergism of the combinations of natural honey and curcuma starch against Rhodotorula mucilaginosa in correlation with total phenolic, flavonoid contents, and diastase activity. The Folin-Ciocalteu test was used to determine the total polyphenols content and the flavonoid content was analyzed using by the aluminum chloride method. The antifungal activity of the natural honey, determined by an agar well diffusion assay and agar incorporation method. Total phenolic content varied from (63.930.11) to (95.366.08) mg GAE/100 g honey as gallic acid equivalent. Total flavonoids content varied from (5.41±0.04) to (9.94±0.54) mg CE/100 g. Diastase activity values were between (7.3±2.8) and (26±2.8). The zone inhibition diameter for the six honey samples without starch ranged between 6 and 20 mm. When starch was mixed with honey and then added to well, a zone inhibition increase diameter 7 and 21 mm. The percentage increase was noticed with each variety and it ranged between 5% and 62.5%. The minimal inhibitory concentrations for the six varieties of honey without starch against Rhodotorula mucilaginosa ranged between 28% and 36% (v/v). When starch was incubated with honey and then added to media, a minimal inhibitory concentration drop has been noticed with each variety. It ranged between 6.66 % and 20% (w/v). No significant correlation was established between diastase activity and bioactive compounds. The mixture of curcuma starch and honey could lead to the development of new combination antibiotics against Rhodotorula infections. Copyright © 2013 Asian Pacific Tropical Biomedical Magazine. Published by Elsevier B.V. All rights reserved.

[Methicillin resistance detection in Staphylococcus aureus: comparison between conventional methods and MRSA-Screen latex agglutination technique].

PubMed

Soloaga, R; Corso, A; Gagetti, P; Faccone, D; Galas, M

2004-01-01

Methicillin-resistant Staphylococcus aureus (MRSA) is a significant pathogen that has emerged over the last four decades, causing both nosocomial and community-acquired infections. Rapid and accurate detection of methicillin resistance in S. aureus is important for the use of appropriate antimicrobial therapy and for the control of nosocomial spread of MRSA strains. We evaluated the efficiency of conventional methods for detection of methicillin resistance such as the disk diffusion, agar dilution, oxacillin agar screen test, and the latex agglutination test MRSA-Screen latex, in 100 isolates of S. aureus, 79 mecA positive and 21 mecA negative. The MRSA-Screen latex (Denka Seiken, Niigata, Japón), is a latex agglutination method that detects the presence of PLP-2a, product of mecA gene in S. aureus. The PCR of the mecA gene was used as the "gold standard" for the evaluation of the different methods tested. The percentages of sensitivity and specificity were as follows: disk difusión 97 and 100%, agar dilution 97 and 95%, oxacillin agar screen test 100 and 100%, and MRSA-Screen latex, 100 and 100 %. All methods presented high sensitivity and specificity, but MRSA-Screen latex had the advantage of giving a reliable result, equivalent to PCR, in only 15 minutes.

Antimicrobial activity and safety evaluation of Enterococcus faecium KQ 2.6 isolated from peacock feces.

PubMed

Zheng, Wei; Zhang, Yu; Lu, Hui-Min; Li, Dan-Ting; Zhang, Zhi-Liang; Tang, Zhen-Xing; Shi, Lu-E

2015-05-12

The objective of this paper was to study antimicrobial activity and safety of Enterococcus faecium KQ 2.6 (E. faecium KQ 2.6) isolated from peacock feces. Agar well diffusion method was adopted in antimicrobial activity assay. Disk diffusion test was used to determine the antibiotic resistance. The identification and virulence potential of E. faecium KQ 2.6 were investigated using PCR amplification. The results indicated that cell free supernatant (CFS) of the strain had the good antimicrobial activity against selected gram-positive and gram-negative bacteria. The biochemical characteristics of antimicrobial substances were investigated. The results indicated that the antimicrobial substances were still active after treatment with catalase and proteinase, respectively. Moreover, the stability of antimicrobial substances did not change after heat treatment at 40, 50, 60, 70 and 80°C for 30 min, respectively. The activity of antimicrobial substances remained stable at 4 and -20°C after long time storage. The antimicrobial activity of CFS was compared with that of the buffer with similar strength and pH. The inhibitory zone of the buffer was apparently smaller than that of CFS, which meant that the acid in CFS was not the only factor that was contributed to antibacterial activity of CFS. The antibiotic resistance and virulence potential were evaluated using disk diffusion test and PCR amplification. The results showed that E. faecium KQ 2.6 did not harbor any tested virulence genes such as gelE, esp, asa1, cylA, efaA and hyl. It was susceptible to most of tested antibiotics except for vancomycin and polymyxin B. E. faecium KQ 2.6 may be used as bio-preservative cultures for the production of fermented foods.

[Effects of stable ANO1 overexpression on biological behaviors of human laryngeal squamous cell carcinoma Hep-2 cells in vitro].

PubMed

Li, Yadong; Zhang, Jinsong; Yang, Kai; Zhang, Fujun; Chen, Rui; Chen, Dan

2014-02-01

To detect the effects of ANO1 overexpression on the biological behaviors of human laryngeal squamous cell carcinoma Hep-2 cells. A Hep-2 cell line stably overexpressing ANO1 were examined with flow cytometry, soft agar assay, wound healing assay, siRNA experiments, and chloride channel block with DIDS to observe the effect of ANO1 overexpression on the growth, migration and invasion of the cells. Flow cytometry revealed a comparable cell percentage in G0/G1 phase between ANO1-overexpressing cells and the control cells (P>0.05). The two cells showed no significant difference in soft agar assay (P>0.05), but in wound healing experiments, ANO1-overexpressing cells showed significantly accelerated migration (P<0.05), whereas siRNA-mediated silencing of ANO1 significantly inhibited the cell migration (P<0.05). Treatment with DIDS resulted in an effective block of the ANO1 chloride channel activity and obviously decreased the migration speed of Hep-2 cells. ANO1 overexpression does not significantly affect the proliferation of cancer cells, but can enhance the migration ability of head and neck squamous cell carcinoma, suggesting the value of ANO1 as a new gene therapy target for head and neck squamous cell carcinoma.

Rapid Quantitative Detection of Lactobacillus sakei in Meat and Fermented Sausages by Real-Time PCR

PubMed Central

Martín, Belén; Jofré, Anna; Garriga, Margarita; Pla, Maria; Aymerich, Teresa

2006-01-01

A quick and simple method for quantitative detection of Lactobacillus sakei in fermented sausages was successfully developed. It is based on Chelex-100-based DNA purification and real-time PCR enumeration using a TaqMan fluorescence probe. Primers and probes were designed in the L. sakei 16S-23S rRNA intergenic transcribed spacer region, and the assay was evaluated using L. sakei genomic DNA and an artificially inoculated sausage model. The detection limit of this technique was approximately 3 cells per reaction mixture using both purified DNA and the inoculated sausage model. The quantification limit was established at 30 cells per reaction mixture in both models. The assay was then applied to enumerate L. sakei in real samples, and the results were compared to the MRS agar count method followed by confirmation of the percentage of L. sakei colonies. The results obtained by real-time PCR were not statistically significantly different than those obtained by plate count on MRS agar (P > 0.05), showing a satisfactory agreement between both methods. Therefore, the real-time PCR assay developed can be considered a promising rapid alternative method for the quantification of L. sakei and evaluation of the implantation of starter strains of L. sakei in fermented sausages. PMID:16957227

Rapid quantitative detection of Lactobacillus sakei in meat and fermented sausages by real-time PCR.

PubMed

Martín, Belén; Jofré, Anna; Garriga, Margarita; Pla, Maria; Aymerich, Teresa

2006-09-01

A quick and simple method for quantitative detection of Lactobacillus sakei in fermented sausages was successfully developed. It is based on Chelex-100-based DNA purification and real-time PCR enumeration using a TaqMan fluorescence probe. Primers and probes were designed in the L. sakei 16S-23S rRNA intergenic transcribed spacer region, and the assay was evaluated using L. sakei genomic DNA and an artificially inoculated sausage model. The detection limit of this technique was approximately 3 cells per reaction mixture using both purified DNA and the inoculated sausage model. The quantification limit was established at 30 cells per reaction mixture in both models. The assay was then applied to enumerate L. sakei in real samples, and the results were compared to the MRS agar count method followed by confirmation of the percentage of L. sakei colonies. The results obtained by real-time PCR were not statistically significantly different than those obtained by plate count on MRS agar (P > 0.05), showing a satisfactory agreement between both methods. Therefore, the real-time PCR assay developed can be considered a promising rapid alternative method for the quantification of L. sakei and evaluation of the implantation of starter strains of L. sakei in fermented sausages.

Characterization, production, and purification of carnocin H, a bacteriocin produced by Carnobacterium 377.

PubMed

Blom, H; Katla, T; Nissen, H; Holo, H

2001-10-01

Carnocin H, a bacteriocin produced by a Carnobacterium sp., inhibited lactic acid bacteria, clostridia, enterococci, and some Staphylococcus aureus strains. Some strains of Listeria and Pediococcus were also sensitive to carnocin H. The bacteriocin was produced during the late stationary growth phase. Carnocin H was purified by cation exchange chromatography and reverse phase chromatography. Amino acid sequence and composition indicate that carnocin H is a novel bacteriocin belonging to the class II bacteriocins. The bacteriocin consists of approximately 75 amino acid residues with a highly cationic N-terminal containing six succeeding lysines. Activity, as measured by agar diffusion zones, was reduced at increased pH values, levels of indicator bacteria, NaCl, agar, and soy oil.

Plating Bacteriophage M13.

PubMed

Green, Michael R; Sambrook, Joseph

2017-10-03

A plaque of bacteriophage M13 derives from infection of a single bacterium by a single virus particle. The progeny particles infect neighboring bacteria, which, in turn, release another generation of daughter virus particles. If the bacteria are growing in semisolid medium (e.g., containing agar or agarose), then the diffusion of the progeny particles is limited. Cells infected with bacteriophage M13 are not killed, but have a longer generation time than uninfected Escherichia coli In consequence, plaques appear as areas of slower-growing cells on a faster-growing lawn of bacterial cells. This protocol describes plating of bacteriophage M13 stocks. Plaques are readily detectable on top agar after 4-8 h of incubation at 37°C. © 2017 Cold Spring Harbor Laboratory Press.

Diffusion of Antimicrobials Across Silicone Hydrogel Contact Lenses.

PubMed

Zambelli, Alison M; Brothers, Kimberly M; Hunt, Kristin M; Romanowski, Eric G; Nau, Amy C; Dhaliwal, Deepinder K; Shanks, Robert M Q

2015-09-01

To measure the diffusion of topical preparations of moxifloxacin, amphotericin B (AmB), and polyhexamethylene biguanide (PHMB) through silicone hydrogel (SH) contact lenses (CLs) in vitro. Using an in vitro model, the diffusion of three antimicrobials through SH CLs was measured. Diffused compounds were measured using a spectrophotometer at set time points over a period of 4 hr. The amount of each diffused antimicrobial was determined by comparing the experimental value with a standard curve. A biological assay was performed to validate the CL diffusion assay by testing antimicrobial activity of diffused material against lawns of susceptible bacteria (Staphylococcus epidermidis) and yeast (Saccharomyces cerevisiae). Experiments were repeated at least two times with a total of at least four independent replicates. Our data show detectable moxifloxacin and PHMB diffusion through SH CLs at 30 min, whereas AmB diffusion remained below the limit of detection within the 4-hr experimental period. In the biological assay, diffused moxifloxacin demonstrated microbial killing starting at 20 min on bacterial lawns, whereas PHMB and AmB failed to demonstrate killing on microbial lawns over the course of the 60-min experiment. In vitro diffusion assays demonstrate limited penetration of certain anti-infective agents through SH CLs. Further studies regarding the clinical benefit of using these agents along with bandage CL for corneal pathologic condition are warranted.

75 FR 61148 - Food and Drug Administration Modernization Act of 1997: Modifications to the List of Recognized...

Federal Register 2010, 2011, 2012, 2013, 2014

2010-10-04

... Evaluation and testing within a risk management process 2-100 ASTM E1372-95 (2003) Standard Test Method...) Standard Title, Type of Test Method for Agar Diffusion Cell standard , Relevant Culture Screening for... Biological evaluation of medical devices - Office(s) and Part 3: Tests for genotoxicity, Division(s...

Mouse Balb/c3T3 cell mutant with low epidermal growth factor receptor activity: induction of stable anchorage-independent growth by transforming growth factor. beta

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kuratomi, Y.; Ono, M.; Yasutake, C.

1987-01-01

A mutant clone (MO-5) was originally isolated as a clone resistant to Na/sup +//K/sup +/ ionophoric antibiotic monensin from mouse Balb/c3T3 cells. MO-5 was found to show low receptor-endocytosis activity for epidermal growth factor (EGF):binding activity for EGF in MO-5 was less than one tenth of that in Balb/c3T3. Anchorage-independent growth of MO-5 was compared to that of Balb/c3T3 when assayed by colony formation capacity in soft agar. Coadministration of EGF and TGF-..beta.. efficiently enhanced anchorage-independent growth of normal rat kidney (NRK) cells, but neither factor alone was competent to promote the anchorage-independent growth. The frequency of colonies appearing inmore » soft agar of MO-5 or Balb/c3T3 was significantly enhanced by TGF-..beta.. while EGF did not further enhance that of MO-5 or Balb/c3T3. Colonies of Balb/c3T3 formed in soft agar in the presence of TGF-..beta.. showed low colony formation capacity in soft agar in the absence of TGF-..beta... Colonies of MO-5 formed by TGF-..beta.. in soft agar, however, showed high colony formation capacity in soft agar in the absence of TGF-..beta... Pretreatment of MO-5 with TGF-..beta.. induced secretion of TGF-..beta..-like activity from the cells, while the treatment of Balb/c3T3 did not induce the secretion of a significant amount of TGF-..beta..-like activity. The loss of EGF-receptor activity in the stable expression and maintenance of the transformed phenotype in MO-5 is discussed.« less

In vitro antifungal sensitivity of fluconazole, clotrimazole and nystatin against vaginal candidiasis in females of childbearing age.

PubMed

Khan, Fouzia; Baqai, Rakhshanda

2010-01-01

Vaginal candidiasis is the most common infection of females. A large variety of antifungal drugs are used for treatment. The objective of this study was isolation and identification of Candida from high vaginal swabs and in vitro antifungal activity of Clotrimazole, Fluconazole and Nystatin against Candida. Two hundred and fifty high vaginal swabs were collected from females reporting at different hospitals of Karachi. Wet mount was performed to observe the budding cells of Candida. Vaginal swabs were cultured on Sabouraud's dextrose agar with added antibiotics. Plates were incubated at room temperature for seven days. Chlamydospores of Candida albicans were identified on corn meal agar. Species of Candida were identified on Biggy agar. In vitro antifungal activity of Clotrimazole, Fluconazole and Nystatin was performed by MIC (Minimum inhibitory concentration), well diffusion method and disc diffusion method. Out of 250 high vaginal swabs, Candida species were isolated in 100 (40%) of cases. Out of 100, C. albican 30 (30%), C. tropicalis 21 (21%), C. parapsillosis 10 (10%), C. parakrusi 8 (8%), C. glabrata 8 (8%), C. krusei 3 (3%) were isolated. In vitro antifungal activity indicated Clotrimazole (MIC 16 and 8 microg/ml) effective against 68 (70%) of Candida SPP, Fluconazole (MIC 64 and 32 microg/ml) effective against 29 (36.2%) and Nystatin disc (100 units) was 51 (63.5%) effective. C. albicans was mainly isolated. Clotrimazole was more effective as compared to Fluconazole and Nystatin. Antifungal susceptibility testing should be determined before therapy to avoid treatment failures.

Mycoflora and mycotoxin of hazelnut (Corylus avellana L.) and walnut (Juglans regia L.) seeds in Egypt.

PubMed

Abdel-Hafez, A I; Saber, S M

1993-03-01

Fifty-one species and 3 varieties appertaining to 20 genera were collected from 20 samples of each of hazelnut and walnut seeds on glucose- and 40% (W/V) sucrose-Czapek's agar at 25 degrees C and 45 degrees C with the most common mesophiles were Aspergillus flavus, A. fumigatus, A. niger, Cladosporium cladosporioides, C. herbarum, Penicillium chrysogenum, P. citrinum and P. oxalicum. Fusarium (represented by F. equiseti, F. moniliforme and F. oxysporum) was recovered from walnut seeds in moderate frequency (on glucose-Czapek's agar). Eurotium (E. amstelodami, E. chevalieri, E. repens and E. rubrum) was completely absent on glucose agar, but it was isolated in high frequency from the two types of seeds on 40% sucrose-Czapek's agar. Aspergillus fumigatus and Rhizomucor pusillus were the most common thermophilic fungi in hazelnut and walnut seeds on glucose agar at 45 degrees C. Humicola grisea var. themoidae and Thermoascus aurantiacus were encountered rarely from walnuts. The nuts samples were assayed for natural occurrence of aflatoxins B1, B2, G1 and G2, citrinin, ochratoxin A, patulin, sterigmatocystin, zearalenone, T-2 toxin and diacetoxyscirpenol by thin layer chromatography analysis. Aflatoxin was detected in 90% of hazelnut samples (25-175 micrograms/kg) and 75% of walnut samples (15-25 micrograms/kg). Zearalenone was detected in one sample of walnut (125 micrograms/kg). This is the first report for the presence of zearalenone in walnut. The other mycotoxins were not detected.

l-Methionine anti-biofilm activity against Pseudomonas aeruginosa is enhanced by the cystic fibrosis transmembrane conductance regulator potentiator, ivacaftor.

PubMed

Cho, Do-Yeon; Lim, Dong-Jin; Mackey, Calvin; Weeks, Christopher G; Peña Garcia, Jaime A; Skinner, Daniel; Grayson, Jessica W; Hill, Harrison S; Alexander, David K; Zhang, Shaoyan; Woodworth, Bradford A

2018-05-01

Biofilms may contribute to refractory chronic rhinosinusitis (CRS), as they lead to antibiotic resistance and failure of effective clinical treatment. l-Methionine is an amino acid with reported biofilm-inhibiting properties. Ivacaftor is a cystic fibrosis transmembrane conductance regulator (CFTR) potentiator with mild antimicrobial activity via inhibition of bacterial DNA gyrase and topoisomerase IV. The objective of this study was to evaluate whether co-treatment with ivacaftor and l-methionine can reduce the formation of Pseudomonas aeruginosa biofilms. P aeruginosa (PAO-1 strain) biofilms were studied in the presence of l-methionine and/or ivacaftor. For static biofilm assays, PAO-1 was cultured in a 48-well plate for 72 hours with stepwise combinations of these agents. Relative biofilm inhibitions were measured according to optical density of crystal violet stain at 590 nm. Live/dead assays (BacTiter-Glo™ assay, Promega) were imaged with laser scanning confocal microscopy. An agar diffusion test was used to confirm antibacterial effects of the drugs. l-Methionine (0.5 μM) significantly reduced PAO-1 biofilm mass (32.4 ± 18.0%; n = 4; p < 0.001) compared with controls. Low doses of ivacaftor alone (4, 8, and 12 μg/mL) had no effect on biofilm formation. When combined with ivacaftor (4 μg/mL), a synergistic anti-biofilm effect was noted at 0.05 μM and 0.5 μM of l-methionine (two-way analysis of variane, p = 0.0415) compared with corresponding concentrations of l-methionine alone. Ivacaftor enhanced the anti-biofilm activity of l-methionine against the PAO-1 strain of P aeruginosa. Further studies evaluating the efficacy of ivacaftor/l-methionine combinations for P aeruginosa sinusitis are planned. © 2018 ARS-AAOA, LLC.

The antimicrobial activity of heterotrophic bacteria isolated from the marine sponge Erylus deficiens (Astrophorida, Geodiidae)

PubMed Central

Graça, Ana Patrícia; Viana, Flávia; Bondoso, Joana; Correia, Maria Inês; Gomes, Luis; Humanes, Madalena; Reis, Alberto; Xavier, Joana R.; Gaspar, Helena; Lage, Olga M.

2015-01-01

Interest in the study of marine sponges and their associated microbiome has increased both for ecological reasons and for their great biotechnological potential. In this work, heterotrophic bacteria associated with three specimens of the marine sponge Erylus deficiens, were isolated in pure culture, phylogenetically identified and screened for antimicrobial activity. The isolation of bacteria after an enrichment treatment in heterotrophic medium revealed diversity in bacterial composition with only Pseudoalteromonas being shared by two specimens. Of the 83 selected isolates, 58% belong to Proteobacteria, 23% to Actinobacteria and 19% to Firmicutes. Diffusion agar assays for bioactivity screening against four bacterial strains and one yeast, revealed that a high number of the isolated bacteria (68.7%) were active, particularly against Candida albicans and Vibrio anguillarum. Pseudoalteromonas, Microbacterium, and Proteus were the most bioactive genera. After this preliminary screening, the bioactive strains were further evaluated in liquid assays against C. albicans, Bacillus subtilis and Escherichia coli. Filtered culture medium and acetone extracts from three and 5 days-old cultures were assayed. High antifungal activity against C. albicans in both aqueous and acetone extracts as well as absence of activity against B. subtilis were confirmed. Higher levels of activity were obtained with the aqueous extracts when compared to the acetone extracts and differences were also observed between the 3 and 5 day-old extracts. Furthermore, a low number of active strains was observed against E. coli. Potential presence of type-I polyketide synthases (PKS-I) and non-ribosomal peptide synthetases (NRPSs) genes were detected in 17 and 30 isolates, respectively. The high levels of bioactivity and the likely presence of associated genes suggest that Erylus deficiens bacteria are potential sources of novel marine bioactive compounds. PMID:25999928

The green synthesis, characterization, and evaluation of the biological activities of silver nanoparticles synthesized from Leptadenia reticulata leaf extract

NASA Astrophysics Data System (ADS)

Kumara Swamy, M.; Sudipta, K. M.; Jayanta, K.; Balasubramanya, S.

2015-01-01

Biosynthesis of silver nanoparticles (Ag Nps) was carried out using methanol leaves extract of L. reticulata. Ag Nps were characterized based on the observations of UV-visible spectroscopy, transmission electron microscopy, and X-ray diffraction (XRD) analysis. These Ag Nps were tested for antimicrobial activity by agar well diffusion method against different pathogenic microorganisms and antioxidant activity was performed using DPPH assay. Further, the in vitro cytotoxic effects of Ag Nps were screened against HCT15 cancer cell line and viability of tumor cells was confirmed using MTT ((3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide, a yellow tetrazole)) assay. The nuclear condensation was studied using the propidium iodide-staining method. The color change from green to dark brown and the absorbance peak at about 420 nm indicated the formation of nanoparticles. XRD pattern showed characteristic peaks indexed to the crystalline planes (111), (200) and (220) of face-centered cubic silver. The nanoparticles were of spherical shape with varying sizes ranging from 50 to 70 nm. Biosynthesized Ag Nps showed potent antibacterial activity and effective radical scavenging activity. MTT assay revealed a dose-dependent decrease in cell viability. Microscopic observations showed distinct cellular morphological changes indicating unhealthy cells, whereas the control appeared normal. Increase in the number of propidium iodide positive cells were observed in maximum concentration. Methanolic leaf extract of L. reticulata acts as an excellent capping agent for the formation of silver nanoparticles and demonstrates immense biological activities. Hence, these Ag NPs can be used as antibacterial, antioxidant as well as cytotoxic agent in treating many medical complications.

Inhibition of quorum sensing regulated bacterial functions by plant essential oils with special reference to clove oil.

PubMed

Khan, M S A; Zahin, M; Hasan, S; Husain, F M; Ahmad, I

2009-09-01

To evaluate quorum sensing (QS) inhibitory activity of plant essential oils using strains of Chromobacterium violaceum (CV12472 and CVO26) and Pseudomonas aeruginosa (PAO1). Inhibition of QS-controlled violacein production in C. violaceum was assayed using disc diffusion and agar well diffusion method. Of the 21 essential oils, four oils showed varying levels of anti-QS activity. Syzygium aromaticum (Clove) oil showed promising anti-QS activity on both wild and mutant strains with zones of pigment inhibition 19 and 17 mm, respectively, followed by activity in cinnamon, lavender and peppermint oils. The effect of clove oil on the extent of violacein production was estimated photometrically and found to be concentration dependent. At sub-MICs of clove oil, 78.4% reduction in violacein production over control and up to 78% reduction in swarming motility in PAO1 over control were recorded. Gas chromatography-mass spectrometry analysis of clove oil indicated presence of many phytocompounds. Eugenol, the major constituent of clove oil could not exhibit anti-QS activity. Presence of anti-QS activity in clove oil and other essential oils has indicated new anti-infective activity. The identification of anti-QS phytoconstituents is needed to assess the mechanism of action against both C. violaceum and Ps. aeruginosa. Essential oils having new antipathogenic drugs principle because of its anti-QS activity might be important in reducing virulence and pathogenicity of drug-resistant bacteria in vivo.

Considering the antibacterial activity of Zataria multiflora Boiss essential oil treated with gamma-irradiation in vitro and in vivo systems

NASA Astrophysics Data System (ADS)

Faezeh, Fatema; Salome, Dini; Abolfazl, Dadkhah; Reza, Zolfaghari Mohammad

2015-01-01

The aim of the present study was to evaluate the antibacterial activities of essential oils (EOs) obtained from the aerial parts of Zataria multiflora Boiss against Bacillus cereus, Pseudomonas aeroginosa, Escherichia coli and Staphylococcus aureus by in vivo and in vitro methods. Also, the effects of gamma-irradiation (0, 10 and 25 kGy) as a new microbial decontamination on the antibacterial activities of Z. multiflora were examined. For this purpose, the collected herbs were exposed to radiation at doses of 0, 10 and 25 kGy following essential oil (EOs) extraction by steam distillation. Then, the in vitro antibacterial potency of the irradiated and non-irradiated oils was determined by using disc diffusion, agar well diffusion and MIC and MBC determination assays. The in vivo antibacterial activity was also studied in sepsis model induced by CLP surgery by Colony forming units (CFUs) determination. The results showed that the extracted oils were discovered to be effective against all the gram positive and gram negative pathogens in vitro system. In addition, the oil significantly diminished the increased CFU count observed in CLP group. Moreover, the irradiated samples were found to possess the antibacterial activities as the non-irradiated ones both in vitro and in vivo systems. These data indicated the potential use of gamma-irradiation as a safe technique for preservation of Z. multiflora as a medicinal plant with effective antibacterial activities.

Rifampicin resistance mutations in the 81 bp RRDR of rpoB gene in Mycobacterium tuberculosis clinical isolates using Xpert® MTB/RIF in Kampala, Uganda: a retrospective study.

PubMed

Mboowa, Gerald; Namaganda, Carolyn; Ssengooba, Willy

2014-09-04

Introduction of Xpert® MTB/RIF assay has revolutionalised the diagnosis of tuberculosis (TB) by simultaneously detecting the bacteria and resistance to rifampicin (rif), a surrogate marker for multi-drug resistant TB (MDR-TB) as well as one of the principal first-line anti-tuberculosis drugs. In general, rpoB mutations can be found in 96.1% of rif-resistant Mycobacterium tuberculosis (MTB) strains worldwide and these mutations usually are located in a region at the 507-533rd amino acid residuals (81 bp) in the MTB rpoB gene, which is referred to as Rifampicin-resistance-determining region (RRDR). In this study, we determined the frequency of MDR-TB in Kampala using Xpert® MTB/RIF in comparison with the agar proportion method using Middlebrook 7H11and further determined the frequency of probes for different rpoB gene mutations using Xpert® MTB/RIF assay in the 81 bp RRDR. A total of 1501 specimens received at Mycobacteriology laboratory, Makerere University for Xpert testing between May 2011 and May 2014 were analysed by Xpert® MTB/RIF assay. Specimens that were positive for both MTB and rifampicin resistance were further subjected to a complete first line anti-mycobacterial drug susceptibility testing using Middlebrook 7H11 agar proportion method (APM). Xpert® MTB/RIF assay detected 313 MTB positive specimens and out of which 12 specimens had both MTB and rifampicin- resistance conferred by four different rpoB gene mutations in the 81 bp-RRDR of MTB, further one (1/12), specimen was found to be rifampicin mono-resistant on APM while the 11 were found to be MDR-TB. Probes associated with the observed rif- resistance were as follows: E (7/12), B (3/12), A (1/12), D (1/12) and no rif-resistance was associated with probe C. No specimen yielded rif-resistance associated with more than one probe failure (mutation combinations). Probe D was associated with rifampicin mono-resistant. MDR-TB was at 3.5% in the studied population. Mutations associated with Probe E (58%) also known as codons 531and 533 are the commonest rpoB gene mutation identified by Xpert® MTB/RIF assay in this setting and mutations identified by probe E of the assay, turned out to be MDR-TB strains by agar proportion method antimicrobial susceptibility testing. No mutation was detected in the codon 522.

Three Strains of Pseudomonas fluorescens Exhibit Differential Toxicity Against Drosophila melanogaster

USDA-ARS?s Scientific Manuscript database

Three strains of Pseudomonas fluorescens were tested for toxicity to Drosophila melanogaster in an insect feeding assay. Insect eggs were placed on the surface of a non-nutritive agar plate supplemented with a food source that was non-inoculated or inoculated with P. fluorescens Pf0-1, SBW25, or Pf-...

Distribution and survival of Pseudomonas sp. on Italian ryegrass and Curly dock in Georgia

USDA-ARS?s Scientific Manuscript database

Yellow bud, caused by Pseudomonas sp. is an emerging bacterial disease of onion. Polymerase chain reaction (PCR) assay based on the coronafacate ligase (cfl) and HrpZ genes were used to detect initial suspected bacteria on weeds. Growth on an agar medium, ability to cause a hypersensitive response i...

Development of a rapid and simple immunochromatographic assay to identify Vibrio parahaemolyticus.

PubMed

Sakata, Junko; Kawatsu, Kentaro; Iwasaki, Tadashi; Kumeda, Yuko

2015-09-01

To rapidly and simply determine whether or not bacterial colonies growing on agar were Vibrio parahaemolyticus, we developed an immunochromatographic assay (VP-ICA) using two different monoclonal antibodies (designated mAb-VP34 and mAb-VP109) against the delta subunit of V. parahaemolyticus-F0F1 ATP synthase. The epitopes recognized by mAb-VP34 and mAb-VP109 were mapped to sequences of eight ((47)LLTSSFSA(54)) and six amino acid residues ((16)FDFAVD(21)), respectively. An amino acid sequence similarity search of the NCBI database using BLASTP showed that both epitopic amino acid sequences were present together only in V. parahaemolyticus. When 124 V. parahaemolyticus strains and 94 strains of 27 other Vibrio species or 35 non-Vibrio species were tested using the VP-ICA, the VP-ICA identified V. parahaemolyticus with 100% accuracy. The VP-ICA rapidly and simply identified the pathogen directly from a single agar colony within 30 min, indicating that VP-ICA will greatly reduce labor and time required to identify V. parahaemolyticus compared with conventional biochemical tests. Copyright © 2015. Published by Elsevier B.V.

Spaceflight exposure effects on transcription, activity, and localization of alcohol dehydrogenase in the roots of Arabidopsis thaliana.

PubMed Central

Porterfield, D M; Matthews, S W; Daugherty, C J; Musgrave, M E

1997-01-01

Although considerable research and speculation have been directed toward understanding a plant's perception of gravity and the resulting gravitropic responses, little is known about the role of gravity-dependent physical processes in normal physiological function. These studies were conducted to determine whether the roots of plants exposed to spaceflight conditions may be experiencing hypoxia. Arabidopsis thaliana (L.) Heynh. plants were grown in agar medium during 6 or 11 d of spaceflight exposure on shuttle missions STS-54 (CHROMEX-03) and STS-68 (CHROMEX-05), respectively. The analysis included measurement of agar redox potential and root alcohol dehydrogenase (ADH) activity, localization, and expression. ADH activity increased by 89% as a result of spaceflight exposure for both CHROMEX-03 and -05 experiments, and ADH RNase protection assays revealed a 136% increase in ADH mRNA. The increase in ADH activity associated with the spaceflight roots was realized by a 28% decrease in oxygen availability in a ground-based study; however, no reduction in redox potential was observed in measurements of the spaceflight bulk agar. Spaceflight exposure appears to effect a hypoxic response in the roots of agar-grown plants that may be caused by changes in gravity-mediated fluid and/or gas behavior. PMID:9085569

Spaceflight exposure effects on transcription, activity, and localization of alcohol dehydrogenase in the roots of Arabidopsis thaliana

NASA Technical Reports Server (NTRS)

Porterfield, D. M.; Matthews, S. W.; Daugherty, C. J.; Musgrave, M. E.

1997-01-01

Although considerable research and speculation have been directed toward understanding a plant's perception of gravity and the resulting gravitropic responses, little is known about the role of gravity-dependent physical processes in normal physiological function. These studies were conducted to determine whether the roots of plants exposed to spaceflight conditions may be experiencing hypoxia. Arabidopsis thaliana (L.) Heynh. plants were grown in agar medium during 6 or 11 d of spaceflight exposure on shuttle missions STS-54 (CHROMEX-03) and STS-68 (CHROMEX-05), respectively. The analysis included measurement of agar redox potential and root alcohol dehydrogenase (ADH) activity, localization, and expression. ADH activity increased by 89% as a result of spaceflight exposure for both CHROMEX-03 and -05 experiments, and ADH RNase protection assays revealed a 136% increase in ADH mRNA. The increase in ADH activity associated with the spaceflight roots was realized by a 28% decrease in oxygen availability in a ground-based study; however, no reduction in redox potential was observed in measurements of the spaceflight bulk agar. Spaceflight exposure appears to effect a hypoxic response in the roots of agar-grown plants that may be caused by changes in gravity-mediated fluid and/or gas behavior.

Nutrient chemotaxis suppression of a diffusive instability in bacterial colony dynamics

NASA Astrophysics Data System (ADS)

Arouh, Scott; Levine, Herbert

2000-07-01

Bacteria grown on a semisolid agar surface have been observed to form branching patterns as the colony envelope propagates outward. The fundamental cause of this instability relates to the need for limited nutrient to diffuse towards the colony. Here, we investigate the effect on this instability of allowing the bacteria to move chemotactically in response to the nutrient gradient. Our results show that this additional effect has a tendency to suppress the instability. Our calculations are done within the context of a simple ``cutoff'' model of colony dynamics, but presumably remain valid for more complex and hence more realistic approaches.

Diffusion of Antimicrobials Across Silicone Hydrogel Contact Lenses

PubMed Central

Zambelli, Alison M.; Brothers, Kimberly M.; Hunt, Kristin M.; Romanowski, Eric G.; Nau, Amy C.; Dhaliwal, Deepinder K.; Shanks, Robert M. Q.

2014-01-01

Objectives To measure the diffusion of topical preparations of moxifloxacin, amphotericin B (AmB), and polyhexamethylene biguanide (PHMB) through silicone hydrogel (SH) contact lenses in vitro. Methods Using an in vitro model, the diffusion of three antimicrobials through SH contact lenses was measured. Diffused compounds were measured using a spectrophotometer at set time points over a period of four hours. The amount of each diffused antimicrobial was determined by comparing the experimental value to a standard curve. A biological assay was performed to validate the contact lens diffusion assay by testing antimicrobial activity of diffused material against lawns of susceptible bacteria (Staphylococcus epidermidis) and yeast (Saccharomyces cerevisiae). Experiments were repeated at least two times with a total of at least 4 independent replicates. Results Our data show detectable moxifloxacin and PHMB diffusion through SH contact lenses at 30 minutes, while amphotericin B diffusion remained below the limit of detection within the 4 hour experimental period. In the biological assay, diffused moxifloxacin demonstrated microbial killing starting at 20 minutes on bacterial lawns, whereas PHMB and amphotericin B failed to demonstrate killing on microbial lawns over the course of the 60 minute experiment. Conclusions In vitro diffusion assays demonstrate limited penetration of certain anti-infective agents through silicone hydrogel contact lenses. Further studies regarding the clinical benefit of using these agents along with bandage contact lens use for corneal pathology are warranted. PMID:25806673

Antimicrobial activity of blended essential oil preparation.

PubMed

Tadtong, Sarin; Suppawat, Supatcha; Tintawee, Anchalee; Saramas, Phanida; Jareonvong, Suchada; Hongratanaworakit, Tapanee

2012-10-01

Antimicrobial activities of two blended essential oil preparations comprising lavender oil, petigrain oil, clary sage oil, ylang ylang oil and jasmine oil were evaluated against various pathogenic microorganisms. Both preparations showed antimicrobial activity in the agar disc diffusion assay against the Gram-positive bacteria, Staphylococcus aureus ATCC6538 and S. epidermidis isolated strain, the fungus, Candida albicans ATCC10231, and the Gram-negative bacterium, Escherichia coli ATCC25922, but showed no activity against Pseudomonas aeruginosa ATCC9027. The minimum inhibitory concentration (MIC) of these preparations was evaluated. By the broth microdilution assay, preparation 1, comprising lavender oil, clary sage oil, and ylang ylang oil (volume ratio 3:4:3), exhibited stronger antimicrobial activity than preparation 2, which was composed of petigrain oil, clary sage oil, and jasmine oil (volume ratio 3:4:3). Moreover, the sum of the fractional inhibitory concentrations (Sigma fic) of preparation 1 expressed a synergistic antimicrobial effect against the tested microorganisms (Sigma fic

Biocompatibility and in vivo degradation of chitosan based hydrogels as potential drug carrier.

PubMed

Su, Feng; Wang, Yuandou; Liu, Xue; Shen, Xin; Zhang, Xingjian; Xing, Quansheng; Wang, Lihong; Chen, Yangsheng

2018-06-07

Carboxymethyl chitosan-graft-polylactide (CMCS-PLA) and carboxymethyl chitosan (CMCS) hydrogels were prepared by using 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride/N-hydroxysuccinimide (EDC/NHS) as crosslinking agent and catalyst at room temperature. The biocompatibility of the hydrogels was evaluated with the aim of assessing their potential as drug carrier. Various aspects of biocompatibility were considered, including MTT assay, agar diffusion test, release of lactate dehydrogenase (LDH), hemolytic test, plasma recalcification time (PRT), and dynamic clotting time. MTT assay showed that the cytotoxicity level of both hydrogels to L-929 cells was 0 or 1. The LDH release of CMCS and CMCS-PLA was 26 and 29%, respectively, which is slightly higher than that of the negative control (21%) and much lower than that of the negative control (87%). The hemolysis ratio of CMCS and CMCS-PLA was 1.4 and 1.7%, respectively, suggesting outstanding anti-hemolysis properties of both materials. The PRT value of CMCS and CMCS-PLA was higher by 77 and 99% than the value of the positive control. All the results revealed that the hydrogels present good cytocompatibility and hemocompatibility in vitro. In vivo degradation and tissue compatibility were evaluated by subcutaneous injection in the dorsal area of rats. CMCS and CMCS-PLA hydrogels were completely degraded and the inflammatory response also completely disappeared around hydrogels after 19 days in vivo. It is thus concluded that hydrogels formed of CMCS and CMCS-PLA with outstanding biocompatibility are promising as potential drug carrier.

A survey of the antibacterial activity of some New Zealand honeys.

PubMed

Allen, K L; Molan, P C; Reid, G M

1991-12-01

To assess the variation in antibacterial activity of honey a survey was carried out on 345 samples of unpasteurized honey obtained from commercial apiarists throughout New Zealand. Most of the honeys were considered to be monofloral, from 26 different floral sources. The honeys were tested against Staphylococcus aureus in an agar well diffusion assay, with reference to phenol as a standard. Antibacterial activity was found to range from the equivalent of less than 2% (w/v) phenol to 58% (w/v) phenol, with a median of 13.6 and a standard deviation of 12.5. Neither the age of the honey samples nor whether they had been processed by the apiarist was associated with lower activity. However, the difference between floral sources in the antibacterial activity was very highly significant. Kanuka (Kunzea ericoides (A. Rich.) J. Thompson. Family: Myrtaceae), manuka (Leptospermum scoparium J. R. et G. Forst. Family: Myrtaceae), ling heather (Calluna vulgaris (L.) Hull. Family: Ericaceae) and kamahi (Weinmannia racemosa Linn. f. Family: Cunoniaceae) were shown to be sources likely to give honey with high antibacterial activity. When antibacterial activity was assayed with catalase added to remove hydrogen peroxide, most of the honeys showed no detectable antibacterial activity. Only manuka and vipers bugloss (Echium vulgare L. Family: Boraginaceae) honeys showed this type of activity in a significant proportion of the samples. The high antibacterial activity of manuka honey was in many cases due entirely to this non-peroxide component.

Determination of apalcillin and its metabolites in human body fluids by high-pressure liquid chromatography.

PubMed Central

Borner, K; Lode, H; Elvers, A

1982-01-01

We describe two methods for the quantitative analysis of apalcillin and its metabolites in serum and urine by reverse-phase high-pressure liquid chromatography (HPLC), a fast isocratic method for the parent drug, and a gradient method that allows the simultaneous assay of two metabolites. Serum was deproteinized with acetonitrile, and urine was diluted with buffer solution. The detection limit was about 0.5 micrograms/ml at a detection wavelength of 254 nm and 1.5 micrograms/ml at 310 nm. Within-batch precision (coefficient of variation) varied from 10.2 to 1.1% for concentrations of 7.8 and 185.3 micrograms/ml of serum, respectively. Recovery rates of 95.1 and 97.7% were found in spiked sera. Results obtained by HPLC correlated well with those from a standard microbiological assay (agar diffusion test); the resulting bivariate regression equation for serum was y-bioassay = 2.5 micrograms/ml + 0.992 X xHPLC, and that for urine was ybioassay = 12.0 micrograms/ml + 1.009 X xHPLC. At a detection wavelength of 315 nm, no interferences were observed in 10 healthy volunteers. Healthy subjects who were given 2 g of apalcillin intravenously excreted 18% of the parent drug within 24 h in the urine. Two inactive compounds were furthermore identified in urine as the isomeric forms of the penicilloic acids. Their excretion within 24 h amounted to 6.9 and 11.2% of the dose. PMID:6818901

Using the overlay assay to qualitatively measure bacterial production of and sensitivity to pneumococcal bacteriocins.

PubMed

Maricic, Natalie; Dawid, Suzanne

2014-09-30

Streptococcus pneumoniae colonizes the highly diverse polymicrobial community of the nasopharynx where it must compete with resident organisms. We have shown that bacterially produced antimicrobial peptides (bacteriocins) dictate the outcome of these competitive interactions. All fully-sequenced pneumococcal strains harbor a bacteriocin-like peptide (blp) locus. The blp locus encodes for a range of diverse bacteriocins and all of the highly conserved components needed for their regulation, processing, and secretion. The diversity of the bacteriocins found in the bacteriocin immunity region (BIR) of the locus is a major contributor of pneumococcal competition. Along with the bacteriocins, immunity genes are found in the BIR and are needed to protect the producer cell from the effects of its own bacteriocin. The overlay assay is a quick method for examining a large number of strains for competitive interactions mediated by bacteriocins. The overlay assay also allows for the characterization of bacteriocin-specific immunity, and detection of secreted quorum sensing peptides. The assay is performed by pre-inoculating an agar plate with a strain to be tested for bacteriocin production followed by application of a soft agar overlay containing a strain to be tested for bacteriocin sensitivity. A zone of clearance surrounding the stab indicates that the overlay strain is sensitive to the bacteriocins produced by the pre-inoculated strain. If no zone of clearance is observed, either the overlay strain is immune to the bacteriocins being produced or the pre-inoculated strain does not produce bacteriocins. To determine if the blp locus is functional in a given strain, the overlay assay can be adapted to evaluate for peptide pheromone secretion by the pre-inoculated strain. In this case, a series of four lacZ-reporter strains with different pheromone specificity are used in the overlay.

Development of a real-time TaqMan assay to detect mendocina sublineage Pseudomonas species in contaminated metalworking fluids.

PubMed

Saha, Ratul; Donofrio, Robert S; Bagley, Susan T

2010-08-01

A TaqMan quantitative real-time polymerase chain reaction (qPCR) assay was developed for the detection and enumeration of three Pseudomonas species belonging to the mendocina sublineage (P. oleovorans, P. pseudoalcaligenes, and P. oleovorans subsp. lubricantis) found in contaminated metalworking fluids (MWFs). These microbes are the primary colonizers and serve as indicator organisms of biodegradation of used MWFs. Molecular techniques such as qPCR are preferred for the detection of these microbes since they grow poorly on typical growth media such as R2A agar and Pseudomonas isolation agar (PIA). Traditional culturing techniques not only underestimate the actual distribution of these bacteria but are also time-consuming. The primer-probe pair developed from gyrase B (gyrB) sequences of the targeted bacteria was highly sensitive and specific for the three species. qPCR was performed with both whole cell and genomic DNA to confirm the specificity and sensitivity of the assay. The sensitivity of the assay was 10(1) colony forming units (CFU)/ml for whole cell and 13.7 fg with genomic DNA. The primer-probe pair was successful in determining concentrations from used MWF samples, indicating levels between 2.9 x 10(3) and 3.9 x 10(6) CFU/ml. In contrast, the total count of Pseudomonas sp. recovered on PIA was in the range of <1.0 x 10(1) to 1.4 x 10(5) CFU/ml for the same samples. Based on these results from the qPCR assay, the designed TaqMan primer-probe pair can be efficiently used for rapid (within 2 h) determination of the distribution of these species of Pseudomonas in contaminated MWFs.

Modeling Antimicrobial Activity of Clorox(R) Using an Agar-Diffusion Test: A New Twist On an Old Experiment.

ERIC Educational Resources Information Center

Mitchell, James K.; Carter, William E.

2000-01-01

Describes using a computer statistical software package called Minitab to model the sensitivity of several microbes to the disinfectant NaOCl (Clorox') using the Kirby-Bauer technique. Each group of students collects data from one microbe, conducts regression analyses, then chooses the best-fit model based on the highest r-values obtained.…

[Study on the antibacterial activity of four kinds of nano-hydroxyapatite composites against Enterococcus faecalis].

PubMed

Liu, Yi; Zhou, Rongjing; Wu, Hongkun

2015-06-01

This study aims to compare and determine a kind of nano-hydroxyapatite composite material with good antibacterial efficacy on Enterococcusfaecalis (E. faecalis) in vitro. We investigated the antimicrobial activity of four kinds of nano-hydroxyapatite composites, namely, silver/hydroxyapatite composite nanoparticles (Ag/nHA), yttrium/hydroxyapatite composite nanoparticles (Yi/nHA), cerium/hydroxyapatite composite nanoparticles (Ce/nHA), and hydroxyapatite nanoparticles (nHA), against E. faecalis in vitro using the agar diffusion and broth dilution method by measuring the growth inhibition zone and the minimum inhibitory concentration (MIC), respectively. The agar diffusion test results showed that Ag/nHA displayed an obvious growth inhibition zone, whereas Yi/nHA, Ce/nHA, and nHA showed no influence on E. faecalis. The MIC value of Ag/nHA was 1.0 g.L-1, and the three other materials had no effect on E.faecalis even at the high concentration of 32.0 g.L-1. Ag/nHA display a potential antimicrobial efficacy to planktonic E.faecalis. Whereas, the three other kinds of nano-hydroxyapatite composites (Yi/nHA, Ce/nHA, nHA) show no influence.

Antigenic analyses of tissues and excretory and secretory products from Strongylus vulgaris.

PubMed Central

Wynne, E; Slocombe, J O; Wilkie, B N

1981-01-01

Rabbit antisera were prepared against veronal buffered saline extracts of L4 and L5 Strongylus vulgaris, adult S. vulgaris and adult Strongylus equinus retrieved from naturally infected horses. In agar gel diffusion with these antisera, adult S vulgaris and S. equinus each appeared to have at least one unique antigen; larval S. vulgaris appeared to have two species-specific and two stage-specific antigens. There were several common antigens. Excretory and secretory products were collected also from L4 and L5 an maintained over several days in tissue culture fluid. In agar gel diffusion against the above rabbit antisera, a stage-specific antigen was found also in excretory and secretory products. In addition, excretory and secretory products had three antigens in common with adult and larval S. vulgaris, but only one of these was common to adult S. equinus. The excretory and secretory products appear, therefore, to have two species-specific and one stage-specific antigens. Images Fig. 1 a and b. Fig. 2 a and b. Fig. 3 a and b. Fig. 4 a and b. Fig. 5 a and b. Fig. 6 a and b. Fig. 7 a and b. Fig. 8 a and b. PMID:6804070

Characterization of the species Malassezia pachydermatis and re-evaluation of its lipid dependence using a synthetic agar medium

PubMed Central

Puig, Laura; Castellá, Gemma

2017-01-01

The genus Malassezia includes lipophilic yeasts, which are part of the skin microbiota of various mammals and birds. Unlike the rest of Malassezia species, M. pachydermatis is described as non-lipid-dependent, as it is able to grow on Sabouraud glucose agar (SGA) without lipid supplementation. In this study we have examined the phenotypic variability within M. pachydermatis and confirmed its lipid-dependent nature using a synthetic agar medium. We used a selection of representative non-lipid-dependent strains from different animal species and three atypical lipid-dependent strains of this species, which were not able to grow after multiple passages on SGA. More than 400 lipid-dependent Malassezia isolates from animals were studied in order to detect the three lipid-dependent strains of M. pachydermatis. The identity of the atypical strains was confirmed by DNA sequencing. On the other hand, we have modified the Tween diffusion test, which is widely used in the characterization of these yeasts, by using a synthetic agar-based medium instead of SGA. This modification has proved to be useful for differentiation of M. pachydermatis strains, providing reproducible results and a straightforward interpretation. The finding of these peculiar lipid-dependent strains exemplifies the large variability within the species M. pachydermatis, which involves rare atypical strains with particular growth requirements. PMID:28586389

Characterization of the species Malassezia pachydermatis and re-evaluation of its lipid dependence using a synthetic agar medium.

PubMed

Puig, Laura; Bragulat, M Rosa; Castellá, Gemma; Cabañes, F Javier

2017-01-01

The genus Malassezia includes lipophilic yeasts, which are part of the skin microbiota of various mammals and birds. Unlike the rest of Malassezia species, M. pachydermatis is described as non-lipid-dependent, as it is able to grow on Sabouraud glucose agar (SGA) without lipid supplementation. In this study we have examined the phenotypic variability within M. pachydermatis and confirmed its lipid-dependent nature using a synthetic agar medium. We used a selection of representative non-lipid-dependent strains from different animal species and three atypical lipid-dependent strains of this species, which were not able to grow after multiple passages on SGA. More than 400 lipid-dependent Malassezia isolates from animals were studied in order to detect the three lipid-dependent strains of M. pachydermatis. The identity of the atypical strains was confirmed by DNA sequencing. On the other hand, we have modified the Tween diffusion test, which is widely used in the characterization of these yeasts, by using a synthetic agar-based medium instead of SGA. This modification has proved to be useful for differentiation of M. pachydermatis strains, providing reproducible results and a straightforward interpretation. The finding of these peculiar lipid-dependent strains exemplifies the large variability within the species M. pachydermatis, which involves rare atypical strains with particular growth requirements.

Extracellular enzyme production and cheating in Pseudomonas fluorescens depend on diffusion rates

PubMed Central

Allison, Steven D.; Lu, Lucy; Kent, Alyssa G.; Martiny, Adam C.

2014-01-01

Bacteria produce extracellular enzymes to obtain resources from complex chemical substrates, but this strategy is vulnerable to cheating by cells that take up reaction products without paying the cost of enzyme production. We hypothesized that cheating would suppress enzyme production in co-cultures of cheater and producer bacteria, particularly under well-mixed conditions. To test this hypothesis, we monitored protease expression and frequencies of Pseudomonas fluorescens producer and cheater genotypes over time in mixed liquid cultures and on agar plates. In mixed culture inoculated with equal frequencies of cheaters and producers, enzyme concentration declined to zero after 20 days, consistent with our hypothesis. We observed a similar decline in cultures inoculated with producers only, suggesting that cheater mutants arose de novo and swept the population. DNA sequencing showed that genetic changes most likely occurred outside the protease operon. In one experimental replicate, the population regained the ability to produce protease, likely due to further genetic changes or population dynamics. Under spatially structured conditions on agar plates, cheaters did not sweep the population. Instead, we observed a significant increase in the variation of enzyme activity levels expressed by clones isolated from the population. Together these results suggest that restricted diffusion favors a diversity of enzyme production strategies. In contrast, well-mixed conditions favor population sweeps by cheater strains, consistent with theoretical predictions. Cheater and producer strategies likely coexist in natural environments with the frequency of cheating increasing with diffusion rate. PMID:24782855

RAPA: a novel in vitro method to evaluate anti-bacterial skin cleansing products.

PubMed

Ansari, S A; Gafur, R B; Jones, K; Espada, L A; Polefka, T G

2010-04-01

Development of efficacious anti-bacterial skin cleansing products has been limited by the availability of a pre-clinical (in vitro) method to predict clinical efficacy adequately. We report a simple and rapid method, designated as rapid agar plate assay (RAPA), that uses the bacteriological agar surface as a surrogate substrate for skin and combines elements of two widely used in vivo (clinical) methods (Agar Patch and Cup Scrub). To simulate the washing of the human hand or forearm skin with the test product, trypticase soy agar plates were directly washed with the test product and rinsed under running tap water. After air-drying the washed plates, test bacteria (Staphylococcus aureus or Escherichia coli) were applied and the plates were incubated at 37 degrees C for 18-24 h. Using S. aureus as the test organism, anti-bacterial bar soap containing triclocarbanilide showed a strong linear relationship (R(2) = 0.97) between bacterial dose and their per cent reduction. A similar dose-response relationship (R(2) = 0.96) was observed for anti-bacterial liquid hand soap against E. coli. RAPA was able to distinguish between anti-bacterial products based on the nature and level of actives in them. In limited comparative tests, results obtained by RAPA were comparable with the results obtained by clinical agar patch and clinical cup scrub methods. In conclusion, RAPA provides a simple, rugged and reproducible in vitro method for testing the relative efficacy of anti-bacterial skin cleansing products with a likelihood of comparable clinical efficacy. Further testing is warranted to improve the clinical predictability of this method.

Broad-range (pan) Salmonella and Salmonella serotype typhi-specific real-time PCR assays: potential tools for the clinical microbiologist.

PubMed

Farrell, John J; Doyle, Laura J; Addison, Rachel M; Reller, L Barth; Hall, Geraldine S; Procop, Gary W

2005-03-01

We describe broad-range salmonellae (ie, Salmonella) and Salmonella serotype Typhi-specific LightCycler (Roche Diagnostics, Indianapolis, IN) real-time polymerase chain reaction assays. We validated these with a battery of 280 bacteria, 108 of which were salmonellae representing 20 serotypes. In addition, 298 isolates from 170 clinical specimens that were suspected to possibly represent Salmonella were tested with the pan- Salmonella assay. Finally, the pan-Salmonella assay also was used to test DNA extracts from 101 archived, frozen stool specimens, 55 of which were culture-positive for salmonellae. Both assays were 100% sensitive and specific when cultured isolates of the battery were tested. The pan- Salmonella assay also characterized correctly all salmonellae on the primary isolation agar and was 96% sensitive (53/55) and 96% specific (49/51) when nucleic acid extracts from direct stool specimens were tested. These assays represent potential tools the clinical microbiologist could use to screen suspect isolates or stool specimens for Salmonella.

Diffusion processes in tumors: A nuclear medicine approach

NASA Astrophysics Data System (ADS)

Amaya, Helman

2016-07-01

The number of counts used in nuclear medicine imaging techniques, only provides physical information about the desintegration of the nucleus present in the the radiotracer molecules that were uptaken in a particular anatomical region, but that information is not a real metabolic information. For this reason a mathematical method was used to find a correlation between number of counts and 18F-FDG mass concentration. This correlation allows a better interpretation of the results obtained in the study of diffusive processes in an agar phantom, and based on it, an image from the PETCETIX DICOM sample image set from OsiriX-viewer software was processed. PET-CT gradient magnitude and Laplacian images could show direct information on diffusive processes for radiopharmaceuticals that enter into the cells by simple diffusion. In the case of the radiopharmaceutical 18F-FDG is necessary to include pharmacokinetic models, to make a correct interpretation of the gradient magnitude and Laplacian of counts images.

Use of culture- and ELISA-based toxin assay for detecting Clostridium Difficile, a neglected pathogen: A single-center study from a tertiary care setting.

PubMed

Lall, Sujata; Nataraj, Gita; Mehta, Preeti

2017-01-01

Clostridium difficile is a Gram-positive spore-bearing anaerobic bacillus increasingly associated with both community- and hospital-acquired colitis and diarrhea. It is the most common identifiable bacterial cause of healthcare-associated diarrhea associated with antibiotic use and one of the most common anaerobic infections. The diagnosis of C. difficile infection includes detection of toxin A/B in stool specimens by direct enzyme immunoassay, culture of pathogen from the stool specimens using a selective agar Cycloserine-Cefoxitin fructose agar (CCFA), tissue culture assay, and detection of glutamate dehydrogenase an enzyme produced by C. difficile. With few reports from India on this disease, the present study was planned to throw more light on the prevalence and utility of laboratory diagnostic methods for C. difficile-associated diarrhea (CDAD). After taking approval from the Ethics Committee, 150 patients with antibiotic-associated diarrhea were taken as a study group and fifty patients with exposure to antibiotics but who did not develop diarrhea were taken as controls. Stool specimen was processed for both culture on CCFA and toxin detection by IVD Tox A + B ELISA. Only four specimens were culture positive, whereas 13 were ELISA positive. All culture-positive isolates were toxigenic. C. difficile was neither isolated nor its toxin detected in the control group. Culture- and toxin-based assays may not detect all cases of CDAD. Based on the results of the present study, culture does not provide any additional yield over toxin assay. Better diagnostic modalities would be required to prove CDAD.

AUTORADIOGRAPHIC ANALYSIS ON AGAR PLATES OF ANTIGENS FROM SUB CELLULAR FRACTIONS OF RAT LIVER SLICES

PubMed Central

Morgan, W. S.; Perlmann, P.; Hultin, T.

1961-01-01

Slices of rat livers were incubated with 14C amino acids, homogenized, and subjected to differential centrifugation. The microsomes were further extracted with the non-ionic detergent Lubrol W and with EDTA. These extracts and the microsome free "cell sap," freed from the pH 5 precipitable fraction, were subsequently reacted with antisera using agar diffusion techniques. The antisera employed were obtained from rabbits injected with different subcellular fractions of rat liver or with rat serum proteins. When the agar diffusion plates were autoradiographed it was found that some of the precipitates were radioactive while others were not. Control experiments indicated that this labeling was due to the specific incorporation of 14C amino acids into various rat liver antigens during incubation of the slices rather than to a non-specific adsorption of radioactive material to the immunological precipitates. When the slices were incubated with the isotope for up to 30 minutes, the serum proteins which could be extracted from the microsomes with the detergent were strongly labeled, as were a number of additional microsomal antigens of unknown significance. In contrast, the serum proteins present in the cell sap were only weakly labeled. Most of the typical cell sap proteins, both those precipitable and those soluble at pH 5, seemed to remain unlabeled. No consistently reproducible results were obtained with the EDTA extracts of the ribosomal residues remaining after extraction of the microsomes with the detergent. Incubation of the liver slices for longer periods (up to 120 minutes) led to a strong labeling of the serum proteins in the cell sap as well as to the appearance of labeling in additional cell sap proteins. The results are discussed with regard to the subcellular site of synthesis and the metabolism of the different antigens. PMID:13772607

Biofilm and metallo beta-lactamase production among the strains of Pseudomonas aeruginosa and Acinetobacter spp. at a Tertiary Care Hospital in Kathmandu, Nepal.

PubMed

Baniya, Bandana; Pant, Narayan Dutt; Neupane, Sanjeev; Khatiwada, Saroj; Yadav, Uday Narayan; Bhandari, Nisha; Khadka, Rama; Bhatta, Sabita; Chaudhary, Raina

2017-11-02

Pseudomonas aeruginosa and Acinetobacter spp. are found to be associated with biofilm and metallo-β-lactamase production and are the common causes of serious infections mainly in hospitalized patients. So, the main aims of this study were to determine the rates of biofilm production and metallo beta-lactamase production (MBL) among the strains of Pseudomonas aeruginosa and Acinetobacter spp. isolated from hospitalized patients. A total of 85 P. aeruginosa isolates and 50 Acinetobacter spp. isolates isolated from different clinical specimens from patients admitted to Shree Birendra Hospital, Kathmandu, Nepal from July 2013 to May 2014 were included in this study. The bacterial isolates were identified with the help of biochemical tests. Modified Kirby-Bauer disc diffusion technique was used for antimicrobial susceptibility testing. Combined disc diffusion technique was used for the detection of MBL production, while Congo red agar method and tube adherence method were used for detection of biofilm production. Around 16.4% of P. aeruginosa isolates and 22% of the strains of Acinetobacter spp. were metallo β-lactamase producers. Out of 85 P. aeruginosa isolates, 23 (27.05%) were biofilm producers according to tube adherence test while, only 13 (15.29%) were biofilm producers as per Congo red agar method. Similarly, out of 50 Acinetobacter spp. 7 (14%) isolates were biofilm producers on the basis of tube adherence test, while only 5 (10%) were positive for biofilm production by Congo red agar method. Highest rates of susceptibility of P. aeruginosa as well as Acinetobacter spp. were seen toward colistin. In our study, biofilm production and metallo beta-lactamase production were observed among Pseudomonas aeruginosa and Acinetobacter spp. However, no statistically significant association could be established between biofilm production and metallo beta-lactamase production.

Antifungal activity of osthol in vitro and enhancement in vivo through Eudragit S100 nanocarriers

PubMed Central

Li, Lin-peng; Wang, Xiao-juan; Zhang, Jin-Yu; Zhang, Lu-lu; Cao, Yong-bing; Gu, Li-qun; Yu, Yi-qun; Yang, Qi-lian; Shen, Chun-ying; Han, Bing; Jiang, Yuan-ying

2018-01-01

ABSTRACT In vitro interaction of osthol (Ost) and fluconazole (FLC) was investigated against 11 fluconazole-resistant clinical isolates of Candida albicans. Synergistic activities were determined using the checkerboard microdilution assay. The results of agar diffusion test confirmed the synergistic interaction. We used an enteric material Eudragit S100 for preparation of Ost nanoparticle (Ost-NP) to improve the oral bioavailability, biological activity of Ost. The physicochemical characteristics of Ost-S100-NP revealed Ost-S100-NP with mean particle size of 55.4±0.4 nm, encapsulation efficiency of 98.95±0.06%, drug loading efficiency of 23.89±0.25%, yield of 98.5±0.1% and a polydispersity index (PDI) of 0.165. As the Ost concentration-time curve showed, Ost-S100-NP can increase the plasma concentration and relative bioavailability of Ost compared with Ost-suspension by oral administration. In vivo, Ost-S100-NP enhanced the therapeutic efficacy of Ost against FLC-resistant C. albicans in immunosuppressed candidiasis mice model. The available information strongly suggests that Ost-S100-NP may be used as a promising compound against drug-resistant fungi. PMID:28795862

1-(2-aminophenyl)-1H-1,2,3-triazole-4-carboxylic acid: activity against Gram-positive and Gram-negative pathogens including Vibrio cholerae

NASA Astrophysics Data System (ADS)

Maji, Krishnendu; Haldar, Debasish

2017-10-01

We report a new synthetic aromatic ε-amino acid containing a triazole moiety with antimicrobial potential against Gram-positive, Gram-negative and pathogenic bacteria including Vibrio cholerae. Structure-property relationship studies revealed that all the functional groups are essential to enhance the antimicrobial activity. The 1-(2-aminophenyl)-1H-1,2,3-triazole-4-carboxylic acid was synthesized by click chemistry. From X-ray crystallography, the amino acid adopts a kink-like structure where the phenyl and triazole rings are perpendicular to each other and the amine and acid groups maintain an angle of 60°. The agar diffusion test shows that the amino acid has significant antibacterial activity. The liquid culture test exhibits that the minimum inhibitory concentration (MIC) value for Bacillus subtilis and Vibrio cholerae is 59.5 µg ml-1. FE-SEM experiments were performed to study the morphological changes of bacterial shape after treatment with compound 1. The antimicrobial activity of the amino acid was further studied by DNA binding and degradation study, protein binding, dye-binding assay and morphological analysis. Moreover, the amino acid does not have any harmful effect on eukaryotes.

Synthesis and characterization of new complexes of nickel (II), palladium (II) and platinum(II) with derived sulfonamide ligand: Structure, DFT study, antibacterial and cytotoxicity activities

NASA Astrophysics Data System (ADS)

Bouchoucha, Afaf; Zaater, Sihem; Bouacida, Sofiane; Merazig, Hocine; Djabbar, Safia

2018-06-01

The synthesis, characterization and biological study of new nickel (II), palladium (II), and platinum (II) complexes with sulfamethoxazole ligand used in pharmaceutical field, were reported. [MLCl2].nH2O is the general formula obtained for Pd(II) and Pt(II) complexes. These complexes have been prepared and characterized by elemental analysis, FTIR, 1HNMR spectral, magnetic measurements, UV-Visible spectra, and conductivity. The DFT calculation was applied to optimize the geometric structure of the Pd(II) and Pt(II) complexes. A new single-crystal X-ray structure of the Ni(II) complex has been determined. It crystallized in monoclinic system with P 21/c space group and Z = 8. The invitro antibacterial activity of ligand and complexes against Escherichia coli, P. aeruginosa, Klebsiella pneumoniae, S. aureus, Bacillus subtilis species has been carried out and compared using agar-diffusion method. The Pd(II) and Pt(II) complexes showed a remarkable inhibition against bacteria tested. The invitro cytotoxicity assay of the complexes against three cell lines chronic myelogenous leukaemia (K562), human colon adenocarcinoma (HT-29) and breast cancer (MCF-7) was also reported.

TRAIL-CM4 fusion protein shows in vitro antibacterial activity and a stronger antitumor activity than solo TRAIL protein.

PubMed

Sang, Ming; Zhang, Jiaxin; Li, Bin; Chen, Yuqing

2016-06-01

A TRAIL-CM4 fusion protein in soluble form with tumor selective apoptosis and antibacterial functions was expressed in the Escherichia coli expression system and isolated through dialysis refolding and histidine-tag Nickel-affinity purification. Fresh Jurkat cells were treated with the TRAIL-CM4 fusion protein. Trypan blue staining and MTT analyses showed that, similar to a TRAIL positive control, Jurkat cell proliferation was significantly inhibited. Flow cytometry analyses using Annexin V-fluorescein revealed that Jurkat cells treated with the TRAIL-CM4 fusion protein exhibited increased apoptosis. Laser confocal microscopy showed that APB-CM4 and the fusion protein TRAIL-CM4 can bind to Jurkat cell membranes and initiate their destruction. ABP-CM4 enhances the antitumor activity of TRAIL by targeting and damaging the tumor cell membrane. In antibacterial experiments, agar well diffusion and bacterial growth inhibition curve assays revealed concentration-dependent TRAIL-CM4 antibacterial activity against Escherichia coli K12D31. The expressed TRAIL-CM4 fusion protein exhibited enhanced antitumor and antibacterial activities. Fusion protein expression allowed the two different proteins to function in combination. Copyright © 2016 Elsevier Inc. All rights reserved.

Analysis of phytochemical profile of Terminalia arjuna bark extract with antioxidative and antimicrobial properties

PubMed Central

Mandal, Shreya; Patra, Arpita; Samanta, Animesh; Roy, Suchismita; Mandal, Arpita; Mahapatra, Tapasi Das; Pradhan, Shrabani; Das, Koushik; Nandi, Dilip Kumar

2013-01-01

Objective To investigate phytochemical screening, antimicrobial activity and qualitative thin layer chromatographic separation of flavonoid components, antioxidant activity and total flavonoid compound of Terminalia arjuna. Methods For phytochemical screening, some common and available standard tests were done. Antimicrobial bioassay was done through agar well diffusion method. Detection of antioxidant activity and flavonoid compounds were done through thin layer chromatography. Total antioxidant activity was measured by 2, 2-diphenyl-1-picrylhydrazyl (DPPH) in colorimetric method. Aluminum chloride colorimetric method was used for total flavonoid determination. Results Phytochemical screening showed the active compounds presence in high concentration, such as phytosterol, lactones, flavonoids, phenolic compounds and tannins and glycosides. The antimicrobial activity of extract showed that greater inhibition zone against Gram negative bacteria than Gram positive bacteria. This methanolic extract showed a promising antioxidant activity, as absorption of DPPH redicles decreased in DPPH free radical scavenging assay. Flavonoids components having antioxidant property present in the methanol extract at a level of 199.00 mg quercetin equivalent/g of dried methanol extract in colorimetric method. Conclusions The Terminalia arjuna bark extract revealed the presence of bio-active constituents which are known to exhibit medicinal as well as physiological activities. PMID:24093787

Antimicrobial activity of natural products against Clostridium difficile in vitro.

PubMed

Roshan, N; Riley, T V; Hammer, K A

2017-05-10

To investigate the antimicrobial activity of various natural products against Clostridium difficile in vitro. The antibacterial activity of 20 natural products was determined by the agar well diffusion and broth microdilution assays against four C. difficile strains, three comparator organisms and four gastrointestinal commensal organisms. Of the raw natural products, garlic juice had the highest activity. The most active processed products were peppermint oil and the four pure compounds trans-cinnamaldehyde, allicin, menthol and zingerone. Furthermore, Bacteroides species had similar susceptibility to C. difficile to most natural products; however, Lactobacillus casei was less susceptible. The combined effect of natural products with vancomycin or metronidazole was determined using the conventional checkerboard titration method and the fractional inhibitory concentration index was calculated. The results showed a possible synergism between trans-cinnamaldehyde and vancomycin and partial synergy between trans-cinnamaldehyde and metronidazole. The study indicates a range of antimicrobial activity of natural products against C. difficile and suggests that they may be useful as alternative or complementary treatments for C. difficile infection (CDI), particularly as most are able to be given orally. This study encourages further investigation of natural products for treatment of CDI. © 2017 The Society for Applied Microbiology.

Aspergillus serology: Have we arrived yet?

PubMed

Richardson, Malcolm D; Page, Iain D

2017-01-01

Aspergillosis presents in various clinical forms, among them chronic pulmonary aspergillosis, which is a spectrum of disease entities including aspergilloma, chronic cavitary pulmonary aspergillosis, and chronic fibrosing pulmonary aspergillosis. Aspergillus also contributes to fungal allergy and sensitization. Analysis of the immune response to Aspergillus and its antigens is an integral part of the diagnosis of these diseases. Over the past half century, the techniques used to determine antibody titers have evolved from testing for precipitating and agglutinating antibodies by agar gel double diffusion and immunolectrophoresis to enzyme-linked immunosorbent assays using recombinant proteins as capture antigens. A resurgence of interest in the detection of immunoglobulins, primarily Aspergillus-specific IgG, has hinted at the possibility of distinguishing between colonization and invasion in immunocompromised patients with invasive aspergillosis. Even though there appears to be a greater degree of discrimination between the clinical forms of aspergillosis there is still a long way to travel. This review presents illustrative examples of where new diagnostic platforms and technologies have been applied to this intriguing spectrum of diseases. © The Authors 2016. Published by Oxford University Press on behalf of The International Society for Human and Animal Mycology. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Comparative evaluation of antimicrobial efficacy of various root canal filling materials along with aloevera used in primary teeth: a microbiological study.

PubMed

Kriplani, R; Thosar, N; Baliga, M S; Kulkarni, P; Shah, N; Yeluri, R

2013-01-01

this study was conducted to evaluate the antimicrobial effectiveness of 6 root canal filling materials and a negative control agent against 18 strains of bacteria isolated from infected root canals of primary molar teeth using agar diffusion assay. Aloevera with sterile water Zinc oxide and Eugenol, Zinc oxide-Eugenol with aloevera, Calcium hydroxide and sterile water, Calcium hydroxide with sterile water and aloevera, Calcium hydroxide and Iodoform (Metapex) and Vaseline (Control). MIC and MBC of aloevera was calculated. All materials except Vaseline showed varied antimicrobial activity against the test bacterias. The zones of inhibition were ranked into 4 inhibition categories based on the proportional distribution of the data. All the 18 bacterial isolates were classified under 2 groups based on Gram positive and Gram negative aerobes. Statistical analysis was carried out to compare the antimicrobial effectiveness between materials tested with each of the bacterial groupings. Aloevera + Sterile Water was found to have superior antimicrobial activity against most of the microorganisms followed by ZOE + Aloevera, calcium hydroxide + Aloevera, ZOE, calcium hydroxide, Metapex in the descending order and Vaseline showed no inhibition.

In vitro synergistic antibacterial activity of the essential oil from Zingiber cassumunar Roxb against extensively drug-resistant Acinetobacter baumannii strains.

PubMed

Boonyanugomol, Wongwarut; Kraisriwattana, Kairin; Rukseree, Kamolchanok; Boonsam, Kraisorn; Narachai, Panchaporn

In this study, we determined the antibacterial and synergistic activities of the essential oil from Zingiber cassumunar against the extensively drug-resistant (XDR) Acinetobacter baumannii strains. The antibacterial and synergistic properties of the essential oil from Z. cassumunar were examined by agar disc diffusion tests. The minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) were evaluated by broth microdilution using the resazurin assay. The in vitro time-kill antibacterial kinetics was analyzed using the plate count technique. We found that the essential oil from Z. cassumunar had antibacterial activity against A. baumannii, with MIC and MBC ranging from 7.00 to 9.24mg/ml. The essential oil could completely inhibit A. baumannii at 1h, and coccoid-shaped bacteria were found after treatment. In addition, the essential oil had a synergistic effect when combined with antibiotics, e.g., aminoglycosides, fluoroquinolones, tetracyclines, and folate pathway inhibitors. Thus, the essential oil from Z. cassumunar has strong antibacterial and synergistic activities against XDR A. baumannii, which may provide the basis for the development of a new therapy against drug-resistant bacteria. Copyright © 2017 The Authors. Published by Elsevier Ltd.. All rights reserved.

Intestinal ameliorative effects of traditional Ogi-tutu, Vernonia amygdalina and Psidium guajava in mice infected with Vibrio cholera.

PubMed

Shittu, Olufunke B; Ajayi, Olusola L; Bankole, Samuel O; Popoola, Temitope Os

2016-06-01

Cholera, a severe acute watery diarrhea caused by Vibrio cholerae is endemic in Nigeria with most cases occurring in the rural areas. In South West Nigeria, some individuals resort to alternative treatments such as Ogi-tutu, Psidium guajava and Vernonia amygdalina during infections. The effectiveness of these alternatives in the prevention and treatment of V. cholerae infection requires experimental investigation. This study was designed to investigate the ameliorative effects of Ogi-tutu, Vernonia amygdalina and Psidium guajava on intestinal histopathology of experimental mice infected with V. cholerae. Preliminary investigation of in vitro vibriocidal activities of these alternatives were carried out using agar cup diffusion assay. For ameliorative effects, adult mice were inoculated with 100 µl (106 cells) of Vibrio cholerae and dosed at 0 h (immediate prevention) and 4 h (treatment of infection) and their intestines were histopathologically evaluated. The histopathological changes were the same irrespective of the treated groups, but the lesions varied in extent and severity. The ameliorative effects in decreasing order were V. amygdalina > P. guajava > Ogi-tutu. V. amygdalina gave the best ameliorative effects in the prevention and treatment of V. cholerae infection.

Chemical composition, antibacterial and antioxidant activities of the essential oils from Thymus satureioides and Thymus pallidus.

PubMed

Ichrak, Ghalbane; Rim, Belaqziz; Loubna, Ait Said; Khalid, Oufdou; Abderrahmane, Romane; Said, El Messoussi

2011-10-01

This study was designed to examine the in vitro antibacterial and antioxidant activities of the essential oils (EOs) of Thymus satureioides (T.s) and T. pallidus (T.p). EOs were isolated by steam distillation and analyzed by capillary gas chromatography and gas chromatography coupled to mass spectrometry (GC-MS). The major constituents of the volatile fraction of T. satureioides were bomeol (29.5%), carvacrol (9.1%), and beta-caryophyllene (8.2%), while those of T. pallidus were camphor (29.8%), dihydrocarvone (17.6%), bomeol (7.6%) and camphene (7.5%). The essential oils were tested against a panel of Gram+ and Gram- bacteria by using agar diffusion and broth dilution methods. The data indicated that the Gram-positive Bacillus subtilis was the most sensitive strain producing an average inhibition zone of 51.7 mm. Furthermore, Pseudomonas aeruginosa, known as a resistant strain, was also sensitive. The samples were also subjected to screening for their possible antioxidant activity by using the 2,2-diphenyl-l-picrylhydrazyl (DPPH) assay. The IC50 values of the oil of T. satureioides and T. pallidus were 0.32 and 11.6 mg/mL, respectively.

Optimization and evaluation of clarithromycin floating tablets using experimental mixture design.

PubMed

Uğurlu, Timucin; Karaçiçek, Uğur; Rayaman, Erkan

2014-01-01

The purpose of the study was to prepare and evaluate clarithromycin (CLA) floating tablets using experimental mixture design for treatment of Helicobacter pylori provided by prolonged gastric residence time and controlled plasma level. Ten different formulations were generated based on different molecular weight of hypromellose (HPMC K100, K4M, K15M) by using simplex lattice design (a sub-class of mixture design) with Minitab 16 software. Sodium bicarbonate and anhydrous citric acid were used as gas generating agents. Tablets were prepared by wet granulation technique. All of the process variables were fixed. Results of cumulative drug release at 8th h (CDR 8th) were statistically analyzed to get optimized formulation (OF). Optimized formulation, which gave floating lag time lower than 15 s and total floating time more than 10 h, was analyzed and compared with target for CDR 8th (80%). A good agreement was shown between predicted and actual values of CDR 8th with a variation lower than 1%. The activity of clarithromycin contained optimizedformula against H. pylori were quantified using well diffusion agar assay. Diameters of inhibition zones vs. log10 clarithromycin concentrations were plotted in order to obtain a standard curve and clarithromycin activity.

Aspects of the antimicrobial efficacy of grapefruit seed extract and its relation to preservative substances contained.

PubMed

von Woedtke, T; Schlüter, B; Pflegel, P; Lindequist, U; Jülich, W D

1999-06-01

The antimicrobial efficacy as well as the content of preservative agents of six commercially available grapefruit seed extracts were examined. Five of the six extracts showed a high growth inhibiting activity against the test germs Bacillus subtilis SBUG 14, Micrococcus flavus SBUG 16, Staphylococcus aureus SBUG 11, Serratia marcescens SBUG 9, Escherichia coli SBUG 17, Proteus mirabilis SBUG 47, and Candida maltosa SBUG 700. In all of the antimicrobial active grapefruit seed extracts, the preservative benzethonium chloride was detected by thin layer chromatography. Additionally, three extracts contained the preserving substances triclosan and methyl parabene. In only one of the grapefruit seed extracts tested no preservative agent was found. However, with this extract as well as with several self-made extracts from seed and juiceless pulp of grapefruits (Citrus paradisi) no antimicrobial activity could be detected (standard serial broth dilution assay, agar diffusion test). Thus, it is concluded that the potent as well as nearly universal antimicrobial activity being attributed to grapefruit seed extract is merely due to the synthetic preservative agents contained within. Natural products with antimicrobial activity do not appear to be present.

Microbial susceptibility to calcium hydroxide pastes and their vehicles.

PubMed

Gomes, Brenda Paula Figueiredo de Almeida; Ferraz, Caio Cezar Randi; Garrido, Fabio Devora; Rosalen, Pedro Luiz; Zaia, Alexandre Augusto; Teixeira, Fabricio Batista; de Souza-Filho, Francisco José

2002-11-01

The aim of this study was to investigate the susceptibility of some microorganisms commonly isolated from root canals to calcium hydroxide in combination with several vehicles by the agar diffusion method. Stainless-steel cylinders were placed on each inoculated agar medium. The test medications and their controls were placed inside the cylinders. The zones of growth inhibition were measured and recorded after the incubation period for each plate, and the results were analyzed statistically. Enterococcus faecalis was most resistant, whereas the anaerobic Porphyromonas endodontalis was more susceptible to all medications, followed by P. gingivalis and Prevotella intermedial intermedia. Ca(OH)2 + CMCP + glycerin showed significantly larger mean zones of inhibition when compared with the other medications. We conclude that anaerobic Gram-negative bacteria are more susceptible to calcium hydroxide pastes than facultative Gram-positive microorganisms.

Behavioral differences of Heterodera glycines and Meloidogyne incognita infective juveniles exposed to root extracts in vitro

USDA-ARS?s Scientific Manuscript database

The in vitro behaviors of infective juveniles (J2) of Heterodera glycines and Meloidogyne incognita were compared in the presence and absence of plant root extracts. In an agar plate attraction-retention assay, H. glycines was 15-fold more responsive to a chemical attractant (CaCl2; P < 0.05) than w...

Interactions between stream fungi and bacteria associated with decomposing leaf litter at different levels of nutrient availability

Treesearch

Vladislav Gulis; Keller Suberkropp

2003-01-01

We examined the potential for interactions between aquatic hyphomycetes and bacteria isolated from leaves decaying in a headwater stream. In agar plate assays, culture filtrates of each of 28 aquatic hyphomycete isolates tested (5 species) inhibited bacterial growth (16 Gram-negative bacterial isolates belonging to 6 colony morphotypes were tested). Inhibition of...

Evaluation of Antibacterial Activity of Three Selected Fruit Juices on Clinical Endodontic Bacterial Strains

PubMed Central

Behera, Subasish; Khetrapal, Prashant; Punia, Sandhya Kapoor; Agrawal, Deepak; Khandelwal, Minal; Lohar, Jitendra

2017-01-01

Introduction: The increasing problem of antibiotic drug resistance by pathogenic microorganisms in the past few decades has recently led to the continuous exploration of natural plant products for new antibiotic agents. Many consumable food materials have good as well as their bad effects, good effect includes their antibacterial effects on different microorganisms present in the oral cavity. Recently, natural products have been evaluated as source of antimicrobial agent with efficacies against a variety of microorganisms. Methodology: The present study describes the antibacterial activity of three selected fruit juices (Apple, Pomegranate and Grape) on endodontic bacterial strains. Antimicrobial activity of fruit juices were tested by wel l diffusion assay by an inhibition zone surrounding the well. The aim of the study was to evaluate the antibacterial activity of three fruit juises on different endodontic strains. Result: Agar well diffusion method was adopted for determining antibacterial potency. Antibacterial activity present on the plates was indicated by an inhibition zone surrounding the well containing the fruit juice. The zone of inhibition was measured by measuring scale in millimeter. Comparision between antibacterial efficacy of all three fruit juices against Enterococcus feacalis and Streptococcus mutans was observed with significant value of P ≤ 0.05. Conclusion: The results obtained in this study clearly demonstrated a significant antimicrobial effect of apple fruit juice against Enterococcus fecalis and Streptococcus mutans. However, preclinical and clinical trials are needed to evaluate biocompatibility & safety before apple can conclusively be recommended in endodontic therapy, but in vitro observation of apple effectiveness appears promising. PMID:29284967

Bacteriocin from Bacillus subtilis as a novel drug against diabetic foot ulcer bacterial pathogens

PubMed Central

Joseph, Baby; Dhas, Berlina; Hena, Vimalin; Raj, Justin

2013-01-01

Objective To isolate and identify Bacillus subtilis (B. subtilis) from soil and to characterize and partially purify the bacteriocin. To evaluate the antimicrobial activity against four diabetic foot ulcer bacterial pathogens. Methods Genotypic identification was done based on Bergey's manual of systemic bacteriology. Antimicrobial susceptibility test was done by Kirby-Bauer disc diffusion method. Colonies were identified by colony morphology and biochemical characterization and also compared with MTCC 121 strain. Further identification was done by 16S rRNA sequencing. Inhibitory activities of partially purified bacteriocin on all the DFU isolates were done by agar well diffusion method. The strain was identified to produce bacteriocin by stab overlay assay. Bacteriocin was extracted by organic solvent extraction using chloroform, further purified by HPLC and physical, and chemical characterization was performed. Results The four isolates showed high level of resistance to amoxyclav and sensitivity to ciprofloxacin. HPLC purification revealed that the extracts are bacteriocin. The phylogenetic tree analysis results showed that the isolate was 99% related to B. subtilis BSF01. The results reveled activity to all the four isolates and high level of activity was seen in case of Klebsiella sp. Conclusions Partially purified bacteriocin was found to have antimicrobial activity against the four diabetic foot ulcer bacterial pathogens, which can thus be applied as a better drug molecule on further studies. The strain B. subtilis are found to be safe for use and these antimicrobial peptides can be used as an antimicrobial in humans to treat DFU bacterial pathogens. PMID:24093784

Bacteriocin from Bacillus subtilis as a novel drug against diabetic foot ulcer bacterial pathogens.

PubMed

Joseph, Baby; Dhas, Berlina; Hena, Vimalin; Raj, Justin

2013-12-01

To isolate and identify Bacillus subtilis (B. subtilis) from soil and to characterize and partially purify the bacteriocin. To evaluate the antimicrobial activity against four diabetic foot ulcer bacterial pathogens. Genotypic identification was done based on Bergey's manual of systemic bacteriology. Antimicrobial susceptibility test was done by Kirby-Bauer disc diffusion method. Colonies were identified by colony morphology and biochemical characterization and also compared with MTCC 121 strain. Further identification was done by 16S rRNA sequencing. Inhibitory activities of partially purified bacteriocin on all the DFU isolates were done by agar well diffusion method. The strain was identified to produce bacteriocin by stab overlay assay. Bacteriocin was extracted by organic solvent extraction using chloroform, further purified by HPLC and physical, and chemical characterization was performed. The four isolates showed high level of resistance to amoxyclav and sensitivity to ciprofloxacin. HPLC purification revealed that the extracts are bacteriocin. The phylogenetic tree analysis results showed that the isolate was 99% related to B. subtilis BSF01. The results reveled activity to all the four isolates and high level of activity was seen in case of Klebsiella sp. Partially purified bacteriocin was found to have antimicrobial activity against the four diabetic foot ulcer bacterial pathogens, which can thus be applied as a better drug molecule on further studies. The strain B. subtilis are found to be safe for use and these antimicrobial peptides can be used as an antimicrobial in humans to treat DFU bacterial pathogens. Copyright © 2013 Asian Pacific Tropical Biomedical Magazine. Published by Elsevier B.V. All rights reserved.

Visualizing repetitive diffusion activity of double-strand RNA binding proteins by single molecule fluorescence assays.

PubMed

Koh, Hye Ran; Wang, Xinlei; Myong, Sua

2016-08-01

TRBP, one of double strand RNA binding proteins (dsRBPs), is an essential cofactor of Dicer in the RNA interference pathway. Previously we reported that TRBP exhibits repetitive diffusion activity on double strand (ds)RNA in an ATP independent manner. In the TRBP-Dicer complex, the diffusion mobility of TRBP facilitates Dicer-mediated RNA cleavage. Such repetitive diffusion of dsRBPs on a nucleic acid at the nanometer scale can be appropriately captured by several single molecule detection techniques. Here, we provide a step-by-step guide to four different single molecule fluorescence assays by which the diffusion activity of dsRBPs on dsRNA can be detected. One color assay, termed protein induced fluorescence enhancement enables detection of unlabeled protein binding and diffusion on a singly labeled RNA. Two-color Fluorescence Resonance Energy Transfer (FRET) in which labeled dsRBPs is applied to labeled RNA, allows for probing the motion of protein along the RNA axis. Three color FRET reports on the diffusion movement of dsRBPs from one to the other end of RNA. The single molecule pull down assay provides an opportunity to collect dsRBPs from mammalian cells and examine the protein-RNA interaction at single molecule platform. Copyright © 2016 Elsevier Inc. All rights reserved.

Soft agar-based selection of spontaneously transformed rat prostate epithelial cells with highly tumorigenic characteristics.

PubMed

Gajdošik, Martina Šrajer; Hixson, Douglas C; Brilliant, Kate E; Yang, DongQin; De Paepe, Monique E; Josić, Djuro; Mills, David R

2018-05-29

The critical molecular and cellular mechanisms involved in the development and progression of prostate cancer remain elusive. In this report, we demonstrate that normal rat prostate epithelial cells (PEC) undergo spontaneous transformation at high passage (p > 85) evidenced by the acquisition of anchorage independent growth when plated on soft agar and tumorigenicity when injected into immunodeficient mice. In addition, we also report the discovery of a minor subpopulation of spontaneously transformed PEC derived from high passage PEC with the ability to migrate through a layer of 1% agar and form expanding colonies on the underlying plastic substratum. Comparison of these soft agar invasive (SAI) cells with low (p < 35), mid (p36-84) and high passage (p > 85) PEC identified marked differences in cell morphology, proliferation and motility. The SAI subpopulation was more tumorigenic than the high passage anchorage independent cultures from which they were isolated, as manifested by a decreased latency period and an increase in the size of tumors arising in immunodeficient mice. In contrast, low and mid passage cells were unable to grow on soft agar and failed to form tumors when injected into immunodeficient mice. Screening with antibody-based signaling arrays identified several differences in the altered expression levels of signaling proteins between SAI-derived cells and low or high passage PEC, including the up-regulation of EGFR and MAPK-related signaling pathways in SAI-selected cells. In summary, these studies suggest that the SAI assay selects for a novel, highly tumorigenic subpopulation of transformed cells that may represent an early step in the progression of slow growing prostatic carcinomas into more rapidly growing and aggressive tumors. Copyright © 2017. Published by Elsevier Inc.

Pharmacological and phytochemical screening of Palestinian traditional medicinal plants Erodium laciniatum and Lactuca orientalis.

PubMed

Jaradat, Nidal; AlMasri, Motasem; Zaid, Abdel Naser; Othman, Dua'a Ghazi

2017-09-01

Various epidemiological studies showed that herbal remedies containing polyphenols may protect against various diseases such as cancers, vascular diseases and inflammatory pathologies. Currently, such groups of bioactive compounds have become a subject of many antimicrobials and antioxidant investigations. Accordingly, the current study aimed to conduct biological and phytochemical screening for two Palestinian traditional medicinal plants, Erodium laciniatum and Lactuca orientalis. Current plants phytoconstituents and their antioxidant activities were evaluated by using standard phytochemical methods; meanwhile, antimicrobial activities were estimated by using several types of American Type Culture Collection and multidrug resistant clinical isolates by using agar diffusion well-variant, agar diffusion disc-variant and broth microdilution methods. Phytochemical screenings showed that L. orientalis and E. laciniatum contain mixtures of secondary and primary metabolites Moreover, total flavonoid, tannins and phenols content in E. laciniatum extract were higher than the L. orientalis extracts with almost the same antioxidant potentials. Additionally, both plants organic and aqueous extracts showed various potentials of antimicrobial activity Conclusions: Overall, the studied species have a mixture of phytochemicals, flavonoids, phenols and tannins also have antioxidant and antimicrobial activities which approved their folk uses in treatments of infectious and Alzheimer diseases and simultaneously can be used as therapeutic agents in the pharmaceutical industries.

Antimicrobial activity of ProRoot MTA in contact with blood

PubMed Central

Farrugia, C.; Baca, P.; Camilleri, J.; Arias Moliz, M. T.

2017-01-01

Dental materials based on Portland cement, which is used in the construction industry have gained popularity for clinical use due to their hydraulic properties, the interaction with tooth tissue and their antimicrobial properties. The antimicrobial properties are optimal in vitro. However in clinical use contact with blood may affect the antimicrobial properties. This study aims to assess whether antimicrobial properties of the Portland cement-based dental cements such as mineral trioxide aggregate (MTA) are also affected by contact with blood present in clinical situations. ProRoot MTA, a Portland cement-based dental cement was characterized following contact with water, or heparinized blood after 1 day and 7 days aging. The antimicrobial activity under the mentioned conditions was assessed using 3 antimicrobial tests: agar diffusion test, direct contact test and intratubular infection test. MTA in contact with blood was severely discoloured, exhibited an additional phosphorus peak in elemental analysis, no calcium hydroxide peaks and no areas of bacterial inhibition growth in the agar diffusion test were demonstrated. ProRoot MTA showed limited antimicrobial activity, in both the direct contact test and intratubular infection test. When aged in water ProRoot MTA showed higher antimicrobial activity than when aged in blood. Antimicrobial activity reduced significantly after 7 days. Further assessment is required to investigate behaviour in clinical situations. PMID:28128328

CHEMICAL CHARACTERIZATION AND EVALUATION OF ANTIBACTERIAL, ANTIFUNGAL, ANTIMYCOBACTERIAL, AND CYTOTOXIC ACTIVITIES OF Talinum paniculatum

PubMed Central

REIS, Luis F.C. DOS; CERDEIRA, Cláudio D.; PAULA, Bruno F. DE; da SILVA, Jeferson J.; COELHO, Luiz F.L.; SILVA, Marcelo A.; MARQUES, Vanessa B.B.; CHAVASCO, Jorge K.; ALVES-DA-SILVA, Geraldo

2015-01-01

SUMMARY In this study, the bioactivity of Talinum paniculatum was evaluated, a plant widely used in folk medicine. The extract from the T. paniculatum leaves (LE) was obtained by percolation with ethanol-water and then subjecting it to liquid-liquid partitions, yielding hexane (HX), ethyl acetate (EtOAc), butanol (BuOH), and aqueous (Aq) fractions. Screening for antimicrobial activity of the LE and its fractions was evaluated in vitro through broth microdilution method, against thirteen pathogenic and non-pathogenic microorganisms, and the antimycobacterial activity was performed through agar diffusion assay. The cytotoxic concentrations (CC90) for LE, HX, and EtOAc were obtained on BHK-21 cells by using MTT reduction assay. The LE showed activity against Serratia marcescens and Staphylococcus aureus, with Minimum Inhibitory Concentration (MIC) values of 250 and 500 µg/mL, respectively. Furthermore, HX demonstrated outstanding activity against Micrococcus luteusand Candida albicans with a MIC of 31.2 µg/mL in both cases. The MIC for EtOAc also was 31.2 µg/mL against Escherichia coli. Conversely, BuOH and Aq were inactive against all tested microorganisms and LE proved inactive against Mycobacterium tuberculosis and Mycobacterium bovis as well. Campesterol, stigmasterol, and sitosterol were the proposed structures as main compounds present in the EF and HX/EtOAc fractions, evidenced by mass spectrometry. Therefore, LE, HX, and EtOAc from T. paniculatum showed potential as possible sources of antimicrobial compounds, mainly HX, for presenting low toxicity on BHK-21 cells with excellent Selectivity Index (SI = CC90/MIC) of 17.72 against C. albicans. PMID:26603226

Biogenic synthesis of silver nanoparticles using guava ( Psidium guajava) leaf extract and its antibacterial activity against Pseudomonas aeruginosa

NASA Astrophysics Data System (ADS)

Bose, Debadin; Chatterjee, Someswar

2016-08-01

Among the various inorganic nanoparticles, silver nanoparticles have received substantial attention in the field of antimicrobial research. For safe and biocompatible use of silver nanoparticles in antimicrobial research, the different biogenic routes are developed to synthesize silver nanoparticles that do not use toxic chemicals. Among those, to synthesize silver nanoparticles, the use of plant part extract becomes an emerging field because plant part acts as reducing as well as capping agent. For large-scale production of antibacterial silver nanoparticles using plant part, the synthesis route should be very simple, rapid, cost-effective and environment friendly based on easy availability and non-toxic nature of plant, stability and antibacterial potential of biosynthesized nanoparticles. In the present study, we report a very simple, rapid, cost-effective and environment friendly route for green synthesis of silver nanoparticles using guava ( Psidium guajava) leaf extract as reducing as well as capping agent. This plant has been opted for the present study for its known medicinal properties, and it is easily available in all seasons and everywhere. The biosynthesized silver nanoparticles are characterized by UV-Vis and TEM analysis. The average particle size is 40 nm in the range of 10-90 nm. The antibacterial activity of these nanoparticles against Pseudomonas aeruginosa MTCC 741 has been measured by disc diffusion method, agar cup assay and serial dilution turbidity measurement assay. The results show that green synthesized silver nanoparticles, using guava ( Psidium guajava) leaf extract, have a potential to inhibit the growth of bacteria.

Antimicrobial resistance, heavy metal resistance and integron content in bacteria isolated from a South African tilapia aquaculture system.

PubMed

Chenia, Hafizah Y; Jacobs, Anelet

2017-11-21

Antibacterial compounds and metals co-select for antimicrobial resistance when bacteria harbour resistance genes towards both types of compounds, facilitating the proliferation and evolution of antimicrobial and heavy metal resistance. Antimicrobial and heavy metal resistance indices of 42 Gram-negative bacteria from a tilapia aquaculture system were determined to identify possible correlations between these phenotypes. Agar dilution assays were carried out to determine susceptibility to cadmium, copper, lead, mercury, chromate and zinc, while susceptibility to 21 antimicrobial agents was investigated by disk diffusion assays. Presence of merA, the mercury resistance gene, was determined by dot-blot hybridizations and PCR. Association of mercury resistance with integrons and transposon Tn21 was also investigated by PCR. Isolates displayed a high frequency of antimicrobial (erythromycin: 100%; ampicillin: 85%; trimethoprim: 78%) and heavy metal (Zn2+: 95%; Cd2+: 91%) resistance. No correlation was established between heavy metal and multiple antibiotic resistance indices. Significant positive correlations were observed between heavy metal resistance profiles, indices, Cu2+ and Cr3+ resistance with erythromycin resistance. Significant positive correlations were observed between merA (24%)/Tn21 (24%) presence and heavy metal resistance profiles and indices; however, significant negative correlations were obtained between integron-associated qacE∆1 (43%) and sulI (26%) gene presence and heavy metal resistance indices. Heavy metal and antimicrobial agents co-select for resistance, with fish-associated, resistant bacteria demonstrating simultaneous heavy metal resistance. Thus, care should be taken when using anti-fouling heavy metals as feed additives in aquaculture facilities.

Effects of hydroethanolic extracts of Balanites aegyptiaca (L.) Delile (Balanitaceae) on some resistant pathogens bacteria isolated from wounds.

PubMed

Anani, Kokou; Adjrah, Yao; Ameyapoh, Yaovi; Karou, Simplice Damintoti; Agbonon, Amegnona; de Souza, Comlan; Gbeassor, Messanvi

2015-04-22

The bark of Balanites aegyptiaca (L.) Delile (Balanitaceae) is widely used in the Togolese folk medicine for the treatment of wounds, malaria and skin diseases. This study aimed to evaluate the antioxidant properties and the activity of the bark extract of this plant against some resistant bacteria isolated from wounds. The antimicrobial activity was assayed using the NCCLS microdilution while the DPPH free radical scavenging and ferric reducing antioxidant power (FRAP) were used for the antioxidant activity. The susceptibility to conventional antibiotics of bacteria was assessed by an agar disk diffusion method using commercial disk. Phenolics were quantified using spectrophotometric methods. The hydroethanolic extracts of the bark inhibited in vitro the growth of Pseudomonas aeruginosa and Staphylococcus aureus in a dose-dependent manner. Bacteriostatic effects were observed on 15 (33%) and 21 (47%) strains of Staphylococcus aureus and Pseudomonas aeruginosa respectively. The MICs and MBCs against Staphylococcus aureus strains ranged respectively from 2.5 to 12.5 µg/mL and 2.5 to 50 µg/mL. Concerning Pseudomonas aeruginosa, these values were equal (12.5 µg/mL). Multidrug resistance was 95% against six antibiotics for Pseudomonas aeruginosa and 91% against five antibiotics for Staphylococcus aureus. The DPPH radical scavenging and the FRAP assays yielded weak antioxidant activity, closely linked with the total phenolic contents. This study indicated that the bark extract of Balanites aegyptiaca possess bioactive compounds implicated in the free radical scavenging and antibacterial activities, justifying the use of the plant in the traditional medicines. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

Negligible fractionation of Kr and Xe isotopes by molecular diffusion in water

NASA Astrophysics Data System (ADS)

Tyroller, Lina; Brennwald, Matthias S.; Busemann, Henner; Maden, Colin; Baur, Heinrich; Kipfer, Rolf

2018-06-01

Molecular diffusion is a key transport process for noble gases in water. Such diffusive transport is often thought to cause a mass-dependent fractionation of noble gas isotopes that is inversely proportional to the square root of the ratio of their atomic mass, referred to as the square root relation. Previous studies, challenged the commonly held assumption that the square root relation adequately describes the behaviour of noble gas isotopes diffusing through water. However, the effect of diffusion on noble gas isotopes has only been determined experimentally for He, Ne and Ar to date, whereas the extent of fractionation of Kr and Xe has not been measured. In the present study the fractionation of Kr and Xe isotopes diffusing through water immobilised by adding agar was quantified through measuring the respective isotope ratio after diffusing through the immobilised water. No fractionation of Kr and Xe isotopes was observed, even using high-precision noble gas analytics. These results complement our current understanding on isotopic fractionation of noble gases diffusing through water. Therefore this complete data set builds a robust basis to describe molecular diffusion of noble gases in water in a physical sound manner which is fundamental to assess the physical aspects of gas dynamics in aquatic systems.

In vitro susceptibility of filamentous fungi to copper nanoparticles assessed by rapid XTT colorimetry and agar dilution method.

PubMed

Ghasemian, E; Naghoni, A; Tabaraie, B; Tabaraie, T

2012-12-01

Metal nanoparticles and their uses in various aspects have recently drawn a great deal of attention. One of the major applications is that it can be used as an antimicrobial agent. They can be considered in approaches targeted to decrease the harms caused by microorganisms, specifically fungi, threatening the medical and industrial areas. The aim of this study was to investigate the antifungal activity of synthesized copper nanoparticles (CuNPs) against four filamentous fungi including Alternaria alternata, Aspergillus flavus, Fusarium solani, and Penicillium chrysogenum. Zerovalent copper nanoparticles of mean size 8nm were synthesized by inert gas condensation (IGC) method. The antifungal activity of these synthesized copper nanoparticles was measured against selected fungi by using two different techniques including agar dilution method and XTT reduction assay. The minimal inhibitory concentrations (MICs) for copper nanoparticles by agar dilution method were less or equal to 40mg/L for P. chrysogenum, less or equal to 60mg/L for A. alternata, less or equal to 60mg/L for F. solani, and less or equal to 80mg/L for A. flavus. And also MICs obtained by XTT reduction assay ranged from 40 to 80mg/L. Our data demonstrated that the copper nanoparticles inhibited fungal growth, but the fungal sensitivity to copper nanoparticles varies depending on the fungal species. Therefore, it is advisable that the minimal inhibitory concentrations (MICs) be examined before using these compounds. It is hoped that, in future, copper nanoparticles could replace some antifungal agents, making them applicable to many different medical devices and antimicrobial control system. Copyright © 2012 Elsevier Masson SAS. All rights reserved.

Enrichment of prostate cancer stem cells from primary prostate cancer cultures of biopsy samples

PubMed Central

Wang, Shunqi; Huang, Shengsong; Zhao, Xin; Zhang, Qimin; Wu, Min; Sun, Feng; Han, Gang; Wu, Denglong

2014-01-01

This study was to enrich prostate cancer stem cells (PrCSC) from primary prostate cancer cultures (PPrCC). Primary prostate cancer cells were amplified in keratinocyte serum-free medium with epidermal growth factor (EGF) and bovine pituitary extract (BPE), supplemented with leukemia inhibitory factor (LIF), stem cell factor (SCF) and cholera toxin. After amplification, cells were transferred into ultra-low attachment dishes with serum-free DMEM/F12 medium, supplemented with EGF, basic fibroblast growth factor (bFGF), bovine serum albumin (BSA), insulin, and N2 nutrition. Expression of cell-type-specific markers was determined by RT-qPCR and immunostaining. Tumorigenicity of enriched PrCSC was determined by soft agar assay and xenograft assay in NOD/SCID mice. Biopsy samples from 19 confirmed prostate cancer patients were used for establishing PPrCC, and 18 cases (95%) succeeded. Both basal marker (CK5) and luminal markers (androgen receptor and CK8) strongly co-expressed in most of PPrCC, indicating their basal epithelial origin. After amplification under adherent culture condition in vitro, transient amplifying cells were the dominant cells. Sphere formation efficiency (SFE) of passaged PPrCC was about 0.5%, which was 27 times lower than SFE of LNCaP (13.67%) in the same condition. Compared with adherent cells from PPrCC, prostasphere from PPrCC showed up regulated stem cell markers and increased tumorigenic potential in soft-agar assay. However, spheroid cells from PPrCC prostasphere failed to initiate tumor in xenograft assay in 6 months. Thus, PPrCC can be established and amplified from prostate cancer biopsy samples. Our modified sphere culture system can enrich PrCSC from PPrCC. PMID:24427338

Serum bactericidal assay for the evaluation of typhoid vaccine using a semi-automated colony-counting method.

PubMed

Jang, Mi Seon; Sahastrabuddhe, Sushant; Yun, Cheol-Heui; Han, Seung Hyun; Yang, Jae Seung

2016-08-01

Typhoid fever, mainly caused by Salmonella enterica serovar Typhi (S. Typhi), is a life-threatening disease, mostly in developing countries. Enzyme-linked immunosorbent assay (ELISA) is widely used to quantify antibodies against S. Typhi in serum but does not provide information about functional antibody titers. Although the serum bactericidal assay (SBA) using an agar plate is often used to measure functional antibody titers against various bacterial pathogens in clinical specimens, it has rarely been used for typhoid vaccines because it is time-consuming and labor-intensive. In the present study, we established an improved SBA against S. Typhi using a semi-automated colony-counting system with a square agar plate harboring 24 samples. The semi-automated SBA efficiently measured bactericidal titers of sera from individuals immunized with S. Typhi Vi polysaccharide vaccines. The assay specifically responded to S. Typhi Ty2 but not to other irrelevant enteric bacteria including Vibrio cholerae and Shigella flexneri. Baby rabbit complement was more appropriate source for the SBA against S. Typhi than complements from adult rabbit, guinea pig, and human. We also examined the correlation between SBA and ELISA for measuring antibody responses against S. Typhi using pre- and post-vaccination sera from 18 human volunteers. The SBA titer showed a good correlation with anti-Vi IgG quantity in the serum as determined by Spearman correlation coefficient of 0.737 (P < 0.001). Taken together, the semi-automated SBA might be efficient, accurate, sensitive, and specific enough to measure functional antibody titers against S. Typhi in sera from human subjects immunized with typhoid vaccines. Copyright © 2016 The Authors. Published by Elsevier Ltd.. All rights reserved.

Simple fluorescence-based high throughput cell viability assay for filamentous fungi.

PubMed

Chadha, S; Kale, S P

2015-09-01

Filamentous fungi are important model organisms to understand the eukaryotic process and have been frequently exploited in research and industry. These fungi are also causative agents of serious diseases in plants and humans. Disease management strategies include in vitro susceptibility testing of the fungal pathogens to environmental conditions and antifungal agents. Conventional methods used for antifungal susceptibilities are cumbersome, time-consuming and are not suitable for a large-scale analysis. Here, we report a rapid, high throughput microplate-based fluorescence method for investigating the toxicity of antifungal and stress (osmotic, salt and oxidative) agents on Magnaporthe oryzae and compared it with agar dilution method. This bioassay is optimized for the resazurin reduction to fluorescent resorufin by the fungal hyphae. Resazurin bioassay showed inhibitory rates and IC50 values comparable to the agar dilution method and to previously reported IC50 or MICs for M. oryzae and other fungi. The present method can screen range of test agents from different chemical classes with different modes of action for antifungal activities in a simple, sensitive, time and cost effective manner. A simple fluorescence-based high throughput method is developed to test the effects of stress and antifungal agents on viability of filamentous fungus Magnaporthe oryzae. This resazurin fluorescence assay can detect inhibitory effects comparable to those obtained using the growth inhibition assay with added advantages of simplicity, time and cost effectiveness. This high throughput viability assay has a great potential in large-scale screening of the chemical libraries of antifungal agents, for evaluating the effects of environmental conditions and hyphal kinetic studies in mutant and natural populations of filamentous fungi. © 2015 The Society for Applied Microbiology.

Inactivation of ferric uptake regulator (Fur) attenuates Helicobacter pylori J99 motility by disturbing the flagellar motor switch and autoinducer-2 production.

PubMed

Lee, Ai-Yun; Kao, Cheng-Yen; Wang, Yao-Kuan; Lin, Ssu-Yuan; Lai, Tze-Ying; Sheu, Bor-Shyang; Lo, Chien-Jung; Wu, Jiunn-Jong

2017-08-01

Flagellar motility of Helicobacter pylori has been shown to be important for the bacteria to establish initial colonization. The ferric uptake regulator (Fur) is a global regulator that has been identified in H. pylori which is involved in the processes of iron uptake and establishing colonization. However, the role of Fur in H. pylori motility is still unclear. Motility of the wild-type, fur mutant, and fur revertant J99 were determined by a soft-agar motility assay and direct video observation. The bacterial shape and flagellar structure were evaluated by transmission electron microscopy. Single bacterial motility and flagellar switching were observed by phase-contrast microscopy. Autoinducer-2 (AI-2) production in bacterial culture supernatant was analyzed by a bioluminescence assay. The fur mutant showed impaired motility in the soft-agar assay compared with the wild-type J99 and fur revertant. The numbers and lengths of flagellar filaments on the fur mutant cells were similar to those of the wild-type and revertant cells. Phenotypic characterization showed similar swimming speed but reduction in switching rate in the fur mutant. The AI-2 production of the fur mutant was dramatically reduced compared with wild-type J99 in log-phase culture medium. These results indicate that Fur positively modulates H. pylori J99 motility through interfering with bacterial flagellar switching. © 2017 John Wiley & Sons Ltd.

Novel alternative to antibiotics in shrimp hatchery: effects of the essential oil of Cinnamosma fragrans on survival and bacterial concentration of Penaeus monodon larvae.

PubMed

Randrianarivelo, R; Danthu, P; Benoit, C; Ruez, P; Raherimandimby, M; Sarter, S

2010-08-01

The activity of two essential oils (EOs) of Cinnamosma fragrans, an endemic plant to Madagascar (B8: linalool-type and B143: 1,8-cineole-type), against bacterial isolates from a shrimp hatchery of Penaeus monodon and their effects on the survival and bacterial concentration of larvae were determined. Minimum inhibitory concentrations were determined using a broth dilution technique. The bacterial concentrations of both larvae and water tank were assessed on Marine agar and Thiosulfate Citrate Bile Sucrose agar. The assays took place in OSO Farming's shrimp hatchery in Madagascar. EOs were directly added to the water tank. Regarding the survival, the assays in larval culture (four replicates each of B8, B143, E and control) showed that B8 oil had a similar effect (P > 0.05) as the antibiotic (Erythromycin) and was more active than B143 (P < 0.05). A negative correlation was observed between the bacterial concentration and the survival of larvae for all assays. Both C. fragrans essential oils, as antibiotic, exhibited significantly higher survival rates and lower bacterial concentrations of the larvae than the control (oil and antibiotic free). The potential of C. fragrans essential oil to control the bacterial load in in vivo conditions, thereby enhancing survival rate of P. monodon larvae, makes it a relevant option for developing a novel alternative to antibiotics in shrimp hatchery culture.

The neoplastic potential of rat tracheal epithelial cell lines induced by dibenzo(a, i)pyrene and 1-nitropyrene

DOE Office of Scientific and Technical Information (OSTI.GOV)

Xiang, M.; Ong, T.; Nath, J.

1997-10-01

The rat tracheal epithelial (RTE) cell transformation system is an important short-term assay for respiratory carcinogenesis. In our laboratories, studies have been performed using this assay system to determine the carcinogenic potential of dibenzo(a,i)pyrene (DBP) and 1-nitropyrene (1-NP), two compounds commonly contaminating occupational and environmental settings. RTE cells were exposed in vivo to DBP or 1-NP by intertracheal instillation. RTE cells were then isolated and plated on a medium for determination of cloning and transformation frequencies. Cell lines established from transformed cells induced by DBP and 1-NP were analyzed for their neoplastic potential with the soft agar cloning and themore » athymic nude mouse tumorigenicity assays. Results showed that: (1) incidence of transformed foci in cultures treated with DBP or 1-NP in vivo was significantly higher than that in the control cultures; (2) 8 and 25 cell lines were established from 28 and 48 transformed foci induced by DBP and 1-NP, respectively; (3) 3 of 5 cell lines from DBP and 5 anchorage independent growth in soft agar; (4) some of the cell lines from DBP and 1-NP induced transformed foci formed tumors after cells were injected in athymic nude mice. These results indicate that in vivo exposure to DBP and 1-NP can induce RTE cell transformation and that transformed cells induced by DBP and 1-NP may have neoplastic potential.« less

Reactivation of Embryonic Nodal Signaling is Associated with Tumor Progression and Promotes the Growth of Prostate Cancer Cells

PubMed Central

Lawrence, Mitchell G.; Margaryan, Naira V.; Loessner, Daniela; Collins, Angus; Kerr, Kris M.; Turner, Megan; Seftor, Elisabeth A.; Stephens, Carson R.; Lai, John; BioResource, APC; Postovit, Lynne-Marie; Clements, Judith A.; Hendrix, Mary J.C.

2011-01-01

Background Nodal is a member of the Transforming Growth Factor β (TGFβ) superfamily that directs embryonic patterning and promotes the plasticity and tumorigenicity of tumor cells, but its role in the prostate is unknown. The goal of this study was to characterize the expression and function of Nodal in prostate cancer and determine whether, like other TGFβ ligands, it modulates androgen receptor (AR) activity. Methods Nodal expression was investigated using immunohistochemistry of tissue microarrays and Western blots of prostate cell lines. The functional role of Nodal was examined using Matrigel and soft agar growth assays. Cross-talk between Nodal and AR signaling was assessed with luciferase reporter assays and expression of endogenous androgen regulated genes. Results Significantly increased Nodal expression was observed in cancer compared with benign prostate specimens. Nodal was only expressed by DU145 and PC3 cells. All cell lines expressed Nodal’s co-receptor, Cripto-1, but lacked Lefty, a critical negative regulator of Nodal signaling. Recombinant human Nodal triggered downstream Smad2 phosphorylation in DU145 and LNCaP cells, and stable transfection of pre-pro-Nodal enhanced the growth of LNCaP cells in Matrigel and soft agar. Finally, Nodal attenuated AR signaling, reducing the activity of a PSA promoter construct in luciferase assays and down-regulating the endogenous expression of androgen regulated genes. Conclusions An aberrant Nodal signaling pathway is re-expressed and functionally active in prostate cancer cells. PMID:21656830

Isolation and characterization of potential antibiotic producing actinomycetes from water and sediments of Lake Tana, Ethiopia

PubMed Central

Gebreyohannes, Gebreselema; Moges, Feleke; Sahile, Samuel; Raja, Nagappan

2013-01-01

Objective To isolate, evaluate and characterize potential antibiotic producing actinomycetes from water and sediments of Lake Tana, Ethiopia. Methods A total of 31 strains of actinomycetes were isolated and tested against Gram positive and Gram negative bacterial strains by primary screening. In the primary screening, 11 promising isolates were identified and subjected to solid state and submerged state fermentation methods to produce crude extracts. The fermented biomass was extracted by organic solvent extraction method and tested against bacterial strains by disc and agar well diffusion methods. The isolates were characterized by using morphological, physiological and biochemical methods. Results The result obtained from agar well diffusion method was better than disc diffusion method. The crude extract showed higher inhibition zone against Gram positive bacteria than Gram negative bacteria. One-way analysis of variance confirmed most of the crude extracts were statistically significant at 95% confidence interval. The minimum inhibitory concentration and minimum bactericidal concentration of crude extracts were 1.65 mg/mL and 3.30 mg/mL against Staphylococcus aureus, and 1.84 mg/mL and 3.80 mg/mL against Escherichia coli respectively. The growth of aerial and substrate mycelium varied in different culture media used. Most of the isolates were able to hydrolysis starch and urea; able to survive at 5% concentration of sodium chloride; optimum temperature for their growth was 30 °C. Conclusions The results of the present study revealed that freshwater actinomycetes of Lake Tana appear to have immense potential as a source of antibacterial compounds. PMID:23730554

QIN DAWG Validation of Gradient Nonlinearity Bias Correction Workflow for Quantitative Diffusion-Weighted Imaging in Multicenter Trials.

PubMed

Malyarenko, Dariya I; Wilmes, Lisa J; Arlinghaus, Lori R; Jacobs, Michael A; Huang, Wei; Helmer, Karl G; Taouli, Bachir; Yankeelov, Thomas E; Newitt, David; Chenevert, Thomas L

2016-12-01

Previous research has shown that system-dependent gradient nonlinearity (GNL) introduces a significant spatial bias (nonuniformity) in apparent diffusion coefficient (ADC) maps. Here, the feasibility of centralized retrospective system-specific correction of GNL bias for quantitative diffusion-weighted imaging (DWI) in multisite clinical trials is demonstrated across diverse scanners independent of the scanned object. Using corrector maps generated from system characterization by ice-water phantom measurement completed in the previous project phase, GNL bias correction was performed for test ADC measurements from an independent DWI phantom (room temperature agar) at two offset locations in the bore. The precomputed three-dimensional GNL correctors were retrospectively applied to test DWI scans by the central analysis site. The correction was blinded to reference DWI of the agar phantom at magnet isocenter where the GNL bias is negligible. The performance was evaluated from changes in ADC region of interest histogram statistics before and after correction with respect to the unbiased reference ADC values provided by sites. Both absolute error and nonuniformity of the ADC map induced by GNL (median, 12%; range, -35% to +10%) were substantially reduced by correction (7-fold in median and 3-fold in range). The residual ADC nonuniformity errors were attributed to measurement noise and other non-GNL sources. Correction of systematic GNL bias resulted in a 2-fold decrease in technical variability across scanners (down to site temperature range). The described validation of GNL bias correction marks progress toward implementation of this technology in multicenter trials that utilize quantitative DWI.

QIN DAWG Validation of Gradient Nonlinearity Bias Correction Workflow for Quantitative Diffusion-Weighted Imaging in Multicenter Trials

PubMed Central

Malyarenko, Dariya I.; Wilmes, Lisa J.; Arlinghaus, Lori R.; Jacobs, Michael A.; Huang, Wei; Helmer, Karl G.; Taouli, Bachir; Yankeelov, Thomas E.; Newitt, David; Chenevert, Thomas L.

2017-01-01

Previous research has shown that system-dependent gradient nonlinearity (GNL) introduces a significant spatial bias (nonuniformity) in apparent diffusion coefficient (ADC) maps. Here, the feasibility of centralized retrospective system-specific correction of GNL bias for quantitative diffusion-weighted imaging (DWI) in multisite clinical trials is demonstrated across diverse scanners independent of the scanned object. Using corrector maps generated from system characterization by ice-water phantom measurement completed in the previous project phase, GNL bias correction was performed for test ADC measurements from an independent DWI phantom (room temperature agar) at two offset locations in the bore. The precomputed three-dimensional GNL correctors were retrospectively applied to test DWI scans by the central analysis site. The correction was blinded to reference DWI of the agar phantom at magnet isocenter where the GNL bias is negligible. The performance was evaluated from changes in ADC region of interest histogram statistics before and after correction with respect to the unbiased reference ADC values provided by sites. Both absolute error and nonuniformity of the ADC map induced by GNL (median, 12%; range, −35% to +10%) were substantially reduced by correction (7-fold in median and 3-fold in range). The residual ADC nonuniformity errors were attributed to measurement noise and other non-GNL sources. Correction of systematic GNL bias resulted in a 2-fold decrease in technical variability across scanners (down to site temperature range). The described validation of GNL bias correction marks progress toward implementation of this technology in multicenter trials that utilize quantitative DWI. PMID:28105469

A Direct Quantitative Agar-Plate Based Assay for Analysis of Pseudomonas protegens PF-5 Degradation of Polyurethane Films (Postprint)

DTIC Science & Technology

2014-10-02

Journal article published in International Biodeterioration & Biodegradation 95 (2014) 311-319. The U.S. Government is joint author of the work and...SUBJECT TERMS pseudomonas biofilms, polyurethane, biodegradation , FTIR spectroscopy, citrate, impranil 16. SECURITY CLASSIFICATION OF: 17. LIMITATION OF...International Biodeterioration & Biodegradation 95 (2014) 311e319Contents lists avaiInternational Biodeterioration & Biodegradation journal homepage

Bacterial Chemotaxis to Naphthalene and Nitroarene Compounds

DTIC Science & Technology

2008-07-31

Qualitative capillary assays showing chemotaxis of succinate-grown 17 (uninduced) and induced (succinate plus salicylate -grown) Acidovorax sp. JS42...succinate plus 2NT- or succinate plus salicylate -grown) wild-type Acidovorax sp. JS42 cells List of Tables Table 1. Summary of chemotaxis...mM salicylate , or naphthalene crystals. Noble agar (1.8%; Difco) was used to solidify MSB medium for plates. For plasmid selection and maintenance

A comparative evaluation of antimicrobial efficacy and flow properties for Epiphany, Guttaflow and AH-Plus sealer.

PubMed

Nawal, R R; Parande, M; Sehgal, R; Naik, A; Rao, N R

2011-04-01

To test the antimicrobial efficacy and flow properties of Guttaflow, Epiphany sealer and AH-Plus sealer. With the use of Enterococcus faecalis ATCC 29212 as a test organism, both the agar diffusion test (ADT) and direct contact test (DCT) were performed. For DCT, sealers were mixed and placed over the bottom of sterile screw-capped test tubes. A 50 μL bacterial suspension was placed on the tested material samples. Bacteria were allowed to directly come in contact with the sealers for 1 h at 37 °C in one group and for 24 h in the other group. The suspensions were then diluted and inoculated over blood agar plates, and bacterial colony counts were determined with the use of a digital colony counter. The data in both 1- and 24-h groups were individually analysed using repeated measures ANOVA. Kruskal Wallis tests were further used to obtain comparison between 1- and 24-h results for all three sealers. In the flow assay, the sealers were placed between two glass slides, and a weight of 500 g was placed on the top of the glass. The diameters of the formed discs were recorded. For both the ADT and DCT tests, Epiphany and AH-Plus sealer reduced the bacterial counts significantly (P = 0.000). Epiphany produced a greater reduction in bacterial counts when compared to AH-Plus in both the tests (P = 0.000). Guttaflow paste failed to show any antibacterial activity in both ADT & DCT. According to the flow test, all root canal sealers flowed; Epiphany sealer had the maximum flow under the given conditions, followed by AH-Plus sealer and Guttaflow paste. Antimicrobial activity of the sealers was greatest for Epiphany followed by AH-Plus sealer and Guttaflow. Epiphany sealer had the maximum flow followed by AH-Plus sealer and Guttaflow. © 2010 International Endodontic Journal.

Comparative evaluation of MRSA nasal colonization epidemiology in the urban and rural secondary school community of Kurdistan, Iraq.

PubMed

Hussein, Nawfal R; Basharat, Zarrin; Muhammed, Ary H; Al-Dabbagh, Samim A

2015-01-01

To study the nasal carriage rate of Staphylococcus aureus (S. aureus) (including methicillin-resistant strains) in secondary school community of the urban and rural districts of the Kurdistan region of Iraq, a cross-sectional population based survey was carried out in the city Duhok and rural areas of Amedya, Akre and Zakho. Nasal swabs were obtained from nostrils of 509 students aged 14-23 years. Resistance to methicillin was assessed by Kirby-Bauer disk diffusion and agar dilution assay. Vancomycin sensitivity was also tested on Muller-Hinton agar. It was found that the frequency of overall S. aureus nasal carriage (SANC) was 17.75% (90/509, CI95, 14.58-21.42%). In urban areas, the carriage rate was 20.59% (49/239, CI95, 15.64-26.29%), whereas it was 15.24% (41/270, CI95, 11.17-20.10%) in rural districts. The frequency of methicillin-resistant S. aureus (MRSA) among the isolated strains was found to be 2.04% (1/49) and 21.95% (9/41) in urban and rural areas respectively. It was found that in urban residents, the odd ratio (OR) of acquiring SANC was 1.44 (CI95, 0.91-2.27%) and risk ratio (RR) was at least 1.35 (CI95, 0.92-1.96%) while OR decreased to 0.12 (CI95, 0.01-0.96%) for MRSA carriage. Hence, the S. aureus carriage rate was higher in urban districts compared to rural areas while more MRSA were found in rural areas compared to urban districts. All studied strains were sensitive to vancomycin. This study provided baseline information for S. aureus nasal colonization in the region. Also, it showed that living in rural areas increased the odds of MRSA colonization. More attention should be paid to control MRSA colonization in rural communities.

Comparative Evaluation of MRSA Nasal Colonization Epidemiology in the Urban and Rural Secondary School Community of Kurdistan, Iraq

PubMed Central

Hussein, Nawfal R.; Basharat, Zarrin; Muhammed, Ary H.; Al-Dabbagh, Samim A.

2015-01-01

Background To study the nasal carriage rate of Staphylococcus aureus (S. aureus) (including methicillin-resistant strains) in secondary school community of the urban and rural districts of the Kurdistan region of Iraq, a cross-sectional population based survey was carried out in the city Duhok and rural areas of Amedya, Akre and Zakho. Methods Nasal swabs were obtained from nostrils of 509 students aged 14-23 years. Resistance to methicillin was assessed by Kirby-Bauer disk diffusion and agar dilution assay. Vancomycin sensitivity was also tested on Muller-Hinton agar. Results It was found that the frequency of overall S. aureus nasal carriage (SANC) was 17.75% (90/509, CI95, 14.58–21.42%). In urban areas, the carriage rate was 20.59% (49/239, CI95, 15.64–26.29%), whereas it was 15.24% (41/270, CI95, 11.17–20.10%) in rural districts. The frequency of methicillin-resistant S. aureus (MRSA) among the isolated strains was found to be 2.04% (1/49) and 21.95% (9/41) in urban and rural areas respectively. It was found that in urban residents, the odd ratio (OR) of acquiring SANC was 1.44 (CI95, 0.91-2.27%) and risk ratio (RR) was at least 1.35 (CI95, 0.92-1.96%) while OR decreased to 0.12 (CI95, 0.01-0.96%) for MRSA carriage. Hence, the S. aureus carriage rate was higher in urban districts compared to rural areas while more MRSA were found in rural areas compared to urban districts. All studied strains were sensitive to vancomycin. Conclusion This study provided baseline information for S. aureus nasal colonization in the region. Also, it showed that living in rural areas increased the odds of MRSA colonization. More attention should be paid to control MRSA colonization in rural communities. PMID:25932644

Microscopic-observation drug susceptibility and thin layer agar assays for the detection of drug resistant tuberculosis: a systematic review and meta-analysis.

PubMed

Minion, Jessica; Leung, Erika; Menzies, Dick; Pai, Madhukar

2010-10-01

Simple, rapid, and affordable tests are needed to detect drug resistance in Mycobacterium tuberculosis. We did a systematic review and meta-analysis to investigate the accuracy of microscopic-observation drug susceptibility (MODS) and thin layer agar (TLA) assays for rapid screening of patients at risk of drug-resistant tuberculosis. In accordance with protocols and methods recommended by the Cochrane Diagnostic Test Accuracy Working Group, we systematically searched PubMed, Embase, and Biosis for reports published between January, 1990, and February, 2009. We included studies investigating detection of drug resistance in M tuberculosis with the MODS or TLA assay, and in which an accepted reference standard was used. Data extracted from the studies were combined by use of bivariate random-effects regression models and hierarchical summary receiver operating characteristic curves to estimate sensitivity and specificity for detection of resistance to specific drugs. We identified 12 studies, of which nine investigated the MODS assay and three investigated the TLA assay. For the MODS assay of rifampicin resistance, pooled estimates were 98·0% (95% CI 94·5-99·3) for sensitivity and 99·4% (95·7-99·9) for specificity. For the MODS assay of isoniazid resistance with a 0·1 μg/mL cutoff, pooled sensitivity was 97·7% (94·4-99·1) and pooled specificity was 95·8% (88·1-98·6), but with a 0·4 μg/mL cutoff, sensitivity decreased to 90·0% (84·5-93·7) and specificity increased to 98·6% (96·9-99·4). All assessments of rifampicin and isoniazid resistance with the TLA assay yielded 100% accuracy. Mean turnaround time was 9·9 days (95% CI 4·1-15·8) for the MODS assay and 11·1 days (10·1-12·0) for the TLA assay. MODS and TLA assays are inexpensive, rapid alternatives to conventional methods for drug susceptibility testing of M tuberculosis. Our data and expert opinion informed WHO's recommendation for use of selected non-commercial drug susceptibility tests, including MODS, as an interim solution until capacity for genotypic or automated liquid culture drug susceptibility testing is developed. Stop TB Department of WHO. Copyright © 2010 Elsevier Ltd. All rights reserved.

Chemical composition and antibacterial activity of selected essential oils and some of their main compounds.

PubMed

Wanner, Juergen; Schmidt, Erich; Bail, Stefanie; Jirovetz, Leopold; Buchbauer, Gerhard; Gochev, Velizar; Girova, Tanya; Atanasova, Teodora; Stoyanova, Albena

2010-09-01

The chemical composition of essential oils of cabreuva (Myrocarpus fastigiatus Allemao, Fabaceae) from Brazil, cedarwood (Juniperus ashei, Cupressaceae) from Texas, Juniper berries (Juniperus communis L., Cupressaceae) and myrrh (Commiphora myrrha (Nees) Engl., Burseraceae) were analyzed using GC/FID and GC/MS. The antimicrobial activity of these essential oils and some of their main compounds were tested against eleven different strains of Gram-positive and Gram-negative bacteria by using agar diffusion and agar serial dilution methods. Animal and plant pathogens, food poisoning and spoilage bacteria were selected. The volatile oils exhibited considerable inhibitory effects against all tested organisms, except Pseudomonas, using both test methods. Higher activity was observed against Gram-positive strains in comparison with Gram-negative bacteria. Cabreuva oil from Brazil showed similar results, but in comparison with the other oils tested, only when higher concentrations of oil were used.

High prevalence of oral colonization by Candida dubliniensis in HIV-positive patients in Argentina.

PubMed

Binolfi, Andrés; Biasoli, Marisa S; Luque, Alicia G; Tosello, María E; Magaró, Hortensia M

2005-08-01

Candida dubliniensis is a recently described yeast species, closely related to Candida albicans. This work represents the first general survey of the carriage of C. dubliniensis in the oral cavities of HIV-positive patients in Argentina. We studied 133 strains isolated from 162 HIV-positive patients, using the following identification tests: chlamydospore production on corn meal agar with Tween 80; colony color on CHROMagar Candida media; differential growth at 45 degrees C on potato dextrose agar; D-xylose assimilation; chlamydospore formation on sunflower seed agar (SSA); carbohydrate assimilation profiles using the API 20 C Aux commercial kit and PCR using primers that hybridize to the class IV intron of the ACT1 gene. Out of the 133 strains, 21 were identified as C. dubliniensis, representing approximately 13% of the 162 patients in this study. From these data, we conclude that although the PCR assay is the most reliable method, clamydospore formation on SSA is an easier and less expensive test for the screening of C. dubliniensis in the routine laboratory. Our results show that C. dubliniensis has a high prevalence among HIV-positive patients in Argentina.

A direct observation technique for evaluating sclerotium germination by Macrophomina phaseolina and effects of biocontrol materials on survival of sclerotia in soil.

PubMed

Pratt, Robert G

2006-08-01

Germination of sclerotia of Macrophomina phaseolina was quantified by direct microscopic observation following application of experimental treatments in vitro and incubation of sclerotia in soil. To assay germination, pieces of agar containing sclerotia were macerated in dilute, liquid cornmeal agar on glass slides; thinly spread; and incubated in a saturated atmosphere for 18-22 h. Germinated sclerotia then were identified by morphological features of germ hyphae. Frequencies of germination were similar in three dilute agar media. Germination was not affected by air-drying sclerotia for 2 weeks, but it was significantly reduced after 4 weeks and greatly reduced or eliminated after 6 or 8 weeks. Survival of sclerotia for 14 days in soil was greatest at 50, 75, and 100% moisture-holding capacity, less at 0 and 25%, and least at 125% (flooded soil). Incorporation of ground poultry litter into soil at 5% by weight reduced survival of sclerotia after 13 days, and incorporation of litter at 10% nearly eliminated it. These results indicate that the direct-observation technique may be used to evaluate animal wastes and other agricultural byproducts for biocontrol activity against sclerotia of M. phaseolina in soil.

A spectrophotometer-based diffusivity assay reveals that diffusion hindrance of small molecules in extracellular matrix gels used in 3D cultures is dominated by viscous effects.

PubMed

Galgoczy, Roland; Pastor, Isabel; Colom, Adai; Giménez, Alicia; Mas, Francesc; Alcaraz, Jordi

2014-08-01

The design of 3D culture studies remains challenging due to the limited understanding of extracellular matrix (ECM)-dependent hindered diffusion and the lack of simple diffusivity assays. To address these limitations, we set up a cost-effective diffusivity assay based on a Transwell plate and the spectrophotometer of a Microplate Reader, which are readily accessible to cell biology groups. The spectrophotometer-based assay was used to assess the apparent diffusivity D of FITC-dextrans with molecular weight (4-70kDa) spanning the physiological range of signaling factors in a panel of acellular ECM gels including Matrigel, fibrin and type I collagen. Despite their technical differences, D data exhibited ∼15% relative difference with respect to FRAP measurements. Our results revealed that diffusion hindrance of small particles is controlled by the enhanced viscosity of the ECM gel in conformance with the Stokes-Einstein equation rather than by geometrical factors. Moreover, we provided a strong rationale that the enhanced ECM viscosity is largely contributed to by unassembled ECM macromolecules. We also reported that gels with the lowest D exhibited diffusion hindrance closest to the large physiologic hindrance of brain tissue, which has a typical pore size much smaller than ECM gels. Conversely, sparse gels (≤1mg/ml), which are extensively used in 3D cultures, failed to reproduce the hindered diffusion of tissues, thereby supporting that dense (but not sparse) ECM gels are suitable tissue surrogates in terms of macromolecular transport. Finally, the consequences of reduced diffusivity in terms of optimizing the design of 3D culture experiments were addressed in detail. Copyright © 2014 Elsevier B.V. All rights reserved.

A highly efficient nonchemical method for isolating live nematodes (Caenorhabditis elegans) from soil during toxicity assays.

PubMed

Kim, Shin Woong; Moon, Jongmin; An, Youn-Joo

2015-01-01

The success of soil toxicity tests using Caenorhabditis elegans may depend in large part on recovering the organisms from the soil. However, it can be difficult to learn the International Organization for Standardization/ASTM International recovery process that uses the colloidal silica flotation method. The present study determined that a soil-agar isolation method provides a highly efficient and less technically demanding alternative to the colloidal silica flotation method. Test soil containing C. elegans was arranged on an agar plate in a donut shape, a linear shape, or a C curve; and microbial food was placed outside the soil to encourage the nematodes to leave the soil. The effects of ventilation and the presence of food on nematode recovery were tested to determine the optimal conditions for recovery. A linear arrangement of soil on an agar plate that was sprinkled with microbial food produced nearly 83% and 90% recovery of live nematodes over a 3-h and a 24-h period, respectively, without subjecting the nematodes to chemical stress. The method was tested using copper (II) chloride dihydrate, and the resulting recovery rate was comparable to that obtained using colloidal silica flotation. The soil-agar isolation method portrayed in the present study enables live nematodes to be isolated with minimal additional physicochemical stress, making it a valuable option for use in subsequent sublethal tests where live nematodes are required. © 2014 SETAC.

Selection of Brain Metastasis-Initiating Breast Cancer Cells Determined by Growth on Hard Agar

PubMed Central

Guo, Lixia; Fan, Dominic; Zhang, Fahao; Price, Janet E.; Lee, Ju-Seog; Marchetti, Dario; Fidler, Isaiah J.; Langley, Robert R.

2011-01-01

An approach that facilitates rapid isolation and characterization of tumor cells with enhanced metastatic potential is highly desirable. Here, we demonstrate that plating GI-101A human breast cancer cells on hard (0.9%) agar selects for the subpopulation of metastasis-initiating cells. The agar-selected cells, designated GI-AGR, were homogeneous for CD44+ and CD133+ and five times more invasive than the parental GI-101A cells. Moreover, mice injected with GI-AGR cells had significantly more experimental brain metastases and shorter overall survival than did mice injected with GI-101A cells. Comparative gene expression analysis revealed that GI-AGR cells were markedly distinct from the parental cells but shared an overlapping pattern of gene expression with the GI-101A subline GI-BRN, which was generated by repeated in vivo recycling of GI-101A cells in an experimental brain metastasis model. Data mining on 216 genes shared between GI-AGR and GI-BRN breast cancer cells suggested that the molecular phenotype of these cells is consistent with that of cancer stem cells and the aggressive basal subtype of breast cancer. Collectively, these results demonstrate that analysis of cell growth in a hard agar assay is a powerful tool for selecting metastasis-initiating cells in a heterogeneous population of breast cancer cells, and that such selected cells have properties similar to those of tumor cells that are selected based on their potential to form metastases in mice. PMID:21514446

Diffusion processes in tumors: A nuclear medicine approach

DOE Office of Scientific and Technical Information (OSTI.GOV)

Amaya, Helman, E-mail: haamayae@unal.edu.co

The number of counts used in nuclear medicine imaging techniques, only provides physical information about the desintegration of the nucleus present in the the radiotracer molecules that were uptaken in a particular anatomical region, but that information is not a real metabolic information. For this reason a mathematical method was used to find a correlation between number of counts and {sup 18}F-FDG mass concentration. This correlation allows a better interpretation of the results obtained in the study of diffusive processes in an agar phantom, and based on it, an image from the PETCETIX DICOM sample image set from OsiriX-viewer softwaremore » was processed. PET-CT gradient magnitude and Laplacian images could show direct information on diffusive processes for radiopharmaceuticals that enter into the cells by simple diffusion. In the case of the radiopharmaceutical {sup 18}F-FDG is necessary to include pharmacokinetic models, to make a correct interpretation of the gradient magnitude and Laplacian of counts images.« less

Carbohydrate hydrogels with stabilized phage particles for bacterial biosensing: bacterium diffusion studies.

PubMed

Balcão, Victor M; Barreira, Sérgio V P; Nunes, Thiago M; Chaud, Marco V; Tubino, Matthieu; Vila, Marta M D C

2014-02-01

Bacteriophage particles have been reported as potentially useful in the development of diagnosis tools for pathogenic bacteria as they specifically recognize and lyse bacterial isolates thus confirming the presence of viable cells. One of the most representative microorganisms associated with health care services is the bacterium Pseudomonas aeruginosa, which alone is responsible for nearly 15% of all nosocomial infections. In this context, structural and functional stabilization of phage particles within biopolymeric hydrogels, aiming at producing cheap (chromogenic) bacterial biosensing devices, has been the goal of a previous research effort. For this, a detailed knowledge of the bacterial diffusion profile into the hydrogel core, where the phage particles lie, is of utmost importance. In the present research effort, the bacterial diffusion process into the biopolymeric hydrogel core was mathematically described and the theoretical simulations duly compared with experimental results, allowing determination of the effective diffusion coefficients of P. aeruginosa in the agar and calcium alginate hydrogels tested.

Application of orange essential oil as an antistaphylococcal agent in a dressing model.

PubMed

Muthaiyan, Arunachalam; Biswas, Debabrata; Crandall, Philip G; Wilkinson, Brian J; Ricke, Steven C

2012-08-16

Staphylococcus aureus is the pathogen most often and prevalently involved in skin and soft tissue infections. In recent decades outbreaks of methicillin-resistant S. aureus (MRSA) have created major problems for skin therapy, and burn and wound care units. Topical antimicrobials are most important component of wound infection therapy. Alternative therapies are being sought for treatment of MRSA and one area of interest is the use of essential oils. With the increasing interest in the use and application of natural products, we screened the potential application of terpeneless cold pressed Valencia orange oil (CPV) for topical therapy against MRSA using an in vitro dressing model and skin keratinocyte cell culture model. The inhibitory effect of CPV was determined by disc diffusion vapor assay for MRSA and vancomycin intermediate-resistant S. aureus (VISA) strains. Antistaphylococcal effect of CPV in an in vitro dressing model was tested on S. aureus inoculated tryptic soya agar plate. Bactericidal effect of CPV on MRSA and VISA infected keratinocyte cells was examined by enumeration of extra- and intra-cellular bacterial cells at different treatment time points. Cytotoxic effects on human skin cells was tested by adding CPV to the keratinocyte (HEK001) cells grown in serum free KSFM media, and observed by phase-contrast microscope. CPV vapour effectively inhibited the MRSA and VISA strains in both disc diffusion vapour assay and in vitro dressing model. Compared to untreated control addition of 0.1% CPV to MRSA infected keratinocyte decreased the viable MRSA cells by 2 log CFU/mL in 1 h and in VISA strain 3 log CFU/mL reduction was observed in 1 h. After 3 h viable S. aureus cells were not detected in the 0.2% CPV treatment. Bactericidal concentration of CPV did not show any cytotoxic effect on the human skin keratinocyte cells in vitro. At lower concentration addition of CPV to keratinocytes infected with MRSA and VISA rapidly killed the bacterial cells without causing any toxic effect to the keratinocytes. Therefore, the results of this study warrant further in vivo study to evaluate the potential of CPV as a topical antistaphylococcal agent.

The high diversity of MRSA clones detected in a university hospital in istanbul.

PubMed

Oksuz, Lutfiye; Dupieux, Celine; Tristan, Anne; Bes, Michele; Etienne, Jerome; Gurler, Nezahat

2013-01-01

To characterize the methicillin-resistant Staphylococcus aureus (MRSA) clones present in Istanbul, 102 MRSA isolates collected during a 5-year period at the Istanbul Medical Faculty Hospital were characterized using microarray analysis and phenotypic resistance profiles. Resistance to methicillin was detected with a cefoxitin disk diffusion assay and confirmed with a MRSA-agar and MRSA detection kit. Antimicrobial susceptibility testing was performed by a disk diffusion assay and interpreted according to the 2012 guidelines of the Antibiogram Committee of the French Society for Microbiology. Decreased susceptibility to glycopeptides was confirmed using the population analysis profile-area under the curve (PAP-AUC) method. The presence of the mecA gene was detected by polymerase chain reaction. Bacterial DNA was extracted according to the manufacturer's recommended protocol using commercial extraction kits. Strains were extensively characterized using the DNA microarray. Isolates were grouped into six clonal complexes. The most frequently detected clone was the Vienna/Hungarian/Brazilian clone (ST239-MRSA-III), which accounted for 53.9% of the isolates. These isolates were resistant to multiple antibiotics, particularly penicillin, tetracycline, rifampicin, kanamycin, tobramycin, gentamicin, levofloxacin, erythromycin, lincomycin and fosfomycin. Furthermore, three isolates were detected by population analysis profile as heterogeneous vancomycin-intermediate S. aureus (hVISA). The UK-EMRSA-15 clone (ST22-MRSA-IV PVL negative) was detected in 9.8% of the isolates and was mainly susceptible to all anti-staphylococcal antibiotics. Seven isolates (6.9%) were positive for PVL genes and were assigned to the CC80-MRSA-IV clone (European CA-MRSA clone, three isolates), ST8-MRSA-IV clone (USA300 clone, two isolates, one ACME-positive) or ST22-MRSA-IV clone ("Regensburg EMRSA" clone, two isolates). All other clones were detected in one to six isolates and corresponded to well-known clones (e.g., Pediatric clone, Dublin EMRSA clone, WA MRSA-54/63, WA MRSA-1/57). This work highlighted both the high prevalence of ST239-MRSA-III clone and the large diversity of the other MRSA clones detected in a university hospital in Istanbul.

The High Diversity of MRSA Clones Detected in a University Hospital in Istanbul

PubMed Central

Oksuz, Lutfiye; Dupieux, Celine; Tristan, Anne; Bes, Michele; Etienne, Jerome; Gurler, Nezahat

2013-01-01

Background: To characterize the methicillin-resistant Staphylococcus aureus (MRSA) clones present in Istanbul, 102 MRSA isolates collected during a 5-year period at the Istanbul Medical Faculty Hospital were characterized using microarray analysis and phenotypic resistance profiles. Methods: Resistance to methicillin was detected with a cefoxitin disk diffusion assay and confirmed with a MRSA-agar and MRSA detection kit. Antimicrobial susceptibility testing was performed by a disk diffusion assay and interpreted according to the 2012 guidelines of the Antibiogram Committee of the French Society for Microbiology. Decreased susceptibility to glycopeptides was confirmed using the population analysis profile-area under the curve (PAP-AUC) method. The presence of the mecA gene was detected by polymerase chain reaction. Bacterial DNA was extracted according to the manufacturer's recommended protocol using commercial extraction kits. Strains were extensively characterized using the DNA microarray. Results: Isolates were grouped into six clonal complexes. The most frequently detected clone was the Vienna/Hungarian/Brazilian clone (ST239-MRSA-III), which accounted for 53.9% of the isolates. These isolates were resistant to multiple antibiotics, particularly penicillin, tetracycline, rifampicin, kanamycin, tobramycin, gentamicin, levofloxacin, erythromycin, lincomycin and fosfomycin. Furthermore, three isolates were detected by population analysis profile as heterogeneous vancomycin-intermediate S. aureus (hVISA). The UK-EMRSA-15 clone (ST22-MRSA-IV PVL negative) was detected in 9.8% of the isolates and was mainly susceptible to all anti-staphylococcal antibiotics. Seven isolates (6.9%) were positive for PVL genes and were assigned to the CC80-MRSA-IV clone (European CA-MRSA clone, three isolates), ST8-MRSA-IV clone (USA300 clone, two isolates, one ACME-positive) or ST22-MRSA-IV clone (“Regensburg EMRSA” clone, two isolates). All other clones were detected in one to six isolates and corresponded to well-known clones (e.g., Pediatric clone, Dublin EMRSA clone, WA MRSA-54/63, WA MRSA-1/57). Conclusions: This work highlighted both the high prevalence of ST239-MRSA-III clone and the large diversity of the other MRSA clones detected in a university hospital in Istanbul. PMID:24151444

Multifunctional polymeric nanoparticles doubly loaded with SPION and ceftiofur retain their physical and biological properties.

PubMed

Solar, Paula; González, Guillermo; Vilos, Cristian; Herrera, Natalia; Juica, Natalia; Moreno, Mabel; Simon, Felipe; Velásquez, Luis

2015-02-13

Advances in nanostructure materials are leading to novel strategies for drug delivery and targeting, contrast media for magnetic resonance imaging (MRI), agents for hyperthermia and nanocarriers. Superparamagnetic iron oxide nanoparticles (SPIONs) are useful for all of these applications, and in drug-release systems, SPIONs allow for the localization, direction and concentration of drugs, providing a broad range of therapeutic applications. In this work, we developed and characterized polymeric nanoparticles based on poly (3-hydroxybutyric acid-co-hydroxyvaleric acid) (PHBV) functionalized with SPIONs and/or the antibiotic ceftiofur. These nanoparticles can be used in multiple biomedical applications, and the hybrid SPION-ceftiofur nanoparticles (PHBV/SPION/CEF) can serve as a multifunctional platform for the diagnosis and treatment of cancer and its associated bacterial infections. Morphological examination using transmission electron microscopy (TEM) showed nanoparticles with a spherical shape and a core-shell structure. The particle size was evaluated using dynamic light scattering (DLS), which revealed a diameter of 243.0 ± 17 nm. The efficiency of encapsulation (45.5 ± 0.6% w/v) of these polymeric nanoparticles was high, and their components were evaluated using spectroscopy. UV-VIS, FTIR and DSC showed that all of the nanoparticles contained the desired components, and these compounds interacted to form a nanocomposite. Using the agar diffusion method and live/dead bacterial viability assays, we demonstrated that these nanoparticles have antimicrobial properties against Escherichia coli, and they retain their magnetic properties as measured using a vibrating sample magnetometer (VSM). Cytotoxicity was assessed in HepG2 cells using live/dead viability assays and MTS, and these assays showed low cytotoxicity with IC50 > 10 mg/mL nanoparticles. Our results indicate that hybrid and multifunctional PHBV/SPION/CEF nanoparticles are suitable as a superparamagnetic drug delivery system that can guide, concentrate and site-specifically release drugs with antibacterial activity.

Anti-inflammatory, antioxidant, and antimicrobial activities of Cocos nucifera var. typica.

PubMed

Silva, Rafaela Ribeiro; Oliveira e Silva, Davi; Fontes, Humberto Rollemberg; Alviano, Celuta Sales; Fernandes, Patricia Dias; Alviano, Daniela Sales

2013-05-16

Teas from the husk fiber of Cocos nucifera are used in the folk medicine to treat arthritis and other inflammatory processes. Some works show that some varieties have biological activities. However, one of the main variety of the species, C. nucifera var. typica, known in Brazil as "gigante", was not studied yet. Thus, this study evaluates if this variety has the anti-inflammatory and antimicrobial activities already reported in other varieties. C. nucifera aqueous crude extract (10, 50, and 100 mg/kg) and the reference drugs morphine (1 mg/kg) and acetylsalicylic acid (100 mg/kg) were evaluated in models of inflammation (formalin-induced licking and subcutaneous air pouch). The antioxidant activity was evaluated by 2,2-diphenyl-1-picryl-hydrazyl-hydrate (DPPH) photometric assay and compared with those of the standards (quercetin, rutin, and ascorbic acid). The extract was also screened against Candida albicans, Escherichia coli, Staphylococcus aureus, and methicillin-resistant Staphylococcus aureus (MRSA), in the agar diffusion method. The minimal inhibitory concentration (MIC) and minimal bactericidal concentration (MBC) were determined by the broth micro-dilution assay. Activities of combinations of the extract and antibiotics (methicillin or vancomycin) against MRSA were evaluated using checkerboard assays. The extract significantly inhibited the time that the animals spent licking the formalin-injected paws (second phase). The extract also inhibited the inflammatory process induced by subcutaneous carrageenan injection by reducing cell migration, protein extravasation, and TNF-α production. Additionally, the extract showed an antioxidant potential in vitro as good as standards in their antioxidant activity. The extract was active only against S. aureus and MRSA. MIC and the bactericidal concentrations were identical (1,024 μg/ml). The extract and methicillin acted synergistically against the clinical MRSA isolate, whereas an indifferent effect was detected when the extract was combined with vancomycin. The extract exhibits anti-inflammatory activity through the inhibition of the cell migration. The mixture of extract constituents and methicillin could lead to the development of a new combination antibiotic against MRSA infections.

Antibacterial activity of Artemisia asiatica essential oil against some common respiratory infection causing bacterial strains and its mechanism of action in Haemophilus influenzae.

PubMed

Huang, Jiehui; Qian, Chao; Xu, Hongjie; Huang, Yanjie

2018-01-01

The main objective of the current study was to investigate the chemical composition of the essential oil of Artemisia asiatica together with investigating the antibacterial effects it exerts on several common respiratory infection causing bacteria including Haemophilus influenzae. Its mechanism of action was studied using various state-of-the-art assays like scanning electron microscopy, DNA, RNA and protein leakage assays, growth curve assays etc. The essential oil was extracted from the leaves of A. asiatica by supercritical CO 2 fluid extraction technology. Chemical composition of essential oils was analyzed by gas chromatography-mass-spectrometry (GC-MS). The antibacterial activity was evaluated against 6 bacteria by the paper disc diffusion method. The minimum inhibitory concentration (MIC) and minimum bactericide concentration (MBC) values of the essential oil were estimated by agar dilution method. The antibacterial mechanism was evaluated by growth curve, the integrity of cell membrane and scanning electronmicroscope (SEM). Gas chromatographic analysis of the A. asiatica essential oil led to the identification of 16 chemical constituents accounting for 97.2% of the total oil composition. The major components were found to be Piperitone, (z)-davanone, p-cymene and 1, 8-cineole. The essential oil showed maximum growth inhibition against Haemophilus influenzae with a zone of inhibition of 24.5 mm and MIC/MBC values of 1.9/4.5 mg/mL respectively. Bacteria treated with the essential oil led to a rapid decrease in the number of viable cells. On adding the essential oil of A. asiatica to the bacterial culture, the constituents of the bacterial cell got released into the medium and this cell constituent release increased with increasing doses of the essential oil. SEM showed that the bacterial cells treated with the essential oil showed damaged cell wall, deformed cell morphology and shrunken cells. Copyright © 2017. Published by Elsevier Ltd.

Near-infrared fluorescence imaging platform for quantifying in vivo nanoparticle diffusion from drug loaded implants.

PubMed

Markovic, Stacey; Belz, Jodi; Kumar, Rajiv; Cormack, Robert A; Sridhar, Srinivas; Niedre, Mark

2016-01-01

Drug loaded implants are a new, versatile technology platform to deliver a localized payload of drugs for various disease models. One example is the implantable nanoplatform for chemo-radiation therapy where inert brachytherapy spacers are replaced by spacers doped with nanoparticles (NPs) loaded with chemotherapeutics and placed directly at the disease site for long-term localized drug delivery. However, it is difficult to directly validate and optimize the diffusion of these doped NPs in in vivo systems. To better study this drug release and diffusion, we developed a custom macroscopic fluorescence imaging system to visualize and quantify fluorescent NP diffusion from spacers in vivo. To validate the platform, we studied the release of free fluorophores, and 30 nm and 200 nm NPs conjugated with the same fluorophores as a model drug, in agar gel phantoms in vitro and in mice in vivo. Our data verified that the diffusion volume was NP size-dependent in all cases. Our near-infrared imaging system provides a method by which NP diffusion from implantable nanoplatform for chemo-radiation therapy spacers can be systematically optimized (eg, particle size or charge) thereby improving treatment efficacy of the platform.

Effects of Starvation on Physiological Activity and Chlorine Disinfection Resistance in Escherichia coli O157:H7

PubMed Central

Lisle, John T.; Broadaway, Susan C.; Prescott, Annette M.; Pyle, Barry H.; Fricker, Colin; McFeters, Gordon A.

1998-01-01

Escherichia coli O157:H7 can persist for days to weeks in microcosms simulating natural conditions. In this study, we used a suite of fluorescent, in situ stains and probes to assess the influence of starvation on physiological activity based on membrane potential (rhodamine 123 assay), membrane integrity (LIVE/DEAD BacLight kit), respiratory activity (5-cyano-2,3-di-4-tolyl-tetrazolium chloride assay), intracellular esterase activity (ScanRDI assay), and 16S rRNA content. Growth-dependent assays were also used to assess substrate responsiveness (direct viable count [DVC] assay), ATP activity (MicroStar assay), and culturability (R2A agar assay). In addition, resistance to chlorine disinfection was assessed. After 14 days of starvation, the DVC values decreased, while the values in all other assays remained relatively constant and equivalent to each other. Chlorine resistance progressively increased through the starvation period. After 29 days of starvation, there was no significant difference in chlorine resistance between control cultures that had not been exposed to the disinfectant and cultures that had been exposed. This study demonstrates that E. coli O157:H7 adapts to starvation conditions by developing a chlorine resistance phenotype. PMID:9835545

Qualitative and Quantitative Assays of Transposition and Homologous Recombination of the Retrotransposon Tf1 in Schizosaccharomyces pombe.

PubMed

Sangesland, Maya; Atwood-Moore, Angela; Rai, Sudhir K; Levin, Henry L

2016-01-01

Transposition and homologous recombination assays are valuable genetic tools to measure the production and integration of cDNA from the long terminal repeat (LTR) retrotransposon Tf1 in the fission yeast (Schizosaccharomyces pombe). Here we describe two genetic assays, one that measures the transposition activity of Tf1 by monitoring the mobility of a drug resistance marked Tf1 element expressed from a multi-copy plasmid and another assay that measures homologous recombination between Tf1 cDNA and the expression plasmid. While the transposition assay measures insertion of full-length Tf1 cDNA mediated by the transposon integrase, the homologous recombination assay measures levels of cDNA present in the nucleus and is independent of integrase activity. Combined, these assays can be used to systematically screen large collections of strains to identify mutations that specifically inhibit the integration step in the retroelement life cycle. Such mutations can be identified because they reduce transposition activity but nevertheless have wild-type frequencies of homologous recombination. Qualitative assays of yeast patches on agar plates detect large defects in integration and recombination, while the quantitative approach provides a precise method of determining integration and recombination frequencies.

Effects of starvation on physiological activity and chlorine disinfection resistance in Escherichia coli O157:H7

NASA Technical Reports Server (NTRS)

Lisle, J. T.; Broadaway, S. C.; Prescott, A. M.; Pyle, B. H.; Fricker, C.; McFeters, G. A.

1998-01-01

Escherichia coli O157:H7 can persist for days to weeks in microcosms simulating natural conditions. In this study, we used a suite of fluorescent, in situ stains and probes to assess the influence of starvation on physiological activity based on membrane potential (rhodamine 123 assay), membrane integrity (LIVE/DEAD BacLight kit), respiratory activity (5-cyano-2,3-di-4-tolyl-tetrazolium chloride assay), intracellular esterase activity (ScanRDI assay), and 16S rRNA content. Growth-dependent assays were also used to assess substrate responsiveness (direct viable count [DVC] assay), ATP activity (MicroStar assay), and culturability (R2A agar assay). In addition, resistance to chlorine disinfection was assessed. After 14 days of starvation, the DVC values decreased, while the values in all other assays remained relatively constant and equivalent to each other. Chlorine resistance progressively increased through the starvation period. After 29 days of starvation, there was no significant difference in chlorine resistance between control cultures that had not been exposed to the disinfectant and cultures that had been exposed. This study demonstrates that E. coli O157:H7 adapts to starvation conditions by developing a chlorine resistance phenotype.

Rapid High-Throughput Assessment of Aerobic Bacteria in Complex Samples by Fluorescence-Based Oxygen Respirometry

PubMed Central

O'Mahony, Fiach C.; Papkovsky, Dmitri B.

2006-01-01

A simple method has been developed for the analysis of aerobic bacteria in complex samples such as broth and food homogenates. It employs commercial phosphorescent oxygen-sensitive probes to monitor oxygen consumption of samples containing bacteria using standard microtiter plates and fluorescence plate readers. As bacteria grow in aqueous medium, at certain points they begin to deplete dissolved oxygen, which is seen as an increase in probe fluorescence above baseline signal. The time required to reach threshold signal is used to either enumerate bacteria based on a predetermined calibration or to assess the effects of various effectors on the growth of test bacteria by comparison with an untreated control. This method allows for the sensitive (down to a single cell), rapid (0.5 to 12 h) enumeration of aerobic bacteria without the need to conduct lengthy (48 to 72 h) and tedious colony counts on agar plates. It also allows for screening a wide range of chemical and environmental samples for their toxicity. These assays have been validated with different bacteria, including Escherichia coli, Micrococcus luteus, and Pseudomonas fluorescens, with the enumeration of total viable counts in broth and industrial food samples (packaged ham, chicken, and mince meat), and comparison with established agar plating and optical-density-at-600-nm assays has been given. PMID:16461677

Three-dimensional characterization of bacterial microcolonies on solid agar-based culture media.

PubMed

Drazek, Laurent; Tournoud, Maud; Derepas, Frédéric; Guicherd, Maryse; Mahé, Pierre; Pinston, Frédéric; Veyrieras, Jean-Baptiste; Chatellier, Sonia

2015-02-01

For the last century, in vitro diagnostic process in microbiology has mainly relied on the growth of bacteria on the surface of a solid agar medium. Nevertheless, few studies focused in the past on the dynamics of microcolonies growth on agar surface before 8 to 10h of incubation. In this article, chromatic confocal microscopy has been applied to characterize the early development of a bacterial colony. This technology relies on a differential focusing depth of the white light. It allows one to fully measure the tridimensional shape of microcolonies more quickly than classical confocal microscopy but with the same spatial resolution. Placing the device in an incubator, the method was able to individually track colonies growing on an agar plate, and to follow the evolution of their surface or volume. Using an appropriate statistical modeling framework, for a given microorganism, the doubling time has been estimated for each individual colony, as well as its variability between colonies, both within and between agar plates. A proof of concept led on four bacterial strains of four distinct species demonstrated the feasibility and the interest of the approach. It showed in particular that doubling times derived from early tri-dimensional measurements on microcolonies differed from classical measurements in micro-dilutions based on optical diffusion. Such a precise characterization of the tri-dimensional shape of microcolonies in their late-lag to early-exponential phase could be beneficial in terms of in vitro diagnostics. Indeed, real-time monitoring of the biomass available in a colony could allow to run well established microbial identification workflows like, for instance, MALDI-TOF mass-spectrometry, as soon as a sufficient quantity of material is available, thereby reducing the time needed to provide a diagnostic. Moreover, as done for pre-identification of macro-colonies, morphological indicators such as three-dimensional growth profiles derived from microcolonies could be used to perform a first pre-identification step, but in a shorten time. Copyright © 2014 Elsevier B.V. All rights reserved.

Isolation and characterization of a c-type lysozyme from the nurse shark.

PubMed

Hinds Vaughan, Nichole; Smith, Sylvia L

2013-12-01

Lysozyme is a ubiquitous antibacterial enzyme that occurs in numerous invertebrate and vertebrate species. Three forms have been described c-type, g-type and i-type which differ in primary structure. Shark lysozyme has not been characterized; here we report on the isolation and characterization of lysozyme from unstimulated shark (Ginglymostoma cirratum) leukocytes and provide amino acid sequence data across the highly conserved active site of the molecule identifying it to be a c-type lysozyme. A leukocyte lysate was applied either (a) to the first of two sequential DE-52 cellulose columns or alternatively, (b) to a DEAE-Sepharose column. Lysozyme activity in lysate and active fractions was identified by zones of lysis of Micrococcus lysodeikticus cell walls on lysoplates and zones of growth inhibition in agar diffusion assays using Planococcus citreus as the target organism. SDS-PAGE analysis revealed a 14 kDa protein which was identified as lysozyme by mass spectroscopic analysis of peptides, reactivity against anti-HEWL antibodies on a Western blot, hydrolysis of M. lysodeikticus cell walls, and inhibition of growth of P. citreus on AU-gel blots in which the area of growth inhibition correlated to a 14 kDa protein. Copyright © 2013 Elsevier Ltd. All rights reserved.

Characterization of a novel antibacterial glycopeptide produced by Penicillium sp. M03.

PubMed

Yang, W H; Zhang, W C; Lu, X M; Jiang, G S; Gao, P J

2009-04-01

To isolate a novel antibiotic termed AF from fermentation broth of Penicillium sp. M03 and to examine its antimicrobial activity, biological properties and structure characteristics. Sephadex LH-20 and HPLC were used to purify AF from fermentation broth of Penicillium sp. M03. The antimicrobial activity of AF was evaluated with the agar diffusion test. Amino acid and monosaccharide composition of AF was analysed by a HITACHI 835 detector and HPLC assay, respectively. Matrix-assisted laser desorption time of flight mass spectrometry, FT-IR and (1)H nuclear magnetic resonance spectra analyses were performed to examine the initial structure of AF. Eighty milligrams of AF was isolated as white powder from 1-l Penicillium sp. M03 fermentation broth. It consists of five amino acid and two monosaccharide residues and the molecular weight of it was 1017, and it was stable to beta-lactamase, heat, acid and alkali. AF showed inhibitory activity to a wide range of bacteria, particularly to multidrug-resistant Staphylococcus aureus. AF was a novel antibacterial glycopeptide with a broad inhibitory spectrum to pathogenic bacteria including multidrug-resistant agents. Furthermore, it is difficult to generate bacteria resistant to AF. Characterization of AF made it a potential antibiotic to fight against antibiotic-resistant bacterial pathogens.

Evaluation of antioxidant, antibacterial, and antidiabetic potential of two traditional medicinal plants of India: Swertia cordata and Swertia chirayita.

PubMed

Roy, Priyanka; Abdulsalam, Fatima I; Pandey, D K; Bhattacharjee, Aniruddha; Eruvaram, Naveen Reddy; Malik, Tabarak

2015-06-01

Swertia cordata and Swertia chirayita are temperate Himalayan medicinal plants used as potent herbal drugs in Indian traditional systems of medicine (Ayurvedic, Unani and Siddha). Assessment of Antioxidant, antibacterial, and antidiabetic potential of Swertia cordata and Swertia chirayita. Phytochemicals of methanolic and aqueous extracts of the two Swertia species were analyzed. The antioxidant potential of all the extracts was assessed by measuring total phenolic content, total flavonoid content and free radical scavenging potential was assessed by 1,1-diphenyl-2-picrilhydrazyl (DPPH) assay, antibacterial activity was assessed against various pathogenic and nonpathogenic bacteria in vitro by Kirby-Bauer agar well diffusion method and antidiabetic activity was assessed by α-amylase inhibition. Methanolic leaf extracts of both the species of Swertia contain significant antibacterial as well as anti-diabetic potential, whereas methanolic root extracts of both species were found to have potential antioxidant activity. However, Swertia chirayita showed better activities than Swertia cordata although both species have good reputation in traditional Indian medicine. Both the species are having high medicinal potential in terms of their antioxidant, antibacterial and antidiabetic activities. Studies are required to further elucidate antioxidant, anti-diabetic and antibacterial potentials using various in-vitro, in-vivo biochemical and molecular biology techniques.

Evaluation of antioxidant, antibacterial, and antidiabetic potential of two traditional medicinal plants of India: Swertia cordata and Swertia chirayita

PubMed Central

Roy, Priyanka; Abdulsalam, Fatima I.; Pandey, D. K.; Bhattacharjee, Aniruddha; Eruvaram, Naveen Reddy; Malik, Tabarak

2015-01-01

Background: Swertia cordata and Swertia chirayita are temperate Himalayan medicinal plants used as potent herbal drugs in Indian traditional systems of medicine (Ayurvedic, Unani and Siddha). Objective: Assessment of Antioxidant, antibacterial, and antidiabetic potential of Swertia cordata and Swertia chirayita. Materials and Methods: Phytochemicals of methanolic and aqueous extracts of the two Swertia species were analyzed. The antioxidant potential of all the extracts was assessed by measuring total phenolic content, total flavonoid content and free radical scavenging potential was assessed by 1,1-diphenyl-2-picrilhydrazyl (DPPH) assay, antibacterial activity was assessed against various pathogenic and nonpathogenic bacteria in vitro by Kirby-Bauer agar well diffusion method and antidiabetic activity was assessed by α-amylase inhibition. Results: Methanolic leaf extracts of both the species of Swertia contain significant antibacterial as well as anti-diabetic potential, whereas methanolic root extracts of both species were found to have potential antioxidant activity. However, Swertia chirayita showed better activities than Swertia cordata although both species have good reputation in traditional Indian medicine. Conclusion: Both the species are having high medicinal potential in terms of their antioxidant, antibacterial and antidiabetic activities. Studies are required to further elucidate antioxidant, anti-diabetic and antibacterial potentials using various in-vitro, in-vivo biochemical and molecular biology techniques. PMID:26109789

Antimicrobial Activity of a Neem Cake Extract in a Broth Model Meat System

PubMed Central

Del Serrone, Paola; Nicoletti, Marcello

2013-01-01

This work reports on the antimicrobial activity of an ethyl acetate extract of neem (Azadirachta indica) cake (NCE) against bacteria affecting the quality of retail fresh meat in a broth model meat system. NCE (100 µg) was also tested by the agar disc diffusion method. It inhibited the growth of all tested microorganisms. The NCE growth inhibition zone (IZ) ranged 11.33–22.67 mm while the ciprofloxacin (10 µg) IZ ranged from 23.41–32.67 mm. There was no significant difference (p ≤ 0.05) between the antimicrobial activity of NCE and ciprofloxacin vs. C. jejuni and Leuconostoc spp. The NCE antibacterial activity was moreover determined at lower concentrations (1:10–1:100,000) in micro-assays. The percent growth reduction ranged from 61 ± 2.08–92 ± 3.21. The higher bacterial growth reduction was obtained at 10 µg concentration of NCE. Species-specific PCR and multiplex PCR with the DNA dye propidium monoazide were used to directly detect viable bacterial cells from experimentally contaminated meat samples. The numbers of bacterial cells never significantly (p ≤ 0.05) exceeded the inocula concentration used to experimentally contaminate the NCE treated meat. This report represents a screening methodology to evaluate the antimicrobial capability of a herbal extract to preserve meat. PMID:23917814

Analysis of phytochemical profile of Terminalia arjuna bark extract with antioxidative and antimicrobial properties.

PubMed

Mandal, Shreya; Patra, Arpita; Samanta, Animesh; Roy, Suchismita; Mandal, Arpita; Mahapatra, Tapasi Das; Pradhan, Shrabani; Das, Koushik; Nandi, Dilip Kumar

2013-12-01

To investigate phytochemical screening, antimicrobial activity and qualitative thin layer chromatographic separation of flavonoid components, antioxidant activity and total flavonoid compound of Terminalia arjuna. For phytochemical screening, some common and available standard tests were done. Antimicrobial bioassay was done through agar well diffusion method. Detection of antioxidant activity and flavonoid compounds were done through thin layer chromatography. Total antioxidant activity was measured by 2, 2-diphenyl-1-picrylhydrazyl (DPPH) in colorimetric method. Aluminum chloride colorimetric method was used for total flavonoid determination. Phytochemical screening showed the active compounds presence in high concentration, such as phytosterol, lactones, flavonoids, phenolic compounds and tannins and glycosides. The antimicrobial activity of extract showed that greater inhibition zone against Gram negative bacteria than Gram positive bacteria. This methanolic extract showed a promising antioxidant activity, as absorption of DPPH redicles decreased in DPPH free radical scavenging assay. Flavonoids components having antioxidant property present in the methanol extract at a level of 199.00 mg quercetin equivalent/g of dried methanol extract in colorimetric method. The Terminalia arjuna bark extract revealed the presence of bio-active constituents which are known to exhibit medicinal as well as physiological activities. Copyright © 2013 Asian Pacific Tropical Biomedical Magazine. Published by Elsevier B.V. All rights reserved.

Antimicrobial activity of lactobacillus strains against uropathogens.

PubMed

Shim, Yoon Hee; Lee, Seung Joo; Lee, Jung Won

2016-10-01

The use of lactobacillus probiotics has been proposed as an alternative to prophylactic antibiotics for preventing urinary tract infection (UTI) in the era of antibiotic resistance. In this study, the antimicrobial activity of lactobacillus strains against uropathogens, was evaluated and compared with that of antibiotics. To evaluate inhibitory activities of lactobacilli against uropathogens, six lactobacillus strains (L. gasseri, L. rhamnosus, L. acidophilus, L. plantarum, L. paracasei, L. acidophilus) and four representative uropathogens of infantile UTI (extended-spectrum beta-lactamase [ESBL](-) Escherichia coli, ESBL(+) E. coli, Proteus vulgaris, Enterococcus fecalis) were selected. Lactobacillus strain in vitro inhibition of each uropathogen was evaluated on MRS agar well diffusion assay and compared with that of commercial antibiotic discs. Average inhibitory zone for each of the six lactobacillus strains against the four uropathogens showed slightly different but consistent inhibition (inhibitory zone diameter, 10.5-20.0 mm). This was different to that of the antibiotic discs, which had a wider range of inhibition (inhibitory zone diameter, <6.0-27.5 mm) depending on the uropathogen resistance pattern. The inhibitory zone of the six lactobacillus strains was between that of sensitive and resistant antibiotics (P < 0.05). Lactobacillus strains had similar moderate antimicrobial activities against uropathogens. Further research is needed to ascertain the strains with the best probiotic potential. © 2016 Japan Pediatric Society.

Antitumor and antimicrobial activities of endophytic fungi from medicinal parts of Aquilaria sinensis.

PubMed

Cui, Jin-long; Guo, Shun-xing; Xiao, Pei-gen

2011-05-01

The purpose of this study was to isolate and characterize endophytic fungi from the stem tissue which can produce fragrant ingredients in Aquilaria sinensis (also called agarwood) to determine their antitumor and antimicrobial activities. Twenty-eight fungal endophytes were isolated from agarwood by strict sterile sample preparation and were classified into 14 genera and 4 taxonomic classes (Sordariomycetes, Dothideomycetes, Saccharomycetes, and Zygomycetes) based on molecular identification. Of the 28 isolates, 13 (46.4%) showed antimicrobial activity against at least one of the test strains by the agar well diffusion method, and 23 isolates (82.1%) displayed antitumor activity against at least one of five cancer cell lines by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. The diameters of inhibition zones of YNAS07, YNAS14, HNAS04, HNAS05, HNAS08, and HNAS11 were equal to or higher than 14.0 mm against Staphylococcus aureus, Escherichia coli, Bacillus subtilis, B. subtilis, Aspergillus fumigatus, and B. subtilis, respectively. The inhibition rates of YNAS06, YNAS08, and HNAS06 were not less than 60% to 293-T, 293-T, and SKVO3 cells, respectively. These results suggest that the endophytic fungi associated with agarwood will provide us with not only useful micro-ecological information, but also potential antimicrobial and antitumor agents.

Antimicrobial activity of Lactobacillus strains of chicken origin against bacterial pathogenss.

PubMed

Dec, Marta; Puchalski, Andrzej; Nowaczek, Anna; Wernicki, Andrzej

2016-03-01

This study was conducted to identify and evaluate the antimicrobial activity of some Lactobacillus isolates of chicken origin. Among 90 isolates 14 Lactobacillus species were distinguished using MALDI-TOF mass spectrometry and 16S-ARDRA. The dominant species was L. salivarius (34.4%), followed by L. johnsonii (23.3%), L. crispatus (13.3%) and L. reuteri (11.1%). All lactobacilli were screened for antimicrobial activity against wild-type strains of Salmonella enterica, Escherichia coli, and Clostridium perfringens. Results from the agar slab method showed that all Lactobacillus isolates were able to produce active compounds on solid media with antagonistic properties against these pathogens. The highest sensitivity to lactobacilli was observed in C. perfringens strains, and the lowest in E. coli. Lactobacillus salivarius exhibited particularly strong antagonism towards all of the indicator bacteria. Strains of L. ingluviei and L. johnsonii and one strain of L. salivarius (10d) selectively inhibited the growth of C. perfringens. No antimicrobial activity of many Lactobacillus isolates was observed when cell-free culture supernatant was used in a well diffusion assay. All Lactobacillus isolates exhibited the ability to produce H2O2 and proved to be hydrophobic (excluding one of L. salivarius). [Int Microbiol 19(1):57-67 (2016)]. Copyright© by the Spanish Society for Microbiology and Institute for Catalan Studies.

Modelling and shadowgraph imaging of cocrystal dissolution and assessment of in vitro antimicrobial activity for sulfadimidine/4-aminosalicylic acid cocrystals.

PubMed

Serrano, Dolores R; Persoons, Tim; D'Arcy, Deirdre M; Galiana, Carolina; Dea-Ayuela, Maria Auxiliadora; Healy, Anne Marie

2016-06-30

The aim of this work was to evaluate the influence of crystal habit on the dissolution and in vitro antibacterial and anitiprotozoal activity of sulfadimidine:4-aminosalicylic acid cocrystals. Cocrystals were produced via milling or solvent mediated processes. In vitro dissolution was carried out in the flow-through apparatus, with shadowgraph imaging and mechanistic mathematical models used to observe and simulate particle dissolution. In vitro activity was tested using agar diffusion assays. Cocrystallisation via milling produced small polyhedral crystals with antimicrobial activity significantly higher than sulfadimidine alone, consistent with a fast dissolution rate which was matched only by cocrystals which were milled following solvent evaporation. Cocrystallisation by solvent evaporation (ethanol, acetone) or spray drying produced flattened, plate-like or quasi-spherical cocrystals, respectively, with more hydrophobic surfaces and greater tendency to form aggregates in aqueous media, limiting both the dissolution rate and in vitro activity. Deviation from predicted dissolution profiles was attributable to aggregation behaviour, supported by observations from shadowgraph imaging. Aggregation behaviour during dissolution of cocrystals with different habits affected the dissolution rate, consistent with in vitro activity. Combining mechanistic models with shadowgraph imaging is a valuable approach for dissolution process analysis. Copyright © 2016 Elsevier B.V. All rights reserved.

Chemical Composition and Biological Activities of the Essential Oil from Leaves and Flowers of Pulicaria incisa sub. candolleana (Family Asteraceae).

PubMed

Shahat, Esraa A; Bakr, Riham O; Eldahshan, Omayma A; Ayoub, Nahla A

2017-04-01

The composition of the essential oil isolated from leaves and flowers of Pulicaria incisa sub. candolleana E. Gamal-Eldin, growing in Egypt, was analysed by GC and GC-MS. Forty-nine and 68 compounds were identified from the oils of the leaves and flowers accounting for 86.69 and 84.29%, respectively of the total detected constituents. Both leaves and flowers oils were characterized by the high content of carvotanacetone with 66.01, 50.87 and chrysanthenone 13.26, 24.3%, respectively. The cytotoxic activity of both essential oils was evaluated against hepatocellular carcinoma cell line HEPG-2, using MTT assay and vinblastine as a reference drug. Leaf oil showed higher activity with IC 50 11.4 μg/ml compared with 37.4 μg/ml for flower oil. The antimicrobial activity of both oils was evaluated using agar well diffusion method towards two representatives for each of Gram positive and Gram negative bacteria as well as four representatives for fungi. The minimum inhibitory concentration of both essential oils against bacterial and fungal strains was obtained in the range of 0.49 - 15.63 μg/ml. © 2017 Wiley-VHCA AG, Zurich, Switzerland.

In vitro evaluation of antimicrobial and antioxidant activities of methanolic extract of Jasminum humile leaves.

PubMed

Nain, Parminder; Kumar, Ashok; Sharma, Sunil; Nain, Jaspreet

2011-10-01

To evaluate in vitro antimicrobial and antioxidant activities of methanolic extract of Jasminum humile (J. humile) leaves extract. Methanolic extract of J. humile was evaluated for its antimicrobial activity by using agar well diffusion method & their possible antioxidant assay by two complementary test systems, namely DPPH and hydrogen peroxide scavenging activity. These various antioxidant activities were compared to standard antioxidants such as ascorbic acid for both the tests. In the DPPH & hydrogen peroxide scavenging activity, the IC(50) value of methanol extract was 70.43 μg/mL & 60.79 μg/mL respectively. Further, the extract showed inhibitory activity for Gram-positive and negative bacteria at different concentrations. The maximum antibacterial activity of extract was exhibited against Staphylococcus aureus (S. aureus) at concentration 50 mg/mL when compared with ciprofloxacin These results clearly indicate that J. humile is effective in scavenging free radicals and has the potential to be a powerful antioxidant. Thus, the results obtained in the present study indicate that J. humile leaves extract could be considered as a potential source of natural antioxidants and that could be used as an effective source against bacterial diseases. Copyright © 2011 Hainan Medical College. Published by Elsevier B.V. All rights reserved.

Antimicrobial properties of lactic acid bacteria isolated from traditional yogurt and milk against Shigella strains.

PubMed

Zare Mirzaei, Elnaze; Lashani, Elahe; Davoodabadi, Abolfazl

2018-01-01

Background: Lactic acid bacteria (LAB) are normal flora of the mouth, intestines and the female genital tract. They are also frequently found in meat, vegetables, and dairy products. Most of probiotic bacteria belong to the LAB group. Some probiotic LAB are useful in prevention and treatment of diarrheal diseases. The aim of this study was to investigate the antimicrobial properties of LAB isolated from traditional yogurt and milk against Shigella strains. Materials and methods: Forty LAB strains were isolated from traditional yogurt and milk. The antimicrobial activity of LAB against Shigella strains (eight S. flexneri , four S. sonnei ) was examined using the agar-well diffusion assay. LAB strains with antimicrobial effect against all Shigella strains were identified by 16S rRNA gene sequencing. Results: Six LAB strains inhibited the growth of all 12 Shigella strains. Lb. paracasei Y1-3, Lb. paracasei Y8-1 and Lb. fermentum Y2-2 were isolated from yogurt. Lb. paracasei M18-1, Lb. parelimentarius M4-3 and Lb. plantarum M19-1 were isolated from milk. Conclusion: This study showed that Lactobacillus strains with good inhibitory activity against S. flexneri and S. sonnei could be isolated from traditional yogurt and milk.

Biofilms of Lactobacillus plantarum and Lactobacillus fermentum: Effect on stress responses, antagonistic effects on pathogen growth and immunomodulatory properties.

PubMed

Aoudia, Nabil; Rieu, Aurélie; Briandet, Romain; Deschamps, Julien; Chluba, Johanna; Jego, Gaëtan; Garrido, Carmen; Guzzo, Jean

2016-02-01

Few studies have extensively investigated probiotic functions associated with biofilms. Here, we show that strains of Lactobacillus plantarum and Lactobacillus fermentum are able to grow as biofilm on abiotic surfaces, but the biomass density differs between strains. We performed microtiter plate biofilm assays under growth conditions mimicking to the gastrointestinal environment. Osmolarity and low concentrations of bile significantly enhanced Lactobacillus spatial organization. Two L. plantarum strains were able to form biofilms under high concentrations of bile and mucus. We used the agar well-diffusion method to show that supernatants from all Lactobacillus except the NA4 isolate produced food pathogen inhibitory molecules in biofilm. Moreover, TNF-α production by LPS-activated human monocytoid cells was suppressed by supernatants from Lactobacillus cultivated as biofilms but not by planktonic culture supernatants. However, only L. fermentum NA4 showed anti-inflammatory effects in zebrafish embryos fed with probiotic bacteria, as assessed by cytokine transcript level (TNF-α, IL-1β and IL-10). We conclude that the biofilm mode of life is associated with beneficial probiotic properties of lactobacilli, in a strain dependent manner. Those results suggest that characterization of isolate phenotype in the biofilm state could be additional valuable information for the selection of probiotic strains. Copyright © 2015 Elsevier Ltd. All rights reserved.

Antiproliferative and antimicrobial activity of traditional Kombucha and Satureja montana L. Kombucha.

PubMed

Cetojevic-Simin, D D; Bogdanovic, G M; Cvetkovic, D D; Velicanski, A S

2008-01-01

To carry out a preliminary investigation of the biological activity of Kombucha beverages from Camellia sinensis L. (black tea) and Satureja montana L. (winter savory tea), that have consuming acidity. Cell growth effect was measured by sulforhodamine B colorimetric assay on HeLa (cervix epithelioid carcinoma), HT-29 (colon adenocarcinoma), and MCF-7 (breast adenocarcinoma). Antimicrobial activity to bacteria, yeasts and moulds was determined by agar-well diffusion method. Consuming Kombuchas had the most expressive antimicrobial activity against all investigated bacteria, except Sarcina lutea, while unfermented tea samples had no activity. Traditional Kombucha showed higher activity against Staphylococcus aureus and Escherichia coli than acetic acid, while both neutralized Kombuchas had bacteriostatic activity on Salmonella enteritidis. Examined Kombuchas did not stimulate cell proliferation of the investigated cell lines. Antiproliferative activity of winter savory tea Kombucha was comparable to that of traditional Kombucha made from black tea. Furthermore, in HeLa cell line Satureja montana L. Kombucha induced cell growth inhibition by 20% (IC20) at lower concentration compared to the activity of water extract of Satureja montana L. obtained in our previous research. Presence of more active antiproliferative component(s) in Satureja montana L. Kombucha compared to Satureja montana L. water extract and antimicrobial component(s) other than acetic acid in both Kombuchas is suggested.

Safety, formulation, and in vitro antiviral activity of the antimicrobial peptide subtilosin against herpes simplex virus type 1

PubMed Central

Torres, Nicolás I.; Noll, Katia Sutyak; Xu, Shiqi; Li, Ji; Huang, Qingrong; Sinko, Patrick J.; Wachsman, Mónica B.; Chikindas, Michael L.

2013-01-01

In the present study the antiviral properties of the bacteriocin subtilosin against Herpes simplex virus type 1 (HSV-1) and the safety and efficacy of a subtilosin-based nanofiber formulation were determined. High concentrations of subtilosin, the cyclical antimicrobial peptide produced by Bacillus amyloliquefaciens, were virucidal against HSV-1. Interestingly, at non-virucidal concentrations, subtilosin inhibited wild type HSV-1 and aciclovir-resistant mutants in a dose-dependent manner. Although the exact antiviral mechanism is not fully understood, time of addition experiments and western blot analysis suggest that subtilosin does not affect viral multiplication steps prior to protein synthesis. Poly(vinyl alcohol) (PVOH)-based subtilosin nanofibers with a width of 278 nm were produced by the electrospinning process. The retained antimicrobial activity of the subtilosin-based fibers was determined via an agar well diffusion assay. The loading capacity of the fibers was 2.4 mg subtilosin/g fiber, and loading efficiency was 31.6%. Furthermore, the nanofibers with and without incorporated subtilosin were shown to be nontoxic to human epidermal tissues using an in vitro human tissue model. Taking together these results subtilosin-based nanofibers should be further studied as a novel alternative method for treatment and/or control of HSV-1 infection. PMID:23637711

Comparative synthesis and antimicrobial action of silver nanoparticles and silver nitrate

NASA Astrophysics Data System (ADS)

Mosselhy, Dina A.; El-Aziz, Mohamed Abd; Hanna, Magdy; Ahmed, Mohamed A.; Husien, Mona M.; Feng, Qingling

2015-12-01

The high wave of antibiotic bacterial resistance has addressed an importance for administration of different antibacterial agents, as silver nanoparticles (Ag NPs). However, many investigators still suffer conflict in the mechanistic antimicrobial action of Ag NPs and Ag+ ions. In this regard, our study investigated the comparative antimicrobial action of different sizes of Ag NPs as 8 (nAg1) and 29 (nAg2) nm, in comparison with silver nitrate (AgNO3) against five different bacterial species; Aeromonas hydrophila ( A. hydrophila), Pseudomonas putida ( Ps. putida), Escherichia coli ( E. coli), Staphylococcus aureus ( S. aureus), and Bacillus subtilis ( B. subtilis) using agar diffusion assay and minimum inhibitory concentration (MIC). The key role of the size of nanomaterials was detected, as the smaller Ag NPs (nAg1) showed more antimicrobial action than the larger particles. Transmission electron microscopy (TEM) studies demonstrated the different mechanistic antibacterial actions of Ag NPs and AgNO3. The effect of combining Ag NPs with antibiotics was also investigated. Synergistic effect of combining Ag NPs with ampicillin was detected against S. aureus, in a size-dependent manner as well. To summarize, our results point towards the major role played by the size of Ag NPs in their antimicrobial effects and the different toxic mechanisms of actions induced by Ag NPs and AgNO3.

Effect of high pressure on the microbiological quality of cooked chicken during storage at normal and abuse refrigeration temperatures.

PubMed

Patterson, Margaret F; McKay, Alan M; Connolly, Malachy; Linton, Mark

2010-04-01

Vacuum-packaged cooked poultry meat was treated at a range of pressures (400-600 MPa) and hold times (1, 2 and 10 min), followed by storage at 4 degrees , 8 degrees or 12 degrees C for up to 35 days. Weissella viridescens was found to be the dominant microorganism in the pressure-treated meat, constituting 100% of the microflora identified at 500 and 600 MPa. None of the pressure-treated samples had obvious signs of spoilage during the 35 day storage period, even when the Weissella count was >7 log(10) cfu/g. Studies on a typical W. viridescens isolate showed it to be relatively pressure-resistant in poultry meat, with <1 log reduction in numbers after a treatment of 2 min at 600 MPa. Agar diffusion assays showed that the isolate also caused the inhibition of a number of Gram-positive and Gram-negative pathogens, including strains of Clostridium botulinum, Listeria monocytogenes, Bacillus cereus and Escherichia coli. The selection of a pressure-resistant organism, such as this Weissella sp. could be advantageous in extending the shelf-life, and also microbiological safety, of the cooked meat, as it could give protection in addition to the pressure treatment itself.

A penicillin- and metronidazole-resistant Clostridium botulinum strain responsible for an infant botulism case.

PubMed

Mazuet, C; Yoon, E-J; Boyer, S; Pignier, S; Blanc, T; Doehring, I; Meziane-Cherif, D; Dumant-Forest, C; Sautereau, J; Legeay, C; Bouvet, P; Bouchier, C; Quijano-Roy, S; Pestel-Caron, M; Courvalin, P; Popoff, M R

2016-07-01

The clinical course of a case of infant botulism was characterized by several relapses despite therapy with amoxicillin and metronidazole. Botulism was confirmed by identification of botulinum toxin and Clostridium botulinum in stools. A C. botulinum A2 strain resistant to penicillins and with heterogeneous resistance to metronidazole was isolated from stool samples up to 110 days after onset. Antibiotic susceptibility was tested by disc agar diffusion and MICs were determined by Etest. Whole genome sequencing allowed detection of a gene cluster composed of blaCBP for a novel penicillinase, blaI for a regulator, and blaR1 for a membrane-bound penicillin receptor in the chromosome of the C. botulinum isolate. The purified recombinant penicillinase was assayed. Resistance to β-lactams was in agreement with the kinetic parameters of the enzyme. In addition, the β-lactamase gene cluster was found in three C. botulinum genomes in databanks and in two of 62 genomes of our collection, all the strains belonging to group I C. botulinum. This is the first report of a C. botulinum isolate resistant to penicillins. This stresses the importance of antibiotic susceptibility testing for adequate therapy of botulism. Copyright © 2016 European Society of Clinical Microbiology and Infectious Diseases. Published by Elsevier Ltd. All rights reserved.

Cloning and expression of antibacterial goat lactoferricin from Escherichia coli AD494(DE3)pLysS expression system.

PubMed

Chen, Gen-Hung; Yin, Li-Jung; Chiang, I-Hua; Jiang, Shann-Tzong

2008-12-01

Goat lactoferricin (GLfcin), an antibacterial peptide, is released from the N terminus of goat lactoferrin by pepsin digestion. Two GLfcin-related cDNAs, GLfcin L and GLfcin S, encoding Ala20-Ser60 and Ser36-Ser60 of goat lactoferrin, respectively, were cloned into the pET-23a(+) expression vector upstream from (His)6-Tag gene and transformed into Escherichia coli AD494(DE3)pLysS expression host. After being induced by isopropyl-beta-D-thiogalactopyranoside (IPTG), two (His)6-Tag fused recombinant lactoferricins, GLfcin L-His*Tag and GLfcin S-His*Tag, were expressed in soluble form within the E. coli cytoplasm. The GLfcin L-His*Tag and GLfcin S-His*Tag were purified using HisTrap affinity chromatography. According to an antibacterial activity assay using the agar diffusion method, GLfcin L-His*Tag had antibacterial activity against E. coli BCRC 11549, Staphylococcus aureus BCRC 25923, and Propionibacterium acnes BCRC 10723, while GLfcin S-His*Tag was able to inhibit the growth of E. coli BCRC 11549 and P. acnes BCRC 10723. These two recombinant lactoferricins behaved as thermostable peptides, which could retain their activity for up to 30 min of exposure at 100 degrees C.

Chemical compositions and antimicrobial potential of Actinodaphne macrophylla leaves oils from East Kalimantan

NASA Astrophysics Data System (ADS)

Putri, A. S.; Purba, F. F.; Kusuma, I. W.; Kuspradini, H.

2018-04-01

Essential oils producing plants comprises about 160-200 species, one of which belongs to Lauraceae family. Actinodaphne macrophylla is a plant of the Lauraceae family and widely spread on Kalimantan island. For humans, essential oils are used in cosmetics industry, food industry, and pharmaceutical industry. This research aimed to analyze the characteristics of essential oil and potential of antimicrobial activity from A. macrophylla leaves oils. Essential oils were obtained by steam distillation method. Antimicrobial activity was assayed using agar diffusion method which compared with two synthetic standards including chlorhexidine and chloramphenicol. Four microorganisms were used in this study were Candida albicans, Staphylococcus aureus, Streptococcus mutans, and Streptococcus sobrinus. The obtained oil was determined for its characteristics including the yield, refractive index, and chemical components. The attained components were analyzed using GC-MS. The results of this study showed that essential oils of A. macrophylla leaves contained 0.1051% of yield, clearless, and refractive index was 1.425. Based on GC-MS analysis result, it showed chemical components including spathulenol, 2-monopalmitin, (+)-sabinene, copaen, camphene, and β-pinene. This plant potentially can inhibit the growth of S. aureus, C. albicans, S. sobrinus, and S. mutans with inhibition zones of 17.22, 20.89, 22.34 and 22.89 mm, respectively.

Catalytically and biologically active silver nanoparticles synthesized using essential oil

NASA Astrophysics Data System (ADS)

Vilas, Vidya; Philip, Daizy; Mathew, Joseph

2014-11-01

There are numerous reports on phytosynthesis of silver nanoparticles and various phytochemicals are involved in the reduction and stabilization. Pure explicit phytosynthetic protocol for catalytically and biologically active silver nanoparticles is of importance as it is an environmentally benign green method. This paper reports the use of essential oil of Myristica fragrans enriched in terpenes and phenyl propenes in the reduction and stabilization. FTIR spectra of the essential oil and the synthesized biogenic silver nanoparticles are in accordance with the GC-MS spectral analysis reports. Nanosilver is initially characterized by an intense SPR band around 420 nm, followed by XRD and TEM analysis revealing the formation of 12-26 nm sized, highly pure, crystalline silver nanoparticles. Excellent catalytic and bioactive potential of the silver nanoparticles is due to the surface modification. The chemocatalytic potential of nanosilver is exhibited by the rapid reduction of the organic pollutant, para nitro phenol and by the degradation of the thiazine dye, methylene blue. Significant antibacterial activity of the silver colloid against Gram positive, Staphylococcus aureus (inhibition zone - 12 mm) and Gram negative, Escherichia coli (inhibition zone - 14 mm) is demonstrated by Agar-well diffusion method. Strong antioxidant activity of the biogenic silver nanoparticles is depicted through NO scavenging, hydrogen peroxide scavenging, reducing power, DPPH and total antioxidant activity assays.

Headspace oxygen as a hurdle to improve the safety of in-pack pasteurized chilled food during storage at different temperatures.

PubMed

Muñoz, Nydia; Bhunia, Kanishka; Zhang, Hongchao; Barbosa-Cánovas, Gustavo V; Tang, Juming; Sablani, Shyam

2017-07-17

This study investigated the use of headspace oxygen in a model food system to prevent the growth of anaerobic pathogenic bacteria in in-pack pasteurized food at various storage temperatures. Three model food formulations prepared with tryptic soy broth and three agar concentrations (0.1, 0.4 and 1.0%) were sealed without removing the air from the package in high oxygen barrier pouches (OTR=0.3cm 3 /m 2 ·day·atm). Important properties influencing bacterial growth, including pH and water activity (a w ) were determined. The oxygen sorption kinetics of each model food were obtained at three different storage temperatures (8, 12, and 20°C) using an OxySense Gen III 300 system. An analytical solution of Fick's second law was used to determine the O 2 diffusion coefficient. Growth challenge studies at 12 and 20°C were conducted at three selected locations (top, center and bottom layers) in model foods containing 1% agar. Model foods were inoculated with Clostridium sporogenes PA 3679 (300spores/mL), and were classified as low-acid (pH>4.5, a w >0.85). When the storage temperature decreased from 20 to 8°C, the oxygen diffusion decreased from 0.82×10 -9 m 2 /s to 0.68×10 -9 m 2 /s. As the agar concentration was increased from 0.1 to 1.0%, the effective oxygen permeability decreased significantly (p=0.007) from 0.88×10 -9 m 2 /s to 0.65×10 -9 m 2 /s. When the inoculated model foods were stored at 12°C for 14days, C. sporogenes PA 3679 was unable to grow. As the storage temperature was increased to 20°C, significant bacterial growth was observed with storage time (p<0.0001), and the C. sporogenes PA 3679 population increased by around 6logCFU/g. However, the location of the food did not influence the growth distribution of C. sporogenes PA 3679. These results demonstrate that oxygen diffusion from the pouch headspace was primarily limited to the food surface. Findings suggest that the air/oxygen present in the package headspace may not be considered as a food safety hurdle in the production of pasteurized packaged food. Copyright © 2017 Elsevier B.V. All rights reserved.

The role of virus dose in experimental bovine leukemia virus infection in sheep.

PubMed

Stirtzinger, T; Valli, V E; Miller, J M

1988-04-01

Twenty-four, six month old lambs were assembled into four groups of five animals each and one group of four animals. All groups were inoculated with lymphocytes from a single donor lamb infected with bovine leukemia virus. The inoculum varied from 250 to 250,000 lymphocytes, in tenfold increments. Animals were exposed by intradermal injection in the neck region immediately anterior to the left shoulder joint. All groups were monitored at 0, 3, 7 and 12 weeks after inoculation using the following procedures: a. Syncytia induction assay for detection of bovine leukemia virus in peripheral blood lymphocytes. b. Agar gel immunodiffusion against the gp51 antigen of bovine leukemia virus for the detection of antibovine leukemia virus gp51 antibody. c. Lymphocyte stimulation test for the assessment of cell-mediated immunity using mitogen, nonfractionated bovine leukemia virus antigen, and partially purified bovine lymphoma tumor-associated antigen for the in vitro activation of lymphocytes from bovine leukemia virus-inoculated and sham-inoculated, control animals. d. Routine hematological techniques for the assessment of total leukocyte and lymphocyte counts. The median infectious dose for lymphocytes from the single bovine leukemia virus-infected donor used in this study was determined to be 2000 cells. The syncytia induction assay detected more infected individuals (13/23) at an earlier time than did the agar gel immunodiffusion assay (10/23). Using either serological or virus isolation techniques, infected animals were first detected at three weeks postinoculation in the group receiving the high-dose inoculum and at seven weeks postinoculation in groups receiving low- or medium-dose inocula.(ABSTRACT TRUNCATED AT 250 WORDS)

Diversity and antimicrobial activity of culturable fungi from fishscale bamboo (Phyllostachys heteroclada) in China.

PubMed

Zhou, Ying-Ke; Shen, Xiao-Ye; Hou, Cheng-Lin

2017-06-01

An important and useful bamboo species, fishscale bamboo (Phyllostachys heteroclada Oliver), is broadly distributed in Southeast China and has multiple purposes, including uses in cuisine, weaving, Chinese medicine and ecological protection. However, no previous studies have focused on the endophytes of this plant. In our article, a total of 127 fungal strains were first isolated from the healthy branches and leaves of common P. heteroclada. These endophytic fungi could be directly categorized into 50 morphotypes according to their culture characteristics, and their internal transcribed spacer (ITS) regions were analyzed for molecular identification. Using the BLAST search tool of the NCBI database and phylogenetic tree analysis, these isolates were divided into two phyla, Ascomycota (95.28%) and Basidiomycota (4.72%), including at least six orders (Xylariales, Capnodiales, Pleosporales, Hypocreales, Chaetothyriales and Polyporales) and fourteen genera (Arthrinium, Pestalotiopsis, Epicoccum, Cladosporium, Nigrospora, Setophoma, Didymella, Calcarisporium, Preussia, Nemania, Creosphaeria, Ophiobolus, Phialophora and Perenniporia). It is fascinating that four genera, Calcarisporium, Preussia, Creosphaeria and Phialophora were isolated from bamboos for the first time. The inhibitory effects against clinical pathogens were also preliminarily screened, and four isolates FB43 (Calcarisporium arbuscula), FB06 (Preussia minima), FB16 (Setophoma sp.) and FB21 (Perenniporia medulla-pains) among the candidate strains displayed broad-spectrum activities according to the agar diffusion method and the disk diffusion assay. Strain FB16 (Setophoma sp.) especially indicated high bioactivity against both clinical bacteria and yeast. This study is the first report on the diversity and antimicrobial activity of the endophytic fungi associated with P. heteroclada, which could be regarded as a potential source of drug precursors and could be used in biocontrol development.

Biological activity of the essential oils from Cinnamodendron dinisii and Siparuna guianensis

PubMed Central

Andrade, Milene Aparecida; Cardoso, Maria das Graças; Gomes, Marcos de Souza; de Azeredo, Camila Maria Oliveira; Batista, Luís Roberto; Soares, Maurilio José; Rodrigues, Leonardo Milani Avelar; Figueiredo, Ana Cristina S.

2015-01-01

This study had analyzed the antibacterial, antifungal and trypanocidal activity of the essential oils from Cinnamodendron dinisii Schwacke (Canellaceae) and Siparuna guianensis Aublet (Siparunaceae). The essential oils were obtained from fresh leaves by hydrodistillation, using a modified Clevenger apparatus. Chemical analysis by gas-liquid chromatography coupled to mass spectrometry (GC-MS) showed that these essential oils are rich in monoterpene and sesquiterpene hydrocarbons. Activity against the pathogenic bacteria Escherichia coli , Listeria monocytogenes , Pseudomonas aeruginosa , Salmonella choleraesuis and Staphylococcus aureus was evaluated with the agar cavity diffusion method, while activity on the filamentous fungi Aspergillus flavus , Aspergillus niger , Aspergillus carbonarius and Penicillium commune was evaluated by the disk diffusion technique. Trypanocidal activity was tested against Trypanosoma cruzi epimastigotes, using the Tetrazolium salt (MTT) colorimetric assay. Both essential oils exhibited low inhibitory effect towards bacteria, showing high MIC values (125–500 μg mL −1 ), with Gram positive bacteria being more susceptible. Better inhibitory effect was obtained for the evaluated fungi, with lower MIC values (7.81–250 μg mL −1 ), being A. flavus the most susceptible species. Both essential oils presented low trypanocidal activity, with IC 50 /24 h values of 209.30 μg mL −1 for S. guianensis and 282.93 μg mL −1 for C. dinisii . Thus, the high values observed for the MIC of evaluated bacteria and for IC 50 /24 h of T. cruzi , suggest that the essential oils have a low inhibitory activity against these microorganisms. In addition, the low MIC values observed for the tested fungi species indicate good inhibitory activity on these microorganisms’s growth. PMID:26221107

Synthesis and antibacterial activity of aromatic and heteroaromatic amino alcohols.

PubMed

de Almeida, Camila G; Reis, Samira G; de Almeida, Angelina M; Diniz, Claudio G; da Silva, Vânia L; Le Hyaric, Mireille

2011-11-01

Two series of aromatic and heteroaromatic amino alcohols were synthesized from alcohols and aldehydes and evaluated for their antibacterial activities. All the octylated compounds displayed a better activity against the four bacteria tested when evaluated by the agar diffusion method and were selected for the evaluation of minimal inhibitory concentration. The best results were obtained for p-octyloxybenzyl derivatives against Staphylococcus epidermidis (minimal inhibitory concentrations = 32 μm). © 2011 John Wiley & Sons A/S.

Standardized methods and quality control limits for agar and broth microdilution susceptibility testing of Mycoplasma pneumoniae, Mycoplasma hominis, and Ureaplasma urealyticum.

PubMed

Waites, Ken B; Duffy, Lynn B; Bébéar, Cécile M; Matlow, Anne; Talkington, Deborah F; Kenny, George E; Totten, Patricia A; Bade, Donald J; Zheng, Xiaotian; Davidson, Maureen K; Shortridge, Virginia D; Watts, Jeffrey L; Brown, Steven D

2012-11-01

An international multilaboratory collaborative study was conducted to develop standard media and consensus methods for the performance and quality control of antimicrobial susceptibility testing of Mycoplasma pneumoniae, Mycoplasma hominis, and Ureaplasma urealyticum using broth microdilution and agar dilution techniques. A reference strain from the American Type Culture Collection was designated for each species, which was to be used for quality control purposes. Repeat testing of replicate samples of each reference strain by participating laboratories utilizing both methods and different lots of media enabled a 3- to 4-dilution MIC range to be established for drugs in several different classes, including tetracyclines, macrolides, ketolides, lincosamides, and fluoroquinolones. This represents the first multilaboratory collaboration to standardize susceptibility testing methods and to designate quality control parameters to ensure accurate and reliable assay results for mycoplasmas and ureaplasmas that infect humans.

Antibacterial and antioxidant activities of Musa sp. leaf extracts against multidrug resistant clinical pathogens causing nosocomial infection

PubMed Central

Karuppiah, Ponmurugan; Mustaffa, Muhammed

2013-01-01

Objective To investigate different Musa sp. leave extracts of hexane, ethyl acetate and methanol were evaluated for antibacterial activity against multi-drug resistant pathogens causing nosocomial infection by agar well diffusion method and also antioxidant activities. Methods The four different Musa species leaves were extracted with hexane, ethyl acetate and methanol. Antibacterial susceptibility test, minimum inhibitory concentration and minimum inhibitory bacterial concentration were determined by agar well diffusion method. Total phenolic content and in vitro antioxidant activity was determined. Results All the Musa sp. extracts showed moderate antibacterial activities expect Musa paradisiaca with the inhibition zone ranging from 8.0 to 18.6 mm. Among four species ethyl acetate extracts of Musa paradisiaca showed highest activity against tested pathogens particularly E. coli, P. aeruginosa and Citrobacter sp. The minimum inhibitory concentrations were within the value of 15.63- 250 µg/mL and minimum bactericidal concentrations were ranging from 31.25- 250 µg/mL. Antioxidant activity of Musa acuminate exhibited maximum activity among other three Musa species. Conclusions The present study concluded that among the different Musa species, Musa paradisiaca displayed efficient antibacterial activity followed by Musa acuminata against multi-drug resistant nosocomial infection causing pathogens. Further, an extensive study is needed to identify the bioactive compounds, mode of action and toxic effect in vivo of Musa sp. PMID:23998016

Antibacterial and antioxidant activities of Musa sp. leaf extracts against multidrug resistant clinical pathogens causing nosocomial infection.

PubMed

Karuppiah, Ponmurugan; Mustaffa, Muhammed

2013-09-01

To investigate different Musa sp. leave extracts of hexane, ethyl acetate and methanol were evaluated for antibacterial activity against multi-drug resistant pathogens causing nosocomial infection by agar well diffusion method and also antioxidant activities. The four different Musa species leaves were extracted with hexane, ethyl acetate and methanol. Antibacterial susceptibility test, minimum inhibitory concentration and minimum inhibitory bacterial concentration were determined by agar well diffusion method. Total phenolic content and in vitro antioxidant activity was determined. All the Musa sp. extracts showed moderate antibacterial activities expect Musa paradisiaca with the inhibition zone ranging from 8.0 to 18.6 mm. Among four species ethyl acetate extracts of Musa paradisiaca showed highest activity against tested pathogens particularly E. coli, P. aeruginosa and Citrobacter sp. The minimum inhibitory concentrations were within the value of 15.63- 250 µg/mL and minimum bactericidal concentrations were ranging from 31.25- 250 µg/mL. Antioxidant activity of Musa acuminate exhibited maximum activity among other three Musa species. The present study concluded that among the different Musa species, Musa paradisiaca displayed efficient antibacterial activity followed by Musa acuminata against multi-drug resistant nosocomial infection causing pathogens. Further, an extensive study is needed to identify the bioactive compounds, mode of action and toxic effect in vivo of Musa sp.

Evaluation of the Antibacterial Activity of Bergamot Essential Oils on Different Listeria Monocytogenes Strains

PubMed Central

Marotta, Stefania M.; Giarratana, Filippo; Parco, Alessio; Neri, Domenico; Ziino, Graziella; Giuffrida, Alessandro; Panebianco, Antonio

2016-01-01

Essential oils are aromatic and volatile substances extracted from plants and characterized by antimicrobial activity. The aim of the present study was to evaluate the antibacterial activity (agar disc-diffusion method) of seven different bergamot essential oils (BEOs) on eight Listeria monocytogenes strains. Minimal inhibitory concentration (MIC) of most efficient BEOs was estimated. Extremely variable results for agar disc-diffusion method for L. monocytogenes strains were reported. One of the tested microorganisms resulted insensible to all the BEOs; 3 strains showed an inhibition from weak to null and the remaining 4 a variable susceptibility. Among the BEOs tested, one showed a strong activity against four pathogenic strains. Four BEOs revealed weak, moderate or null activity in all the 7 sensitive strains, while for two oils only a weak or no activity was reported. MIC values were 0.625 μL/mL for the most efficient BEO, 2.5 and 5 μL/mL for the other samples that showed moderate inhibition. Experiment results are significantly related to the strains tested (P<0.01), rather than the BEO employed (P>0.01). In conclusion, we can consider BEO as a natural technological hurdle for Listeria monocytogenes in combination with other preservation strategies. Finally, this study underlines the necessity to evaluate the antimicrobial activity of EOs on a significant strains number of the same bacteria. PMID:28058251

Evaluation of the Antibacterial Activity of Bergamot Essential Oils on Different Listeria Monocytogenes Strains.

PubMed

Marotta, Stefania M; Giarratana, Filippo; Parco, Alessio; Neri, Domenico; Ziino, Graziella; Giuffrida, Alessandro; Panebianco, Antonio

2016-09-20

Essential oils are aromatic and volatile substances extracted from plants and characterized by antimicrobial activity. The aim of the present study was to evaluate the antibacterial activity (agar disc-diffusion method) of seven different bergamot essential oils (BEOs) on eight Listeria monocytogenes strains. Minimal inhibitory concentration (MIC) of most efficient BEOs was estimated. Extremely variable results for agar disc-diffusion method for L. monocytogenes strains were reported. One of the tested microorganisms resulted insensible to all the BEOs; 3 strains showed an inhibition from weak to null and the remaining 4 a variable susceptibility. Among the BEOs tested, one showed a strong activity against four pathogenic strains. Four BEOs revealed weak, moderate or null activity in all the 7 sensitive strains, while for two oils only a weak or no activity was reported. MIC values were 0.625 μL/mL for the most efficient BEO, 2.5 and 5 μL/mL for the other samples that showed moderate inhibition. Experiment results are significantly related to the strains tested (P<0.01), rather than the BEO employed (P>0.01). In conclusion, we can consider BEO as a natural technological hurdle for Listeria monocytogenes in combination with other preservation strategies. Finally, this study underlines the necessity to evaluate the antimicrobial activity of EOs on a significant strains number of the same bacteria.

Differential action and differential expression of DNA polymerase I during Escherichia coli colony development.

PubMed Central

Shapiro, J A

1992-01-01

A mini-Tn10 insertion in the polA cistron (polA2099) was isolated in a search for mutations that affect patterned Mudlac replication in colonies. The polA2099 mutation had a dramatic effect on cell morphogenesis during the first few hours of microcolony development. Abnormal microcolonies containing filamentous cells were produced as a result of SOS induction. Despite gross abnormalities in early microcolonies, mature polA2099 colonies after 2 to 4 days were morphologically indistinguishable from Pol+ colonies, and 44-h polA2099 colonies displayed a cell size distribution very similar to that of Pol+ colonies. These results suggested the involvement of a protective factor produced during colony growth that compensated for the polA deficiency. The action of a diffusible substance that stimulates growth of polA2099 microcolonies was shown by spotting dilute polA2099 cultures next to established colonies. Differential transcription of polA during colony development was visualized by growing colonies containing polA-lacZ fusions on beta-galactosidase indicator agar. When polA-lacZ colonies were inoculated next to established colonies, a diffusible factor was seen to inhibit polA transcription during the earliest stages of colony development. These results show that a basic housekeeping function, DNA polymerase I, is subject to multicellular control by the changing conditions which the bacteria create as they proliferate on agar. Images PMID:1331025

[A new zoonosis--investigation of Gardnerella vaginalis disease of fox. V. Studies serotype of Gardnerella vaginalis in fox].

PubMed

Yan, X; Yan, Z; Yan, X; Luan, F; Wang, C

1996-10-01

145 strains Gardnerella vaginalis isolated in foxes were isolated from 13 main farms raising foxes in six provinces (regions), China, after antigenicity and immunogenicity of the strains were measured, 1-3 appropriate strains were selected from each farm raising foxes for serotype studies. Cross agglutinin absorption test confirmed that selected 26 strains Gardnerella vaginalis were divided into three serotypes and then the representing strains were used to produce typing serum. Among remaining 119 strain, 108 strains were typable with the typing sera, and 11 strains can't be set. Among three serotypes, serotype I made up 79.1% of the strains. It was shown that serotype I was the principal serotype of Gardnerella vaginalis of fox in China. The test also confirmed that 5 strains of Gardnerella vaginalis isolated from racoon dog, 4 strain Gardnerella vaginalis from mink and 2 strains Gardnerella vaginalis from canine also belonged to serotype I. Supersonic antigben was produced with three serotypes, representative strains. By agar immuno-diffusion test, it confirmed that the antigens of three serotypes formed a obvious blending precipitating line with the homologous or heterologous serotype antiserum. It indicated common antigen existed among all serotypes. The agar immuno-diffusion test results revealed that the precipitating line of the homologous serotype completely blended. It is our opinion that the method of serotyping is reliable.

Development of a high-throughput liquid state assay for lipase activity using natural substrates and rhodamine B.

PubMed

Zottig, Ximena; Meddeb-Mouelhi, Fatma; Beauregard, Marc

2016-03-01

A fluorescence-based assay for the determination of lipase activity using rhodamine B as an indicator, and natural substrates such as olive oil, is described. It is based on the use of a rhodamine B-natural substrate emulsion in liquid state, which is advantageous over agar plate assays. This high-throughput method is simple and rapid and can be automated, making it suitable for screening and metagenomics application. Reaction conditions such as pH and temperature can be varied and controlled. Using triolein or olive oil as a natural substrate allows monitoring of lipase activity in reaction conditions that are closer to those used in industrial settings. The described method is sensitive over a wide range of product concentrations and offers good reproducibility. Copyright © 2015 Elsevier Inc. All rights reserved.

Eliminating Size-Associated Diffusion Constraints for Rapid On-Surface Bioassays with Nanoparticle Probes.

PubMed

Li, Junwei; Zrazhevskiy, Pavel; Gao, Xiaohu

2016-02-24

Nanoparticle probes enable implementation of advanced on-surface assay formats, but impose often underappreciated size-associated constraints, in particular on assay kinetics and sensitivity. The present study highlights substantially slower diffusion-limited assay kinetics due to the rapid development of a nanoprobe depletion layer next to the surface, which static incubation and mixing of bulk solution employed in conventional assay setups often fail to disrupt. In contrast, cyclic solution draining and replenishing yields reaction-limited assay kinetics irrespective of the probe size. Using common surface bioassays, enzyme-linked immunosorbent assays and immunofluorescence, this study shows that this conceptually distinct approach effectively "erases" size-dependent diffusion constraints, providing a straightforward route to rapid on-surface bioassays employing bulky probes and procedures involving multiple labeling cycles, such as multicycle single-cell molecular profiling. For proof-of-concept, the study demonstrates that the assay time can be shortened from hours to minutes with the same probe concentration and, at a typical incubation time, comparable target labeling can be achieved with up to eight times lower nanoprobe concentration. The findings are expected to enable realization of novel assay formats and stimulate development of rapid on-surface bioassays with nanoparticle probes. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Phytochemical, Antimicrobial, and Toxicological Evaluation of Traditional Herbs Used to Treat Sore Throat.

PubMed

Mehreen, Arifa; Waheed, Muzzamil; Liaqat, Iram; Arshad, Najma

2016-01-01

The in vitro antibacterial activities of 29 traditional medicinal plants used in respiratory ailments were assessed on multidrug resistant Gram-positive and Gram-negative bacteria isolated from the sore throat patients and two reference strains. The methanolic, n-hexane, and aqueous extracts were screened by the agar well diffusion assay. Bioactive fractions of effective extracts were identified on TLC coupled with bioautography, while their toxicity was determined using haemolytic assay against human erythrocytes. Qualitative and quantitative phytochemical analysis of effective extracts was also performed. Methanolic extract of 18 plants showed antimicrobial activity against test strains. Adhatoda vasica (ZI = 17-21 mm, MIC: 7.12-62.5 μg/mL), Althaea officinalis (ZI = 16-20 mm, MIC: 15.62-31.25 μg/mL), Cordia latifolia (ZI = 16-20 mm, MIC: 12.62-62.5 μg/mL), Origanum vulgare (ZI = 20-22 mm, MIC: 3-15.62 μg/mL), Thymus vulgaris (ZI = 21-25 mm, MIC: 7.81-31.25 μg/mL), and Ziziphus jujuba (ZI = 14-20 mm, MIC: 7.81-31.25 μg/mL) showed significant antibacterial activity. Alkaloid fractions of Adhatoda vasica, Cordia latifolia, and Origanum vulgare and flavonoid fraction of the Althaea officinalis, Origanum vulgare, Thymus Vulgaris, and Ziziphus jujuba exhibited antimicrobial activity. Effective plant extracts show 0.93-0.7% erythrocyte haemolysis. The results obtained from this study provide a scientific rationale for the traditional use of these herbs and laid the basis for future studies to explore novel antimicrobial compounds.

Phytochemical, Antimicrobial, and Toxicological Evaluation of Traditional Herbs Used to Treat Sore Throat

PubMed Central

Mehreen, Arifa; Waheed, Muzzamil; Liaqat, Iram; Arshad, Najma

2016-01-01

The in vitro antibacterial activities of 29 traditional medicinal plants used in respiratory ailments were assessed on multidrug resistant Gram-positive and Gram-negative bacteria isolated from the sore throat patients and two reference strains. The methanolic, n-hexane, and aqueous extracts were screened by the agar well diffusion assay. Bioactive fractions of effective extracts were identified on TLC coupled with bioautography, while their toxicity was determined using haemolytic assay against human erythrocytes. Qualitative and quantitative phytochemical analysis of effective extracts was also performed. Methanolic extract of 18 plants showed antimicrobial activity against test strains. Adhatoda vasica (ZI = 17–21 mm, MIC: 7.12–62.5 μg/mL), Althaea officinalis (ZI = 16–20 mm, MIC: 15.62–31.25 μg/mL), Cordia latifolia (ZI = 16–20 mm, MIC: 12.62–62.5 μg/mL), Origanum vulgare (ZI = 20–22 mm, MIC: 3–15.62 μg/mL), Thymus vulgaris (ZI = 21–25 mm, MIC: 7.81–31.25 μg/mL), and Ziziphus jujuba (ZI = 14–20 mm, MIC: 7.81–31.25 μg/mL) showed significant antibacterial activity. Alkaloid fractions of Adhatoda vasica, Cordia latifolia, and Origanum vulgare and flavonoid fraction of the Althaea officinalis, Origanum vulgare, Thymus Vulgaris, and Ziziphus jujuba exhibited antimicrobial activity. Effective plant extracts show 0.93–0.7% erythrocyte haemolysis. The results obtained from this study provide a scientific rationale for the traditional use of these herbs and laid the basis for future studies to explore novel antimicrobial compounds. PMID:27429983

Evaluation of Antimicrobial and Antifungal efficacy of Chitosan as endodontic irrigant against Enterococcus Faecalis and Candida Albicans Biofilm formed on tooth substrate.

PubMed

Yadav, Pankaj; Chaudhary, Sarika; Saxena, Rajendra K; Talwar, Sangeeta; Yadav, Sudha

2017-03-01

Bacterial biofilms formed on the root canal wall are often difficult to remove. This study aimed to evaluate the cytotoxic effect and antibacterial efficacy of chitosan when used as root canal irrigant against E. Faecalis and Candida albicans biofilm formed on tooth substrate. The present study evaluated antibacterial effect of 0.25% Chitosan, 0.5% Chitosan, 2% chlorhexidine and 3% sodium hypochlorite against Enterococcus faecalis and Candida Albicans . Agar-well diffusion methods, minimal inhibitory concentration tests and biofilm susceptibility assays were used to determine antibacterial activity. Teeth specimens were sectioned to obtain a standardized tooth length of 12mm. Specimens were inoculated with 10 mL of the freshly prepared E. Faecalis suspension and Candida albicans for 4 weeks. The specimens were then instrumented with ProTaper rotary files F3 size. After irrigation with test solution, three sterile paper points were placed into one canal, left for 60 s and transferred to a test tube containing 1 mL of reduced transport fluid. The number of CFU in 1 mL was determined. 3-week biofilm qualitative assay showed complete inhibition of bacterial growth with 3% Sodium hypochlorite, 2% Chlorhexidine and Chitosan except saline, which showed presence of bacterial growth. Significant reduction of colony forming units (CFU)/mL was observed for the chitosan groups and the antibacterial activity of the chitosan groups was at par with 3% NaOCl and 2% Chlorhexidine. It was observed that the chitosan showed no cytotoxicity at 3mg/ml and 10% cytotoxicity at 6mg/ml. The use of chitosan as a root canal irrigant might be an alternative considering the various undesirable properties of NaOCl and chlorhexidine. Key words: Biofilm, Candida albicans, Chitosan, Cytotoxicity, Enterococcus faecalis.

Long-Term Fungal Inhibition by Pisum sativum Flour Hydrolysate during Storage of Wheat Flour Bread.

PubMed

Rizzello, Carlo Giuseppe; Lavecchia, Anna; Gramaglia, Valerio; Gobbetti, Marco

2015-06-15

In order to identify antifungal compounds from natural sources to be used as ingredients in the bakery industry, water/salt-soluble extracts (WSE) from different legume flour hydrolysates obtained by the use of a fungal protease were assayed against Penicillium roqueforti DPPMAF1. The agar diffusion assays allowed the selection of the pea (Pisum sativum) hydrolysate as the most active. As shown by the hyphal radial growth rate, the WSE had inhibitory activity towards several fungi isolated from bakeries. The MIC of the WSE was 9.0 mg/ml. Fungal inhibition was slightly affected by heating and variations in pH. The antifungal activity was attributed to three native proteins (pea defensins 1 and 2 and a nonspecific lipid transfer protein [nsLTP]) and a mixture of peptides released during hydrolysis. The three proteins have been reported previously as components of the defense system of the plant. Five peptides were purified from WSE and were identified as sequences encrypted in leginsulin A, vicilin, provicilin, and the nsLTP. To confirm antifungal activity, the peptides were chemically synthesized and tested. Freeze-dried WSE were used as ingredients in leavened baked goods. In particular, breads made by the addition of 1.6% (wt/wt) of the extract and fermented by baker's yeast or sourdough were characterized for their main chemical, structural, and sensory features, packed in polyethylene bags, stored at room temperature, and compared to controls prepared without pea hydrolysate. Artificially inoculated slices of a bread containing the WSE did not show contamination by fungi until at least 21 days of storage and behaved like the bread prepared with calcium propionate (0.3%, wt/wt). Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Chemical composition and in vitro antioxidant and antibacterial activity of Heracleum transcaucasicum and Heracleum anisactis roots essential oil

PubMed Central

Torbati, Mohammadali; Nazemiyeh, Hossein; Lotfipour, Farzaneh; Nemati, Mahboob; Asnaashari, Solmaz; Fathiazad, Fatemeh

2014-01-01

Introduction: In vitro antioxidant and antibacterial activity and volatile compositions of two Heracleum species (Apiaceae) including Heracleum transcaucasicum and Heracleum anisactis roots Essential Oil (EO) were investigated. Methods: The volatile compositions of EOs were analyzed by GC/Mass spectroscopy. To detect the antioxidant activity of essential oils TLC-bioautography and DPPH radical scavenging assay by spectrophotometry was performed. Additionally, the antibacterial activity of two essential oils were studied and compared against four pathogenic bacteria by agar disc diffusion method and MIC values of the EOs were determined using the broth dilution method. Results: Myristicin was the dominant component in both EOs. It was identified as 96.87% and 95.15% of the essential oil composition of H. transcaucasicum and H. anisactis roots, respectively. The TLC-bioautography showed antioxidant spots in both EOs and IC50 of H. anisactis and H. transcaucasicum EO was found to be 54 μg × ml (-1) and 77 μg × ml (-1), respectively. Regarding the antimicrobial assay, H. anisactis EO exhibited weak to moderate antibacterial activity against gram-positive bacteria and also Escherichia coli, whereas the essential oil from H. transcaucasicum was inactive. Conclusion: Based on the results from this study, both tested EOs mainly consist of myristicin. Despite the presence of myristicin with known antibacterial property, the EO from H. transcacausicum showed no antibacterial activity. Thus it is supposed that the biological activity of plants is remarkably linked to the extracts’ chemical profile and intercomponents’ synergistic or antagonistic effect could play a crucial role in bioactivity of EOs and other plant extracts. PMID:25035849

Antioxidant and biocidal activities of Carum nigrum (seed) essential oil, oleoresin, and their selected components.

PubMed

Singh, Gurdip; Marimuthu, Palanisamy; de Heluani, Carola S; Catalan, Cesar A N

2006-01-11

In the present study, chemical constituents of the essential oil and oleoresin of the seed from Carum nigrum obtained by hydrodistillation and Soxhlet extraction using acetone, respectively, have been studied by GC and GC-MS techniques. The major component was dillapiole (29.9%) followed by germacrene B (21.4%), beta-caryophyllene (7.8%), beta-selinene (7.1%), and nothoapiole (5.8%) along with many other components in minor amounts. Seventeen components were identified in the oleoresin (Table 2) with dillapiole as a major component (30.7%). It also contains thymol (19.1%), nothoapiole (15.2.3%), and gamma-elemene (8.0%). The antioxidant activity of both the essential oil and oleoresin was evaluated in mustard oil by monitoring peroxide, thiobarbituric acid, and total carbonyl and p-anisidine values of the oil substrate. The results showed that both the essential oil and oleoresin were able to reduce the oxidation rate of the mustard oil in the accelerated condition at 60 degrees C in comparison with synthetic antioxidants such as butylated hydroxyanisole and butylated hydroxytoluene at 0.02%. In addition, individual antioxidant assays such as linoleic acid assay, DPPH scavenging activity, reducing power, hydroxyl radical scavenging, and chelating effects have been used. The C. nigrum seed essential oil exhibited complete inhibition against Bacillus cereus and Pseudomonas aeruginosa at 2000 and 3000 ppm, respectively, by agar well diffusion method. Antifungal activity was determined against a panel of foodborne fungi such as Aspergillus niger, Penicillium purpurogenum, Penicillium madriti, Acrophialophora fusispora, Penicillium viridicatum, and Aspergillus flavus. The fruit essential oil showed 100% mycelial zone inhibition against P. purpurogenum and A. fusispora at 3000 ppm in the poison food method. Hence, both oil and oleoresin could be used as an additive in food and pharmaceutical preparations after screening.

Amish burn ointment and burdock leaf dressings: assessments of antimicrobial and cytotoxic activities.

PubMed

Rieman, Mary T; Neely, Alice N; Boyce, Steven T; Kossenjans, William J; Durkee, Paula J; Zembrodt, Jacquelyn M; Puthoff, Barbara K; Kagan, Richard J

2014-01-01

Amish burn wound ointment (ABO) contains honey, lanolin, oils, glycerin, bees wax, and other natural additives. Although there are many anecdotal reports that this ointment covered with a burdock leaf (BL) dressing promotes burn wound healing, little scientific testing of this treatment has occurred. The goal of this study was to evaluate in vitro some of the components of this treatment modality for antimicrobial and cytotoxic activities. The ABO was tested for sterility using standard microbiological techniques. Because of the semisolid, lipid-based nature of the salve, the at-use product could not be tested in bioassays. Samples of BL and the dry ingredients (DI) used in the ointment were provided by the Amish vendor. Aqueous extracts of the DI and of the BL were prepared and freeze dried. The freeze-dried extracts were reconstituted, filtered, and tested separately on keratinocyte and fibroblast cell cultures for cytotoxicity (growth inhibition assay) and against a panel of susceptible and resistant microbes for antimicrobial activity (Nathan's agar-well diffusion assay) in a series of concentrations (% wt/vol). Neither DI nor BL extracts demonstrated antimicrobial activity against any of organisms tested. The DI extract inhibited growth of both keratinocytes and fibroblasts at the 0.1% concentration. The 0.1 and 0.03% concentrations of the BL extract were cytotoxic to both keratinocytes and fibroblasts. Although tests for microbial growth from the at-use preparation of the ABO were negative, extracts of the DI and BL did not demonstrate any antimicrobial activity. Additionally, both extracts inhibited the growth of skin cells in vitro at higher concentrations. These results suggest caution in the use of ABO and BL dressings if there is more than a minimal risk of complications from the burn injury.

Antimicrobial, antioxidant and cytotoxic properties of essential oil from Dictamnus angustifolius.

PubMed

Sun, Jianbo; Wang, Xinzhi; Wang, Pei; Li, Linzhen; Qu, Wei; Liang, Jingyu

2015-01-15

Dictamnus angustifolius (Rutaceae) has been used as an alternative for folk medicine, Dictamnus dasycarpus in the treatment of rheumatism, bleeding, itching, jaundice, chronic hepatitis, and skin diseases in Xinjiang Province of China. The aim of this study was to evaluate the antimicrobial and antioxidant activities of essential oil (EO) from Dictamnus angustifolius, correlated with their chemical composition and evaluate their cytotoxicity. The EO were extracted by water-distillation using a Clevenger-type apparatus. The chemical composition of EO was identified by GC-MS analysis. The antimicrobial activity was evaluated against five microorganisms by the agar disc diffusion method and minimal inhibitory concentration (MIC) assay. The antioxidant activity was measured by employing DPPH and FRAP assays. The cytotoxic activity was evaluated in the mammalian cells lines A549, MCF7, B16 and LoVo using the MTT method to assess cell viability. 52 compounds representing the 97.2% of the total oil were identified by GC/MS. The major constituents of the oil were tetramethylenecyclobutane (42.07%) and fraxinellone (19.06%). The antimicrobial activity showed that the EO possess significant inhibition in Monilia albican ATCC 10231 and Staphylococcus aureus ATCC 6538. The antioxidant activity suggested that the EO possess significant reducing power. The cytotoxic activity of the EO in MCF7 and LoVo cells was significant stronger than in the other cell lines. This study is the first characterization of the chemical composition and biological activities of EO from Dictamnus angustifolius. All experimental data indicate that the EO have not only remarkable antioxidant properties but also potential antimicrobial activity and cytotoxic activities. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

Improved protease stability of the antimicrobial peptide Pin2 substituted with D-amino acids.

PubMed

Carmona, G; Rodriguez, A; Juarez, D; Corzo, G; Villegas, E

2013-08-01

Cationic antimicrobial peptides (AMPs) have attracted a great interest as novel class of antibiotics that might help in the treatment of infectious diseases caused by pathogenic bacteria. However, some AMPs with high antimicrobial activities are also highly hemolytic and subject to proteolytic degradation from human and bacterial proteases that limit their pharmaceutical uses. In this work a D-diastereomer of Pandinin 2, D-Pin2, was constructed to observe if it maintained antimicrobial activity in the same range as the parental one, but with the purpose of reducing its hemolytic activity to human erythrocytes and improving its ability to resist proteolytic cleavage. Although, the hydrophobic and secondary structure characteristics of L- and D-Pin2 were to some extent similar, an important reduction in D-Pin2 hemolytic activity (30-40 %) was achieved compared to that of L-Pin2 over human erythrocytes. Furthermore, D-Pin2 had an antimicrobial activity with a MIC value of 12.5 μM towards Staphylococcus aureus, Escherichia coli, Streptococcus agalactiae and two strains of Pseudomonas aeruginosa in agar diffusion assays, but it was half less potent than that of L-Pin2. Nevertheless, the antimicrobial activity of D-Pin2 was equally effective as that of L-Pin2 in microdilution assays. Yet, when D- and L-Pin2 were incubated with trypsin, elastase and whole human serum, only D-Pin2 kept its antimicrobial activity towards all bacteria, but in diluted human serum, L- and D-Pin2 maintained similar peptide stability. Finally, when L- and D-Pin2 were incubated with proteases from P. aeruginosa DFU3 culture, a clinical isolated strain, D-Pin2 kept its antibiotic activity while L-Pin2 was not effective.

Purification and characterization of the bacteriocin Thuricin Bn1 produced by Bacillus thuringiensis subsp. kurstaki Bn1 isolated from a hazelnut pest.

PubMed

Ugras, Serpil; Sezen, Kazim; Kati, Hatice; Demirbag, Zihni

2013-02-01

A novel bioactive molecule produced by Bacillus thuringiensis subsp. kurstaki Bn1 (Bt-Bn1), isolated from a common pest of hazelnut, Balaninus nucum L. (Coleoptera: Curculionidae), was determined, purified, and characterized in this study. The Bt-Bn1 strain was investigated for antibacterial activity with an agar spot assay and well diffusion assay against B. cereus, B. weinhenstephenensis, L. monocytogenes, P. savastanoi, P. syringae, P. lemoignei, and many other B. thuringiensis strains. The production of bioactive molecule was determined at the early logarithmic phase in the growth cycle of strain Bt-Bn1 and its production continued until the beginning of the stationary phase. The mode of action of this molecule displayed bacteriocidal or bacteriolytic effect depending on the concentration. The bioactive molecule was purified 78-fold from the bacteria supernatant with ammonium sulfate precipitation, dialysis, ultrafiltration, gel filtration chromatography, and HPLC, respectively. The molecular mass of this molecule was estimated via SDS-PAGE and confirmed by the ESI-TOFMS as 3,139 Da. The bioactive molecule was also determined to be a heat-stable, pH-stable (range 6-8), and proteinase K sensitive antibacterial peptide, similar to bacteriocins. Based on all characteristics determined in this study, the purified bacteriocin was named as thuricin Bn1 because of the similarities to the previously identified thuricin-like bacteriocin produced by the various B. thuringiensis strains. Plasmid elution studies showed that gene responsible for the production of thuricin Bn1 is located on the chromosome of Bt-Bn1. Therefore, it is a novel bacteriocin and the first recorded one produced by an insect originated bacterium. It has potential usage for the control of many different pathogenic and spoilage bacteria in the food industry, agriculture, and various other areas.

Comparative studies on polyphenolic profile and antimicrobial activity of propolis samples selected from distinctive geographical areas of Hungary.

PubMed

Molnár, Szabolcs; Mikuska, Kata; Patonay, Katalin; Sisa, Krisztina; Daood, Hussein G; Némedi, Erzsébet; Kiss, Attila

2017-06-01

The present paper reports about a comparative survey on the chemical composition, antioxidant activity and in vitro antimicrobial activity of selected propolis samples collected in Hungary. The total levels of polyphenolic compounds including flavonoids in ethanolic extracts of propolis were assessed. The major constituents of ethanolic extracts of propolis were analysed by gas chromatography/mass spectrometry analysis. Total phenolic content was determined spectrophotometrically using a Folin-Ciocalteu reagent. Free radical scavenging activities were evaluated by means of 2,2-diphenyl-1-picrylhydrazyl assay. In vitro inhibitory activity was investigated against eight different bacterial strains by agar well diffusion assay. An extensive comparison was carried out regarding general parameters and specific polyphenolic components. The experimental data led to the observation that there is considerable variability in terms of the quality and the biological value of the distinctive propolis samples. These findings confirm the hypothesis of the study; versatile experimental results are required for proper, well-reasoned, balanced and standardised industrial applications. The major flavonoid components were found to be chrysin and pinocembrin; however, versatile minor components were also detected. The total polyphenol content of ethanolic extracts of propolis ranged between 104.6 mg/g and 286.9 mg/g (gallic acid equivalent). The radical scavenging activity of ethanolic extracts of propolis varied between 101.7 mg/g and 286.9 mg/g (ascorbic acid equivalent). As the quality of propolis depends on the season, vegetation and the area of collection, marked differences were found among the different products examined in terms of both composition and general characteristics. The studied samples exhibited significant differences in term of antimicrobial activities.

Long-Term Fungal Inhibitory Activity of Water-Soluble Extracts of Phaseolus vulgaris cv. Pinto and Sourdough Lactic Acid Bacteria during Bread Storage▿

PubMed Central

Coda, Rossana; Rizzello, Carlo G.; Nigro, Franco; De Angelis, Maria; Arnault, Philip; Gobbetti, Marco

2008-01-01

The antifungal activity of proteinaceous compounds from different food matrices was investigated. In initial experiments, water-soluble extracts of wheat sourdoughs, cheeses, and vegetables were screened by agar diffusion assays with Penicillium roqueforti DPPMAF1 as the indicator fungus. Water-soluble extracts of sourdough fermented with Lactobacillus brevis AM7 and Phaseolus vulgaris cv. Pinto were selected for further study. The crude water-soluble extracts of L. brevis AM7 sourdough and P. vulgaris cv. Pinto had a MIC of 40 mg of peptide/ml and 30.9 mg of protein/ml, respectively. MICs were markedly lower when chemically synthesized peptides or partially purified protein fractions were used. The water-soluble extract of P. vulgaris cv. Pinto showed inhibition toward a large number of fungal species isolated from bakeries. Phaseolin alpha-type precursor, phaseolin, and erythroagglutinating phytohemagglutinin precursor were identified in the water-soluble extract of P. vulgaris cv. Pinto by nano liquid chromatography-electrospray ionization-tandem mass spectrometry. When the antifungal activity was assayed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, all three proteins were inhibitory. A mixture of eight peptides was identified from the water-soluble extract of sourdough L. brevis AM7, and five of these exhibited inhibitory activity. Bread was made at the pilot plant scale by sourdough fermentation with L. brevis AM7 and addition of the water-soluble extract (27%, vol/wt; 5 mg of protein/ml) of P. vulgaris cv. Pinto. Slices of bread packed in polyethylene bags did not show contamination by fungi until at least 21 days of storage at room temperature, a level of protection comparable to that afforded by 0.3% (wt/wt) calcium propionate. PMID:18849463

Long-term fungal inhibitory activity of water-soluble extracts of Phaseolus vulgaris cv. Pinto and sourdough lactic acid bacteria during bread storage.

PubMed

Coda, Rossana; Rizzello, Carlo G; Nigro, Franco; De Angelis, Maria; Arnault, Philip; Gobbetti, Marco

2008-12-01

The antifungal activity of proteinaceous compounds from different food matrices was investigated. In initial experiments, water-soluble extracts of wheat sourdoughs, cheeses, and vegetables were screened by agar diffusion assays with Penicillium roqueforti DPPMAF1 as the indicator fungus. Water-soluble extracts of sourdough fermented with Lactobacillus brevis AM7 and Phaseolus vulgaris cv. Pinto were selected for further study. The crude water-soluble extracts of L. brevis AM7 sourdough and P. vulgaris cv. Pinto had a MIC of 40 mg of peptide/ml and 30.9 mg of protein/ml, respectively. MICs were markedly lower when chemically synthesized peptides or partially purified protein fractions were used. The water-soluble extract of P. vulgaris cv. Pinto showed inhibition toward a large number of fungal species isolated from bakeries. Phaseolin alpha-type precursor, phaseolin, and erythroagglutinating phytohemagglutinin precursor were identified in the water-soluble extract of P. vulgaris cv. Pinto by nano liquid chromatography-electrospray ionization-tandem mass spectrometry. When the antifungal activity was assayed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, all three proteins were inhibitory. A mixture of eight peptides was identified from the water-soluble extract of sourdough L. brevis AM7, and five of these exhibited inhibitory activity. Bread was made at the pilot plant scale by sourdough fermentation with L. brevis AM7 and addition of the water-soluble extract (27%, vol/wt; 5 mg of protein/ml) of P. vulgaris cv. Pinto. Slices of bread packed in polyethylene bags did not show contamination by fungi until at least 21 days of storage at room temperature, a level of protection comparable to that afforded by 0.3% (wt/wt) calcium propionate.

Endophytic fungi isolated from wheat (Triticum durum Desf.): evaluation of their antimicrobial activity, antioxidant activity and host growth promotion.

PubMed

Harzallah, Daoud; Sadrati, Nouari; Zerroug, Amina; Dahamna, Saliha; Bouharati, Saddek

2012-01-01

The emergence of antibiotic-resistant micro-organisms calls for inventive research and development strategies. The screening for antimicrobial compounds from endophytes is a promising way to meet the increasing threat of drug-resistant strains of human and plant pathogens. Endophytes may be defined as "microbes that colonize living, internal tissues of plants without causing any immediate, overt negative effects". Endophytes are relatively unstudied as potential sources of novel natural products for exploitation in medicine, agriculture, and industry. The purpose of this study was to evaluate several isolated fungi from wheat (Triticum durum Desf.) Mohamed Ben Bachir variety and to select endophytic fungi for further evaluation of its antimicrobial, antioxidant activities and host growth promotion. A total of 20 endophytic fungi have been isolated. Antimicrobial activity was evaluated for crude ethyl acetate extracts using an agar diffusion assay. All extracts showed inhibitory activity on at least one or more pathogenic microorganism, with an average zone of inhibition varied between 7 mm to 25 mm, a large zone of 23 and 25mm against candida albicans and Escherichia coli respectively. The antioxidant capacity of the extracts was evaluated by beta-carotene/linoleic acid assay. Results showed that 70% of these extracts have antioxidant activity, exhibiting 50, 57% to 78, 96% inhibitions. While 30% from them, their inhibitory activity for oxidation of linoleic acid Were less than 50%. Growth promotion ability of these endophytes was tested on seed germination among ten isolates tested, two isolates showed significant growth promotion effects on wheat seeds. From the present work we can conclude that these microorganisms could be promising source of bioactive compounds, growth promotion and warrant further study.

Long-Term Fungal Inhibition by Pisum sativum Flour Hydrolysate during Storage of Wheat Flour Bread

PubMed Central

Lavecchia, Anna; Gramaglia, Valerio; Gobbetti, Marco

2015-01-01

In order to identify antifungal compounds from natural sources to be used as ingredients in the bakery industry, water/salt-soluble extracts (WSE) from different legume flour hydrolysates obtained by the use of a fungal protease were assayed against Penicillium roqueforti DPPMAF1. The agar diffusion assays allowed the selection of the pea (Pisum sativum) hydrolysate as the most active. As shown by the hyphal radial growth rate, the WSE had inhibitory activity towards several fungi isolated from bakeries. The MIC of the WSE was 9.0 mg/ml. Fungal inhibition was slightly affected by heating and variations in pH. The antifungal activity was attributed to three native proteins (pea defensins 1 and 2 and a nonspecific lipid transfer protein [nsLTP]) and a mixture of peptides released during hydrolysis. The three proteins have been reported previously as components of the defense system of the plant. Five peptides were purified from WSE and were identified as sequences encrypted in leginsulin A, vicilin, provicilin, and the nsLTP. To confirm antifungal activity, the peptides were chemically synthesized and tested. Freeze-dried WSE were used as ingredients in leavened baked goods. In particular, breads made by the addition of 1.6% (wt/wt) of the extract and fermented by baker's yeast or sourdough were characterized for their main chemical, structural, and sensory features, packed in polyethylene bags, stored at room temperature, and compared to controls prepared without pea hydrolysate. Artificially inoculated slices of a bread containing the WSE did not show contamination by fungi until at least 21 days of storage and behaved like the bread prepared with calcium propionate (0.3%, wt/wt). PMID:25862230

Characterisation of biofilm formation by a Streptococcus suis meningitis isolate.

PubMed

Grenier, Daniel; Grignon, Louis; Gottschalk, Marcelo

2009-02-01

Biofilm formation by a strain of Streptococcus suis serotype 2 isolated from a case of meningitis in pigs was characterised. Using a polystyrene microtitre plate assay, S. suis 95-8242 produced a dense biofilm when glucose, fructose or sucrose was used as the carbohydrate source, whereas no biofilm formed in the presence of lactose. Polysaccharide production by the biofilm-forming strain was demonstrated by the Congo red agar assay. Transmission electron microscopy revealed that bacterial cells were surrounded by a thick layer of polycationic ferritin-labelled material. S. suis 95-8242 was more resistant to both penicillin G and ampicillin in biofilms than in planktonic cultures on the basis of minimal inhibitory and minimal bactericidal concentrations.

Comparison of the antibacterial activity of essential oils and extracts of medicinal and culinary herbs to investigate potential new treatments for irritable bowel syndrome.

PubMed

Thompson, Aiysha; Meah, Dilruba; Ahmed, Nadia; Conniff-Jenkins, Rebecca; Chileshe, Emma; Phillips, Chris O; Claypole, Tim C; Forman, Dan W; Row, Paula E

2013-11-28

Irritable bowel syndrome (IBS) is a common functional gastrointestinal disorder, which may result from alteration of the gastrointestinal microbiota following gastrointestinal infection, or with intestinal dysbiosis or small intestinal bacterial overgrowth. This may be treated with antibiotics, but there is concern that widespread antibiotic use might lead to antibiotic resistance. Some herbal medicines have been shown to be beneficial, but their mechanism(s) of action remain incompletely understood. To try to understand whether antibacterial properties might be involved in the efficacy of these herbal medicines, and to investigate potential new treatments for IBS, we have conducted a preliminary study in vitro to compare the antibacterial activity of the essential oils of culinary and medicinal herbs against the bacterium, Esherichia coli. Essential oils were tested for their ability to inhibit E. coli growth in disc diffusion assays and in liquid culture, and to kill E. coli in a zone of clearance assay. Extracts of coriander, lemon balm and spearmint leaves were tested for their antibacterial activity in the disc diffusion assay. Disc diffusion and zone of clearance assays were analysed by two-tailed t tests whereas ANOVA was performed for the turbidometric assays. Most of the oils exhibited antibacterial activity in all three assays, however peppermint, lemon balm and coriander seed oils were most potent, with peppermint and coriander seed oils being more potent than the antibiotic rifaximin in the disc diffusion assay. The compounds present in these oils were identified by gas chromatography mass spectrometry. Finally, extracts were made of spearmint, lemon balm and coriander leaves with various solvents and these were tested for their antibacterial activity against E. coli in the disc diffusion assay. In each case, extracts made with ethanol and methanol exhibited potent antibacterial activity. Many of the essential oils had antibacterial activity in the three assays, suggesting that they would be good candidates for testing in clinical trials. The observed antibacterial activity of ethanolic extracts of coriander, lemon balm and spearmint leaves suggests a mechanistic explanation for the efficacy of a mixture of coriander, lemon balm and mint extracts against IBS in a published clinical trial.

Evaluation of the Biomic V3 Microbiology System for Identification of Selected Species on BBL CHROMagar Orientation Agar and CHROMagar MRSA Medium ▿

PubMed Central

Baron, Ellen Jo; D'Souza, Holly; Qi Wang, Andrew; Gibbs, David L.

2008-01-01

The Biomic V3 microbiology system identifies bacteria by reading the color of colonies selected by the user. For CHROMagar orientation, Biomic results agreed with conventional methods for 94% of the strains assayed. For CHROMagar MRSA, Biomic correctly identified 100% of the strains tested and did not misidentify two methicillin-susceptible Staphylococcus aureus strains growing on the plates. PMID:18701661

University of Texas MD Anderson: Phenotypic Examination of PIK3CA Allelic Series using In Vitro/In Vivo Sensor Platforms | Office of Cancer Genomics

Cancer.gov

The CTD2 Center at the University of Texas MD Anderson Cancer Center utilized an established and operational MCF10A normal breast epithelial cell model to assess the ability of candidate driver aberrations to promote cell grow in anchorage-independent conditions (soft agar assay) and proliferate in the absence of insulin and epidermal growth factor (EGF).

University of Texas MD Anderson Cancer Center: Phenotypic Examination of PIK3CA Allelic Series using In Vitro/In Vivo Sensor Platforms | Office of Cancer Genomics

Cancer.gov

The CTD2 Center at the University of Texas MD Anderson Cancer Center utilized an established and operational MCF10A normal breast epithelial cell model to assess the ability of candidate driver aberrations to promote cell grow in anchorage-independent conditions (soft agar assay) and proliferate in the absence of insulin and epidermal growth factor (EGF).

A.D.A.M. test (Antibiofilm Dressing's Activity Measurement) - Simple method for evaluating anti-biofilm activity of drug-saturated dressings against wound pathogens.

PubMed

Junka, Adam F; Żywicka, Anna; Szymczyk, Patrycja; Dziadas, Mariusz; Bartoszewicz, Marzena; Fijałkowski, Karol

2017-12-01

In the present article, we propose a simple Antibiofilm Dressing's Activity Measurement (A.D.A.M.) test that allows to check in vitro a dressing's suitability against biofilm-related wound infections. To perform the test, three agar discs are covered with biofilm formed by the tested pathogen after which they are assembled one over another in the form of an agar plug and placed in the well of a 24-well plate. The top disc is covered with the analyzed dressing and the entire set is incubated for 24h. During this time, the investigated antimicrobial substance is released from the dressing and penetrates to subsequent biofilm-covered agar discs. Biofilm reduction is measured using 2,3,5-triphenyl-2H-tetrazolium chloride (TTC) spectrometric assay and the results are compared to untreated control samples (agar plug covered with biofilm and without the dressing/or with a passive dressing placed on the top disc). Furthermore, in order to standardize the differences in penetrability of the drugs released from active dressings the results can be expressed as a dimensionless value referred to as the Penetrability Index. In summary, A.D.A.M. test is simple, cheap, can be performed practically in every clinical laboratory and takes no more time than routine microbiological diagnostics. Apart from measuring the released drug's activity, the A.D.A.M. test allows to assess drug penetrability (across three agar discs), reflecting real wound conditions, where microbes are frequently hidden under the necrotic tissue or cloth. In conclusion, the A.D.A.M. test produces a high volume of data that, when analyzed, can provide a researcher with a valuable hint concerning the applicability of active dressings against specific biofilm pathogens in a particular setting. Copyright © 2017 Elsevier B.V. All rights reserved.

Comparison of the BBL CHROMagar Staph aureus Agar Medium to Conventional Media for Detection of Staphylococcus aureus in Respiratory Samples

PubMed Central

Flayhart, Diane; Lema, Clara; Borek, Anita; Carroll, Karen C.

2004-01-01

Screening for Staphylococcus aureus has become routine in certain patient populations. This study is the first clinical evaluation of the BBL CHROMagar Staph aureus agar (CSA) medium (BD Diagnostics, Sparks, Md.) for detection of S. aureus in nasal surveillance cultures and in respiratory samples from cystic fibrosis (CF) patients. S. aureus colonies appear mauve on CSA. Other organisms are inhibited or produce a distinctly different colony color. S. aureus was identified from all media by slide coagulase, exogenous DNase, and mannitol fermentation assays. Susceptibility testing was performed using the agar dilution method. A total of 679 samples were evaluated. All samples were inoculated onto CSA. Nasal surveillance cultures were inoculated onto sheep blood agar (SBA) (BD Diagnostics), and samples from CF patients were inoculated onto mannitol salt agar (MSA) (BD Diagnostics). Of the 679 samples cultured, 200 organisms produced a mauve color on CSA (suspicious for S. aureus) and 180 were positive for S. aureus on SBA or MSA. Of 200 CSA-positive samples 191 were identified as S. aureus. Nine mauve colonies were slide coagulase negative and were subsequently identified as Staphylococcus lugdunensis (one), Staphylococcus epidermidis (three), Staphylococcus haemolyticus (one), and Corynebacterium species (four). CSA improved the ability to detect S. aureus by recovering 12 S. aureus isolates missed by conventional media. Of the 192 S. aureus isolates recovered, 122 were methicillin susceptible and 70 were methicillin resistant. Overall, the sensitivity and specificity of CSA in this study were 99.5 and 98%, respectively. There was no difference in the performance of the slide coagulase test or in susceptibility testing performed on S. aureus recovered from CSA compared to SBA or MSA. Our data support the use of CSA in place of standard culture media for detection of S. aureus in heavily contaminated respiratory samples. PMID:15297498

Biodegradative potential of fungal isolates from sacral ambient: In vitro study as risk assessment implication for the conservation of wall paintings

PubMed Central

Dimkić, Ivica; Stupar, Miloš; Stanković, Slaviša; Vukojević, Jelena; Ljaljević Grbić, Milica

2018-01-01

The principal purpose of the study was to evaluate in vitro the potential ability of fungal isolates obtained from the painted layer of frescoes and surrounding air to induce symptoms of fresco deterioration, associated with their growth and metabolism, so that the risk of such deterioration can be precisely assessed and appropriate conservation treatments formulated. Biodegradative properties of the tested microfungi were qualitatively characterized through the use of a set of special agar plates: CaCO3 glucose agar (calcite dissolution), casein nutrient agar (casein hydrolysis), Czapek-Dox minimal medium (pigment secretion); and Czapek-Dox minimal broth (acid and alkali production). Most of the tested isolates (71.05%) demonstrated at least one of the degradative properties, with Penicillium bilaiae as the most potent, since it tested positive in all four. The remaining isolates (28.95%) showed no deterioration capabilities and were hence considered unlikely to partake in the complex process of fungal deterioration of murals via the tested mechanisms. The obtained results clearly indicate that utilization of fast and simple plate assays can provide insight into the biodegradative potential of deteriogenic fungi and allow for their separation from allochthonous transients, a prerequisite for precise assessment of the amount of risk posed by a thriving mycobiota to mural paintings. PMID:29309432

Effect of permeable flow on cyclic layering in solidifying magma bodies: Insights from an analog experiment of diffusion-precipitation systems

NASA Astrophysics Data System (ADS)

Toramaru, A.; Yamauchi, S.

2012-04-01

Characteristic structures such as rhythmic layering, cress cumulate, cross bedding, perpendicular feldspar rock etc, are commonly observed in layered intrusion or shallow magmatic intrusions. These structures result from complex processes including thermal and compositional diffusions, crystallization, crystal settling, convection and interaction among three phases (crystals, bubble, melt). In order to understand how the differentiation proceeds in solidifying magma bodies from each characteristic structure together with chemical signatures, it is necessary to evaluate the relative importance among these elemental processes on structures. As an attempt to evaluate the effect of advection on a diffusion-related structure, we carried out an analog experiment of Liesegang system using lead-iodide (PbI2) crystallization in agar media which have been normally used to prohibit convection. In the ordinary Liesegang band formation experiments including only diffusion and crystallization kinetics without any advection and convection, the precipitation bands develop with regular spacing following a geometric progression due to two-component diffusion and reaction with supersaturation. This type of banding structure has been advocated as the same type of cyclic layering or vesicle layering (a sort of rhythmic layering) in dykes or sills. In order to see the effect of one-directional advection on Liesegang band, we apply the electric field (5 V to 25 V for a distance 15 cm) along the concentration gradient in agar media, thereby counteracting flows of lead anion Pb2+ and iodide ion I- are driven at constant velocities. The flows of anions and ions are equivalent to the permeable flows in porous media of crystal mush. The resultant precipitation structures exhibit very curious banding structure in which band spacings do not change with distance, are nearly constant and quite narrow, depending on the voltage, unlike those in ordinary Liesegang bands in which band spacings increase with distance following geometric progression. Further interestingly each band consists of a lot of very tiny irregular-shaped crystal aggregates. From experimental results and scaling arguments, with regard to the effect of one directional permeable flow on band spacing of cyclic layering, we propose a hypothesis of constant Peclet number that Peclet number (ratio of flow velocity to diffusive velocity) is nearly unity. By applying the hypothesis to natural examples, we can estimate a value of permeable flow velocity of interstitial melts in differentiating magma bodies from values of a band spacing and diffusivity data.

Growth and antimicrobial studies of γ-glycine crystal grown using CuSO4

NASA Astrophysics Data System (ADS)

Vijayalakshmi, V.; Dhanasekaran, P.

2018-05-01

In the current work single crystals of pure and 1M of CuSO4-added glycine were grown by slow evaporation method and its optical and antimicrobial properties were studied. The Polymorph of glycine transforms from a-glycine to γ-glycine due to the incorporation of CuSO4 on glycine was affirmed by the PXRD and FTIR studies. The impact of CuSO4 on the antimicrobial action of the grown samples was deliberate by utilizing the agar diffusion method.

Identification of Burkholderia spp. in the Clinical Microbiology Laboratory: Comparison of Conventional and Molecular Methods

PubMed Central

van Pelt, Cindy; Verduin, Cees M.; Goessens, Wil H. F.; Vos, Margreet C.; Tümmler, Burkhard; Segonds, Christine; Reubsaet, Frans; Verbrugh, Henri; van Belkum, Alex

1999-01-01

Cystic fibrosis (CF) predisposes patients to bacterial colonization and infection of the lower airways. Several species belonging to the genus Burkholderia are potential CF-related pathogens, but microbiological identification may be complicated. This situation is not in the least due to the poorly defined taxonomic status of these bacteria, and further validation of the available diagnostic assays is required. A total of 114 geographically diverse bacterial isolates, previously identified in reference laboratories as Burkholderia cepacia (n = 51), B. gladioli (n = 14), Ralstonia pickettii (n = 6), B. multivorans (n = 2), Stenotrophomonas maltophilia (n = 3), and Pseudomonas aeruginosa (n = 11), were collected from environmental, clinical, and reference sources. In addition, 27 clinical isolates putatively identified as Burkholderia spp. were recovered from the sputum of Dutch CF patients. All isolates were used to evaluate the accuracy of two selective growth media, four systems for biochemical identification (API 20NE, Vitek GNI, Vitek NFC, and MicroScan), and three different PCR-based assays. The PCR assays amplify different parts of the ribosomal DNA operon, either alone or in combination with cleavage by various restriction enzymes (PCR-restriction fragment length polymorphism [RFLP] analysis). The best system for the biochemical identification of B. cepacia appeared to be the API 20NE test. None of the biochemical assays successfully grouped the B. gladioli strains. The PCR-RFLP method appeared to be the optimal method for accurate nucleic acid-mediated identification of the different Burkholderia spp. With this method, B. gladioli was also reliably classified in a separate group. For the laboratory diagnosis of B. cepacia, we recommend parallel cultures on blood agar medium and selective agar plates. Further identification of colonies with a Burkholderia phenotype should be performed with the API 20NE test. For final confirmation of species identities, PCR amplification of the small-subunit rRNA gene followed by RFLP analysis with various enzymes is recommended. PMID:10364579

[Inhibitory effects of butyl alcohol extract of Baitouweng decoction on yeast-to-hyphae transition of Candida albicans isolates from VVC in alkaline pH environment].

PubMed

Zhang, Meng-xiang; Xia, Dan; Shi, Gao-xiang; Shao, Jing; Wang, Tian-ming; Tang, Chuan-chao; Wang, Chang-zhong

2015-02-01

To investigate the effects of butyl alcohol extract of Baitouweng decoction ( BAEB) on yeast-to-hyphae transition of Candida albicans isolates from vulvovaginal candidiasis (VVC) in alkaline pH. Serial 2-fold dilution assay was used to determine the minimal inhibitory concentrations (MICs) of Baitouweng decoction extracts against C. albicans isolates from VVC, XTT assay was applied to determine the metabolic activity of C. albicans hypha treated by BAEB for 6 h. The morphological change of C. albicans treated by BAEB was inspected at different pH by inverted microscope, fluorescence microscope, scanning electron microscopy (SEM). Solid agar plate and semi-solid agar were utilized to evaluate colony morphology and invasive growth of C. albicans, respectively. Quantitative Real-time PCR (qRT-PCR) was adopted to observe the expressions of hyphae-specific genes including HWP1, ALS3, CSH1, SUN41 and CaPDE2. The MIC of BAEB against C. albicans is less than that of other extracts; hyphae grow best at pH 8. 0; 512 mg · L(-1) and 1,024 mg · L(-1) BAEB could inhibit formation of hyphae and influence colony morphology. When treated by 512 mg · L(-1) and 1,024 mg · L(-1) BAEB, the colonies became smooth; while by 0 and 256 mg · L(-1) BAEB, the colonies became wrinkled. In semi-solid agar, the length of hyphae decreased steadily as the concentration of BAEB lowered. The expression of HWP1, ALS3, CSHl, SUN41 were downregulated by 5.12, 4.26, 3.2 and 2.74 folds, and CaPDE2 was upregulated by 2.38 fold. BAEB could inhibit yeast-to-hyphae transition of C. albicans isolates from VVC in alkaline pH.

Protease and lipase activities of fungal and bacterial strains derived from an artisanal raw ewe's milk cheese.

PubMed

Ozturkoglu-Budak, Sebnem; Wiebenga, Ad; Bron, Peter A; de Vries, Ronald P

2016-11-21

We previously identified the microbiota present during cheese ripening and observed high protease and lipase activity in Divle Cave cheese. To determine the contribution of individual isolates to enzyme activities, we investigated a range of species representing this microbiota for their proteolytic and lipolytic ability. In total, 17 fungal, 5 yeast and 18 bacterial strains, previously isolated from Divle Cave cheese, were assessed. Qualitative protease and lipase activities were performed on skim-milk agar and spirit-blue lipase agar, respectively, and resulted in a selection of strains for quantitative assays. For the quantitative assays, the strains were grown on minimal medium containing irradiated Divle Cave cheese, obtained from the first day of ripening. Out of 16 selected filamentous fungi, Penicillium brevicompactum, Penicillium cavernicola and Penicillium olsonii showed the highest protease activity, while Mucor racemosus was the best lipase producer. Yarrowia lipolytica was the best performing yeast with respect to protease and lipase activity. From the 18 bacterial strains, 14 and 11 strains, respectively showed protease and lipase activity in agar plates. Micrococcus luteus, Bacillus stratosphericus, Brevibacterium antiquum, Psychrobacter glacincola and Pseudomonas proteolytica displayed the highest protease and lipase activity. The proteases of yeast and filamentous fungi were identified as mainly aspartic protease by specific inhibition with Pepstatin A, whereas inhibition by PMSF (phenylmethylsulfonyl fluoride) indicated that most bacterial enzymes belong to serine type protease. Our results demonstrate that aspartic proteases, which usually have high milk clotting activity, are predominantly derived from fungal strains, and therefore fungal enzymes appear to be more suitable for use in the cheese industry. Microbial enzymes studied in this research might be alternatives for rennin (chymosin) from animal source because of their low cost and stable availability. Future studies will aim to purify these enzymes to test their suitability for use in similar artisanal cheeses or in large scale commercial cheeses. Copyright © 2016 Elsevier B.V. All rights reserved.

Enhanced bioactivity of ZnO nanoparticles—an antimicrobial study

PubMed Central

Padmavathy, Nagarajan; Vijayaraghavan, Rajagopalan

2008-01-01

In this study, we investigate the antibacterial activity of ZnO nanoparticles with various particle sizes. ZnO was prepared by the base hydrolysis of zinc acetate in a 2-propanol medium and also by a precipitation method using Zn(NO3)2 and NaOH. The products were characterized by x-ray diffraction (XRD) analysis, transmission electron microscopy (TEM) and photoluminescence (PL) spectroscopy. Bacteriological tests such as minimum inhibitory concentration (MIC) and disk diffusion were performed in Luria-Bertani and nutrient agar media on solid agar plates and in liquid broth systems using different concentrations of ZnO by a standard microbial method for the first time. Our bacteriological study showed the enhanced biocidal activity of ZnO nanoparticles compared with bulk ZnO in repeated experiments. This demonstrated that the bactericidal efficacy of ZnO nanoparticles increases with decreasing particle size. It is proposed that both the abrasiveness and the surface oxygen species of ZnO nanoparticles promote the biocidal properties of ZnO nanoparticles. PMID:27878001

Enhanced bioactivity of ZnO nanoparticles—an antimicrobial study

NASA Astrophysics Data System (ADS)

Padmavathy, Nagarajan; Vijayaraghavan, Rajagopalan

2008-07-01

In this study, we investigate the antibacterial activity of ZnO nanoparticles with various particle sizes. ZnO was prepared by the base hydrolysis of zinc acetate in a 2-propanol medium and also by a precipitation method using Zn(NO3)2 and NaOH. The products were characterized by x-ray diffraction (XRD) analysis, transmission electron microscopy (TEM) and photoluminescence (PL) spectroscopy. Bacteriological tests such as minimum inhibitory concentration (MIC) and disk diffusion were performed in Luria-Bertani and nutrient agar media on solid agar plates and in liquid broth systems using different concentrations of ZnO by a standard microbial method for the first time. Our bacteriological study showed the enhanced biocidal activity of ZnO nanoparticles compared with bulk ZnO in repeated experiments. This demonstrated that the bactericidal efficacy of ZnO nanoparticles increases with decreasing particle size. It is proposed that both the abrasiveness and the surface oxygen species of ZnO nanoparticles promote the biocidal properties of ZnO nanoparticles.

Multiple Echo Diffusion Tensor Acquisition Technique (MEDITATE) on a 3T clinical scanner

PubMed Central

Baete, Steven H.; Cho, Gene; Sigmund, Eric E.

2013-01-01

This paper describes the concepts and implementation of an MRI method, Multiple Echo Diffusion Tensor Acquisition Technique (MEDITATE), which is capable of acquiring apparent diffusion tensor maps in two scans on a 3T clinical scanner. In each MEDITATE scan, a set of RF-pulses generates multiple echoes whose amplitudes are diffusion-weighted in both magnitude and direction by a pattern of diffusion gradients. As a result, two scans acquired with different diffusion weighting strengths suffice for accurate estimation of diffusion tensor imaging (DTI)-parameters. The MEDITATE variation presented here expands previous MEDITATE approaches to adapt to the clinical scanner platform, such as exploiting longitudinal magnetization storage to reduce T2-weighting. Fully segmented multi-shot Cartesian encoding is used for image encoding. MEDITATE was tested on isotropic (agar gel), anisotropic diffusion phantoms (asparagus), and in vivo skeletal muscle in healthy volunteers with cardiac-gating. Comparisons of accuracy were performed with standard twice-refocused spin echo (TRSE) DTI in each case and good quantitative agreement was found between diffusion eigenvalues, mean diffusivity, and fractional anisotropy derived from TRSE-DTI and from the MEDITATE sequence. Orientation patterns were correctly reproduced in both isotropic and anisotropic phantoms, and approximately so for in vivo imaging. This illustrates that the MEDITATE method of compressed diffusion encoding is feasible on the clinical scanner platform. With future development and employment of appropriate view-sharing image encoding this technique may be used in clinical applications requiring time-sensitive acquisition of DTI parameters such as dynamical DTI in muscle. PMID:23828606

An immortalized goat mammary epithelial cell line induced with human telomerase reverse transcriptase (hTERT) gene transfer.

PubMed

He, Y L; Wu, Y H; He, X N; Liu, F J; He, X Y; Zhang, Y

2009-06-01

Although mammary epithelial cell lines can provide a rapid and reliable indicator of gene expression efficiency of transgenic animals, their short lifespan greatly limits this application. To provide stable and long lifespan cells, goat mammary epithelial cells (GMECs) were transduced with pLNCX2-hTERT by retrovirus-mediated gene transfer. Transduced GMECs were evaluated by reverse transcriptase polymerase chain reaction (RT-PCR), proliferation assays, karyotype analysis, telomerase activity assay, western blotting, soft agar assay, and injection into nude mice. Non-transduced GMECs were used as a control. The hTERT-GMECs had higher telomerase activity and extended proliferative lifespan compared to non-transfected GMECs; even after Passage 50, hTERT-GMECs had a near diploid complement of chromosomes. Furthermore, they did not gain the anchorage-independent growth property and were not associated with a malignant phenotype in vitro or in vivo.

Comparative evaluation of antimicrobial effect of herbal root canal irrigants (Morinda citrifolia, Azadirachta indica, Aloe vera) with sodium hypochlorite: An in vitro study.

PubMed

Babaji, Prashant; Jagtap, Kiran; Lau, Himani; Bansal, Nandita; Thajuraj, S; Sondhi, Priti

2016-01-01

Successful root canal treatment involves the complete elimination of microorganism from the root canal and the three-dimensional obturation of the canal space. Enterococcus faecalis is the most commonly found bacteria in failed root canal. Chemical irrigation of canals along with biomechanical preparation helps in the elimination of microorganisms. The present study was aimed to evaluate the antimicrobial effect of herbal root canal irrigants (Morinda citrifolia, Azadirachta indica extract, Aloe vera) with sodium hypochlorite (NaOCl). The bacterial E. faecalis (ATCC) culture was grown overnight in brain heart infusion (BHI) broth and inoculated in Mueller-Hinton agar plates. Antibacterial inhibition was assessed using agar well diffusion method. All five study irrigants were added to respective wells in agar plates and incubated at 37°C for 24 h. Bacterial inhibition zone around each well was recorded. Results were tabulated and statistically analyzed using Statistical Package for the Social Sciences software for Windows, version 19.0. (IBM Corp., Armonk, NY. Highest inhibitory zone against E. faecalis was seen in NaOCl fallowed by M. citrifolia and A. indica extract, and the least by A. vera extract. Tested herbal medicine (A. indica extract, M. citrifolia, A. vera) showed inhibitory zone against E. faecalis. Hence, these irrigants can be used as root canal irrigating solutions.

Evaluating the Risk of Tumors Diseases Based on Measurement of Urinary and Serumal Antioxidants Using the New Agar Diffusion Methods

PubMed Central

Zhou, Ying; Chen, Jing; Wang, Zhen

2017-01-01

Objectives. To discuss the characteristics of the amount of urinary total antioxidants in tumor diseases and the possibility of utilizing the changing regulation of urinary antioxidants to diagnose tumor diseases. Method. Urine and serum specimens from 130 healthy people were used to investigate the variation of antioxidant capacity against age. Urine and serum specimens from 44 unselected patients with tumors and 44 healthy people with same age background were used to explore the significance of urinary antioxidant capacity in clinic to diagnose tumor diseases. Potassium permanganate agar method and iodine starch method were used to determine the amount of total antioxidants. Results. In healthy people, more antioxidants in urine were measured in older people, while the results were opposite in serum. More antioxidants were found in urine of tumor patients than in healthy people with same age-range. Conclusions. According to the results of 130 measurements, the amount of antioxidants in urine varies by age. By using agar methods to measure antioxidants, the effect of age is required to be considered. Antioxidants levels from tumor patients were significantly higher than healthy individuals in urine. The combination of urine and serum to determine total antioxidants can better diagnose tumor diseases based on iodine starch method, with area under the receiver operating characteristics curve at 0.787. PMID:28458777

Mammalian cell-transforming potential of traffic-linked ultrafine particulate matter PM0.056 in urban roadside atmosphere.

PubMed

Verma, Mukesh K; Poojan, Shiv; Sultana, Sarwat; Kumar, Sushil

2014-09-01

We examined the clastogenic and cell-transforming potential of ultrafine particulate matter fraction PM0.056 of urban ambient aerosol using mammalian cells. PM1.0, PM0.56 and PM0.056 fractions were sampled from roadside atmosphere of an urban area using the cascade impactor MOUDI-NR-110. The potential to induce cytotoxicity, DNA damage and micronuclei formation was examined at the test concentrations of 3, 6, 12.5, 25, 50 and 100 μg/ml using the 3-4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay, the plasmid relaxation assay and the C3H10T1/2 (10T1/2) cells. The cell-transforming potential was investigated in vitro using 10T1/2 cell transformation assay and the soft agar assay. PM1, PM0.56 and PM0.056 fractions were found to be toxic in dose-dependent manner. These induced cytotoxicity at five test concentrations, the ultrafine particle fraction PM0.056 showed greater cytotoxic potential. PM0.056 induced micronucleus formation in 10T1/2 cells. The effect was statistically significant. The DNA-damaging potential was measured in a plasmid relaxation assay. Both fine and ultrafine particle fraction PM0.56 and PM0.056 displayed greater effect as compared to larger PM1 fraction. DNA damage was found to be dependent on particulate matter intrinsic pro-oxidant chemicals. The ability of the ultrafine particle fraction PM0.056 to induce morphological cell transformation was demonstrated by significant and dose-dependent increases in type III focus formation by morphologically transformed cells in culture flasks and their clonal expansion in soft agar. It is concluded that the traffic-linked ultrafine particle fraction PM0.056 in the atmosphere by the roadside of an urban area is clastogenic and able to induce morphological transformation of mammalian cells. © The Author 2014. Published by Oxford University Press on behalf of the UK Environmental Mutagen Society. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Detection of Shiga toxin-producing Escherichia coli (STEC) O157:H7, O26, O45, O103, O111, O121, and O145, and Salmonella in retail raw ground beef using the DuPont™ BAX® system.

PubMed

Wasilenko, Jamie L; Fratamico, Pina M; Sommers, Christopher; DeMarco, Daniel R; Varkey, Stephen; Rhoden, Kyle; Tice, George

2014-01-01

Shiga toxin-producing Escherichia coli (STEC) and Salmonella are food-borne pathogens commonly associated with beef, and reliable methods are needed to determine their prevalence in beef and to ensure food safety. Retail ground beef was tested for the presence of E. coli O157:H7, STEC serogroups O26, O45, O103, O111, O121, and O145, and Salmonella using the DuPont™ BAX® system method. Ground beef (325 g) samples were enriched in 1.5 L of TSB with 2 mg/L novobiocin at 42°C for 18 h, and then evaluated using the BAX® System real-time PCR assays for E. coli O157:H7 and STEC suite, and the BAX® System standard PCR assays for E. coli O157:H7 MP and Salmonella. Samples positive for STEC target genes by the BAX® System assays were subjected to immunomagnetic separation (IMS) and plating onto modified Rainbow Agar O157. Enrichments that were PCR positive for Salmonella were inoculated into RV broth, incubated for 18 h at 42°C, and then plated onto XLT-4 agar. Presumptive positive STEC and Salmonella colonies were confirmed using the BAX® System assays. Results of the BAX® System STEC assays showed 20/308 (6.5%) of samples positive for both the Shiga toxin (stx) and intimin (eae) genes; 4 (1.3%) for stx, eae, and O26; 1 (0.3%) for stx, eae, and O45; 3 (1%) for stx, eae, and O103; and 1 (0.3%) for stx, eae, and O145. There were also 3 samples positive for stx, eae, and more than one STEC serogroup. Three (1.0%) of the samples were positive using the BAX® System real-time E. coli O157:H7 assay, and 28 (9.1%) were positive using the BAX® System Salmonella assay. STEC O103 and E. coli O157:H7 were isolated from 2/6 and 2/3 PCR positive samples, respectively. Salmonella isolates were recovered and confirmed from 27 of the 28 Salmonella PCR positive samples, and a portion of the isolates were serotyped and antibiotic resistance profiles determined. Results demonstrate that the BAX® System assays are effective for detecting STEC and Salmonella in beef.

Impedance Measurements Could Accelerate Phage-Based Identification of Bacillus anthracis and Other Bacteria

DTIC Science & Technology

2016-09-01

The Bacillus-inoculated NSM agar plates were incubated at 35°C for at least 48 h until Gram stains revealed the presence of > 90% Bacillus spores in...longer visible in Gram stained samples. Finally, centrifugation was used to remove soluble debris from the preparation and spore concentrations were...minutes post treatment. Gram Stains . Gram stains were used to track the emergence of vegetative Bacillus cells from spores. In this assay, bacterial

University of Texas MD Anderson Cancer Center (UT-MDACC): Phenotypic Examination of PIK3CA Allelic Series using In Vitro/In Vivo Sensor Platforms | Office of Cancer Genomics

Cancer.gov

The CTD2 Center at the University of Texas MD Anderson Cancer Center utilized an established and operational MCF10A normal breast epithelial cell model to assess the ability of candidate driver aberrations to promote cell grow in anchorage-independent conditions (soft agar assay) and proliferate in the absence of insulin and epidermal growth factor (EGF).

Cadmium-Aluminum Layered Double Hydroxide Microspheres for Photocatalytic CO2 Reduction.

PubMed

Saliba, Daniel; Ezzeddine, Alaa; Sougrat, Rachid; Khashab, Niveen M; Hmadeh, Mohamad; Al-Ghoul, Mazen

2016-04-21

We report the synthesis of cadmium-aluminum layered double hydroxide (CdAl LDH) using the reaction-diffusion framework. As the hydroxide anions diffuse into an agar gel matrix containing the mixture of aluminum and cadmium salts at a given ratio, they react to give the LDH. The LDH self-assembles inside the pores of the gel matrix into a unique spherical-porous shaped microstructure. The internal and external morphologies of the particles are studied by electron microscopy and tomography revealing interconnected channels and a high surface area. This material is shown to exhibit a promising performance in the photoreduction of carbon dioxide using solar light. Moreover, the palladium-decorated version shows a significant improvement in its reduction potential at room temperature. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Antimicrobial activity of lactic acid bacteria isolated from bekasam against staphylococcus aureus ATCC 25923, escherichia coli ATCC 25922, and salmonella sp

NASA Astrophysics Data System (ADS)

Sari, Melia; Suryanto, Dwi; Yurnaliza

2018-03-01

Bekasam is an Indonesian fermented food made of fish. As a fermented food, this food may contain some beneficial bacteria like lactic acid bacteria (LAB), which usually have antimicrobial properties such as organic acid, hydrogen peroxide, and a bacteriocin. A study on antimicrobial activity of LAB isolated from bekasam against some pathogenic bacteria has been conducted. The purpose of this study was to know the ability of crude bacteriocin produced LAB of bekasam against Staphylococcus aureus ATCC 25923, Escherichia coli ATCC 25922, and Salmonella sp. Bekasam sample was taken from South Sumatera. LAB isolation was done using de Man Rogosa and Sharpe agar. A bacterial colony with clear zone was selected and purified to get a single colony. The antagonistic assay of the LAB was conducted in Muller-Hinton agar Selected isolates with higher clearing zone were assayed for antibacterial effect of their crude bacteriocin of different culture incubation time of 6, 9, and 12 hours. The results showed that the crude extract bacteriocin of isolate MS2 of 9 hours culture incubation time inhibited more in Staphylococcus aureus ATCC 25923 with inhibition zone of 13.1 mm, whereas isolate MS9 of 9 hours culture incubation time inhibited more in Escherichia coli ATCC 25922 and Salmonella sp. with inhibition zone of 12.7 and 7.3 mm, respectively.

Eckol suppresses maintenance of stemness and malignancies in glioma stem-like cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hyun, Kyung-Hwan; Yoon, Chang-Hwan; Kim, Rae-Kwon

A subpopulation of cancer cells with stem cell properties is responsible for tumor maintenance and progression, and may contribute to resistance to anticancer treatments. Thus, compounds that target cancer stem-like cells could be usefully applied to destroy cancer. In this study, we investigated the effect of Eckol, a phlorotannin compound, on stemness and malignancies in glioma stem-like cells. To determine whether Eckol targets glioma stem-like cells, we examined whether Eckol treatment could change the expression levels of glioma stem-like cell markers and self-renewal-related proteins as well as the sphere forming ability, and the sensitivity to anticancer treatments. Alterations in themore » malignant properties of sphere-derived cells by Eckol were also investigated by soft-agar colony forming assay, by xenograft assay in nude mice, and by cell invasion assay. Treatment of sphere-forming glioma cells with Eckol effectively decreased the sphere formation as well as the CD133{sup +} cell population. Eckol treatment suppressed expression of the glioma stem-like cell markers and the self-renewal-related proteins without cell death. Moreover, treatment of glioma stem-like cells with Eckol significantly attenuated anchorage-independent growth on soft agar and tumor formation in xenograft mice. Importantly, Eckol treatment effectively reduced the resistance of glioma stem-like cells to ionizing radiation and temozolomide. Treatment of glioma stem-like cells with Eckol markedly blocked both phosphoinositide 3-kinase-Akt and Ras-Raf-1-Erk signaling pathways. These results indicate that the natural phlorotannin Eckol suppresses stemness and malignancies in glioma stem-like cells, and thereby makes glioma stem-like cells more sensitive to anticancer treatments, providing novel therapeutic strategies targeting specifically cancer stem-like cells.« less

Growth of Serratia liquefaciens under 7 mbar, 0°C, and CO2-Enriched Anoxic Atmospheres

PubMed Central

Ulrich, Richard; Berry, Bonnie J.; Nicholson, Wayne L.

2013-01-01

Abstract Twenty-six strains of 22 bacterial species were tested for growth on trypticase soy agar (TSA) or sea-salt agar (SSA) under hypobaric, psychrophilic, and anoxic conditions applied singly or in combination. As each factor was added to multi-parameter assays, the interactive stresses decreased the numbers of strains capable of growth and, in general, reduced the vigor of the strains observed to grow. Only Serratia liquefaciens strain ATCC 27592 exhibited growth at 7 mbar, 0°C, and CO2-enriched anoxic atmospheres. To discriminate between the effects of desiccation and hypobaria, vegetative cells of Bacillus subtilis strain 168 and Escherichia coli strain K12 were grown on TSA surfaces and simultaneously in liquid Luria-Bertani (LB) broth media. Inhibition of growth under hypobaria for 168 and K12 decreased in similar ways for both TSA and LB assays as pressures were reduced from 100 to 25 mbar. Results for 168 and K12 on TSA and LB are interpreted to indicate a direct low-pressure effect on microbial growth with both species and do not support the hypothesis that desiccation alone on TSA was the cause of reduced growth at low pressures. The growth of S. liquefaciens at 7 mbar, 0°C, and CO2-enriched anoxic atmospheres was surprising since S. liquefaciens is ecologically a generalist that occurs in terrestrial plant, fish, animal, and food niches. In contrast, two extremophiles tested in the assays, Deinococcus radiodurans strain R1 and Psychrobacter cryohalolentis strain K5, failed to grow under hypobaric (25 mbar; R1 only), psychrophilic (0°C; R1 only), or anoxic (<0.1% ppO2; both species) conditions. Key Words: Habitable zone—Hypobaria—Extremophiles—Special regions—Planetary protection. Astrobiology 13, 115–131. PMID:23289858

Towards a phenotypic screening strategy for emerging β-lactamases in Gram-negative bacilli.

PubMed

Willems, Elise; Verhaegen, Jan; Magerman, Koen; Nys, Sita; Cartuyvels, Reinoud

2013-02-01

The purpose of this manuscript was to review recent literature and guidelines regarding phenotypic detection of emerging β-lactamases [extended-spectrum β-lactamases (ESBLs), AmpC β-lactamases and carbapenemases] in Gram-negative bacilli (GNB) in order to formulate recommendations on best practice to screen for them. We conclude that chromogenic ESBL screening agar plates are suitable to screen for ESBL-producing Enterobacteriaceae directly from clinical samples. Furthermore, ceftazidime (CAZ) and ceftriaxone or cefotaxime (CTX) are the indicator antimicrobial agents of choice for ESBL detection in GNB. In non-inducible Enterobacteriaceae, the combined double-disk synergy test (CDDST) with at least CTX and CAZ and additionally cefepime as indicators is the preferred ESBL confirmation assay. The two most suitable ESBL confirmation strategies in AmpC co-producing Enterobacteriaceae are adapted CDDSTs: (i) with addition of 3-aminophenylboronic acid to CTX and CAZ disks; and (ii) with addition of cloxacillin (CLOX) to Mueller-Hinton agar. Reduced cefoxitin susceptibility and decreased susceptibility to cefotetan are regarded as suitable screening tests for plasmid-mediated and derepressed AmpC production. A CLOX-based CDDST with CTX and CAZ as indicators is considered to be the best AmpC confirmation assay. Finally, in Enterobacteriaceae isolates we suggest to screen for carbapenemases with a 0.5 μg/mL meropenem screening breakpoint. For class A carbapenemase confirmation, the home-prepared as well as the commercially available boronic acid-based CDDST can be considered. For metallo-β-lactamase confirmation, ethylene diamine tetra-acetic-acid-based home-prepared assays are recommended. The most suitable method (CDDST or DDST) and indicator antimicrobial agent(s) vary depending on the bacterial genus. Copyright © 2012 Elsevier B.V. and the International Society of Chemotherapy. All rights reserved.

Screening for antimicrobial and proteolytic activities of lactic acid bacteria isolated from cow, buffalo and goat milk and cheeses marketed in the southeast region of Brazil.

PubMed

Tulini, Fabricio L; Hymery, Nolwenn; Haertlé, Thomas; Le Blay, Gwenaelle; De Martinis, Elaine C P

2016-02-01

Lactic acid bacteria (LAB) can be isolated from different sources such as milk and cheese, and the lipolytic, proteolytic and glycolytic enzymes of LAB are important in cheese preservation and in flavour production. Moreover, LAB produce several antimicrobial compounds which make these bacteria interesting for food biopreservation. These characteristics stimulate the search of new strains with technological potential. From 156 milk and cheese samples from cow, buffalo and goat, 815 isolates were obtained on selective agars for LAB. Pure cultures were evaluated for antimicrobial activities by agar antagonism tests and for proteolytic activity on milk proteins by cultivation on agar plates. The most proteolytic isolates were also tested by cultivation in skim milk followed by sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) analysis of the fermented milk. Among the 815 tested isolates, three of them identified as Streptococcus uberis (strains FT86, FT126 and FT190) were bacteriocin producers, whereas four other ones identified as Weissella confusa FT424, W. hellenica FT476, Leuconostoc citreum FT671 and Lactobacillus plantarum FT723 showed high antifungal activity in preliminary assays. Complementary analyses showed that the most antifungal strain was L. plantarum FT723 that inhibited Penicillium expansum in modified MRS agar (De Man, Rogosa, Sharpe, without acetate) and fermented milk model, however no inhibition was observed against Yarrowia lipolytica. The proteolytic capacities of three highly proteolytic isolates identified as Enterococcus faecalis (strains FT132 and FT522) and Lactobacillus paracasei FT700 were confirmed by SDS-PAGE, as visualized by the digestion of caseins and whey proteins (β-lactoglobulin and α-lactalbumin). These results suggest potential applications of these isolates or their activities (proteolytic activity or production of antimicrobials) in dairy foods production.

Comparing Diagnostic Accuracy of Kato-Katz, Koga Agar Plate, Ether-Concentration, and FLOTAC for Schistosoma mansoni and Soil-Transmitted Helminths

PubMed Central

Glinz, Dominik; Silué, Kigbafori D.; Knopp, Stefanie; Lohourignon, Laurent K.; Yao, Kouassi P.; Steinmann, Peter; Rinaldi, Laura; Cringoli, Giuseppe; N'Goran, Eliézer K.; Utzinger, Jürg

2010-01-01

Background Infections with schistosomes and soil-transmitted helminths exert a considerable yet underappreciated economic and public health burden on afflicted populations. Accurate diagnosis is crucial for patient management, drug efficacy evaluations, and monitoring of large-scale community-based control programs. Methods/Principal Findings The diagnostic accuracy of four copromicroscopic techniques (i.e., Kato-Katz, Koga agar plate, ether-concentration, and FLOTAC) for the detection of Schistosoma mansoni and soil-transmitted helminth eggs was compared using stool samples from 112 school children in Côte d'Ivoire. Combined results of all four methods served as a diagnostic ‘gold’ standard and revealed prevalences of S. mansoni, hookworm, Trichuris trichiura, Strongyloides stercoralis and Ascaris lumbricoides of 83.0%, 55.4%, 40.2%, 33.9% and 28.6%, respectively. A single FLOTAC from stool samples preserved in sodium acetate-acetic acid-formalin for 30 or 83 days showed a higher sensitivity for S. mansoni diagnosis (91.4%) than the ether-concentration method on stool samples preserved for 40 days (85.0%) or triplicate Kato-Katz using fresh stool samples (77.4%). Moreover, a single FLOTAC detected hookworm, A. lumbricoides and T. trichiura infections with a higher sensitivity than any of the other methods used, but resulted in lower egg counts. The Koga agar plate method was the most accurate diagnostic assay for S. stercoralis. Conclusion/Significance We have shown that the FLOTAC method holds promise for the diagnosis of S. mansoni. Moreover, our study confirms that FLOTAC is a sensitive technique for detection of common soil-transmitted helminths. For the diagnosis of S. stercoralis, the Koga agar plate method remains the method of choice. PMID:20651931

FIZICS: fluorescent imaging zone identification system, a novel macro imaging system.

PubMed

Skwish, Stephen; Asensio, Francisco; King, Greg; Clarke, Glenn; Kath, Gary; Salvatore, Michael J; Dufresne, Claude

2004-12-01

Constantly improving biological assay development continues to drive technological requirements. Recently, a specification was defined for capturing white light and fluorescent images of agar plates ranging in size from the NUNC Omni tray (96-well footprint, 128 x 85 mm) to the NUNC Bio Assay Dish (245 x 245 mm). An evaluation of commercially available products failed to identify any system capable of fluorescent macroimaging with discrete wavelength selection. To address the lack of a commercially available system, a custom imaging system was designed and constructed. This system provides the same capabilities of many commercially available systems with the added ability to fluorescently image up to a 245 x 245 mm area using wavelengths in the visible light spectrum.

[Short-term screening of anticarcinogenic ingredients of tea by cell biology assays].

PubMed

Liu, L; Han, C; Chen, J

1998-01-01

By using a panel of short term cell biology assays, several ingredients of tea (tea pigments, caffeine, tea polysaccharide, tea polyphenols tablet and mixed tea) were screened in order to investigate their anticarcinogenic effects. The cytokinesis block micronuclei test in V79 cells induced by mitomycin, the test of metabolic cooperation between V79 and M cells and the test of growth ability of Hela cells in soft agar were used in the screening. The results showed that the six kinds of tea ingredients tested were effective in the test involved in different stages of carcinogenesis, i.e. initiation, promotion and progression. The effects of mixed tea and tea pigments were the strongest among the ingredients tested.

Comparison of two antifungal susceptibility testing of Candida sp. isolates using agar diffusion method: Neo-sensitabs® tablets and Bio-rad® disks.

PubMed

Uwingabiye, J; Iken, M; Zohoun, A G; Boumhil, L; Lemkhente, Z; Naoui, H; Bouchrik, M; Lmimouni, B

2016-03-01

The aim of our study was to evaluate the concordance between the two antifungal susceptibility testing of Candida sp. isolates using agar diffusion method: Neo-Sensitabs(®) tablets and Bio-Rad(®) disks. This is a prospective study conducted in the Laboratory of Parasitology and Mycology of the Mohammed V military teaching hospital from February to August 2012. Upon receiving blood cultures and peripheral sites samples, the identification of Candida isolates performed using routine phenotypic standard tests and the realization of the antifungal susceptibility was carried out on Neo-sensitabs(®) tablets and Bio-Rad(®) disks. A total of 38 Candida strains were isolated: 15 C. albicans (39%), 13 C. glabrata (34%), 5 C. tropicalis (13%), 4 C. krusei (11%) and 1 C. dubliniensis (3%). There were no significant difference (P>0.05) in susceptibility rate between both methods for all antifungal agents tested except for 5-fluorocytosine. The concordance percentage between two methods was 100% for amphotericin B, 97.4% for fluconazole, 94.7% for voriconazole and 73% for 5-fluorocytosine. Both methods are easy to perform, rapid and cost effective. Our results showed the best agreement between the two methods for testing the susceptibility of Candida isolates to amphotericin B, fluconazole and voriconazole while for the 5-fluorocytosine, the concordance rate was low. Copyright © 2015 Elsevier Masson SAS. All rights reserved.

Antimicrobial effects of Citrus sinensis peel extracts against dental caries bacteria: An in vitro study

PubMed Central

Shetty, Sapna B.; Mahin-Syed-Ismail, Prabu; Varghese, Shaji; Thomas-George, Bibin; Kandathil- Thajuraj, Pathinettam; Baby, Deepak; Haleem, Shaista; Sreedhar, Sreeja

2016-01-01

Background Ethnomedicine is gaining admiration since years but still there is abundant medicinal flora which is unrevealed through research. The study was conducted to assess the in vitro antimicrobial potential and also determine the minimum inhibitory concentration (MIC) of Citrus sinensis peel extracts with a view of searching a novel extract as a remedy for dental caries pathogens. Material and Methods Aqueous and ethanol (cold and hot) extracts prepared from peel of Citrus sinensis were screened for in vitro antimicrobial activity against Streptococcus mutans and Lactobacillus acidophilus, using agar well diffusion method. The lowest concentration of every extract considered as the minimal inhibitory concentration (MIC) values were determined for both test organisms. One way ANOVA with Post Hoc Bonferroni test was applied for statistical analysis. Confidence level and level of significance were set at 95% and 5% respectively. Results Dental caries pathogens were inhibited most by hot ethanolic extract of Citrus sinensispeel followed by cold ethanolic extract. Aqueous extracts were effective at very high concentrations. Minimum inhibitory concentration of hot and cold ethanolic extracts of Citrus sinensis peel ranged between 12-15 mg/ml against both the dental caries pathogens. Conclusions Citrus sinensispeels extract was found to be effective against dental caries pathogens and contain compounds with therapeutic potential. Nevertheless, clinical trials on the effect of these plants are essential before advocating large-scale therapy. Key words:Agar well diffusion, antimicrobial activity, dental caries, Streptococcus mutans, Lactobacillus acidophilus. PMID:26855710

Differentiation between probiotic and wild-type Bacillus cereus isolates by antibiotic susceptibility test and Fourier transform infrared spectroscopy (FT-IR).

PubMed

Mietke, Henriette; Beer, W; Schleif, Julia; Schabert, G; Reissbrodt, R

2010-05-30

Animal feed often contains probiotic Bacillus strains used as feed additives. Spores of the non-pathogenic B. cereus var. toyoi (product name Toyocerin) are used. Distinguishing between toxic wild-type Bacillus cereus strains and this probiotic strain is essential for evaluating the quality and risk of feed. Bacillus cereus CIP 5832 (product name Paciflor was used as probiotic strain until 2001. The properties of the two probiotic strains are quite similar. Differentiating between probiotic strains and wild-type B. cereus strains is not easy. ss-lactam antibiotics such as penicillin and cefamandole exhibit an inhibition zone in the agar diffusion test of probiotic B. cereus strains which are not seen for wild-type strains. Therefore, performing the agar diffusion test first may make sense before FT-IR testing. When randomly checking these strains by Fourier transform infrared spectroscopy (FT-IR), the probiotic B. cereus strains were separated from wild-type B. cereus/B. thuringiensis/B. mycoides/B. weihenstephanensis strains by means of hierarchical cluster analysis. The discriminatory information was contained in the spectral windows 3000-2800 cm(-1) ("fatty acid region"), 1200-900 cm(-1) ("carbohydrate region") and 900-700 cm(-1) ("fingerprint region"). It is concluded that FT-IR spectroscopy can be used for the rapid quality control and risk analysis of animal feed containing probiotic B. cereus strains. (c) 2010 Elsevier B.V. All rights reserved.

Spectral Induced Polarization of Disseminated Pyrite Particles in Soil

NASA Astrophysics Data System (ADS)

Slater, L. D.; Kessouri, P.; Seleznev, N. V.

2017-12-01

Disseminated metallic particles in soil, particularly pyrite, occur naturally or are enhanced by anthropogenic activities. Detecting their presence and quantifying their concentration and location is of interest for numerous applications such as remediation of hydrocarbon contamination, mine tailings assessment, detection of oil traps, and archaeological studies. Because pyrite is a semiconductor, spectral induced polarization (SIP) is a promising geophysical method for sensing it in porous media. Previous studies have identified relations between pyrite properties (e.g., volumetric content, grain size) and SIP parameters (e.g., chargeability, relaxation time). However, the effect of pyrite grains in porous media on the SIP response is not fully understood over the entire low-frequency range. We tested the relationship between the presence of pyrite grains and the change in electrical properties of the medium through an extended series of laboratory measurements: (1) variation of grain size, (2) variation of grain concentration, (3) variation of electrolyte conductivity, (4) change in the diffusion properties of the host medium. For the fourth set of measurements, we compared sand columns to agar gel columns. Our experimental design included more than 20 different samples with multiple repeats to ensure representative results. We confirm the strong relation between grain size and relaxation time and that between grain concentration and chargeability in both the sand and agar gel samples. Furthermore, our results shed light on the significance of the diffusion coefficient and the recently hypothesized role of pyrite grains as resistors at frequencies lower than the relaxation frequency.

Substrate Diffusion Heterogeneity Controls Bacterial Competition and Coexistence

NASA Astrophysics Data System (ADS)

Dechesne, A.; Or, D.; Smets, B. F.

2005-12-01

Diffusion has long been recognized as a key process affecting bacterial physiological functions ranging from nutrient uptake to removal of metabolic waste products. In the vadose zone, significant convective flows are limited and bacteria rely primarily on diffusion for nutrient supply. Even under relatively "wet" conditions (e.g. matric potentials -20 J/kg), soil water is fragmented and exists as thin liquid films or held in crevices imposing constraints on substrate diffusion. Our objective was to investigate the role of diffusion on soil microbial diversity, by focusing on one of the processes that shapes the structure of bacterial communities: competitive interactions. We used a simplified setup, in which the substrate (citrate) fluxes were controlled by different agar gels thicknesses and spatially heterogeneous diffusive pathways were created by an impermeable film with prescribed hole sizes and patterns. Our competition experiments involved two soil bacteria: Burkholderia xenovorans LB400 and Pseudomonas putida KT2440, which were tagged with different constitutive fluorescent markers, allowing for their on line microscopic detection. The growth parameters on citrate of these strains were thoroughly assessed. B. xenovorans LB400 is the weaker competitor. As a result, this strain was outcompeted by KT2440 under high substrate diffusivity and homogeneous conditions. Conversely, the disadvantage of the weakest competitor was not so marked under low substrate diffusivity condition. These results suggest that dry conditions in soil would provide conditions allowing the sustaining of weak bacterial competitors, resulting in the maintenance of high bacterial diversity.

Comparison of the antibacterial activity of essential oils and extracts of medicinal and culinary herbs to investigate potential new treatments for irritable bowel syndrome

PubMed Central

2013-01-01

Background Irritable bowel syndrome (IBS) is a common functional gastrointestinal disorder, which may result from alteration of the gastrointestinal microbiota following gastrointestinal infection, or with intestinal dysbiosis or small intestinal bacterial overgrowth. This may be treated with antibiotics, but there is concern that widespread antibiotic use might lead to antibiotic resistance. Some herbal medicines have been shown to be beneficial, but their mechanism(s) of action remain incompletely understood. To try to understand whether antibacterial properties might be involved in the efficacy of these herbal medicines, and to investigate potential new treatments for IBS, we have conducted a preliminary study in vitro to compare the antibacterial activity of the essential oils of culinary and medicinal herbs against the bacterium, Esherichia coli. Methods Essential oils were tested for their ability to inhibit E. coli growth in disc diffusion assays and in liquid culture, and to kill E. coli in a zone of clearance assay. Extracts of coriander, lemon balm and spearmint leaves were tested for their antibacterial activity in the disc diffusion assay. Disc diffusion and zone of clearance assays were analysed by two-tailed t tests whereas ANOVA was performed for the turbidometric assays. Results Most of the oils exhibited antibacterial activity in all three assays, however peppermint, lemon balm and coriander seed oils were most potent, with peppermint and coriander seed oils being more potent than the antibiotic rifaximin in the disc diffusion assay. The compounds present in these oils were identified by gas chromatography mass spectrometry. Finally, extracts were made of spearmint, lemon balm and coriander leaves with various solvents and these were tested for their antibacterial activity against E. coli in the disc diffusion assay. In each case, extracts made with ethanol and methanol exhibited potent antibacterial activity. Conclusions Many of the essential oils had antibacterial activity in the three assays, suggesting that they would be good candidates for testing in clinical trials. The observed antibacterial activity of ethanolic extracts of coriander, lemon balm and spearmint leaves suggests a mechanistic explanation for the efficacy of a mixture of coriander, lemon balm and mint extracts against IBS in a published clinical trial. PMID:24283351

Generation and precise control of dynamic biochemical gradients for cellular assays

NASA Astrophysics Data System (ADS)

Saka, Yasushi; MacPherson, Murray; Giuraniuc, Claudiu V.

2017-03-01

Spatial gradients of diffusible signalling molecules play crucial roles in controlling diverse cellular behaviour such as cell differentiation, tissue patterning and chemotaxis. In this paper, we report the design and testing of a microfluidic device for diffusion-based gradient generation for cellular assays. A unique channel design of the device eliminates cross-flow between the source and sink channels, thereby stabilizing gradients by passive diffusion. The platform also enables quick and flexible control of chemical concentration that makes highly dynamic gradients in diffusion chambers. A model with the first approximation of diffusion and surface adsorption of molecules recapitulates the experimentally observed gradients. Budding yeast cells cultured in a gradient of a chemical inducer expressed a reporter fluorescence protein in a concentration-dependent manner. This microfluidic platform serves as a versatile prototype applicable to a broad range of biomedical investigations.

Cytotoxic and antibacterial activity of the mixture of olive oil and lime cream in vitro conditions.

PubMed

Sumer, Zeynep; Yildirim, Gulay; Sumer, Haldun; Yildirim, Sahin

2013-01-01

The mixture of olive oil and lime cream has been traditionally used to treat external burns in the region of Hatay/Antakya and middle Anatolia. Olive oil and lime cream have been employed by many physicians to treat many ailments in the past. A limited number of studies have shown the antibacterial effect of olive oil and that it does not have any toxic effect on the skin. But we did not find any reported studies on the mixture of olive oil and lime cream. The aim of this paper is to investigate the cytotoxic and antibacterial activity of olive oil and lime cream individually or/and in combination in vitro conditions, by using disk-diffusion method and in cell culture. The main purpose in using this mixture is usually to clear burns without a trace. Agar overlay, MTT (Cytotoxicity assay) and antibacterial susceptibility tests were used to investigate the cytotoxic and antibacterial activity of olive oil and lime cream. We found that lime cream has an antibacterial activity but also cytotoxic on the fibroblasts. On the other hand olive oil has limited or no antibacterial effect and it has little or no cytotoxic on the fibroblasts. When we combined lime cream and olive oil, olive oil reduced its cytotoxic impact. These results suggest that mixture of olive oil and lime cream is not cytotoxic and has antimicrobial activity.

Ibuprofen reverts antifungal resistance on Candida albicans showing overexpression of CDR genes.

PubMed

Ricardo, Elisabete; Costa-de-Oliveira, Sofia; Dias, Ana Silva; Guerra, José; Rodrigues, Acácio Gonçalves; Pina-Vaz, Cidália

2009-06-01

Several mechanisms may be associated with Candida albicans resistance to azoles. Ibuprofen was described as being able to revert resistance related to efflux activity in Candida. The aim of this study was to uncover the molecular base of antifungal resistance in C. albicans clinical strains that could be reverted by ibuprofen. Sixty-two clinical isolates and five control strains of C. albicans were studied: the azole susceptibility phenotype was determined according to the Clinical Laboratory for Standards Institute, M27-A2 protocol and minimal inhibitory concentration values were recalculated with ibuprofen (100 microg mL(-1)); synergistic studies between fluconazole and FK506, a Cdr1p inhibitor, were performed using an agar disk diffusion assay and were compared with ibuprofen results. Gene expression was quantified by real-time PCR, with and without ibuprofen, regarding CDR1, CDR2, MDR1, encoding for efflux pumps, and ERG11, encoding for azole target protein. A correlation between susceptibility phenotype and resistance gene expression profiles was determined. Ibuprofen and FK506 showed a clear synergistic effect when combined with fluconazole. Resistant isolates reverting to susceptible after incubation with ibuprofen showed CDR1 and CDR2 overexpression especially of the latter. Conversely, strains that did not revert displayed a remarkable increase in ERG11 expression along with CDR genes. Ibuprofen did not alter resistance gene expression significantly (P>0.05), probably acting as a Cdrp blocker.

Biological Properties and Characterization of ASL50 Protein from Aged Allium sativum Bulbs.

PubMed

Kumar, Suresh; Jitendra, Kumar; Singh, Kusum; Kapoor, Vaishali; Sinha, Mou; Xess, Immaculata; Das, Satya N; Sharma, Sujata; Singh, Tej P; Dey, Sharmistha

2015-08-01

Allium sativum is well known for its medicinal properties. The A. sativum lectin 50 (ASL50, 50 kDa) was isolated from aged A. sativum bulbs and purified by gel filtration chromatography on Sephacryl S-200 column. Agar well diffusion assay were used to evaluate the antimicrobial activity of ASL50 against Candida species and bacteria then minimal inhibitory concentration (MIC) was determined. The lipid A binding to ASL50 was determined by surface plasmon resonance (SPR) technology with varying concentrations. Electron microscopic studies were done to see the mode of action of ASL50 on microbes. It exerted antimicrobial activity against clinical Candida isolates with a MIC of 10-40 μg/ml and clinical Pseudomonas aeruginosa isolates with a MIC of 10-80 μg/ml. The electron microscopic study illustrates that it disrupts the cell membrane of the bacteria and cell wall of fungi. It exhibited antiproliferative activity on oral carcinoma KB cells with an IC50 of 36 μg/ml after treatment for 48 h and induces the apoptosis of cancer cells by inducing 2.5-fold higher caspase enzyme activity than untreated cells. However, it has no cytotoxic effects towards HEK 293 cells as well as human erythrocytes even at higher concentration of ASL50. Biological properties of ASL50 may have its therapeutic significance in aiding infection and cancer treatments.

Inhibitory effects of antiseptic mouthrinses on Streptococcus mutans, Streptococcus sanguinis and Lactobacillus acidophilus.

PubMed

Evans, A; Leishman, S J; Walsh, L J; Seow, W K

2015-06-01

Oral antiseptics are valuable in controlling oral infections caused by cariogenic bacteria. The aim of this study was to investigate the effects of mouthrinses and pure antiseptic compounds on Streptococcus mutans and non-mutans bacteria (Streptococcus sanguinis and Lactobacillus acidophilus). The agar diffusion assay was employed to determine bacterial growth inhibition. Commercial mouthrinses containing chlorhexidine gluconate (0.2%), cetylpyridinium chloride (0.05%) and sodium fluoride (0.05%) produced statistically similar growth inhibition of S. mutans, S. sanguinis and L. acidophilus (with zones of inhibition ranging from 7.56 ± 0.52 mm to 7.39 ± 0.53 mm, 17.44 ± 0.94 mm to 18.31 ± 0.62 mm and 8.61 ± 1.43 to 8.67 ± 1.43 mm respectively, p > 0.05). The chlorhexidine mouthwash produced the greatest mean growth inhibition of S. sanguinis and S. mutans compared to all other mouthrinses tested (p < 0.01). The minimum concentrations at which inhibition against S. mutans could be detected were chlorhexidine gluconate at 0.005% (wt/vol), cetylpyridinium chloride 0.01% (wt/ vol), povidone iodine 10% (wt/vol) and sodium hypochlorite 0.5% (vol/vol). Chlorhexidine (0.01%), cetylpyridinium chloride (0.01%), povidone iodine (10%) and sodium hypochlorite (0.5%) are effective at inhibiting the growth of S. mutans, S. sanguinis and L. acidophilus. © 2015 Australian Dental Association.

Higher efficiency soluble prokaryotic expression, purification, and structural analysis of antimicrobial peptide G13.

PubMed

Che, Yuanyuan; Lu, Yinghu; Zha, Xiangdong; Huang, Huoqing; Yang, Peilong; Ma, Lijuan; Xu, Xuejiao

2016-03-01

G13 is a 19-residue cationic antimicrobial peptide derived from granulysin. In order to achieve high-level expression of G13 in Escherichia coli cells, and to reduce downstream processing costs, we introduced an Asp-Pro acid labile bond between the His-Patch thioredoxin and G13 and constructed the recombinant plasmid pThiohisA-DP-G13. The plasmid was transformed into E. coli BL21 (DE3). After induction with isopropyl-β-d-thiogalactopyranoside for 5 h, the fusion protein accumulated up to 200 mg/L in soluble form. The fusion protein was released by a high pressure homogenizer, cleaved using 13% acetic acid at 50 °C hydrolysis for 72 h. The recombinant G13 (r-G13) was then successively purified by fractional precipitation with ammonium sulfate and trichloroacetic acid, followed by one-step cation exchange chromatography. The purified r-G13 displayed a single band (about 2.2 kDa) as analyzed by Tris-Tricine buffered SDS-PAGE, and its precise molecular weight was confirmed using tandem mass spectrometry. Analysis of r-G13 by circular dichroism (CD) indicated that r-G13 contained predominantly β-sheet and random coil. Agar plate diffusion assay revealed that the r-G13 exhibited antibacterial activity against both Bacillus subtilis and E. coli. Copyright © 2015 Elsevier Inc. All rights reserved.

Structure elucidation and in silico docking studies of a novel furopyrimidine antibiotics synthesized by endolithic bacterium Actinomadura sp. AL2.

PubMed

Bhattacharjee, Kaushik; Kumar, Shakti; Palepu, Narasinga Rao; Patra, Pradeep Kumar; Rao, Kollipara Mohan; Joshi, Santa Ram

2017-09-20

On screening of endolithic actinobacteria from a granite rock sample of Meghalaya for antibacterial compound, a novel antibacterial compound CCp1 was isolated from the fermentation broth of Actinomadura sp. AL2. On purification of the compound based on chromatographic techniques followed by characterization with FT-IR, UV-visible, 1 H NMR, 13 C NMR and mass spectrometry, the molecular formula of the compound was generated as C 20 H 17 N 3 O 2 , a furopyrimidine derivative. In vitro antibacterial activity of the compound was evaluated against both Gram positive and negative bacteria by agar well diffusion assay. The compound had lowest MIC (2.00 µg/ml) for Bacillus subtilis and highest MIC (> 64 µg/ml) for Staphylococcus epidermidis and Pseudomonas aeruginosa. The study revealed that the compound has potential antibacterial activity. The mode of action of the antibacterial compound was evaluated through in silico studies for its ability to bind DNA gyrase, 30S RNA molecules, OmpF porins and N-Acetylglucosamine-1-phosphate uridyltransferase (GlmU). The antibacterial compound demonstrated more favorable docking with DNA gyrase, 30S RNA molecules and OmpF porins than GlmU which support the antibacterial compound CCp1 can be as a promising broad spectrum antibiotic agent with "multitarget" characteristics.

Structural characterization and biomedical properties of sulfated polysaccharide from the gladius of Sepioteuthis lessoniana (Lesson, 1831).

PubMed

Seedevi, Palaniappan; Moovendhan, Meivelu; Vairamani, Shanmugam; Shanmugam, Annian

2016-04-01

Sulfated polysaccharide was extracted from the internal shell (gladius) of Sepioteuthis lessoniana. The sulfated polysaccharide contained 61.3% of carbohydrate, 0.8% of protein, 28.2% of ash and 1.33% of moisture respectively. The elemental composition was analyzed using CHNS/O analyzer. The molecular weight of sulfated polysaccharide determined through PAGE was found to be as 66 kDa. Monosaccharides analysis revealed that sulfated polysaccharide was composed of rhamnose, galactose, xylose and glucose. The structural features of sulfated polysaccharide were analyzed by FT-IR and NMR spectroscopy. Further the sulfated polysaccharide was evaluated for its antibacterial activity against selected human clinical pathogens, namely Staphylococcus aureus, Klebsiella pneumoniae, Salmonella typhi, Vibrio cholerae, Klebsiella oxytoca, Escherichia coli, Salmonella paratyphi, Proteus mirabilis, Vibrio parahaemolyticus and Streptococcus pyogenes using agar well diffusion method. The polysaccharide has showed good antibacterial activity and MIC and MBC have also been evaluated. The anticancer activity was tested against HeLa cell line by MTT assay. The Cytotoxic Concentration (CC50) was observed as 700 μg/ml and the maximum anticancer activity of 62.89% was recorded at 200 μg/ml; whereas, the lowest of 9.87% was observed at 25 μg/ml. In conclusion, the sulfated polysaccharide is an alternate, non-toxic and cheap source of substance that showed good antibacterial and anticancer acitivity. Copyright © 2015 Elsevier B.V. All rights reserved.

Inhibitory effect of essential oils against Trichosporon ovoides causing Piedra Hair Infection.

PubMed

Saxena, Seema; Uniyal, Veena; Bhatt, R P

2012-10-01

Piedra, is an asymptomatic fungal infection of the hair shaft, resulting in the formation of nodules of different hardness on the infected hair. The infection also known as Trichomycosis nodularis is a superficial fungal infection arising from the pathogen being restricted to the stratum corneum with little or no tissue reaction. The nodules are a concretion of hyphae and fruiting bodies of the fungus. Two varieties of Piedra may be seen, Black Piedra and White Piedra. The fungus Trichosporon ovoides is involved in the occurrence of both types of Piedras. The purpose of this study was to examine the effectiveness of selected essential oils for the control of growth of the fungus and to determine whether the antifungal effect was due to the major compounds of the oils. Two screening methods viz. Agar well diffusion assay and Minimum Inhibitory Concentration were adopted for the study. MIC and MFC were determined by tube dilution method. Essential oils from Eucalyptus, Ocimum basilicum, Mentha piperita, Cymbopogon flexuosus, Cymbopogon winterians, Trachyspermum ammi, Zingiber officinalis, Citrus limon, Cinnamomon zeylanicum, Salvia sclarea, Citrus aurantifolia, Melaleuca alternifolia, Citrus aurantium, Citrus bergamia, Pogostemon pathchouli, Cedrus atlantica, Jasminum officinale, Juniperus communis, Abelmoschus moschatus, Cyperus scariosus, Palargonium graveolens, Boswellia carterii, Rosa damascene, Veteveria zizanoides and Commiphora myrrha were evaluated. The essential oils of Cymbopogon winterians, Mentha piperita, Cinnamomum zeylanicum, Melaleuca alternifolia and Eucalyptus globulus were proved to be most effective against the fungus Trichosporon ovoides.

Chemical composition and biological activity of the essential oil from Thymus lanceolatus.

PubMed

Khadir, Abdelmounaim; Sobeh, Mansour; Gad, Haidy A; Benbelaid, Fethi; Bendahou, Mourad; Peixoto, Herbenya; Sporer, Frank; Ashour, Mohamed L; Wink, Michael

2016-01-01

Thymus lanceolatus is a rare species, which grows wild in Algeria and Tunis. It is used traditionally as a drink and to flavor and preserve meat and poultry. The composition of the essential oil was determined by GLC/FID and GLC/MS. Forty-nine components were identified and quantified, accounting for 96.75% of the total detected components in the oil. The oxygenated monoterpenes (74.85%) constitute the major class of volatile secondary metabolites in the oil. Thymol was the most abundant constituent (69.61%) followed by γ-terpinene (8.38%). The antioxidant activity was evaluated using both diphenylpicrylhydrazyl (DPPH˙) reduction and 2-deoxyribose (2-DR) degradation prevention methods. The oil showed a very potent antioxidant activity with IC(50) values of 0.20 ± 0.07 and 4.96 ± 0.39 μg/mL for the DPPH˙ and 2-DR methods, respectively. The antimicrobial activity of the oil was assessed using the agar diffusion method, and the in vitro cytotoxicity on five different cancer cells was examined using the MTT assay. The oil revealed promising inhibitory activity against Gram positive bacteria, especially Bacillus subtilis and Streptococcus pyogenes with an MIC value of 62.5 μg/mL. Additionally, the highest cytotoxic activity was observed against the HL-60 cells with an IC(50) of 113.5 μg/mL. These results validate some of their traditional uses in food preservation.

Comparative Study of Hydroalcoholic Extracts of Momordica charantia L. against Foodborne Pathogens

PubMed Central

Rakholiya, Kalpna; Vaghela, P.; Rathod, T.; Chanda, Sumitra

2014-01-01

The antimicrobial effect of 24 different hydroalcoholic extracts (100, 75, 50 and 25% methanol and water) obtained from four parts (leaf+stem (aerial), peel, pulp and seed) of Momordica charantia L. were investigated against five Gram-positive, six Gram-negative and four fungal strains. The extraction was done by individual cold percolation method using hexane, different hydroalcoholic solvent (100, 75, 50 and 25% methanol) and water. The antimicrobial activity was done by agar well diffusion assay. The extracts, which showed >15 mm zone of inhibition, were further screened to determine minimum inhibitory concentration and minimum bactericidal concentration using a broth dilution method performed in 96-well microtitre plate. The extractive yield was highest in aqueous extracts of all the four parts closely followed by 25% methanol. Micrococcus flavus was the most susceptible Gram-positive bacteria and Pseudomonas testosteroni was the most susceptible Gram-negative bacteria. The highest antibacterial activity was shown by 100% methanol. The Gram-negative Pseudomonas spp. was more susceptible towards all the extracts than the Gram-positive bacteria or fungal strains investigated. One hundred percent and 50% methanol extracts of seed showed lowest minimum inhibitory concentration and minimum bactericidal concentration values, that is <39 and 625 μg/ml, respectively, against Pseudomonas pictorum. Therefore, these extracts would be of interest in the control of Pseudomonas spp. in food industry as well as used for therapeutic purposes. PMID:24843188

Elucidating pharmacodynamic interaction of silver nanoparticle - topical deliverable antibiotics.

PubMed

Thirumurugan, G; Seshagiri Rao, J V L N; Dhanaraju, M D

2016-07-18

In order to exploit the potential benefits of antimicrobial combination therapy, we need a better understanding of the circumstances under which pharmacodynamic interactions expected. In this study, Pharmacodynamic interactions between silver nanoparticle (SNP) and topical antibiotics such as Cefazolin (CEF), Mupirocin (MUP), Gentamycin (GEN), Neomycin (NEO), Tetracycline (TET), Vancomycin (VAN) were investigated using the MIC test, Combination assay followed by Fractional Inhibitory concentration Index and Agar well diffusion method. SNP + MUP, SNP + NEO, SNP + VAN combinations showed Synergism (SN) and SNP + CEF, SNP + GEN, SNP + TET showed Partial synergism (PS) against Staphylococcus aureus. Four combinations (SNP + CEF, SNP + MUP, SNP + GEN, SNP + VAN) showed SN, SNP + TET showed PS and Indifferent effect (ID) were observed for SNP + NEO against Pseudomonas aeruginosa. SN was observed for SNP + CEF, SNP + GEN, SNP + NEO, SNP + TET and SNP + MUP showed ID, SNP + VAN showed PS against Escherichia coli. In addition, we elucidated the possible mechanism involved in the pharmacodynamic interaction between SNP-topical antibiotics by increased ROS level, membrane damage following protein release, K(+) leakage and biofilm inhibition. Thus, our findings support that conjugation of the SNP with topical antibiotics have great potential in the topical formulation when treating complex resistant bacterial infections and where there is a need of more concentration to kill pathogenic bacteria.

Evaluation of extracts and essential oil from Callistemon viminalis leaves: antibacterial and antioxidant activities, total phenolic and flavonoid contents.

PubMed

Salem, Mohamed Z M; Ali, Hayssam M; El-Shanhorey, Nader A; Abdel-Megeed, Ahmed

2013-10-01

To investigate antioxidant and antibacterial activities of Callistemon viminalis (C. viminalis) leaves. The essential oil of C. viminalis leaves obtained by hydro-distillation was analyzed by GC/MS. Different extracts were tested for total phenolic and flavonoid contents and in vitro antioxidant (DPPH assay) and antibacterial (agar disc diffusion and 96-well micro-plates methods) actives. Fourteen components were identified in the essential oil, representing 98.94% of the total oil. The major components were 1,8-cineole (64.53%) and α-pinene (9.69%). Leaf essential oil exhibited the highest antioxidant activity of (88.60±1.51)% comparable to gallic acid, a standard compound [(80.00±2.12)%]. Additionally, the biggest zone of inhibitions against the studied bacterial strains was observed by the essential oil when compared to the standard antibiotic (tetracycline). The crude methanol extract and ethyl acetate fraction had a significant antibacterial activity against the tested bacterial strains. It can be suggested that C. viminalis is a great potential source of antibacterial and antioxidant compounds useful for new antimicrobial drugs from the natural basis. The present study revealed that the essential oil as well as the methanol extracts and ethyl acetate fraction of C. viminalis leaves exhibited highly significant antibacterial activity against the tested bacterial strains. Copyright © 2013 Hainan Medical College. Published by Elsevier B.V. All rights reserved.

Antitumour and antimicrobial activities of endophytic streptomycetes from pharmaceutical plants in rainforest.

PubMed

Li, J; Zhao, G-Z; Chen, H-H; Wang, H-B; Qin, S; Zhu, W-Y; Xu, L-H; Jiang, C-L; Li, W-J

2008-12-01

The aim of this study was to screen antitumour and antimicrobial activities of endophytic actinomycetes isolated from pharmaceutical plants in rainforest in Yunnan province, China. Antitumour activity was studied by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay and antimicrobial activity was determined by agar well diffusion method. The high bioactive endophytic isolates were identified and further investigated for the presence of polyketide synthases (PKS-I, PKS-II) and nonribosomal peptide synthetases (NRPS) sequences by specific amplification. The molecular identification confirmed that the 41 isolates showed significant activities were members of the genus Streptomyces. Among them, 31.7% of endophytic streptomycete cultures were cytotoxic against A549 cells, 29.3% against HL-60 cells, 85.4% against BEL-7404 cells, 90.2% against P388D1 cells, 65.9% were active against Escherichia coli, 24.4% against Staphylococcus aureus, 31.7% against Staphylococcus epidermidis, 12.2% against Candida albicans and no strain displayed antagonistic activity against Klebsiella pneumoniae. High frequencies of positive PCR amplification were obtained for PKS-I (34.1%), PKS-II (63.4%) and NRPS (61.0%) biosynthetic systems. Many endophytic streptomycetes isolated from pharmaceutical plants in rainforest possess remarkable and diverse antitumour and antimicrobial bioactivities. These endophytic streptomycetes are precious resources obtained from rainforests, and they could be a promising source for bioactive agents.

Elucidating pharmacodynamic interaction of silver nanoparticle - topical deliverable antibiotics

NASA Astrophysics Data System (ADS)

Thirumurugan, G.; Seshagiri Rao, J. V. L. N.; Dhanaraju, M. D.

2016-07-01

In order to exploit the potential benefits of antimicrobial combination therapy, we need a better understanding of the circumstances under which pharmacodynamic interactions expected. In this study, Pharmacodynamic interactions between silver nanoparticle (SNP) and topical antibiotics such as Cefazolin (CEF), Mupirocin (MUP), Gentamycin (GEN), Neomycin (NEO), Tetracycline (TET), Vancomycin (VAN) were investigated using the MIC test, Combination assay followed by Fractional Inhibitory concentration Index and Agar well diffusion method. SNP + MUP, SNP + NEO, SNP + VAN combinations showed Synergism (SN) and SNP + CEF, SNP + GEN, SNP + TET showed Partial synergism (PS) against Staphylococcus aureus. Four combinations (SNP + CEF, SNP + MUP, SNP + GEN, SNP + VAN) showed SN, SNP + TET showed PS and Indifferent effect (ID) were observed for SNP + NEO against Pseudomonas aeruginosa. SN was observed for SNP + CEF, SNP + GEN, SNP + NEO, SNP + TET and SNP + MUP showed ID, SNP + VAN showed PS against Escherichia coli. In addition, we elucidated the possible mechanism involved in the pharmacodynamic interaction between SNP-topical antibiotics by increased ROS level, membrane damage following protein release, K+ leakage and biofilm inhibition. Thus, our findings support that conjugation of the SNP with topical antibiotics have great potential in the topical formulation when treating complex resistant bacterial infections and where there is a need of more concentration to kill pathogenic bacteria.

Probiotic screening and safety evaluation of Lactobacillus strains from plants, artisanal goat cheese, human stools, and breast milk.

PubMed

Gotteland, Martin; Cires, Maria Jose; Carvallo, Claudia; Vega, Natalia; Ramirez, Maria Antonieta; Morales, Pamela; Rivas, Patricia; Astudillo, Fernanda; Navarrete, Paola; Dubos, Céline; Figueroa, Alvaro; Troncoso, Miriam; Ulloa, Carolina; Mizgier, Maria Luisa; Carrasco-Pozo, Catalina; Speisky, Hernan; Brunser, Oscar; Figueroa, Guillermo

2014-04-01

The aim of this study was to select autochthonous strains of Lactobacillus from stools of healthy infants and adults, human milk, artisanal goat cheese, and fruits and vegetables according to their probiotic properties and safety. From 421 strains of Lactobacillus isolated, 102 (24.2%) were shown to be tolerant to gastric pH and bile salts; they were used to determine their anti-Helicobacter pylori (agar diffusion assay), antioxidant (oxygen radical absorption capacity), and anti-inflammatory (inhibition of interleukin-8 release by tumor necrosis factor-α-stimulated HT-29 cells) activities as well as their ability to adhere to intestinal (Caco-2) and gastric (AGS) epithelial cells. Results obtained were compared with three commercial probiotic Lactobacillus rhamnosus GG, L. plantarum 299v, and L. johnsonii NCC533. The five strains most efficient according to these activities were subsequently identified by sequencing their 16S rRNA gene, their susceptibility to antibiotics was determined, and their safety evaluated in mice. One strain of L. plantarum was discarded due to the higher prevalence of liver bacterial translocation observed in the animals fed this strain. In conclusion, four autochthonous strains of L. rhamnosus were finally selected with probiotic properties and safety allowing their eventual use in human studies. These results contribute to increase the diversity of probiotic strains available for the development of nutraceuticals and functional foods.

One-pot facile green synthesis of biocidal silver nanoparticles

NASA Astrophysics Data System (ADS)

Nudrat Hazarika, Shabiha; Gupta, Kuldeep; Shamin, Khan Naseem Ahmed Mohammed; Bhardwaj, Pushpender; Boruah, Ratan; Yadav, Kamlesh K.; Naglot, Ashok; Deb, P.; Mandal, M.; Doley, Robin; Veer, Vijay; Baruah, Indra; Namsa, Nima D.

2016-07-01

The plant root extract mediated green synthesis method produces monodispersed spherical shape silver nanoparticles (AgNPs) with a size range of 15-30 nm as analyzed by atomic force and transmission electron microscopy. The material showed potent antibacterial and antifungal properties. Synthesized AgNPs display a characteristic surface plasmon resonance peak at 420 nm in UV-Vis spectroscopy. X-ray diffractometer analysis revealed the crystalline and face-centered cubic geometry of in situ prepared AgNPs. Agar well diffusion and a colony forming unit assay demonstrated the potent biocidal activity of AgNPs against Staphylococcus aureus, Escherichia coli, Bacillus subtilis, Klebsiella pneumoniae, Pseudomonas diminuta and Mycobacterium smegmatis. Intriguingly, the phytosynthesized AgNPs exhibited activity against pathogenic fungi, namely Trichophyton rubrum, Aspergillus versicolor and Candida albicans. Scanning electron microscopy observations indicated morphological changes in the bacterial cells incubated with silver nanoparticles. The genomic DNA isolated from the bacteria was incubated with an increasing concentration of AgNPs and the replication fidelity of 16S rDNA was observed by performing 18 and 35 cycles PCR. The replication efficiency of small (600 bp) and large (1500 bp) DNA fragments in the presence of AgNPs were compromised in a dose-dependent manner. The results suggest that the Thalictrum foliolosum root extract mediated synthesis of AgNPs could be used as a promising antimicrobial agent against clinical pathogens.

Biosurfactants production potential of native strains of Bacillus cereus and their antimicrobial, cytotoxic and antioxidant activities.

PubMed

Basit, Madiha; Rasool, Muhammad Hidayat; Naqvi, Syed Ali Raza; Waseem, Muhammad; Aslam, Bilal

2018-01-01

Present study was designed to evaluate the biosurfactant production potential by native strains of Bacillus cereus as well as determine their antimicrobial and antioxidant activities. The strains isolated from garden soil were characterized as B. cereus MMIC 1, MMIC 2 and MMIC 3. Biosurfactants were extracted as grey white precipitates. Optimum conditions for biosurfactant production were 37°C, the 7th day of incubation, 0.5% NaCl, pH 7.0. Moreover, corn steep liquor was the best carbon source. Biuret test, Thin Layer Chromatography (TLC), agar double diffusion and Fourier Transform Infrared Spectroscopy (FTIR) characterized the biosurfactants as cationic lipopeptides. Biosurfactants exhibited significant antibacterial and antifungal activity against S. aureus, E. coli, P. aeruginosa, K. pneumoniae, A. niger and C. albicans at 30 mg/ml. Moreover, they also possessed antiviral activity against NDV at 10 mg/ml. Cytotoxicity assay in BHK-21 cell lines revealed 63% cell survival at 10 mg/ml of biosurfactants and thus considered as safe. They also showed very good antioxidant activity by ferric-reducing activity and DPPH scavenging activity at 2 mg/ml. Consequently, the study offers an insight for the exploration of new bioactive molecules from the soil. It was concluded that lipopeptide biosurfactants produced from native strains of B. cereus may be recommended as safe antimicrobial, emulsifier and antioxidant agent.