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Sample records for broth microdilution reference

  1. Validation of EUCAST zone diameter breakpoints against reference broth microdilution.

    PubMed

    Bengtsson, S; Bjelkenbrant, C; Kahlmeter, G

    2014-06-01

    The European Committee on Antimicrobial Susceptibility Testing (EUCAST) began harmonizing clinical breakpoints in Europe 2002. In 2009, work to develop a disc diffusion method began and the first disc diffusion breakpoints calibrated to EUCAST clinical MIC breakpoints were published in December 2009. In this study we validated EUCAST clinical zone diameter breakpoints against the International Standard Organization (ISO) reference broth microdilution. A collection of 544 isolates (238 Gram-negative and 306 Gram-positive) were tested against a panel of antimicrobial agents. Antimicrobial susceptibility testing was performed with broth microdilution as described by ISO and disc diffusion in accordance with EUCAST methodology. Inhibition zone diameters and MIC values were interpreted and categorized (S, I and R) according to EUCAST clinical breakpoint table version 2.0. Categorical agreement (CA) as well as minor (mD), major (MD) and very major (VMD) discrepancies were determined. There was in general good correlation between susceptibility test results obtained with disc diffusion and broth microdilution. Overall CA was 97.3% for all combinations of organisms and antimicrobial agents (n = 5231) and the overall discrepancy rates were 110 (2.1%) mD, 24 (0.5%) MD and 7 (0.1%) VMD. The overall CA for Gram-positive and Gram-negative organisms were 98.7% (2346 tests) and 96.2% (2942 tests), respectively. Seven VMD were observed, five for Gram-positive organisms (coagulase negative staphylococci (n = 2) and Staphylococcus aureus (n = 3)) and two for Gram-negative organisms (Pseudomonas aeruginosa). Minor discrepancies were mainly observed in Gram-negatives and were related to different antimicrobial agents and species.

  2. A Reference Broth Microdilution Method for Dalbavancin In Vitro Susceptibility Testing of Bacteria that Grow Aerobically.

    PubMed

    Koeth, Laura M; DiFranco-Fisher, Jeanna M; McCurdy, Sandra

    2015-09-09

    Antimicrobial susceptibility testing (AST) is performed to assess the in vitro activity of antimicrobial agents against various bacteria. The AST results, which are expressed as minimum inhibitory concentrations (MICs) are used in research for antimicrobial development and monitoring of resistance development and in the clinical setting for antimicrobial therapy guidance. Dalbavancin is a semi-synthetic lipoglycopeptide antimicrobial agent that was approved in May 2014 by the Food and Drug Administration (FDA) for the treatment of acute bacterial skin and skin structure infections caused by Gram-positive organisms. The advantage of dalbavancin over current anti-staphylococcal therapies is its long half-life, which allows for once-weekly dosing. Dalbavancin has activity against Staphylococcus aureus (including both methicillin-susceptible S. aureus [MSSA] and methicillin-resistant S. aureus [MRSA]), coagulase-negative staphylococci, Streptococcus pneumoniae, Streptococcus anginosus group, β-hemolytic streptococci and vancomycin susceptible enterococci. Similar to other recent lipoglycopeptide agents, optimization of CLSI and ISO broth susceptibility test methods includes the use of dimethyl sulfoxide (DMSO) as a solvent when preparing stock solutions and polysorbate 80 (P80) to alleviate adherence of the agent to plastic. Prior to the clinical studies and during the initial development of dalbavancin, susceptibility studies were not performed with the use of P-80 and MIC results tended to be 2-4 fold higher and similarly higher MIC results were obtained with the agar dilution susceptibility method. Dalbavancin was first included in CLSI broth microdilution methodology tables in 2005 and amended in 2006 to clarify use of DMSO and P-80. The broth microdilution (BMD) procedure shown here is specific to dalbavancin and is in accordance with the CLSI and ISO methods, with step-by-step detail and focus on the critical steps added for clarity.

  3. Revised Reference Broth Microdilution Method for Testing Telavancin: Effect on MIC Results and Correlation with Other Testing Methodologies

    PubMed Central

    Mendes, Rodrigo E.; Rhomberg, Paul R.; Jones, Ronald N.

    2014-01-01

    The reference broth microdilution (BMD) antimicrobial susceptibility testing method for telavancin was revised to include dimethyl sulfoxide (DMSO) as a solvent and diluent for frozen-form panel preparation, following the CLSI recommendations for water-insoluble agents. Polysorbate 80 (P-80) was also added to the test medium to minimize proven drug losses associated with binding to plastic surfaces. Four hundred sixty-two Gram-positive isolates, including a challenge set of organisms with reduced susceptibilities to comparator agents, were selected and tested using the revised method for telavancin, and the MIC results were compared with those tested by the previously established method and several Sensititre dry-form BMD panel formulations. The revised method provided MIC results 2- to 8-fold lower than the previous method when tested against staphylococci and enterococci, resulting in MIC50 values of 0.03 to 0.06 μg/ml for staphylococci and 0.03 and 0.12 μg/ml for Enterococcus faecium and Enterococcus faecalis, respectively. Less-significant MIC decreases (1 to 2 log2 dilution steps) were observed when testing streptococci in broth supplemented with blood, which showed similar MIC50 values for both methods. However, Streptococcus pneumoniae had MIC50 results of 0.008 and 0.03 μg/ml when tested by the revised and previous methods, respectively. Highest essential agreement rates (≥94.0%) were noted for one candidate dry-form panel formulation compared to the revised test. The revised BMD method provides lower MIC results for telavancin, especially when tested against staphylococci and enterococci. This is secondary to the use of DMSO for panel production and the presence of P-80, which ensure the proper telavancin testing concentration and result in a more accurate MIC determination. Moreover, earlier studies where the previous method was applied underestimated the in vitro drug potency. PMID:25022579

  4. Quality control and reference guidelines for CLSI broth microdilution susceptibility method (M 38-A document) for amphotericin B, itraconazole, posaconazole, and voriconazole.

    PubMed

    Espinel-Ingroff, A; Fothergill, A; Ghannoum, M; Manavathu, E; Ostrosky-Zeichner, L; Pfaller, M; Rinaldi, M; Schell, W; Walsh, T

    2005-10-01

    Although standard conditions are available for testing the susceptibilities of filamentous fungi to antifungal agents by the Clinical and Laboratory Standards Institute (CLSI; formerly National Committee for Clinical Laboratory Standards) broth microdilution assay, quality control (QC) MIC limits have not been established for any mold-agent combination. This multicenter (eight-center) study documented the reproducibility of tests for one isolate of Paecilomyces variotii ATCC MYA-3630 and 11 other mold isolates (three isolates of Aspergillus fumigatus; two isolates of A. terreus; one isolate each of A. flavus, A. nidulans, Fusarium moniliforme, and F. solani; and two isolates of Scedosporium apiospermum) by the CLSI reference broth microdilution method (M 38-A document). Control limits (amphotericin B, 1 to 4 microg/ml; itraconazole, 0.06 to 0.5 microg/ml; posaconazole, 0.03 to 0.25 microg/ml; voriconazole, 0.015 to 0.12 microg/ml) for the selected QC P. variotii ATCC MYA-3630 were established by the analysis of replicate MIC results. Reference isolates and corresponding MIC ranges were also established for 6 of the 12 molds evaluated. MIC limits were not proposed for the other five molds tested due to low testing reproducibility for these isolates.

  5. Comparison of a highly automated 5-h susceptibility testing system, the Cobas-Bact, with two reference methods: Kirby-Bauer disk diffusion and broth microdilution.

    PubMed

    Murray, P R; Niles, A C; Heeren, R L

    1987-12-01

    The results of susceptibility tests performed with the Cobas-Bact system were compared with those of the Kirby-Bauer disk diffusion and the broth microdilution methods. The evaluation included tests with 24 antibiotics against 250 isolates of the family Enterobacteriaceae and 13 antibiotics against 100 gram-positive cocci. Complete agreements between the Cobas-Bact and Kirby-Bauer methods were 82.8 and 84.5% for gram-positive cocci and gram-negative bacilli, respectively. Agreements between the Cobas-Bact and broth microdilution methods were 76.7% for gram-positive cocci and 84.8% for gram-negative bacilli. Complete agreements between the Kirby-Bauer and broth microdilution methods were 87.0% for gram-positive cocci and 92.2% for gram-negative bacilli. Despite generally satisfactory results with most organism-antibiotic combinations tested, additional modifications of the Cobas-Bact system are required to reduce the number of major and very major discrepancies, as well as to permit testing of Pseudomonas spp. and other gram-negative nonfermentative bacilli.

  6. Comparison of a highly automated 5-h susceptibility testing system, the Cobas-Bact, with two reference methods: Kirby-Bauer disk diffusion and broth microdilution.

    PubMed Central

    Murray, P R; Niles, A C; Heeren, R L

    1987-01-01

    The results of susceptibility tests performed with the Cobas-Bact system were compared with those of the Kirby-Bauer disk diffusion and the broth microdilution methods. The evaluation included tests with 24 antibiotics against 250 isolates of the family Enterobacteriaceae and 13 antibiotics against 100 gram-positive cocci. Complete agreements between the Cobas-Bact and Kirby-Bauer methods were 82.8 and 84.5% for gram-positive cocci and gram-negative bacilli, respectively. Agreements between the Cobas-Bact and broth microdilution methods were 76.7% for gram-positive cocci and 84.8% for gram-negative bacilli. Complete agreements between the Kirby-Bauer and broth microdilution methods were 87.0% for gram-positive cocci and 92.2% for gram-negative bacilli. Despite generally satisfactory results with most organism-antibiotic combinations tested, additional modifications of the Cobas-Bact system are required to reduce the number of major and very major discrepancies, as well as to permit testing of Pseudomonas spp. and other gram-negative nonfermentative bacilli. PMID:3429627

  7. Evaluation of CLSI agar dilution method and Trek Sensititre broth microdilution panel for determining antimicrobial susceptibility of Streptococcus pneumoniae.

    PubMed

    Zhang, Sean X; Rawte, Prasad; Brown, Shirley; Lo, Steven; Siebert, Heather; Pong-Porter, Sylvia; Low, Donald E; Jamieson, Frances B

    2011-02-01

    Both the CLSI agar dilution method and Trek Sensititre broth microdilution panel for Streptococcus pneumoniae antimicrobial susceptibility testing were evaluated against the reference CLSI broth microdilution method using the most recently published CLSI breakpoints. While agar dilution was not an optimal method, the commercial panel appeared to be an acceptable method, with minor errors encountered for ceftriaxone, penicillin, and meropenem.

  8. Recommendation for a Standardised Method of Broth Microdilution Susceptibility Testing for Porcine Bordetella bronchiseptica

    PubMed Central

    Prüller, Sandra; Frömke, Cornelia; Kaspar, Heike; Klein, Günter; Kreienbrock, Lothar; Kehrenberg, Corinna

    2015-01-01

    The objective was to establish and standardise a broth microdilution susceptibility testing method for porcine Bordetella (B.) bronchiseptica. B. bronchiseptica isolates from different geographical regions and farms were genotyped by macrorestriction analysis and subsequent pulsed-field gel electrophoresis. One reference and one type strain plus two field isolates of B. bronchiseptica were chosen to analyse growth curves in four different media: cation-adjusted Mueller-Hinton broth (CAMHB) with and without 2% lysed horse blood, Brain-Heart-Infusion (BHI), and Caso broth. The growth rate of each test strain in each medium was determined by culture enumeration and the suitability of CAMHB was confirmed by comparative statistical analysis. Thereafter, reference and type strain and eight epidemiologically unrelated field isolates of B. bronchiseptica were used to test the suitability of a broth microdilution susceptibility testing method following CLSI-approved performance standards given in document VET01-A4. Susceptibility tests, using 20 antimicrobial agents, were performed in five replicates, and data were collected after 20 and 24 hours incubation and statistically analysed. Due to the low growth rate of B. bronchiseptica, an incubation time of 24 hours resulted in significantly more homogeneous minimum inhibitory concentrations after five replications compared to a 20-hour incubation. An interlaboratory comparison trial including susceptibility testing of 24 antimicrobial agents revealed a high mean level of reproducibility (97.9%) of the modified method. Hence, in a harmonization for broth microdilution susceptibility testing of B. bronchiseptica, an incubation time of 24 hours in CAMHB medium with an incubation temperature of 35°C and an inoculum concentration of approximately 5 x 105 cfu/ml was proposed. PMID:25910232

  9. In Vitro Antibiotic Susceptibilities of Burkholderia mallei (Causative Agent of Glanders) Determined by Broth Microdilution and E-Test

    PubMed Central

    Heine, Henry S.; England, Marilyn J.; Waag, David M.; Byrne, W. Russell

    2001-01-01

    In vitro susceptibilities to 28 antibiotics were determined for 11 strains of Burkholderia mallei by the broth microdilution method. The B. mallei strains demonstrated susceptibility to aminoglycosides, macrolides, quinolones, doxycycline, piperacillin, ceftazidime, and imipenem. For comparison and evaluation, 17 antibiotic susceptibilities were also determined by the E-test. E-test values were always lower than the broth dilution values. Establishing and comparing antibiotic susceptibilities of specific B. mallei strains will provide reference information for assessing new antibiotic agents. PMID:11408233

  10. Comparative evaluation of Etest and sensititre yeastone panels against the Clinical and Laboratory Standards Institute M27-A2 reference broth microdilution method for testing Candida susceptibility to seven antifungal agents.

    PubMed

    Alexander, Barbara D; Byrne, Terry C; Smith, Kelly L; Hanson, Kimberly E; Anstrom, Kevin J; Perfect, John R; Reller, L Barth

    2007-03-01

    To assess their utility for antifungal susceptibility testing in our clinical laboratory, the Etest and Sensititre methods were compared with the Clinical and Laboratory Standards Institute (CLSI) M27-A2 reference broth microdilution method. Fluconazole (FL), itraconazole (I), voriconazole (V), posaconazole (P), flucytosine (FC), caspofungin (C), and amphotericin B (A) were tested with 212 Candida isolates. Reference MICs were determined after 48 h of incubation, and Etest and Sensititre MICs were determined after 24 h and 48 h of incubation. Overall, excellent essential agreement (EA) between the reference and test methods was observed for Etest (95%) and Sensititre (91%). Etest showed an >or=92% EA for MICs for all drugs tested; Sensititre showed a >or=92% EA for MICs for I, FC, A, and C but 82% for FL and 85% for V. The overall categorical agreement (CA) was 90% for Etest and 88% for Sensititre; minor errors accounted for the majority of all categorical errors for both systems. Categorical agreement was lowest for Candida glabrata and Candida tropicalis with both test systems. Etest and Sensititre provided better CA at 24 h compared to 48 h for C. glabrata; however, CA for C. glabrata was <80% for FL with both test systems despite MIC determination at 24 h. Agreement between technologists for both methods was >or=98% for each agent against all organisms tested. Overall, Etest and Sensititre methods compared favorably with the CLSI reference method for determining the susceptibility of Candida. However, further evaluation of their performance for determining the MICs of azoles, particularly for C. glabrata, is warranted.

  11. Activities of Tedizolid and Linezolid Determined by the Reference Broth Microdilution Method against 3,032 Gram-Positive Bacterial Isolates Collected in Asia-Pacific, Eastern Europe, and Latin American Countries in 2014

    PubMed Central

    Pfaller, Michael A.; Flamm, Robert K.; Jones, Ronald N.; Farrell, David J.

    2016-01-01

    Tedizolid and linezolid in vitro activities against 3,032 Gram-positive pathogens collected in Asia-Pacific, Eastern European, and Latin American medical centers during 2014 were assessed. The isolates were tested for susceptibility by the current reference broth microdilution methods. Due to concern over the effect of MIC endpoint criteria on the results of testing the oxazolidinones tedizolid and linezolid, MIC endpoint values were read by two methods: (i) reading the MIC at the first well where the trailing began without regard for pinpoint trailing, according to CLSI M07-A10 and M100-S26 document instructions for reading linezolid (i.e., 80% inhibition of growth; these reads were designated tedizolid 80 and linezolid 80), and (ii) at 100% inhibition of growth (designated tedizolid 100 and linezolid 100). All Staphylococcus aureus, Streptococcus pyogenes, Streptococcus agalactiae, Streptococcus anginosus group, and Enterococcus faecalis isolates were inhibited at tedizolid 80 and 100 MIC values of 0.25 and 0.5, 0.25 and 0.25, 0.25 and 0.5, 0.12 and 0.25, and 0.5 and 1 μg/ml, respectively. Generally, MIC50 and MIC90 results for tedizolid 80 and linezolid 80 were one doubling dilution lower than those read at 100% inhibition. Tedizolid was 4- to 8-fold more potent than linezolid against all the isolates tested regardless of the MIC endpoint criterion used. Despite the differences in potency, >99.9% of isolates tested in this survey were susceptible to both linezolid and tedizolid using CLSI and EUCAST interpretive criteria. In conclusion, tedizolid demonstrated greater in vitro potency than linezolid against Gram-positive pathogens isolated from patients in medical centers across the Asia-Pacific region, Eastern Europe, and Latin America. PMID:27353270

  12. Comparison of cation-adjusted Mueller-Hinton broth with Iso-Sensitest broth for the NCCLS broth microdilution method.

    PubMed

    Koeth, L M; King, A; Knight, H; May, J; Miller, L A; Phillips, I; Poupard, J A

    2000-09-01

    Comparison of MIC results obtained in different parts of the world is currently difficult because of variations in methods. In this study, cation-adjusted Mueller-Hinton broth, the NCCLS-recommended medium, was compared with Iso-Sensitest broth, which is widely used in Europe. Microbroth dilution testing, using the NCCLS procedure, was performed on 124 Gram-positive (staphylococci and enterococci) and Gram-negative (Enterobacteriaceae and Pseudomonas aeruginosa) isolates from the CDC reference set, with the only variable being the medium used. Twelve antimicrobial agents were tested: amoxycillin-clavulanic acid, ampicillin, ciprofloxacin, erythromycin, gentamicin, imipenem, levofloxacin, oxacillin, gemifloxacin, trimethoprim- sulphamethoxazole, tetracycline and vancomycin. Vancomycin, erythromycin and oxacillin were only evaluated for the Gram-positive organisms. Trimethoprim-sulphamethoxazole was only evaluated for a subset of Gram-negative organisms because of off-scale results. The 124 isolates were tested in one American and one UK laboratory with two batches of cation-adjusted Mueller-Hinton broth and two of Iso-Sensitest broth. A statistical evaluation of the data used a 24 fully specified factorial analysis to determine if there were significant differences in results owing to Gram reaction, site of testing and type and/or batch of broth. In addition, the cumulative results for each antimicrobial agent in each broth were plotted against the range of MIC dilutions tested. MICs of ciprofloxacin, levofloxacin, gemifloxacin, gentamicin and tetracycline were slightly higher (half a doubling dilution) with Iso-Sensitest broth than with Mueller-Hinton broth. MIC results for the other antimicrobial agents were equivalent. Essential and category agreement rates were comparable for all agents (88.4-100% and 88.2-99.0%, respectively).

  13. In Vitro Antibiotic Susceptibilities of Yersinia pestis Determined by Broth Microdilution following CLSI Methods

    PubMed Central

    Hershfield, Jeremy; Marchand, Charles; Miller, Lynda; Halasohoris, Stephanie; Purcell, Bret K.; Worsham, Patricia L.

    2015-01-01

    In vitro susceptibilities to 45 antibiotics were determined for 30 genetically and geographically diverse strains of Yersinia pestis by the broth microdilution method at two temperatures, 28°C and 35°C, following Clinical and Laboratory Standards Institute (CLSI) methods. The Y. pestis strains demonstrated susceptibility to aminoglycosides, quinolones, tetracyclines, β-lactams, cephalosporins, and carbapenems. Only a 1-well shift was observed for the majority of antibiotics between the two temperatures. Establishing and comparing antibiotic susceptibilities of a diverse but specific set of Y. pestis strains by standardized methods and establishing population ranges and MIC50 and MIC90 values provide reference information for assessing new antibiotic agents and also provide a baseline for use in monitoring any future emergence of resistance. PMID:25583720

  14. In Vitro Antibiotic Susceptibilities of Francisella tularensis Determined by Broth Microdilution following CLSI Methods.

    PubMed

    Heine, Henry S; Miller, Lynda; Halasohoris, Stephanie; Purcell, Bret K

    2017-09-01

    In vitro susceptibilities for 47 antibiotics were determined in 30 genetic diverse strains of Francisella tularensis by the broth microdilution method following Clinical and Laboratory Standards Institute (CLSI) methods. The F. tularensis strains demonstrated susceptibility to aminoglycosides, fluoroquinolones, and tetracyclines. There was a distinct difference in macrolide susceptibilities between A and B type strains, as has been noted previously. The establishment and comparison of antibiotic susceptibilities of a diverse but specific set of F. tularensis strains by standardized methods and the establishment of population ranges and MIC50/90 values provide reference information for assessing new antibiotic agents and a baseline to monitor any future emergence of resistance, whether natural or intentional. Copyright © 2017 American Society for Microbiology.

  15. In vitro antibiotic susceptibilities of Yersinia pestis determined by broth microdilution following CLSI methods.

    PubMed

    Heine, Henry S; Hershfield, Jeremy; Marchand, Charles; Miller, Lynda; Halasohoris, Stephanie; Purcell, Bret K; Worsham, Patricia L

    2015-04-01

    In vitro susceptibilities to 45 antibiotics were determined for 30 genetically and geographically diverse strains of Yersinia pestis by the broth microdilution method at two temperatures, 28°C and 35°C, following Clinical and Laboratory Standards Institute (CLSI) methods. The Y. pestis strains demonstrated susceptibility to aminoglycosides, quinolones, tetracyclines, β-lactams, cephalosporins, and carbapenems. Only a 1-well shift was observed for the majority of antibiotics between the two temperatures. Establishing and comparing antibiotic susceptibilities of a diverse but specific set of Y. pestis strains by standardized methods and establishing population ranges and MIC50 and MIC90 values provide reference information for assessing new antibiotic agents and also provide a baseline for use in monitoring any future emergence of resistance. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  16. Verification of an Automated, Digital Dispensing Platform for At-Will Broth Microdilution-Based Antimicrobial Susceptibility Testing.

    PubMed

    Smith, Kenneth P; Kirby, James E

    2016-09-01

    With rapid emergence of multidrug-resistant bacteria, there is often a need to perform susceptibility testing for less commonly used or newer antimicrobial agents. Such testing can often be performed only by using labor-intensive, manual dilution methods and lies outside the capacity of most clinical labs, necessitating reference laboratory testing and thereby delaying the availability of susceptibility data. To address the compelling clinical need for microbiology laboratories to perform such testing in-house, we explored a novel, automated, at-will broth microdilution-based susceptibility testing platform. Specifically, we used the modified inkjet printer technology in the HP D300 digital dispensing system to dispense, directly from stock solutions into a 384-well plate, the 2-fold serial dilution series required for broth microdilution testing. This technology was combined with automated absorbance readings and data analysis to determine MICs. Performance was verified by testing members of the Enterobacteriaceae for susceptibility to ampicillin, cefazolin, ciprofloxacin, colistin, gentamicin, meropenem, and tetracycline in comparison to the results obtained with a broth microdilution reference standard. In precision studies, essential and categorical agreement levels were 96.8% and 98.3%, respectively. Furthermore, significantly fewer D300-based measurements were outside ±1 dilution from the modal MIC, suggesting enhanced reproducibility. In accuracy studies performed using a panel of 80 curated clinical isolates, rates of essential and categorical agreement and very major, major, and minor errors were 94%, 96.6%, 0%, 0%, and 3.4%, respectively. Based on these promising initial results, it is anticipated that the D300-based methodology will enable hospital-based clinical microbiology laboratories to perform at-will broth microdilution testing of antimicrobials and to address a critical testing gap.

  17. Verification of an Automated, Digital Dispensing Platform for At-Will Broth Microdilution-Based Antimicrobial Susceptibility Testing

    PubMed Central

    Smith, Kenneth P.

    2016-01-01

    With rapid emergence of multidrug-resistant bacteria, there is often a need to perform susceptibility testing for less commonly used or newer antimicrobial agents. Such testing can often be performed only by using labor-intensive, manual dilution methods and lies outside the capacity of most clinical labs, necessitating reference laboratory testing and thereby delaying the availability of susceptibility data. To address the compelling clinical need for microbiology laboratories to perform such testing in-house, we explored a novel, automated, at-will broth microdilution-based susceptibility testing platform. Specifically, we used the modified inkjet printer technology in the HP D300 digital dispensing system to dispense, directly from stock solutions into a 384-well plate, the 2-fold serial dilution series required for broth microdilution testing. This technology was combined with automated absorbance readings and data analysis to determine MICs. Performance was verified by testing members of the Enterobacteriaceae for susceptibility to ampicillin, cefazolin, ciprofloxacin, colistin, gentamicin, meropenem, and tetracycline in comparison to the results obtained with a broth microdilution reference standard. In precision studies, essential and categorical agreement levels were 96.8% and 98.3%, respectively. Furthermore, significantly fewer D300-based measurements were outside ±1 dilution from the modal MIC, suggesting enhanced reproducibility. In accuracy studies performed using a panel of 80 curated clinical isolates, rates of essential and categorical agreement and very major, major, and minor errors were 94%, 96.6%, 0%, 0%, and 3.4%, respectively. Based on these promising initial results, it is anticipated that the D300-based methodology will enable hospital-based clinical microbiology laboratories to perform at-will broth microdilution testing of antimicrobials and to address a critical testing gap. PMID:27335151

  18. Comparison of E-Test and Broth Microdilution Methods for Antifungal Drug Susceptibility Testing of Molds

    PubMed Central

    Szekely, Adrien; Johnson, Elizabeth M.; Warnock, David W.

    1999-01-01

    We compared the E test with a broth microdilution method, performed according to National Committee for Clinical Laboratory Standards document M27-A guidelines, for determining the in vitro susceptibilities of 90 isolates of pathogenic molds (10 Absidia corymbifera, 10 Aspergillus flavus, 10 Aspergillus fumigatus, 10 Aspergillus niger, 10 Aspergillus terreus, 10 Exophiala dermatitidis, 10 Fusarium solani, 10 Scedosporium apiospermum, 5 Scedosporium prolificans, and 5 Scopulariopsis brevicaulis). Overall, there was 71% agreement between the results of the two methods for amphotericin B (E-test MICs within ±2 log2 dilutions of broth microdilution MICs) and 88% agreement with the results for itraconazole. The overall levels of agreement (within ±2 log2 dilutions) were ≥80% for 5 of the 10 species tested against amphotericin B and 8 of the 10 species tested against itraconazole. The best agreement between the results was seen with A. fumigatus and A. terreus (100% of results for both agents within ±2 log2 dilutions). The poorest agreement was seen with S. apiospermum, S. prolificans, and S. brevicaulis tested against amphotericin B (20% of results within ±2 log2 dilutions). In every instance, this low level of agreement was due to isolates for which the broth microdilution MICs were low but for which the E-test MICs were much higher. The E test appears to be a suitable alternative procedure for testing the susceptibility of Aspergillus spp. and some other molds to amphotericin B or itraconazole. PMID:10203509

  19. Quality control ranges for testing broth microdilution susceptibility of Flavobacterium columnare and F. psychrophilium to nine antimicrobials

    USDA-ARS?s Scientific Manuscript database

    A multi-laboratory broth microdilution method trial was performed to standardize the specialized test conditions required for fish pathogens Flavobacterium columnare and F. pyschrophilum. Nine laboratories tested the quality control (QC) strains Escherichia coli ATCC 25922 and Aeromonas salmonicid...

  20. Comparative evaluation of the Vitek 2 yeast susceptibility test and CLSI broth microdilution reference method for testing antifungal susceptibility of invasive fungal isolates in Italy: the GISIA3 study.

    PubMed

    Borghi, Elisa; Iatta, Roberta; Sciota, Rita; Biassoni, Caterina; Cuna, Teresa; Montagna, Maria Teresa; Morace, Giulia

    2010-09-01

    The newly available AST-YS01 Vitek 2 cards were evaluated, and the results were compared with those obtained by the CLSI M27-A2 microdilution reference method. Clinical fungal isolates, including 614 isolates of Candida spp., 10 Cryptococcus neoformans isolates, 1 Geotrichum capitatum isolate, and 2 quality control strains, were tested for their susceptibilities to amphotericin B, fluconazole, and voriconazole using both methods. The majority of fungal isolates were susceptible to all antifungal agents tested: the MIC(90) values determined by the Vitek 2 and CLSI methods were 0.5 and 1 microg/ml, respectively, for amphotericin B; 8 and 16 microg/ml, respectively, for fluconazole; and <0.12 and 0.25 microg/ml, respectively, for voriconazole. Overall there was excellent categorical agreement (CA) between the methods (99.5% for amphotericin B, 92% for fluconazole, 98.2% for voriconazole), but discrepancies were observed within species. The CAs for fluconazole were low for Candida glabrata and Candida krusei when the results of the CLSI method at 48 h were considered. Moreover, the fully automated commercial system did not detect the susceptibility of Cryptococcus neoformans to voriconazole. The Vitek 2 system can be considered a valid support for antifungal susceptibility testing of fungi, but testing of susceptibility to agents not included in the system (e.g., echinocandins and posaconazole) should be performed with other methods.

  1. In vitro susceptibility testing of Geotrichum capitatum: comparison of the E-test, disk diffusion, and Sensititre colorimetric methods with the NCCLS M27-A2 broth microdilution reference method.

    PubMed

    Girmenia, C; Pizzarelli, G; D'Antonio, D; Cristini, F; Martino, P

    2003-12-01

    The in vitro activities of amphotericin B, flucytosine, fluconazole, itraconazole, and voriconazole against 23 isolates of Geotrichum capitatum were determined by the National Committee for Clinical Laboratory Standards (NCCLS) M27-A2 microdilution method and the Sensititre and agar diffusion methods. Amphotericin B and voriconazole appeared to be the more active drugs. Sensititre showed the highest rates of agreement with the NCCLS M27-A2 method.

  2. Comparison of E-Test, disk diffusion and a modified CLSI broth microdilution (M 38-A) method for in vitro testing of itraconazole, fluconazole and voriconazole against dermatophytes.

    PubMed

    Méndez, Carmen Castro; Serrano, Maria Carmen; Valverde, Anastasio; Pemán, Javier; Almeida, Carmen; Martín-Mazuelos, Estrella

    2008-03-01

    We compared two agar-based methods, the E-test and the disk diffusion method with the Clinical Laboratory Standards Institute (CLSI) reference broth microdilution method (CLSI M38-A; MD). Forty six dermatophytes strains including 30 Trichophyton mentagrophytes, 8 T. rubrum and 8 M. gypseum were tested against three antifungal agents, i.e., fluconazole (FLC), itraconazole (ITC) and voriconazole (VRC). The level of agreement between the E-test and MD (+/-2 dilutions) was 45.6% for fluconazole, 19.5% for itraconazole and 52.1% for voriconazole. The results obtained with disk diffusion had low correlation with the results obtained by the CLSI broth microdilution reference method with azoles.

  3. Towards a Standardized Method for Broth Microdilution Susceptibility Testing of Haemophilus parasuis.

    PubMed

    Prüller, Sandra; Turni, Conny; Blackall, Patrick J; Beyerbach, Martin; Klein, Günter; Kreienbrock, Lothar; Strutzberg-Minder, Katrin; Kaspar, Heike; Meemken, Diana; Kehrenberg, Corinna

    2017-01-01

    Currently, there is no agreed method available for broth microdilution susceptibility testing of Haemophilus parasuis, one of the most important bacterial pathogens in pig production. Therefore, the aim of this study was to develop a method that could be easily performed by diagnostic laboratories and that appears suitable for a harmonized susceptibility testing. Growth determinations using one type strain and three field isolates revealed no visible growth of H. parasuis in media which have proven to be suitable for susceptibility testing of fastidious organisms. Therefore, a new medium, cation-adjusted Mueller-Hinton broth (CAMHB) plus NADH and sterile filtered heat-inactivated chicken serum, was developed. The reproducibility of MICs obtained in this medium was evaluated and statistically analyzed, considering a model with two different variables (precondition of five identical MICs and MIC mode accepting a deviation of ±1 dilution step, respectively). No significant differences for both variables were seen between two time points investigated and between results obtained with the recently proposed test medium broth (TMB). Nearly all MICs of quality control strains were in the acceptable range. Subsequently, 47 H. parasuis isolates representing 13 serovars were tested with the newly developed medium and TMB. Statistical analysis of all isolates and 15 antimicrobial agents and antimicrobial combinations showed no significant difference between MICs obtained in supplemented CAMHB and TMB. Because of a simplified implementation in routine diagnostic and a lower chance of interference between medium components and antimicrobial agents, supplemented CAMHB is recommended with an incubation time of 24 h.

  4. Early detection of oxacillin-resistant staphylococcal strains with hypertonic broth diluent for microdilution panels.

    PubMed Central

    Dillon, L K; Howe, S E

    1984-01-01

    A total of 292 coagulase-positive and 111 coagulase-negative staphylococcal strains were tested in microdilution MIC panels containing 16 to 0.13 microgram of oxacillin per ml diluted in cation-supplemented Mueller-Hinton broth with and without an additional 2% NaCl. All strains were tested using the stationary-phase inoculum procedure with an incubation temperature of 35 degrees C. Test results were recorded after 16 to 20 h of incubation; staphylococcal strains susceptible to oxacillin (less than or equal to 2 micrograms/ml) were reincubated for 20 to 24 h, and endpoints were determined again. Oxacillin resistance was found in 27 (9%) of the 292 coagulase-positive strains and 39 (35%) of the 111 coagulase-negative strains. Of these resistant strains, 5 (19%) of the 27 coagulase-positive strains and 13 (33%) of the 39 coagulase-negative strains were detected 24 h earlier in cation-supplemented Mueller-Hinton broth with 2% NaCl than in cation-supplemented Mueller-Hinton broth without the additional NaCl. However, 9 (33%) of the 27 resistant coagulase-positive strains and 10 (26%) of the 39 resistant coagulase-negative strains were detected only after an additional 24 h of incubation. Oxacillin MICs for the 265 coagulase-positive susceptible strains and 72 coagulase-negative susceptible strains were not affected by the additional 2% NaCl. These results support the utility of adding 2% NaCl to the broth diluent for the early detection of oxacillin-resistant staphylococcal strains and the necessity of extended incubation for those strains which initially appear to be susceptible to oxacillin after only 16 to 20 h of incubation. PMID:6562124

  5. Evaluation of agar dilution and broth microdilution methods to determine the disinfectant susceptibility.

    PubMed

    Wu, Guoyan; Yang, Qianru; Long, Mei; Guo, Lijuan; Li, Bei; Meng, Yue; Zhang, Anyun; Wang, Hongning; Liu, Shuliang; Zou, Likou

    2015-11-01

    A variety of disinfectants have been widely used in veterinary hygiene, food industries and environments, which could induce the development of bacterial resistance to disinfectants. The methods used to investigate antimicrobial effects of disinfectant vary considerably among studies, making comparisons difficult. In this study, agar dilution and broth microdilution methods were used to compare the antimicrobial activities of four quaternary ammonium compounds (QACs) against foodborne and zoonotic pathogens. The potential relationship between the presence of QACs resistance genes and phenotypic resistance to QACs was also investigated. Our results indicated that the minimum inhibitory concentrations (MICs) determined by two methods might be different depended upon different QACs and bacteria applied. Regardless of the testing methods, Klebsiella pneumoniae was more tolerant among Gram-negative strains to four QACs, followed by Salmonella and Escherichia coli. The agreement between MICs obtained by the two methods was good, for benzalkonium chloride (78.15%), didecyldimethylammonium chloride (DDAC) (82.35%), cetylpyridinium chloride (CTPC) (97.48%) and cetyltrimethylammonium bromide (CTAB) (99.16%), respectively. Among all Gram-negative bacteria, 94.55% (n=52) of qacEΔ1-positive strains showed higher MICs (512 mg l(-1)) to CTAB. The qacEΔ1 gene was highly associated (P<0.05) with the high MICs of QACs (⩾512 mg l(-1)). In addition, DDAC remained as the most effective disinfectant against both Gram-positive and Gram-negative bacteria. This is the first study that compared the agar dilution and broth microdilution methods to assess the antimicrobial activity of QACs. The study demonstrated the need to standardize method that would be used in evaluating QACs antimicrobial properties in the future.

  6. Daptomycin Bactericidal Activity and Correlation between Disk and Broth Microdilution Method Results in Testing of Staphylococcus aureus Strains with Decreased Susceptibility to Vancomycin

    PubMed Central

    Sader, Helio S.; Fritsche, Thomas R.; Jones, Ronald N.

    2006-01-01

    A total of 207 Staphylococcus aureus strains, including 105 well-characterized strains with decreased susceptibility to vancomycin (17 vancomycin-intermediate S. aureus [VISA] and 88 heteroresistant VISA [hVISA] strains) and 102 wild-type methicillin-resistant S. aureus (MRSA-WT) strains were tested by reference/standardized broth microdilution and disk diffusion methods, as well as by Etest (AB BIODISK, Solna, Sweden), against daptomycin and vancomycin. The lowest concentration of antimicrobial agent that killed ≥99.9% of the initial inoculum was defined as the minimum bactericidal concentration (MBC) endpoint, and time-kill curves were performed in selected strains to further evaluate bactericidal activity. All MRSA-WT and hVISA strains were inhibited by ≤1 μg/ml of daptomycin, while the VISA strains showed slightly higher daptomycin MICs (range, 0.5 to 4 μg/ml). All daptomycin MBC results were at the MIC or twofold higher. In contrast, 14.7% of MRSA-WT, 69.3% of hVISA, and all VISA strains showed a vancomycin MBC/MIC ratio of ≥32 or an MBC of ≥16 μg/ml (tolerant). The correlation coefficients between broth microdilution and disk diffusion method results were low for daptomycin (0.07) and vancomycin (0.11). Eight (3.8%) strains (all hVISA or VISA) were “nonsusceptible” to daptomycin by broth microdilution methods but susceptible by the disk diffusion method. For vancomycin, 35 (16.9%) strains were nonsusceptible by broth microdilution methods but susceptible by disk diffusion methods. In conclusion, daptomycin was highly bactericidal against S. aureus strains, and its bactericidal activity was not affected by decreased susceptibility to vancomycin. In contrast, many (one in seven) contemporary MRSA-WT, the majority of hVISA, and all VISA strains showed vancomycin MBC/MIC ratios consistent with tolerance, a predictor of poor clinical response. Disk diffusion tests generally failed to detect strains categorized as nonsusceptible to daptomycin or

  7. Posaconazole susceptibility testing against Candida species: comparison of broth microdilution and E-test methods.

    PubMed

    Sóczó, G; Kardos, G; McNicholas, P M; Falusi, E; Gergely, L; Majoros, L

    2007-05-01

    Posaconazole (POS) is a newer triazole with activity against yeasts and moulds. POS and fluconazole were tested in vitro against 32 Candida albicans, 30 C. glabrata, 21 C. tropicalis, 29 C. krusei, 28 C. parapsilosis, 50 C. inconspicua, 13 C. kefyr and 5 C. famata isolates using CLSI broth microdilution method (BMD). We compared E-test and a modified BMD using polyethylene-glycol (PEG) as solvent to the CLSI method. BMDs and E-test were performed according to CLSI and the manufacturer's instructions respectively. Geometric means of POS MICs using BMD were 0.71, 0.22 and 0.21 microg ml(-1) against C. glabrata, C. krusei and C. inconspicua, respectively, and remained below 0.1 microg ml(-1) against all other species tested. One of two C. albicans and two of three C. glabrata isolates resistant to fluconazole showed MICs above 8 microg ml(-1) to POS. The impact of using PEG instead of DMSO had only a minor effect (agreements above 95% with the exception of C. parapsilosis). E-tests read after 24 h showed good agreement with the BMD. POS exhibited excellent in vitro activity against Hungarian Candida strains. E-test showed good correlation with the CLSI method, but to facilitate the comparability of results we believe that DMSO should be used as solvent in the BMD.

  8. Etest® versus broth microdilution for ceftaroline MIC determination with Staphylococcus aureus: results from PREMIUM, a European multicentre study.

    PubMed

    Cantón, Rafael; Livermore, David M; Morosini, María Isabel; Díaz-Regañón, Jazmín; Rossolini, Gian Maria

    2017-02-01

    To compare the concordance of ceftaroline MIC values by reference broth microdilution (BMD) and Etest (bioMérieux, France) for MSSA and MRSA isolates obtained from PREMIUM (D372SL00001), a European multicentre study. Ceftaroline MICs were determined by reference BMD and by Etest for 1242 MSSA and MRSA isolates collected between February and May 2012 from adult patients with community-acquired pneumonia or complicated skin and soft tissue infections; tests were performed across six European laboratories. Selected isolates with ceftaroline resistance in broth (MIC >1 mg/L) were retested in three central laboratories to confirm their behaviour. Overall concordance between BMD and Etest was good, with >97% essential agreement and >95% categorical agreement. Nevertheless, 12 of the 26 MRSA isolates found resistant by BMD scored as susceptible by Etest, with MICs ≤1 mg/L, thus counting as very major errors, whereas only 5 of 380 MRSA isolates found ceftaroline susceptible in BMD were miscategorized as resistant by Etest. Twenty-one of the 26 isolates with MICs of 2 mg/L by BMD were then retested twice by each of three central laboratories: BMD MICs of 2 mg/L were consistently found for 19 of the 21 isolates. Among 147 Etest results for these 21 isolates (original plus six repeats per isolate) 112 were >1 mg/L. BMD and Etest have good overall agreement for ceftaroline against Staphylococcus aureus; nevertheless, reliable Etest-based discrimination of the minority of ceftaroline-resistant (MIC 2 mg/L) MRSA is extremely challenging, requiring careful reading of strips, ideally with duplicate testing. © The Author 2016. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy.

  9. Comparison of Disk Diffusion, VITEK 2, and Broth Microdilution Antimicrobial Susceptibility Test Results for Unusual Species of Enterobacteriaceae▿

    PubMed Central

    Stone, Nimalie D.; O'Hara, Caroline M.; Williams, Portia P.; McGowan, John E.; Tenover, Fred C.

    2007-01-01

    We compared the antimicrobial susceptibility testing results generated by disk diffusion and the VITEK 2 automated system with the results of the Clinical and Laboratory Standards Institute (CLSI) broth microdilution (BMD) reference method for 61 isolates of unusual species of Enterobacteriaceae. The isolates represented 15 genera and 26 different species, including Buttiauxella, Cedecea, Kluyvera, Leminorella, and Yokenella. Antimicrobial agents included aminoglycosides, carbapenems, cephalosporins, fluoroquinolones, penicillins, and trimethoprim-sulfamethoxazole. CLSI interpretative criteria for Enterobacteriaceae were used. Of the 12 drugs tested by BMD and disk diffusion, 10 showed >95% categorical agreement (CA). CA was lower for ampicillin (80.3%) and cefazolin (77.0%). There were 3 very major errors (all with cefazolin), 1 major error (also with cefazolin), and 26 minor errors. Of the 40 isolates (representing 12 species) that could be identified with the VITEK 2 database, 36 were identified correctly to species level, 1 was identified to genus level only, and 3 were reported as unidentified. VITEK 2 generated MIC results for 42 (68.8%) of 61 isolates, but categorical interpretations (susceptible, intermediate, and resistant) were provided for only 22. For the 17 drugs tested by both BMD and VITEK 2, essential agreement ranged from 80.9 to 100% and CA ranged from 68.2% (ampicillin) to 100%; thirteen drugs exhibited 100% CA. In summary, disk diffusion provides a reliable alternative to BMD for testing of unusual Enterobacteriaceae, some of which cannot be tested, or produce incorrect results, by automated methods. PMID:17135429

  10. Evaluation of disk diffusion and Etest compared to broth microdilution for antifungal susceptibility testing of posaconazole against clinical isolates of filamentous fungi.

    PubMed

    Messer, Shawn A; Diekema, Daniel J; Hollis, Richard J; Boyken, Linda B; Tendolkar, Shailesh; Kroeger, Jennifer; Pfaller, Michael A

    2007-04-01

    We performed Etest, disk diffusion, and broth microdilution susceptibility testing of posaconazole against 146 clinical isolates of filamentous fungi. By using provisional breakpoints for comparison purposes only, categorical agreement between the results of the agar-based methods and those of broth microdilution were 96 to 98%, with no very major errors. These agar-based methods hold promise as simple and reliable methods for determining the posaconazole susceptibilities of filamentous fungi.

  11. In Vitro Activity of Fosfomycin against a Collection of Clinical Pseudomonas aeruginosa Isolates from 16 Spanish Hospitals: Establishing the Validity of Standard Broth Microdilution as Susceptibility Testing Method

    PubMed Central

    Díez-Aguilar, María; del Campo, Rosa; García-Castillo, María; Zamora, Javier; Cantón, Rafael

    2013-01-01

    The broth microdilution method for fosfomycin and Pseudomonas aeruginosa was assessed and compared with the approved agar dilution method in 206 genetically unrelated P. aeruginosa clinical isolates. Essential agreement between the two methods was 84%, and categorical agreement was 89.3%. Additionally, Etest and disk diffusion assays were performed. Results validate broth microdilution as a reliable susceptibility testing method for fosfomycin against P. aeruginosa. Conversely, unacceptable concordance was established between Etest and disk diffusion results with agar dilution results. PMID:23939889

  12. Underestimation of Vancomycin and Teicoplanin MICs by Broth Microdilution Leads to Underdetection of Glycopeptide-Intermediate Isolates of Staphylococcus aureus▿ †

    PubMed Central

    Vaudaux, Pierre; Huggler, Elzbieta; Bernard, Louis; Ferry, Tristan; Renzoni, Adriana; Lew, Daniel P.

    2010-01-01

    Broth microdilution was compared with tube macrodilution and a simplified population analysis agar method for evaluating vancomycin and teicoplanin MICs and detecting glycopeptide-intermediate isolates of Staphylococcus aureus. Modal vancomycin and teicoplanin MICs recorded by tube macrodilution and the agar plate assay, which both used inocula of 106 CFU, were significantly higher (2 μg/ml) against a panel of borderline glycopeptide-susceptible and glycopeptide-intermediate methicillin-resistant S. aureus (MRSA) bloodstream isolates compared to broth microdilution (1 μg/ml). Vancomycin and teicoplanin MIC distributions by tube macrodilution and agar testing were also markedly different from those evaluated by broth microdilution. The 20-fold-lower inoculum size used for broth microdilution compared to macrodilution and agar MIC assays explained in part, but not entirely, the systematic trend toward lower vancomycin and teicoplanin MICs by microdilution compared to other methods. Broth microdilution assay led to underdetection of the vancomycin-intermediate S. aureus (VISA) phenotype, yielding only three VISA isolates, for which vancomycin MICs were 4 μg/ml compared to 8 and 19 VISA isolates detected by macrodilution and agar testing, respectively. While macrodilution and agar testing detected 7 and 22 isolates with elevated teicoplanin MICs (8 μg/ml), respectively, broth microdilution failed to detect such isolates. Detection rates of isolates with elevated vancomycin and teicoplanin MICs by macrodilution and agar testing assays were higher at 48 h than at 24 h. In conclusion, the sensitivity of broth microdilution MIC testing is questionable for reliable detection and epidemiological surveys of glycopeptide-intermediate resistance in S. aureus isolates. PMID:20547791

  13. In vitro susceptibility testing of dermatophytes isolated in Goiania, Brazil, against five antifungal agents by broth microdilution method.

    PubMed

    Araújo, Crystiane Rodrigues; Miranda, Karla Carvalho; Fernandes, Orionalda de Fatima Lisboa; Soares, Ailton José; Silva, Maria do Rosário Rodrigues

    2009-01-01

    The antifungal activities of fluconazole, itraconazole, ketoconazole, terbinafine and griseofulvin were tested by broth microdilution technique, against 60 dermatophytes isolated from nail or skin specimens from Goiania city patients, Brazil. In this study, the microtiter plates were incubated at 28 masculineC allowing a reading of the minimal inhibitory concentration (MIC) after four days of incubation for Trichophyton mentagrophytes and five days for T. rubrum and Microsporum canis. Most of the dermatophytes had uniform patterns of susceptibility to the antifungal agents tested. Low MIC values as 0.03 microg/mL were found for 33.3%, 31.6% and 15% of isolates for itraconazole, ketoconazole and terbinafine, respectively.

  14. Oxacillin susceptibility testing of Staphylococcus saprophyticus using disk diffusion, agar dilution, broth microdilution, and the Vitek GPS-105 card.

    PubMed

    Ramotar, K; Woods, W; Toye, B

    2001-08-01

    Eighty-three mecA negative isolates of S. saprophyticus had oxacillin zone diameters broth microdilution, or the Vitek GPS-105 card. Greater than 90% of these isolates would be considered resistant using NCCLS M7-A5, M100-S10 criteria. These results suggest that the current NCCLS MIC and zone diameter breakpoints for oxacillin resistance in coagulase-negative Staphylococci are not appropriate for S. saprophyticus as they do not correlate with the presence of the mecA gene.

  15. Vancomycin MIC for Methicillin-Resistant Coagulase-Negative Staphylococcus Isolates: Evaluation of the Broth Microdilution and Etest Methods ▿

    PubMed Central

    Paiva, Rodrigo M.; Mombach Pinheiro Machado, Alice B.; Zavascki, Alexandre P.; Barth, Afonso L.

    2010-01-01

    Vancomycin MIC results were determined by the broth microdilution (BMD) method and by Etest using 130 methicillin-resistant coagulase-negative staphylococcus bloodstream isolates obtained from a tertiary hospital. The majority (98.5%) of MIC results determined by BMD were ≤1 μg/ml, in contrast to MIC results determined by Etest (72.3% were ≥1.5 μg/ml). The MICs obtained by Etest were, in general, 1- to 2-fold higher than the MICs obtained by BMD. PMID:20861345

  16. Broth microdilution testing of Pseudomonas aeruginosa and aminoglycosides: need for employing dilutions differing by small arithmetic increments.

    PubMed Central

    Woolfrey, B F; Fox, J M; Lally, R T; Quall, C O

    1982-01-01

    The use of dilutions differing by small arithmetic increments was studied as a means for improving the definition and measurement of minimum inhibitory concentrations and precision parameters for testing Pseudomonas aeruginosa versus the aminoglycosides by the broth microdilution test. For five strains of P. aeruginosa versus gentamicin, tobramycin, and amikacin, comparisons were made of minimum inhibitory concentrations which were replicated in parallel by using three microdilution systems: small increment panels prepared by us, modified twofold dilution panels prepared by us, and similar modified twofold dilution panels obtained commercially. The small increment dilutions were prepared to differ by concentrations of 1.0 microgram/ml for gentamicin and tobramycin and by 2.0 micrograms/ml for amikacin. Use of the small increment dilutions resulted in the ability to measure minimum inhibitory concentrations at more closely spaced intervals than those dictated by modified twofold dilution schemes, and confidence limits were significantly improved. The average coefficient of variation for the small increment microdilution test results was 9.5%, with 99.5% of minimum inhibitory concentrations falling within +/- 2 small increment dilutions from their modal values. PMID:6818244

  17. Evaluating the resistance to posaconazole by E-test and CLSI broth microdilution methodologies of Candida spp. and pathogenic moulds.

    PubMed

    Araujo, R; Costa-de-Oliveira, S; Coutinho, I; Rodrigues, A G; Pina-Vaz, C

    2009-09-01

    E-test methodology was compared with Clinical and Laboratory Standards Institute (CLSI) broth microdilution, particularly concerning the detection of resistance to posaconazole among clinical fungal isolates. The susceptibility of a large set of fungal strains (n = 300) was evaluated following 24 and 48 h in two different culture media (RPMI 1640 and Sabouraud agar). Fungal strains were highly susceptible to posaconazole; however, few less susceptible strains were found, mostly regarding Candida albicans, Candida glabrata, Acremonium sp., Cladosporium sp. and Scedosporium apiospermum. Broth microdilution and E-test methods provided similar results for posaconazole-susceptible strains, while the less susceptible fungal strains (10.3% of the strains showed MIC > or =2 microg/mL) resulted in higher discrepancies between the two methodologies, particularly concerning Candida spp. E-test susceptibility values were critically affected by the pH of the culture media. Sabouraud medium provided similar susceptibility results for moulds to those for RPMI, soon after 24 h. Posaconazole resistance was rare in this study, but routine susceptibility methods, such as the E-test, should be able to detect fungal strains with reduced susceptibility. E-test methodology still needs improvements to recognise accurately strains less susceptible to posaconazole.

  18. A comparative study of M.I.C evaluator test with the broth microdilution method for antimicrobial susceptibility testing of Enterobacter cloacae isolated from cooked food.

    PubMed

    Nyenje, Mirriam Ethel; Tanih, Nicoline Fri; Ndip, Roland Ndip

    2014-01-01

    Agar dilution and broth microdilution are widely recommended quantitative antimicrobial susceptibility test methods, but they are tedious and time consuming to implement as routine tests in many clinical laboratories. Therefore, this study aimed at comparing the broth microdilution and the M.I.C Evaluator method which has been validated for its high accuracy and easy performance for routine diagnostic use. Twenty Enterobacter cloacae strains were isolated following microbiological procedures and confirmation of the isolates used the API 20E test. The strains were evaluated for their susceptibility to seven antimicrobials using the broth microdilution and MIC Evaluator methods. The doubling dilution difference (essential agreement) in the MIC result was derived from: log2 (MIC by BMD) -log2 (MIC by M.I.C Evaluator method). The categorical agreement, interpreted as breakpoints of sensitive and resistance strains was also noted. Categorical agreement between M.I.C Evaluator strip and broth microdilution for amoxicillin, metronidazole and erythromycin was 100%: while categorical agreement for ciprofloxacin was 90%. The essential agreement for erythromycin, ciprofloxacin and tetracycline were 90%, 70% and 15% respectively. Results indicate a high efficiency of the M.I.C Evaluator strip method in determination of minimum inhibitory concentration as compared to broth microdilution method. However, further analysis regarding the suitability of the M.I.C Evaluator for testing Enterobacter cloacae is warranted considering that no consensus guidelines exist for the use of this method with the organism.

  19. 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.

  20. Are E-test and Vitek2 good choices for tigecycline susceptibility testing when comparing broth microdilution for MDR and XDR Acinetobacter baumannii?

    PubMed

    Grandesso, Stefano; Sapino, Barbara; Amici, Gianpaolo; Mazzucato, Sandra; Solinas, Maria; Gion, Massimo

    2014-10-01

    This study reports the results of antimicrobial susceptibility testing of 10 MDR and 74 XDR Acinetobacter bauman- nii clinical isolates from our hospital routine. We used three different methods: two automated systems (Sensititre and VITEK 2) and one standardized manual method (E-test). Since many published papers refer to in vitro tests performed by E-test, the aim of this study was to test if this method is reliable for testing tigecycline. The results obtained show that E-test significantly overestimates the MIC of the broth microdilution (reference test), thus ob- taining a significant number of major errors (resistant instead of sensitive). VITEK 2 also shows the same problem, but it is less critical. We therefore conclude that these methods do not seem to be very reliable in the performance of susceptibility testing of MDR and XDR Acinetobacter baumannii against tigecycline.

  1. In vitro activity of imazalil against Penicillium expansum: comparison of the CLSI M38-A broth microdilution method with traditional techniques.

    PubMed

    Cabañas, R; Abarca, M L; Bragulat, M R; Cabañes, F J

    2009-01-31

    Penicillium expansum is one of the most important pathogens that cause blue mold in stored apples and is regarded as the major producer of the mycotoxin patulin. Imazalil is one of the fungicides used in Spain to control postharvest blue mold, but development of fungal resistance has been reported in P. digitatum and P. italicum. The most common used methods to detect antifungal susceptibility of fungal crop pathogens in vitro, are direct-plating isolates in media amended with various concentrations of fungicide and determining inhibition of growth and/or spore germination. These techniques are time- and labor-intensive and are not suitable if a large number of isolates has to be evaluated. On the other hand, the broth microdilution method M38-A is the reference method developed by the Clinical and Laboratory Standards Institute (CLSI) for antifungal susceptibility testing in some clinical fungi, but Penicillium spp. are not included. Due to the lack of a standard method, the aim of this work is to evaluate the suitability of an adaptation of the CLSI M38-A method to monitor P. expansum susceptibility to imazalil in comparison with other techniques. A total of 128 P. expansum strains have been studied (118 isolates from apples and pears, 5 from grapes and 5 reference strains). Imazalil has shown to be highly active in vitro against all the P. expansum isolates tested, as all the evaluated parameters were in the range reported for imazalil sensitive Penicillium spp. The mean minimum inhibitory concentration determined by broth microdilution method and by agar dilution method (48-72 h readings) was 0.0625 microg/ml and 0.11-0.12 microg/ml respectively. The mean concentration that inhibited the size of colonies (48-72 h) and spore germination by 50% was 0.05-0.06 and 0.04 microg/ml respectively. Our results highlight that the broth microdilution method CLSI M38-A is a good alternative to be used in screening the in vitro activity of imazalil against a large number of

  2. Comparison of disk diffusion, E-test and broth microdilution test in determination of susceptibility of Aspergillus species to amphotericin B, itraconazole and voriconazole.

    PubMed

    Ozkutuk, A; Ergon, C; Metin, D Y; Yucesoy, M; Polat, S H

    2008-02-01

    In response to the recent increase in Aspergillus infections, new antifungal agents have become available accompanied by studies on antifungal susceptibility tests for epidemiological follow-up. The aim of this study was to compare the efficacy of Clinical Laboratory Standards Institute (CLSI) M 38-A broth microdilution test with the disk diffusion and E-test in determining the susceptibility of Aspergillus spp. to amphotericin B, itraconazole and voriconazole. The study was carried out on 18 A. fumigatus, 7 A. flavus, 5 A. niger and 2 A. versicolor strains isolated from clinical samples. The microdilution method was performed by following the instructions of CLSI M 38-A. The E-test and disk diffusion tests were performed according to the instructions of their manufacturers. The percent agreement between the E-test and CLSI M38-A broth microdilution test at 24 (48) h within +/- 2 dilutions was, respectively, 81% (69%) for amphotericin B, 75% (72%) for itraconazole and 85% (81%) for voriconazole. The disk diffusion test showed good correlation with the E-test but poor correlation with the broth microdilution test for the three antifungal agents we tested. In conclusion, E-test and disk diffusion test have their advantages such as ease of application and interpretation, but their correlation with the broth microdilution should be improved.

  3. Comparative Analysis of Disc Diffusion and E-test with Broth Micro-dilution for Susceptibility Testing of Clinical Candida Isolates Against Amphotericin B, Fluconazole, Voriconazole and Caspofungin

    PubMed Central

    Bhattacharyya, Sayan; Gupta, Prashant; Banerjee, Gopa; Singh, Mastan

    2015-01-01

    Background Antifungal susceptibility testing remains an area of intense interest because of the increasing number of clinical isolates resistant to antifungal therapy. Clinical and Laboratory Standards Institute has proposed reference broth micro dilution (BMD) method for susceptibility testing. The reference method is time-consuming and poorly suited for the routine clinical laboratory setting. Agar-based susceptibility testing methods, disk diffusion (DD) method and the E-test method can be an easier, reliable and less time consuming alternative for the BMD method. Aim To compare the results of Amphotericin B, fluconazole, voriconazole, and Caspofungin susceptibility testing by DD, and the E-test method with the CLSI reference method for clinical Candida isolates. Materials and Methods Broth Microdilution (BMD), E-test and Disk diffusion testing of the various clinical Candida isolates was performed in accordance with CLSI documents. The results obtained were analysed and compared. Results The categorical agreement for Amphotericin B, fluconazole, voriconazole, and Caspofungin susceptibility results by E-test and DD method was 65.2%, 67.4%; 100%, 82.6%; 100%, 100%; 100%, 97.8% respectively. Conclusion The agar-based E-test and disk diffusion methods are reliable alternatives to the BMD method for Candida isolates when test susceptible to fluconazole, voriconazole, and Caspofungin, however the susceptibility testing results must be interpreted with caution in case of Amphotericin B. PMID:26675415

  4. Comparative Analysis of Disc Diffusion and E-test with Broth Micro-dilution for Susceptibility Testing of Clinical Candida Isolates Against Amphotericin B, Fluconazole, Voriconazole and Caspofungin.

    PubMed

    Kumar, Deepak; Bhattacharyya, Sayan; Gupta, Prashant; Banerjee, Gopa; Singh, Mastan

    2015-11-01

    Antifungal susceptibility testing remains an area of intense interest because of the increasing number of clinical isolates resistant to antifungal therapy. Clinical and Laboratory Standards Institute has proposed reference broth micro dilution (BMD) method for susceptibility testing. The reference method is time-consuming and poorly suited for the routine clinical laboratory setting. Agar-based susceptibility testing methods, disk diffusion (DD) method and the E-test method can be an easier, reliable and less time consuming alternative for the BMD method. To compare the results of Amphotericin B, fluconazole, voriconazole, and Caspofungin susceptibility testing by DD, and the E-test method with the CLSI reference method for clinical Candida isolates. Broth Microdilution (BMD), E-test and Disk diffusion testing of the various clinical Candida isolates was performed in accordance with CLSI documents. The results obtained were analysed and compared. The categorical agreement for Amphotericin B, fluconazole, voriconazole, and Caspofungin susceptibility results by E-test and DD method was 65.2%, 67.4%; 100%, 82.6%; 100%, 100%; 100%, 97.8% respectively. The agar-based E-test and disk diffusion methods are reliable alternatives to the BMD method for Candida isolates when test susceptible to fluconazole, voriconazole, and Caspofungin, however the susceptibility testing results must be interpreted with caution in case of Amphotericin B.

  5. Development of Similar Broth Microdilution Methods to Determine the Antimicrobial Susceptibility of Flavobacterium columnare and F. psychrophilum.

    PubMed

    Gieseker, Charles M; Crosby, Tina C; Mayer, Tamara D; Bodeis, Sonya M; Stine, Cynthia B

    2016-03-01

    Flavobacterium columnare and F. psychrophilum are major fish pathogens that cause diseases that may require antimicrobial therapy. Choice of appropriate treatment is dependent upon determining the antimicrobial susceptibility of isolates. Therefore we optimized methods for broth microdilution testing of F. columnare and F. psychrophilum to facilitate standardizing an antimicrobial susceptibility test. We developed adaptations to make reproducible broth inoculums and confirmed the proper incubation time and media composition. We tested the stability of potential quality-control bacteria and compared test results between different operators. Log phase occurred at 48 h for F. columnare and 72-96 h for F. psychrophilum, confirming the test should be incubated at 28°C for approximately 48 h and at 18°C for approximately 96 h, respectively. The most consistent susceptibility results were achieved with plain, 4-g/L, dilute Mueller-Hinton broth supplemented with dilute calcium and magnesium. Supplementing the broth with horse serum did not improve growth. The quality-control strains, Escherichia coli ATCC 25922 and Aeromonas salmonicida subsp. salmonicida ATCC 33658, yielded stable minimal inhibitory concentrations (MIC) against all seven antimicrobials tested after 30 passes at 28°C and 15 passes at 18°C. In comparison tests, most MICs of the isolates agreed 100% within one drug dilution for ampicillin, florfenicol, and oxytetracycline. The agreement was lower with the ormetoprim-sulfdimethoxine combination, but there was at least 75% agreement for all but one isolate. These experiments have provided methods to help standardize antimicrobial susceptibility testing of these nutritionally fastidious aquatic bacteria. Received June 24, 2015; accepted October 2, 2015.

  6. Evaluation of antibacterial potential and toxicity of plant volatile compounds using new broth microdilution volatilization method and modified MTT assay.

    PubMed

    Houdkova, Marketa; Rondevaldova, Johana; Doskocil, Ivo; Kokoska, Ladislav

    2017-04-01

    With aim to develop effective proof-of-concept approach which can be used in a development of new preparations for the inhalation therapy, we designed a new screening method for simple and rapid simultaneous determination of antibacterial potential of plant volatiles in the liquid and the vapour phase at different concentrations. In addition, EVA (ethylene vinyl acetate) capmat™ as vapour barrier cover was used as reliable modification of thiazolyl blue tetrazolium bromide (MTT) assay for cytotoxicity testing of volatiles on microtiter plates. Antibacterial activity of carvacrol, cinnamaldehyde, eugenol, 8-hydroxyquinoline, thymol and thymoquinone was determined against Haemophilus influenzae, Staphylococcus aureus, and Streptococcus pneumoniae using new broth microdilution volatilization method. The cytotoxicity of these compounds was evaluated using MTT test in lung fibroblast cells MRC-5. The most effective antibacterial agents were 8-hydroxyquinoline and thymoquinone with the lowest minimum inhibitory concentrations (MICs) ranging from 2 to 128μg/mL, but they also possessed the highest toxicity in lung cell lines with half maximal inhibitory concentration (IC50) values 0.86-2.95μg/mL. The lowest cytotoxicity effect was identified for eugenol with IC50 295.71μg/mL, however this compound produced only weak antibacterial potency with MICs 512-1024μg/mL. The results demonstrate validity of our novel broth microdilution volatilization method, which allows cost and labour effective high-throughput antimicrobial screening of volatile agents without need of special apparatus. In our opinion, this assay can also potentially be used for development of various medicinal, agricultural, and food applications that are based on volatile antimicrobials. Copyright © 2017 Elsevier B.V. All rights reserved.

  7. Thymoquinone vapor significantly affects the results of Staphylococcus aureus sensitivity tests using the standard broth microdilution method.

    PubMed

    Novy, Pavel; Kloucek, Pavel; Rondevaldova, Johana; Havlik, Jaroslav; Kourimska, Lenka; Kokoska, Ladislav

    2014-04-01

    The broth microdilution (BMD) method is widely used for the determination of minimum inhibitory concentrations of antimicrobial agents, including volatile oils and their components. In this series of various experiments, we have demonstrated the influence of thymoquinone (TQ) vapor on the results of the BMD test performed with Staphylococcus aureus as a model organism. The spread of vapor from the TQ containing wells (32-512 μg/mL) caused the complete inhibition of staphylococcal growth in adjoining wells initially containing bacterium-inoculated pure Mueller-Hinton broth only and thus produced false positive results of the test. The ability of TQ to pass into the adjoined wells was subsequently confirmed by gas chromatography-mass spectrometry, whereas TQ at concentrations up to 84 μg/mL was detected in these wells after five hours. Based on these results, we suppose that vapors of TQ as well as of other naturally occurring volatile compounds and their mixtures (for example essential oils and plant extracts) can significantly influence results of the standard BMD assay. These observations, therefore, call for development of new appropriate BMD method suitable for assessment of antimicrobial activity of volatile substances. Copyright © 2014 Elsevier B.V. All rights reserved.

  8. In vitro susceptibility of Burkholderia cepacia complex isolates: Comparison of disk diffusion, Etest®, agar dilution, and broth microdilution methods.

    PubMed

    Fehlberg, Lorena Cristina Corrêa; Nicoletti, Adriana Gianinni; Ramos, Ana Carolina; Rodrigues-Costa, Fernanda; de Matos, Adriana Pereira; Girardello, Raquel; Marques, Elizabeth Andrade; Gales, Ana Cristina

    2016-12-01

    Broth microdilution, agar dilution, Etest® and disk diffusion techniques were compared to evaluate the susceptibility profile of 82 Bcc clinical isolates against six antimicrobials as recommended by CLSI. Broth microdilution was considered the "gold standard" method. The regression analysis was applied to determine the essential (EA) and categorical (CA) agreement rates. STX (MIC50, 1 mg/L) was the most potent antimicrobial tested against Bcc isolates. The worst in vitro activity was observed for chloramphenicol (MIC50, 16 mg/L) and ticarcillin-clavulanic acid (MIC50, >256 mg/L). The EA among broth microdilution and agar dilution results was good for the majority of antimicrobial tested. When comparing broth microdilution and Etest®, ceftazidime, SXT and chloramphenicol exhibited EA rates below 90%. SXT showed an excellent CA (100%) when dilution methodologies were compared. However, a low CA rate was found for this agent between dilution and disk diffusion methodologies resulting in unacceptable very major and minor error rates.

  9. E-Test Method for Testing Susceptibilities of Aspergillus spp. to the New Triazoles Voriconazole and Posaconazole and to Established Antifungal Agents: Comparison with NCCLS Broth Microdilution Method

    PubMed Central

    Espinel-Ingroff, Ana; Rezusta, A.

    2002-01-01

    NCCLS document M38-P describes standard parameters for testing the fungistatic activities (MICs) of established agents against filamentous fungi (molds). This study evaluated the in vitro susceptibilities of 15 Aspergillus flavus isolates, 62 A. fumigatus isolates, and 10 isolates each of A. niger, A. nidulans, and A. terreus to voriconazole, posaconazole, itraconazole, and amphotericin B by the E-test and NCCLS M38-P microdilution methods. The agreement (within 3 dilutions) between methods for voriconazole was independent of the E-test incubation time (93.3 to 100% for four of five species at both incubation times). In contrast, with amphotericin B, itraconazole, and posaconazole, E-test results were more dependent on the incubation time for certain species. For A. fumigatus, posaconazole E-test MICs had better concordance with reference values after 48 h (95.2%) than after 24 h (90%), while the highest agreement for itraconazole MICs was after 24 h (90.3 versus 74.2%) of incubation. Better agreement between the methods was also obtained with 24-h E-test amphotericin B MICs for A. flavus (73.3 versus 26.7%) and A. fumigatus (96.7 versus 64.5%). E-test MICs of the four agents had the lowest percentages of agreement with reference values for A. nidulans (60 to 80%). For isolates for which high MICs were obtained for the four agents by the reference method, high MICs were also obtained by E-test at both 24 and 48 h. The utility of in vitro results of either the E-test or the NCCLS broth microdilution (M38-P) method for Aspergillus spp. needs to be established in clinical trials. PMID:12037072

  10. Standardized broth microdilution antimicrobial susceptibility testing of Francisella tularensis subsp. holarctica strains from Europe and rare Francisella species.

    PubMed

    Georgi, Enrico; Schacht, Erik; Scholz, Holger C; Splettstoesser, Wolf D

    2012-10-01

    Tularaemia is a widespread zoonosis in Europe caused by Francisella tularensis subsp. holarctica. Because of a lack of standardized CLSI-approved antibiotic susceptibility data from European Francisella strains, the antibiotic susceptibilities of a selection of F. tularensis subsp. holarctica isolates originating from Germany, Austria, France, Spain and other European countries were determined. Rarely isolated species and subspecies of Francisella such as Francisella philomiragia, F. tularensis subsp. novicida and F. tularensis subsp. mediasiatica as well as the type strain of Francisella hispaniensis were included in this study. MIC data were obtained using cation-adjusted Mueller-Hinton broth with a 2% growth supplement. The broth microdilution testing system comprised 14 antibiotics, including gentamicin, streptomycin, ciprofloxacin and tetracycline. All of the 91 strains tested were susceptible to aminoglycosides, quinolones, tetracycline and chloramphenicol. The antimicrobial susceptibility of rare Francisellae was similar to the antibiotic profile of F. tularensis subsp. holarctica strains. For erythromycin, we detected two geographically distinct groups of F. tularensis subsp. holarctica isolates in western Europe. One group was resistant and the other one was susceptible. Both groups overlapped in a small region in Germany. Being performed in accordance with CLSI criteria, this study provides reliable data on antibiotic susceptibility patterns of European Francisella isolates. The standardized methodology of this study can be used for testing of suspicious colonies from clinical specimens for therapeutic guidance. Based on the results, aminoglycosides or quinolones are recommended as first-choice antibiotics for the therapy of F. hispaniensis, F. philomiragia or F. tularensis subsp. novicida infections in immunocompromised patients.

  11. Comparison of agar dilution and antibiotic gradient strip test with broth microdilution for susceptibility testing of swine Brachyspira species.

    PubMed

    Mirajkar, Nandita S; Gebhart, Connie J

    2016-03-01

    Production-limiting diseases in swine caused by Brachyspira are characterized by mucohemorrhagic diarrhea (B. hyodysenteriae and "B. hampsonii") or mild colitis (B. pilosicoli), while B. murdochii is often isolated from healthy pigs. Emergence of novel pathogenic Brachyspira species and strains with reduced susceptibility to commonly used antimicrobials has reinforced the need for standardized susceptibility testing. Two methods are currently used for Brachyspira susceptibility testing: agar dilution (AD) and broth microdilution (BMD). However, these tests have primarily been used for B. hyodysenteriae and rarely for B. pilosicoli. Information on the use of commercial susceptibility testing products such as antibiotic gradient strips is lacking. Our main objective was to validate and compare the susceptibility results, measured as the minimum inhibitory concentration (MIC), of 6 antimicrobials for 4 Brachyspira species (B. hyodysenteriae, "B. hampsonii", B. pilosicoli, and B. murdochii) by BMD and AD (tiamulin, valnemulin, lincomycin, tylosin, and carbadox) or antibiotic gradient strip (doxycycline) methods. In general, the results of a high percentage of all 4 Brachyspira species differed by ±1 log2 dilution or less by BMD and AD for tiamulin, valnemulin, lincomycin, and tylosin, and by BMD and antibiotic gradient strip for doxycycline. The carbadox MICs obtained by BMD were 1-5 doubling dilutions different than those obtained by AD. BMD for Brachyspira was quicker to perform with less ambiguous interpretation of results when compared with AD and antibiotic gradient strip methods, and the results confirm the utility of BMD in routine diagnostics.

  12. Comparative Evaluation of PASCO and National Committee for Clinical Laboratory Standards M27-A Broth Microdilution Methods for Antifungal Drug Susceptibility Testing of Yeasts

    PubMed Central

    Arthington-Skaggs, Beth A.; Motley, Milwood; Warnock, David W.; Morrison, Christine J.

    2000-01-01

    The PASCO antifungal susceptibility test system, developed in collaboration with a commercial company, is a broth microdilution assay which is faster and easier to use than the reference broth microdilution test performed according to the National Committee for Clinical Laboratory Standards (NCCLS) document M27-A guidelines. Advantages of the PASCO system include the system's inclusion of quality-controlled, premade antifungal panels containing 10, twofold serial dilutions of drugs and a one-step inoculation system whereby all wells are simultaneously inoculated in a single step. For the prototype panel, we chose eight antifungal agents for in vitro testing (amphotericin B, flucytosine, fluconazole, ketoconazole, itraconazole, clotrimazole, miconazole, and terconazole) and compared the results with those of the NCCLS method for testing 74 yeast isolates (14 Candida albicans, 10 Candida glabrata, 10 Candida tropicalis, 10 Candida krusei, 10 Candida dubliniensis, 10 Candida parapsilosis, and 10 Cryptococcus neoformans isolates). The overall agreements between the methods were 91% for fluconazole, 89% for amphotericin B and ketoconazole, 85% for itraconazole, 80% for flucytosine, 77% for terconazole, 66% for miconazole, and 53% for clotrimazole. In contrast to the M27-A reference method, the PASCO method classified as resistant seven itraconazole-susceptible isolates (9%), two fluconazole-susceptible isolates (3%), and three flucytosine-susceptible isolates (4%), representing 12 major errors. In addition, it classified two fluconazole-resistant isolates (3%) and one flucytosine-resistant isolate (1%) as susceptible, representing three very major errors. Overall, the agreement between the methods was greater than or equal to 80% for four of the seven species tested (C. dubliniensis, C. glabrata, C. krusei, and C. neoformans). The lowest agreement between methods was observed for miconazole and clotrimazole and for C. krusei isolates tested against terconazole. When the

  13. Comparative evaluation of PASCO and national committee for clinical laboratory standards M27-A broth microdilution methods for antifungal drug susceptibility testing of yeasts.

    PubMed

    Arthington-Skaggs, B A; Motley, M; Warnock, D W; Morrison, C J

    2000-06-01

    The PASCO antifungal susceptibility test system, developed in collaboration with a commercial company, is a broth microdilution assay which is faster and easier to use than the reference broth microdilution test performed according to the National Committee for Clinical Laboratory Standards (NCCLS) document M27-A guidelines. Advantages of the PASCO system include the system's inclusion of quality-controlled, premade antifungal panels containing 10, twofold serial dilutions of drugs and a one-step inoculation system whereby all wells are simultaneously inoculated in a single step. For the prototype panel, we chose eight antifungal agents for in vitro testing (amphotericin B, flucytosine, fluconazole, ketoconazole, itraconazole, clotrimazole, miconazole, and terconazole) and compared the results with those of the NCCLS method for testing 74 yeast isolates (14 Candida albicans, 10 Candida glabrata, 10 Candida tropicalis, 10 Candida krusei, 10 Candida dubliniensis, 10 Candida parapsilosis, and 10 Cryptococcus neoformans isolates). The overall agreements between the methods were 91% for fluconazole, 89% for amphotericin B and ketoconazole, 85% for itraconazole, 80% for flucytosine, 77% for terconazole, 66% for miconazole, and 53% for clotrimazole. In contrast to the M27-A reference method, the PASCO method classified as resistant seven itraconazole-susceptible isolates (9%), two fluconazole-susceptible isolates (3%), and three flucytosine-susceptible isolates (4%), representing 12 major errors. In addition, it classified two fluconazole-resistant isolates (3%) and one flucytosine-resistant isolate (1%) as susceptible, representing three very major errors. Overall, the agreement between the methods was greater than or equal to 80% for four of the seven species tested (C. dubliniensis, C. glabrata, C. krusei, and C. neoformans). The lowest agreement between methods was observed for miconazole and clotrimazole and for C. krusei isolates tested against terconazole. When the

  14. Comparative Evaluation of Disc Diffusion and E-test with Broth Micro-dilution in Susceptibility testing of Amphotericin B, Voriconazole and Caspofungin against Clinical Aspergillus isolates

    PubMed Central

    Khare, Vineeta; Kumar, Deepak; Ahmad, Abrar; Banerjee, Gopa; Singh, Mastan

    2015-01-01

    Background: Clinical importance of Aspergillus has increased over the past few decades because of rise in immunosuppressive drugs and immune-modulating diseases. Antifungal susceptibility of Aspergillus is rarely performed by clinical laboratories because of lack of easier method. This study has investigated and compared susceptibility pattern of Aspergillus isolates by disc diffusion, E-test and broth micro-dilution for amphotericin B, voriconazole and caspofungin. Materials and Methods: Disk diffusion (DD) method of antifungal susceptibility (AFS) was evaluated for three different classes of antifungals: amphotericin B (AMB), voriconazole (VCZ) and caspofungin (CAS). Forty four clinical isolates of Aspergillus were selected; these included 34 A.fumigatus, 8 A.flavus and 2 A. terreus. AFS by DD and E-test was done on non-supplemented Mueller Hinton Agar (MHA) and was compared to Clinical Laboratory Standard Institute(CLSI) broth micro-dilution (BMD) method of AFS. Results: Disk diffusion method for amphotericin B showed 87.5% agreement while E-test showed 93.8% agreement with broth micro-dilution. The agreement with broth micro-dilution was similar for both disk diffusion and E-test in case of voriconazole (93.8%) and caspofungin (100%). 31.8% and 9.1% Aspergillus isolates were found to have amphotericin B and voriconazole MIC values above epidemiological cut off value (ECV) respectively. All isolates were within ECV for caspofungin. Conclusion: CLSI method of DD promises to be easier, reproducible and cost effective method of susceptibility testing, but this method must be interpreted with caution in case of amphotericin B susceptibility testing. E-test correlates better than DD with BMD. PMID:25737984

  15. Comparative Evaluation of Disc Diffusion and E-test with Broth Micro-dilution in Susceptibility testing of Amphotericin B, Voriconazole and Caspofungin against Clinical Aspergillus isolates.

    PubMed

    Gupta, Prashant; Khare, Vineeta; Kumar, Deepak; Ahmad, Abrar; Banerjee, Gopa; Singh, Mastan

    2015-01-01

    Clinical importance of Aspergillus has increased over the past few decades because of rise in immunosuppressive drugs and immune-modulating diseases. Antifungal susceptibility of Aspergillus is rarely performed by clinical laboratories because of lack of easier method. This study has investigated and compared susceptibility pattern of Aspergillus isolates by disc diffusion, E-test and broth micro-dilution for amphotericin B, voriconazole and caspofungin. Disk diffusion (DD) method of antifungal susceptibility (AFS) was evaluated for three different classes of antifungals: amphotericin B (AMB), voriconazole (VCZ) and caspofungin (CAS). Forty four clinical isolates of Aspergillus were selected; these included 34 A.fumigatus, 8 A.flavus and 2 A. terreus. AFS by DD and E-test was done on non-supplemented Mueller Hinton Agar (MHA) and was compared to Clinical Laboratory Standard Institute(CLSI) broth micro-dilution (BMD) method of AFS. Disk diffusion method for amphotericin B showed 87.5% agreement while E-test showed 93.8% agreement with broth micro-dilution. The agreement with broth micro-dilution was similar for both disk diffusion and E-test in case of voriconazole (93.8%) and caspofungin (100%). 31.8% and 9.1% Aspergillus isolates were found to have amphotericin B and voriconazole MIC values above epidemiological cut off value (ECV) respectively. All isolates were within ECV for caspofungin. CLSI method of DD promises to be easier, reproducible and cost effective method of susceptibility testing, but this method must be interpreted with caution in case of amphotericin B susceptibility testing. E-test correlates better than DD with BMD.

  16. In Vitro Amphotericin B Susceptibility of Malassezia pachydermatis Determined by the CLSI Broth Microdilution Method and Etest Using Lipid-Enriched Media

    PubMed Central

    Álvarez-Pérez, Sergio; Peláez, Teresa; Cutuli, Maite; García, Marta E.

    2014-01-01

    We determined the in vitro amphotericin B susceptibility of 60 Malassezia pachydermatis isolates by the CLSI broth microdilution method and the Etest using lipid-enriched media. All isolates were susceptible at MICs of ≤1 μg/ml, confirming the high activity of amphotericin B against this yeast species. Overall, the essential agreement between the tested methods was high (80% and 96.7% after 48 h and 72 h, respectively), and all discrepancies were regarded as nonsubstantial. PMID:24752258

  17. In vitro susceptibility of bovine mastitis pathogens to a combination of penicillin and framycetin: development of interpretive criteria for testing by broth microdilution and disk diffusion.

    PubMed

    Pillar, C M; Stoneburner, A; Shinabarger, D L; Abbeloos, E; Goby, L; Bradley, Andrew J

    2014-10-01

    Dry cow therapy is an important part of mastitis control. This therapy typically consists of an antibiotic or antibiotics administered at a single dose by intramammary infusion at dry off to treat or prevent infection by prevalent mastitis pathogens. A combination dry cow therapy consisting of the active components penicillin and framycetin is currently used in several countries. Despite its use, standardized methods for the susceptibility testing of this combination against mastitis pathogens have not been established. In this study, which used Clinical and Laboratory Standards Institute methodology, preliminary interpretive criteria for the broth microdilution minimum inhibitory concentration (MIC) testing of mastitis pathogens to penicillin combined with framycetin (2:1 wt/wt) were established based on the amount of drug achieved and maintained postadministration in the udder. Based on resulting MIC distributions of recent veterinary field isolates and a subset of isolates preselected for resistance to β-lactams or aminoglycosides and concentrations achieved postadministration, criteria for broth microdilution testing of the combination (susceptible, intermediate, resistant in micrograms per milliliter) were set as follows: Escherichia coli ≤8/4, 16/8, ≥32/16; Staphylococcus spp. ≤2/1, 4/2-8/4, >16/8; Streptococcus uberis and Streptococcus dysgalactiae <0.25/0.12, 0.5/0.25-2/1, >4/2. A disk diffusion test using disks containing 100 μg of framycetin and 10 IU of penicillin was also developed, and preliminary interpretive criteria (susceptible, intermediate, resistant in millimeters) were set based on correlation to broth MIC values and the minimization of interpretive errors between isolates tested concurrently by broth microdilution and disk diffusion as follows: E. coli ≥18, 16-17, ≤15; Staphylococcus spp. ≥21, 18-20, ≤17; Strep. uberis and Strep. dysgalactiae ≥21, 19-20, ≤18. In addition, ranges for the quality control of the testing of

  18. Comparison of the WST-8 colorimetric method and the CLSI broth microdilution method for susceptibility testing against drug-resistant bacteria.

    PubMed

    Tsukatani, Tadayuki; Suenaga, Hikaru; Shiga, Masanobu; Noguchi, Katsuya; Ishiyama, Munetaka; Ezoe, Takatoshi; Matsumoto, Kiyoshi

    2012-09-01

    The minimum inhibitory concentrations (MICs) obtained from the susceptibility testing of various bacteria to antibiotics were determined by a colorimetric microbial viability assay based on reduction of a tetrazolium salt {2-(2-methoxy-4-nitrophenyl)-3-(4-nitrophenyl)-5-(2,4-disulfophenyl)-2H-tetrazolium, monosodium salt (WST-8)} via 2-methyl-1,4-napthoquinone as an electron mediator and compared with those obtained by the broth microdilution methods approved by the Clinical and Laboratory Standard Institute (CLSI). Especially for drug-resistant bacteria, the CLSI method at an incubation time of 24h tended to give lower MICs. The extension of incubation time was necessary to obtain consistent MICs for drug-resistant bacteria such as methicillin-resistant Staphylococcus aureus (MRSA), vancomycin-resistant enterococi (VRE) and multi-drug resistant Pseudomonas aeruginosa (MDRP) in the broth microdilution method. There was excellent agreement between the MICs determined after 24h using the WST-8 colorimetric method and those obtained after 48-96 h using the broth microdilution method. The results suggest that the WST-8 colorimetric assay is a useful method for rapid determination of consistent MICs for drug-resistant bacteria. Copyright © 2012 Elsevier B.V. All rights reserved.

  19. In vitro activity of a new oral glucan synthase inhibitor (MK-3118) tested against Aspergillus spp. by CLSI and EUCAST broth microdilution methods.

    PubMed

    Pfaller, Michael A; Messer, Shawn A; Motyl, Mary R; Jones, Ronald N; Castanheira, Mariana

    2013-02-01

    MK-3118, a glucan synthase inhibitor derived from enfumafungin, and comparator agents were tested against 71 Aspergillus spp., including itraconazole-resistant strains (MIC, ≥ 4 μg/ml), using CLSI and EUCAST reference broth microdilution methods. The CLSI 90% minimum effective concentration (MEC(90))/MIC(90) values (μg/ml) for MK-3118, amphotericin B, and caspofungin, respectively, were as follows: 0.12, 2, and 0.03 for Aspergillus flavus species complex (SC); 0.25, 2, and 0.06 for Aspergillus fumigatus SC; 0.12, 2, and 0.06 for Aspergillus terreus SC; and 0.06, 1, and 0.03 for Aspergillus niger SC. Essential agreement between the values found by CLSI and EUCAST (± 2 log(2) dilution steps) was 94.3%. MK-3118 was determined to be a potent agent regardless of the in vitro method applied, with excellent activity against contemporary wild-type and itraconazole-resistant strains of Aspergillus spp.

  20. In Vitro Activity of a New Oral Glucan Synthase Inhibitor (MK-3118) Tested against Aspergillus spp. by CLSI and EUCAST Broth Microdilution Methods

    PubMed Central

    Messer, Shawn A.; Motyl, Mary R.; Jones, Ronald N.; Castanheira, Mariana

    2013-01-01

    MK-3118, a glucan synthase inhibitor derived from enfumafungin, and comparator agents were tested against 71 Aspergillus spp., including itraconazole-resistant strains (MIC, ≥4 μg/ml), using CLSI and EUCAST reference broth microdilution methods. The CLSI 90% minimum effective concentration (MEC90)/MIC90 values (μg/ml) for MK-3118, amphotericin B, and caspofungin, respectively, were as follows: 0.12, 2, and 0.03 for Aspergillus flavus species complex (SC); 0.25, 2, and 0.06 for Aspergillus fumigatus SC; 0.12, 2, and 0.06 for Aspergillus terreus SC; and 0.06, 1, and 0.03 for Aspergillus niger SC. Essential agreement between the values found by CLSI and EUCAST (±2 log2 dilution steps) was 94.3%. MK-3118 was determined to be a potent agent regardless of the in vitro method applied, with excellent activity against contemporary wild-type and itraconazole-resistant strains of Aspergillus spp. PMID:23229479

  1. Antifungal susceptibility of Malassezia furfur, Malassezia sympodialis, and Malassezia globosa to azole drugs and amphotericin B evaluated using a broth microdilution method.

    PubMed

    Rojas, Florencia D; Sosa, María de los A; Fernández, Mariana S; Cattana, María E; Córdoba, Susana B; Giusiano, Gustavo E

    2014-08-01

    We studied the in vitro activity of fluconazole (FCZ), ketoconazole (KTZ), miconazole (MCZ), voriconazole (VCZ), itraconazole (ITZ) and amphotericin B (AMB) against the three major pathogenic Malassezia species, M. globosa, M. sympodialis, and M. furfur. Antifungal susceptibilities were determined using the broth microdilution method in accordance with Clinical and Laboratory Standards Institute reference document M27-A3. To support lipid-dependent yeast development, glucose, peptone, ox bile, malt extract, glycerol, and Tween supplements were added to Roswell Park Memorial Institute RPMI 1640 medium. The supplemented medium allowed good growth of all three species studied. The minimal inhibitory concentrations (MICs) were recorded after 72 h of incubation at 32ºC. The three species showed different susceptibility profiles for the drugs tested. Malassezia sympodialis was the most susceptible and M. furfur the least susceptible species. KTZ, ITZ, and VCZ were the most active drugs, showing low variability among isolates of the same species. FCZ, MCZ, and AMB showed high MICs and wide MIC ranges. Differences observed emphasize the need to accurately identify and evaluate antifungal susceptibility of Malassezia species. Further investigations and collaborative studies are essential for correlating in vitro results with clinical outcomes since the existing limited data do not allow definitive conclusions.

  2. 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

  3. Antifungal susceptibility of 205 Candida spp. isolated primarily during invasive Candidiasis and comparison of the Vitek 2 system with the CLSI broth microdilution and Etest methods.

    PubMed

    Bourgeois, N; Dehandschoewercker, L; Bertout, S; Bousquet, P-J; Rispail, P; Lachaud, L

    2010-01-01

    Infections due to Candida spp. are frequent, particularly in immunocompromised and intensive care unit patients. Antifungal susceptibility tests are now required to optimize antifungal treatment given the emergence of acquired antifungal resistance in some Candida species. An antifungal susceptibility automated method, the Vitek 2 system (VK2), was evaluated. VK2 was compared to the CLSI broth microdilution reference method and the Etest procedure. For this purpose, 205 clinical isolates of Candida spp., including 11 different species, were tested for fluconazole, voriconazole, and amphotericin B susceptibility. For azoles, essential agreement ranged from 25% to 100%, depending on the method used and the Candida species tested. Categorical agreements for all of the species averaged 92.2% and ranged from 14.3 to 100%, depending on the 24-h or 48-h MIC reading by the Etest and CLSI methods and on the Candida species. Results obtained for Candida albicans showed excellent categorical and essential agreements with the two comparative methods. For Candida glabrata, the essential agreement was high with the CLSI method but low with the Etest method, and several very major errors in interpretation were observed between VK2 and the Etest method for both azoles. Low MICs of fluconazole were obtained for all of the Candida krusei isolates, but the VK2 expert software corrected all of the results obtained to resistant. Amphotericin B results showed MICs of < or = 1 mg/liter for 201 (VK2), 190 (CLSI), and 202 (Etest) isolates. The AST-YS01 Vitek 2 card system (bioMérieux) is a reliable and practical standardized automated antifungal susceptibility test. Nevertheless, more assays are needed to better evaluate C. glabrata fluconazole sensitivity.

  4. Antifungal Susceptibility of 205 Candida spp. Isolated Primarily during Invasive Candidiasis and Comparison of the Vitek 2 System with the CLSI Broth Microdilution and Etest Methods▿

    PubMed Central

    Bourgeois, N.; Dehandschoewercker, L.; Bertout, S.; Bousquet, P.-J.; Rispail, P.; Lachaud, L.

    2010-01-01

    Infections due to Candida spp. are frequent, particularly in immunocompromised and intensive care unit patients. Antifungal susceptibility tests are now required to optimize antifungal treatment given the emergence of acquired antifungal resistance in some Candida species. An antifungal susceptibility automated method, the Vitek 2 system (VK2), was evaluated. VK2 was compared to the CLSI broth microdilution reference method and the Etest procedure. For this purpose, 205 clinical isolates of Candida spp., including 11 different species, were tested for fluconazole, voriconazole, and amphotericin B susceptibility. For azoles, essential agreement ranged from 25% to 100%, depending on the method used and the Candida species tested. Categorical agreements for all of the species averaged 92.2% and ranged from 14.3 to 100%, depending on the 24-h or 48-h MIC reading by the Etest and CLSI methods and on the Candida species. Results obtained for Candida albicans showed excellent categorical and essential agreements with the two comparative methods. For Candida glabrata, the essential agreement was high with the CLSI method but low with the Etest method, and several very major errors in interpretation were observed between VK2 and the Etest method for both azoles. Low MICs of fluconazole were obtained for all of the Candida krusei isolates, but the VK2 expert software corrected all of the results obtained to resistant. Amphotericin B results showed MICs of ≤1 mg/liter for 201 (VK2), 190 (CLSI), and 202 (Etest) isolates. The AST-YS01 Vitek 2 card system (bioMérieux) is a reliable and practical standardized automated antifungal susceptibility test. Nevertheless, more assays are needed to better evaluate C. glabrata fluconazole sensitivity. PMID:19889902

  5. Invitro antifungal susceptibilities of Candida species to liposomal amphotericin B, determined using CLSI broth microdilution, and amphotericin B deoxycholate, measured using the Etest.

    PubMed

    Lovero, Grazia; De Giglio, Osvalda; Rutigliano, Serafina; Diella, Giusy; Caggiano, Giuseppina; Montagna, Maria Teresa

    2017-03-01

    The antifungal susceptibilities of 598 isolates of Candida spp. (bloodstream and other sterile sites) to liposomal amphotericin B (L-AmB) versus amphotericin B (AmB) were determined. MICs were calculated using the Clinical and Laboratory Standards Institute broth microdilution (M27-A3) method for L-AmB and the Etest method for AmB. The MIC50/MIC90 (µg ml-1) values for L-AmB broth microdilution and AmB Etest were 0.25/1 and 0.19/0.5, respectively. The overall essential agreement (±2 dilutions) was 91.5 %, ranging from 37.5 % (Candida lusitaniae) to 100 % (Candida glabrata and Candida krusei). Categorical agreement between the two methods was categorized based on a previously published breakpoint (susceptible/resistant MIC cut-off of 1 µg ml-1). The overall categorical agreement at the 48 h reading was 97.3 %, ranging from 72.7 % (C. krusei) to 100 % (Candida albicans). Major and very major discrepancies occurred in 2.3 and 0.3 %, respectively. Spearman's ρ was 0.48 (P<0.0001). These results demonstrate the utility of the AmB Etest as a surrogate marker to predict the sensibility and resistance of Candida spp. to L-AmB and thus to support its use in antifungal treatment.

  6. Activities of E1210 and Comparator Agents Tested by CLSI and EUCAST Broth Microdilution Methods against Fusarium and Scedosporium Species Identified Using Molecular Methods

    PubMed Central

    Duncanson, Frederick P.; Diekema, Daniel J.; Guarro, Josep; Jones, Ronald N.; Pfaller, Michael A.

    2012-01-01

    Fusarium (n = 67) and Scedosporium (n = 63) clinical isolates were tested by two reference broth microdilution (BMD) methods against a novel broad-spectrum (active against both yeasts and molds) antifungal, E1210, and comparator agents. E1210 inhibits the inositol acylation step in glycophosphatidylinositol (GPI) biosynthesis, resulting in defects in fungal cell wall biosynthesis. Five species complex organisms/species of Fusarium (4 isolates unspeciated) and 28 Scedosporium apiospermum, 7 Scedosporium aurantiacum, and 28 Scedosporium prolificans species were identified by molecular techniques. Comparator antifungal agents included anidulafungin, caspofungin, itraconazole, posaconazole, voriconazole, and amphotericin B. E1210 was highly active against all of the tested isolates, with minimum effective concentration (MEC)/MIC90 values (μg/ml) for E1210, anidulafungin, caspofungin, itraconazole, posaconazole, voriconazole, and amphotericin B, respectively, for Fusarium of 0.12, >16, >16, >8, >8, 8, and 4 μg/ml. E1210 was very potent against the Scedosporium spp. tested. The E1210 MEC90 was 0.12 μg/ml for S. apiospermum, but 1 to >8 μg/ml for other tested agents. Against S. aurantiacum, the MEC50 for E1210 was 0.06 μg/ml versus 0.5 to >8 μg/ml for the comparators. Against S. prolificans, the MEC90 for E1210 was only 0.12 μg/ml, compared to >4 μg/ml for amphotericin B and >8 μg/ml for itraconazole, posaconazole, and voriconazole. Both CLSI and EUCAST methods were highly concordant for E1210 and all comparator agents. The essential agreement (EA; ±2 doubling dilutions) was >93% for all comparisons, with the exception of posaconazole and F. oxysporum species complex (SC) (60%), posaconazole and S. aurantiacum (85.7%), and voriconazole and S. aurantiacum (85.7%). In conclusion, E1210 exhibited very potent and broad-spectrum antifungal activity against azole- and amphotericin B-resistant strains of Fusarium spp. and Scedosporium spp. Furthermore, in vitro

  7. Progress in Antifungal Susceptibility Testing of Candida spp. by Use of Clinical and Laboratory Standards Institute Broth Microdilution Methods, 2010 to 2012

    PubMed Central

    Pfaller, M. A.

    2012-01-01

    Antifungal susceptibility testing of Candida has been standardized and refined and now may play a useful role in managing Candida infections. Important new developments include validation of 24-h reading times for all antifungal agents and the establishment of species-specific epidemiological cutoff values (ECVs) for the systemically active antifungal agents and both common and uncommon species of Candida. The clinical breakpoints (CBPs) for fluconazole, voriconazole, and the echinocandins have been revised to provide species-specific interpretive criteria for the six most common species. The revised CBPs not only are predictive of clinical outcome but also provide a more sensitive means of identifying those strains with acquired or mutational resistance mechanisms. This brief review serves as an update on the new developments in the antifungal susceptibility testing of Candida spp. using Clinical and Laboratory Standards Institute (CLSI) broth microdilution (BMD) methods. PMID:22740712

  8. Determination of Legionella pneumophila susceptibility to Melaleuca alternifolia Cheel (tea tree) oil by an improved broth micro-dilution method under vapour controlled conditions.

    PubMed

    Mondello, Francesca; Girolamo, Antonietta; Scaturro, Maria; Ricci, Maria Luisa

    2009-05-01

    The aim of this study was to determine the in vitro activity of Melaleuca alternifolia Cheel (tea tree) oil (TTO) against 22 strains of Legionella pneumophila of different serogroup and source of isolation. Both a standard broth micro-dilution method, with slight modifications, and a micro-atmosphere diffusion method were used. Furthermore, we have established a simple sealing procedure in the micro-dilution method to determine the antibacterial activity of TTO against Legionella in aqueous phase. The results showed that L. pneumophila, quite irrespective of serogroup and source of isolation, is exquisitely sensitive to TTO, with minimal inhibitory concentration (MIC) ranging from 0.125 to 0.5% v/v, and a bactericidal activity at 0.5% v/v. In addition, we show here that TTO vapours exert critical activity, that must be controlled for reproducible MIC determinations. Overall, our data suggest that TTO could be active as anti-Legionella disinfectant, for control of water system contamination, especially in spas, in small waterlines or in particular respiratory medical devices.

  9. [Comparison of disc diffusion, E-test, and broth microdilution methods for the determination of resistance to colistin, polymyxin B, and tigecycline in multi-resistant Acinetobacter baumannii isolates].

    PubMed

    Akin, F Ebru Ozgür; Bayram, Ayşen; Balci, Iclal

    2010-04-01

    The increased rate of antimicrobial resistance in Acinetobacter baumannii made it necessary to reconsider old antibiotics such as polymyxins and develop new drugs such as tigecycline. The aim of this study was to investigate the susceptibility rates of multi-drug resistant A. baumannii clinical isolates to colistin, polymyxin B, and tigecycline by three different methods in microbiology laboratory of Gaziantep University Research Hospital, between September 2006 and April 2008. A total of 200 A. boumannii strains isolated from various clinical samples (tracheal aspirate, blood, sputum, surgical wound, catheter, pleural fluid, urine, and others) were included to the study. Identification of bacteria was performed by conventional microbiological methods and by an automatized identification system (Vitek 2, bioMerieux, France). Antimicrobial susceptibility pattern of A. baumannii isolates was determined by disc diffusion method and 95 multiple resistant A. baumannii isolates were identified. Susceptibilities of these multiple resistant bacteria to colistin, polymyxin B, and tigecycline were tested with disc diffusion, E-test, and broth microdilution methods. All of the isolates (100%) were sensitive to colistin with all three methods. Ninety-two (96.8%) of them were sensitive to polymyxin B with both disc diffusion and broth microdilution methods, and 90 (94.7%) of them were sensitive to polymyxin B with E-test. Eighty-three (87.4%) of them were sensitive to tigecycline by disc diffusion method, 78 (82.1%) by E-test, and 90 (94.7%) by broth microdilution method. No statistically significant difference was detected for the three methods in terms of susceptibility testing for polymyxin B (p > 0.05). However, while no significant difference wa detected for tigecycline susceptibility testing by disk diffusion and broth microdilution (p > 0.05), statistically significant difference was determined for broth microdilution and E-test methods (p = 0.000). In conclusion

  10. Comparison of the Vitek 2 Antifungal Susceptibility System with the Clinical and Laboratory Standards Institute (CLSI) and European Committee on Antimicrobial Susceptibility Testing (EUCAST) Broth Microdilution Reference Methods and with the Sensititre YeastOne and Etest Techniques for In Vitro Detection of Antifungal Resistance in Yeast Isolates ▿ ‖

    PubMed Central

    Cuenca-Estrella, Manuel; Gomez-Lopez, Alicia; Alastruey-Izquierdo, Ana; Bernal-Martinez, Leticia; Cuesta, Isabel; Buitrago, Maria J.; Rodriguez-Tudela, Juan L.

    2010-01-01

    The commercial technique Vitek 2 system for antifungal susceptibility testing of yeast species was evaluated. A collection of 154 clinical yeast isolates, including amphotericin B- and azole-resistant organisms, was tested. Results were compared with those obtained by the reference procedures of both the CLSI and the European Committee on Antimicrobial Susceptibility Testing (EUCAST). Two other commercial techniques approved for clinical use, the Etest and the Sensititre YeastOne, were included in the comparative exercise as well. The average essential agreement (EA) between the Vitek 2 system and the reference procedures was >95%, comparable with the average EAs observed between the reference procedures and the Sensititre YeastOne and Etest. The EA values were >97% for Candida spp. and stood at 92% for Cryptococcus neoformans. Intraclass correlation coefficients (ICC) between the commercial techniques and the reference procedures were statistically significant (P < 0.01). Percentages of very major errors were 2.6% between Vitek 2 and the EUCAST technique and 1.6% between Vitek 2 and the CLSI technique. The Vitek 2 MIC results were available after 14 to 18 h of incubation for all Candida spp. (average time to reading, 15.5 h). The Vitek 2 system was shown to be a reliable technique to determine antifungal susceptibility testing of yeast species and a more rapid and easier alternative for clinical laboratories than the procedures developed by either the CLSI or EUCAST. PMID:20220169

  11. Comparison of the Vitek 2 antifungal susceptibility system with the clinical and laboratory standards institute (CLSI) and European Committee on Antimicrobial Susceptibility Testing (EUCAST) Broth Microdilution Reference Methods and with the Sensititre YeastOne and Etest techniques for in vitro detection of antifungal resistance in yeast isolates.

    PubMed

    Cuenca-Estrella, Manuel; Gomez-Lopez, Alicia; Alastruey-Izquierdo, Ana; Bernal-Martinez, Leticia; Cuesta, Isabel; Buitrago, Maria J; Rodriguez-Tudela, Juan L

    2010-05-01

    The commercial technique Vitek 2 system for antifungal susceptibility testing of yeast species was evaluated. A collection of 154 clinical yeast isolates, including amphotericin B- and azole-resistant organisms, was tested. Results were compared with those obtained by the reference procedures of both the CLSI and the European Committee on Antimicrobial Susceptibility Testing (EUCAST). Two other commercial techniques approved for clinical use, the Etest and the Sensititre YeastOne, were included in the comparative exercise as well. The average essential agreement (EA) between the Vitek 2 system and the reference procedures was >95%, comparable with the average EAs observed between the reference procedures and the Sensititre YeastOne and Etest. The EA values were >97% for Candida spp. and stood at 92% for Cryptococcus neoformans. Intraclass correlation coefficients (ICC) between the commercial techniques and the reference procedures were statistically significant (P<0.01). Percentages of very major errors were 2.6% between Vitek 2 and the EUCAST technique and 1.6% between Vitek 2 and the CLSI technique. The Vitek 2 MIC results were available after 14 to 18 h of incubation for all Candida spp. (average time to reading, 15.5 h). The Vitek 2 system was shown to be a reliable technique to determine antifungal susceptibility testing of yeast species and a more rapid and easier alternative for clinical laboratories than the procedures developed by either the CLSI or EUCAST.

  12. [Comparison of broth microdilution and E-test methods for the antifungal susceptibility testing of Candida spp. strains isolated from blood cultures].

    PubMed

    Ozcan, Sema Keçeli; Mutlu, Birsen; Dündar, Devrim; Willke, Ayşe

    2010-04-01

    The incidence of serious fungal infections, particularly invasive Candida infections exhibit an increasing trend in the last decades since the number of patients receiving immunosuppressive treatment is increasing. This situation eventually results in an increment in resistance to antifungal agents. The aim of this study was to compare the standard broth microdilution (BMD) and E-test methods for antifungal susceptibility testing of Candida species isolated from blood cultures in our hospital, against fluconazole, voriconazole, caspofungin and amphotericin B. A total of 46 Candida strains isolated from the blood cultures by BACTEC 9000 (Becton Dickinson, USA) and identified by conventional techniques and API 20C AUX (BioMerieux, France) during January 2006-December 2007, were included into this study. The identification results of the isolates were as follows: C. albicans (23), C. parapsilosis (10), C. tropicalis (5), C. krusei (3), C. famata (2), C. glabrata (1), C. guilliermondii (1), C. kefyr (1). The antifungal susceptibilities were determined by BMD method described in Clinical and Laboratory Standards Institute M27-A3 document and E-test. Only two isolates (C. albicans and C. globrata) were found to be resistant to fluconazole with E-test but susceptible with BMD. The minimal inhibitory concentration (MIC) values of caspofungin were higher (MIC = 1-2 microg/ml) in C. parapsilosis compared to other Candida species using E-test. Only one C. albicans was resistant to voriconazole by E-test (MIC = 4 microg/ml), but it was susceptible by BMD (MIC = 0.08 microg/ml). Since definite resistance breakpoints do not yet exist for amphotericin B, MIC values were considered for amphotericin B and it was found that all strains had identical low MIC values (< 0.002-0.5). When E-test results were compared with the standard BMD results, MIC values were in agreement 80.4% for fluconazole, 84.7% for amphotericin B, 95.6% for voriconazole and 93.4% for caspofungin. These results

  13. Wild-Type MIC Distributions and Epidemiological Cutoff Values for Amphotericin B and Aspergillus spp. for the CLSI Broth Microdilution Method (M38-A2 Document)▿

    PubMed Central

    Espinel-Ingroff, A.; Cuenca-Estrella, M.; Fothergill, A.; Fuller, J.; Ghannoum, M.; Johnson, E.; Pelaez, T.; Pfaller, M. A.; Turnidge, J.

    2011-01-01

    Although clinical breakpoints have not been established for mold testing, epidemiological cutoff values (ECVs) are available for Aspergillus spp. versus the triazoles and caspofungin. Wild-type (WT) MIC distributions (organisms in a species-drug combination with no acquired resistance mechanisms) were defined in order to establish ECVs for six Aspergillus spp. and amphotericin B. Two sets (CLSI/EUCAST broth microdilution) of available MICs were evaluated: those for A. fumigatus (3,988/833), A. flavus (793/194), A. nidulans (184/69), A. niger (673/140), A. terreus (545/266), and A. versicolor (135/22). Three sets of data were analyzed: (i) CLSI data gathered in eight independent laboratories in Canada, Europe, and the United States; (ii) EUCAST data from a single laboratory; and (iii) the combined CLSI and EUCAST data. ECVs, expressed in μg/ml, that captured 95%, 97.5%, and 99% of the modeled wild-type population (CLSI and combined data) were as follows: for A. fumigatus, 2, 2, and 4; for A. flavus, 2, 4, and 4; for A. nidulans, 4, 4, and 4; for A. niger, 2, 2, and 2; for A. terreus, 4, 4, and 8; and for A. versicolor, 2, 2, and 2. Similar to the case for the triazoles and caspofungin, amphotericin B ECVs may aid in the detection of strains with acquired mechanisms of resistance to this agent. PMID:21876047

  14. In vitro Degradation of Antimicrobials during Use of Broth Microdilution Method Can Increase the Measured Minimal Inhibitory and Minimal Bactericidal Concentrations

    PubMed Central

    Lallemand, Elodie A.; Lacroix, Marlène Z.; Toutain, Pierre-Louis; Boullier, Séverine; Ferran, Aude A.; Bousquet-Melou, Alain

    2016-01-01

    The antibacterial activity of some antimicrobials may be under-estimated during in vitro microbiological susceptibility tests, due to their instability under such conditions. The in vitro degradation of seven widely used antimicrobials (amoxicillin, cephalexin monohydrate, cefotaxime sodium salt, ciprofloxacin, erythromycin hydrate, clarithromycin, and doxycycline hyclate) and its effect on MIC and MBC determinations was studied using the broth microdilution method, considered as the gold standard for MIC determinations. In vitro concentrations of antimicrobials, over a 24 h incubation period in the medium tested without bacteria, decreased from 33% for ciprofloxacin to 69% for clarithromycin. For cephalexin, cefotaxime, clarithromycin, and doxycycline which were the most degraded drugs, MIC and MBC values for one strain of E. coli and one strain of S. aureus were compared using the standard method or after ad-hoc drug complementation aiming at maintaining constant drug concentration. Abiotic degradation during the standard method was associated with a significant increase of the MIC (2 antibiotics) and MBC (3 antibiotics). However, the observed discrepancy (less than one twofold dilution), even for the most degraded drug for which the concentration at 24 h was reduced by two thirds, suggests that this would only be clinically significant in special cases such as slow-growing bacteria. PMID:28066372

  15. Assessment of telavancin minimal inhibitory concentrations by revised broth microdilution method in phase 3 complicated skin and skin-structure infection clinical trial isolates.

    PubMed

    Smart, Jennifer I; Corey, G Ralph; Stryjewski, Martin E; Wang, Whedy; Barriere, Steven L

    2017-03-01

    The broth microdilution (BMD) MIC testing method for telavancin was recently revised BMD (rBMD) to improve accuracy and reproducibility. Staphylococcus aureus isolates from telavancin phase 3 complicated skin and skin-structure infection (cSSSI) studies were tested using the rBMD method. Retesting of 1132 isolates produced MICs ranging from ≤0.015 to 0.12μg/mL that were 8-fold lower than the original method. All isolates tested remained susceptible to telavancin at the revised susceptibility breakpoint of 0.12μg/mL. The clinical cure and microbiological eradication rates were 90% (368/409) and 89% (366/409) for telavancin-treated patients, and were similar for patients with methicillin-susceptible and -resistant S. aureus isolates and S. aureus isolates with elevated vancomycin MICs (≥1μg/mL). The data presented here are aimed to update the literature and better inform clinicians and clinical microbiologists about the revised telavancin MICs, as well as the corresponding clinical and microbiological cure rates for cSSSI patients.

  16. Wild-type MIC distributions and epidemiological cutoff values for amphotericin B and Aspergillus spp. for the CLSI broth microdilution method (M38-A2 document).

    PubMed

    Espinel-Ingroff, A; Cuenca-Estrella, M; Fothergill, A; Fuller, J; Ghannoum, M; Johnson, E; Pelaez, T; Pfaller, M A; Turnidge, J

    2011-11-01

    Although clinical breakpoints have not been established for mold testing, epidemiological cutoff values (ECVs) are available for Aspergillus spp. versus the triazoles and caspofungin. Wild-type (WT) MIC distributions (organisms in a species-drug combination with no acquired resistance mechanisms) were defined in order to establish ECVs for six Aspergillus spp. and amphotericin B. Two sets (CLSI/EUCAST broth microdilution) of available MICs were evaluated: those for A. fumigatus (3,988/833), A. flavus (793/194), A. nidulans (184/69), A. niger (673/140), A. terreus (545/266), and A. versicolor (135/22). Three sets of data were analyzed: (i) CLSI data gathered in eight independent laboratories in Canada, Europe, and the United States; (ii) EUCAST data from a single laboratory; and (iii) the combined CLSI and EUCAST data. ECVs, expressed in μg/ml, that captured 95%, 97.5%, and 99% of the modeled wild-type population (CLSI and combined data) were as follows: for A. fumigatus, 2, 2, and 4; for A. flavus, 2, 4, and 4; for A. nidulans, 4, 4, and 4; for A. niger, 2, 2, and 2; for A. terreus, 4, 4, and 8; and for A. versicolor, 2, 2, and 2. Similar to the case for the triazoles and caspofungin, amphotericin B ECVs may aid in the detection of strains with acquired mechanisms of resistance to this agent.

  17. Multicenter Study of Isavuconazole MIC Distributions and Epidemiological Cutoff Values for Aspergillus spp. for the CLSI M38-A2 Broth Microdilution Method

    PubMed Central

    Chowdhary, A.; Gonzalez, G. M.; Lass-Flörl, C.; Martin-Mazuelos, E.; Meis, J.; Peláez, T.; Pfaller, M. A.; Turnidge, J.

    2013-01-01

    Epidemiological cutoff values (ECVs) were established for the new triazole isavuconazole and Aspergillus species wild-type (WT) MIC distributions (organisms in a species-drug combination with no detectable acquired resistance mechanisms) that were defined with 855 Aspergillus fumigatus, 444 A. flavus, 106 A. nidulans, 207 A. niger, 384 A. terreus, and 75 A. versicolor species complex isolates; 22 Aspergillus section Usti isolates were also included. CLSI broth microdilution MIC data gathered in Europe, India, Mexico, and the United States were aggregated to statistically define ECVs. ECVs were 1 μg/ml for the A. fumigatus species complex, 1 μg/ml for the A. flavus species complex, 0.25 μg/ml for the A. nidulans species complex, 4 μg/ml for the A. niger species complex, 1 μg/ml for the A. terreus species complex, and 1 μg/ml for the A. versicolor species complex; due to the small number of isolates, an ECV was not proposed for Aspergillus section Usti. These ECVs may aid in detecting non-WT isolates with reduced susceptibility to isavuconazole due to cyp51A (an A. fumigatus species complex resistance mechanism among the triazoles) or other mutations. PMID:23716059

  18. Comparison between E-test and CLSI broth microdilution method for antifungal susceptibility testing of Candida albicans oral isolates.

    PubMed

    Koga-Ito, Cristiane Yumi; Lyon, Juliana Pereira; Resende, Maria Aparecida de

    2008-01-01

    Thirty Candida albicans isolated from oral candidosis patients and 30 C. albicans isolated from control individuals were studied. In vitro susceptibility tests were performed for amphotericin B, fluconazole, 5-flucytosine and itraconazole through the Clinical and Laboratorial Standards Institute (CLSI) reference method and E test system. The results obtained were analyzed and compared. MIC values were similar for the strains isolated from oral candidosis patients and control individuals. The agreement rate for the two methods was 66.67% for amphotericin B, 53.33% for fluconazole, 65% for flucytosine and 45% for itraconazole. According to our data, E test method could be an alternative to trial routine susceptibility testing due to its simplicity. However, it can not be considered a substitute for the CLSI reference method.

  19. Comparative analysis of the Vitek 2 antifungal susceptibility system and E-test with the CLSI M27-A3 broth microdilution method for susceptibility testing of Indian clinical isolates of Cryptococcus neoformans.

    PubMed

    Tewari, Aarti; Behera, Bijayani; Mathur, Purva; Xess, Immaculata

    2012-06-01

    The emergence of antifungal resistance among Cryptococcus neoformans isolates is a matter of great concern. The Clinical and Laboratory Standards Institute (CLSI) broth microdilution reference method (BMD) for antifungal susceptibility testing of C. neoformans is tedious and time-consuming. Consequently, there is a greater need for a reproducible in vitro susceptibility testing method for use in clinical microbiology laboratories. By random amplified polymorphic DNA analysis, the 62 Indian clinical isolates were characterized as Cryptococcus neoformans var. grubii. We evaluated the susceptibilities of these isolates for amphotericin B (AMB) and fluconazole (FLC) by two commercial techniques, i.e., Vitek 2 and E-test against the CLSI M27-A3 BMD. The essential agreement (EA) between the Vitek 2 and E-test with the reference procedure for FLC was similar (82.2%). For AMB, EA of 92 and 76% was obtained with E-test and Vitek 2. Excellent categorical agreement (CA) (98.3% and 100% by Vitek 2 and E-test, respectively) was obtained for AMB. The CA for FLC was 81 and 77.4% by Vitek 2 and E-test. We conclude that both E-test and Vitek 2 system have acceptable levels of accuracy for susceptibility testing of both the drugs. Both of them could identify fluconazole-resistant strains. Vitek 2 could be used for testing susceptibility of voriconazole and 5-flucytosine also at the same time.

  20. Carbapenem Susceptibility Testing Errors Using Three Automated Systems, Disk Diffusion, Etest, and Broth Microdilution and Carbapenem Resistance Genes in Isolates of Acinetobacter baumannii-calcoaceticus Complex▿

    PubMed Central

    Markelz, Ana Elizabeth; Mende, Katrin; Murray, Clinton K.; Yu, Xin; Zera, Wendy C.; Hospenthal, Duane R.; Beckius, Miriam L.; Calvano, Tatjana; Akers, Kevin S.

    2011-01-01

    The Acinetobacter baumannii-calcoaceticus complex (ABC) is associated with increasing carbapenem resistance, necessitating accurate resistance testing to maximize therapeutic options. We determined the accuracy of carbapenem antimicrobial susceptibility tests for ABC isolates and surveyed them for genetic determinants of carbapenem resistance. A total of 107 single-patient ABC isolates from blood and wound infections from 2006 to 2008 were evaluated. MICs of imipenem, meropenem, and doripenem determined by broth microdilution (BMD) were compared to results obtained by disk diffusion, Etest, and automated methods (the MicroScan, Phoenix, and Vitek 2 systems). Discordant results were categorized as very major errors (VME), major errors (ME), and minor errors (mE). DNA sequences encoding OXA beta-lactamase enzymes (blaOXA-23-like, blaOXA-24-like, blaOXA-58-like, and blaOXA-51-like) and metallo-β-lactamases (MBLs) (IMP, VIM, and SIM1) were identified by PCR, as was the KPC2 carbapenemase gene. Imipenem was more active than meropenem and doripenem. The percentage of susceptibility was 37.4% for imipenem, 35.5% for meropenem, and 3.7% for doripenem. Manual methods were more accurate than automated methods. blaOXA-23-like and blaOXA-24-like were the primary resistance genes found. blaOXA-58-like, MBLs, and KPC2 were not present. Both automated testing and manual testing for susceptibility to doripenem were very inaccurate, with VME rates ranging between 2.8 and 30.8%. International variability in carbapenem breakpoints and the absence of CLSI breakpoints for doripenem present a challenge in susceptibility testing. PMID:21807971

  1. Carbapenem susceptibility testing errors using three automated systems, disk diffusion, Etest, and broth microdilution and carbapenem resistance genes in isolates of Acinetobacter baumannii-calcoaceticus complex.

    PubMed

    Markelz, Ana Elizabeth; Mende, Katrin; Murray, Clinton K; Yu, Xin; Zera, Wendy C; Hospenthal, Duane R; Beckius, Miriam L; Calvano, Tatjana; Akers, Kevin S

    2011-10-01

    The Acinetobacter baumannii-calcoaceticus complex (ABC) is associated with increasing carbapenem resistance, necessitating accurate resistance testing to maximize therapeutic options. We determined the accuracy of carbapenem antimicrobial susceptibility tests for ABC isolates and surveyed them for genetic determinants of carbapenem resistance. A total of 107 single-patient ABC isolates from blood and wound infections from 2006 to 2008 were evaluated. MICs of imipenem, meropenem, and doripenem determined by broth microdilution (BMD) were compared to results obtained by disk diffusion, Etest, and automated methods (the MicroScan, Phoenix, and Vitek 2 systems). Discordant results were categorized as very major errors (VME), major errors (ME), and minor errors (mE). DNA sequences encoding OXA beta-lactamase enzymes (bla(OXA-23-like), bla(OXA-24-like), bla(OXA-58-like), and bla(OXA-51-like)) and metallo-β-lactamases (MBLs) (IMP, VIM, and SIM1) were identified by PCR, as was the KPC2 carbapenemase gene. Imipenem was more active than meropenem and doripenem. The percentage of susceptibility was 37.4% for imipenem, 35.5% for meropenem, and 3.7% for doripenem. Manual methods were more accurate than automated methods. bla(OXA-23-like) and bla(OXA-24-like) were the primary resistance genes found. bla(OXA-58-like), MBLs, and KPC2 were not present. Both automated testing and manual testing for susceptibility to doripenem were very inaccurate, with VME rates ranging between 2.8 and 30.8%. International variability in carbapenem breakpoints and the absence of CLSI breakpoints for doripenem present a challenge in susceptibility testing.

  2. Multicenter study of anidulafungin and micafungin MIC distributions and epidemiological cutoff values for eight Candida species and the CLSI M27-A3 broth microdilution method.

    PubMed

    Pfaller, M A; Espinel-Ingroff, A; Bustamante, B; Canton, E; Diekema, D J; Fothergill, A; Fuller, J; Gonzalez, G M; Guarro, J; Lass-Flörl, C; Lockhart, S R; Martin-Mazuelos, E; Meis, J F; Ostrosky-Zeichner, L; Pelaez, T; St-Germain, G; Turnidge, J

    2014-01-01

    Since epidemiological cutoff values (ECVs) using CLSI MICs from multiple laboratories are not available for Candida spp. and the echinocandins, we established ECVs for anidulafungin and micafungin on the basis of wild-type (WT) MIC distributions (for organisms in a species-drug combination with no detectable acquired resistance mechanisms) for 8,210 Candida albicans, 3,102 C. glabrata, 3,976 C. parapsilosis, 2,042 C. tropicalis, 617 C. krusei, 258 C. lusitaniae, 234 C. guilliermondii, and 131 C. dubliniensis isolates. CLSI broth microdilution MIC data gathered from 15 different laboratories in Canada, Europe, Mexico, Peru, and the United States were aggregated to statistically define ECVs. ECVs encompassing 97.5% of the statistically modeled population for anidulafungin and micafungin were, respectively, 0.12 and 0.03 μg/ml for C. albicans, 0.12 and 0.03 μg/ml for C. glabrata, 8 and 4 μg/ml for C. parapsilosis, 0.12 and 0.06 μg/ml for C. tropicalis, 0.25 and 0.25 μg/ml for C. krusei, 1 and 0.5 μg/ml for C. lusitaniae, 8 and 2 μg/ml for C. guilliermondii, and 0.12 and 0.12 μg/ml for C. dubliniensis. Previously reported single and multicenter ECVs defined in the present study were quite similar or within 1 2-fold dilution of each other. For a collection of 230 WT isolates (no fks mutations) and 51 isolates with fks mutations, the species-specific ECVs for anidulafungin and micafungin correctly classified 47 (92.2%) and 51 (100%) of the fks mutants, respectively, as non-WT strains. These ECVs may aid in detecting non-WT isolates with reduced susceptibility to anidulafungin and micafungin due to fks mutations.

  3. Comparison of EUCAST and CLSI broth microdilution methods for the susceptibility testing of 10 systemically active antifungal agents when tested against Candida spp.

    PubMed

    Pfaller, Michael A; Castanheira, Mariana; Messer, Shawn A; Rhomberg, Paul R; Jones, Ronald N

    2014-06-01

    The antifungal broth microdilution (BMD) method of the European Committee on Antimicrobial Susceptibility Testing (EUCAST) was compared with Clinical and Laboratory Standards Institute (CLSI) BMD method M27-A3 for amphotericin B, flucytosine, anidulafungin, caspofungin, micafungin, fluconazole, isavuconazole, itraconazole, posaconazole, and voriconazole susceptibility testing of 357 isolates of Candida. The isolates were selected from global surveillance collections to represent both wild-type (WT) and non-WT MIC results for the azoles (12% of fluconazole and voriconazole results were non-WT) and the echinocandins (6% of anidulafungin and micafungin results were non-WT). The study collection included 114 isolates of Candida albicans, 73 of C. glabrata, 76 of C. parapsilosis, 60 of C. tropicalis, and 34 of C. krusei. The overall essential agreement (EA) between EUCAST and CLSI results ranged from 78.9% (posaconazole) to 99.6% (flucytosine). The categorical agreement (CA) between methods and species of Candida was assessed using previously determined CLSI epidemiological cutoff values. The overall CA between methods was 95.0% with 2.5% very major (VM) and major (M) discrepancies. The CA was >93% for all antifungal agents with the exception of caspofungin (84.6%), where 10% of the results were categorized as non-WT by the EUCAST method and WT by the CLSI method. Problem areas with low EA or CA include testing of amphotericin B, anidulafungin, and isavuconazole against C. glabrata, itraconazole, and posaconazole against most species, and caspofungin against C. parapsilosis, C. tropicalis, and C. krusei. We confirm high level EA and CA (>90%) between the 2 methods for testing fluconazole, voriconazole, and micafungin against all 5 species. The results indicate that the EUCAST and CLSI methods produce comparable results for testing the systemically active antifungal agents against the 5 most common species of Candida; however, there are several areas where additional

  4. Multicenter Study of Anidulafungin and Micafungin MIC Distributions and Epidemiological Cutoff Values for Eight Candida Species and the CLSI M27-A3 Broth Microdilution Method

    PubMed Central

    Pfaller, M. A.; Bustamante, B.; Canton, E.; Diekema, D. J.; Fothergill, A.; Fuller, J.; Gonzalez, G. M.; Guarro, J.; Lass-Flörl, C.; Lockhart, S. R.; Martin-Mazuelos, E.; Meis, J. F.; Ostrosky-Zeichner, L.; Pelaez, T.; St-Germain, G.; Turnidge, J.

    2014-01-01

    Since epidemiological cutoff values (ECVs) using CLSI MICs from multiple laboratories are not available for Candida spp. and the echinocandins, we established ECVs for anidulafungin and micafungin on the basis of wild-type (WT) MIC distributions (for organisms in a species-drug combination with no detectable acquired resistance mechanisms) for 8,210 Candida albicans, 3,102 C. glabrata, 3,976 C. parapsilosis, 2,042 C. tropicalis, 617 C. krusei, 258 C. lusitaniae, 234 C. guilliermondii, and 131 C. dubliniensis isolates. CLSI broth microdilution MIC data gathered from 15 different laboratories in Canada, Europe, Mexico, Peru, and the United States were aggregated to statistically define ECVs. ECVs encompassing 97.5% of the statistically modeled population for anidulafungin and micafungin were, respectively, 0.12 and 0.03 μg/ml for C. albicans, 0.12 and 0.03 μg/ml for C. glabrata, 8 and 4 μg/ml for C. parapsilosis, 0.12 and 0.06 μg/ml for C. tropicalis, 0.25 and 0.25 μg/ml for C. krusei, 1 and 0.5 μg/ml for C. lusitaniae, 8 and 2 μg/ml for C. guilliermondii, and 0.12 and 0.12 μg/ml for C. dubliniensis. Previously reported single and multicenter ECVs defined in the present study were quite similar or within 1 2-fold dilution of each other. For a collection of 230 WT isolates (no fks mutations) and 51 isolates with fks mutations, the species-specific ECVs for anidulafungin and micafungin correctly classified 47 (92.2%) and 51 (100%) of the fks mutants, respectively, as non-WT strains. These ECVs may aid in detecting non-WT isolates with reduced susceptibility to anidulafungin and micafungin due to fks mutations. PMID:24277027

  5. Comparison of a spectrophotometric microdilution method with RPMI-2% glucose with the National Committee for Clinical Laboratory Standards reference macrodilution method M27-P for in vitro susceptibility testing of amphotericin B, flucytosine, and fluconazole against Candida albicans.

    PubMed Central

    Rodríguez-Tudela, J L; Berenguer, J; Martínez-Suárez, J V; Sanchez, R

    1996-01-01

    The National Committee for Clinical Laboratory Standards has proposed a reference broth macrodilution method for in vitro antifungal susceptibility testing of yeasts (the M27-P method). This method is cumbersome and time-consuming and includes MIC endpoint determination by visual and subjective inspection of growth inhibition after 48 h of incubation. An alternative microdilution procedure was compared with the M27-P method for determination of the amphotericin B, flucytosine, and fluconazole susceptibilities of 8 American Type Culture Collection strains (6 of them were quality control or reference strains) and 50 clinical isolates of candida albicans. This microdilution method uses as culture medium RPMI 1640 supplemented with 18 g of glucose per liter (RPMI-2% glucose). Preparation of drugs, basal medium, and inocula was done by following the recommendations of the National Committee for Clinical Laboratory Standards. The MIC endpoint was calculated objectively from the turbidimetric data read at 24 h. Increased growth of C. albicans in RPMI-2% glucose and its spectrophotometric reading allowed for the rapid (24 h) and objective calculation of MIC endpoints compared with previous microdilution methods with standard RPMI 1640. Nevertheless, good agreement was shown between the M27-P method and this microdilution test. The MICs obtained for the quality control or reference strains by the microdilution method were in the ranges published for those strains. For clinical isolates, the percentages of agreement were 100% for amphotericin B and fluconazole and 98.1% for flucytosine. These data suggest that this microdilution method may serve as a less subjective and more rapid alternative to the M27-P method for antifungal susceptibility testing of yeasts. PMID:8878570

  6. Use of fluconazole as a surrogate marker to predict susceptibility and resistance to voriconazole among 13,338 clinical isolates of Candida spp. Tested by clinical and laboratory standards institute-recommended broth microdilution methods.

    PubMed

    Pfaller, M A; Messer, S A; Boyken, L; Rice, C; Tendolkar, S; Hollis, R J; Diekema, D J

    2007-01-01

    Clinical laboratories frequently face the problem of delayed availability of commercially prepared approved reagents for performing susceptibility testing of new antimicrobials. Although this problem is encountered more often with antibacterial agents, it is also an issue with antifungal agents. A current example is voriconazole, a new triazole antifungal with an expanded spectrum and potency against Candida spp., Aspergillus spp., and other opportunistic fungal pathogens. The present study addresses the use of fluconazole as a surrogate marker to predict the susceptibility of Candida spp. to voriconazole. Reference broth microdilution MIC results for 13,338 strains of Candida spp. isolated from more than 200 medical centers worldwide were used. Voriconazole MICs and interpretive categories (susceptible, < or =1 microg/ml; susceptible dose dependent, 2 microg/ml; resistant, > or =4 microg/ml) were compared with those of fluconazole by regression statistics and error rate bounding analyses. For all 13,338 isolates, the absolute categorical agreement was 91.6% (false susceptible or very major error [VME], 0.0%). Since voriconazole is 16- to 32-fold more potent than fluconazole, the performance of fluconazole as a surrogate marker for voriconazole susceptibility was improved by designating those isolates with fluconazole MICs of < or =32 microg/ml as being susceptible to voriconazole, resulting in a categorical agreement of 97% with 0.1% VME. Clinical laboratories performing antifungal susceptibility testing of fluconazole against Candida spp. can reliably use these results as surrogate markers until commercial FDA-approved voriconazole susceptibility tests become available.

  7. Comparison of the Antifungal Susceptibility Testing Subcommittee of the European Committee on Antibiotic Susceptibility Testing Proposed Standard and the E-Test with the NCCLS Broth Microdilution Method for Voriconazole and Caspofungin Susceptibility Testing of Yeast Species

    PubMed Central

    Chryssanthou, Erja; Cuenca-Estrella, Manuel

    2002-01-01

    The proposed standard of the Antifungal Susceptibility Testing Subcommittee of the European Committee on Antibiotic Susceptibility Testing (AFST-EUCAST) and the E-test procedures were compared with the NCCLS reference broth microdilution method for voriconazole and caspofungin susceptibility testing of 102 clinical Candida species and Saccharomyces cerevisiae isolates. The voriconazole MIC at which 50% of strains were inhibited (MIC50) was ≤0.125 mg/liter for all yeast species except for Candida glabrata and Candida krusei, which yielded MIC50 values of 0.25 to 1 mg/liter depending on the method. Caspofungin exhibited in vitro activity (MIC50 of ≤0.125 to 2 mg/liter) against all yeast species except for Candida guilliermondii. The agreements between MICs within ±2 dilutions obtained by the NCCLS method and the EUCAST standard were 97% for voriconazole and 96% for caspofungin. Intraclass correlation coefficients were statistically significant (P < 0.05). The agreements between voriconazole MICs provided by the E-test and the NCCLS and between the E-test and the AFST-EUCAST method were 100 and 90%, respectively. Because of lower caspofungin MICs provided by the E-test, the agreement was slightly poorer with the NCCLS method (89%) than with the AFST-EUCAST procedure (94%). Both the EUCAST and the E-test procedures can be reliable techniques for susceptibility testing of yeasts to voriconazole and caspofungin. PMID:12354895

  8. The effect of carbon dioxide on susceptibility testing of azithromycin, clarithromycin and roxithromycin against clinical isolates of Streptococcus pneumoniae and Streptococcus pyogenes by broth microdilution and the Etest: Artemis Project-first-phase study.

    PubMed

    Johnson, Jack; Bouchillon, Sam; Pontani, Dennis

    1999-06-01

    OBJECTIVE: To evaluate the effect of carbon dioxide on the susceptibility testing, using broth microdilution and the Etest (AB Biodisk, Solna, Sweden), of azithromycin, clarithromycin and roxithromycin against Streptococcus pneumoniae and Streptococcus pyogenes. METHODS: Fresh clinical isolates collected from 36 hospital laboratories in 12 countries were evaluated using the Etest in the presence of carbon dioxide. The isolates were retested under ambient conditions (absence of carbon dioxide) using broth microdilution and/or the Etest. RESULTS: Carbon dioxide falsely elevated azithromycin, clarithromycin and roxithromycin MIC90S for S. pneumoniae, determined by the Etest, approximately 12-fold. Also, the azithromycin MIC90 for S. pyogenes was increased fourfold; the effect was less marked for clarithromycin and roxithromycin. When isolates were retested in the absence of carbon dioxide, using the Etest or microdilution, susceptibilities to azithromycin were comparable to those to clarithromycin (S. pneumoniae, 93.4% versus 91.3%; S. pyogenes, 96.4% versus 95.8%). Both organisms were less susceptible to roxithromycin (S. pneumoniae, 71.3%; S. pyogenes, 85.7%). An internal standard control, consisting of 50 isolates each of S. pneumoniae, S. pyogenes and Haemophilus influenzae, confirmed that azithromycin susceptibility testing resulted in falsely elevated MICs. CONCLUSIONS: Carbon dioxide falsely elevated azithromycin MICs for S. pneumoniae and S. pyogenes, with an apparent reduction in susceptibility. When the in vitro activity of azithromycin and other macrolides against S. pneumoniae and S. pyogenes is being evaluated, awareness of the pH effect is essential.

  9. Comparison of four reading methods of broth microdilution based on the clinical and laboratory standards institute M27-A3 method for Candida spp.

    PubMed

    Morishige, Hiromi; Mano, Yoko; Oguri, Toyoko; Furuya, Nobuhiko

    2012-10-01

    This study aimed to compare the susceptibilities of 5 reference strains and 28 isolates of Candida spp., to micafungin, amphotericin B, flucytosine, fluconazole, itraconazole, voriconazole, and miconazole, obtained by visually determined minimum inhibitory concentration (MIC) using the agitation method (V-A), as described in the Clinical and Laboratory Standards Institute M27-A3 document; visual determinations without agitation (V-NA); and spectrophotometric determinations for the presence or absence of agitation (SP-A and SP-NA, respectively). Our results indicate that when the V-NA, SP-A, and SP-NA-the 3 alternative microdilution procedures for MIC endpoint determinations-were compared with the V-A, excellent agreements were observed between the V-NA and V-A rather than with the spectrophotometric methods (between the SP-A or SP-NA, and V-A). Furthermore, many errors occurred while using the SP-A method in the presence of agitation and some isolates showed major errors. Three of 5 isolates that showed very major errors between the spectrophotometric SP-A or SP-NA, and the reference V-A method were trailing isolates. Therefore, it was suggested that the MICs of Candida spp. obtained by the V-NA method were more precise than those by the conventional SP-A method.

  10. Reproducible measurement of vancomycin MICs within the susceptible range in Staphylococcus aureus by a broth microdilution method with a "quasi-continuum" gradient of antibiotic concentrations.

    PubMed

    Falcón, R; Mateo, E M; Talaya, A; Giménez, E; Vinuesa, V; Clari, M Á; Navarro, D

    2017-07-25

    The availability of reproducible broth microdilution (BMD) methods including inter log2 antibiotic dilutions for measuring Staphylococcus aureus (SA) vancomycin minimum inhibitory concentrations (MICs) within the susceptible range is needed to elucidate the impact of vancomycin MICs on clinical outcomes of invasive SA infections. Here, we report on the development of a very precise BMD method that incorporates the following incremental antibiotic concentrations: 0.50, 0.62, 0.75, 0.87, 1.0, 1.25, 1.40, 1.50, 1.60, 1.75, and 2.0 μg/mL. The intra- and inter-assay coefficients of variation of this method were around 20%. The mean of the differences in MIC values for all isolates obtained across two independent runs performed at one center was 0.04 μg/mL [95% confidence interval (CI), 0.011-0.07 μg/mL] and that for ten isolates measured at two different centers was 0.04 μg/mL (95% CI, 0-13 μg/mL). Vancomycin MIC values differed by less than 0.1 μg/mL between runs for most isolates. Storage of isolates at -20 °C for up to 3 months had no impact on the vancomycin MIC values. The mean vancomycin MIC values obtained by the Etest using a standard inoculum (0.5 McFarland) were significantly higher (p ≤ 0.001) than those measured by BMD and the MIC values measured by the two methods correlated poorly (Rho, 0.319; p = 0.148). Nevertheless, the mean MIC values measured by the Etest using lower inocula (10(7) or 10(6) CFU/mL) and those measured by BMD were comparable and correlated significantly (p = 0.004 for 10(7) CFU/mL and p = 0.029 for 10(6) CFU/mL).

  11. Comparison of agar dilution, broth microdilution, disk diffusion, E-test, and BACTEC radiometric methods for antimicrobial susceptibility testing of clinical isolates of the Nocardia asteroides complex.

    PubMed Central

    Ambaye, A; Kohner, P C; Wollan, P C; Roberts, K L; Roberts, G D; Cockerill, F R

    1997-01-01

    An evaluation was undertaken to determine the optimal method for the in vitro susceptibility testing of 26 Nocardia asteroides complex isolates to the following antimicrobial agents: amikacin, ampicillin, amoxicillin-clavulanate, ceftriaxone, ciprofloxacin, erythromycin, imipenem, minocycline, and trimethoprim-sulfamethoxazole. Five testing methods were studied including the agar dilution, broth microdilution, and disk diffusion methods, the epsilometer test (E-test), and the BACTEC radiometric method. Results for each antimicrobial agent and each testing method were interpreted as indicating susceptibility, intermediate susceptibility, or resistance according to current guidelines of the National Committee for Clinical Laboratory Standards (NCCLS) for bacteria that grow aerobically and were then compared to a "gold standard" susceptibility test result. The gold standard result for each Nocardia isolate was established by a consensus of the results of the majority of testing methods used in the study. When the results were combined for all antimicrobial agents tested against all Nocardia isolates by all methods, the BACTEC radiometric method produced the highest level of agreement (97.9%) with the consensus results and had the fewest very major (n = 1), major (n = 2), and minor (n = 2) errors. In contrast, the results of the agar dilution method were in least agreement (93.2%) with the consensus results, and this method also produced the most very major (n = 8), major (n = 4), and, along with the disk diffusion method, minor (n = 6) errors. For all test methods, interpretive errors were most frequent when testing ampicillin or amoxicillin-clavulanate. Moreover, for all Nocardia nova isolates tested, ampicillin susceptibility results by any of the testing methods were not in agreement with the results of testing for beta-lactamase by the nitrocefin (Cefinase) disk method. We conclude that among the methods evaluated, the BACTEC radiometric method appeared to be the

  12. Comparison of European Committee on Antimicrobial Susceptibility Testing (EUCAST) and Etest Methods with the CLSI Broth Microdilution Method for Echinocandin Susceptibility Testing of Candida Species▿

    PubMed Central

    Pfaller, M. A.; Castanheira, M.; Diekema, D. J.; Messer, S. A.; Moet, G. J.; Jones, R. N.

    2010-01-01

    The antifungal broth microdilution (BMD) method of the European Committee on Antibiotic Susceptibility Testing (EUCAST) and the Etest agar diffusion method were compared with the Clinical and Laboratory Standards Institute (CLSI) BMD method M27-A3 for anidulafungin, caspofungin, and micafungin susceptibility testing of 133 clinical isolates of Candida species. The isolates were characterized for the presence or absence of fks1 and/or fks2 gene mutations and included 34 isolates of C. glabrata (4 mutant strains), 32 of C. albicans (1 mutant strain), 25 of C. parapsilosis, 19 of C. guilliermondii, 12 of C. tropicalis (2 mutant strains), and 11 of C. krusei. Excellent essential agreement (EA; within 2 dilutions) between the CLSI and EUCAST and CLSI and Etest MIC results was observed. The overall EA between the EUCAST and CLSI results ranged from 89.5% (caspofungin) to 99.2% (micafungin), whereas the EA between the Etest and CLSI results ranged from 90.2% (caspofungin) to 93.2% (anidulafungin). The categorical agreement (CA) between methods for each antifungal agent was assessed using previously determined epidemiological cutoff values (ECVs). Excellent CA (>90%) was observed for all comparisons between the EUCAST and CLSI results with the exceptions of C. glabrata and caspofungin (85.3%) and C. krusei and caspofungin (54.5%). The CA between the Etest and CLSI results was also excellent for all comparisons, with the exception of C. krusei and caspofungin (81.8%). All three methods were able to differentiate wild-type (WT) strains from those with fks mutations. With anidulafungin as the test reagent, the CLSI method identified 5 of 7 mutant strains, whereas the EUCAST method and the Etest identified 6 of 7 mutant strains. With either caspofungin or micafungin as the test reagent, the CLSI method identified all 7 mutant strains and the EUCAST method identified 6 of 7 mutant strains. The Etest identified all 7 mutant strains using caspofungin as the reagent. All three

  13. Comparison of European Committee on Antimicrobial Susceptibility Testing (EUCAST) and Etest methods with the CLSI broth microdilution method for echinocandin susceptibility testing of Candida species.

    PubMed

    Pfaller, M A; Castanheira, M; Diekema, D J; Messer, S A; Moet, G J; Jones, R N

    2010-05-01

    The antifungal broth microdilution (BMD) method of the European Committee on Antibiotic Susceptibility Testing (EUCAST) and the Etest agar diffusion method were compared with the Clinical and Laboratory Standards Institute (CLSI) BMD method M27-A3 for anidulafungin, caspofungin, and micafungin susceptibility testing of 133 clinical isolates of Candida species. The isolates were characterized for the presence or absence of fks1 and/or fks2 gene mutations and included 34 isolates of C. glabrata (4 mutant strains), 32 of C. albicans (1 mutant strain), 25 of C. parapsilosis, 19 of C. guilliermondii, 12 of C. tropicalis (2 mutant strains), and 11 of C. krusei. Excellent essential agreement (EA; within 2 dilutions) between the CLSI and EUCAST and CLSI and Etest MIC results was observed. The overall EA between the EUCAST and CLSI results ranged from 89.5% (caspofungin) to 99.2% (micafungin), whereas the EA between the Etest and CLSI results ranged from 90.2% (caspofungin) to 93.2% (anidulafungin). The categorical agreement (CA) between methods for each antifungal agent was assessed using previously determined epidemiological cutoff values (ECVs). Excellent CA (>90%) was observed for all comparisons between the EUCAST and CLSI results with the exceptions of C. glabrata and caspofungin (85.3%) and C. krusei and caspofungin (54.5%). The CA between the Etest and CLSI results was also excellent for all comparisons, with the exception of C. krusei and caspofungin (81.8%). All three methods were able to differentiate wild-type (WT) strains from those with fks mutations. With anidulafungin as the test reagent, the CLSI method identified 5 of 7 mutant strains, whereas the EUCAST method and the Etest identified 6 of 7 mutant strains. With either caspofungin or micafungin as the test reagent, the CLSI method identified all 7 mutant strains and the EUCAST method identified 6 of 7 mutant strains. The Etest identified all 7 mutant strains using caspofungin as the reagent. All three

  14. Comparison of two alternative microdilution procedures with the National Committee for Clinical Laboratory Standards reference macrodilution method M27-P for in vitro testing of fluconazole-resistant and -susceptible isolates of Candida albicans.

    PubMed Central

    Espinel-Ingroff, A; Rodríguez-Tudela, J L; Martínez-Suárez, J V

    1995-01-01

    The National Committee for Clinical Laboratory Standards has proposed a reference broth macrodilution method for in vitro antifungal susceptibility testing of yeasts (the M27-P method). This method is cumbersome and time-consuming and includes MIC endpoint determination by the visual and subjective inspection of growth inhibition after 48 h of incubation. Two alternative microdilution procedures for MIC endpoint determination, a spectrophotometric MIC endpoint test that evaluates 80% growth inhibition by the drug and a colorimetric method with an oxidation-reduction indicator (Alamar Blue), were compared with the M27-P method for fluconazole susceptibility testing of 45 susceptible and resistant isolates of Candida albicans. The spectrophotometric method was performed with RPMI 1640 medium with 2% glucose, and the other two tests were performed with plain RPMI 1640 medium. All tests were incubated at 35 degrees C. Excellent agreement was demonstrated between the M27-P method and both 24-h microdilution tests (97.7%) as well as between the two microdilution tests (95.5%). Also, there was agreement in the detection in vivo of fluconazole resistance by the three methods. These preliminary data indicate that both microdilution methods may serve as less subjective alternatives to the M27-P method for the determination of fluconazole MIC endpoints. PMID:8586692

  15. Interlaboratory Study of Quality Control Isolates for a Broth Microdilution Method (Modified CLSI M38-A) for Testing Susceptibilities of Dermatophytes to Antifungals▿

    PubMed Central

    Ghannoum, M. A.; Arthington-Skaggs, B.; Chaturvedi, V.; Espinel-Ingroff, A.; Pfaller, M. A.; Rennie, R.; Rinaldi, M. G.; Walsh, T. J.

    2006-01-01

    The Clinical and Laboratory Standards Institute (CLSI; formerly National Committee for Clinical Laboratory Standards, or NCCLS) M38-A standard for the susceptibility testing of filamentous fungi does not specifically address the testing of dermatophytes. In 2003, a multicenter study investigated the reproducibility of the microdilution method developed at the Center for Medical Mycology, Cleveland, Ohio, for testing the susceptibility of dermatophytes. Data from that study supported the introduction of this method for testing dermatophytes in the future version of the CLSI M38-A standard. In order for the method to be accepted by CLSI, appropriate quality control isolates needed to be identified. To that end, an interlaboratory study, involving the original six laboratories plus two additional sites, was conducted to evaluate potential candidates for quality control isolates. These candidate strains included five Trichophyton rubrum strains known to have elevated MICs to terbinafine and five Trichophyton mentagrophytes strains. Antifungal agents tested included ciclopirox, fluconazole, griseofulvin, itraconazole, posaconazole, terbinafine, and voriconazole. Based on the data generated, two quality control isolates, one T. rubrum isolate and one T. mentagrophytes isolate, were identified and submitted to the American Type Culture Collection (ATCC) for inclusion as reference strains. Ranges encompassing 95.2 to 97.9% of all data points for all seven drugs were established. PMID:17050812

  16. Detection of amphotericin B resistance in Candida haemulonii and closely related species by use of the Etest, Vitek-2 yeast susceptibility system, and CLSI and EUCAST broth microdilution methods.

    PubMed

    Shin, Jong Hee; Kim, Mi-Na; Jang, Sook Jin; Ju, Min Young; Kim, Soo Hyun; Shin, Myung Geun; Suh, Soon Pal; Ryang, Dong Wook

    2012-06-01

    The emerging fungal pathogens Candida haemulonii and Candida pseudohaemulonii often show high-level resistance to amphotericin B (AMB). We compared the utilities of five antifungal susceptibility testing methods, i.e., the Etest using Mueller-Hinton agar supplemented with glucose and methylene blue (Etest-MH), the Etest using RPMI agar supplemented with glucose (Etest-RPG), the Vitek-2 yeast susceptibility system, and the Clinical and Laboratory Standards Institute (CLSI) and European Committee on Antimicrobial Susceptibility Testing (EUCAST) broth microdilution methods, for the detection of AMB-resistant isolates of C. haemulonii and closely related species. Thirty-eight clinical isolates (8 C. haemulonii, 10 C. pseudohaemulonii, and 20 Candida auris isolates) were analyzed. Of the 18 C. haemulonii and C. pseudohaemulonii isolates, 18, 15, 18, 10, and 9 exhibited AMB MICs of >1 μg/ml by the Etest-MH, Etest-RPG, Vitek-2, CLSI, and EUCAST methods, respectively. All 20 C. auris isolates showed AMB MICs of ≤1 μg/ml by all five methods. Of the methods, the Etest-MH generated the broadest distribution of AMB MICs for all 38 isolates and showed the best discrimination between the C. haemulonii and C. pseudohaemulonii isolates (4 to 32 μg/ml) and those of C. auris (0.125 to 0.5 μg/ml). Taking the Etest-MH as the reference method, the essential agreements (within two dilutions) for the Etest-RPG, Vitek-2, CLSI, and EUCAST methods were 84, 92, 55, and 55%, respectively; the categorical agreements were 92, 92, 79, and 76%, respectively. This study provides the first data on the efficacy of the Etest-MH and its excellent agreement with Vitek-2 for discriminating AMB-resistant from AMB-susceptible isolates of these Candida species.

  17. Wild-type MIC distributions and epidemiological cutoff values for posaconazole and voriconazole and Candida spp. as determined by 24-hour CLSI broth microdilution.

    PubMed

    Pfaller, M A; Boyken, L; Hollis, R J; Kroeger, J; Messer, S A; Tendolkar, S; Diekema, D J

    2011-02-01

    We tested 16,191 strains of Candida against posaconazole and voriconazole, using the CLSI M27-A3 broth microdilution (BMD) method (24-h incubation), in order to define wild-type (WT) populations and epidemiological cutoff values (ECVs). From 2001 to 2009, 8,619 isolates of Candida albicans, 2,415 isolates of C. glabrata, 2,278 isolates of C. parapsilosis, 1,895 isolates of C. tropicalis, 508 isolates of C. krusei, 205 isolates of C. lusitaniae, 177 isolates of C. guilliermondii, and 93 isolates of C. kefyr were obtained from over 100 centers worldwide. The modal MICs (μg/ml) for posaconazole and voriconazole, respectively, were as follows: for C. albicans, 0.016 and 0.007; for C. glabrata, 0.5 and 0.06; for C. parapsilosis, 0.06 and 0.007; for C. tropicalis, 0.03 and 0.015; for C. krusei, 0.25 and 0.12; for C. lusitaniae, 0.03 and 0.007; for C. guilliermondii, 0.12 and 0.03; and for C. kefyr, 0.06 and 0.007. The ECVs (μg/ml [% of isolates that had MICs equal to or less than the ECV]) for posaconazole and voriconazole, respectively, were as follows: 0.06 (98.5) and 0.03 (98.9) for C. albicans, 2 (96.2) and 0.5 (90.4%) for C. glabrata, 0.25 (99.3) and 0.12 (97.9) for C. parapsilosis, 0.12 (97.6) and 0.06 (97.2) for C. tropicalis, 0.5 (99.8) and 0.5 (99.4) for C. krusei, 0.12 (95.6) and 0.03 (96.6) for C. lusitaniae, 0.5 (98.9) and 0.25 (98.3) for C. guilliermondii, and 0.25 (100.0) and 0.015 (100.0) for C. kefyr. In the absence of clinical breakpoints (CBPs) for posaconazole, these WT distributions and ECVs will be useful in surveillance for emergence of reduced susceptibility to posaconazole among Candida spp. Whereas a CBP for susceptibility of ≤ 1 μg/ml has been established for voriconazole and all species of Candida, it is notable that ECVs for this agent range from 10- to >100-fold lower than the CBP, depending on the species of Candida. The CBP is inadequate in detecting the emergence of voriconazole resistance among most Candida species encountered

  18. Wild-Type MIC Distributions and Epidemiological Cutoff Values for Amphotericin B, Flucytosine, and Itraconazole and Candida spp. as Determined by CLSI Broth Microdilution

    PubMed Central

    Espinel-Ingroff, A.; Canton, E.; Castanheira, M.; Cuenca-Estrella, M.; Diekema, D. J.; Fothergill, A.; Fuller, J.; Ghannoum, M.; Jones, R. N; Lockhart, S. R.; Martin-Mazuelos, E.; Melhem, M. S. C.; Ostrosky-Zeichner, L.; Pappas, P.; Pelaez, T.; Peman, J.; Rex, J.; Szeszs, M. W.

    2012-01-01

    Clinical breakpoints (CBPs) and epidemiological cutoff values (ECVs) have been established for several Candida spp. and the newer triazoles and echinocandins but are not yet available for older antifungal agents, such as amphotericin B, flucytosine, or itraconazole. We determined species-specific ECVs for amphotericin B (AMB), flucytosine (FC) and itraconazole (ITR) for eight Candida spp. (30,221 strains) using isolates from 16 different laboratories in Brazil, Canada, Europe, and the United States, all tested by the CLSI reference microdilution method. The calculated 24- and 48-h ECVs expressed in μg/ml (and the percentages of isolates that had MICs less than or equal to the ECV) for AMB, FC, and ITR, respectively, were 2 (99.8)/2 (99.2), 0.5 (94.2)/1 (91.4), and 0.12 (95.0)/0.12 (92.9) for C. albicans; 2 (99.6)/2 (98.7), 0.5 (98.0)/0.5 (97.5), and 2 (95.2)/4 (93.5) for C. glabrata; 2 (99.7)/2 (97.3), 0.5 (98.7)/0.5 (97.8), and 05. (99.7)/0.5 (98.5) for C. parapsilosis; 2 (99.8)/2 (99.2), 0.5 (93.0)/1 (90.5), and 0.5 (97.8)/0.5 (93.9) for C. tropicalis; 2 (99.3)/4 (100.0), 32 (99.4)/32 (99.3), and 1 (99.0)/2 (100.0) for C. krusei; 2 (100.0)/4 (100.0), 0.5 (95.3)/1 (92.9), and 0.5 (95.8)/0.5 (98.1) for C. lusitaniae; −/2 (100.0), 0.5 (98.8)/0.5 (97.7), and 0.25 (97.6)/0.25 (96.9) for C. dubliniensis; and 2 (100.0)/2 (100.0), 1 (92.7)/−, and 1 (100.0)/2 (100.0) for C. guilliermondii. In the absence of species-specific CBP values, these wild-type (WT) MIC distributions and ECVs will be useful for monitoring the emergence of reduced susceptibility to these well-established antifungal agents. PMID:22461672

  19. A Modified Christensen's Urea and CLSI Broth Microdilution Method for Testing Susceptibilities of Six Malassezia Species to Voriconazole, Itraconazole, and Ketoconazole

    PubMed Central

    Rincón, S.; Cepero de García, M. C.; Espinel-Ingroff, A.

    2006-01-01

    Two supplemented broths (Christensen's urea with 0.1% Tween 80 and 0.5% Tween 40 and RPMI 1640 with 1% glycerol, 1% peptone, 1.8% glucose, and 0.05% Tween 80) were evaluated to determine voriconazole, itraconazole, and ketoconazole MICs for 200 Malassezia sp. isolates. Malassezia globosa and M. restricta were the least susceptible species (MICs at which 90% of the isolates tested were inhibited, 1 to ≥8 μg/ml versus 0.25 to 1 μg/ml). PMID:16954293

  20. A modified Christensen's urea and CLSI broth microdilution method for testing susceptibilities of six Malassezia species to voriconazole, itraconazole, and ketoconazole.

    PubMed

    Rincón, S; Cepero de García, M C; Espinel-Ingroff, A

    2006-09-01

    Two supplemented broths (Christensen's urea with 0.1% Tween 80 and 0.5% Tween 40 and RPMI 1640 with 1% glycerol, 1% peptone, 1.8% glucose, and 0.05% Tween 80) were evaluated to determine voriconazole, itraconazole, and ketoconazole MICs for 200 Malassezia sp. isolates. Malassezia globosa and M. restricta were the least susceptible species (MICs at which 90% of the isolates tested were inhibited, 1 to >or=8 microg/ml versus 0.25 to 1 microg/ml).

  1. Comparison of 24-hour and 48-hour voriconazole MICs as determined by the Clinical and Laboratory Standards Institute broth microdilution method (M27-A3 document) in three laboratories: results obtained with 2,162 clinical isolates of Candida spp. and other yeasts.

    PubMed

    Espinel-Ingroff, Ana; Canton, E; Peman, J; Rinaldi, M G; Fothergill, A W

    2009-09-01

    We evaluated the performance of the 24-h broth microdilution voriconazole MIC by obtaining MICs for 2,162 clinical isolates of Candida spp. and other yeasts; the 24-h results were compared to 48-h reference MICs to assess essential, as well as categorical, agreement. Although the overall essential agreement was 88.6%, it ranged from 96.4 to 100% for 6 of the 11 species or groups of yeasts tested. The overall categorical agreement was 93.2%, and it was above 90% for eight species. However, unacceptable percentages of very major errors (false susceptibility) were observed for Candida albicans (2.7%), C. glabrata (4.1%), C. tropicalis (9.7%), and other less common yeast species (9.8%). Since it is essential to identify potentially resistant isolates and breakpoints are based on 48-h MICs, it appears that the 24-h MIC is not as clinically useful as the 48-h reference MIC. However, further characterization of these falsely susceptible MICs for three of the four common Candida spp. is needed to understand whether these errors are due to trailing misinterpretation or if the 48-h incubation is required to detect voriconazole resistance. Either in vivo versus in vitro correlations or the determination of resistance mechanisms should be investigated.

  2. Comparison of the Broth Microdilution (BMD) Method of the European Committee on Antimicrobial Susceptibility Testing with the 24-Hour CLSI BMD Method for Testing Susceptibility of Candida Species to Fluconazole, Posaconazole, and Voriconazole by Use of Epidemiological Cutoff Values▿

    PubMed Central

    Pfaller, M. A.; Espinel-Ingroff, A.; Boyken, L.; Hollis, R. J.; Kroeger, J.; Messer, S. A.; Tendolkar, S.; Diekema, D. J.

    2011-01-01

    The antifungal broth microdilution (BMD) method of the European Committee on Antimicrobial Susceptibility Testing (EUCAST) was compared with CLSI BMD method M27-A3 for fluconazole, posaconazole, and voriconazole susceptibility testing of 1,056 isolates of Candida. The isolates were obtained in 2009 from more than 60 centers worldwide and included 560 isolates of C. albicans, 175 of C. glabrata, 162 of C. parapsilosis, 124 of C. tropicalis, and 35 of C. krusei. The overall essential agreement (EA) between EUCAST and CLSI results ranged from 96.9% (voriconazole) to 98.6% (fluconazole). The categorical agreement (CA) between methods and species of Candida was assessed using previously determined epidemiological cutoff values (ECVs). The ECVs (expressed as μg/ml) for fluconazole, posaconazole, and voriconazole, respectively, were as follows: 0.12, 0.06, and 0.03 for C. albicans; 32, 2, and 0.5 for C. glabrata; 2, 0.25, and 0.12 for C. parapsilosis; 2, 0.12, and 0.06 for C. tropicalis; 64, 0.5, and 0.5 for C. krusei. Excellent CA was observed for all comparisons between the EUCAST and CLSI results for fluconazole, posaconazole, and voriconazole, respectively, for each species: 98.9%, 93.6%, and 98.6% for C. albicans; 96.0%, 98.9%, and 93.7% for C. glabrata; 90.8%, 98.1%, and 98.1% for C. parapsilosis; 99.2%, 99.2%, and 96.8% for C. tropicalis; 97.1%, 97.1%, and 97.1% for C. krusei. We demonstrate high levels of EA and CA between the CLSI and EUCAST BMD methods for testing of triazoles against Candida when the MICs were determined after 24 h and ECVs were used to differentiate wild-type (WT) from non-WT strains. These results provide additional data in favor of the harmonization of these two methods. PMID:21227994

  3. Comparison of the broth microdilution (BMD) method of the European Committee on Antimicrobial Susceptibility Testing with the 24-hour CLSI BMD method for testing susceptibility of Candida species to fluconazole, posaconazole, and voriconazole by use of epidemiological cutoff values.

    PubMed

    Pfaller, M A; Espinel-Ingroff, A; Boyken, L; Hollis, R J; Kroeger, J; Messer, S A; Tendolkar, S; Diekema, D J

    2011-03-01

    The antifungal broth microdilution (BMD) method of the European Committee on Antimicrobial Susceptibility Testing (EUCAST) was compared with CLSI BMD method M27-A3 for fluconazole, posaconazole, and voriconazole susceptibility testing of 1,056 isolates of Candida. The isolates were obtained in 2009 from more than 60 centers worldwide and included 560 isolates of C. albicans, 175 of C. glabrata, 162 of C. parapsilosis, 124 of C. tropicalis, and 35 of C. krusei. The overall essential agreement (EA) between EUCAST and CLSI results ranged from 96.9% (voriconazole) to 98.6% (fluconazole). The categorical agreement (CA) between methods and species of Candida was assessed using previously determined epidemiological cutoff values (ECVs). The ECVs (expressed as μg/ml) for fluconazole, posaconazole, and voriconazole, respectively, were as follows: 0.12, 0.06, and 0.03 for C. albicans; 32, 2, and 0.5 for C. glabrata; 2, 0.25, and 0.12 for C. parapsilosis; 2, 0.12, and 0.06 for C. tropicalis; 64, 0.5, and 0.5 for C. krusei. Excellent CA was observed for all comparisons between the EUCAST and CLSI results for fluconazole, posaconazole, and voriconazole, respectively, for each species: 98.9%, 93.6%, and 98.6% for C. albicans; 96.0%, 98.9%, and 93.7% for C. glabrata; 90.8%, 98.1%, and 98.1% for C. parapsilosis; 99.2%, 99.2%, and 96.8% for C. tropicalis; 97.1%, 97.1%, and 97.1% for C. krusei. We demonstrate high levels of EA and CA between the CLSI and EUCAST BMD methods for testing of triazoles against Candida when the MICs were determined after 24 h and ECVs were used to differentiate wild-type (WT) from non-WT strains. These results provide additional data in favor of the harmonization of these two methods.

  4. Wild-Type MIC Distributions and Epidemiological Cutoff Values for the Triazoles and Six Aspergillus spp. for the CLSI Broth Microdilution Method (M38-A2 Document)▿

    PubMed Central

    Espinel-Ingroff, A.; Diekema, D. J.; Fothergill, A.; Johnson, E.; Pelaez, T.; Pfaller, M. A.; Rinaldi, M. G.; Canton, E.; Turnidge, J.

    2010-01-01

    Clinical breakpoints have not been established for mold testing. Wild-type (WT) MIC distributions (organisms in a species/drug combination with no detectable acquired resistance mechanisms) were defined in order to establish epidemiologic cutoff values (ECVs) for five Aspergillus spp. and itraconazole, posaconazole, and voriconazole. Also, we have expanded prior ECV data for Aspergillus fumigatus. The number of available isolates varied according to the species/triazole combination as follows: 1,684 to 2,815 for A. fumigatus, 323 to 592 for A. flavus, 131 to 143 for A. nidulans, 366 to 520 for A. niger, 330 to 462 for A. terreus, and 45 to 84 for A. versicolor. CLSI broth microdilution MIC data gathered in five independent laboratories in Europe and the United States were aggregated for the analyses. ECVs expressed in μg/ml were as follows (percentages of isolates for which MICs were equal to or less than the ECV are in parentheses): A. fumigatus, itraconazole, 1 (98.8%); posaconazole, 0.5 (99.2%); voriconazole, 1 (97.7%); A. flavus, itraconazole, 1 (99.6%); posaconazole, 0.25 (95%); voriconazole, 1 (98.1%); A. nidulans, itraconazole, 1 (95%); posaconazole, 1 (97.7%); voriconazole, 2 (99.3%); A. niger, itraconazole, 2 (100%); posaconazole, 0.5 (96.9%); voriconazole, 2 (99.4%); A. terreus, itraconazole, 1 (100%); posaconazole, 0.5 (99.7%); voriconazole, 1 (99.1%); A. versicolor, itraconazole, 2 (100%); posaconazole, 1 (not applicable); voriconazole, 2 (97.5%). Although ECVs do not predict therapy outcome as clinical breakpoints do, they may aid in detection of azole resistance (non-WT MIC) due to cyp51A mutations, a resistance mechanism in some Aspergillus spp. These ECVs should be considered for inclusion in the future CLSI M38-A2 document revision. PMID:20592159

  5. Telavancin Demonstrates Activity against Methicillin-Resistant Staphylococcus aureus Isolates with Reduced Susceptibility to Vancomycin, Daptomycin, and Linezolid in Broth Microdilution MIC and One-Compartment Pharmacokinetic/Pharmacodynamic Models

    PubMed Central

    Smith, Jordan R.; Barber, Katie E.; Hallesy, Jessica; Raut, Animesh

    2015-01-01

    Methicillin-resistant Staphylococcus aureus (MRSA) isolates have arisen with reduced susceptibility to several anti-MRSA agents. Telavancin (TLV), a novel anti-MRSA agent, retains low MICs against these organisms. Our objective was to determine the MICs for TLV, daptomycin (DAP), vancomycin (VAN), and linezolid (LZD) against daptomycin-nonsusceptible (DNS) S. aureus, vancomycin-intermediate S. aureus (VISA), heteroresistant VISA (hVISA), and linezolid-resistant (LZDr) S. aureus. We also evaluated these agents against each phenotype in pharmacokinetic/pharmacodynamic (PK/PD) models. Seventy DNS, 100 VISA, 180 hVISA, and 25 LZDr MRSA isolates were randomly selected from our library and tested to determine their MICs against TLV, DAP, VAN, and LZD via broth microdilution and a Trek panel. Four isolates were randomly selected for 168-h in vitro models to evaluate treatment with TLV at 10 mg/kg of body weight/day, DAP at 10 mg/kg/day, VAN at 1 g every 12 h (q12h), and LZD at 600 mg q12h. The MIC50/90 for TLV, DAP, VAN, and LZD against 70 DNS S. aureus isolates were 0.06/0.125 μg/ml, 2/4 μg/ml, 1/2 μg/ml, and 2/2 μg/ml, respectively. Against 100 VISA isolates, the MIC50/90 were 0.06/0.125 μg/ml, 1/1 μg/ml, 4/8 μg/ml, and 1/2 μg/ml, respectively. Against 170 hVISA isolates, the MIC50/90 were 0.06/0.125 μg/ml, 0.5/1 μg/ml, 1/2 μg/ml, and 1/2 μg/ml, respectively. Against 25 LZDr isolates, the MIC50/90 were 0.03/0.06 μg/ml, 1/1 μg/ml, 2/2 μg/ml, and 8/8 μg/ml, respectively. The TLV MIC was >0.125 μg/ml for 10/365 (2.7%) isolates. In PK/PD models, TLV was universally bactericidal at 168 h and statistically superior to all antibiotics against DNS S. aureus strain R2334. These data further establish the potency of TLV against resistant MRSA. The model data demonstrate in vitro bactericidal activity of TLV against hVISA, VISA, DNS S. aureus, and LZDr S. aureus strains. Further clinical research is warranted. PMID:26124162

  6. Telavancin demonstrates activity against methicillin-resistant Staphylococcus aureus isolates with reduced susceptibility to vancomycin, daptomycin, and linezolid in broth microdilution MIC and one-compartment pharmacokinetic/pharmacodynamic models.

    PubMed

    Smith, Jordan R; Barber, Katie E; Hallesy, Jessica; Raut, Animesh; Rybak, Michael J

    2015-09-01

    Methicillin-resistant Staphylococcus aureus (MRSA) isolates have arisen with reduced susceptibility to several anti-MRSA agents. Telavancin (TLV), a novel anti-MRSA agent, retains low MICs against these organisms. Our objective was to determine the MICs for TLV, daptomycin (DAP), vancomycin (VAN), and linezolid (LZD) against daptomycin-nonsusceptible (DNS) S. aureus, vancomycin-intermediate S. aureus (VISA), heteroresistant VISA (hVISA), and linezolid-resistant (LZD(r)) S. aureus. We also evaluated these agents against each phenotype in pharmacokinetic/pharmacodynamic (PK/PD) models. Seventy DNS, 100 VISA, 180 hVISA, and 25 LZD(r) MRSA isolates were randomly selected from our library and tested to determine their MICs against TLV, DAP, VAN, and LZD via broth microdilution and a Trek panel. Four isolates were randomly selected for 168-h in vitro models to evaluate treatment with TLV at 10 mg/kg of body weight/day, DAP at 10 mg/kg/day, VAN at 1 g every 12 h (q12h), and LZD at 600 mg q12h. The MIC50/90 for TLV, DAP, VAN, and LZD against 70 DNS S. aureus isolates were 0.06/0.125 μg/ml, 2/4 μg/ml, 1/2 μg/ml, and 2/2 μg/ml, respectively. Against 100 VISA isolates, the MIC50/90 were 0.06/0.125 μg/ml, 1/1 μg/ml, 4/8 μg/ml, and 1/2 μg/ml, respectively. Against 170 hVISA isolates, the MIC50/90 were 0.06/0.125 μg/ml, 0.5/1 μg/ml, 1/2 μg/ml, and 1/2 μg/ml, respectively. Against 25 LZD(r) isolates, the MIC50/90 were 0.03/0.06 μg/ml, 1/1 μg/ml, 2/2 μg/ml, and 8/8 μg/ml, respectively. The TLV MIC was >0.125 μg/ml for 10/365 (2.7%) isolates. In PK/PD models, TLV was universally bactericidal at 168 h and statistically superior to all antibiotics against DNS S. aureus strain R2334. These data further establish the potency of TLV against resistant MRSA. The model data demonstrate in vitro bactericidal activity of TLV against hVISA, VISA, DNS S. aureus, and LZD(r) S. aureus strains. Further clinical research is warranted. Copyright © 2015, American Society for

  7. Comparison of microdilution method and E-test procedure in susceptibility testing of caspofungin against Candida non-albicans species.

    PubMed

    Serefko, Anna; Los, Renata; Biernasiuk, Anna; Malm, Anna

    2008-04-01

    It is widely known that systemic and mucosal candidiasis caused by Candida non-albicans strains endangers the lives of hospitalised patients since these pathogens are extremely difficult to defeat by commonly used antifungal agents. The present study determined the in vitro activities of a novel antimicotic drug - caspofungin - against 76 Candida non-albicans isolates by means of the CLSI reference method for broth dilution antifungal susceptibility testing of yeasts and the E-test procedure for comparison. Caspofungin was efficacious against the majority of strains tested, with the average MICs90 evaluated by the microdilution method and E-tests amounting to 1 mg/l and 0.5 mg/l, respectively. Since the agreement between MICs within +/-2 dilutions obtained by these two techniques was 92% (Kappa coefficient of 0.92), the E-test procedure seems to be a reliable alternative to the broth microdilution method and may provide another choice for clinical laboratories.

  8. Validation of 24-hour flucytosine MIC determination by comparison with 48-hour determination by the Clinical and Laboratory Standards Institute M27-A3 broth microdilution reference method.

    PubMed

    Lockhart, Shawn R; Bolden, Carol B; Iqbal, Naureen; Kuykendall, Randall J

    2011-12-01

    Flucytosine and itraconazole are the only antifungal agents for which the Clinical Laboratory and Standards Institute recommendations include MIC breakpoint readings at 48 h only. Here we show good essential and categorical agreement between the flucytosine MIC readings at 48 and 24 h for Candida species.

  9. [Comparative study of the susceptibility to daptomycin and other antimicrobials against Staphylococcus spp. resistant to methicillin and Enterococcus spp. using Wider, E-test, and microdilution methods].

    PubMed

    Gómez-Garcés, J L; López-Fabal, F; Burillo, A; Gil, Y

    2010-06-01

    The human and material costs of inappropriate antimicrobial therapy are high. This study was designed to search for a rapid, simple and effective antimicrobial susceptibility test capable of identifying the best treatment strategy against microorganisms causing hospital infections showing resistance or reduced susceptibility to the more traditional antibiotics. The tests compared were the E-test, an automated test (Wider) and broth microdilution ( as the reference test), to determine the susceptibility to vancomycin, teicoplanin, linezolid and daptomycin of clinical isolates of methicillin resistant Staphylococcus aureus (MRSA), methicillin resistant coagulase negative Staphylococcus spp. and Enterococccus spp. The E-test and Wider methods showed good agreement with the reference method indicating their reliability for routine susceptibility testing of staphylococci and enterococci against vancomycin, teicoplanin, linezolid and daptomycin. Notwithstanding, when faced with a serious enterococcal infection, the MIC of daptomycin should be more accurately determined using a reference technique such as broth microdilution.

  10. Correlation between E-test, disk diffusion, and microdilution methods for antifungal susceptibility testing of fluconazole and voriconazole.

    PubMed

    Matar, Madonna J; Ostrosky-Zeichner, Luis; Paetznick, Victor L; Rodriguez, Jose R; Chen, Enuo; Rex, John H

    2003-05-01

    The activities of fluconazole and voriconazole against isolates of Candida spp. (n = 400) were tested by the E-test, disk diffusion, and the National Committee for Clinical Laboratory Standards (NCCLS) M27-A2 broth microdilution-based reference methods. More than 96% of isolates found to be susceptible to fluconazole by the reference method were identified as susceptible by the agar-based methods. Lesser degrees of correlation with the reference method were seen for isolates identified as resistant by the agar-based methods. Interpretive categories are not available for voriconazole, but results qualitatively similar to those for fluconazole were seen. The agar-based E-test and disk diffusion methods are reliable alternatives to the NCCLS M27-A2 reference microdilution method for isolates that test susceptible to fluconazole.

  11. Multidrug-Resistant Candida auris Misidentified as Candida haemulonii: Characterization by Matrix-Assisted Laser Desorption Ionization–Time of Flight Mass Spectrometry and DNA Sequencing and Its Antifungal Susceptibility Profile Variability by Vitek 2, CLSI Broth Microdilution, and Etest Method

    PubMed Central

    Kathuria, Shallu; Singh, Pradeep K.; Sharma, Cheshta; Prakash, Anupam; Masih, Aradhana; Kumar, Anil

    2015-01-01

    Candida auris is a multidrug-resistant yeast that causes a wide spectrum of infections, especially in intensive care settings. We investigated C. auris prevalence among 102 clinical isolates previously identified as Candida haemulonii or Candida famata by the Vitek 2 system. Internal transcribed spacer region (ITS) sequencing confirmed 88.2% of the isolates as C. auris, and matrix-assisted laser desorption ionization–time of flight mass spectrometry (MALDI-TOF MS) easily separated all related species, viz., C. auris (n = 90), C. haemulonii (n = 6), C. haemulonii var. vulnera (n = 1), and Candida duobushaemulonii (n = 5). The in vitro antifungal susceptibility was determined using CLSI broth microdilution (CLSI-BMD), the Vitek 2 antifungal susceptibility test, and the Etest method. C. auris isolates revealed uniformly elevated fluconazole MICs (MIC50, 64 μg/ml), and an alarming percentage of isolates (37%) exhibited elevated caspofungin MICs by CLSI-BMD. Notably, 34% of C. auris isolates had coexisting elevated MICs (≥2 μg/ml) for both fluconazole and voriconazole, and 10% of the isolates had elevated coexisting MICs (≥2 μg/ml) to two additional azoles, i.e., posaconazole and isavuconazole. In contrast to reduced amphotericin B MICs by CLSI-BMD (MIC50, 1 μg/ml) for C. auris, elevated MICs were noted by Vitek 2 (MIC50, 8 μg/ml), which were statistically significant. Candida auris remains an unnoticed pathogen in routine microbiology laboratories, as 90% of the isolates characterized by commercial identification systems are misidentified as C. haemulonii. MALDI-TOF MS proved to be a more robust diagnostic technique for rapid identification of C. auris. Considering that misleading elevated MICs of amphotericin B by the Vitek AST-YS07 card may lead to the selection of inappropriate therapy, a cautionary approach is recommended for laboratories relying on commercial systems for identification and antifungal susceptibility testing of rare yeasts. PMID:25809970

  12. [In vitro susceptibility of isolates of Paracoccidioides spp complex to systemic antifungals using the microdilution method].

    PubMed

    Cermehol, Julman R; Alvarado, Primavera; Mendoza, Mireya; Herndndez, Isabel; Cuestal, De

    2015-09-01

    Broth microdilution, the reference method recommended by the Clinical Laboratory Standards Institute (CLSI), is not available for use with dimorphic fungi, such as those of the Paracoccidioides genus. In this work, in vitro susceptibility of the Paracoccidioides complex (n=19) to systemic antifungals: amphotericin B, 5-flucytosine, ketoconazole, itraconazole, fluconazole, voriconazole and caspofungin, was evaluated using the microdilution method (Document M27-A3, M27-S3), with some modifications such as: culture time in Sabouraud dextrose agar (7-10 days), RPMI 1640 medium supplemented with 2% glucose and the incubation time (7, 8 and 18 days). The sensitivity in vitro was variable; the majority of Paracoccidioides isolates was susceptible to ketoconazol (73.7%), followed by voriconazole (68.4%), itraconazole (63.1%), amphotericin B (52.6%), fluconazole (47.4%), 5-flucytosine (42.1%) and caspofungin (5%). The overall resistance was mainly to caspofungin (94.7%), followed by 5-flucytosine (52.6%) and amphotericin B (47.4%). Fifty-three percent of the isolates were susceptible-dose dependent to fluconazole followed by itraconazole (15.7%) and 5-fluorocytosine (5.3%). Amphotericin B, itraconazole and voriconazole were the most potent antifungal drugs against Paracoccidioides spp (CMI: 0.03-1 microg/mL). Based on these results, we tentatively propose a microdilution assay protocol for susceptibility testing of Paracoccidioides spp to antifungal drugs. This method may be clinically useful to predict resistance, even though further studies are needed.

  13. Multidrug-Resistant Candida auris Misidentified as Candida haemulonii: Characterization by Matrix-Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry and DNA Sequencing and Its Antifungal Susceptibility Profile Variability by Vitek 2, CLSI Broth Microdilution, and Etest Method.

    PubMed

    Kathuria, Shallu; Singh, Pradeep K; Sharma, Cheshta; Prakash, Anupam; Masih, Aradhana; Kumar, Anil; Meis, Jacques F; Chowdhary, Anuradha

    2015-06-01

    Candida auris is a multidrug-resistant yeast that causes a wide spectrum of infections, especially in intensive care settings. We investigated C. auris prevalence among 102 clinical isolates previously identified as Candida haemulonii or Candida famata by the Vitek 2 system. Internal transcribed spacer region (ITS) sequencing confirmed 88.2% of the isolates as C. auris, and matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) easily separated all related species, viz., C. auris (n = 90), C. haemulonii (n = 6), C. haemulonii var. vulnera (n = 1), and Candida duobushaemulonii (n = 5). The in vitro antifungal susceptibility was determined using CLSI broth microdilution (CLSI-BMD), the Vitek 2 antifungal susceptibility test, and the Etest method. C. auris isolates revealed uniformly elevated fluconazole MICs (MIC50, 64 μg/ml), and an alarming percentage of isolates (37%) exhibited elevated caspofungin MICs by CLSI-BMD. Notably, 34% of C. auris isolates had coexisting elevated MICs (≥2 μg/ml) for both fluconazole and voriconazole, and 10% of the isolates had elevated coexisting MICs (≥2 μg/ml) to two additional azoles, i.e., posaconazole and isavuconazole. In contrast to reduced amphotericin B MICs by CLSI-BMD (MIC50, 1 μg/ml) for C. auris, elevated MICs were noted by Vitek 2 (MIC50, 8 μg/ml), which were statistically significant. Candida auris remains an unnoticed pathogen in routine microbiology laboratories, as 90% of the isolates characterized by commercial identification systems are misidentified as C. haemulonii. MALDI-TOF MS proved to be a more robust diagnostic technique for rapid identification of C. auris. Considering that misleading elevated MICs of amphotericin B by the Vitek AST-YS07 card may lead to the selection of inappropriate therapy, a cautionary approach is recommended for laboratories relying on commercial systems for identification and antifungal susceptibility testing of rare yeasts. Copyright

  14. Genetic profiles of Shiga toxin and intimin genes found in stool broth cultures: a 2-year reference laboratory study.

    PubMed

    Michel, Pierre A; Kase, Julie A

    2009-10-01

    Shiga toxin (Stx)-producing Escherichia coli (STEC) are associated with potentially serious illness in humans. STEC detection is often based on the presence of Stxs, Stx(1) and/or Stx(2), and intimin, encoded by the eae gene. A 2-year collection of stool broth cultures was tested for variants of stx(1), stx(2), and eae. Approximately 80% (138 of 174) were positive for stx(1) and/or stx(2), with stx(1) as the most prevalent (66%). Of the stx(1) variants, stx(1) was the most common (76%) followed by stx(1c) (22%). Analysis of stx(2)-positive isolates found 20 (53%) stx(2), 13 (34%) stx(2)/stx(2v-ha), 3 (8%) stx(2v-ha), 1 (3%) stx(2v-hb), and 1 (3%) stx(2d-activatable). Findings of stx(2)/stx(2v-ha) and stx(2d-activatable) are noteworthy given associations with hemolytic uremic syndrome and increased cytotoxicity, respectively. Of the Stx positive, 94 (68%) were eae positive with 31 (33%) eae(varepsilon1), 19 (20%) eae(gamma1), and 18 (19%) eae(beta1). A predominance of eae(varepsilon1) may suggest a new pathogenic significance because, reportedly, eae(beta1) is one of the most widespread variants.

  15. [Helicobacter pylori antibiotic sensitivity by microdilution].

    PubMed

    Rivas, F; Rivera, P; Hernández, F; Hevia, F; Guillén, F; Tamayo, G

    2000-01-01

    The gastric pathogen Helicobacter pylori has been recognized as the major aetiologic agent of chronic gastritis and peptic ulcers and also a risk factor for gastric cancer; eradication of H pylori prevents peptic ulcer recurrence and may also decrease the prevalence of gastric cancer in high risk populations around the world. Currently the only accepted indication for treatment is ulcer disease and maltosa, infected with Helicobacter pilory. However treatment is difficult and easily develops resistance. The elaboration of an antibiotic profile is recommended after a treatment failure. There is a lack of information in developing countries so the aim of this work was to determine the antibiotic profile of 51 strains isolated from patients gastric biopsies attended at Hospital San Juan de Dios in Costa Rica, using egg yolk broth and finding a resistance of 63.0% to metronidazole with a breakpoint of 8.0 microg/ml and 20.0% resistance to tetracycline (MIC1.0 microg/ml), 6.0% to clarithromicyn with a MIC of 0.125 microg/ml. There was no resistance to amoxicilin (MIC 0.015 microg/ml). The microdilution technique is very laborious, but highly reproducible with results accordingly to previous work, and we recommended it for the designing of therapeutical scheme.

  16. Comparative Evaluation of Disk Diffusion with Microdilution Assay in Susceptibility Testing of Caspofungin against Aspergillus and Fusarium Isolates

    PubMed Central

    Arikan, Sevtap; Paetznick, Victor; Rex, John H.

    2002-01-01

    We compared the disk diffusion and broth microdilution methods for susceptibility testing of caspofungin against Aspergillus (n = 78) and Fusarium (n = 22) isolates. Microdilution testing followed the NCCLS M-38P guidelines but was performed in antibiotic medium 3 supplemented to 2% glucose (AM3). Disk diffusion assays were performed on AM3 agar plates with a 2-μg caspofungin disk. By both methods, caspofungin showed favorable activity against Aspergillus isolates and no activity against Fusarium isolates. In the disk-based format, intrazonal growth that was not influenced by the drug concentration gradient was consistently observed for all of the Aspergillus isolates tested. PMID:12183278

  17. Screening of new bioactive materials from microbial extracts of soil microorganism (I). Antimicrobial activity from 200 samples using microdilution assay.

    PubMed

    Jung, S O; Kim, J; Chang, I M; Ryu, J C

    1998-06-01

    The microdilution assay recommended by NCCLS (National Committee for Clinical Laboratory Standards) is one of the standardized methods of antibiotic susceptibility test. This method has been widely used clinically to obtain MIC values of antibiotics on pathogenic microorganisms. It is more convenient, rapid and simple to test many samples than other test methods such as agar diffusion assay and broth macrodilution assay. The screening of antimicrobial agents from microbial extracts is too laborious in its process. Therefore, a number of screening methods having more simple procedure have been developed. In our laboratory, we applied microdilution assay for screening the antimicrobial agents. This assay showed dose-response results and was more sensitive than disc diffusion assay in our system. We tested 200 samples of microbial extracts originated from 100 microbial strains and selected several samples as potential candidates. In this report, we show that the microdilution assay is more convenient method in screening of antibiotic susceptibility than those previously reported.

  18. A new rapid resazurin-based microdilution assay for antimicrobial susceptibility testing of Neisseria gonorrhoeae.

    PubMed

    Foerster, Sunniva; Desilvestro, Valentino; Hathaway, Lucy J; Althaus, Christian L; Unemo, Magnus

    2017-07-01

    Rapid, cost-effective and objective methods for antimicrobial susceptibility testing of Neisseria gonorrhoeae would greatly enhance surveillance of antimicrobial resistance. Etest, disc diffusion and agar dilution methods are subjective, mostly laborious for large-scale testing and take ∼24 h. We aimed to develop a rapid broth microdilution assay using resazurin (blue), which is converted into resorufin (pink fluorescence) in the presence of viable bacteria. The resazurin-based broth microdilution assay was established using 132 N. gonorrhoeae strains and the antimicrobials ceftriaxone, cefixime, azithromycin, spectinomycin, ciprofloxacin, tetracycline and penicillin. A regression model was used to estimate the MICs. Assay results were obtained in ∼7.5 h. The EC 50 of the dose-response curves correlated well with Etest MIC values (Pearson's r  = 0.93). Minor errors resulting from misclassifications of intermediate strains were found for 9% of the samples. Major errors (susceptible strains misclassified as resistant) occurred for ceftriaxone (4.6%), cefixime (3.3%), azithromycin (0.6%) and tetracycline (0.2%). Only one very major error was found (a ceftriaxone-resistant strain misclassified as susceptible). Overall the sensitivity of the assay was 97.1% (95% CI 95.2-98.4) and the specificity 78.5% (95% CI 74.5-82.9). A rapid, objective, high-throughput, quantitative and cost-effective broth microdilution assay was established for gonococci. For use in routine diagnostics without confirmatory testing, the specificity might remain suboptimal for ceftriaxone and cefixime. However, the assay is an effective low-cost method to evaluate novel antimicrobials and for high-throughput screening, and expands the currently available methodologies for surveillance of antimicrobial resistance in gonococci.

  19. In vitro evaluation of Malassezia pachydermatis susceptibility to azole compounds using E-test and CLSI microdilution methods.

    PubMed

    Cafarchia, Claudia; Figueredo, Luciana A; Iatta, Roberta; Colao, Valeriana; Montagna, Maria T; Otranto, Domenico

    2012-11-01

    Dermatitis caused by Malassezia spp., one of most common skin disease in dogs, requires prolonged therapy and/or high doses of antifungal agents. In the present study, the antifungal susceptibility of M. pachydermatis to ketoconazole (KTZ), fluconazole (FLZ), itraconazole (ITZ), posaconazole (POS) and voriconazole (VOR) was evaluated in vitro using both CLSI reference broth microdilution (CLSI BMD) and E-test. A total of 62 M. pachydermatis strains from dogs with and without skin lesions were tested. M. pachydermatis strains were susceptible to ITZ, KTZ and POS using both test methods, with the highest MIC found in tests of FLZ. Essential agreement between the two methods ranged from 87.1% (VOR) to 91.9% (ITZ), and categorical agreement from 74.2% (FLZ) to 96.8% (ITZ). Minor error discrepancies were observed between the two methods, with major discrepancies observed for KTZ. A higher MIC(50) value for FLZ was noted with M. pachydermatis genotype B. The MICs(50) of M. pachydermatis genotype B for KTZ, VOR and POS were higher in isolates from dogs with skin lesions than those in isolates from animals without skin lesions. The results suggest a link between genotypes of M. pachydermatis and in vitro drug susceptibility. The categorical agreement for both E-test and CLSI BMD methods found in this investigation confirms the E-test as a reliable diagnostic method for routine use in clinical mycology laboratories.

  20. In-vitro Activity of 10 Antifungal Agents against 320 Dermatophyte Strains Using Microdilution Method in Tehran.

    PubMed

    Adimi, Parvaneh; Hashemi, Seyed Jamal; Mahmoudi, Mahmood; Mirhendi, Hossein; Shidfar, Mohammad Reza; Emmami, Masood; Rezaei-Matehkolaei, Ali; Gramishoar, Mohsen; Kordbacheh, Parivash

    2013-01-01

    Dermatophyte fungi are the etiologic agents of skin infections commonly referred to as ringworm. These infections are not dangerous but as a chronic cutaneous infections they may be difficult to treat and can also cause physical discomfort for patients. They are considered important as a public health problem as well. No information is available regarding the efficacy of antifungal agents against dermatophytes in Tehran. Therefore, in this study we evaluated the efficacy of 10 systemic and topical antifungal medications using CLSI broth microdilution method (M38-A). The antifungal agents used included griseofulvin, terbinafine, itraconazole, ketoconazole, fluconazole, voriconazole, clotrimazole, ciclopirox olamine, amorolfine and naftifine.Fifteen different species of dermatophytes which were mostly clinical isolates were used as follows; T. mentagrophytes, T. rubrum, E. floccosum, M. canis, T. verrucosum, T. tonsurans, M. gypseum, T. violaceum, M. ferruginum, M. fulvum, T. schoenleinii, M. racemosum, T. erinacei, T. eriotrephon and Arthroderma benhamiae. The mean number of fungi particles (conidia) inoculated was 1.25 ×10⁴ CFU/mL. Results were read after 7 days of incubation at 28 °C. According to the obtained results,itraconazole and terbinafine showed the lowest and fluconazole had the greatest MIC values for the most fungi tested. Based on the results, it is necessary to do more research and design a reliable standard method for determination of antifungal susceptibility to choose proper antibiotics with fewer side effects and decrease antifungal resistance and risk of treatment failure.

  1. In-vitro Activity of 10 Antifungal Agents against 320 Dermatophyte Strains Using Microdilution Method in Tehran

    PubMed Central

    Adimi, Parvaneh; Hashemi, Seyed Jamal; Mahmoudi, Mahmood; Mirhendi, Hossein; Shidfar, Mohammad Reza; Emmami, Masood; Rezaei-Matehkolaei, Ali; Gramishoar, Mohsen; Kordbacheh, Parivash

    2013-01-01

    Dermatophyte fungi are the etiologic agents of skin infections commonly referred to as ringworm. These infections are not dangerous but as a chronic cutaneous infections they may be difficult to treat and can also cause physical discomfort for patients. They are considered important as a public health problem as well. No information is available regarding the efficacy of antifungal agents against dermatophytes in Tehran. Therefore, in this study we evaluated the efficacy of 10 systemic and topical antifungal medications using CLSI broth microdilution method (M38-A). The antifungal agents used included griseofulvin, terbinafine, itraconazole, ketoconazole, fluconazole, voriconazole, clotrimazole, ciclopirox olamine, amorolfine and naftifine.Fifteen different species of dermatophytes which were mostly clinical isolates were used as follows; T. mentagrophytes, T. rubrum, E. floccosum, M. canis, T. verrucosum, T. tonsurans, M. gypseum, T. violaceum, M. ferruginum, M. fulvum, T. schoenleinii, M. racemosum, T. erinacei, T. eriotrephon and Arthroderma benhamiae. The mean number of fungi particles (conidia) inoculated was 1.25 ×10⁴ CFU/mL. Results were read after 7 days of incubation at 28 °C. According to the obtained results,itraconazole and terbinafine showed the lowest and fluconazole had the greatest MIC values for the most fungi tested. Based on the results, it is necessary to do more research and design a reliable standard method for determination of antifungal susceptibility to choose proper antibiotics with fewer side effects and decrease antifungal resistance and risk of treatment failure. PMID:24250660

  2. Microbiological culture broth designed from food waste.

    PubMed

    Chalón, Miriam C; Terán, Victoria; Arena, Mario E; Oliszewki, Rubén; González, Silvia N

    2013-01-30

    The current trend of increasing air, water, and soil pollution is, in part, due to inadequate management of municipal solid waste (MSW). The relationship between public health and the collection, storage and improper disposal of solid waste has encouraged several studies and the results were attributed to the spread of over twenty human and animal diseases due to this interrelationship. The term "single cell protein" (SCP) refers to microbial biomass used as a dietary additive. It has high nutritional value because of its high content of vitamins, lipids, and proteins of biological quality (the presence of all essential amino acids) (Lal, 2005). The aim of this work was to design a culture media for microbiological assays and to produce SCP for animal feeding, using nutrients contained in organic waste. In order to compare the effectiveness of food waste (FW) and LAPTg media, different strains of Lactobacillus, Enterococcus, Staphylococcus, Shigella, Salmonella, Saccharomyces and Schizosaccharomyces were studied. In all cases, the growth obtained from FW and LAPTg culture media were not significantly different (p > 0.05). In addition, the growth of Saccharomyces cerevisiae was studied in order to produce SCP for animal feeding. Comparative experiments involving molasses broth, FW broth, and basal broth were carried out. The biomass yield calculated at 24 h from FW broth was 13% lower than from molasses broth. The FW broth provided a significantly lower biomass yield; however, it can be very useful in areas where molasses are not available. FW broth can be elaborated at low cost, in any populated region of the world because its ingredients are wastes generated by humans. It has great versatility, allowing the development of a wide variety of microorganisms, both Gram negative and Gram positive bacteria as well as yeasts. The production of safe protein additives, with high biological quality and low cost, is necessary due to the increasing global demand for food

  3. [Comparison of microdilution and disk diffusion methods for the detection of fluconazole and voriconazole susceptibility against clinical Candida glabrata isolates and determination of changing susceptibility with new CLSI breakpoints].

    PubMed

    Hazırolan, Gülşen; Sarıbaş, Zeynep; Arıkan Akdağlı, Sevtap

    2016-07-01

    Candida albicans is the most frequently isolated species as the causative agent of Candida infections. However, in recent years, the isolation rate of non-albicans Candida species have increased. In many centers, Candida glabrata is one of the commonly isolated non-albicans species of C.glabrata infections which are difficult-to-treat due to decreased susceptibility to fluconazole and cross-resistance to other azoles. The aims of this study were to determine the in vitro susceptibility profiles of clinical C.glabrata isolates against fluconazole and voriconazole by microdilution and disk diffusion methods and to evaluate the results with both the previous (CLSI) and current species-specific CLSI (Clinical and Laboratory Standards Institute) clinical breakpoints. A total of 70 C.glabrata strains isolated from clinical samples were included in the study. The identification of the isolates was performed by morphologic examination on cornmeal Tween 80 agar and assimilation profiles obtained by using ID32C (BioMérieux, France). Broth microdilution and disk diffusion methods were performed according to CLSI M27-A3 and CLSI M44-A2 documents, respectively. The results were evaluated according to CLSI M27-A3 and M44-A2 documents and new vs. species-specific CLSI breakpoints. By using both previous and new CLSI breakpoints, broth microdilution test results showed that voriconazole has greater in vitro activity than fluconazole against C.glabrata isolates. For the two drugs tested, very major error was not observed with disk diffusion method when microdilution method was considered as the reference method. Since "susceptible" category no more exists for fluconazole vs. C.glabrata, the isolates that were interpreted as susceptible by previous breakpoints were evaluated as susceptible-dose dependent by current CLSI breakpoints. Since species-specific breakpoints remain yet undetermined for voriconazole, comparative analysis was not possible for this agent. The results obtained

  4. Activity of retapamulin against Streptococcus pyogenes and Staphylococcus aureus evaluated by agar dilution, microdilution, E-test, and disk diffusion methodologies.

    PubMed

    Pankuch, Glenn A; Lin, Gengrong; Hoellman, Dianne B; Good, Caryn E; Jacobs, Michael R; Appelbaum, Peter C

    2006-05-01

    The in vitro activity of retapamulin against 106 Staphylococcus aureus isolates and 109 Streptococcus pyogenes isolates was evaluated by the agar dilution, broth microdilution, E-test, and disk diffusion methodologies. Where possible, the tests were performed by using the CLSI methodology. The results of agar dilution, broth microdilution, and E-test (all with incubation in ambient air) for S. aureus yielded similar MICs, in the range of 0.03 to 0.25 microg/ml. These values corresponded to zone diameters between 25 and 33 mm by the use of a 2-microg retapamulin disk. Overall, 99% of the agar dilution results and 95% of E-test results for S. aureus were within +/-1 dilution of the microdilution results. For S. pyogenes, the MICs obtained by the agar and broth microdilution methods (both after incubation in ambient air) were in the range of 0.008 to 0.03 microg/ml, and E-test MICs (with incubation in ambient air) were 0.016 to 0.06 microg/ml. For S. pyogenes, 100% of the agar dilution MIC results were within +/-1 dilution of the broth microdilution results. E-test MICs (after incubation in ambient air) were within +/-1 and +/-2 dilutions of the broth microdilution results for 76% and 99% of the isolates, respectively. E-test MICs for S. pyogenes strains in CO(2) were up to 4 dilutions higher than those in ambient air. Therefore, it is recommended that when retapamulin MICs are determined by E-test, incubation be done in ambient air and not in CO(2), due to the adverse effect of CO(2) on the activity of this compound. Diffusion zones (with incubation in CO(2)) for S. pyogenes were 18 to 24 mm. Retapamulin MICs for all strains by all methods (with incubation in ambient air) were < or =0.25 microg/ml. These results demonstrate that S. pyogenes (including macrolide-resistant strains) and S. aureus (including methicillin-resistant and vancomycin-nonsusceptible strains) are inhibited by very low concentrations of retapamulin and that all four testing methods are

  5. Activity of Retapamulin against Streptococcus pyogenes and Staphylococcus aureus Evaluated by Agar Dilution, Microdilution, E-Test, and Disk Diffusion Methodologies

    PubMed Central

    Pankuch, Glenn A.; Lin, Gengrong; Hoellman, Dianne B.; Good, Caryn E.; Jacobs, Michael R.; Appelbaum, Peter C.

    2006-01-01

    The in vitro activity of retapamulin against 106 Staphylococcus aureus isolates and 109 Streptococcus pyogenes isolates was evaluated by the agar dilution, broth microdilution, E-test, and disk diffusion methodologies. Where possible, the tests were performed by using the CLSI methodology. The results of agar dilution, broth microdilution, and E-test (all with incubation in ambient air) for S. aureus yielded similar MICs, in the range of 0.03 to 0.25 μg/ml. These values corresponded to zone diameters between 25 and 33 mm by the use of a 2-μg retapamulin disk. Overall, 99% of the agar dilution results and 95% of E-test results for S. aureus were within ±1 dilution of the microdilution results. For S. pyogenes, the MICs obtained by the agar and broth microdilution methods (both after incubation in ambient air) were in the range of 0.008 to 0.03 μg/ml, and E-test MICs (with incubation in ambient air) were 0.016 to 0.06 μg/ml. For S. pyogenes, 100% of the agar dilution MIC results were within ±1 dilution of the broth microdilution results. E-test MICs (after incubation in ambient air) were within ±1 and ±2 dilutions of the broth microdilution results for 76% and 99% of the isolates, respectively. E-test MICs for S. pyogenes strains in CO2 were up to 4 dilutions higher than those in ambient air. Therefore, it is recommended that when retapamulin MICs are determined by E-test, incubation be done in ambient air and not in CO2, due to the adverse effect of CO2 on the activity of this compound. Diffusion zones (with incubation in CO2) for S. pyogenes were 18 to 24 mm. Retapamulin MICs for all strains by all methods (with incubation in ambient air) were ≤0.25 μg/ml. These results demonstrate that S. pyogenes (including macrolide-resistant strains) and S. aureus (including methicillin-resistant and vancomycin-nonsusceptible strains) are inhibited by very low concentrations of retapamulin and that all four testing methods are satisfactory for use for susceptibility

  6. Clinical evaluation of the ASTY colorimetric microdilution panel for antifungal susceptibility testing.

    PubMed

    Pfaller, M A; Arikan, S; Lozano-Chiu, M; Chen, Y; Coffman, S; Messer, S A; Rennie, R; Sand, C; Heffner, T; Rex, J H; Wang, J; Yamane, N

    1998-09-01

    A method using a commercially prepared colorimetric microdilution panel (ASTY; Kyokuto Pharmaceutical Industrial Co., Ltd.) was compared in four different laboratories with the National Committee for Clinical Laboratory Standards (NCCLS) reference microdilution method by testing 802 clinical isolates of Candida spp. (C. albicans, C. glabrata, C. tropicalis, C. parapsilosis, C. krusei, C. lusitaniae, C. guilliermondii, C. lipolytica, C. rugosa, and C. zeylanoides) against amphotericin B, 5-fluorocytosine (5FC), fluconazole, and itraconazole. Reference MIC endpoints were established after 48 h of incubation, and ASTY endpoints were established after 24 and 48 h of incubation. ASTY endpoints were determined to be the time at which the color of the first well changed from red (indicating growth) to purple (indicating growth inhibition) or blue (indicating no growth). Excellent agreement (within 2 dilutions) between the reference and colorimetric MICs was observed. Overall agreement was 93% at 24 h and 96% at 48 h. Agreement ranged from 90% with itraconazole and 5FC to 96% with amphotericin B at 24 h and from 92% with itraconazole to 99% with amphotericin B and 5FC at 48 h. The ASTY colorimetric microdilution panel method appears to be comparable to the NCCLS reference method for testing the susceptibilities of Candida spp. to a variety of antifungal agents.

  7. [In vitro antifungal activity of azoles and amphotericin B against Malassezia furfur by the CLSI M27-A3 microdilution and Etest(®) methods].

    PubMed

    Galvis-Marín, Juan Camilo; Rodríguez-Bocanegra, María Ximena; Pulido-Villamarín, Adriana Del Pilar; Castañeda-Salazar, Rubiela; Celis-Ramírez, Adriana Marcela; Linares-Linares, Melva Yomary

    Malassezia furfur is a human skin commensal yeast that can cause skin and opportunistic systemic infections. Given its lipid dependant status, the reference methods established by the Clinical and Laboratory Standards Institute (CLSI) to evaluate antifungal susceptibility in yeasts are not applicable. To evaluate the in vitro susceptibility of M. furfur isolates from infections in humans to antifungals of clinical use. The susceptibility profile to amphotericin B, itraconazole, ketoconazole and voriconazole of 20 isolates of M. furfur, using the broth microdilution method (CLSI M27-A3) and Etest(®), was evaluated. Itraconazole and voriconazole had the highest antifungal activity against the isolates tested. The essential agreement between the two methods for azoles antifungal activity was in the region of 60-85% and the categorical agreement was around 70-80%, while the essential and categorical agreement for amphotericin B was 10%. The azoles were the compounds that showed the highest antifungal activity against M. furfur, as determined by the two techniques used; however more studies need to be performed to support that Etest(®) is a reliable method before its implementation as a routine clinical laboratory test. Copyright © 2016 Asociación Española de Micología. Publicado por Elsevier España, S.L.U. All rights reserved.

  8. Reliability of high-content disks and modified broth dilution tests for detecting staphylococcal resistance to the penicillinase-resistant penicillins.

    PubMed Central

    Barry, A L; Jones, R N

    1987-01-01

    In vitro susceptibility tests were performed with 271 isolates of Staphylococcus species (204 Staphylococcus aureus), including 110 strains resistant to the penicillinase-resistant penicillins. Disks containing 5 or 10 micrograms of methicillin, 1 or 4 micrograms of oxacillin, and 1 or 4 micrograms of nafcillin were evaluated. After a full 24 h of incubation at 35 degrees C, tests with 1-microgram oxacillin disks provided optimal results. Use of the more potent oxacillin, nafcillin, or methicillin disks only increased the number of false-susceptible test results. For broth microdilution tests, 2% NaCl should be added to cation-supplemented Mueller-Hinton broth, and MICs should be recorded after a full 24 h at 35 degrees C. Microdilution tests with oxacillin in broth with 2% NaCl were more reliable than similar tests with methicillin. PMID:3499451

  9. The agar microdilution method - a new method for antimicrobial susceptibility testing for essential oils and plant extracts.

    PubMed

    Golus, J; Sawicki, R; Widelski, J; Ginalska, G

    2016-11-01

    To develop a new agar microdilution technique suitable for the assessment of the antimicrobial activity of natural plant products such as essential oils or plant extracts as well as to evaluate the antimicrobial effect of several essential oils and plant extracts. The proposed agar microdilution method was evolved on the basis of the CLSI agar dilution method, approved for conventional antimicrobials. However, this new method combines convenience and time/cost effectiveness typical for microtitre methods with the advantages of the agar dilution of hydrophobic or coloured substances. Different concentrations of the tested agents were added to Eppendorf tubes with molten Mueller-Hinton agar, vortexed and dispensed into the 96-well microplate in a small volume of 100 μl per well which allows for rapid, easy and economical preparation of samples as well as providing a uniform and stable dispersion without separation of the oil-water phases which occurs in methods with liquid medium. Next, the agar microdilution plates were inoculated with four reference bacterial strains. The results of our study demonstrated that the minimal inhibitory concentrations (MICs) were successfully determined using the agar microdilution method even with hydrophobic essential oils or strongly coloured plant extracts. The new agar microdilution method avoids the problems associated with testing of water insoluble, oily or strongly coloured plant natural products. Moreover, it enables the reliable, cheap and easy MIC determination of such agents. In the era of increasing antibiotic resistance high hopes are associated with new drugs of plant origin. However, the lack of standardized and reliable testing methods for assessing antibacterial activity of plant natural products causes impediment to research into this area. This study demonstrated that the agar microdilution method can be used successfully for testing oily and coloured substances. © 2016 The Society for Applied Microbiology.

  10. Comparison of the Sensititre YeastOne® dilution method with the Clinical Laboratory Standards Institute (CLSI) M27-A3 microbroth dilution reference method for determining MIC of eight antifungal agents on 102 yeast strains.

    PubMed

    Bertout, S; Dunyach, C; Drakulovski, P; Reynes, J; Mallié, M

    2011-02-01

    The Clinical Laboratory Standards Institute ([CLSI] formerly NCCLS) reference broth microdilution testing method (protocol M27-A3) was compared with a commercially available methods (Sensititre YeastOne(®)) by testing two quality control strains and 102 isolates of Candida sp. and Cryptococcus sp. against fluconazole, itraconazole, ketoconazole, posaconazole, voriconazole, flucytosin, amphotericin B and caspofungin. Minimal inhibitory concentrations (MIC) endpoints were determined after 24h of incubation for Sensititre YeastOne(®) and after 24 and 48 h for CLSI microdilution method. Essential agreements between methods vary from 70.6 to 92.2%. Categorical agreements vary from 94.1% for 5FC to 72.6% for AMB. Sensititre YeastOne(®) reading appears to be useful for avoiding very major errors and this confirms the interest of this method for evaluating new antifungals activity in vitro.

  11. [Comparison of microdilution method and Phoenix automated system for testing antimicrobial susceptibilities of Enterococcus strains].

    PubMed

    Gülmez, Dolunay; Hasçelik, Gülşen

    2011-01-01

    Enterococcus spp. are important pathogens which are intrinsically resistant to most of the commonly used antimicrobial agents such as aminoglycosides and cephalosporins. Accurate determination of resistance is important to ensure appropriate antimicrobial therapy. This study was undertaken to compare the susceptibility results obtained by Phoenix system (Becton Dickinson, USA) with reference microdilution method. We included 1248 Enterococcus spp. (903 Enterococcus faecalis, 345 Enterococcus faecium) strains isolated from clinical samples between 2005-2007 in routine microbiology laboratory of Hacettepe University Hospital. The strains were identified and the antimicrobial susceptibilities were determined by the Phoenix system. Antimicrobial susceptibilities to ampicillin, teicoplanin, vancomycin, gentamicin and streptomycin were also studied by microdilution method according to Clinical and Laboratory Standards Institute (CLSI) guidelines. Evaluation revealed excellent agreement for all of the antibiotics with category agreement rates of > 97%. Among 1248 strains, 76 revealed discordant results. Very major error rates were 1.5% for ampicillin, 1.3% for gentamicin, and 0.9% for streptomycin. Major error rates were 1.4% for streptomycin, 0.6% for ampicillin and vancomycin and 0.3% for gentamicin. Minor error rates were found as 0.2% for vancomycin, and 0.1% for teicoplanin. Resistance rates obtained by microdilution were as follows; high level streptomycin 44%, high level gentamicin 29.7%, ampicillin 25.6%, vancomycin 2.2% and teicoplanin 2.2%. Resistance rates were higher in E.faecium than E.faecalis and 96.4% of the vancomycin resistant enterococcus isolates were identified as E.faecium. In conclusion, based on the data obtained, Phoenix system is reliable for testing susceptibilities of Enterococcus spp. to these antimicrobials. Since isolation of vancomycin resistant enterococci has an important impact in terms of hospital infection control, vancomycin

  12. Antimicrobial susceptibility and MIC distribution of 41 drugs against clinical isolates from China and reference strains of nontuberculous mycobacteria.

    PubMed

    Li, Guilian; Pang, Hui; Guo, Qian; Huang, Mingxiang; Tan, Yanhong; Li, Chao; Wei, Jianhao; Xia, Yuanzhi; Jiang, Yi; Zhao, Xiuqin; Liu, Haican; Zhao, Li-Li; Liu, Zhiguang; Xu, Donglei; Wan, Kanglin

    2017-03-01

    To treat nontuberculous mycobacteria (NTM) infections more optimally, further research pertaining to mycobacterial susceptibility to antimicrobial agents is required. A total of 82 species of NTM reference strains and 23 species of NTM clinical isolates were included. Minimum inhibitory concentrations (MICs) for 41 drugs were determined using the microdilution method in cation-adjusted Mueller-Hinton broth. The results showed that most of the NTM were susceptible to aminoglycosides, quinolones, three macrolides (clarithromycin, azithromycin and roxithromycin), cefmetazole, linezolid and capreomycin. Rapidly growing mycobacterium strains were additionally susceptible to cefoxitin, clofazimine, rifapentine, doxycycline, minocycline, tigecycline, meropenem and sulfamethoxazole, whereas slowly growing mycobacterium strains were additionally susceptible to rifabutin. This study on the susceptibility of NTM includes the largest sample size of Chinese clinical isolates and reference strains. NTM species-specific drug susceptibility patterns suggested that it is urgent to identify the species of NTM, to normalise the treatment of NTM infectious disease and to clarify the resistance mechanisms of NTM.

  13. Inactivation of Penicillins by Thiol Broth

    PubMed Central

    Murray, Patrick R.; Niles, Ann C.

    1982-01-01

    Thiol broth with sodium polyanetholesulfonate inactivated penicillin G, carbenicillin, nafcillin, oxacillin, and gentamicin, but had no effect on cephalothin, cefoxitin, clindamycin, chloramphenicol, erythromycin, and tetracycline. Only Thiol broth was capable of this inactivation, which was not influenced by the presence of blood. PMID:7153352

  14. Ultrafiltration of hemicellulose hydrolysate fermentation broth

    NASA Astrophysics Data System (ADS)

    Kresnowati, M. T. A. P.; Desiriani, Ria; Wenten, I. G.

    2017-03-01

    Hemicelulosic material is often used as the main substrate to obtain high-value products such as xylose. The five carbon sugar, xylose, could be further processed by fermentation to produce xylitol. However, not only the hemicellulose hydrolysate fermentation broth contains xylitol, but also metabolite products, residual substances, biomass and mineral salts. Therefore, in order to obtain the end products, various separation processes are required to separate and purify the desired product from the fermentation broth. One of the most promising downstream processing methods of fermentation broth clarification is ultrafiltration due to its potential for energy saving and higher purity. In addition, ultrafiltration membrane has a high performance in separating inhibitory components in the fermentation broth. This paper assesses the influence of operating conditions; including trans-membrane pressure, velocity, pH of the fermentation broth solutions, and also to the xylitol concentration in the product. The challenges of the ultrafiltration process will be pointed out.

  15. Hypertonic sabouraud broth as a simple and powerful test for Candida dubliniensis screening.

    PubMed

    Alves, Sydney Hartz; Milan, Eveline Pipolo; de Laet Sant'Ana, Priscilla; Oliveira, Loiva O; Santurio, Janio M; Colombo, Arnaldo Lopes

    2002-05-01

    We developed a new screening test for C. dubliniensis based on its inability to grow on Sabouraud dextrose broth with 6.5% NaCl. A total of 266 clinical yeast isolates and 3 reference strains were tested, including 250 C. albicans and 19 C. dubliniensis strains. All C. albicans isolates tested exhibited significant growth on hypertonic Sabouraud broth up to 96 h, while, all C. dubliniensis isolates did not exhibit any visually detectable growth during the same period.

  16. Pervaporation of ethanol from lignocellulosic fermentation broth.

    PubMed

    Gaykawad, Sushil S; Zha, Ying; Punt, Peter J; van Groenestijn, Johan W; van der Wielen, Luuk A M; Straathof, Adrie J J

    2013-02-01

    Pervaporation can be applied in ethanol production from lignocellulosic biomass. Hydrophobic pervaporation, using a commercial PDMS membrane, was employed to concentrate the ethanol produced by fermentation of lignocellulosic hydrolysate. To our knowledge, this is the first report describing this. Pervaporation carried out with three different lignocellulosic fermentation broths reduced the membrane performance by 17-20% as compared to a base case containing only 3 wt.% ethanol in water. The membrane fouling caused by these fermentation broths was irreversible. Solutions containing model lignocellulosic components were tested during pervaporation at the same conditions. A total flux decrease of 12-15%, as compared to the base case, was observed for each component except for furfural. Catechol was found to be most fouling component whereas furfural permeated through the membrane and increased the total flux. The membrane selectivity increased in the presence of fermentation broth but remained unchanged for all selected components.

  17. Species of Genus Ganoderma (Agaricomycetes) Fermentation Broth: A Novel Antioxidant and Antimicrobial Agent.

    PubMed

    Cilerdzic, Jasmina; Kosanic, Marijana; Stajić, Mirjana; Vukojevic, Jelena; Ranković, Branislav

    2016-01-01

    The bioactivity of Ganoderma lucidum basidiocarps has been well documented, but there are no data on the medicinal properties of its submerged cultivation broth nor on the other species of the genus Ganoderma. Thus the aim of this study was to test the potential antimicrobial and antioxidant activity of fermentation broth obtained after submerged cultivation of G. applanatum, G. carnosum, and G. lucidum. DPPH· scavenging ability, total phenols, and flavonoid contents were measured to determine the antioxidative potential of Ganoderma spp. fermentation filtrates, whereas their antimicrobial potential was studied using the microdilution method. DPPH· scavenging activity of G. lucidum fermentation filtrates was significantly higher than that of G. applanatum and G. carnosum, with the maximum (39.67%) obtained from strain BEOFB 432. This filtrate also contained the highest concentrations of phenols (134.89 μg gallic acid equivalents/mL) and flavonoids (42.20 μg quercetin equivalent/mL). High correlations between the activity and phenol content in the extracts showed that these compounds were active components of the antioxidative activity. G. lucidum strain BEOFB 432 was the most effective antibacterial agent, whereas strain BEOFB 434 has proven to be the most effective antifungal agent. The study showed that Ganoderma spp. fermentation filtrates are novel potent antioxidative and antimicrobial agents that could be obtained more quickly and cheaper than basidiocarps.

  18. Recovery of succinic acid from fermentation broth.

    PubMed

    Kurzrock, Tanja; Weuster-Botz, Dirk

    2010-03-01

    Succinic acid is of high interest as bio-feedstock for the chemical industry. It is a precursor for a variety of many other chemicals, e.g. 1,4-butandiol, tetrahydrofuran, biodegradable polymers and fumaric acid. Besides optimized production strains and fermentation processes it is indispensable to develop cost-saving and energy-effective downstream processes to compete with the current petrochemical production process. Various methods such as precipitation, sorption and ion exchange, electrodialysis, and liquid-liquid extraction have been investigated for the recovery of succinic acid from fermentation broth and are reviewed critically here.

  19. Carbapenem Susceptibility Testing Errors Using Three Automated Systems, Disk Diffusion, Etest, and Broth Microdilution and Carbapenem Resistance Genes in Isolates of Acinetobacter baumannii-calcoaceticus Complex

    DTIC Science & Technology

    2011-10-01

    Phoenix, and Vitek 2 systems). Discordant results were categorized as very major errors (VME), major errors (ME), and minor errors (mE). DNA sequences...01 OCT 2011 2 . REPORT TYPE N/A 3. DATES COVERED - 4. TITLE AND SUBTITLE Carbapenem Susceptibility Testing Errors Using Three Automated...FDA standards required for device approval (11). The Vitek 2 method was the only automated susceptibility method in our study that satisfied FDA

  20. Comparison of the EUCAST and CLSI Broth Microdilution Methods for Testing Isavuconazole, Posaconazole, and Amphotericin B against Molecularly Identified Mucorales Species

    PubMed Central

    Singh, Pradeep Kumar; Kathuria, Shallu

    2015-01-01

    We compared EUCAST and CLSI antifungal susceptibility testing (AFST) methods for triazoles and amphotericin B against 124 clinical Mucorales isolates. The EUCAST method yielded MIC values 1- to 3-fold dilutions higher than those of the CLSI method for amphotericin B. The essential agreements between the two methods for triazoles were high, i.e., 99.1% (voriconazole), 98.3% (isavuconazole), and 87% (posaconazole), whereas it was significantly lower for amphotericin B (66.1%). Strategies for harmonization of the two methods for Mucorales AFST are warranted. PMID:26438489

  1. Comparison of anidulafungin MICs determined by the clinical and laboratory standards institute broth microdilution method (M27-A3 document) and Etest for Candida species isolates.

    PubMed

    Espinel-Ingroff, Ana; Canton, E; Peman, J; Martín-Mazuelo, E

    2010-03-01

    Anidulafungin Etest and CLSI MICs were compared for 143 Candida sp. isolates to assess essential (within 2 log(2) dilutions) and categorical agreements (according to three susceptibility breakpoints). Based on agreement percentages, our data indicated that Etest is not suitable to test anidulafungin against Candida parapsilosis and C. guilliermondii (54.4 to 82.4% essential and categorical agreements) but is more suitable for C. albicans, C. glabrata, C. krusei, and C. tropicalis (87.9 to 100% categorical agreement).

  2. International Evaluation of MIC Distributions and Epidemiological Cutoff Value (ECV) Definitions for Fusarium Species Identified by Molecular Methods for the CLSI Broth Microdilution Method.

    PubMed

    Espinel-Ingroff, A; Colombo, A L; Cordoba, S; Dufresne, P J; Fuller, J; Ghannoum, M; Gonzalez, G M; Guarro, J; Kidd, S E; Meis, J F; Melhem, T M S C; Pelaez, T; Pfaller, M A; Szeszs, M W; Takahaschi, J P; Tortorano, A M; Wiederhold, N P; Turnidge, J

    2016-02-01

    The CLSI epidemiological cutoff values (ECVs) of antifungal agents are available for various Candida spp., Aspergillus spp., and the Mucorales. However, those categorical endpoints have not been established for Fusarium spp., mostly due to the difficulties associated with collecting sufficient CLSI MICs for clinical isolates identified according to the currently recommended molecular DNA-PCR-based identification methodologies. CLSI MIC distributions were established for 53 Fusarium dimerum species complex (SC), 10 F. fujikuroi, 82 F. proliferatum, 20 F. incarnatum-F. equiseti SC, 226 F. oxysporum SC, 608 F. solani SC, and 151 F. verticillioides isolates originating in 17 laboratories (in Argentina, Australia, Brazil, Canada, Europe, Mexico, and the United States). According to the CLSI guidelines for ECV setting, ECVs encompassing ≥97.5% of pooled statistically modeled MIC distributions were as follows: for amphotericin B, 4 μg/ml (F. verticillioides) and 8 μg/ml (F. oxysporum SC and F. solani SC); for posaconazole, 2 μg/ml (F. verticillioides), 8 μg/ml (F. oxysporum SC), and 32 μg/ml (F. solani SC); for voriconazole, 4 μg/ml (F. verticillioides), 16 μg/ml (F. oxysporum SC), and 32 μg/ml (F. solani SC); and for itraconazole, 32 μg/ml (F. oxysporum SC and F. solani SC). Insufficient data precluded ECV definition for the other species. Although these ECVs could aid in detecting non-wild-type isolates with reduced susceptibility to the agents evaluated, the relationship between molecular mechanisms of resistance (gene mutations) and MICs still needs to be investigated for Fusarium spp. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

  3. International and multicenter comparison of EUCAST and CLSI M27-A2 broth microdilution methods for testing susceptibilities of Candida spp. to fluconazole, itraconazole, posaconazole, and voriconazole.

    PubMed

    Espinel-Ingroff, A; Barchiesi, F; Cuenca-Estrella, M; Pfaller, M A; Rinaldi, M; Rodriguez-Tudela, J L; Verweij, P E

    2005-08-01

    The aim of this study was to compare MICs of fluconazole, itraconazole, posaconazole, and voriconazole obtained by the European Committee on Antibiotic Susceptibility Testing (EUCAST) and CLSI (formerly NCCLS) methods in each of six centers for 15 Candida albicans (5 fluconazole-resistant and 4 susceptible-dose-dependent [S-DD] isolates), 10 C. dubliniensis, 7 C. glabrata (2 fluconazole-resistant isolates), 5 C. guilliermondii (2 fluconazole-resistant isolates), 10 C. krusei, 9 C. lusitaniae, 10 C. parapsilosis, and 5 C. tropicalis (1 fluconazole-resistant isolate) isolates. CLSI MICs were obtained visually at 24 and 48 h and spectrophotometric EUCAST MICs at 24 h. The agreement (within a 3-dilution range) between the methods was species, drug, and incubation time dependent and due to lower EUCAST than CLSI MICs: overall, 94 to 95% with fluconazole and voriconazole and 90 to 91% with posaconazole and itraconazole when EUCAST MICs were compared against 24-h CLSI results. The agreement was lower (85 to 94%) against 48-h CLSI endpoints. The overall interlaboratory reproducibility by each method was > or =92%. When the comparison was based on CLSI breakpoint categorization, the agreement was 68 to 76% for three of the four species that included fluconazole-resistant and S-DD isolates; 9% very major discrepancies (< or =8 microg/ml versus > or =64 microg/ml) were observed among fluconazole-resistant isolates and 50% with voriconazole (< or =1 microg/ml versus > or =4 microg/ml). Similar results were observed with itraconazole for seven of the eight species evaluated (28 to 77% categorical agreement). Posaconazole EUCAST MICs were also substantially lower than CLSI MIC modes (0.008 to 1 microg/ml versus 1 to > or =8 microg/ml) for some of these isolates. Therefore, the CLSI breakpoints should not be used to interpret EUCAST MIC data.

  4. In vitro activity of a novel broad-spectrum antifungal, E1210, tested against Aspergillus spp. determined by CLSI and EUCAST broth microdilution methods.

    PubMed

    Pfaller, Michael A; Duncanson, Frederick; Messer, Shawn A; Moet, Gary J; Jones, Ronald N; Castanheira, Mariana

    2011-11-01

    E1210 is a first-in-class broad-spectrum antifungal that suppresses hyphal growth by inhibiting fungal glycophosphatidylinositol (GPI) biosynthesis. In the present study, we extend these findings by examining the activity of E1210 and comparator antifungal agents against Aspergillus spp. by using the methods of the Clinical and Laboratory Standards Institute (CLSI) and the European Committee for Antimicrobial Susceptibility Testing (EUCAST) to test wild-type (WT) as well as amphotericin B (AMB)-resistant (-R) and azole-R strains (as determined by CLSI methods). Seventy-eight clinical isolates of Aspergillus were tested including 20 isolates of Aspergillus flavus species complex (SC), 22 of A. fumigatus SC, 13 of A. niger SC, and 23 of A. terreus SC. The collection included 15 AMB-R (MIC, ≥ 2 μg/ml) isolates of A. terreus SC and 10 itraconazole-R (MIC, ≥ 4 μg/ml) isolates of A. fumigatus SC (7 isolates), A. niger SC (2 isolates), and A. terreus SC (1 isolate). Comparator antifungal agents included anidulafungin, caspofungin, amphotericin B, itraconazole, posaconzole, and voriconazole. Both CLSI and EUCAST methods were highly concordant for E1210 and all comparators. The essential agreement (EA; ± 2 log(2) dilution steps) was 100% for all comparisons with the exception of posaconazole versus A. terreus SC (EA = 91.3%). The minimum effective concentration (MEC)/MIC(90) values (μg/ml) for E1210, anidulafungin, caspofungin, itraconazole, posaconazole, and voriconazole, respectively, were as follows for each species: for A. flavus SC, 0.03, ≤ 0.008, 0.12, 1, 1, and 1; for A. fumigatus SC, 0.06, 0.015, 0.12, >8, 1, and 4; for A. niger SC, 0.015, 0.03, 0.12, 4, 1, and 2; and for A. terreus SC, 0.06, 0.015, 0.12, 1, 0.5, and 1. E1210 was very active against AMB-R strains of A. terreus SC (MEC range, 0.015 to 0.06 μg/ml) and itraconazole-R strains of A. fumigatus SC (MEC range, 0.03 to 0.12 μg/ml), A. niger SC (MEC, 0.008 μg/ml), and A. terreus SC (MEC, 0.015 μg/ml). In conclusion, E1210 was a very potent and broad-spectrum antifungal agent regardless of in vitro method applied, with excellent activity against AMB-R and itraconazole-R strains of Aspergillus spp.

  5. In Vitro Activity of a Novel Broad-Spectrum Antifungal, E1210, Tested against Aspergillus spp. Determined by CLSI and EUCAST Broth Microdilution Methods ▿

    PubMed Central

    Pfaller, Michael A.; Duncanson, Frederick; Messer, Shawn A.; Moet, Gary J.; Jones, Ronald N.; Castanheira, Mariana

    2011-01-01

    E1210 is a first-in-class broad-spectrum antifungal that suppresses hyphal growth by inhibiting fungal glycophosphatidylinositol (GPI) biosynthesis. In the present study, we extend these findings by examining the activity of E1210 and comparator antifungal agents against Aspergillus spp. by using the methods of the Clinical and Laboratory Standards Institute (CLSI) and the European Committee for Antimicrobial Susceptibility Testing (EUCAST) to test wild-type (WT) as well as amphotericin B (AMB)-resistant (-R) and azole-R strains (as determined by CLSI methods). Seventy-eight clinical isolates of Aspergillus were tested including 20 isolates of Aspergillus flavus species complex (SC), 22 of A. fumigatus SC, 13 of A. niger SC, and 23 of A. terreus SC. The collection included 15 AMB-R (MIC, ≥2 μg/ml) isolates of A. terreus SC and 10 itraconazole-R (MIC, ≥4 μg/ml) isolates of A. fumigatus SC (7 isolates), A. niger SC (2 isolates), and A. terreus SC (1 isolate). Comparator antifungal agents included anidulafungin, caspofungin, amphotericin B, itraconazole, posaconzole, and voriconazole. Both CLSI and EUCAST methods were highly concordant for E1210 and all comparators. The essential agreement (EA; ±2 log2 dilution steps) was 100% for all comparisons with the exception of posaconazole versus A. terreus SC (EA = 91.3%). The minimum effective concentration (MEC)/MIC90 values (μg/ml) for E1210, anidulafungin, caspofungin, itraconazole, posaconazole, and voriconazole, respectively, were as follows for each species: for A. flavus SC, 0.03, ≤0.008, 0.12, 1, 1, and 1; for A. fumigatus SC, 0.06, 0.015, 0.12, >8, 1, and 4; for A. niger SC, 0.015, 0.03, 0.12, 4, 1, and 2; and for A. terreus SC, 0.06, 0.015, 0.12, 1, 0.5, and 1. E1210 was very active against AMB-R strains of A. terreus SC (MEC range, 0.015 to 0.06 μg/ml) and itraconazole-R strains of A. fumigatus SC (MEC range, 0.03 to 0.12 μg/ml), A. niger SC (MEC, 0.008 μg/ml), and A. terreus SC (MEC, 0.015 μg/ml). In conclusion, E1210 was a very potent and broad-spectrum antifungal agent regardless of in vitro method applied, with excellent activity against AMB-R and itraconazole-R strains of Aspergillus spp. PMID:21844312

  6. International and Multicenter Comparison of EUCAST and CLSI M27-A2 Broth Microdilution Methods for Testing Susceptibilities of Candida spp. to Fluconazole, Itraconazole, Posaconazole, and Voriconazole

    PubMed Central

    Espinel-Ingroff, A.; Barchiesi, F.; Cuenca-Estrella, M.; Pfaller, M. A.; Rinaldi, M.; Rodriguez-Tudela, J. L.; Verweij, P. E.

    2005-01-01

    The aim of this study was to compare MICs of fluconazole, itraconazole, posaconazole, and voriconazole obtained by the European Committee on Antibiotic Susceptibility Testing (EUCAST) and CLSI (formerly NCCLS) methods in each of six centers for 15 Candida albicans (5 fluconazole-resistant and 4 susceptible-dose-dependent [S-DD] isolates), 10 C. dubliniensis, 7 C. glabrata (2 fluconazole-resistant isolates), 5 C. guilliermondii (2 fluconazole-resistant isolates), 10 C. krusei, 9 C. lusitaniae, 10 C. parapsilosis, and 5 C. tropicalis (1 fluconazole-resistant isolate) isolates. CLSI MICs were obtained visually at 24 and 48 h and spectrophotometric EUCAST MICs at 24 h. The agreement (within a 3-dilution range) between the methods was species, drug, and incubation time dependent and due to lower EUCAST than CLSI MICs: overall, 94 to 95% with fluconazole and voriconazole and 90 to 91% with posaconazole and itraconazole when EUCAST MICs were compared against 24-h CLSI results. The agreement was lower (85 to 94%) against 48-h CLSI endpoints. The overall interlaboratory reproducibility by each method was ≥92%. When the comparison was based on CLSI breakpoint categorization, the agreement was 68 to 76% for three of the four species that included fluconazole-resistant and S-DD isolates; 9% very major discrepancies (≤8 μg/ml versus ≥64 μg/ml) were observed among fluconazole-resistant isolates and 50% with voriconazole (≤1 μg/ml versus ≥4 μg/ml). Similar results were observed with itraconazole for seven of the eight species evaluated (28 to 77% categorical agreement). Posaconazole EUCAST MICs were also substantially lower than CLSI MIC modes (0.008 to 1 μg/ml versus 1 to ≥8 μg/ml) for some of these isolates. Therefore, the CLSI breakpoints should not be used to interpret EUCAST MIC data. PMID:16081926

  7. Comparison of the EUCAST and CLSI Broth Microdilution Methods for Testing Isavuconazole, Posaconazole, and Amphotericin B against Molecularly Identified Mucorales Species.

    PubMed

    Chowdhary, Anuradha; Singh, Pradeep Kumar; Kathuria, Shallu; Hagen, Ferry; Meis, Jacques F

    2015-12-01

    We compared EUCAST and CLSI antifungal susceptibility testing (AFST) methods for triazoles and amphotericin B against 124 clinical Mucorales isolates. The EUCAST method yielded MIC values 1- to 3-fold dilutions higher than those of the CLSI method for amphotericin B. The essential agreements between the two methods for triazoles were high, i.e., 99.1% (voriconazole), 98.3% (isavuconazole), and 87% (posaconazole), whereas it was significantly lower for amphotericin B (66.1%). Strategies for harmonization of the two methods for Mucorales AFST are warranted. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  8. Wild-type MIC distributions, epidemiological cutoff values and species-specific clinical breakpoints for fluconazole and Candida: time for harmonization of CLSI and EUCAST broth microdilution methods.

    PubMed

    Pfaller, M A; Andes, D; Diekema, D J; Espinel-Ingroff, A; Sheehan, D

    2010-12-01

    Both the Clinical and Laboratory Standards Institute (CLSI) and the European Committee on Antimicrobial Susceptibility Testing (EUCAST) have MIC clinical breakpoints (CBPs) for fluconazole (FLU) and Candida. EUCAST CBPs are species-specific, and apply only to C. albicans, C. tropicalis and C. parapsilosis, while CLSI CBPs apply to all species. We reassessed the CLSI CBPs for FLU and Candida in light of recent data. We examined (1) molecular mechanisms of resistance and cross-resistance profiles, (2) wild-type (WT) MICs and epidemiological cutoff values (ECVs) for FLU and major Candida species by both CLSI and EUCAST methods, (3) determination of essential (EA) and categorical agreement (CA) between CLSI and EUCAST methods, (4) correlation of MICs with outcomes from previously published data using CLSI and EUCAST methods, and (5) pharmacokinetic and pharmacodynamic considerations. We applied these findings to propose new species-specific CLSI CBPs for FLU and Candida. WT distributions from large collections of Candida revealed similar ECVs by both CLSI and EUCAST methods (0.5-1 mcg/ml for C. albicans, 2 mcg/ml for C. parapsilosis and C. tropicalis, 32 mcg/ml for C. glabrata, and 64-128 for C. krusei). Comparison of CLSI and EUCAST MICs reveal EA and CA of 95% and 96%, respectively. Datasets correlating CLSI and EUCAST FLU MICs with outcomes revealed decreased response rates when MICs were > 4 mcg/ml for C. albicans, C. tropicalis and C. parapsilosis, and > 16 mcg/ml for C. glabrata. Adjusted CLSI CBPs for FLU and C. albicans, C. parapsilosis, C. tropicalis (S, ≤ 2 mcg/ml; SDD, 4 mcg/ml; R, ≥ 8 mcg/ml), and C. glabrata (SDD, ≤ 32 mcg/ml; R, ≥ 64 mcg/ml) should be more sensitive for detecting emerging resistance among common Candida species and provide consistency with EUCAST CBPs. Copyright © 2010 Elsevier Ltd. All rights reserved.

  9. International Evaluation of MIC Distributions and Epidemiological Cutoff Value (ECV) Definitions for Fusarium Species Identified by Molecular Methods for the CLSI Broth Microdilution Method

    PubMed Central

    Colombo, A. L.; Cordoba, S.; Dufresne, P. J.; Fuller, J.; Ghannoum, M.; Gonzalez, G. M.; Guarro, J.; Kidd, S. E.; Melhem, T. M. S. C.; Pelaez, T.; Pfaller, M. A.; Szeszs, M. W.; Takahaschi, J. P.; Wiederhold, N. P.; Turnidge, J.

    2015-01-01

    The CLSI epidemiological cutoff values (ECVs) of antifungal agents are available for various Candida spp., Aspergillus spp., and the Mucorales. However, those categorical endpoints have not been established for Fusarium spp., mostly due to the difficulties associated with collecting sufficient CLSI MICs for clinical isolates identified according to the currently recommended molecular DNA-PCR-based identification methodologies. CLSI MIC distributions were established for 53 Fusarium dimerum species complex (SC), 10 F. fujikuroi, 82 F. proliferatum, 20 F. incarnatum-F. equiseti SC, 226 F. oxysporum SC, 608 F. solani SC, and 151 F. verticillioides isolates originating in 17 laboratories (in Argentina, Australia, Brazil, Canada, Europe, Mexico, and the United States). According to the CLSI guidelines for ECV setting, ECVs encompassing ≥97.5% of pooled statistically modeled MIC distributions were as follows: for amphotericin B, 4 μg/ml (F. verticillioides) and 8 μg/ml (F. oxysporum SC and F. solani SC); for posaconazole, 2 μg/ml (F. verticillioides), 8 μg/ml (F. oxysporum SC), and 32 μg/ml (F. solani SC); for voriconazole, 4 μg/ml (F. verticillioides), 16 μg/ml (F. oxysporum SC), and 32 μg/ml (F. solani SC); and for itraconazole, 32 μg/ml (F. oxysporum SC and F. solani SC). Insufficient data precluded ECV definition for the other species. Although these ECVs could aid in detecting non-wild-type isolates with reduced susceptibility to the agents evaluated, the relationship between molecular mechanisms of resistance (gene mutations) and MICs still needs to be investigated for Fusarium spp. PMID:26643334

  10. Caspofungin Etest endpoint for Aspergillus isolates shows poor agreement with the reference minimum effective concentration.

    PubMed

    Fuller, Jeff; Schofield, Adam; Jiwa, Safeer; Sand, Crystal; Jansen, Brad; Rennie, Robert

    2010-02-01

    The Clinical and Laboratory Standards Institute (CLSI) M38-A2 reference broth microdilution (BMD) method for the antifungal susceptibility testing of filamentous fungi now includes guidelines for testing echinocandin activity using the minimum effective concentration (MEC) as the endpoint measurement. In this study, we compared the caspofungin Etest MIC on RPMI agar and Mueller-Hinton agar (supplemented with glucose and methylene blue [MGM]) to the BMD MEC for 345 clinical Aspergillus isolates, including A. flavus, A. fumigatus, A. nidulans, A. niger, and A. terreus. The essential agreement (+/-1 log(2) dilution) of the Etest on MGM and RPMI agar with the reference BMD MEC was 18 and 26%, respectively. The geometric mean values for BMD MEC and MGM Etest were 0.137 and 0.024 microg/ml, respectively, and the geometric mean values for BMD and RPMI agar were 0.128 and 0.031 microg/ml, respectively. Comparatively, 91% of paired MGM and RPMI Etest results were within 2 log(2) dilutions of each other and consistently produced clearly defined endpoints. In conclusion, the caspofungin Etest MIC, like the BMD MEC, is a reproducible endpoint but is markedly lower than the reference BMD. In anticipation of susceptibility breakpoint assignments, optimization studies will be required to improve the concordance of these two assays so that the potential for underreporting echinocandin resistance in Aspergillus is mitigated.

  11. Recovery of actinorhodin from fermentation broth.

    PubMed

    Güzeltunç, E; Ulgen, K O

    2001-04-20

    In the present work, a new method of purification for actinorhodin was developed using an expanded bed chromatography technique in which antibiotic capture, feedstock clarification, centrifugation, dialysis and concentration are done in one step. The cation-exchanger (P-11) resulted in 26% adsorption and 2% recovery whereas the anion-exchanger (DE-52) resulted in 99% adsorption and 56% recovery of adsorbed antibiotic using methanol buffer and 2 M NH4Cl as eluting agent. Streamline DEAE anion-exchanger, which is especially designed for EBA applications, yields 82% adsorption and 50% elution of actinorhodin fed into the chromatography column directly from the fermentation broth. Isocratic elution resulted in extremely efficient yield compared to linear gradient elution, i.e. 13.5-fold more recovery in the column with an aspect ratio (L:D) of 4. Expansion by 150% of settled bed resulted in the best recovery of actinorhodin among 100 and 200% expansions. A comparison of breakthrough profiles in packed and expanded bed adsorption showed that the performance of the expanded bed is better (by 33%) at allowing more volume of the fermentation broth to pass through the chromatography column.

  12. Comparison of microdilution and disc diffusion methods in assessing the in vitro activity of fluconazole and Melaleuca alternifolia (tea tree) oil against vaginal Candida isolates.

    PubMed

    Ergin, A; Arikan, S

    2002-10-01

    The in vitro activity of fluconazole and Melaleuca alternifolia (tea tree) oil was evaluated against 99 vaginal Candida strains by the broth microdilution and disc diffusion methods. The microdilution method was performed in accordance with NCCLS-M27A guidelines. An investigational method was used for the disc diffusion test. Fluconazole and tea tree oil minimum inhibitory concentrations (MICs) obtained at 48 h tended to increase 1- to 2-fold or remain the same compared to 24 h readings for most of the isolates tested. C. krusei and C. norvegensis had significantly higher MICs and smaller inhibition zones for fluconazole compared to other species. Tea tree oil MICs were found to be similar, in general, for all Candida spp. tested. The geometric mean MIC of tea tree oil for all isolates was 2.2% (range, 0.25-4%) at 24 h and 3.0% (range, 1-8%) at 48 h. Tea tree oil mean inhibition zone diameter was 24 mm (range, 14-42 mm) at 24 h and 15.8 mm (range, 10-35 mm) at 48 h. In vitro activity of tea tree oil against fluconazole-resistant Candida strains was of particular interest. The isolates had similar tea tree oil MICs and inhibition zone diameters regardless of their fluconazole susceptibility profile. Tea tree oil MIC ranges (inhibition zone diameter ranges) were 2-4% (12-21 mm) and 2% (35 mm) at 48 h for C. krusei and C. norvegensis, respectively. These results suggest that tea tree oil MICs of the fluconazole-resistant isolates are comparable to those of fluconazole-susceptible isolates. This in vitro finding is promising for potential use of topical tea tree oil formulations in the treatment of candidiasis due to fluconazole-resistant strains.

  13. Preparative separation of echinocandin B from Aspergillus nidulans broth using macroporous resin adsorption chromatography.

    PubMed

    Zou, Shu-Ping; Liu, Miao; Wang, Qiu-Liang; Xiong, Yan; Niu, Kun; Zheng, Yu-Guo; Shen, Yin-Chu

    2015-01-26

    Echinocandin B (ECB), an echinocandin type of lipopeptide antibiotic produced by Aspergillus nidulans, is a precursor for the synthesis of novel anti-fungal drug - anidulafungin. In this work, a separation strategy involving one-step macroporous resin adsorption chromatography was established for ECB purification from Aspergillus nidulans CCTCC M 2010275 fermentation broth. Among nine macroporous resin adsorbents tested, the non-polar resin HP-20 had the best adsorption and desorption performance. The static equilibrium adsorption data fitted well with the Langmuir equation, and the adsorption kinetic followed the pseudo-second order model. The separation parameters of ECB from broth were optimised by dynamic adsorption/desorption experiments with the column packed with HP-20 resin. Under optimal conditions, the purity increased by 3.8-fold from 23.2% in broth to 88.5% in eluent with 87.1% recovery yield by a one-step treatment. Our study provided a one-step and effective method for large-scale production of ECB, and offered references for separating other echinocandins from broth. Copyright © 2014 Elsevier B.V. All rights reserved.

  14. Performance of three chromogenic VRE screening agars, two Etest(®) vancomycin protocols, and different microdilution methods in detecting vanB genotype Enterococcus faecium with varying vancomycin MICs.

    PubMed

    Klare, Ingo; Fleige, Carola; Geringer, Uta; Witte, Wolfgang; Werner, Guido

    2012-10-01

    Frequencies of vanB-type Enterococcus faecium increased in Europe during the last years. VanB enterococci show various levels of vancomycin MICs even below the susceptible breakpoint challenging a reliable diagnostics. The performance of 3 chromogenic vancomycin-resistant enterococci (VRE) screening agars, 2 Etest® vancomycin protocols, and different microdilution methods to detect 129 clinical vanB E. faecium strains was investigated. Altogether, 112 (87%) were correctly identified as VanB-type Enterococcus by microdilution MICs. An Etest® macromethod protocol was more sensitive than the standard protocol while keeping sufficient specificity in identifying 15 vanA/vanB-negative strains. Three chromogenic VRE agars performed similarly with 121 (94%), 123 (95%), and 124 (96%) vanB isolates that grew on Brilliance™ VRE Agar, CHROMagar™ VRE, and chromID™ VRE agar, respectively. Using identical media and conditions, we did not identify different growth behaviour on agar and in broth. A few vanB strains showed growth of microcolonies inside the Etest® vancomycin inhibition zones, suggesting a VanB heteroresistance phenotype. Copyright © 2012 Elsevier Inc. All rights reserved.

  15. Correlation between microdilution, E-test, and disk diffusion methods for antifungal susceptibility testing of posaconazole against Candida spp.

    PubMed

    Sims, Charles R; Paetznick, Victor L; Rodriguez, Jose R; Chen, Enuo; Ostrosky-Zeichner, Luis

    2006-06-01

    Agar-based antifungal susceptibility testing is an attractive alternative to the microdilution method. We examined the correlation between the microdilution, E-test, and disk diffusion methods for posaconazole against Candida spp. A total of 270 bloodstream isolates of Candida spp. with a broad range of posaconazole MICs were tested using the CLSI M27-A2 method for microdilution, as well as the M-44A method and E-test methods for agar-based testing on Mueller-Hinton agar supplemented with 2% glucose and 0.5 microg of methylene blue. MICs and inhibitory zone diameters at the prominent growth reduction endpoint were recorded at 24 and 48 h. The Candida isolates included Candida albicans (n = 124), C. parapsilosis (n = 44), C. tropicalis (n = 41), C. glabrata (n = 36), C. krusei (n = 20), C. lusitaniae (n = 3), and C. dubliniensis (n = 2). The overall concordance (i.e., the percentage of isolates within two dilutions) between the E-test and microdilution was 64.8% at 24 h and 82.6% at 48 h. When we considered an arbitrary breakpoint of < or = 1 microg/ml, the agreement between the E-test and microdilution methods was 87.8% at 24 h and 93.0% at 48 h. The correlation of MICs with disk diffusion zone diameters was better for the E-test than the microdilution method. Zone correlation for diameters produced by the disks of two manufacturers was high, with a Pearson test value of 0.941 at 24 h. The E-test and microdilution MICs show good concordance and interpretative agreement. The disk diffusion zone diameters are highly reproducible and correlate well with both the E-test and the microdilution method, making agar-based methods a viable alternative to microdilution for posaconazole susceptibility testing.

  16. Steady-state shear characteristics of Aspergillus niger broths

    SciTech Connect

    Svihla, C.K.; Dronawat, S.N.; Hanley, T.R.

    1995-12-31

    It can be difficult to obtain reliable rheological data for filamentous fermentation broths using conventional instruments. One common approach is to measure the torque drawn by an impeller rotating in the suspension. Many previous workers have assumed that the applicable shear rate in such a device is related to the impeller speed by a fluid-independent constant determined by calibration with Newtonian and non-Newtonian fluids. The rheology of Aspergillus niger broths have been characterized using the impeller viscometer approach. The changes in the broth rheology were measured, and used to interpret the growth of biomass and the evolution of the microorganism morphology.

  17. Antifungal susceptibility of the aspergillus species by Etest and CLSI reference methods.

    PubMed

    Badiee, Parisa; Alborzi, Abdolvahab; Moeini, Mahsa; Haddadi, Pedram; Farshad, Shohreh; Japoni, Aziz; Ziyaeyan, Maziar

    2012-07-01

    Because resistance to antifungal drugs is seen in patients, susceptibility testing of these drugs aids in choosing the appropriate drug and respective epidemiology. This study has investigated and compared susceptibility patterns of the Aspergillus species isolated from patients by the Clinical and Laboratory Standards Institute (CLSI) reference broth microdilution (MD) assay and Etest method. The minimum inhibitory concentrations (MICs) of various antifungal agents (amphotericin B, ketoconazole, itraconazole, and voriconazole) for 108 Aspergillus species isolated from patients were determined by CLSI M38-A broth MD and Etest. The isolates were obtained from clinical samples that included tissues, sputum, bronchoalveolar lavage, abdominal tap, and cerebrospinal fluid. As revealed by the MD method, 63.9% of the isolates were sensitive to amphotericin B and 36.1% were resistant.Etest revealed that 61.1% were sensitive to amphotericin B and 38.9% were resistant. As for ketoconazole, 108 isolates (100%) were shown to be sensitive through the MD method; while the Etest revealedan 88.9% sensitivity and 11.1% were resistant. All species were susceptible to voriconazole, according to both methods. The measure of agreement (Kappa Index) for these three drugs was satisfactory (≥0.6). According to the MD method, 69.4% of the species were susceptible to itraconazole, whereas 30.6% were not.For this drug, the Etest showed 86.1% susceptible and 13.9% resistant. Voriconazole was the most effective agent against isolates. Using RPMI agar, we found the Etest to be helpful, readily available, and easy to use for determining invitro susceptibilities of Aspergillus species to voriconazole, amphotericin B, ketoconazole, and itraconazole in the region of this study.

  18. Determination of Hydrogen Sulfide in Fermentation Broths Containing SO21

    PubMed Central

    Acree, T. E.; Sonoff, Elisabeth P.; Splittstoesser, D. F.

    1971-01-01

    A procedure for the determination of hydrogen sulfide in fermentation broths containing up to 100 μg of SO2 per ml is described. The method involves the sparging of H2S from the broth into a cadmium hydroxide absorption solution, the formation of methylene blue from the absorbed sulfide, and the measuring of this color spectrophotometrically. The use of cadmium hydroxide instead of zinc acetate, the common absorbent, substantially reduced the interference of SO2 with the analysis. PMID:5111300

  19. Evaluation of TA10 Broth for Recovery of Heat- and Freeze-Injured Salmonella from Beef

    USDA-ARS?s Scientific Manuscript database

    The Bacteriological Analytical Manual (BAM) Salmonella pre-enrichment broth (lactose [LAC] broth), buffered peptone water (BPW), and universal preenrichment (UP) broth were compared with TA10 broth, developed in our laboratory, for recovery of heat- and freeze-injured Salmonella (55ºC for 2-20 min a...

  20. Mechanistic modeling of broth temperature in outdoor photobioreactors.

    PubMed

    Béchet, Quentin; Shilton, Andy; Fringer, Oliver B; Muñoz, Raul; Guieysse, Benoit

    2010-03-15

    This study presents the first mechanistic model describing broth temperature in column photobioreactors as a function of static (location, reactor geometry) and dynamic (light irradiance, air temperature, wind velocity) parameters. Based on a heat balance on the liquid phase the model predicted temperature in a pneumatically agitated column photobioreactor (1 m(2) illuminated area, 0.19 m internal diameter, 50 L gas-free cultivation broth) operated outdoor in Singapore to an accuracy of 2.4 °C at the 95% confidence interval over the entire data set used (104 measurements from 7 different batches). Solar radiation (0 to 200 W) and air convection (-30 to 50 W)were the main contributors to broth temperature change. The model predicted broth temperature above 40 °C will be reached during summer months in the same photobioreactor operated in California, a value well over the maximum temperature tolerated by most commercial algae species. Accordingly, 18,000 and 5500 GJ year(-1) ha(-1) of heat energy must be removed to maintain broth temperature at or below 25 and 35 °C, respectively, assuming a reactor density of one reactor per square meter. Clearly, the significant issue of temperature control must be addressed when evaluating the technical feasibility, costs, and sustainability of large-scale algae production.

  1. Salmonella in foods: new enrichment procedure for TECRA Salmonella Visual Immunoassay using a single rv(R10) only, TT only, or dual rv(R10) and TT selective enrichment broths (AOAC official method 998.09): collaborative study.

    PubMed

    Hughes, Denise; Dailianis, Angela E; Hill, Louise; Curiale, Michael S; Gangar, Vidhya

    2003-01-01

    A collaborative study was conducted to compare a new enrichment procedure for the TECRA Salmonella Visual Immunoassay (TSVIA) with the reference method given in the U.S. Food and Drug Administration's Bacteriological Analytical Manual (7th Ed.). Three food types (milk powder, pepper, and soy flour) were analyzed in Australia and 3 food types (milk chocolate, dried egg, and raw turkey) were analyzed in the United States. Thirty-eight collaborators participated in the study. The TECRA method was evaluated using both Rappaport-Vassiliadis R10 (RV(R10)) and tetrathionate (TT) broths for selective enrichment. M broth cultures arising from each of the 2 selective enrichment broths were tested in the TSVIA using 2 individual wells, one for each selective broth, and a single well to test the pooled selective enrichment broths. The results for the pooled enrichment broths were reported elsewhere. This study presents the results for the use of single enrichment broths, i.e., RV(R10) only or TT only, with the TSVIA. No significant differences (p > 0.05) were observed for the pairwise comparison of the proportion of positive samples for either RV(R10) or TT used as a single enrichment broth for the TSVIA with that for the reference method.

  2. Reverse osmosis processing of organic model compounds and fermentation broths.

    PubMed

    Diltz, Robert A; Marolla, Theodore V; Henley, Michael V; Li, Lixiong

    2007-02-01

    Post-treatment of an anaerobic fermentation broth was evaluated using a 150 gal/day, single cartridge prototype reverse osmosis (RO) system. Baseline tests were conducted at 25 degrees C using six organic model compounds representing key species found in the fermentation broth: ethanol, butanol, acetic acid, oxalic acid, lactic acid, and butyric acid. Correlations of the rejection and recovery efficiencies for these organic species, individually and in simulated mixtures, were obtained as a function of feed pressure with and without recirculation of the retentate. The actual fermentation broth obtained from a continuous-flow biohydrogen process was treated by the RO system under the operating conditions similar to those used in the baseline tests, resulting in greater than 95% removal of total organic carbon. These results are encouraging and useful for further studies on the feasibility of incorporating the RO technology into an integrated and field deployable wastewater management and water recovery system.

  3. Neuraminidase Inhibitors from the Fermentation Broth of Phellinus linteus

    PubMed Central

    Hwang, Byung Soon; Lee, Myeong-Seok; Lee, Seung Woong; Lee, In-Kyoung; Seo, Geon-Sik; Choi, Hwa Jung

    2014-01-01

    During a search for neuraminidase inhibitors derived from medicinal fungi, we found that the fermentation broth of Phellinus linteus exhibited potent neuraminidase inhibitory activity. Through bioassay-guided fractionation, two active compounds were purified from the ethyl acetate-soluble portion of the fermentation broth of P. linteus. These structures were identified as inotilone (1) and 4-(3,4-dihydroxyphenyl)-3-buten-2-one (2) by spectroscopic methods. Compounds 1 and 2 inhibited H1N1 neuraminidase activity with IC50 values of 29.1 and 125.6 µM, respectively, in a dose-dependent manner. They also exhibited an antiviral effect in a viral cytopathic effect reduction assay using MDCK cells. These results suggest that compounds 1 and 2 from the culture broth of P. linteus would be good candidates for the prevention and therapeutic strategies towards viral infections. PMID:25071390

  4. Neuraminidase Inhibitors from the Fermentation Broth of Phellinus linteus.

    PubMed

    Hwang, Byung Soon; Lee, Myeong-Seok; Lee, Seung Woong; Lee, In-Kyoung; Seo, Geon-Sik; Choi, Hwa Jung; Yun, Bong-Sik

    2014-06-01

    During a search for neuraminidase inhibitors derived from medicinal fungi, we found that the fermentation broth of Phellinus linteus exhibited potent neuraminidase inhibitory activity. Through bioassay-guided fractionation, two active compounds were purified from the ethyl acetate-soluble portion of the fermentation broth of P. linteus. These structures were identified as inotilone (1) and 4-(3,4-dihydroxyphenyl)-3-buten-2-one (2) by spectroscopic methods. Compounds 1 and 2 inhibited H1N1 neuraminidase activity with IC50 values of 29.1 and 125.6 µM, respectively, in a dose-dependent manner. They also exhibited an antiviral effect in a viral cytopathic effect reduction assay using MDCK cells. These results suggest that compounds 1 and 2 from the culture broth of P. linteus would be good candidates for the prevention and therapeutic strategies towards viral infections.

  5. Evaluation of the AutoSCAN-3, a device for reading microdilution trays.

    PubMed Central

    DeGirolami, P C; Eichelberger, K A; Salfity, L C; Rizzo, M F

    1983-01-01

    The AutoSCAN-3 (American MicroScan, Mahwah, N.J.) is an instrument capable of automated reading of commercially available microdilution trays for identification and quantitative susceptibility testing of rapidly growing bacteria. This study compared the results of visual and automated reading of microdilution trays for determination and interpretation of minimum inhibitory concentrations of 471 selected gram-negative and gram-positive clinical bacterial isolates. Visual and automated readings were performed in a double-blind fashion, and all discrepancies were examined by a referee. A quantitative comparison of minimum inhibitory concentrations was performed for 201 organisms, yielding 2,472 drug-organism combinations. After exclusion of off-scale values, complete quantitative agreement was obtained in 94% of 959 on-scale combinations, and agreement within +/- 1 well was obtained in 99.3%. Considering the minimum inhibitory concentration interpretations routinely furnished by the instrument, a qualitative comparison was performed for all 471 organisms. Complete agreement in interpretation was obtained in 97.6% of 5,843 drugs-organism combinations, with very major discrepancies accounting for only 0.1% and major discrepancies accounting for 0.2% of all combinations tested. PMID:6361050

  6. Metronidazole susceptibility testing for Helicobacter pylori: comparison of disk, broth, and agar dilution methods and their clinical relevance.

    PubMed Central

    DeCross, A J; Marshall, B J; McCallum, R W; Hoffman, S R; Barrett, L J; Guerrant, R L

    1993-01-01

    Since the methods for metronidazole susceptibility testing of Helicobacter pylori have not been standardized or validated, we compared three methods that are used to test the metronidazole susceptibilities of 25 isolates of H. pylori. Specifically, we examined the methods of Steer's replicator agar dilution, tube broth microdilution, and modified Kirby-Bauer disk diffusion. The metronidazole disk zone sizes obtained by the disk diffusion method correlated well (r = 0.74) with the MICs obtained by the agar dilution method. Afterward, the disk diffusion method was used to characterize the metronidazole susceptibilities of 44 isolates of H. pylori. Dual therapy (bismuth and metronidazole) proved to be highly effective against metronidazole-susceptible strains (81.6% eradication rate) but fared poorly against resistant strains (16.7% eradication rate; P < 0.01). Using agar dilution testing, we validated the modified Kirby-Bauer disk diffusion method for metronidazole susceptibility testing of H. pylori and conclude that it is practical, accurate, and clinically applicable. PMID:8370723

  7. Evaluation of TA10 Broth for Recovery of Listeria monocytogenes from Ground Beef.

    PubMed

    Kamisaki-Horikoshi, Naoko; Okada, Yukio; Takeshita, Kazuko; Takada, Makoto; Kawamoto, Shinichi; Kawasaki, Susumu

    2017-03-01

    In 2009, the enrichment broth TA10 was released for simultaneous recovery of Salmonella spp., Listeria monocytogenes, and Escherichia coli O157:H7. This medium was compared with other Salmonella enrichment broths [lactose (LAC) broth, buffered peptone water (BPW), and universal pre-enrichment (UP) broth] for the recovery of heat- and freeze-injured Salmonella spp. in beef by the conventional culture method. There was a significant difference between TA10 and LAC enrichment broths for detecting injured Salmonella spp. In this study, the International Organization for Standardization Listeria pre-enrichment broth (Half-Fraser/Fraser) was compared with TA10 broth for the recovery of L. monocytogenes from ground beef. Ground beef samples were contaminated with single Listeria serovars at levels of 0.096 to 0.001 most probable number/g. Twenty 25 g test portions of the contaminated ground beef were pre-enriched in each broth, and the ISO-11290-1 Listeria official isolation protocol was used thereafter. There was a significant difference between TA10 broth (48 h) and Half-Fraser/Fraser broth (72 h) in the recovery of L. monocytogenes. In addition, the incubation time for TA10 broth was shorter than for Half-Fraser/Fraser broth. The results indicate that TA10 broth should be used instead of Half-Fraser/Fraser broth for analysis of beef that may be contaminated with very low levels of L. monocytogenes.

  8. Recovery of butanol from fermentation broth by pervaporation

    USDA-ARS?s Scientific Manuscript database

    Butanol can be produced by fermentation from corn, molasses or lignocellulosic biomass for use as a chemical or superior biofuel. However, butanol’s production is hampered by its toxicity to the microbial culture that produces it. In fermentation broths, final butanol concentrations typically range ...

  9. Antipneumococcal Activities of Levofloxacin and Clarithromycin as Determined by Agar Dilution, Microdilution, E-Test, and Disk Diffusion Methodologies

    PubMed Central

    Clark, Catherine L.; Jacobs, Michael R.; Appelbaum, Peter C.

    1998-01-01

    The activities of levofloxacin and clarithromycin against 199 penicillin- and macrolide-susceptible and -resistant pneumococci were tested by agar and microdilution methods in air and by disk diffusion and E-test methods in air and CO2. For levofloxacin, ≥99.0% of strains were susceptible at ≤2.0 μg/ml with zone diameters of ≥17 mm, regardless of incubation in air or CO2. Although zone sizes were smaller and E-test MICs were higher for clarithromycin in CO2 than those in air, category differences were minor, and susceptibility rates for clarithromycin were similar to those obtained by agar and microdilution in air (range, 76.9 to 80.9% by all methods). For clarithromycin, adjustment of breakpoints based upon distribution of results resulted in susceptibility rates which were similar by all methods (75.8 to 76.9% susceptible, 0 to 1.5% intermediate, 22.6 to 23.1% resistant). Minor discrepancies were obtained with levofloxacin for one strain (0.5%) by microdilution and two strains (1.0%) by disk diffusion in CO2. For clarithromycin, minor discrepancies were found in three strains (1.5%) by microdilution, seven strains (3.5%) by agar dilution, four strains (2.0%) by E-test in air, six strains (3.0%) by disk diffusion in air, and five strains (2.5%) by disk diffusion in CO2. Major discrepancies occurred with levofloxacin in one strain (0.5%) by microdilution but were not found with clarithromycin. Very major discrepancies were not seen with levofloxacin, but occurred with clarithromycin in five strains (2.5%) by microdilution, three strains (1.5%) by agar dilution, two strains (1.0%) by E-test in air, eight strains (4.0%) by disk diffusion in air, and one strain (0.5%) by disk diffusion in CO2. PMID:9817876

  10. Inhibition of Nanobacteria by Antimicrobial Drugs as Measured by a Modified Microdilution Method

    PubMed Central

    Çíftçíoglu, N.; Miller-Hjelle, M. A.; Hjelle, J. T.; Kajander, E. O.

    2002-01-01

    Compounds from 16 classes of antimicrobial drugs were tested for their abilities to inhibit the in vitro multiplication of nanobacteria (NB), a newly discovered infectious agent found in human kidney stones and kidney cyst fluids from patients with polycystic kidney disease (PKD). Because NB form surface calcifications at physiologic levels of calcium and phosphate, they have been hypothesized to mediate the formation of tissue calcifications. We describe a modified microdilution inhibitory test that accommodates the unique growth conditions and long multiplication times of NB. This modified microdilution method included inoculation of 96-well plates and determination of inhibition by periodic measurement of the absorbance for 14 days in cell culture medium under cell culture conditions. Bactericidal or bacteriostatic drug effects were distinguished by subsequent subculture in drug-free media and monitoring for increasing absorbance. NB isolated from fetal bovine serum (FBS) were inhibited by tetracycline HCl, nitrofurantoin, trimethoprim, trimethoprim-sulfamethoxazole, and ampicillin at levels achievable in serum and urine; all drugs except ampicillin were cidal. Tetracycline also inhibited multiplication of isolates of NB from human kidney stones and kidney cyst fluids from patients with PKD. The other antibiotics tested against FBS-derived NB either had no effect or exhibited an inhibitory concentration above clinically achievable levels; the aminoglycosides and vancomycin were bacteriostatic. Antibiotic-induced morphological changes to NB were observed by electron microscopy. Bisphosphonates, aminocaproic acid, potassium citrate-citric acid solutions, and 5-fluorouracil also inhibited the multiplication of NB in a cidal manner. Insights into the nature of NB, the action(s) of these drugs, and the role of NB in calcifying diseases may be gained by exploiting this in vitro inhibition test system. PMID:12069958

  11. Separation of gamma-aminobutyric acid from fermented broth.

    PubMed

    Li, Haixing; Qiu, Ting; Chen, Yan; Cao, Yusheng

    2011-12-01

    Gamma-aminobutyric acid (GABA) is a non-proteinaceous amino acid that is widely distributed in nature and acts as the major inhibitory neurotransmitter in the mammalian brain. This study aimed to find a separation method for getting high-purity GABA from a fermented broth. Firstly, a fermented broth with a high content of GABA (reaching 997 ± 51 mM) was prepared by fermentation with Lactobacillus brevis NCL912. GABA purification was conducted by successive centrifugation, filtration, decoloration, desalination, ion-exchange chromatography (IEC), and crystallization. Inorganic salt (Na₂SO₄) was removed from the both by desalination with 70% ethanol solution. A ninhydrin test strip was designed for the real-time detection of GABA during IEC. The recovery rate for the whole purification process was about 50%. The purified product was characterized by thin-layer chromatography and HPLC, and its purity reached 98.66 ± 2.36%.

  12. Green synthesis of gold nanoparticles using Cinnamomum zeylanicum leaf broth.

    PubMed

    Smitha, S L; Philip, Daizy; Gopchandran, K G

    2009-10-15

    Development of biologically inspired experimental processes for the synthesis of nanoparticles is an important branch of nanotechnology. The synthesis of gold nanoparticles using Cinnamomum zeylanicum leaf broth as the reducing agent is reported. The morphology of the particles formed consists of a mixture of gold nanoprisms and spheres with fcc (111) structure of gold. At lower concentrations of the extract, formation of prism shaped Au particles dominates, while at higher concentrations almost spherical particles alone are observed. Good crystallinity of the nanoparticles with fcc phase is evident from XRD patterns, clear lattice fringes in the high resolution TEM image and bright circular rings in the SAED pattern. Au nanoparticles grown are observed to be photoluminescent and the intensity of photoemission is found to increase with increase in leaf broth concentration. The ability to modulate the shape of nanoparticles as observed in this study for gold nanoparticles opens up the exciting possibility of developing further synthetic routes employing ecofriendly sources.

  13. Reverse Osmosis Processing of Organic Model Compounds and Fermentation Broths

    DTIC Science & Technology

    2006-04-01

    key species found in the fermentation broth: ethanol, butanol, acetic acid, oxalic acid, lactic acid, and butyric acid. Correlations of the rejection...into the feed tank of the RO system prior to the RO experiment. Ethanol, butanol, acetic acid, lactic acid, oxalic acid and butyric acid were used as...into a plastic syringe and filtered through a cartridge filter (Lida Manufacturing Corp. 0.45 lm hydrophilic cellulose acetate membrane) into a TOC

  14. Enhanced recovery of Salmonella from apple cider and apple juice with universal preenrichment broth.

    PubMed

    Hammack, Thomas S; Johnson, Mildred L; Jacobson, Andrew P; Andrews, Wallace H

    2002-01-01

    A comparison was made of the relative efficiencies of Universal Preenrichment (UP) broth and lactose broth for the recovery of a variety of Salmonella serovars from pasteurized and unpasteurized apple cider and pasteurized apple juice. Bulk portions of juice were contaminated with single Salmonella serovars at high and low levels of 0.4 and 0.04 CFU/mL, respectively. The juice was aged for a minimum of 5 days at 2-5 degrees C. On the day analysis was initiated, each of 20 test portions (25 mL) of the contaminated juice was preenriched in UP broth and in lactose broth. The Bacteriological Analytical Manual Salmonella culture method was followed thereafter. For pasteurized apple cider, UP broth recovered significantly (p < 0.05) more Salmonella-positive test portions than did lactose broth (112 and 75, respectively). For unpasteurized apple cider, UP broth recovered significantly more Salmonella-positive test portions than did lactose broth (326 and 221, respectively). For pasteurized apple juice, UP broth recovered more Salmonella-positive test portions than did lactose broth (93 and 81, respectively). However, this difference was not statistically significant. These results indicate that UP broth should replace lactose broth for the analysis of pasteurized and unpasteurized apple cider and pasteurized apple juice.

  15. Selective enumeration of propionibacteria in Emmental-type cheese using Petrifilm™ aerobic count plates added to lithium glycerol broth.

    PubMed

    de Freitas, Rosângela; Luiz, Lívia M Pinheiro; Alves, Maura Pinheiro; Valence-Bertel, Florence; Nero, Luís Augusto; de Carvalho, Antônio Fernandes

    2013-08-01

    Propionibacteria derived from dairy products are relevant starter cultures for the production of Swiss and Emmental-type cheeses, and the monitoring of which is mandatory for proper quality control. This study aimed to evaluate an alternative procedure to enumerate propionibacteria, in order to develop a reliable and practical methodology to be employed by dairy industries. 2,3,5-triphenyltetrazolium chloride (TTC) inhibitory activity was tested against five reference strains (CIRM 09, 38, 39, 40 and 116); TTC at 0·0025% (w/v) was not inhibitory, with the exception of one strain (CIRM 116). Subsequently, the four TTC-resistant strains, three commercial starter cultures (PS-1, PB-I, and CHOO) and twelve Emmental-type cheese samples were subjected to propionibacteria enumeration using Lithium Glycerol (LG) agar, and Petrifilm™ Aerobic Count (AC) plates added to LG broth (anaerobic incubation at 30 °C for 7 d). Petrifilm™ AC added to LG broth presented high counts than LG agar (P<0·05) for only two reference strains (CIRM 39, and 40) and for all commercial starter cultures. Cheese sample counts obtained by both procedures did not show significant differences (P<0·05). Significant correlation indexes were observed between the counts recorded by both methods (P<0·05). These results demonstrate the reliability of Petrifilm™ AC plates added to LG broth in enumerating select Propionibacterium spp., despite some limitations observed for specific commercial starter cultures.

  16. Removing Bacillus subtilis from fermentation broth using alumina nanoparticles.

    PubMed

    Mu, Dashuai; Mu, Xin; Xu, Zhenxing; Du, Zongjun; Chen, Guanjun

    2015-12-01

    In this study, an efficient separation technology using Al2O3 nanoparticles (NPs) was developed for removing Bacillus subtilis from fermentation broth. The dosage of alumina nanoparticles used for separating B. subtilis increased during the culture process and remained stable in the stationary phase of the culture process. The pH of the culture-broth was also investigated for its effects on flocculation efficiency, and showed an acidic pH could enhance the flocculation efficiency. The attachment mechanisms of Al2O3 NPs to the B. subtilis surface were investigated, and the zeta potential analysis showed that Al2O3 NPs could attach to B. subtilis via electrostatic attachment. Finally, the metabolite content and the antibacterial effect of the fermentation supernatants were detected and did not significantly differ between alumina nanoparticle separation and centrifugation separation. Together, these results indicate a great potential for a highly efficient and economical method for removing B. subtilis from fermentation broth using alumina nanoparticles. Copyright © 2015 Elsevier Ltd. All rights reserved.

  17. An improved method for the recovery of Salmonella serovars from orange juice using universal preenrichment broth.

    PubMed

    Hammack, T S; Amaguaña, R M; Andrews, W H

    2001-05-01

    The relative effectiveness of three methods for the recovery of Salmonella serovars from orange juice was determined. One method, a modified Bacteriological Analytical Manual (BAM) procedure consisted of preenrichment in lactose broth at 35 degrees C for 24 h, selective enrichment, and selective plating. Another method, a National Centers for Disease Control and Prevention (CDC 1) procedure, consisted of direct enrichment in tetrathionate broth at 35 degrees C for 24 and 48 h, followed by selective plating. The third method (also from CDC and designated CDC 2) consisted of preenrichment in Universal Preenrichment (UP) broth at 35 degrees C for 24 h, selective enrichment, and selective plating. In 10 experiments encompassing five different Salmonella serovars and 200 test portions per broth, the CDC 1 method recovered 141 Salmonella-positive test portions, the BAM method recovered 151, and the CDC 2 method recovered 171. In 2 of the 10 experiments, with two different Salmonella serovars, the BAM recovered significantly fewer (P < 0.05) Salmonella-positive test portions than did the CDC 2 method. On the basis of the above results, the second phase of this study focused on a comparison of the effectiveness of the BAM-recommended lactose broth and the CDC 2-recommended UP broth as preenrichment media for the recovery of Salmonella serovars from pasteurized and unpasteurized orange juice. Subsequent culture treatment of the two preenrichments was identical so that the effect of other variables (e.g., different selective enrichment media, various incubation temperatures, and different selective plating agars) on the relative performance of these two preenrichment media was excluded. In one of nine experiments, with pasteurized orange juice, lactose broth recovered significantly fewer (P < 0.05) Salmonella-positive test portions than did UP broth. For the combined results of the nine pasteurized orange juice experiments (180 test portions per broth), lactose broth

  18. Paradigm Diagnostics Salmonella Indicator Broth (PDX-SIB) for detection of Salmonella on selected environmental surfaces.

    PubMed

    Olstein, Alan; Griffith, Leena; Feirtag, Joellen; Pearson, Nicole

    2013-01-01

    The Paradigm Diagnostics Salmonella Indicator Broth (PDX-SIB) is intended as a single-step selective enrichment indicator broth to be used as a simple screening test for the presence of Salmonella spp. in environmental samples. This method permits the end user to avoid multistep sample processing to identify presumptively positive samples, as exemplified by standard U.S. reference methods. PDX-SIB permits the outgrowth of Salmonella while inhibiting the growth of competitive Gram-negative and -positive microflora. Growth of Salmonella-positive cultures results in a visual color change of the medium from purple to yellow when the sample is grown at 37 +/- 1 degree C. Performance of PDX-SIB has been evaluated in five different categories: inclusivity-exclusivity, methods comparison, ruggedness, lot-to-lot variability, and shelf stability. The inclusivity panel included 100 different Salmonella serovars, 98 of which were SIB-positive during the 30 to 48 h incubation period. The exclusivity panel included 33 different non-Salmonella microorganisms, 31 of which were SIB-negative during the incubation period. Methods comparison studies included four different surfaces: S. Newport on plastic, S. Anatum on sealed concrete, S. Abaetetuba on ceramic tile, and S. Typhimurium in the presence of 1 log excess of Citrobacter freundii. Results of the methods comparison studies demonstrated no statistical difference between the SIB method and the U.S. Food and Drug Administration-Bacteriological Analytical Manual reference method, as measured by the Mantel-Haenszel Chi-square test. Ruggedness studies demonstrated little variation in test results when SIB incubation temperatures were varied over a 34-40 degrees C range. Lot-to-lot consistency results suggest no detectable differences in manufactured goods using two reference Salmonella serovars and one non-Salmonella microorganism.

  19. Production of 1,3-propanediol by Klebsiella pneumoniae from glycerol broth.

    PubMed

    Cheng, Ke-Ke; Zhang, Jian-An; Liu, De-Hua; Sun, Yan; Yang, Ming-De; Xu, Jing-Ming

    2006-11-01

    Broth containing 152 g glycerol l(-1) from Candida krusei culture was converted to 1,3-propanediol by Klebsiella pneumoniae. Residual glucose in the broth promoted growth of K. pneumoniae while acetate was inhibitory. After desalination treatment of glycerol broth by electrodialysis, the acetate in the broth was removed. A fed-batch culture with electrodialytically pretreated broth as substrate was developed giving 53 g 1,3-propanediol l(-1) with a yield of 0.41 g g(-1) glycerol and a productivity of 0.94 g l(-1) h(-1).

  20. Novel Microdilution Method to Assess Double and Triple Antibiotic Combination Therapy In Vitro.

    PubMed

    El-Azizi, Mohamed

    2016-01-01

    An in vitro microdilution method was developed to assess double and triple combinations of antibiotics. Five antibiotics including ciprofloxacin, amikacin, ceftazidime, piperacillin, and imipenem were tested against 10 clinical isolates of Pseudomonas aeruginosa. Each isolate was tested against ten double and nine triple combinations of the antibiotics. A 96-well plate was used to test three antibiotics, each one alone and in double and triple combinations against each isolate. The minimum bacteriostatic and bactericidal concentrations in combination were determined with respect to the most potent antibiotic. An Interaction Code (IC) was generated for each combination, where a numerical value was designated based on the 2-fold increase or decrease in the MICs with respect to the most potent antibiotic. The results of the combinations were verified by time-kill assay at constant concentrations of the antibiotics and in a chemostat. Only 13% of the double combinations were synergistic, whereas 5% showed antagonism. Forty-three percent of the triple combinations were synergistic with no antagonism observed, and 100% synergism was observed in combination of ciprofloxacin, amikacin, and ceftazidime. The presented protocol is simple and fast and can help the clinicians in the early selection of the effective antibiotic therapy for treatment of severe infections.

  1. On-line extraction of ethanol from fermentation broths using hydrophobic adsorbents

    SciTech Connect

    Lencki, R.W.; Robinson, C.W.; Moo-Young, M.

    1983-01-01

    The effect of removing ethanol during fermentation on the batch kinetics of glucose utilization by the yeast Saccharomyces cerevisiae was examined using several hydrophobic adsorbents. The addition of the molecular sieve, silicalite, to fermentation broths greatly reduced the concentration of ethanol present, but did not increase the glucose utilization rate to the extent predicted by product-inhibition kinetic models obtained from adsorbent-free continuous culture experiments. Addition of two polymeric adsorbents (XAD-4 and XAD-7) greatly inhibited cell growth. This was due to nutrient and cell adsorption by the resin. Adsorption systems were compared to other on-line schemes (vacuferm, solvent extraction) and it was concluded that all of these currently proposed systems would not appear to confer any added advantage if integrated into a high biomass concentration fermenter system. 19 references, 7 figures, 1 table.

  2. Bromothymol blue broth: improved medium for detection of Ureaplasma urealyticum (T-strain mycoplasma).

    PubMed

    Robertson, J A

    1978-02-01

    Bromothymol blue (B) broth for the cultivation, detection, and identification of Ureaplasma urealyticum is described. In this medium, strains Cook and 960 had shorter generation times (60 min or less) and reached higher populations (over 10(8)) than have yet been reported for this species. Furthermore, the indicator changes color before the end of logarithmic growth, and the cultures retain viability for at least 1 day thereafter, greatly simplifying the handling of the organism. When the populations in cultures of these two strains and seven new isolates were determined, growth was detected earlier and proceeded to higher final titers in B broth than in urease test color medium (U-9 broth). The inclusion of antibiotics in B broth for use in clinical laboratories (B/NL broth) made the medium selective, specific, and more sensitive for the isolation of U. urealyticum. Comparison of B/NL broth with genital mycoplasma (GM) agar and U-9 broth for the primary isolation of U. urealyticum was made with 183 urethral swabs. All 70 isolates were detected on B/NL broth, but only 66 and 63 isolates were detected on GM agar and in U-9 broth, respectively. Moreover, the cultures in B/NL broth were pure and at titers that generally showed good correlation with colony counts on GM agar.

  3. Evaluation of TA10 broth for recovery of heat- and freeze-injured Salmonella from beef.

    PubMed

    Kamisaki-Horikoshi, Naoko; Okada, Yukio; Takeshita, Kazuko; Sameshima, Takashi; Kawasaki, Susumu; Kawamoto, Shinichi; Fratamico, Pina M

    2011-01-01

    The Bacteriological Analytical Manual (BAM) Salmonella pre-enrichment broth [lactose (LAC) broth], buffered peptone water, and universal pre-enrichment (UP) broth were compared with TA10 broth, developed in our laboratory, for recovery of heat- and freeze-injured Salmonella (55 degrees C for 2-20 min and -20 degrees C for 2 months, respectively) from beef. Beef samples were contaminated with single Salmonella serovars, and contamination levels of 0.44 to <0.001 most probable number (MPN)/g and 0.74 to 0.14 MPN/g were used for heat- and freezing-induced injury studies, respectively. Twenty test portions (25 g) of the contaminated beef were pre-enriched in each broth, and the BAM Salmonella culture method was used thereafter. There was a significant difference (chi2 = 7.73) in recovery of heat-injured Salmonella between TA10 broth and LAC broth, 189 (67.5%) versus 156 (55.7%) positive samples, respectively, determined by plating onto selective agars and identification by biochemical tests. For the recovery of freeze-injured Salmonella, there was a significant difference (chi2 = 24.7) between TA10 and LAC broth, 189 (72.7%) versus 133 (51.2%) positive samples, respectively. TA10 broth was more effective than LAC broth and UP broth for recovery of freeze-injured Salmonella. The results indicate that TA10 broth should be used instead of LAC broth for testing of beef that may be contaminated with heat- and freeze-injured Salmonella spp.

  4. The visual assessment of broth cultures for tissue bank samples.

    PubMed

    Varettas, Kerry

    2017-01-05

    The bioburden screening process of allograft musculoskeletal tissue samples received at the South Eastern Area Laboratory Services includes the routine use of solid agar and cooked meat (CM) broth media. CM has been routinely sub-cultured onto solid agar plates after aerobic incubation at 35 °C. This study will evaluate whether a visual assessment of CM can replace sub-culture by an in vitro inoculation and a prospective study. Eight challenge organisms were serially diluted and inoculated into CM. The average inoculum of 0.5-5.5 CFU produced visible turbidity of CM after 24-h incubation for 7 of the challenge organisms with one organism producing turbidity after 48-h incubation. The prospective study evaluated 222 CM of which 213 were visually clear and no-growth on sub-culture and 9 turbid CM which were culture positive. Broth cultures are an integral part of the bioburden screening process of allograft musculoskeletal tissue and swab samples and visual assessment of CM can replace sub-culture.

  5. Microbiologic and clinical value of primary broth cultures of wound specimens collected with swabs.

    PubMed Central

    Silletti, R P; Ailey, E; Sun, S; Tang, D

    1997-01-01

    In order to assess the microbiologic and clinical value of primary broth culture of wound specimens collected with swabs and submitted to the laboratory in transport medium, we compared the results of primary agar culture with the results of a corresponding primary broth culture for 344 aerobic specimens and 176 anaerobic specimens. While 8.7% (45 of 520) of the specimens yielded organisms from the primary broth culture that were not recovered from the corresponding primary agar culture, only 5.0% (26 of 520) of the specimens yielded organisms from the primary broth culture other than Staphylococcus epidermidis, viridans group streptococci, and Corynebacterium spp. Moreover, the primary broth culture of only 0.6% (3 of 520) of the specimens yielded organisms not recovered from the primary agar culture that caused a change in the therapy of the patient. Our conclusion is that primary broth cultures are unnecessary for the processing of wound specimens properly collected with swabs. PMID:9230370

  6. Recovery of Salmonella by Using Selenite Brilliant Green Sulfa Enrichment Broth

    PubMed Central

    Chang, Chiao-tang; Yuo, Chung-Yee; Shen, Hui-Ching; Li, A-Mai; Chen, Chao-yu; Chou, Jui-ling; Huang, Shiao-ping

    1999-01-01

    The efficacy and sensitivity of selenite brilliant green sulfa enrichment (SBG) broth for the isolation of Salmonella from fecal specimens were evaluated by using both clinical and artificially infected (artificial) fecal specimens. An examination of 1,588 clinical fecal specimens found Salmonella in 296 specimens, including 89 cases detected by the direct-plating xylose-lysine-desoxycholate method and an additional 207 cases detected after enrichment with SBG broth. Therefore, the recovery of Salmonella with SBG broth is increased 3.3-fold over that by the direct-plating method alone. Furthermore, the isolation rate of Salmonella is higher when using SBG broth than when using gram-negative (GN) broth or GN broth supplemented with sodium selenite. To determine the sensitivity for the recovery of Salmonella, artificial specimens containing various amounts of Salmonella were prepared and analyzed. The results indicated that the sensitivity is also higher with SBG broth than with GN broth. Moreover, the optimal incubation period for SBG broth can be extended to 24 h. In conclusion, the SBG enrichment method provides a higher recovery rate of Salmonella from fecal specimens. PMID:10565941

  7. Antimicrobial activity of broth fermented with kefir grains.

    PubMed

    Silva, Karoline R; Rodrigues, Sheila A; Filho, Lauro Xavier; Lima, Alvaro S

    2009-02-01

    Kefir grains originate from the Caucasus region and are used for preparing beverages using sugar solution, milk, and fruit juice. As long as they are formed by a microbial consortium useful in the intestine, the produced drinks can be called probiotics. The aim of this study was to determine the antimicrobial activity during kefir fermentation in sugar broth. Fermentations with three kinds of carbohydrates (molasses, demerara sugar, and brown sugar) as carbon source were carried out. Brown sugar promoted the greatest antimicrobial activities, producing inhibition halos corresponding to 35, 14, 12, 14, and 14 mm for Candida albicans, Salmonella typhi, Shigella sonnei, Staphylococcus aureus, and Escherichia coli, respectively. Different carbon source concentrations and the time of fermentation influenced the size of the inhibition halos of the pathogenic microorganisms.

  8. Glycerol Monolaurate Antibacterial Activity in Broth and Biofilm Cultures

    PubMed Central

    Schlievert, Patrick M.; Peterson, Marnie L.

    2012-01-01

    Background Glycerol monolaurate (GML) is an antimicrobial agent that has potent activity against gram-positive bacteria. This study examines GML antibacterial activity in comparison to lauric acid, in broth cultures compared to biofilm cultures, and against a wide range of gram-positive, gram-negative, and non-gram staining bacteria. Methodology/Principal Findings GML is ≥200 times more effective than lauric acid in bactericidal activity, defined as a ≥3 log reduction in colony-forming units (CFU)/ml, against Staphylococcus aureus and Streptococcus pyogenes in broth cultures. Both molecules inhibit superantigen production by these organisms at concentrations that are not bactericidal. GML prevents biofilm formation by Staphylococcus aureus and Haemophilus influenzae, as representative gram-positive and gram-negative organisms, tested in 96 well microtiter plates, and simultaneously is bactericidal for both organisms in mature biofilms. GML is bactericidal for a wide range of potential bacterial pathogens, except for Pseudomonas aeruginosa and Enterobacteriaceae. In the presence of acidic pH and the cation chelator ethylene diamine tetraacetic acid, GML has greatly enhanced bactericidal activity for Pseudomonas aeruginosa and Enterobacteriaceae. Solubilization of GML in a nonaqueous delivery vehicle (related to K-Y Warming®) enhances its bactericidal activity against S. aureus. Both R and S, and 1 and 2 position lauric acid derivatives of GML exhibit bactericidal activity. Despite year-long passage of Staphylococcus aureus on sub-growth inhibitory concentrations of GML (0.5 x minimum bactericidal concentration), resistance to GML did not develop. Conclusions/Significance GML may be useful as a broad-spectrum human or animal topical microbicide and may be useful as an environmental surface microbicide for management of bacterial infections and contamination. PMID:22808139

  9. Rheology and hydrodynamic properties of Tolypocladium inflatum fermentation broth and its simulation.

    PubMed

    Benchapattarapong, N; Anderson, W A; Bai, F; Moo-Young, M

    2005-07-01

    A physico-chemical, two phase simulated pseudoplastic fermentation (SPF) broth was investigated in which Solka Floc cellulose fibre was used to simulate the filamentous biomass, and a mixture of 0.1% (w/v) carboxymethyl cellulose (CMC) and 0.15 M aqueous sodium chloride was used to simulate the liquid fraction of the fermentation broth. An investigation of the rheological behaviour and hydrodynamic properties of the SPF broth was carried out, and compared to both a fungal Tolypocladium inflatum fermentation broth and a CMC solution in a 50 L stirred tank bioreactor equipped with conventional Rushton turbines. The experimental data confirmed the ability of the two phase SPF broth to mimic both the T. inflatum broth bulk rheology as well as the mixing and mass transfer behaviour. In contrast, using a homogeneous CMC solution with a similar bulk rheology to simulate the fermentation resulted in a significant underestimation of the mass transfer and mixing times. The presence of the solid phase and its microstructure in the SPF broth appear to play a significant role in gas holdup and bubble size, thus leading to the different behaviours. The SPF broth seems to be a more accurate simulation fluid that can be used to predict the bioreactor mixing and mass transfer performance in filamentous fermentations, in comparison with CMC solutions used in some previous studies.

  10. Membrane fouling mechanism in ultrafiltration of succinic acid fermentation broth.

    PubMed

    Wang, Caixia; Li, Qiang; Tang, Huang; Yan, Daojiang; Zhou, Wei; Xing, Jianmin; Wan, Yinhua

    2012-07-01

    The membrane fouling mechanism was studied in treating succinic acid fermentation broth during dead-end ultrafiltration. Different membranes were used and two models were applied to analyze the fouling mechanism. Resistance-in-series model was applied to determine the main factor that caused the operation resistance. Results indicated that most membranes tended to be fouled by cake layer or concentration polarization. Hermia's model, which is composed of four individual sub-models, was used to analyze the predominant fouling mechanism. Results showed that the fouling of RC 10 kDa and PES 30 kDa was controlled by the complete blocking mechanism, while PES 100 kDa was controlled by the intermediate blocking and PES 10 kDa was controlled by cake layer. This conclusion was also proved by SEM photos. Membrane characteristics were monitored before and after ultrafiltration by AFM and goniometer. Both contact angle and roughness of most membranes increased after ultrafiltration. Copyright © 2012 Elsevier Ltd. All rights reserved.

  11. Filtration of a bacterial fermentation broth: harvest conditions effects on cake hydraulic resistance.

    PubMed

    Meireles, M; Lavoute, E; Bacchin, P

    2003-03-01

    The hydraulic resistance of cakes formed during the ultrafiltration of Streptomyces pristinaespiralis broths has been investigated for different harvesting conditions. S. pristinaespiralis broth was harvested after the point of microorganism activity declines (0-h aged broth) and afterwards held for different durations of up to 16 h (16 aged broths). Aging behavior occurring between the end of microorganism activity and harvest was compared for different acidification procedures (pH) and the mechanisms for which the hydraulic resistance of the cake is affected by aging have been investigated. For broths harvested under conditions where the acidification is fixed at pH 2 or 3, hydraulic resistance associated with cake build-up is directly determined by the interactions between the cells. Holding broths beyond 5 h contributes to a release of a soluble component from the cell surface. Enhanced cell surface interactions then turn the cake structure into a more open one and reduce the specific hydraulic resistance. For broths harvested under conditions where the acidification is fixed at pH 4, hydraulic resistance associated with cake build-up is both determined by cell interactions and cell morphology. The cause of the increase in specific hydraulic resistance with aging is due to the binding of a soluble component released by the microorganisms, which decreases the cell surface interactions.

  12. Supplementation of Bolton broth with triclosan improves detection of Campylobacter jejuni and Campylobacter coli in chicken carcass rinse.

    PubMed

    Chon, Jung-Whan; Kim, Young-Ji; Kim, Hong-Seok; Kim, Dong-Hyeon; Kim, Hyunsook; Song, Kwang-Young; Seo, Kun-Ho

    2014-07-02

    We compared Bolton enrichment broth supplemented with antimicrobial triclosan (T-Bolton broth) and normal Bolton broth for the isolation of Campylobacter jejuni (C. jejuni) and Campylobacter coli (C. coli) from chicken carcass rinse. Whole chickens were rinsed with buffered peptone water prior to enrichment in normal Bolton broth or T-Bolton broth, followed by inoculation onto modified charcoal-cefoperazone-deoxycholate agar (mCCDA). Suspect colonies were confirmed by PCR. We observed a significantly higher number of C. jejuni or C. coli-positive samples in the T-Bolton broth (71.3%) than in the normal Bolton broth (27.5%) (p<0.05). Furthermore, the number of contaminated mCCDA plates was lower after enrichment in T-Bolton broth (3.8%) than in the normal Bolton broth (75%) (p<0.05), indicating that T-Bolton broth has higher selectivity. Finally, we identified extended-spectrum β-lactamase-producing Escherichia coli as the predominant competing flora in normal Bolton broth. In conclusion, the use of T-Bolton broth results in significant elimination of competing bacteria. Copyright © 2014 Elsevier B.V. All rights reserved.

  13. Effect of fermented broth from lactic acid bacteria on pathogenic bacteria proliferation.

    PubMed

    Gutiérrez, S; Martínez-Blanco, H; Rodríguez-Aparicio, L B; Ferrero, M A

    2016-04-01

    In this study, the effect that 5 fermented broths of lactic acid bacteria (LAB) strains have on the viability or proliferation and adhesion of 7 potentially pathogenic microorganisms was tested. The fermented broth from Lactococcus lactis C660 had a growth inhibitory effect on Escherichia coli K92 that reached of 31%, 19% to Pseudomonas fluorescens, and 76% to Staphylococcus epidermidis. The growth of Staph. epidermidis was negatively affected to 90% by Lc. lactis 11454 broth, whereas the growth of P. fluorescens (25%) and both species of Staphylococcus (35% to Staphylococcus aureus and 76% to Staph. epidermidis) were inhibited when they were incubated in the presence of Lactobacillus casei 393 broth. Finally, the fermented broth of Lactobacillus rhamnosus showed an inhibitory effect on growth of E. coli K92, Listeria innocua, and Staph. epidermidis reached values of 12, 28, and 76%, respectively. Staphylococcus epidermidis was the most affected strain because the effect was detected from the early stages of growth and it was completely abolished. The results of bacterial adhesion revealed that broths from Lc. lactis strains, Lactobacillus paracasei, and Lb. rhamnosus caused a loss of E. coli K92 adhesion. Bacillus cereus showed a decreased of adhesion in the presence of the broths of Lc. lactis strains and Lb. paracasei. Listeria innocua adhesion inhibition was observed in the presence of Lb. paracasei broth, and the greatest inhibitory effect was registered when this pathogenic bacterium was incubated in presence of Lc. lactis 11454 broth. With respect to the 2 Pseudomonas, we observed a slight adhesion inhibition showed by Lactobacillus rhamnosus broth against Pseudomonas putida. These results confirm that the effect caused by the different LAB assayed is also broth- and species-specific and reveal that the broth from LAB tested can be used as functional bioactive compounds to regulate the adhesion and biofilm synthesis and ultimately lead to preventing food and

  14. Succinic acid adsorption from fermentation broth and regeneration.

    PubMed

    Davison, Brian H; Nghiem, Nhuan P; Richardson, Gerald L

    2004-01-01

    More than 25 sorbents were tested for uptake of succinic acid from aqueous solutions. The best resins were then tested for successive loading and regeneration using hot water. The key desired properties for an ideal sorbent are high capacity, complete stable regenerability, and specificity for the product. The best resins have a stable capacity of about 0.06 g of succinic acid/g of resin at moderate concentrations (1-5 g/L) of succinic acid. Several sorbents were tested more exhaustively for uptake of succinic acid and for successive loading and regeneration using hot water. One resin, XUS 40285, has a good stable isotherm capacity, prefers succinate over glucose, and has good capacities at both acidic and neutral pH. Succinic acid was removed from simulated media containing salts, succinic acid, acetic acid, and sugar using a packed column of sorbent resin, XUS 40285. The fermentation byproduct, acetate, was completely separated from succinate. A simple hot water regeneration successfully concentrated succinate from 10 g/L (inlet) to 40-110 g/L in the effluent. If successful, this would lower separation costs by reducing the need for chemicals for the initial purification step. Despite promising initial results of good capacity (0.06 g of succinic/g of sorbent), 70% recovery using hot water, and a recovered concentration of >100 g/L, this regeneration was not stable over 10 cycles in the column. Alternative regeneration schemes using acid and base were examined. Two (XUS 40285 and XFS-40422) showed both good stable capacities for succinic acid over 10 cycles and >95% recovery in a batch operation using a modified extraction procedure combining acid and hot water washes. These resins showed comparable results with actual broth.

  15. Comparative performance of isolation methods using Preston broth, Bolton broth and their modifications for the detection of Campylobacter spp. from naturally contaminated fresh and frozen raw poultry meat.

    PubMed

    Seliwiorstow, T; De Zutter, L; Houf, K; Botteldoorn, N; Baré, J; Van Damme, I

    2016-10-03

    The performance of different isolation methods was evaluated for the detection of Campylobacter from naturally contaminated raw poultry meat. Therefore, fresh and frozen poultry meat samples were analysed using the standard procedure (ISO 10272-1:2006), enrichment in Preston broth, and enrichment in modified Bolton broth (supplemented with (i) potassium clavulanate (C-BB), (ii) triclosan (T-BB), (iii) polymyxin B (P-BB)). The enrichment cultures were streaked onto both modified charcoal cefoperazone deoxycholate agar (mCCDA) and RAPID'Campylobacter agar (RCA). Moreover, direct plating on mCCDA and RCA was performed to quantify Campylobacter. In total, 33 out of 59 fresh retail meat samples (55.9%) were Campylobacter positive. For both fresh and frozen poultry meat samples, enrichment in Bolton broth (ISO 10272-1:2006) resulted in a higher number of positive samples than enrichment in Preston broth. Supplementation of Bolton broth with potassium clavulanate (C-BB) and triclosan (T-BB) enhanced the Campylobacter recovery from fresh poultry meat compared to non-supplemented Bolton broth, although the use of C-BB was less applicable than T-BB for Campylobacter recovery from frozen samples. Additionally, the use of RCA resulted in a higher isolation rate compared to mCCDA. The present study demonstrates the impact of culture medium on the recovery of Campylobacter from fresh and frozen naturally contaminated poultry meat samples and can support laboratories in choosing the most appropriate culturing method to detect Campylobacter. Copyright © 2016 Elsevier B.V. All rights reserved.

  16. [Kinetic simulation of enhanced biological phosphorus removal with fermentation broth as carbon source].

    PubMed

    Zhang, Chao; Chen, Yin-Guang

    2013-07-01

    As a high-quality carbon source, fermentation broth could promote the phosphorus removal efficiency in enhanced biological phosphorus removal (EBPR). The transformation of substrates in EBPR fed with fermentation broth was well simulated using the modified activated sludge model No. 2 (ASM2) based on the carbon source metabolism. When fermentation broth was used as the sole carbon source, it was found that heterotrophic bacteria acted as a promoter rather than a competitor to the phosphorus accumulating organisms (PAO). When fermentation broth was used as a supplementary carbon source of real municipal wastewater, the wastewater composition was optimized for PAO growth; and the PAO concentration, which was increased by 3.3 times compared to that in EBPR fed with solely real municipal wastewater, accounting for about 40% of the total biomass in the reactor.

  17. Energy efficient recovery and dehydration of ethanol from fermentation broths by Membrane Assisted Vapor Stripping technology

    EPA Science Inventory

    Distillation combined with molecular sieve dehydration is the current state of the art for fuel grade ethanol production from fermentation broths. To improve the sustainability of bioethanol production, energy efficient separation alternatives are needed, particularly for lower ...

  18. Separation technologies for the recovery and dehydration of alcohols from fermentation broths

    EPA Science Inventory

    Multi-column distillation followed by molecular sieve adsorption is currently the standard method for producing fuel grade ethanol from dilute fermentation broths in modern corn-to-ethnol facilities. As the liquid biofuels industry transitions to lignocellulosic feedstocks, expan...

  19. Membrane-based recovery and dehydration of alcohols from fermentation broths - of materials and modules

    EPA Science Inventory

    Distillation combined with molecular sieve dehydration is the current state of the art for fuel grade ethanol production from fermentation broths. As the liquid biofuels industry transitions to lignocellulosic feedstocks, expands the end product portfolio to include other alcoho...

  20. Membrane-based recovery and dehydration of alcohols from fermentation broths - of materials and modules

    EPA Science Inventory

    Distillation combined with molecular sieve dehydration is the current state of the art for fuel grade ethanol production from fermentation broths. As the liquid biofuels industry transitions to lignocellulosic feedstocks, expands the end product portfolio to include other alcoho...

  1. Separation technologies for the recovery and dehydration of alcohols from fermentation broths

    EPA Science Inventory

    Multi-column distillation followed by molecular sieve adsorption is currently the standard method for producing fuel grade ethanol from dilute fermentation broths in modern corn-to-ethnol facilities. As the liquid biofuels industry transitions to lignocellulosic feedstocks, expan...

  2. Energy efficient recovery and dehydration of ethanol from fermentation broths by Membrane Assisted Vapor Stripping technology

    EPA Science Inventory

    Distillation combined with molecular sieve dehydration is the current state of the art for fuel grade ethanol production from fermentation broths. To improve the sustainability of bioethanol production, energy efficient separation alternatives are needed, particularly for lower ...

  3. SST broth, a new serum free germ tube induction medium for identification of Candida albicans.

    PubMed

    Raghunath, Pendru; Seshu Kumari, K; Subbannayya, K

    2014-07-01

    Three serum free media viz, sucrose solution, starch solution and SST broth have been formulated. The objective of the present study was to evaluate these three different serum free media for induction of germ tubes by Candida albicans and to compare their efficacy with the pooled human serum. Out of 50 C. albicans isolates 47 (94%) and 49 (98%) produced germ tubes in pooled human serum and SST broth, respectively. Germ tube production was positive in 40 (80%) and 36 (72%) isolates, respectively in sucrose solution and starch solution. This study reports SST broth as a new stable and less expensive germ tube induction medium, which requires less time for preparation and can be used without any safety concerns. SST broth is found to be more effective than pooled human serum for induction of germ tubes by C. albicans isolates.

  4. Slide Coagglutination for Salmonella typhi Antigens in Broths Inoculated with Feces from Typhoid Fever Patients

    DTIC Science & Technology

    1981-12-01

    SLIDE COAGGLUTINATION FOR SALMONELLA TYPHI ANTIGENS IN BROTHS INOCULATED WITH FECES FROM TYPHOID FEVER PATIENTS R. C. Rockhill, L. W. Rumans and M...BROTHS INOCULATED WITH FECES FROM TYPHOID FEVER PATIENTS ROBERT C. ROCKMiILL, LARRY W. RUMANS* and MURAD LESMANA U.S. Naval Medical Research Unit No. 2...METHODS The method of bacterial coagglutination Patients: Individuals suspected of having was developed by Kronvall for serotyping typhoid fever were

  5. Comparative analysis of mixing distribution in aerobic stirred bioreactor for simulated yeasts and fungus broths.

    PubMed

    Cascaval, Dan; Galaction, Anca-Irina; Turnea, Marius

    2007-01-01

    The study on mixing distribution for an aerobic stirred bioreactor and simulated (solutions of carboxymethylcellulose sodium salt), yeasts (S. cerevisiae) and fungus (P. chrysogenum pellets and free mycelia) broths indicated the significant variation of mixing time on the bioreactor height. The experiments suggested the possibility to reach a uniform mixing in whole bulk of the real broths for a certain value of rotation speed or biomass concentration domain. For S. cerevisiae broths the optimum rotation speed increased to 500 rpm with the biomass accumulation from 40 to 150 g/l d.w. Irrespective of their morphology, for fungus cultures the existence of optimum rotation speed (500 rpm) has been recorded only for biomass concentration below 24 g/l d.w. The influence of aeration rate depends on the apparent viscosity/biomass concentration and on the impellers and sparger positions. By increasing the apparent viscosity for simulated broths, or biomass amount for real broths, the shape of the curves describing the mixing time variation is significantly changed for all the considered positions. The intensification of the aeration induced the increase of mixing time, which reached a maximum value, decreasing then, due to the flooding phenomena. This variation became more pronounced at higher viscosities for simulated broths, at higher yeasts concentration, and at lower pellets or filamentous fungus concentration, respectively. By means of the experimental data and using MATLAB software, some mathematical correlations for mixing time have been proposed for each broth and considered position inside the bioreactor. These equations offer a good agreement with the experiment, the maximum deviation being +/-7.3% for S. cerevisiae broths.

  6. Efficient recovery of gamma-poly (glutamic acid) from highly viscous culture broth.

    PubMed

    Do, J H; Chang, H N; Lee, S Y

    2001-11-01

    An efficient strategy for the separation and recovery of gamma-polyglutamic acid (gamma-PGA) from highly viscous broth was developed. This strategy was divided into two processes: The first was to separate gamma-PGA from highly viscous culture broth; the second was to concentrate gamma-PGA solution by ultrafiltration for the reduction of the amount of alcohol required during recovery process with precipitation. By lowering the pH value of culture broth to 3, the viscosity of culture broth and the zeta potential of cell could be reduced to a sixth of the original value at 35 degrees C and a third, respectively. After the acidification of culture broth the energy demand for the separation of gamma-PGA from culture broth by centrifugation could be reduced to 17% of that without it when the centrifugal force was 22,000g. The amount of alcohol required for precipitation could be reduced to a fourth of that generally used without concentration by concentrating 20 g gamma-PGA/L solution to 60 g gamma-PGA/L at pH 5 by ultrafiltration with hollow-fiber membrane cartridge (MWCO 500,000). Copyright 2001 John Wiley & Sons, Inc.

  7. Comparative analysis of oxygen transfer rate distribution in stirred bioreactor for simulated and real fermentation broths.

    PubMed

    Caşcaval, Dan; Galaction, Anca-Irina; Turnea, Marius

    2011-09-01

    Study of the distribution of the oxygen mass transfer coefficient, k (l) a, for a stirred bioreactor and simulated (pseudoplastic solutions of carboxymethylcellulose sodium salt) bacterial (P. shermanii), yeast (S. cerevisiae), and fungal (P. chrysogenum free mycelia) broths indicated significant variation of transfer rate with bioreactor height. The magnitude of the influence of the considered factors differed from one region to another. As a consequence of cell adsorption to bubble surface, the results indicated the impossibility of achieving a uniform oxygen transfer rate throughout the whole bulk of the microbial broth, even when respecting the conditions for uniform mixing. Owing to the different affinity of biomass for bubble surface, the positive influence of power input on k (l) a is more important for fungal broths, while increasing aeration is favorable only for simulated, bacterial and yeast broths. The influence of the considered factors on k (l) a were included in mathematical correlations established based on experimental data. For all considered positions, the proposed equations for real broths have the general expression [Formula in text] exhibiting good agreement with experimental results (with maximum deviations of ± 10.7% for simulated broths, ± 8.4% for P. shermanii, ± 9.3% for S. cerevisiae, and ± 6.6% for P. chrysogenum).

  8. Clinical comparison of isolator, Septi-Chek, nonvented tryptic soy broth, and direct agar plating combined with thioglycolate broth for diagnosing spontaneous bacterial peritonitis.

    PubMed Central

    Hay, J E; Cockerill, F R; Kaese, D; Vetter, E A; Wollan, P C; Rakela, J; Wilhelm, M P

    1996-01-01

    Spontaneous bacterial peritonitis is a life-threatening complication of cirrhotic ascites. Optimal patient management depends on the isolation of the causal organism from ascitic fluid. To evaluate culture techniques for the diagnosis of spontaneous bacterial peritonitis, we prospectively compared three blood culture system, the Isolator system, a lysis-centrifugation system, the Septi-Chek system, a biphasic culture system, and a nonvented tryptic soy broth system, all inoculated at the bedside, and our standard method of direct inoculation of specimens after transport to the laboratory onto agar plates and into thioglycolate broth. The results showed that the Septi-Chek and nonvented tryptic soy broth systems each recovered statistically significantly more pathogens than either the Isolator system (P = 0.0084) or the standard method (P = 0.00098). The Isolator system recovered more pathogens than the standard plate method, but this difference was not statistically significant. Both the Isolator system and the standard plate method recovered more contaminating microorganisms than the Septi-Chek or nonvented tryptic soy broth system. The Isolator system required the most processing time compared with the processing times required by any other method. PMID:8748267

  9. Comparison of the Sensititre YeastOne colorimetric antifungal panel with CLSI microdilution for antifungal susceptibility testing of the echinocandins against Candida spp., using new clinical breakpoints and epidemiological cutoff values.

    PubMed

    Pfaller, M A; Chaturvedi, V; Diekema, D J; Ghannoum, M A; Holliday, N M; Killian, S B; Knapp, C C; Messer, S A; Miskou, A; Ramani, R

    2012-08-01

    A commercially prepared dried colorimetric microdilution panel (Sensititre Yeast One, TREK Diagnostic Systems, Cleveland, OH, USA) was compared in 3 different laboratories with the Clinical and Laboratory Standards Institute (CLSI) reference microdilution method by testing 2 quality control strains, 25 reproducibility strains, and 404 isolates of Candida spp. against anidulafungin, caspofungin, and micafungin. Reference CLSI BMD MIC end points and YeastOne colorimetric end points were read after 24 h of incubation. Excellent (100%) essential agreement (within 2 dilutions) between the reference and colorimetric MICs was observed. Categorical agreement (CA) between the 2 methods was assessed using the new species-specific clinical breakpoints (CBPs): susceptible (S), ≤0.25 μg/mL; intermediate (I), 0.5 μg/mL; and resistant (R), ≥1 μg/mL, for C. albicans, C. tropicalis, and C. krusei, and ≤2 μg/mL (S), 4 μg/mL (I), and ≥8 μg/mL (R) for C. parapsilosis and all 3 echinocandins. The new CBPs for anidulafungin and caspofungin and C. glabrata are ≤0.12 μg/mL (S), 0.25 μg/mL (I), and ≥0.5 μg/mL (R), whereas those for micafungin are ≤0.06 μg/mL (S), 0.12 μg/mL (I), and ≥0.25 μg/mL (R). Due to the lack of CBPs for any of the echinocandins and C. lusitaniae, the epidemiological cutoff values (ECVs) were used for this species to categorize the isolates as wild-type (WT; MIC ≤ECV) and non-WT (MIC >ECV), respectively, for anidulafungin (≤2 μg/mL/>2 μg/mL), caspofungin (≤1 μg/mL/>1 μg/mL), and micafungin (≤0.5 μg/mL/>0.5 μg/mL). CA ranged from 93.6% (caspofungin) to 99.6% (micafungin) with less than 1% very major or major errors. The YeastOne colorimetric method remains comparable to the CLSI BMD reference method for testing the susceptibility of Candida spp. to the echinocandins when using the new (lower) CBPs and ECVs. Further study using defined fks mutant strains of Candida is warranted.

  10. Comparison of different preenrichment broths, egg:preenrichment broth ratios, and surface disinfection for the detection of Salmonella enterica ssp. enterica serovar Enteritidis in shell eggs.

    PubMed

    Zhang, G; Brown, E W; Hammack, T S

    2013-11-01

    Salmonella enterica ssp. enterica serovar Enteritidis is the leading reported cause of Salmonella infections. Most Salmonella Enteritidis infections are associated with whole shell eggs and egg products. This project attempted to lay the foundation for improving the Food and Drug Administration's current Bacteriological Analytical Manual method for the detection of Salmonella Enteritidis in shell eggs. Two Salmonella Enteritidis isolates were used for comparisons among different preenrichment and enrichment media and for the evaluation of egg:preenrichment broth ratios for the detection of Salmonella Enteritidis in shell eggs. The effect of surface disinfection on the detection of Salmonella Enteritidis in shell eggs was also investigated. The results indicated that tryptic soy broth (TSB) was similar to TSB plus ferrous sulfate, but significantly (α = 0.05) better than nutrient broth, Universal Preenrichment broth, and buffered peptone water when used for preenrichment of Salmonella in shell eggs. Salmonella Enteritidis populations after enrichment with Rappaport-Vassiliadis broth were 0.40 to 1.11 log cfu/mL of culture lower than those in preenrichment cultures. The reduction was statistically significant (α = 0.05). Egg:broth ratios at 1:9 and 1:2 produced significantly (α = 0.05) higher Salmonella Enteritidis populations after preenrichment with TSB with inoculum levels at 4 cfu/100 g of eggs and 40 cfu/1,000 g of eggs than the ratio at 1:1. Salmonella Enteritidis populations in TSB preenrichment cultures of shell eggs surface-disinfected with 70% alcohol:iodine/potassium iodide solution and untreated control were 9.11 ± 0.11 and 9.18 ± 0.05 log cfu/mL, respectively, for SE 13-2, and 9.20 ± 0.04 and 9.16 ± 0.05 log cfu/mL, respectively, for SE CDC_2010K_1543. Surface disinfection of eggs did not reduce the sensitivity of detection of Salmonella Enteritidis in liquid eggs. These results could improve the Food and Drug Administration's current

  11. Broth versus solid agar culture of swab samples of cadaveric allograft musculoskeletal tissue.

    PubMed

    Varettas, Kerry

    2013-12-01

    As part of the donor assessment protocol, bioburden assessment must be performed on allograft musculoskeletal tissue samples collected at the time of tissue retrieval. Swab samples of musculoskeletal tissue allografts from cadaveric donors are received at the microbiology department of the South Eastern Area Laboratory Services (Australia) to determine the presence of bacteria and fungi. This study will review the isolation rate of organisms from solid agar and broth culture of swab samples of cadaveric allograft musculoskeletal tissue over a 6-year period, 2006-2011. Swabs were inoculated onto horse blood agar (anaerobic, 35 °C) and chocolate agar (CO2, 35 °C) and then placed into a cooked meat broth (aerobic, 35 °C). A total of 1,912 swabs from 389 donors were received during the study period. 557 (29.1 %) swabs were culture positive with the isolation of 713 organisms, 249 (34.9 %) from solid agar culture and an additional 464 (65.1 %) from broth culture only. This study has shown that the broth culture of cadaveric allograft musculoskeletal swab samples recovered a greater amount of organisms than solid agar culture. Isolates such as Clostridium species and Staphylococcus aureus would not have been isolated from solid agar culture alone. Broth culture is an essential part of the bioburden assessment protocol of swab samples of cadaveric allograft musculoskeletal tissue in this laboratory.

  12. Supplementing chicken broth with monosodium glutamate reduces hunger and desire to snack but does not affect energy intake in women.

    PubMed

    Carter, Brett E; Monsivais, Pablo; Perrigue, Martine M; Drewnowski, Adam

    2011-11-01

    The effect of monosodium glutamate (MSG) supplementation in soup or broth on satiety is not well understood. In the present study, the relative effects of four chicken broths with or without added MSG on motivational ratings and energy intakes at the next meal were compared using a double-blinded, within-subject design. A total of thirty-five normal-weight women, aged 20-40 years, took part in four study sessions. The four broths were base chicken broth (63 kJ), broth with added MSG (1.19 g) and nucleotides (0.03 g), broth with added MSG (1.22 g), and broth with added fat (BAF; 681 kJ). The preloads were presented twice at 09.00 and 11.15 hours for a maximum cumulative dose of 2.44 g MSG. Motivational ratings were collected before and at 15 min intervals post-ingestion for a total of 210 min. A test lunch meal was served at 12.00 hours, and plate waste was measured. The addition of MSG to chicken broth did not increase energy intakes at lunch or affect motivational ratings over the entire testing session. Both hunger and desire to snack between the second preload exposure and the test meal were significantly reduced in the MSG condition relative to the base broth condition (both, P = 0.03). However, only the BAF significantly suppressed energy intakes at lunch compared with the base broth control condition. Supplementing chicken broth with MSG can increase subjective ratings for satiety but does not alter energy intake at the next meal relative to an equal energy broth without added MSG.

  13. Role of transparent exopolymeric particles in membrane fouling: Chlorella vulgaris broth filtration.

    PubMed

    Discart, V; Bilad, M R; Vandamme, D; Foubert, I; Muylaert, K; Vankelecom, I F J

    2013-02-01

    Recent reports show strong evidence for the involvement of transparent exopolymer particles (TEPs), mainly produced by microalgae in natural environments, in membrane fouling in a wide range of membrane filtration processes. The objective of this study is to fundamentally investigate the direct role of TEPs on membrane fouling by using different Chlorella vulgaris broth solutions and different fractions of such broth (the soluble and bound fractions, the cells separated from these fractions and the cells with their bound sugars, separated from the soluble fraction) as filtration feed. The relation between the feed properties and their filterability over three membranes was determined. Scanning electron microscopy and light microscopy showed that the foulant types differed for each broth fraction and confirmed the role of TEPs in the fouling of microfiltration membranes. In addition, this study contributes to the role of TEPs in the filtration of microalgae cultivated for commercial reasons.

  14. Solvent screening study and conceptual extractive distillation process to produce anhydrous ethanol from fermentation broth

    SciTech Connect

    Lee, F-M.; Pahl, R.H.

    1985-01-01

    This solvent screening study indicates that glycols are very selective solvents for producing anhydrous ethanol from fermentation broth by extractive distillation. The promising solvents are glycerin, ethylene glycol, tetraethylene glycol, and triethylene glycol. An improved extractive distillation process for producing anhydrous ethanol was conceptually developed which was based on the analysis of the experimental solvent screening data. The basic idea of this process is to add an extractive solvent only to the ethanol-rich portion of the fractionator to eliminate the ethanol-water azeotrope and to change the undesirable shape of the vapor-liquid equilibrium curve to one more favorable for distillation. With a suitable solvent, such as ethylene glycol, anhydrous ethanol could be produced from the fermentation broth in a column with only 18 theoretical trays. A low reflux ratio of 1.5 and a low solvent-to-feed ratio of 0.27 (based on fermentation broth) would be needed for the separation.

  15. Chemical Constituents of the Culture Broth of Phellinus linteus and Their Antioxidant Activity.

    PubMed

    Lee, Myeong-Seok; Hwang, Byung Soon; Lee, In-Kyoung; Seo, Geon-Sik; Yun, Bong-Sik

    2015-03-01

    The medicinal fungus Phellinus linteus, in the family Hymenochaetaceae, has been used as a traditional medicine for the treatment of various diseases. In this study, the chemical constituents of the culture broth of P. linteus were investigated. P. linteus was cultured in potato dextrose broth medium, and the culture broth was extracted with ethyl acetate. The ethyl acetate-soluble portion was concentrated and subjected to ODS column chromatography, followed by Sephadex LH-20 column chromatography. Six compounds (1~6) were purified by preparative reversed-phase high-performance liquid chromatography. Spectroscopic methods identified their structures as caffeic acid (1), inotilone (2), 4-(3,4-dihydroxyphenyl)-3-buten-2-one (3), phellilane H (4), (2E,4E)-(+)-4'-hydroxy-γ-ionylideneacetic acid (5), and (2E,4E)-γ-ionylideneacetic acid (6). Compounds 1, 2, and 3 exhibited potent dose-dependent antioxidant activity.

  16. Chemical Constituents of the Culture Broth of Phellinus linteus and Their Antioxidant Activity

    PubMed Central

    Lee, Myeong-Seok; Hwang, Byung Soon; Lee, In-Kyoung; Seo, Geon-Sik

    2015-01-01

    The medicinal fungus Phellinus linteus, in the family Hymenochaetaceae, has been used as a traditional medicine for the treatment of various diseases. In this study, the chemical constituents of the culture broth of P. linteus were investigated. P. linteus was cultured in potato dextrose broth medium, and the culture broth was extracted with ethyl acetate. The ethyl acetate-soluble portion was concentrated and subjected to ODS column chromatography, followed by Sephadex LH-20 column chromatography. Six compounds (1~6) were purified by preparative reversed-phase high-performance liquid chromatography. Spectroscopic methods identified their structures as caffeic acid (1), inotilone (2), 4-(3,4-dihydroxyphenyl)-3-buten-2-one (3), phellilane H (4), (2E,4E)-(+)-4'-hydroxy-γ-ionylideneacetic acid (5), and (2E,4E)-γ-ionylideneacetic acid (6). Compounds 1, 2, and 3 exhibited potent dose-dependent antioxidant activity. PMID:25892914

  17. Antibiotic Fermentation Broth Treatment by a pilot upflow anaerobic sludge bed reactor and kinetic modeling.

    PubMed

    Coskun, T; Kabuk, H A; Varinca, K B; Debik, E; Durak, I; Kavurt, C

    2012-10-01

    In this study, an upflow anaerobic sludge blanket (UASB) mesophilic reactor was used to remove antibiotic fermentation broth wastewater. The hydraulic retention time was held constant at 13.3 days. The volumetric organic loading value increased from 0.33 to 7.43 kg(COD)m(-3)d(-1) using antibiotic fermentation broth wastewater gradually diluted with various ratios of domestic wastewater. A COD removal efficiency of 95.7% was obtained with a maximum yield of 3,700 L d(-1) methane gas production. The results of the study were interpreted using the modified Stover-Kincannon, first-order, substrate mass balance and Van der Meer and Heertjes kinetic models. The obtained kinetic coefficients showed that antibiotic fermentation broth wastewater can be successfully treated using a UASB reactor while taking COD removal and methane production into account. Copyright © 2012 Elsevier Ltd. All rights reserved.

  18. Enumeration of starter cultures during yogurt production using Petrifilm AC plates associated with acidified MRS and M17 broths.

    PubMed

    Gonçalves, Marília M; Freitas, Rosangela; Nero, Luís A; Carvalho, Antônio F

    2009-05-01

    The efficiency of Petrifilm AC (3M Microbiology, St. Paul, MN, USA) associated with the broths M17 and de Mann-Rogosa-Sharpe (MRS) at pH 5.4 was evaluated to enumerate Streptococcus salivarius subsp. thermophilus and Lactobacillus delbrueckii subsp. bulgaricus during the yogurt production. Commercial and reference strains of these microorganisms were experimentally inoculated in nonfat milk and incubated at 42 degrees C for 4 h for yogurt production. At the moment of inoculation and after incubation, aliquots were collected, submitted to dilution using the broths M17 and MRS at pH 5.4, and plated for Strep. salivarius and Lb. bulgaricus enumeration according ISO 9232 and at Petrifilm AC plates, respectively. M17 plates were incubated at 42 degrees C, and MRS plates were incubated at 35 degrees C under anaerobiosis. After 48 h, the formed colonies were enumerated and the counts were compared by correlation and analysis of variance (P<0.05). In addition, colonies were randomly selected from all plates and characterized according to Gram staining and morphology. The obtained results indicated that Petrifilm AC plates associated to M17 and MRS at pH 5.4 can be considered as a suitable alternative for Strep. salivarius and Lb. bulgaricus enumeration during yogurt production, with slight interferences due to the acidity of MRS at the moment of inoculation, and due to the acidity of yogurt at the end of fermentation process. It was also observed that the MRS at pH 5.4 was not sufficiently selective for Lb. delbrueckii enumeration, despite it is indicated by the official protocol from ISO 9232.

  19. Preconcentrating (within the broth) secreted extracellular proteins during a bakers' yeast fermentation

    SciTech Connect

    Effler, W.T. Jr.; Pandey, N.K.; Tanner, R.D.; Malaney, G.W.; Scott, C.D.

    1986-01-01

    Proteins secreted by yeast during the fermentation process are spacially fractionated (concentrated at a particular vertical position) within the fermentation vessel due to the phenomenon of bubble fractionation, despite moderately vigorous mixing. The degree of fractionation is influenced by the conditions in which the fermentation takes place. The broth pH strongly influences the extent of fractionation of specific proteins. In addition fractionation is enhanced under anaerobic conditions, presumably because there are more CO2 bubbles present for hydrophobic protein adsorption. The addition of moderate levels of salt to the broth reduces the fractionation for most (but not all) of the proteins.

  20. Comparison of EUCAST and CLSI Reference Microdilution MICs of Eight Antifungal Compounds for Candida auris and Associated Tentative Epidemiological Cutoff Values.

    PubMed

    Arendrup, M C; Prakash, Anupam; Meletiadis, Joseph; Sharma, Cheshta; Chowdhary, Anuradha

    2017-06-01

    Candida auris is an emerging multidrug-resistant yeast. So far, all but two susceptibility testing studies have examined ≤50 isolates, mostly with the CLSI method. We investigated CLSI and EUCAST MICs for 123 C. auris isolates and eight antifungals and evaluated various methods for epidemiological cutoff (ECOFF) determinations. MICs (in milligrams per liter) were determined using CLSI method M27-A3, and the EUCAST E.Def 7.3. ANOVA analysis of variance with Bonferroni's multiple-comparison test and Pearson analysis were used on log2 MICs (significance at P values of <0.05). The percent agreement (within ±0 to ±2 2-fold dilutions) between the methods was calculated. ECOFFs were determined visually, statistically (using the ECOFF Finder program and MicDat1.23 software with 95% to 99% endpoints), and via the derivatization method (dECOFFs). The CLSI and EUCAST MIC distributions were wide, with several peaks for all compounds except amphotericin B, suggesting possible acquired resistance. Modal MIC, geometric MIC, MIC50, and MIC90 values were ≤1 2-fold dilutions apart, and no significant differences were found. The quantitative agreement was best for amphotericin B (80%/97% within ±1/±2 dilutions) and lowest for isavuconazole and anidulafungin (58%/76% to 75% within ±1/±2 dilutions). We found that 90.2%/100% of the isolates were amphotericin B susceptible based on CLSI/EUCAST methods, respectively (i.e., with MICs of ≤1 mg/liter), and 100%/97.6% were fluconazole nonsusceptible by CLSI/EUCAST (MICs > 2). The ECOFFs (in milligrams per liter) were similar across the three different methods for itraconazole (ranges for CLSI/EUCAST, 0.25 to 0.5/0.5 to 1), posaconazole (0.125/0.125 to 0.25), amphotericin B (0.25 to 0.5/1 to 2), micafungin (0.25 to 0.5), and anidulafungin (0.25 to 0.5/0.25 to 1). In contrast, the estimated ECOFFs were dependent on the method applied for voriconazole (1 to 32) and isavuconazole (0.125 to 4). CLSI and EUCAST MICs were remarkably similar and confirmed uniform fluconazole resistance and variable acquired resistance to the other agents. Copyright © 2017 American Society for Microbiology.

  1. Identification of Candida species and susceptibility testing with Sensititre YeastOne microdilution panel to 9 antifungal agents.

    PubMed

    Kucukates, Emine; Gultekin, Nuh N; Alisan, Zeynep; Hondur, Nur; Ozturk, Recep

    2016-07-01

    To determine the species incidence and susceptibility pattern to 9 antifungal agents of yeasts isolated from various clinical specimens of colonized or infected patients treated in the coronary and surgical intensive care units (ICU).  A total of 421 ICU patients were treated at the Cardiology Institute, Istanbul University, Istanbul, Turkey between June 2013 and May 2014, and 44 Candida species were isolated from blood, urine, endotracheal aspiration fluid, sputum, and wounds of 16 ICU patients. Identification of Candida was performed using CHROMagar. Antifungal susceptibility was determined by a Sensititre YeastOne colorimetric microdilution panel.  Candida albicans (C. albicans) was the most commonly observed microorganism 23 (54%); the other microorganisms isolated were Candida tropicalis 12 (27%), Candida glabrata 5 (11%), Candida parapsilosis 1 (2%), Candida lusitaniae 1 (2%), Candida sake 1 (2%), and Geotrichum capitatum 1 (2%). All isolates were susceptible to amphotericin B and 5-flucytosine. Geotrichum capitatum excepted, the other isolates were also susceptible to anidulafungin, micafungin, and caspofungin. Candida parapsilosis was found to be susceptible to all the studied antifungals. High MIC rates for azole group of antifungal drugs were found for C. albicans, C. tropicalis, and C. glabrata. The rate of colonisation was 3.8% (16/421). Only 0.7% (3/421) patients out of a total of 421 developed candidemia.  We found that the yeast colonization and infection rates of patients in our ICUs are very low. Candida albicans is still the most common species. We detected a decreasing susceptibility to azole compounds.

  2. Development of a multi-pathogen enrichment broth for simultaneous growth of five common foodborne pathogens.

    PubMed

    Chen, Juan; Tang, Junni; Bhunia, Arun K; Tang, Cheng; Wang, Changting; Shi, Hui

    2015-01-01

    The objective of the present study was to formulate a multi-pathogen enrichment broth which could support the simultaneous growth of five common foodborne pathogens (Salmonella enterica, Staphylococcus aureus, Shigella flexneri, Listeria monocytogenes and Escherichia coli O157:H7). The formulated broth SSSLE was composed of potassium tellurite, bile salt, lithium chloride, and sodium chloride as growth-inhibitors; glucose, esculin, mannitol and sodium pyruvate as growth-promoters. Compared with the respective specific selective enrichment broths, the individual growth pattern of each target pathogen in SSSLE was equal, or even better, except in the case of S. flexneri. In mixed-culture experiments, the gram-negative bacteria showed higher growth capabilities than the gram-positive bacteria after 8-h enrichment; however, the cell numbers after 24-h enrichment indicated that SSSLE could support the concurrent growth of five target pathogens irrespective of whether pathogens were inoculated initially at equal or unequal levels. For natural food samples under the high background flora, the final cell numbers enriched in SSSLE for five targets were enough to be detected by multiplex PCR. In conclusion, SSSLE was capable of supporting the growth of five target pathogens concurrently. The new broth formulated in this study has the potential of saving time, efforts and costs in multi-pathogen enrichment procedures.

  3. Simple and efficient isolation of cordycepin from culture broth of a Cordyceps militaris mutant.

    PubMed

    Masuda, Mina; Hatashita, Masanori; Fujihara, Shinya; Suzuki, Yu; Sakurai, Akihiko

    2015-12-01

    Isolation of cordycepin from the culture broth of Cordyceps militaris mutant was investigated. Based on the solubility curve, three crystallizing processes, temperature shift (process I), pH shift (process II), and pH shift followed by temperature shift (process III) were carried out. Process III was the most promising method regarding both purity and yield.

  4. Evaluation of a newly developed triple buffered peptone broth for detection of Salmonella in broiler feed

    USDA-ARS?s Scientific Manuscript database

    Lactose broth (LB) and buffered peptone (BP) are used as pre-enrichment media to recover Salmonella from feed. Bacterial utilization of feed carbohydrates results in the production of acidic byproducts causing a drop in the media pH which can injure or kill Salmonella and yield false negative resul...

  5. Use of Mueller-Hinton broth and agar in the germ tube test.

    PubMed

    Mattei, Antonella Souza; Alves, Sydney Hartz; Severo, Cecília Bittencourt; Guazzelli, Luciana da Silva; Oliveira, Flávio de Mattos; Severo, Luiz Carlos

    2014-01-01

    Candida albicans is often isolated from clinical samples, thus its presumptive differentiation from other species of the same genus can be based on its ability to form the germ tube in human serum. Nevertheless, there are two other species that share this characteristic: C. dubliniensis and C. africana. The aim of this study was to compare four different substrates to perform the germ tube (GT) test. The Candida spp. isolates were identified using a manual system (135 C. albicans, 24 C. tropicalis and one C. dubliniensis). The germ tube test was performed with fresh, previously frozen serum and Mueller-Hinton (MH) broth and agar. GT was observed in 96% (130/136) of the isolates through the fresh serum technique, 94% (128/136) through previously frozen serum, 92% (125/136) in MH agar, and 90% (122/136) in MH broth. The sensitivity of each test was higher than 90%, with 100% specificity. Both the MH agar and broth were able to identify the true positives, and false positives were not found. However, some C. albicans isolates were not identified. MH agar and broth may be used in laboratory for the rapid presumptive identification of C. albicans, as an alternative method for germ tube test.

  6. Use of liquid/supercritical CO2 extraction process for butanol recovery from fermentation broth

    USDA-ARS?s Scientific Manuscript database

    In order for butanol fermentation to be a viable option, it is essential to recover it from fermentation broth using economical alternate in-situ product recovery techniques such as liquid/supercritical CO2 extraction as compared to distillation. This technique (liquid CO2 extraction & supercritical...

  7. Dithiolopyrrolone antibiotic formation induced by adding valeric acid to the culture broth of Saccharothrix algeriensis.

    PubMed

    Merrouche, Rabiâa; Bouras, Noureddine; Coppel, Yannick; Mathieu, Florence; Monje, Marie-Carmen; Sabaou, Nasserdine; Lebrihi, Ahmed

    2010-06-25

    Three new antibiotics were isolated from the fermentation broth of Saccharothrix algeriensis NRRL B-24137 and characterized as the dithiolopyrrolone derivatives valerylpyrrothine (1), isovalerylpyrrothine (2), and formylpyrrothine (3) as well as the known antibiotic aureothricin. The production of the dithiolopyrrolone derivatives was induced by adding valeric acid to the culture medium. The compounds exhibited moderate antimicrobial activity in vitro.

  8. Multi-Probe Real-Time PCR Identification of Common Mycobacterium Species in Blood Culture Broth

    PubMed Central

    Foongladda, Suporn; Pholwat, Suporn; Eampokalap, Boonchuay; Kiratisin, Pattarachai; Sutthent, Ruengpung

    2009-01-01

    Mycobacterium tuberculosis complex, M. avium, and M. intracellulare are the most common causes of systemic bacterial infection in AIDS patients. To identify these mycobacterial isolates in primary blood culture broths, we developed a multiple hybridization probe-based real-time PCR assay using the LightCycler system. The primers were designed to amplify a 320-bp fragment of Mycobacterium 16S rRNA genes. Reaction specificity was evaluated using PCR amplification curves along with specific melting temperatures of probes on DNA extracted from 13 Mycobacterium species. In this study, results showed 100% accuracy for the selected bacterial panel. Detection limits were 350, 600, and 650 colony-forming unit (CFU)/ml blood culture broths for M. tuberculosis complex, M. avium, and M. intracellulare, respectively (1 to 2 CFU/reaction). To evaluate clinical applicability, 341 acid-fast bacilli in blood culture broths were analyzed. In total, 327 (96%) were positively identified: 54.5% M. tuberculosis complex, 37.5% M. avium, and 3.8% M. intracellulare. Results can be available within 3 hours of receiving a broth sample, which makes this rapid and simple assay an attractive diagnostic tool for clinical use. PMID:19095775

  9. Production of cell-free xanthan fermentation broth by cell adsorption on fibers

    PubMed

    Yang; Lo; Chattopadhyay

    1998-03-01

    Xanthan gum is a microbial polysaccharide widely used in food and oil-drilling industries. Xanthan gum produced from the current commercial fermentation process usually contains cells and cell debris, which lower the filterability of the xanthan solution and limit its applications. The production of cell-free xanthan gum fermentation broth is thus desirable. The feasibility of removing cells from the xanthan fermentation broth by cell adsorption to various woven fibrous materials was studied. It was found that both cotton and polyester fibers could be used to adsorb Xanthomonas campestris cells present in the fermentation broth either during batch fermentation or after the fermentation. Almost all cells were removed from the fermentation broth by adsorption to fibers. Cotton terry cloth had rough surfaces and was the preferred material for cell adsorption. Cell adsorption to cotton was faster than to polyester fibers. The adsorption kinetics can be modeled by a first-order rate equation. The adsorption rate constants were 30-40% higher for cotton than for polyester. Cell adsorption was not efficient in the absence of xanthan gum, suggesting that the exopolysaccharide, xanthan gum, was important for efficient cell adsorption to fibers.

  10. Analysis of lard in meatball broth using Fourier transform infrared spectroscopy and chemometrics.

    PubMed

    Kurniawati, Endah; Rohman, Abdul; Triyana, Kuwat

    2014-01-01

    Meatball is one of the favorite foods in Indonesia. For the economic reason (due to the price difference), the substitution of beef meat with pork can occur. In this study, FTIR spectroscopy in combination with chemometrics of partial least square (PLS) and principal component analysis (PCA) was used for analysis of pork fat (lard) in meatball broth. Lard in meatball broth was quantitatively determined at wavenumber region of 1018-1284 cm(-1). The coefficient of determination (R(2)) and root mean square error of calibration (RMSEC) values obtained were 0.9975 and 1.34% (v/v), respectively. Furthermore, the classification of lard and beef fat in meatball broth as well as in commercial samples was performed at wavenumber region of 1200-1000 cm(-1). The results showed that FTIR spectroscopy coupled with chemometrics can be used for quantitative analysis and classification of lard in meatball broth for Halal verification studies. The developed method is simple in operation, rapid and not involving extensive sample preparation.

  11. Recovery of butanol from Clostridium beijerinckii P260 fermentation broth by supercritical CO

    USDA-ARS?s Scientific Manuscript database

    Butanol is a superior biofuel to ethanol because of its blend properties and higher energy density. However, its recovery by distillation from the fermentation broth is energy intensive. For this reason, we studied butanol recovery by supercritical CO2 extraction from simulated and actual fermentati...

  12. Studies on the interaction of fermentation and microfiltration operations: erythromycin recovery from Saccharopolyspora erythraea fermentation broths.

    PubMed

    Davies, J L; Baganz, F; Ison, A P; Lye, G J

    2000-08-20

    Changes in fermentation media not only affect the performance of the fermentation itself (with regard to the kinetics of biomass and product formation and the yields obtained) but also the initial product-recovery operations downstream of the fermentor. In this work, microfiltration experiments to remove Saccharopolyspora erythraea biomass from fermentation broth and to recover erythromycin were carried out using two fundamentally different media; a soluble complex medium (SCM) and an oil-based process medium (OBM). Small-scale batch fermentations of 14-L working volume were carried out in triplicate using both media. Broth samples were taken from each fermentation at regular intervals from the end of the exponential-growth phase onwards. These were then processed using a Minitan II (acrylic), tangential crossflow-filtration module, fitted with a single 60 cm(2) Durapore hydrophilic 0.2 microm membrane, operated in concentration mode. The OBM fermentations produced higher titers of erythromycin but required longer fermentation times due to increased lag phases and slower maximum-growth rates. The OBM also increased the loading on the membrane; at maximum product titers residual oil concentrations of 3 g. L(-1), antifoam concentrations of 2 g. L(-1) and flour concentrations estimated at approximately 10 g/L(-1) were typical. It was found that both the permeate flux and erythromycin transmission were affected by the choice of medium. The OBM had significantly lower values for both parameters (12.8 Lm(-2) h(-1) and 89.6% respectively) than the SCM (35.9 Lm(-2) h(-1) and 96.7% respectively) when the fermentations were harvested at maximum erythromycin titers. Transmission of erythromycin stayed approximately constant as a function of fermentation time for both media, however, for the OBM the permeate flux decreased with time which correlated with an increase in broth viscosity. The relatively poor microfiltration performance of the OBM medium was, however, offset by

  13. In vitro antagonistic growth effects of Lactobacillus fermentum and lactobacillus salivarius and their fermentative broth on periodontal pathogens.

    PubMed

    Chen, Ling-Ju; Tsai, Hsiu-Ting; Chen, Wei-Jen; Hsieh, Chu-Yang; Wang, Pi-Chieh; Chen, Chung-Shih; Wang, Lina; Yang, Chi-Chiang

    2012-10-01

    As lactobacilli possess an antagonistic growth property, these bacteria may be beneficial as bioprotective agents for infection control. However, whether the antagonistic growth effects are attributed to the lactobacilli themselves or their fermentative broth remains unclear. The antagonistic growth effects of Lactobacillus salivarius and Lactobacillus fermentum as well as their fermentative broth were thus tested using both disc agar diffusion test and broth dilution method, and their effects on periodontal pathogens, including Streptococcus mutans, Streptococcus sanguis, and Porphyromonas gingivalis in vitro at different concentrations and for different time periods were also compared. Both Lactobacillus salivarius and Lactobacillus fermentum and their concentrated fermentative broth were shown to inhibit significantly the growth of Streptococcus mutans, Streptococcus sanguis, and Porphyromonas gingivalis, although different inhibitory effects were observed for different pathogens. The higher the counts of lactobacilli and the higher the folds of concentrated fermentative broth, the stronger the inhibitory effects are observed. The inhibitory effect is demonstrated to be dose-dependent. Moreover, for the lactobacilli themselves, Lactobacillus fermentum showed stronger inhibitory effects than Lactobacillus salivarius. However, the fermentative broth of Lactobacillus fermentum showed weaker inhibitory effects than that of Lactobacillus salivarius. These data suggested that lactobacilli and their fermentative broth exhibit antagonistic growth activity, and consumption of probiotics or their broth containing lactobacilli may benefit oral health.

  14. Effect of dried-bonito broth on mood states: a pooled analysis of four randomized controlled human trials.

    PubMed

    Kuroda, Motonaka; Nozawa, Yoshizu

    2008-08-01

    Dried-bonito broth is commonly employed as a soup and sauce base in Japanese cuisine and is considered to be a nutritional supplement that promotes recovery from fatigue. Previous human trials have indicated that the ingestion of dried-bonito broth improves several mood states including fatigue, however, the effects in these studies have differed. The aim of this study was to clarify the effects of daily ingestion of dried-bonito broth on mood states by a pooled analysis of the randomized placebo-controlled trials. Five comparisons in four trials were selected for the pooled analysis (n = 159). The ingestion of dried-bonito broth significantly decreased the scores for fatigue (P = 0.032) and increased those for vigor (P = 0.027) compared to the placebo ingestion, suggesting that the dried-bonito broth improved fatigue felt in daily life. Furthermore, the ingestion of dried-bonito broth decreased the scores for tension-anxiety (P = 0.004) and confusion (P = 0.008) compared to the placebo ingestion. The ingestion of dried-bonito broth also significantly decreased the scores for total mood disturbance (P = 0.005). These results suggest that the ingestion of dried-bonito broth improves mood states especially fatigue, vigor, tension-anxiety and confusion.

  15. Dried-bonito aroma components enhance salivary hemodynamic responses to broth tastes detected by near-infrared spectroscopy.

    PubMed

    Matsumoto, Tomona; Saito, Kana; Nakamura, Akio; Saito, Tsukasa; Nammoku, Takashi; Ishikawa, Masashi; Mori, Kensaku

    2012-01-25

    To elucidate the effects of aroma from dried bonito (katsuo-bushi) on broth tastes caused by the central integration of flavor, optical imaging of salivary hemodynamic responses was conducted using near-infrared spectroscopy (NIRS). A reconstituted dried bonito flavored broth produced a significantly larger hemodynamic response than the odorless broth taste solutions for 5 of the 10 panelists, who felt that the combination of the aroma with the tastes was congruent. In the remaining 5 panelists who felt the combination incongruent, the flavored broth did not cause the enhancement of response. Moreover, when the odor-active smoky parts were removed from the flavoring, the reconstituted flavoring did not enhance the response in the former five panelists. These results indicate that NIRS offers a sensitive method to detect the effect of specific congruent aroma components from dried-bonito broth on the taste-related salivary hemodynamic responses, dependent on the perceptual experience of the combination of aromas and tastes.

  16. Reference Services.

    ERIC Educational Resources Information Center

    Bunge, Charles A.

    1999-01-01

    Discusses library reference services. Topics include the historical development of reference services; instruction in library use, particularly in college and university libraries; guidance; information and referral services and how they differ from traditional question-answering service; and future concerns, including user fees and the planning…

  17. Reference Assessment

    ERIC Educational Resources Information Center

    Bivens-Tatum, Wayne

    2006-01-01

    This article presents interesting articles that explore several different areas of reference assessment, including practical case studies and theoretical articles that address a range of issues such as librarian behavior, patron satisfaction, virtual reference, or evaluation design. They include: (1) "Evaluating the Quality of a Chat Service"…

  18. Reference Assessment

    ERIC Educational Resources Information Center

    Bivens-Tatum, Wayne

    2006-01-01

    This article presents interesting articles that explore several different areas of reference assessment, including practical case studies and theoretical articles that address a range of issues such as librarian behavior, patron satisfaction, virtual reference, or evaluation design. They include: (1) "Evaluating the Quality of a Chat Service"…

  19. Reference Services.

    ERIC Educational Resources Information Center

    Bunge, Charles A.

    1999-01-01

    Discusses library reference services. Topics include the historical development of reference services; instruction in library use, particularly in college and university libraries; guidance; information and referral services and how they differ from traditional question-answering service; and future concerns, including user fees and the planning…

  20. Reference Revolutions.

    ERIC Educational Resources Information Center

    Mason, Marilyn Gell

    1998-01-01

    Describes developments in Online Computer Library Center (OCLC) electronic reference services. Presents a background on networked cataloging and the initial implementation of reference services by OCLC. Discusses the introduction of OCLC FirstSearch service, which today offers access to over 65 databases, future developments in integrated…

  1. Growth of healthy and sanitizer-injured Salmonella cells on mung bean sprouts in different commercial enrichment broths.

    PubMed

    Zheng, Qianwang; Mikš-Krajnik, Marta; D'Souza, Craig; Yang, Yishan; Heo, Da-Jeong; Kim, Si-Kyung; Lee, Seung-Cheol; Yuk, Hyun-Gyun

    2015-12-01

    The ability of nine commercial broths to enrich healthy and 90% sanitizer-injured Salmonella Typhimurium and Salmonella cocktail on mung bean sprouts was evaluated to select an optimum broth for detection. Results showed that S. Typhimurium multiplied faster and reached a higher population in buffered peptone water (BPW), Salmonella AD media (AD) and ONE broth-Salmonella (OB), compared with other broths. Healthy and 90% sanitizer-injured Salmonella at low concentrations increased by 4.0 log CFU/ml in these three broths. However, no Salmonella growth was observed in lactose broth (LB). Further investigation showed that during incubation, pH of LB dropped from 6.7 to 4.2, due to production of lactic (66 mM) and acetic acids (62 mM) by lactic acid bacteria that were identified as dominant microbiota in bean sprouts. Though no cell membrane damage was detected by propidium monoazide combined with real-time PCR, it was found that LB inhibited Salmonella growth, especially from low inoculum levels. This study suggests that in consideration of effectiveness and cost, BPW would be a suitable enrichment broth to use for isolating and detecting Salmonella on mung bean sprouts, while using LB might cause false negative results in Salmonella detection by either PCR or standard cultural method. Copyright © 2015 Elsevier Ltd. All rights reserved.

  2. Crude oil biodegradation aided by biosurfactants from Pseudozyma sp. NII 08165 or its culture broth.

    PubMed

    Sajna, Kuttuvan Valappil; Sukumaran, Rajeev Kumar; Gottumukkala, Lalitha Devi; Pandey, Ashok

    2015-09-01

    The aim of this work was to evaluate the biosurfactants produced by the yeast Pseudozyma sp. NII 08165 for enhancing the degradation of crude oil by a model hydrocarbon degrading strain, Pseudomonas putida MTCC 1194. Pseudozyma biosurfactants were supplemented at various concentrations to the P. putida culture medium containing crude oil as sole carbon source. Supplementation of the biosurfactants enhanced the degradation of crude oil by P. putida; the maximum degradation of hydrocarbons was observed with a 2.5 mg L(-1) supplementation of biosurfactants. Growth inhibition constant of the Pseudozyma biosurfactants was 11.07 mg L(-1). It was interesting to note that Pseudozyma sp. NII 08165 alone could also degrade diesel and kerosene. Culture broth of Pseudozyma containing biosurfactants resulted up to ∼46% improvement in degradation of C10-C24 alkanes by P. putida. The enhancement in degradation efficiency of the bacterium with the culture broth supplementation was even more pronounced than that with relatively purer biosurfactants.

  3. Application of electrodialysis to glycerate recovery from a glycerol containing model solution and culture broth.

    PubMed

    Habe, Hiroshi; Fukuoka, Tokuma; Kitamoto, Dai; Sakaki, Keiji

    2009-04-01

    Glyceric acid is produced by the conversion of glycerol via bioprocesses. The glycerate recovery from model solutions and from real culture broth was demonstrated by a desalting electrodialysis (ED) method. The addition of several impurities in glycerate model solutions, such as polypepton or yeast extract, did not have significant adverse effects on the whole ED process, and more than 93% of the glycerol added in the model solutions (50-150 g/l) was excluded. Using culture broth of Acetobacter tropicalis containing 14.6 g/l D-glycerate, the D-glycerate recovery and the energy consumption were 99.4% and 0.24 kWh/kg, respectively.

  4. Nanofiltration, bipolar electrodialysis and reactive extraction hybrid system for separation of fumaric acid from fermentation broth.

    PubMed

    Prochaska, Krystyna; Staszak, Katarzyna; Woźniak-Budych, Marta Joanna; Regel-Rosocka, Magdalena; Adamczak, Michalina; Wiśniewski, Maciej; Staniewski, Jacek

    2014-09-01

    A novel approach based on a hybrid system allowing nanofiltration, bipolar electrodialysis and reactive extraction, was proposed to remove fumaric acid from fermentation broth left after bioconversion of glycerol. The fumaric salts can be concentrated in the nanofiltration process to a high yield (80-95% depending on pressure), fumaric acid can be selectively separated from other fermentation components, as well as sodium fumarate can be conversed into the acid form in bipolar electrodialysis process (stack consists of bipolar and anion-exchange membranes). Reactive extraction with quaternary ammonium chloride (Aliquat 336) or alkylphosphine oxides (Cyanex 923) solutions (yield between 60% and 98%) was applied as the final step for fumaric acid recovery from aqueous streams after the membrane techniques. The hybrid system permitting nanofiltration, bipolar electrodialysis and reactive extraction was found effective for recovery of fumaric acid from the fermentation broth.

  5. Impact of boiling conditions on the molecular and sensory profile of a vegetable broth.

    PubMed

    Mougin, Alice; Mauroux, Olivier; Matthey-Doret, Walter; Barcos, Eugenia Maria; Beaud, Fernand; Bousbaine, Ahmed; Viton, Florian; Smarrito-Menozzi, Candice

    2015-02-11

    Low-pressure cooking has recently been identified as an alternative to ambient and high-pressure cooking to provide food with enhanced organoleptic properties. This work investigates the impact of the cooking process at different pressures on the molecular and sensory profile of a vegetable broth. Experimental results showed similar sensory and chemical profiles of vegetable broths when boiling at 0.93 and 1.5 bar, while an enhancement of sulfur volatile compounds correlated with a greater leek content and savory aroma was observed when boiling at low pressure (80 °C/0.48 bar). Thus, low-pressure cooking would allow preserving the most labile volatiles likely due to the lower water boiling temperature and the reduced level of oxygen. This study evidenced chemical and sensory impact of pressure during cooking and demonstrated that the flavor profile of culinary preparations can be enhanced by applying low-pressure conditions.

  6. Isolation of ATP from a yeast fermentation broth using a cryogel column at high flow velocities.

    PubMed

    Yan, Chen; Shen, Shaochuan; Yun, Junxian; Wang, Lianghua; Yao, Kejian; Yao, Shan-Jing

    2008-12-01

    This communication presents an effective method for isolating adenosine triphosphate (ATP) from a yeast fermentation broth using an anion-exchange supermacroporous cryogel column at high flow velocities. The breakthrough and elution behaviors of pure ATP in the cryogel bed were investigated at flow velocities of 2, 5, and 10 cm/min and the ATP binding capacities were determined. Then the ATP-containing yeast fermentation broth was employed as the test feedstock and various chromatographic runs were conducted to isolate ATP by the cryogel at different high flow velocities. The ATP samples obtained were analyzed quantitatively by HPLC. The results showed that even at a flow velocity of 5 or 10 cm/min, a product purity of 97.4 or 98.0% can be achieved, illustrating the potential of the present method for separation of high-purity ATP directly from fermentation feedstock at high flow velocities.

  7. Efficient extraction of intracellular reduced glutathione from fermentation broth of Saccharomyces cerevisiae by ethanol.

    PubMed

    Xiong, Zhi-Qiang; Guo, Mei-Jin; Guo, Yuan-Xin; Chu, Ju; Zhuang, Ying-Ping; Zhang, Si-Liang

    2009-01-01

    Reduced glutathione (GSH) from fermentation broth of Saccharomyces cerevisiae was extracted with ethanol without disruption of the cells. The effects of ethanol concentration, extraction temperature and extraction time were assessed by using 2(3) full factorial designs (FFD). Preliminary studies showed that ethanol concentration had the most influence on GSH yield by ethanol extraction, based on the first order regression coefficients derived using MINITAB software, and an optimal ethanol concentration (25%, v/v) was obtained. However, compared to the conventional extraction technique (hot water extraction), there was no significant advantage in yield of GSH from yeast cells using ethanol extraction under these optimized conditions. But ethanol extraction has several advantages, such as lower energy consumption and lower protein concentration of extraction broth, which may reduce the complexity and cost of the purification process. Hence, ethanol extraction which does not disrupt yeast cells could be an inexpensive, simple and efficient alternative to conventional extraction techniques in the GSH industry.

  8. Chromatographic separation of cytidine triphosphate from fermentation broth of yeast using anion-exchange cryogel.

    PubMed

    Wang, Lianghua; Shen, Shaochuan; Yun, Junxian; Yao, Kejian; Yao, Shan-Jing

    2008-03-01

    A novel separation method was developed to isolate directly cytidine triphosphate (CTP) from fermentation broth of yeast using anion-exchange supermacroporous cryogel. The anion-exchange cryogel with tertiary amine groups was prepared by graft polymerization. The breakthrough characteristics and elution performance of pure CTP in the cryogel bed were investigated experimentally and the CTP binding capacity was determined. Then the separation experiments of CTP from crude fermentation broth of yeast using the cryogel column were carried out using deionized water and 0.01 M HCl as washing buffer, respectively. The chromatographic behavior was monitored and analyzed. The purity and concentration of the obtained CTP in these processes were determined quantitatively by HPLC. The maximal purity of CTP obtained at the condition of 0.01 M HCl as washing buffer and 0.5 M NaCl in 0.01 M HCl as elution buffer reached 93%.

  9. In situ separation of lactic acid from fermentation broth using ion exchange resins.

    PubMed

    Ataei, Seyed Ahmad; Vasheghani-Farahani, Ebrahim

    2008-11-01

    Lactic acid fermentation is an end product inhibited reaction. In situ separation of lactic acid from fermentation broth using ion exchange resins was investigated and compared with conventional fermentation system. Amberlite resin (IRA-400, Cl-) was used to separate lactic acid from fermentation broth and pH was controlled online with an automatic pH controller. The effect of process variables on lactic acid production by Lactobacillus casei in whey permeate was studied. The maximum productivity was obtained at pH=6.1, T=37 degrees C and impeller speed=200 rpm. The maximum concentration of lactic acid at optimum condition was found to be 37.4 g/L after 38 h of fermentation using in situ separation system. The productivity of in situ separation system was five times increased in comparison with conventional system.

  10. Comparison of enrichment broths for the recovery of healthy and heat-injured Salmonella typhimurium on raw duck wings.

    PubMed

    Zheng, Qianwang; Bustandi, Caroline; Yang, Yishan; Schneider, Keith R; Yuk, Hyun-Gyun

    2013-11-01

    This study was performed to optimiz eSalmonella Typhimurium recovery from raw duck wings with five nonselective broths (buffered peptone water, tryptic soy broth, lactose broth, universal preenrichment broth, nutrient broth) and four selective broths (selenite broth, BAX System MP media [MP], Salmonella AD media [AD], ONE broth-Salmonella [OB]). Healthy or heat-injured (50 and 85% injury) cells were inoculated at a level of 10(2), 10(1), or 10(0) CFU/25 g on raw duck wings. Growth was modeled using DMfit with four growth parameters: lag-phase duration, maximum growth rate, doubling time, and maximum population density. Most enrichments were able to recover Salmonella Typhimurium to greater than 6 log CFU/ml. AD, MP, and OB had significantly (P < 0.05) higher maximum growth rate (0.9 to 1.0/h) and lower doubling time (0.7 to 0.8 h). Buffered peptone water, AD, MP, and OB recovered healthy and 50%-injured cells at low inoculum levels to more than 6.0 log CFU/ml; OB achieved the greatest recovery (7.6 and 7.9 log CFU/ml), following 24 h of incubation. The 85%-injured cells at 10(0) and 10(1) CFU/25 g, however, were only recovered in OB, reaching 7.3 and 7.5 log CFU/ml, respectively. These results suggest that OB may be an appropriate enrichment broth for the recovery of Salmonella Typhimurium from raw duck wings in standard diagnostic tests or other rapid detection methods, to avoid false-negative results.

  11. Diagnostic assays for identification of microorganisms and antimicrobial resistance determinants directly from positive blood culture broth.

    PubMed

    Pence, Morgan A; McElvania TeKippe, Erin; Burnham, Carey-Ann D

    2013-09-01

    The detection of blood stream infections is one of the most important functions of the clinical microbiology laboratory. Sepsis is a clinical emergency, and mortality increases if commencement of appropriate antimicrobial therapy is delayed. Automated blood culture systems are the most sensitive approach for detection of the causative agent of sepsis. Several laboratory methods have been developed to expedite identification of organisms directly from positive blood culture broth. The principle and analytical performance characteristics of these methods are described in this review.

  12. Antitumor Compounds from the Stout Camphor Mushroom Taiwanofungus camphoratus (Higher Basidiomycetes) Spent Culture Broth.

    PubMed

    Jia, Wei; Bai, Yan-Yan; Zhang, Zhong; Feng, Na; Feng, Jie; Yan, Meng-Qiu; Zhu, Li-Na; Jia, Xin-Cheng; Wang, Ming-Dao; Zhang, Jing-song; Fan, Hua

    2015-01-01

    A known compound, 5-(hydroxymethyl) furan-2-carbaldehyde, and a novel compound, 3-isobutyl-1-methoxy-4-(4'-(3-methylbut-2-enyloxy)phenyl)-1H-pyrrole-2,5-dione were isolated from spent broth from submerged cultures of Taiwanofungus camphoratus. Their structures were elucidated by nuclear magnetic resonance (1H, 13C, and 2D) and mass spectra. These compounds inhibited the proliferation of K562 and HepG2 tumor cells in vitro.

  13. Expression of food-grade phytase in Lactococcus lactis from optimized conditions in milk broth.

    PubMed

    Miao, Yuzhi; Xu, Hui; Fei, Baojin; Qiao, Dairong; Cao, Yi

    2013-07-01

    The major objective of this study was to engineer lactic acid bacteria to produce the enzyme phytase from a gene native to Bacillus subtilis GYPB04. The phytase gene (phyC) of B. subtilis GYPB04 was cloned into the plasmid pMG36e for expression in Lactococcus lactis. The enzyme activity in L. lactis cultured in GM17 broth was 20.25 U/mL at 36°C. The expressed phytase was characterized as active in a pH range of 2.0-9.0 at a temperature range of 20-80°C, with an optimum pH of 5.5-6.5 and temperature of 60°C. When cultured in food-grade milk broth, the transformed L. lactis grew to an OD(600 nm) value of 1.05 and had a phytase yield of 13.58 U/mL. In same broth under optimized conditions for cell growth and phytase production, the transformant reached an OD(600 nm) value of 1.68 and a phytase yield of 42.12 U/mL, representing approximately 1.6-fold and 3.1-fold increases, respectively, compared to growth in natural milk broth. Fermentation was scaled to 5 L under optimized conditions, and product analysis revealed a final OD(600 nm) value of 1.89 and an extracellular enzyme activity of 24.23 U/mL. The results of this study may be used in the dairy fermentation industry for the development of functional, healthy yogurts and other fermented dairy foods that provide both active phytase and viable probiotics to the consumer.

  14. Xanthan gum recovery from fermentation broth using ultrafiltration: Kinetics and process evaluation

    SciTech Connect

    Lo, Y.M.; Yang, S.T.; Min, D.B.

    1995-12-01

    Ultrafiltration of xanthan gum solution as an alternative method to alcohol precipitation for xanthan gum recovery from dilute fermentation broth was studied. A polysulfone membrane (with 500,000 MWCO) hollow fiber (106 mil fiber diameter) tubular cartridge was used to concentrate xanthan broth from less than 3 (w/v) % to {approximately}13.5 (w/v) %, with the xanthan recovery yield of {approximately}95 % or higher. During ultrafiltration, the filtrate flux was one order of magnitude lower for xanthan broth than for water, However, the flux remained almost constant for xanthan concentrations up to {approximately}8%. It was then reduced dramatically as the xanthan concentration increased beyond 8%. The reduced filtrate flux was caused by the reduced pumping (shear) rate and higher viscosities at higher xanthan concentrations. At constant xanthan concentration, the filtrate flux remained almost unchanged for the entire period studied, suggesting that the process is not subject to membrane fouling. In general, the filtrate flux decreased with increasing the xanthan concentration and increased with increasing the pumping (shear) rate and the trans-membrane pressure difference. Changing the solution pH had a slight effect on the viscosity of xanthan solution, but did not affect the filtration performance. Even under high-shear-rate conditions, ultrafiltration did not give any adverse effects on the rheological properties and molecular weight of the xanthan polymer. Thus, ultra filtration can be used to concentrate xanthan broth from fermentation by a factor of four or higher and to reduce the subsequent alcohol recovery costs by at least 75 %.

  15. A new bisabolane-type sesquiterpenoid from the fermentation broth of fungus Antrodiella gypsea.

    PubMed

    Zhao, Zhen-Zhu; He, Li-Qiang; Chen, He-Ping; Li, Zheng-Hui; Dong, Ze-Jun; Feng, Tao; Liu, Ji-Kai

    2016-01-01

    Studies of the fermentation broth of fungus Antrodiella gypsea led to the isolation of a new bisabolane-type sesquiterpenoid that was named gypseatriol (1), together with the known compound 2,10-dodecadiene-1,6,7-triol (2). The structure of this new metabolite was assigned by analysis of 2D NMR and HR-EI-MS. Absolute configuration was assigned by single crystal X-ray diffraction analysis. Compound 1 was evaluated for its antifungal activity on Candida albicans.

  16. Chemical constituents of the fermentation broth of the marine-derived fungus Penicillium roqueforti.

    PubMed

    Mioso, Roberto; Marante, Francisco Javier Toledo; Laguna, Irma Herrera Bravo de

    2015-01-01

    The filamentous fungus Penicillium roqueforti is a well-known multifunctional cell factory of high added-value biomolecules. The objective of this work was to carry out a detailed analysis of the metabolites present in the culture broth of a new marine-derived Penicillium roqueforti strain isolated in the Canary Islands, Spain. The fungal biomass production was carried out in liquid-state fermentation, and after 10-12 days of incubation at 22-25°C, the supernatant mycelia was separated by filtration, and the culture broth (12l) was stored in a refrigerator at 4°C for a subsequent liquid-liquid extraction with dichloromethane (3×), in accordance with the modified Kupchan method. The volatile and semi-volatile organic compounds were separated by chromatography and analyzed using GC-MS and NMR spectroscopy analyses. Several volatile organic compounds involved in the fatty acid pathway were identified: a terpenoid, a cyclic dipeptide, phthalates, and an alkyl adipate. In addition, three categories of non-volatile compounds (alkanes, fatty acids and 1-alkanols) were identified by spectroscopy. The results show that the fermented broth of this fungal strain has no mycotoxins under the culture conditions applied. It is hoped that this chemo-specific information will offer critical input for improving the biotechnological applications of this filamentous fungus. Copyright © 2013 Revista Iberoamericana de Micología. Published by Elsevier Espana. All rights reserved.

  17. Recovery of ammonium lactate and removal of hardness from fermentation broth by nanofiltration.

    PubMed

    Kang, Sang Hyeon; Chang, Yong Keun; Chang, Ho Nam

    2004-01-01

    Nanofiltration (NF) was investigated as an alternative to desalting electrodialysis (ED) and ion exchange for the recovery of ammonium lactate from fermentation broth. Three commercial NF membranes, NF45, NF70, and NTR-729HF, were characterized with 50 mM NaCl, MgSO(4), and glucose solutions. NF45 membrane was selected because it showed the lowest rejection of monovalent ion, the highest rejection of divalent ion, and the highest rejection of nonpolar molecule. Effects of the operating pressure were investigated in a range of 100-400 psig, on the flux, lactate recovery, and glucose and magnesium removal from a real fermentation broth containing about 1.0 M of ammonium lactate. The flux and recovery rate increased linearly with the pressure. However, lactate rejection also increased with the pressure, lowering the recovery yield. More magnesium ions and glucose were rejected as the pressure was increased, and at 400 psig, for example, magnesium ion was almost completely rejected, highlighting the chance of obviating the necessity of ion exchange to remove hardness, by using NF instead of desalting ED. Membrane fouling was not so severe as expected, considering the complex nature and a rather high concentration of the fermentation broth treated.

  18. Direct measurement of the yield stress of filamentous fermentation broths with the rotating vane technique.

    PubMed

    Leong-Poi, L; Allen, D G

    1992-07-01

    The existence of a yield stress in filamentous fermentation broths has important transport phenomena implications in the design and operation of bioreactors. In this study, the constant shear rate vane method was assessed for directly measuring the yield stress of filamentous Aspergillus niger fermentation broths, as well as model fluids (ketchup, yogurt, and pulp suspensions). The method involved rotating 4-, 6-, and 8-bladed vanes (7.2 cm < or = height < or = 15 cm; 4.0 cm < or = dia. < or= 6 cm) at speeds of 0.01 to 0.64 rpm in the fluid and plotting the torque as a function of time. Based on visual observations, the consistency of the results with vane type and speed and comparison with previous work on nonbiological samples, it was concluded that the method is an effective and consistent technique for yield stress measurements on filamentous fermentation broths. Based on comparisons with concentric cylinder viscometer results, it was also concluded that the value determined via the vane method was a "static" yield stress (values of up to 28 Pa) which was much greater than the extrapolated (or "dynamic") yield stress determined via the concentric cylinder viscometer. (c) 1992 John Wiley & Sons, Inc.

  19. Sugaring-out extraction of acetoin from fermentation broth by coupling with fermentation.

    PubMed

    Dai, Jian-Ying; Ma, Lin-Hui; Wang, Zhuang-Fei; Guan, Wen-Tian; Xiu, Zhi-Long

    2017-03-01

    Acetoin is a natural flavor and an important bio-based chemical which could be separated from fermentation broth by solvent extraction, salting-out extraction or recovered in the form of derivatives. In this work, a novel method named as sugaring-out extraction coupled with fermentation was tried in the acetoin production by Bacillus subtilis DL01. The effects of six solvents on bacterial growth and the distribution of acetoin and glucose in different solvent-glucose systems were explored. The operation parameters such as standing time, glucose concentration, and volume ratio of ethyl acetate to fermentation broth were determined. In a system composed of fermentation broth, glucose (100%, m/v) and two-fold volume of ethyl acetate, nearly 100% glucose was distributed into bottom phase, and 61.2% acetoin into top phase without coloring matters and organic acids. The top phase was treated by vacuum distillation to remove solvent and purify acetoin, while the bottom phase was used as carbon source to produce acetoin in the next batch of fermentation.

  20. Characterization of dye decolorization in cell-free culture broth of Trametes versicolor CBR43.

    PubMed

    Ryu, Hyun; Ryu, Hee Wook; Cho, Kyung Suk

    2016-09-23

    The dye decolorization rate in a cell-free culture broth of the white rot fungus Trametes versicolor CBR43 was studied, including the effects of inhibitors of NaCl, Zn(II), and Cd(II) on dye decolorization activity. The maximum rates of dye decolorization in cell-free culture broth were 1410, 44.7, 41.2, and 0.19 μmol L(-1) min(-1) for Acid Blue 62, Acid Black 175, Reactive Blue 4, and Acid Red 114, respectively. The inhibition effects of NaCl, Zn(II), and Cd(II) on dye decolorization were quantitatively compared using the half maximal inhibition concentration (IC50), which indicates the concentration of an inhibitor required for 50% inhibition. Based on IC50 values, dye decolorization in the cell-free culture broth of CBR43 was most potently inhibited by Cd(II), whereas the inhibitory effect of NaCl was relatively low. The dye decolorization rates and IC50 data can be used in the design and development of a dye-wastewater treatment process using T. versicolor CBR43 and its operating factors.

  1. Use of potato extract broth for culturing root-nodule bacteria.

    PubMed

    Martyniuk, Stefan; Oroń, Jadwiga

    2011-01-01

    Liquid media containing potato extract and 1% of glucose or sucrose were used to culture root-nodule bacteria (rhizobia) in shaken Erlenmeyer flasks. For comparison, these bacteria were also cultured in yeast extract-mannitol broth (YEMB) as a standard medium. Proliferation of rhizobia was monitored by measuring optical densities (OD550) of the cultures and by plate counting of the viable cells (c.f.u) of the bacteria. In general, multiplication of the rhizobia in potato extract-glucose broth (PEGB) and potato extract-sucrose broth (PESB) was markedly faster, as indicated by higher values of OD550, than in YEMB. The numbers of R. leguminosarum by. vicae GGL and S. meliloti 330 in PEGB and PEGB were high and ranged from 1.2 x 10(10) to 4.9 x 10(10) mL(-1) after 48 h of incubation at 28 degrees C. B. japonicum B3S culture in PEGB contained 6.4 x 10(9) c.f.u. ml(-1) after 72 h of incubation. PEGB and YEMB cultures of the rhizobia were similar with respect to their beneficial effects on nodulation of the host-plants of these bacteria.

  2. Volatile flavor constituents in the pork broth of black-pig.

    PubMed

    Zhao, Jian; Wang, Meng; Xie, Jianchun; Zhao, Mengyao; Hou, Li; Liang, Jingjing; Wang, Shi; Cheng, Jie

    2017-07-01

    Pork of black-pig in China is well known for its quality and preferred by consumers. However, there is a lack of research on its flavors. By solvent assisted flavor evaporation combined with GC-MS, 104 volatile compounds in the stewed pork broth of black-pig were identified with the dominant amounts of fatty acids, alcohols, and esters. By aroma extract dilution analysis-GC-O method, 27 odor-active compounds were characterized, including 2-methyl-3-furanthiol, 3-(methylthio)propanal, 2-furfurylthiol, γ-decalactone, nonanal, (E)-2-nonenal, and (E,E)-2,4-decadienal that had high FD factors. Compared to the common white-pig, the aroma compounds in both pork broths were almost the same, but the aroma profile of potent odorants for the black-pig pork broth showed less fatty and more roasted notes, which were partially attributed to the higher monounsaturated fatty acids and lower polyunsaturated fatty acids in meat. With aid of authentic chemicals and selected reaction monitoring mode of GC-MS/MS, 19 aroma compounds were quantitated. Copyright © 2017 Elsevier Ltd. All rights reserved.

  3. Modeling of direct recovery of lactic acid from whole broths by ion exchange adsorption.

    PubMed

    Sosa, A V; Ochoa, J; Perotti, N I

    2000-01-01

    Lactic acid fermentation process with L. casei CRL 686 was performed. The static adsorption isotherm over a strong anionic exchange resin, Amberlite IRA-400 was measured, and the static binding capacity parameters were quantified. Early recovery of lactic acid from this lactate producer from unclarified culture broth was performed in a liquid solid fluidized bed, with the resin as the solid adsorbent, and the dynamic adsorption capacity was calculated. Good agreement was found between static and dynamic binding capacity values. The fluidized bed height was twice the settled bed height and the overall process was controlled by the liquid solid mass transfer. This operation was also simulated by continuously well stirred tanks arranged in series and superficial solid deactivation as in a gas solid catalytic reactor. The deactivation process takes into account liquid channeling and agglomerations of solid induced by the viscosity of the broth and also by the cells during the adsorption. These patterns were also verified by experimental observations, and are in agreement with the results found in the literature. The breakthrough data together with others from previous works were satisfactorily fitted until the 90% dimensionless concentration was reached for both culture broths. The model could be used in future studies on predictions about the liquid solid fluidized bed behavior and other different operating conditions.

  4. UV-Heat Treatments for the Control of Foodborne Microbial Pathogens in Chicken Broth

    PubMed Central

    Gouma, M.; Gayán, E.; Raso, J.; Condón, S.; Álvarez, I.

    2015-01-01

    This investigation established the process criteria for using UV-C light and mild heat (UV-H treatment) to inactivate 5-Log10 cycles (performance criterion) of common foodborne pathogen populations, Escherichia coli, Salmonella Typhimurium, Listeria monocytogenes, and Staphylococcus aureus, when inoculated in chicken broth. To define the target microorganism and the proper UV-H treatment conditions (including UV dose, treatment time, and temperature) that would achieve the stated performance criterion, mathematical equations based on Geeraerd's model were developed for each microorganism. For the sake of comparison, inactivation equations for heat treatments were also performed on the same chicken broth and for the same microorganisms. L. monocytogenes was the most UV-H resistant microorganism at all temperatures, requiring a UV dose between 6.10 J/mL (5.6 min) and 2.26 J/mL (2.09 min) to achieve 5-Log10 reductions. In comparison with UV treatments at room temperatures, the combination of UV and mild heat allowed both the UV dose and treatment time to be reduced by 30% and 63% at 55°C and 60°C, respectively. Compared to heat treatments, the UV-H process reduced the heating time for 5-Log10 reductions of all the investigated microorganisms in chicken broth from 20-fold to 2-fold when the operating temperature varied from 53 to 60°C. PMID:26539493

  5. Separation of salvianic acid A from the fermentation broth of engineered Escherichia coli using macroporous resins.

    PubMed

    Bai, Chen-Long; Zhao, Guang-Rong

    2015-08-01

    Salvianic acid A (also known as danshensu) is a plant-derived polyphenolic acid, and has a variety of physiological and pharmacological activities. Our laboratory previously constructed an unprecedented artificial biosynthetic pathway in Escherichia coli and established the fermentation process to produce salvianic acid A. Here, we developed an efficient method for separating salvianic acid A from the fermentation broth of engineered Escherichia coli by macroporous resins. Among ten tested macroporous resins, the static and dynamic adsorption/desorption experiments demonstrated that X5 resin was the best to separate salvianic acid A from fermentation broth. Other parameters during static and dynamic procedures were also investigated. Under the optimum separation conditions, the average adsorption capacity of SAA were 10.66±0.54 mg/g dry resin and the desorption ratio was 85.6±4.1%. The purity and recovery yield of salvianic acid A in the final dry product were 90.2±1.5 and 81.5±2.3%, respectively. The results show that adsorption separation with macroporous resin X5 was an efficient method to prepare salvianic acid A from fermentation broth. This work will benefit the development and application of plant-derived salvianic acid A and its derivatives. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  6. Separation and purification of γ-aminobutyric acid from fermentation broth by flocculation and chromatographic methodologies.

    PubMed

    Gao, Qiang; Duan, Qiang; Wang, Depei; Zhang, Yunze; Zheng, Chunyang

    2013-02-27

    To date, the multifunctional γ-aminobutyric acid (GABA) is mainly produced by microbial fermentation in industry. The purpose of this study was to find an effective method for separation and purification of 31.2 g/L initial GABA from the fermentation broth of Enterococcus raffinosus TCCC11660. To remove the impurities from fermentation broth, flocculation pretreatment using chitosan and sodium alginate was first implemented to facilitate subsequent filtration. Ultrafiltration followed two discontinuous diafiltration steps to effectively remove proteins and macromolecular pigments, and the resulting permeate was further decolored by DA201-CII resin at a high decoloration ratio and GABA recovery. Subsequently, ion exchange chromatography (IEC) with Amberlite 200C resin and gradient elution were applied for GABA separation from glutamate and arginine. Finally, GABA crystals of 99.1% purity were prepared via warm ethanol precipitation twice. Overall, our results reveal that the successive process including flocculation, filtration, ultrafiltration, decoloration, IEC, and crystallization is promising for scale-up GABA extraction from fermentation broth.

  7. HPLC-ELSD determination of kanamycin B in the presence of kanamycin A in fermentation broth.

    PubMed

    Zhang, Yong; He, Hui-Min; Zhang, Jin; Liu, Feng-Jiao; Li, Chao; Wang, Bing-Wu; Qiao, Ren-Zhong

    2015-03-01

    A novel method for the direct determination of kanamycin B in the presence of kanamycin A in fermentation broth using high performance liquid chromatography with evaporative light scattering detector (HPLC-ELSD) was developed. An Agilent Technologies C18 column was utilized, evaporation temperature of 40°C and nitrogen pressure of 3.5 bar, the optimized mobile phase was water-acetonitrile (65:35, v/v), containing 11.6 mm heptafluorobutyric acid (isocratic elution with flow rate of 0.5 mL/min) with the gain 11. Kanamycin B was eluted at 5.6 min with an asymmetry factor of 1.827. The method showed good linearity over the concentration range of 0.05 to 0.80 mg/mL for the kanamycin B (r(2) = 0.9987). The intra-day and inter-day coefficients of variation obtained from kanamycin B were less than 4.3%. Mean recovery of kanamycin B from spiked fermentation broth was 95%. The developed method was applied to the determination of kanamycin B without any interference from other constituents in the fermentation broth. This method offers simple, rapid and quantitative detection of kanamycin B. Copyright © 2014 John Wiley & Sons, Ltd.

  8. Superparamagnetic poly(methyl methacrylate) beads for nattokinase purification from fermentation broth.

    PubMed

    Yang, Chengli; Xing, Jianmin; Guan, Yueping; Liu, Huizhou

    2006-09-01

    An effective method for purification of nattokinase from fermentation broth using magnetic poly(methyl methacrylate) (PMMA) beads immobilized with p-aminobenzamidine was proposed in this study. Firstly, magnetic PMMA beads with a narrow size distribution were prepared by spraying suspension polymerization. Then, they were highly functionalized via transesterification reaction with polyethylene glycol. The surface hydroxyl-modified magnetic beads obtained were further modified with chloroethylamine to transfer the surface amino-modified magnetic functional beads. The morphology and surface functionality of the magnetic beads were examined by scanning electron microscopy and Fourier transform infrared. An affinity ligand, p-aminobenzamidine was covalently immobilized to the amino-modified magnetic beads by the glutaraldehyde method for nattokinase purification directly from the fermentation broth. The purification factor and the recovery of the enzyme activity were found to be 8.7 and 85%, respectively. The purification of nattokinase from fermentation broth by magnetic beads only took 40 min, which shows a very fast purification of nattokinase compared to traditional purification methods.

  9. Ready Reference.

    ERIC Educational Resources Information Center

    Koltay, Emery

    1999-01-01

    Includes the following ready reference information: "Publishers' Toll-Free Telephone Numbers"; "How to Obtain an ISBN (International Standard Book Number)"; "How to Obtain an ISSN (International Standard Serial Number)"; and "How to Obtain an SAN (Standard Address Number)". (AEF)

  10. Highly specific separation for antitumor Spiropreussione A from endophytic fungal [Preussia sp.] fermentation broth by one-step macroporous resins AB-8 treatment.

    PubMed

    Li, Bing; Wang, Chunlan; Chen, Xiaomei; Lyu, Jing; Guo, Shunxing

    2013-11-01

    It is attractive to pharmaceutical works to seek useful material from endophytic fungi. Spiropreussione A (SA) which is isolated from endophytic fungus Preussia sp. is a novel anti-tumor product. Since previous preparation method cannot provide enough SA, and considering the large volume of broth and low concentration of the target product, macroporous resins were introduced to separate SA in our study. Four kinds of macroporous resins ADS-8, H103, X-5 and AB-8 were applied for separating SA, and AB-8 was selected as the optimal resin according to its performances through static and dynamic measurements. HPLC was used to analyze SA in all samples. Under optimal conditions, the specific SA adsorption capacity of AB-8 resin was 15.23mg/g, and the purity increased by 2.5-fold from 35.0% in broth to 90.0% in eluent with 70.0% recovery yield by a one-step treatment. Conclusively, our study achieved the goal of separating and purifying SA in high efficiency, and offered references for further fermentation works. Copyright © 2013 Elsevier B.V. All rights reserved.

  11. Significant decrease of broth viscosity and glucose consumption in erythromycin fermentation by dynamic regulation of ammonium sulfate and phosphate.

    PubMed

    Chen, Yong; Wang, Zejian; Chu, Ju; Zhuang, Yingping; Zhang, Siliang; Yu, Xiaoguang

    2013-04-01

    In this study, the effects of nitrogen sources on broth viscosity and glucose consumption in erythromycin fermentation were investigated. By controlling ammonium sulfate concentration, broth viscosity and glucose consumption were decreased by 18.2% and 61.6%, respectively, whereas erythromycin biosynthesis was little affected. Furthermore, erythromycin A production was increased by 8.7% still with characteristics of low broth viscosity and glucose consumption through the rational regulations of phosphate salt, soybean meal and ammonium sulfate. It was found that ammonium sulfate could effectively control proteinase activity, which was correlated with the utilization of soybean meal as well as cell growth. The pollets formation contributed much to the decrease of broth viscosity. The accumulation of extracellular propionate and succinate under the new regulation strategy indicated that higher propanol consumption might increase the concentration of methylmalonyl-CoA and propionyl-CoA and thus could increase the flux leading to erythromycin A. Copyright © 2013 Elsevier Ltd. All rights reserved.

  12. Antifungal susceptibility profile in vitro of Sporothrix schenckii in two growth phases and by two methods: microdilution and E-test.

    PubMed

    Gutierrez-Galhardo, Maria C; Zancopé-Oliveira, Rosely M; Monzón, Araceli; Rodriguez-Tudela, Juan L; Cuenca-Estrella, Manuel

    2010-05-01

    The susceptibility profile of 91 Sporothrix schenckii isolates in both growth phases was determined by microdilution test (Antifungal Susceptibility Testing of the European Committee for Antimicrobial Susceptibility Testing; AFST-EUCAST). Amphotericin B (AMB), itraconazole (ITC), posaconazole, ravuconazole and terbinafine were found active in vitro against both phases but minimum inhibitory concentrations values for mycelial phase were significantly higher. Fluconazole (FLC) and voriconazole (VRC) were inactive in vitro against both phases. The E-test technique was also performed with 41 representative isolates for AMB, FLC, ITC and VRC. Average agreement rates between yeast phase microdilution results and E-test results were high for AMB (77.5%) and FLC (87.8%), but low for ITC and VRC with rates of 56.4% and 54.5%, respectively. AFST-EUCAST is not the most recommended test to perform drug susceptibility testing of S. schenckii in clinical laboratories, and E-test could be an alternative methodology for this purpose, mainly when the activity in vitro of antifungal agents of AMB and FLC are evaluated.

  13. Commensal symbiosis between a Lactococcus lactis strain and an Enterococcus mundtii strain increases cell yield in constituted broth.

    PubMed

    Kimoto-Nira, H; Ohmori, H; Suzuki, C

    2012-11-01

    To exert their beneficial effects, probiotics need to survive in the stringent conditions of the gastrointestinal tract. Symbiosis between different bacteria is a potential way of enhancing this survival. In developing new probiotic cultures, we investigated the synergic effect between Enterococcus mundtii IFO 13712 and 7 strains of Lactococcus lactis, many of which are widely used as starter bacteria for making dairy products and have probiotic properties. The growth yield of a mixed culture of L. lactis strain Y and IFO 13712 in de Man, Rogosa, and Sharpe broth was greater than that of a single culture. Supernatant from culture of strain IFO 13712 enhanced the growth of strain Y, but that of strain Y did not enhance the growth of strain IFO 13712. This commensalism phenomenon was confirmed by using a simpler tryptone-yeast extract-glucose (TYG) broth. Increased cell yield in mixed culture of the 2 strains compared with single cultures was observed in TYG broth in the presence of both Tween 80 and citrate but not in TYG broth alone or TYG broth containing either Tween 80 or citrate. Thus, the Tween 80 and citrate in the broth contributed to the commensalism. Metabolite analysis revealed that ethanol production in the co-metabolism of glucose and citrate by strain Y was suppressed by mixed culture in TYG broth containing Tween 80 and citrate, compared with that in TYG broth containing citrate alone. The mechanism supporting the observed commensal symbiosis between strains Y and IFO 13712 was the increase in availability of glucose for lactate production by strain Y because, in glycolysis, the pathway from glucose to lactate is energic, whereas the pathway from glucose to ethanol is not. Whether growth stimulation of strain Y by mixing it with IFO 13712 in milk products will enhance the survival of strain Y in the intestine remains to be elucidated.

  14. Effect of dried-bonito broth on mental fatigue and mental task performance in subjects with a high fatigue score.

    PubMed

    Kuroda, Motonaka; Ishizaki, Taichi; Maruyama, Tomoaki; Takatsuka, Yoji; Kuboki, Tomifusa

    2007-12-05

    Dried-bonito broth is commonly employed as a soup and sauce base in Japanese cuisine and is considered to be a nutritional supplement that promotes recovery from fatigue. Previous human trials suggest that the ingestion of dried-bonito broth improves several mood states; however, its effect on fatigue has not yet been clarified. The aim of this study was to clarify the effects of daily ingestion of dried-bonito broth on fatigue and cognitive parameters by a placebo-controlled double blind crossover trial. Forty-eight subjects with fatigue symptoms ingested the dried-bonito broth or a placebo solution every day for 4 weeks. Mood states were evaluated by the Profile of Mood States (POMS), and mental task performance was evaluated by the Uchida-Kraepelin psychodiagnostic (UKP) test. Fatigue and total mood disturbance (TMD) scores on the POMS test decreased significantly during the dried-bonito broth ingestion (p<0.05), but did not change significantly during placebo ingestion. The change in vigor score during dried-bonito broth ingestion was significantly higher than that during placebo ingestion at 2 weeks (p<0.05). The results of the UKP test indicate that the numbers of both total answers and correct answers significantly increased during dried-bonito broth ingestion (p<0.05), while no significant changes were observed in the placebo ingestion. These results suggest that the daily ingestion of dried-bonito broth may improve the mood states, may reduce mental fatigue and may increase performance on a simple calculation task.

  15. Esterification synthesis of ethyl oleate in solvent-free system catalyzed by lipase membrane from fermentation broth.

    PubMed

    Li, Wei-Na; Chen, Bi-Qiang; Tan, Tian-Wei

    2011-01-01

    In this study, the immobilized lipase was prepared by fabric membrane adsorption in fermentation broth. The lipase immobilization method in fermentation broth was optimized on broth activity units and pH adjustments. The viscose fermentation broth can be used with a certain percentage of dilution based on the original broth activity units. The fermentation broth can be processed directly without pH adjustment. In addition, the oleic acid ethyl ester production in solvent-free system catalyzed by the immobilized lipase was optimized. The molar ratio of ethanol to oil acid, the enzyme amount, the molecular amount, and the temperature were 1:1, 12% (w/w), 9% (w/w)(based the total amount of reaction mixture), and 30 °C, respectively. Finally, the optimal condition afforded at least 19 reuse numbers with esterification rate above 80% under stepwise addition of ethanol. Due to simple lipase immobilization preparation, acceptable esterification result during long-time batch reactions and lower cost; the whole process was suitable for industrial ethyl oleate production.

  16. Poroelastic references

    SciTech Connect

    Morency, Christina

    2014-12-12

    This file contains a list of relevant references on the Biot theory (forward and inverse approaches), the double-porosity and dual-permeability theory, and seismic wave propagation in fracture porous media, in RIS format, to approach seismic monitoring in a complex fractured porous medium such as Brady?s Geothermal Field.

  17. Reference Roundup.

    ERIC Educational Resources Information Center

    Silver, Linda; And Others

    1982-01-01

    Briefly describes the nature and availability of reference books for children and adolescents and then reviews some recent publications of this type, including works of a general nature and works on social science, science, the arts, language, history and geography, and biography. (JL)

  18. Ready Reference.

    ERIC Educational Resources Information Center

    Koltay, Emery

    2001-01-01

    Includes four articles that relate to ready reference, including a list of publishers' toll-free telephone numbers and Web sites; how to obtain an ISBN (International Standard Book Number) and an ISSN (International Standard Serial Number); and how to obtain an SAN (Standard Address Number), for organizations that are involved in the book…

  19. Production of staphylococcal enterotoxins in microbial broth and milk by Staphylococcus aureus strains harboring seh gene.

    PubMed

    Schubert, Justyna; Podkowik, Magdalena; Bystroń, Jarosław; Bania, Jacek

    2016-10-17

    Twenty Staphylococcus aureus strains harboring seh gene, including one carrying also sec gene and 11 sea gene, were grown in BHI+YE broth and milk and were tested for SEA, SEC and SEH production. All strains decreased pH of BHI+YE broth at 24h and increased them at 48h. Seventeen S. aureus strains grown in milk changed pH for no >0.3 unit until 48h. Three other S. aureus strains significantly decreased pH during growth in milk. All S. aureus produced SEH in BHI+YE broth in amounts ranging from 95 to 1292ng/ml, and from 170 to 4158ng/ml at 24 and 48h, respectively. SEH production in milk by 17 strains did not exceed 23ng/ml at 24h and 36ng/ml at 48h. Three S. aureus strains able to decrease milk pH produced 107-3029ng/ml and 320-4246ng/ml of SEH in milk at 24 and 48h, respectively. These strains were grown in milk and BHI+YE broth with pH stabilized at values near neutral leading to a significant decrease of SEH production. Representative weak SEH producers were grown in milk at reduced pH resulting in moderate increase in SEH production. SEA was produced in milk by 10S. aureus strains at 24-151ng/ml at 24h, and 31-303ng/ml at 48h. SEA production in milk was higher or comparable as in BHI+YE broth in 3 strains and lower for remaining strains. Production of SEC by sec-positive S. aureus strains was lower in milk than in BHI+YE broth, ranging from 131 to 2319ng/ml at 24 and 48h in milk and 296-30,087ng/ml in BHI+YE at 24 and 48h. Both lacE and lacG transcripts involved in lactose metabolism were significantly up-regulated in milk in strong SEH producers. In these strains hld, rot and sarA transcripts were up-regulated in milk as compared to weak SEH producers. Stabilization of milk pH at a value of raw milk significantly down-regulated hld, rot and sarA RNA in strong SEH producers. Milk was generally found unfavorable for enterotoxin production. However, certain S. aureus strains were not restricted in SEH and SEA expression in milk, unlike SEC which remained down

  20. Automated harvesting and 2-step purification of unclarified mammalian cell-culture broths containing antibodies.

    PubMed

    Holenstein, Fabian; Eriksson, Christer; Erlandsson, Ioana; Norrman, Nils; Simon, Jill; Danielsson, Åke; Milicov, Adriana; Schindler, Patrick; Schlaeppi, Jean-Marc

    2015-10-30

    Therapeutic monoclonal antibodies represent one of the fastest growing segments in the pharmaceutical market. The growth of the segment has necessitated development of new efficient and cost saving platforms for the preparation and analysis of early candidates for faster and better antibody selection and characterization. We report on a new integrated platform for automated harvesting of whole unclarified cell-culture broths, followed by in-line tandem affinity-capture, pH neutralization and size-exclusion chromatography of recombinant antibodies expressed transiently in mammalian human embryonic kidney 293T-cells at the 1-L scale. The system consists of two bench-top chromatography instruments connected to a central unit with eight disposable filtration devices used for loading and filtering the cell cultures. The staggered parallel multi-step configuration of the system allows unattended processing of eight samples in less than 24h. The system was validated with a random panel of 45 whole-cell culture broths containing recombinant antibodies in the early profiling phase. The results showed that the overall performances of the preparative automated system were higher compared to the conventional downstream process including manual harvesting and purification. The mean recovery of purified material from the culture-broth was 66.7%, representing a 20% increase compared to that of the manual process. Moreover, the automated process reduced by 3-fold the amount of residual aggregates in the purified antibody fractions, indicating that the automated system allows the cost-efficient and timely preparation of antibodies in the 20-200mg range, and covers the requirements for early in vitro and in vivo profiling and formulation of these drug candidates. Copyright © 2015 The Authors. Published by Elsevier B.V. All rights reserved.

  1. Shock Compression and Recovery of Microorganism-Loaded Broths and AN Emulsion

    NASA Astrophysics Data System (ADS)

    Hazell, P. J.; Beveridge, C.; Groves, K.; Stennett, C.

    2009-12-01

    The microorganisms Escherichia coli, Enterococcus faecalis and Zygosaccharomyces bailii and an oil-based emulsion, have been subjected to shock compression using the flyer-plate technique to initial pressures of 0.8 GPa (in the suspension). In each experiment, a stainless steel capsule was used to contain the broths and allow for recovery without contamination. Where cavitation was mostly suppressed by virtue of simultaneous shock and dynamic compression, no kill was observed. By introducing an air gap behind the suspension, limited kill was measured in the yeast. Results also suggest that stable emulsification occurs in coarse oil-based emulsions that are subjected to shock.

  2. Shock compression and recovery of microorganism-loaded broths and an emulsion

    NASA Astrophysics Data System (ADS)

    Hazell, Paul; Beveridge, Cliff; Groves, Kathy

    2009-06-01

    The microorganisms Escherichia coli, Enterococcus feacalis and Zygosaccharomyces bailii and an oil-based emulsion, have been subjected to shock compression using the flyer-plate technique to initial pressures of 0.8 GPa (in the suspension). In each experiment, a stainless steel capsule was used to contain the broths and allow for recovery without contamination. Where cavitation was suppressed by virtue of simultaneous shock and quasi-static compression, no kill was observed. By introducing an air gap behind the suspension, limited kill was measured in the yeast. Results also suggest that emulsification occurs in oil-based emulsions that are subjected to shock.

  3. [Determination of organic acids in fermentation broth of spiramycin by high performance liquid chromatography].

    PubMed

    Li, You-yuan; Chen, Chang-hua; Tao, Ping

    2002-01-01

    A method for determining organic acids in spiramycin fermentation broth by high performance liquid chromatography is described. The operating conditions were Zorbax 300-SB C18 column (5 microns, 4.6 mm i.d. x 15 cm) at 35 degrees C, 0.01 mol/L phosphoric acid buffer solution (pH 2.32) and methanol as mobile phase at a flow rate of 0.6 mL/min and UV detection at 210 nm. The relative standard deviations were 0.33%-0.10% and the recoveries were 99.95%-100.08%. It's a simple, rapid and accurate method.

  4. [Analysis of oxygen transfer in bioreactors for fungus broths. 2. Suspensions of P. chrysogenum mycelial associations].

    PubMed

    Caşcaval, D; Galaction, Anca-Irina; Cămăruţ, Stefănica

    2009-01-01

    Unlike the P. shermanii and S. cerevisiae cultures, the study on the distribution of oxygen transfer in stirred bioreactor for P. chrysogenum pellets broths indicated that this process is controlled mainly by the deposition tendency of the biomass. Similar to the previously studied systems, the analysis of k(1)a distribution indicated its heterogeneity on the bioreactor height, the oxygen transfer rate increasing from position 1 to 4. Contrary to the bacterial and yeasts cultures, the intensification of aeration promoted the initial reduction of k(1)a, which reached a minimum level, followed by its increase, due to the flooding phenomenon.

  5. Antibacterial activity composition of the fermentation broth of Streptomyces djakartensis NW35.

    PubMed

    Zhang, Wenjuan; Wei, Shaopeng; Zhang, Jiwen; Wu, Wenjun

    2013-03-01

    The new compound Z-4-2 was isolated from the fermentation broth of Streptomyces djakartensis NW35, together with the known compound N-acetyltryptamine (Z-9-2) by bioassay-guided fractionation. Its chemical structure was elucidated as (E)-2-methoxy-1,4 naphthoquinone-1-oxime (Z-4-2) mainly by NMR analyses and MS spectral data. Their antibacterial activities against bacteria were evaluated by the filter paper method. The results of indicated that these compounds possess significant antibacterial activities.

  6. Quality, antioxidative ability, and cell proliferation-enhancing activity of fermented black soybean broths with various supplemental culture medium.

    PubMed

    Lin, Chih-Chien; Wu, Pey-Shiuan; Liang, David Woei-Ming; Kwan, Chang-Chin; Chen, Yi-Shyan

    2012-01-01

    The fermented soybean-based foods have played an important role in traditional diets around the world for many centuries, and Bacillus subtilis is typically used in the fermentation of soybean-based foods. The fermentation process may improve not only the flavor but also the nutritional value of food, and substances produced in this fermented broth were affected by many factors including culture medium and the selected soybeans. In this study, we use 3 potential culture mediums in the fermentation of black soybean and the fermented black soybean broths were used for the examination of amino acid composition, total phenolics content, flavonoids and anthocyanins contents, the antioxidant properties, and cytotoxicity. Our results indicated that the fermented black soybean broth, fermentation III, have the most abundant essential amino acid (79.77 mg/g), phenolics (19.33 mg/g), flavonoids (46.01 mg/g), and anthocyanins (1.06 mg/g). Besides, all of the fermented black soybean broths exhibited the significant antioxidative abilities with 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging effect, reducing power and ferrous ion chelating effect. In addition, the fermented black soybean broths demonstrated the cell proliferation-enhancing activity in Detroit 551 cells. The cells were augmented up to the maximum value of 183.6% (compared with control) at 10 mg/mL of the fermentation I. Therefore, the different supplemental culture medium fermented black soybean broths may be used as a functional ingredient in the products of nutritional drinks and health foods. The present study illustrated the potential of various supplemental culture medium fermented black soybean broths in the application of functional ingredient for nutritional drinks and health foods. © 2011 Institute of Food Technologists®

  7. Rapid determination of lovastatin in the fermentation broth of Aspergillus terreus using dual-wavelength UV spectrophotometry.

    PubMed

    Li, Shi-Weng; Song, Hong-Ping; Leng, Yan

    2014-01-01

    Lovastatin, a hypocholesterolemic drug, is produced by submerged fermentation of Aspergillus terreus Thom (Trichocomaceae). High performance liquid chromatography is usually used to determine lovastatin in samples of the fermentation broth. However, this method is inconvenient and costly, especially in the context of high-throughput sample analysis. A direct and simple dual-wavelength ultraviolet spectrophotometric method for quantifying lovastatin in the fermentation broth of A. terreus was developed. A. terreus Z15-7 was used for all experiments. The liquid fermentation was conducted at 30 °C in a rotary shaker at 150 rpm for 15 d. Silica gel and neutral alumina column chromatography were used for the separation and purification of lovastatin from the fermentation broth. The limits of detection of lovastatin were 0.320 μg/ml in the lovastatin standard solution and 0.490 μg/ml in the fermentation broth sample and the limits of quantification of lovastatin were 1.265 μg/ml in the lovastatin standard solution and 3.955 μg/ml in the fermentation broth sample. The amounts of lovastatin in the fermentation broth ranged from 876.614 to 911.967 μg/ml, with relative standard deviations from 1.203 to 1.709%. The mean recoveries of lovastatin using silica gel and neutral alumina column chromatography were 84.2 ± 0.82 and 87.2 ± 0.21%, respectively. Dual-wavelength UV spectrophotometry is a rapid, sensitive, accurate, and convenient method for quantifying lovastatin in fermentation broth. Neutral alumina column chromatography is more efficient than silica gel column chromatography for the purification and determination lovastatin using the developed dual-wavelength UV spectrophotometry method.

  8. Activity of a long-acting echinocandin, CD101, determined using CLSI and EUCAST reference methods, against Candida and Aspergillus spp., including echinocandin- and azole-resistant isolates.

    PubMed

    Pfaller, Michael A; Messer, Shawn A; Rhomberg, Paul R; Jones, Ronald N; Castanheira, Mariana

    2016-10-01

    The objective of this study was to evaluate the in vitro activity of CD101, a novel echinocandin with a long serum elimination half-life, and comparator (anidulafungin and caspofungin) antifungal agents against a collection of Candida and Aspergillus spp. isolates. CD101 and comparator agents were tested against 106 Candida spp. and 67 Aspergillus spp. isolates, including 27 isolates of Candida harbouring fks hotspot mutations and 12 itraconazole non-WT Aspergillus, using CLSI and EUCAST reference susceptibility broth microdilution (BMD) methods. Against WT and fks mutant Candida albicans, Candida glabrata and Candida tropicalis, the activity of CD101 [MIC90 = 0.06, 0.12 and 0.03 mg/L, respectively (CLSI method values)] was comparable to that of anidulafungin (MIC90 = 0.03, 0.12 and 0.03 mg/L, respectively) and caspofungin (MIC90 = 0.12, 0.25 and 0.12 mg/L, respectively). WT Candida krusei isolates were very susceptible to CD101 (MIC = 0.06 mg/L). CD101 activity (MIC50/90 = 1/2 mg/L) was comparable to that of anidulafungin (MIC50/90 = 2/2 mg/L) against Candida parapsilosis. CD101 (MIC mode = 0.06 mg/L for C. glabrata) was 2- to 4-fold more active against fks hotspot mutants than caspofungin (MIC mode = 0.5 mg/L). CD101 was active against Aspergillus fumigatus, Aspergillus terreus, Aspergillus niger and Aspergillus flavus (MEC90 range = ≤0.008-0.03 mg/L). The essential agreement between CLSI and EUCAST methods for CD101 was 92.0%-100.0% among Candida spp. and 95.0%-100.0% among Aspergillus spp. The activity of CD101 is comparable to that of other members of the echinocandin class for the prevention and treatment of serious fungal infections. Similar results for CD101 activity versus Candida and Aspergillus spp. may be obtained with either CLSI or EUCAST BMD methods. © The Author 2016. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy.

  9. Activity of a long-acting echinocandin, CD101, determined using CLSI and EUCAST reference methods, against Candida and Aspergillus spp., including echinocandin- and azole-resistant isolates

    PubMed Central

    Pfaller, Michael A.; Messer, Shawn A.; Rhomberg, Paul R.; Jones, Ronald N.; Castanheira, Mariana

    2016-01-01

    Objectives The objective of this study was to evaluate the in vitro activity of CD101, a novel echinocandin with a long serum elimination half-life, and comparator (anidulafungin and caspofungin) antifungal agents against a collection of Candida and Aspergillus spp. isolates. Methods CD101 and comparator agents were tested against 106 Candida spp. and 67 Aspergillus spp. isolates, including 27 isolates of Candida harbouring fks hotspot mutations and 12 itraconazole non-WT Aspergillus, using CLSI and EUCAST reference susceptibility broth microdilution (BMD) methods. Results Against WT and fks mutant Candida albicans, Candida glabrata and Candida tropicalis, the activity of CD101 [MIC90 = 0.06, 0.12 and 0.03 mg/L, respectively (CLSI method values)] was comparable to that of anidulafungin (MIC90 = 0.03, 0.12 and 0.03 mg/L, respectively) and caspofungin (MIC90 = 0.12, 0.25 and 0.12 mg/L, respectively). WT Candida krusei isolates were very susceptible to CD101 (MIC = 0.06 mg/L). CD101 activity (MIC50/90 = 1/2 mg/L) was comparable to that of anidulafungin (MIC50/90 = 2/2 mg/L) against Candida parapsilosis. CD101 (MIC mode = 0.06 mg/L for C. glabrata) was 2- to 4-fold more active against fks hotspot mutants than caspofungin (MIC mode = 0.5 mg/L). CD101 was active against Aspergillus fumigatus, Aspergillus terreus, Aspergillus niger and Aspergillus flavus (MEC90 range = ≤0.008–0.03 mg/L). The essential agreement between CLSI and EUCAST methods for CD101 was 92.0%–100.0% among Candida spp. and 95.0%–100.0% among Aspergillus spp. Conclusions The activity of CD101 is comparable to that of other members of the echinocandin class for the prevention and treatment of serious fungal infections. Similar results for CD101 activity versus Candida and Aspergillus spp. may be obtained with either CLSI or EUCAST BMD methods. PMID:27287236

  10. Effect of Eleutherine americana Merr. extract on enzymatic activity and enterotoxin production of Staphylococcus aureus in broth and cooked pork.

    PubMed

    Ifesan, Beatrice O T; Voravuthikunchai, Supayang P

    2009-01-01

    Crude ethanolic extract from the bulb of Eleutherine americana was investigated for its inhibitory activities against lipase and protease enzymes and enterotoxin production by Staphylococcus aureus. Eleven isolates that demonstrated high enzyme activity with three reference strains were selected to study the effect of extract on enzyme production. Exposure of the isolates to subminimal inhibitory concentrations, (1/2) minimum inhibitory concentration (MIC) (125 microg/mL), and (1/4)MIC (62.5 microg/mL) of the crude extract resulted in both partial and total inhibition of lipase and protease enzymes. About 15% of the 106 isolates were positive for enterotoxin production with staphylococcal enterotoxin A (11.3%), enterotoxin B (3.7%), and enterotoxin C (10.3%), and no enterotoxin D was produced. The production of staphylococcal enterotoxins A-D in the presence or absence of the crude extract was carried out. In the broth system, the extract reduced enterotoxin production at subminimal inhibitory concentrations compared with the control. At MIC, total enterotoxin inhibition was observed for enterotoxin C production, whereas synthesis of enterotoxins A, B, and D was totally eliminated at 2MIC. The food system study revealed that the extract could delay production of enterotoxins A, B, and C compared with the control. The extract at 2 mg/mL delayed production of toxins A and C for 8 and 4 h, while toxin B was not detected in the pork at 48 h. The ability of E. americana extract to inhibit lipase and protease enzymes and to delay enterotoxin production in food could present it as a novel food additive to combat the growth of S. aureus in food.

  11. Comparative evaluation of the micro-media system, sceptor, and MIC-2000 microdilution methods for testing Pseudomonas aeruginosa against gentamicin, tobramycin, and amikacin.

    PubMed Central

    Woolfrey, B F; Lally, R T; Quall, C O

    1983-01-01

    Minimum inhibitory concentrations (MICs) for selected strains of Pseudomonas aeruginosa versus gentamicin, tobramycin, and amikacin were replicated in parallel with MMS (Micro-Media Systems, Potomac, Md). Sceptor (BBL Microbiology Systems, Cockeysville, Md), and matching MIC-2000 (Dynatech Laboratories, Inc., Alexandria, Va.) twofold dilution panels and with MIC-2000 panels with dilutions differing by small arithmetic increments. The three microdilution systems produced comparable modal MICs. However, dispersion about modal values was greater for MMS than for either Sceptor or comparable MIC-2000 twofold panels. MICs were best define by MIC-2000 panels with dilutions differing by small arithmetic increments, for which 72.9% of MICs were modal, 95.4% were one small increment step or less from the modal value, and 100% were two small-increment steps or less form the modal value. The similarity of MIC replication for Sceptor and MIC-2000 twofold panels suggests the possibility of using small increment dilutions by Sceptor. PMID:6403574

  12. Comparative evaluation of the micro-media system, sceptor, and MIC-2000 microdilution methods for testing Pseudomonas aeruginosa against gentamicin, tobramycin, and amikacin.

    PubMed

    Woolfrey, B F; Lally, R T; Quall, C O

    1983-02-01

    Minimum inhibitory concentrations (MICs) for selected strains of Pseudomonas aeruginosa versus gentamicin, tobramycin, and amikacin were replicated in parallel with MMS (Micro-Media Systems, Potomac, Md). Sceptor (BBL Microbiology Systems, Cockeysville, Md), and matching MIC-2000 (Dynatech Laboratories, Inc., Alexandria, Va.) twofold dilution panels and with MIC-2000 panels with dilutions differing by small arithmetic increments. The three microdilution systems produced comparable modal MICs. However, dispersion about modal values was greater for MMS than for either Sceptor or comparable MIC-2000 twofold panels. MICs were best define by MIC-2000 panels with dilutions differing by small arithmetic increments, for which 72.9% of MICs were modal, 95.4% were one small increment step or less from the modal value, and 100% were two small-increment steps or less form the modal value. The similarity of MIC replication for Sceptor and MIC-2000 twofold panels suggests the possibility of using small increment dilutions by Sceptor.

  13. Lethal paralytic shellfish poisoning from consumption of green mussel broth, Western Samar, Philippines, August 2013.

    PubMed

    Ching, Paola Katrina; Ramos, Ruth Alma; de los Reyes, Vikki Carr; Sucaldito, Ma Nemia; Tayag, Enrique

    2015-01-01

    In July 2013, the Philippines' Event-Based Surveillance & Response Unit received a paralytic shellfish poisoning (PSP) report from Tarangnan, Western Samar. A team from the Department of Health conducted an outbreak investigation to identify the implicated source and risk factors in coastal villages known for green mussel production and exportation. A case was defined as a previously well individual from Tarangan, Western Samar who developed gastrointestinal symptoms and any motor and/or sensory symptoms after consumption of shellfish from 29 June to 4 July 2013 in the absence of any known cause. The team reviewed medical records, conducted active case finding and a case-control study. Relatives of cases who died were interviewed. Sera and urine specimens, green mussel and seawater samples were tested for saxitoxin levels using high performance liquid chromatography. Thirty-one cases and two deaths were identified. Consumption of > 1 cup of green mussel broth was associated with being a case. Seawater sample was positive for Pyrodinium bahamense var. compressum and green mussel samples were positive for saxitoxin. Inspection revealed villagers practice open defecation and improper garbage disposal. This PSP outbreak was caused by the consumption of the green mussel broth contaminated by saxitoxin. As a result of this outbreak, dinoflagellate and saxitoxin surveillance was established, and since the outbreak, there have been no harmful algal blooms event or PSP case reported since. A "Save Cambatutay Bay" movement, focusing on proper waste disposal practice and clean-up drives has been mobilized.

  14. Isolation of brefeldin A from Eupenicillium brefeldianum broth using macroporous resin adsorption chromatography.

    PubMed

    Wang, Ya-Jun; Wu, Ye-Fei; Xue, Feng; Wu, Zhi-Xian; Xue, Ya-Ping; Zheng, Yu-Guo; Shen, Yin-Chu

    2012-05-01

    Brefeldin A (BFA) is a macrolide lactone antibiotic, possessing antitumor, antiviral, antifungal activities. In this work, a separation strategy involving one-step macroporous resin adsorption chromatography combined with crystallization was established for BFA purification from Eupenicillium brefeldianum CCTCC M 208113 fermentation broth. Among six macroporous resin adsorbents tested, the non-polar resin HZ830 had the best adsorption and desorption performance. The static equilibrium adsorption data fitted well with the Freundlich equation, and the adsorption kinetic followed the pseudo-second order model. Through experimental optimization of column adsorption and desorption, BFA in purity of 90.4% (w/w), 92.1% (w/w) yield was obtained by a one-step macroporous resin adsorption chromatography, using a stepwise elution protocol. Furthermore, high purity (>99%, w/w) of BFA crystals were prepared from E. brefeldianum CCTCC M 208113 fermentation broth in an overall recovery of 67.0% (w/w), using a combination of adsorption chromatography packed with non-polar macroporous adsorbent HZ830 and crystallization in acetone. Copyright © 2012 Elsevier B.V. All rights reserved.

  15. Effects of solution environment on mammalian cell fermentation broth properties: enhanced impurity removal and clarification performance.

    PubMed

    Westoby, Matthew; Chrostowski, James; de Vilmorin, Philippe; Smelko, John Paul; Romero, Jonathan K

    2011-01-01

    The processing of recombinant proteins from high cell density, high product titer cell cultures containing mammalian cells is commonly performed using tangential flow microfiltration (MF). However, the increased cellular debris present in these complex feed streams can prematurely foul the membrane, adversely impacting MF capacity and throughput. In addition, high cell density cell culture streams introduce elevated levels of process-related impurities, which increase the burden on subsequent purification operations to remove these complex media components and impurities. To address this challenge, an evaluation of mammalian cell culture broth buffer properties was examined to determine if enhanced impurity removal and clarification performance could be achieved. A framework is presented here for establishing optimized mammalian cell culture buffer conditions, involving trade-offs between product recovery and purification and improved clarification at manufacturing-scale production. A reduction in cell culture broth pH to 4.7-5.0 induced flocculation and impurity precipitation which increased the average feed particle-size. These conditions led to enhanced impurity removal and improved MF throughput and filter capacity for several mammalian systems. Feed conditions were further optimized by controlling ionic composition along with pH to improve product recovery from high cell density/high product titer cell cultures. © 2010 Wiley Periodicals, Inc.

  16. Downstream extraction process development for recovery of organic acids from a fermentation broth.

    PubMed

    Bekatorou, Argyro; Dima, Agapi; Tsafrakidou, Panagiotia; Boura, Konstantina; Lappa, Katerina; Kandylis, Panagiotis; Pissaridi, Katerina; Kanellaki, Maria; Koutinas, Athanasios A

    2016-11-01

    The present study focused on organic acids (OAs) recovery from an acidogenic fermentation broth, which is the main problem regarding the use of OAs for production of ester-based new generation biofuels or other applications. Specifically, 10 solvents were evaluated for OAs recovery from aqueous media and fermentation broths. The effects of pH, solvent/OAs solution ratios and application of successive extractions were studied. The 1:1 solvent/OAs ratio showed the best recovery rates in most cases. Butyric and isobutyric acids showed the highest recovery rates (80-90%), while lactic, succinic, and acetic acids were poorly recovered (up to 45%). The OAs recovery was significantly improved by successive 10-min extractions. Alcohols presented the best extraction performance. The process using repeated extractions with 3-methyl-1-butanol led to the highest OAs recovery. However, 1-butanol can be considered as the most cost-effective option taking into account its price and availability. Copyright © 2016 Elsevier Ltd. All rights reserved.

  17. High pressure inactivation of Pseudomonas in black truffle - comparison with Pseudomonas fluorescens in tryptone soya broth

    NASA Astrophysics Data System (ADS)

    Ballestra, Patricia; Verret, Catherine; Cruz, Christian; Largeteau, Alain; Demazeau, Gerard; El Moueffak, Abdelhamid

    2010-03-01

    Pseudomonas is one of the most common genera in black Perigord truffle. Its inactivation by high pressure (100-500 MPa/10 min) applied on truffles at sub-zero or low temperatures was studied and compared with those of Pseudomonas fluorescens in tryptone soya broth. Pressurization of truffles at 300 MPa/4 °C reduced the bacterial count of Pseudomonas by 5.3 log cycles. Higher pressures of 400 or 500 MPa, at 4 °C or 20 °C, allowed us to slightly increase the level of destruction to the value of ca. 6.5 log cycles but did not permit us to completely inactivate Pseudomonas. The results showed a residual charge of about 10 CFU/g. Pressure-shift freezing of truffles, which consists in applying a pressure of 200 MPa/-18 °C for 10 min and then quickly releasing this pressure to induce freezing, reduced the population of Pseudomonas by 3.3 log cycles. The level of inactivation was higher than those obtained with conventional freezing. Endogenous Pseudomonas in truffle was shown to be more resistant to high pressure treatments than P. fluorescens used for inoculation of broths.

  18. Antibacterial activity of 11 essential oils against Bacillus cereus in tyndallized carrot broth.

    PubMed

    Valero, M; Salmerón, M C

    2003-08-15

    The antibacterial activity of 11 essential oils from aromatic plants against the strain INRA L2104 of the foodborne pathogen Bacillus cereus grown in carrot broth at 16 degrees C was studied. The quantity needed by the essential oils of nutmeg, mint, clove, oregano, cinnamon, sassafras, sage, thyme or rosemary to produce 14-1110% relative extension of the lag phase was determined. Total growth inhibition of bacterial spores was observed for some of the antimicrobial agents assayed. The addition of 5 microl cinnamon essential oil per 100 ml of broth in combination with refrigeration temperatures of

  19. 2,3-Butanediol recovery from fermentation broth by alcohol precipitation and vacuum distillation.

    PubMed

    Jeon, Sangjun; Kim, Duk-Ki; Song, Hyohak; Lee, Hee Jong; Park, Sunghoon; Seung, Doyoung; Chang, Yong Keun

    2014-04-01

    This study presents a new and effective downstream process to recover 2,3-butanediol (2,3-BD) from fermentation broth which is produced by a recombinant Klebsiella pneumoniae strain. The ldhA-deficient K. pneumoniae strain yielded about 90 g/L of 2,3-BD, along with a number of by-products, such as organic acids and alcohols, in a 65 h fed-batch fermentation. The pH-adjusted cell-free fermentation broth was firstly concentrated until 2,3-BD reached around 500 g/L by vacuum evaporation at 50°C and 50 mbar vacuum pressure. The concentrated solution was further treated using light alcohols, including methanol, ethanol, and isopropanol, for the precipitation of organic acids and inorganic salts. Isopropanol showed the highest removal efficiency, in which 92.5% and 99.8% of organic acids and inorganic salts were precipitated, respectively. At a final step, a vacuum distillation process enabled the recovery of 76.2% of the treated 2,3-BD, with 96.1% purity, indicating that fermentatively produced 2,3-BD is effectively recovered by a simple alcohol precipitation and vacuum distillation.

  20. Development of a Novel Listeria Enrichment Broth for the Isolation of Pathogenic Listeria.

    PubMed

    Liu, Dongxin; Wang, Yan; Wang, Yi; Zhang, Lu; Luo, Lijuan; Liu, Kai; Ye, Changyun

    2017-10-01

    Listeriosis, the disease caused by pathogenic Listeria species, can present severe symptoms in susceptible people. The goal of this study was to develop a novel enrichment broth, Listeria allose enrichment broth (LAEB), to improve isolation of Listeria monocytogenes and Listeria ivanovii from samples through incorporating a specific carbohydrate and reducing inhibitor concentrations. Other coexisting bacteria, particularly Listeria innocua, can interfere with the isolation of pathogenic Listeria in such ways as overgrowth of L. innocua and the generation of inhibitory metabolites. The incorporation of allose into the novel LAEB was effective for slowing the growth of L. innocua and other nontarget microorganisms. We determined that 35°C and pH 7.0 under aerobic conditions are optimal for Listeria growth in this medium. The novelty of the use of LAEB is the single enrichment procedure at 35°C for 24 h, obviating the need for a secondary enrichment medium. In 50 simulated samples, the sensitivity of the LAEB method (86%) was higher than that of the International Organization for Standardization (EN ISO) method (70%). In 142 naturally contaminated samples tested, the isolation rate for pathogenic Listeria with the LAEB method was 26.0% (37 of 142 samples), which was significantly higher than the 17.6% (25 of 142 samples) for the EN ISO method. Higher isolation rates and a quicker and easier protocol make the novel LAEB method an appropriate alternative for the isolation of pathogenic Listeria.

  1. Rapid broth macrodilution method for determination of MICs for Mycobacterium avium isolates.

    PubMed Central

    Siddiqi, S H; Heifets, L B; Cynamon, M H; Hooper, N M; Laszlo, A; Libonati, J P; Lindholm-Levy, P J; Pearson, N

    1993-01-01

    A multicenter study was done to investigate the accuracy and reproducibility of a method for determining the MICs of antimicrobial agents against the Mycobacterium avium complex in 7H12 broth with the BACTEC system. In phase I, with eight drugs and 10 strains, intralaboratory reproducibility was 95.7 to 100%, allowing a 1-dilution difference upon repeat testing. The results of phase II testing with 41 additional strains were consistent with those obtained in phase I, with good interlaboratory reproducibility. The radiometric method was validated by sampling and plating of the same broth cultures and determining, by the number of CFU per milliliter, the lowest drug concentration that inhibited more than 99% of the initial bacterial population. Three test concentrations of each drug and the tentative interpretation of results are proposed. Radiometric MIC determination has the potential to become the method of choice for clinical microbiology laboratories and evaluation of new agents for the treatment of M. avium infections, both pulmonary and disseminated. Images PMID:8408551

  2. The effect of diets supplemented with fish broth and fish oil on the health of weaners.

    PubMed

    Bakuła, T; Lis, Ł; Iwaniuk, Z; Ordyński, Z

    2011-01-01

    The aim of this study was to determine the effect of fish-based feed materials, as a source of readily available protein contained in fish broth and essential polyunsaturated fatty acids (PUFAs) found in fish oil, on the health of piglets and rearing results. The experiment was conducted on a commercial pig fattening farm. The study involved a total of 80 weaners with an approximate body weight of 15 kg. The experiment was carried out over a period of 40 days. Feed samples were subjected to laboratory analyses. Blood samples were collected from experimental group animals to determine serum biochemical and immunological parameters. The body weight gains of weaners, mortality rates and average feed intake per animal were calculated for the entire experimental period. The addition of fish broth and fish oil significantly improved the n3:n6 fatty acid ratio in diets. The presence of EPA and DHA in the experimental diet could have had a positive health effect on piglets, comparable with that exerted by therapeutic doses of zinc often administered to pigs of this age group. During the experiment, feed conversion ratio (FCR) gain was considerably reduced in the experimental group, with similar daily gains in the control and experimental group.

  3. A microfluidic device for antimicrobial susceptibility testing based on a broth dilution method.

    PubMed

    Lee, Wen-Bin; Fu, Chien-Yu; Chang, Wen-Hsin; You, Huey-Ling; Wang, Chih-Hung; Lee, Mel S; Lee, Gwo-Bin

    2017-01-15

    Bacterial resistance to antimicrobial compounds is increasing at a faster rate than the development of new antibiotics; this represents a critical challenge for clinicians worldwide. Normally, the minimum inhibitory concentration of an antibiotic, the dosage at which bacterial growth is thwarted, provides an effective quantitative measure for antimicrobial susceptibility testing, and determination of minimum inhibitory concentration is conventionally performed by either a serial broth dilution method or with the commercially available Etest(®) (Biomerieux, France) kit. However, these techniques are relatively labor-intensive and require a significant amount of training. In order to reduce human error and increase operation simplicity, a simple microfluidic device that can perform antimicrobial susceptibility testing automatically via a broth dilution method to accurately determine the minimum inhibitory concentration was developed herein. As a proof of concept, wild-type (ATCC 29212) and vancomycin-resistant Enterococcus cells were incubated at five different vancomycin concentrations on-chip, and the sample injection, transport, and mixing processes occurred within five reaction chambers and three reagent chambers via the chip's automatic dispensation and dilution functions within nine minutes. The minimum inhibitory concentration values measured after 24h of antibiotic incubation were similar to those calculated using Etest(®). With its high flexibility, reliability, and portability, the developed microfluidic device provides a simple method for antimicrobial susceptibility testing in an automated format that could be implemented for clinical and point-of-care applications.

  4. Biological Pretreatment of Chicken Feather and Biogas Production from Total Broth.

    PubMed

    Patinvoh, Regina J; Feuk-Lagerstedt, Elisabeth; Lundin, Magnus; Sárvári Horváth, Ilona; Taherzadeh, Mohammad J

    2016-12-01

    Chicken feathers are available in large quantities around the world causing environmental challenges. The feathers are composed of keratin that is a recalcitrant protein and is hard to degrade. In this work, chicken feathers were aerobically pretreated for 2-8 days at total solid concentrations of 5, 10, and 20 % by Bacillus sp. C4, a bacterium that produces both α- and β-keratinases. Then, the liquid fraction (feather hydrolysate) as well as the total broth (liquid and solid fraction of pretreated feathers) was used as substrates for biogas production using anaerobic sludge or bacteria granules as inoculum. The biological pretreatment of feather waste was productive; about 75 % of feather was converted to soluble crude protein after 8 days of degradation at initial feather concentration of 5 %. Bacteria granules performed better during anaerobic digestion of untreated feathers, resulting in approximately two times more methane yield (i.e., 199 mlCH4/gVS compared to 105 mlCH4/gVS when sludge was used). Pretreatment improved methane yield by 292 and 105 % when sludge and granules were used on the hydrolysate. Bacteria granules worked effectively on the total broth, yielded 445 mlCH4/gVS methane, which is 124 % more than that obtained with the same type of inoculum from untreated feather.

  5. The antileukaemic cell cycle regulatory activities of swainsonine purified from Metarhizium anisopliae fermentation broth.

    PubMed

    Singh, Digar; Kaur, Gurvinder

    2014-01-01

    Swainsonine is a Metarhizium secondary metabolite known differentially for its specific mannosidase inhibitory, toxic and therapeutic activities. Here, the standard and purified swainsonine from Metarhizium anisopliae fermentation broth were comparatively evaluated for their in situ antileukaemic activities in human promyelocytic cell line, HL-60. Both the standard (IC50 = 6.96 μM) and purified (IC50 = 9.50 μM) compounds inhibited the leukaemic cell proliferation without inflicting cell membrane disruption at 48 h of post-treatment incubation. The DNA cell cycle analysis showed approximately 48.81% and 60.72% of the treated cells arrested in the synthetic phase (S-phase) at 36 and 48 h, respectively, upon treatment with IC50 concentration of the purified swainsonine. However, only 29.62% of cells were arrested in S-phase with standard swainsonine at 48 h, suggesting the comprehensive action of certain other metabolites sharing the similar paradigm of antiproliferative properties in Metarhizium broth extract.

  6. Two-stage electrodialytic concentration of glyceric acid from fermentation broth.

    PubMed

    Habe, Hiroshi; Shimada, Yuko; Fukuoka, Tokuma; Kitamoto, Dai; Itagaki, Masayuki; Watanabe, Kunihiko; Yanagishita, Hiroshi; Sakaki, Keiji

    2010-12-01

    The aim of this research was the application of a two-stage electrodialysis (ED) method for glyceric acid (GA) recovery from fermentation broth. First, by desalting ED, glycerate solutions (counterpart is Na+) were concentrated using ion-exchange membranes, and the glycerate recovery and energy consumption became more efficient with increasing the initial glycerate concentration (30 to 130 g/l). Second, by water-splitting ED, the concentrated glycerate was electroconverted to GA using bipolar membranes. Using a culture broth of Acetobacter tropicalis containing 68.6 g/l of D-glycerate, a final D-GA concentration of 116 g/l was obtained following the two-stage ED process. The total energy consumption for the D-glycerate concentration and its electroconversion to D-GA was approximately 0.92 kWh per 1 kg of D-GA. Copyright © 2010 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

  7. Preparation of maltotriose from fermentation broth by hydrolysis of pullulan using pullulanase.

    PubMed

    Wu, Sheng-Jun; Chen, Jing

    2014-07-17

    In this study, we prepared maltotriose from the fermentation broth of Auerobasidium Pullulans CJ001 isolated from the sea mud by hydrolysis of pullulan with pullulanase. The fermentation broth was centrifuged to remove the microorganisms and then hydrolysed by pullulanase. The optimal hydrolysis conditions were obtained as follows: time, 9.40 h; pH, 4.92; temperature, 47.88°C; pullulanase, 10ASPU/g. Under these optimum hydrolysis conditions, the maximum dextrose equivalent value reached 31.86. The hydrolysates were filtrated through a filter membrane to separate any particle with molecular weight higher than 1,000 Da, concentrated to ∼20%, and precipitated with 8 volumes of absolute ethanol. The precipitate was dried at 80°C for 2h to yield the maltotriose product. The maltotriose content in the product and the yield of maltotriose were 92.13% and 90.23%, respectively. The results indicate that this was a promising way of maltotriose production. Copyright © 2014 Elsevier Ltd. All rights reserved.

  8. Influence of Different Media, Incubation Times, and Temperatures for Determining the MICs of Seven Antifungal Agents against Paracoccidioides brasiliensis by Microdilution

    PubMed Central

    Cruz, R. C.; Werneck, S. M. C.; Oliveira, C. S.; Santos, P. C.; Soares, B. M.; Santos, D. A.

    2013-01-01

    MIC assays with Paracoccidioides brasiliensis, the etiological agent of paracoccidioidomycosis, had been conducted with variable protocols, employing both macrodilution and microdilution tests and including differences in inoculum preparation, media used, incubation periods, and temperatures. Twenty-one clinical and environmental isolates of Paracoccidioides were tested using amphotericin B, itraconazole, ketoconazole, fluconazole, sulfamethoxazole, sulfamethoxazole-trimethoprim, and terbinafine, according to the National Committee for Clinical Laboratory Standards (National Committee for Clinical Laboratory Standards, document M27-A2, 2002), with modifications such as three medium formulations (RPMI 1640 medium, McVeigh and Morton [MVM] medium, and modified Mueller-Hinton [MMH] medium), two incubation temperatures (room temperature [25 to 28°C] and 37°C), and three incubation periods (7, 10, and 15 days). The antifungal activities were also classified as fungicidal or fungistatic. The best results were obtained after 15 days of incubation, which was chosen as the standard incubation time. The MICs for most individual isolates grown for the same length of time at the same temperature varied with the different media used (P < 0.05). Of the isolates, 81% showed transition from the yeast to the mycelial form in RPMI 1640 medium at 37°C, independent of the presence of antifungals. MMH medium appears to be a suitable medium for susceptibility testing of antifungal drugs with P. brasiliensis, except for sulfamethoxazole and the combination of sulfamethoxazole-trimethoprim, for which the MVM medium yielded better results. The incubation temperature influenced the MICs, with, in general, higher MICs at 25°C (mycelial form) than at 37°C (P < 0.05). Based on our results, we tentatively propose a microdilution assay protocol for susceptibility testing of antifungal drugs against Paracoccidioides. PMID:23175254

  9. Producing cell-free culture broth of rhamnolipids as a cost-effective fungicide against plant pathogens.

    PubMed

    Sha, Ruyi; Jiang, Lifang; Meng, Qin; Zhang, Guoliang; Song, Zhirong

    2012-08-01

    Biosurfactants of rhamnolipids have been enthusiastically investigated for substitutes of synthetic agrochemicals against plant pathogens. However, all such studies have been conducted on rhamnolipids with high purity which have limitations due to high costs. This paper focused on the applicability of rhamnolipid-containing cell-free culture broth. It was found that rhamnolipids in cell-free culture broth of Pseudomonas aeruginosa ZJU211 were largely composed of di-rhamnolipid and mono-rhamnolipid with the ratio varying with culture time. After 96 h of fermentation, the mass ratio of di-rhamnolipid over mono-rhamnolipid increased to 2.6:1. Crude rhamnolipids in a form of cell-free culture broth showed high antifungal activity against colony growth and biomass accumulation of seven plant pathogens comprising two Oomycetes, three Ascomycota and two Mucor spp. fungi, among which three plant pathogens were firstly reported in this paper showing inhibition to rhamnolipids. Particularly, rhamnolipids showed potent activity against two Oomycetes that acquire resistance to commercial compound of metalaxyal. Furthermore, di-rhamnolipid was elucidated to dominate the antifungal activity of crude rhamnolipids by in vitro studies. At last, the efficacy and safety of cell-free culture broth was preliminarily illustrated on plants in vivo. So cell-free culture broth as a crude rhamnolipid product could be served as a potential cost-effective and environmental-friendly fungicide in agriculture.

  10. Reliability of the E-Test Method for Detection of Colistin Resistance in Clinical Isolates of Acinetobacter baumannii

    PubMed Central

    Arroyo, L. A.; García-Curiel, A.; Pachón-Ibañez, M. E.; Llanos, A. C.; Ruiz, M.; Pachón, J.; Aznar, J.

    2005-01-01

    We compared the E-test to the broth microdilution method for testing the susceptibility of 115 clinical isolates of Acinetobacter baumannii to colistin. Twenty-two (19.1%) strains were resistant to colistin and 93 (80.8%) strains were susceptible according to the reference broth microdilution method. A categorical agreement of 98.2% was found, with only two (1.7%) very major errors. Agreement within 1 twofold dilution between the E-test and the broth microdilution was 16.5%. Complete agreement was found for the strains for which MICs fell within the range of 0.25 to 1 μg of colistin/ml. However, there was poor concordance, particularly in extreme dilutions with higher MICs by the E-test method. PMID:15695701

  11. Reliability of the E-test method for detection of colistin resistance in clinical isolates of Acinetobacter baumannii.

    PubMed

    Arroyo, L A; García-Curiel, A; Pachón-Ibañez, M E; Llanos, A C; Ruiz, M; Pachón, J; Aznar, J

    2005-02-01

    We compared the E-test to the broth microdilution method for testing the susceptibility of 115 clinical isolates of Acinetobacter baumannii to colistin. Twenty-two (19.1%) strains were resistant to colistin and 93 (80.8%) strains were susceptible according to the reference broth microdilution method. A categorical agreement of 98.2% was found, with only two (1.7%) very major errors. Agreement within 1 twofold dilution between the E-test and the broth microdilution was 16.5%. Complete agreement was found for the strains for which MICs fell within the range of 0.25 to 1 microg of colistin/ml. However, there was poor concordance, particularly in extreme dilutions with higher MICs by the E-test method.

  12. Synergistic bactericidal effect of carvacrol, cinnamaldehyde or thymol and refrigeration to inhibit Bacillus cereus in carrot broth.

    PubMed

    Valero, M; Francés, E

    2006-02-01

    Possible use of three different essential oil components as natural food preservatives was studied by examining their influence in the kinetics of growth from activated spores of four Bacillus cereus strains in tyndallized carrot broth over the temperature range 5-16 degrees C. Selected low concentrations of carvacrol, cinnamaldehyde, or thymol showed a clear antibacterial activity against B. cereus in the vegetable substrate. The addition of 2 microl cinnamaldehyde or 20 mg thymol to 100 ml of broth in combination with refrigeration temperatures (broth, however, were unable to inhibit bacterial growth at 8 degrees C.

  13. Antilisterial activity of hops beta acids in broth with or without other antimicrobials.

    PubMed

    Shen, C; Sofos, J N

    2008-11-01

    Hops beta acids (HBA) are parts of hops flowers used to preserve wort and provide flavor in beer, and are reported as having antimicrobial properties. This study evaluated the antilisterial activity of HBA alone or in combination with other known antimicrobials in a culture broth medium. Listeria monocytogenes (10-strain mixture) was inoculated (2.6 to 2.8 log CFU/mL) into tryptic soy broth supplemented with 0.6% yeast extract (TSBYE) without (control) or with HBA (0.5 to 5.0 microg/mL), potassium lactate (1.0%), sodium diacetate (0.25%), or acetic acid (0.1%), alone or in combination with HBA (0.5 to 3.0 microg/mL). Survival/growth of the pathogen during storage at 4 degrees C (35 d), 10 degrees C (20 d), or 25 degrees C (2 d) was periodically monitored by spiral plating onto tryptic soy agar plus 0.6% yeast extract. As expected, TSBYE without antimicrobials (control) supported rapid pathogen growth with growth rates of 0.40, 2.88, and 9.58 log CFU/mL/d at 4, 10, and 25 degrees C, respectively; corresponding Y(end) values exceeded 9.0 log CFU/mL at 35, 20, and 2 d storage. HBA used alone (1.0 to 5.0 microg/mL) inhibited growth of L. monocytogenes at all 3 temperatures, with inhibition being more pronounced at higher concentrations and at the lower storage temperature (4 degrees C). The antilisterial activity of HBA (0.5 to 3.0 microg/mL) was enhanced when combined with sodium diacetate, acetic acid, or potassium lactate, achieving complete inhibition at 4 degrees C when 3.0 microg/mL HBA were used in combination with each of the above antimicrobials. Overall, HBA exhibited promising antilisterial activity in a broth medium and further studies are needed to investigate its potential antilisterial effects in food products.

  14. Lytic enzyme production optimization using low-cost substrates and its application in the clarification of xanthan gum culture broth.

    PubMed

    da Silva, Cíntia Reis; Silva, Marilia Lordelo Cardoso; Kamida, Helio Mitoshi; Goes-Neto, Aristoteles; Koblitz, Maria Gabriela Bello

    2014-07-01

    Lytic enzymes are widely used in industrial biotechnology as they are able to hydrolyze the bacterial cell wall. One application of these enzymes is the clarification of the culture broth for the production of xanthan gum, because of its viability in viscous media and high specificity. The screening process for filamentous fungi producing lytic enzymes, the optimization of production of these enzymes by the selected microorganism, and the optimization of the application of the enzymes produced in the clarification of culture broth are presented in this article. Eleven fungal isolates were tested for their ability to produce enzymes able to increase the transmittance of the culture broth containing cells of Xanthomonas campestris. To optimize the secretion of lytic enzymes by the selected microorganism the following variables were tested: solid substrate, initial pH, incubation temperature, and addition of inducer (gelatin). Thereafter, secretion of the enzymes over time of incubation was assessed. To optimize the clarification process a central composite rotational design was applied in which the pH of the reaction medium, the dilution of the broth, and the reaction temperature were evaluated. The isolate identified as Aspergillus tamarii was selected for increasing the transmittance of the broth from 2.1% to 54.8%. The best conditions for cultivation of this microorganism were: use of coconut husk as solid substrate, with 90% moisture, at 30°C for 20 days. The lytic enzymes produced thereby were able to increase the transmittance of the culture broth from 2.1% to 70.6% at 65°C, without dilution and without pH adjustment.

  15. Lytic enzyme production optimization using low-cost substrates and its application in the clarification of xanthan gum culture broth

    PubMed Central

    da Silva, Cíntia Reis; Silva, Marilia Lordelo Cardoso; Kamida, Helio Mitoshi; Goes-Neto, Aristoteles; Koblitz, Maria Gabriela Bello

    2014-01-01

    Lytic enzymes are widely used in industrial biotechnology as they are able to hydrolyze the bacterial cell wall. One application of these enzymes is the clarification of the culture broth for the production of xanthan gum, because of its viability in viscous media and high specificity. The screening process for filamentous fungi producing lytic enzymes, the optimization of production of these enzymes by the selected microorganism, and the optimization of the application of the enzymes produced in the clarification of culture broth are presented in this article. Eleven fungal isolates were tested for their ability to produce enzymes able to increase the transmittance of the culture broth containing cells of Xanthomonas campestris. To optimize the secretion of lytic enzymes by the selected microorganism the following variables were tested: solid substrate, initial pH, incubation temperature, and addition of inducer (gelatin). Thereafter, secretion of the enzymes over time of incubation was assessed. To optimize the clarification process a central composite rotational design was applied in which the pH of the reaction medium, the dilution of the broth, and the reaction temperature were evaluated. The isolate identified as Aspergillus tamarii was selected for increasing the transmittance of the broth from 2.1% to 54.8%. The best conditions for cultivation of this microorganism were: use of coconut husk as solid substrate, with 90% moisture, at 30°C for 20 days. The lytic enzymes produced thereby were able to increase the transmittance of the culture broth from 2.1% to 70.6% at 65°C, without dilution and without pH adjustment. PMID:25473487

  16. [Antimycoplasmic Activity of Fermentation Broth of Trichoderma harzianum Rifai F-180, an Organism Producing L-Lysine-α-Oxidase, an Antitumor and Antiviral Enzyme].

    PubMed

    Smirnova, I P; Rakovskaya, I V

    2014-01-01

    A concentrate of the fermentation broth of Trichoderma harzianum Rifai F-180, an organism producing L-lysine-α-oxidase, an antitumor and antiviral enzyme, with the activity in the fermentation broth of 0.54-0.56 U/mI was recovered. The effect of the concentrate on the mycoplasmas growth was investigated for the first time. Two representatives of Mycoplasmafaceae, i.e. Mycoplasma hominis and Mycoplasma fermentans and one representative of Aholeplasmataceae. i. e. Aholeplasma laidlawii were used. It was shown that the fermentation broth inhibited the growth of Mycoplasma hominis after the preliminary exposure. The inhibition rate depended on the mycoplasma inoculation dose and the fermentation broth concentration.

  17. Determination of Ideal Broth Formulations Needed to Prepare Hydrous Hafnium Oxide Microspheres via the Internal Gelation Process

    SciTech Connect

    Collins, Jack Lee; Hunt, Rodney Dale; Simmerman, S. G.

    2009-02-01

    A simple test-tube methodology was used to determine optimum process parameters for preparing hydrous hafnium oxide microspheres by the internal gelation process. Broth formulations of hafnyl chloride [HfOCl{sub 2}], hexamethylenetetramine, and urea were found that can be used to prepare hydrous hafnium oxide gel spheres in the temperature range of 70-90 C. A few gel-forming runs were made in which microspheres were prepared with some of these formulations in order to equate the test-tube gelation times with actual gelation times. These preparations confirmed that the test-tube methodology is reliable for determining the ideal broths.

  18. Mueller-Hinton broth undergoes visible oxidative color changes in the presence of peroxidase and hydrogen peroxide.

    PubMed Central

    Galeazzi, L; Groppa, G; Giunta, S

    1990-01-01

    In the presence of peroxidase and hydrogen peroxide, Mueller-Hinton broth undergoes a slow but clearly detectable color change from pale yellow to dark yellow or brown. An investigation of this phenomenon led to the conclusion that it is the result of the oxidation of tyrosine, a major component of the broth. Indeed, tyrosine has long been known to oxidize upon treatment with peroxidase and hydrogen peroxide. The observations reported here, besides being curious for the clinical microbiologist, might deserve attention for the possible implications in the medium color darkening which sometimes happens during microbial growth. Images PMID:2172301

  19. Determination of Ideal Broth Formulations Needed to Prepare Hydrous Aluminum Oxide Microspheres via the Internal Gelation Process

    SciTech Connect

    Collins, Jack Lee; Pye, S. L.

    2009-02-01

    A simple test-tube methodology was used to determine optimum process parameters for preparing hydrous aluminum oxide microspheres by the internal gelation process. Broth formulations of aluminum, hexamethylenetetramine, and urea were found that can be used to prepare hydrous aluminum oxide gel spheres in the temperature range of 60-90 C. A few gel-forming runs were made in which microspheres were prepared with some of these formulations in order to equate the test-tube gelation times with actual gelation times. These preparations confirmed that the test-tube methodology is reliable for determining the ideal broths.

  20. Recovery and purification of rapamycin from the culture broth of Mtcc 5681.

    PubMed

    Rani, Polavarapu Baby; Kumar, Battula Suneel; Rao, A K S Bhujanga; Sreenivasrao, S; Narasu, Mangamoori Lakshmi

    2013-03-01

    In this study of the recovery and purification of rapamycin from the culture broth of an actinomycetes strain MTCC 5681, we investigated various factors such as biomass separation, suitable solvents for extraction, normal phase and flash chromatographic conditions and selective precipitation to obtain rapamycin in substantially pure form of the product. Adsorption chromatography particularly with normal phase and flash chromatography, in combination with centrifugal decantation is found to be the most suitable for separation as well as purification of rapamycin. Centrifugal decantation technique is likely to emerge as an efficient, industrially scalable, high yielding and economical process for biomass separation. The purity of rapamycin obtained through the method described was 99.4% which has not been reported so far.

  1. Energy-efficient recovery of butanol from model solutions and fermentation broth by adsorption.

    PubMed

    Qureshi, N; Hughes, S; Maddox, I S; Cotta, M A

    2005-07-01

    This article discusses the separation of butanol from aqueous solutions and/or fermentation broth by adsorption. Butanol fermentation is also known as acetone butanol ethanol (ABE) or solvent fermentation. Adsorbents such as silicalite, resins (XAD-2, XAD-4, XAD-7, XAD-8, XAD-16), bone charcoal, activated charcoal, bonopore, and polyvinylpyridine have been studied. Use of silicalite appears to be the more attractive as it can be used to concentrate butanol from dilute solutions (5 to 790-810 g L(-1)) and results in complete desorption of butanol (or ABE). In addition, silicalite can be regenerated by heat treatment. The energy requirement for butanol recovery by adsorption-desorption processes has been calculated to be 1,948 kcal kg(-1) butanol as compared to 5,789 kcal kg(-1) butanol by steam stripping distillation. Other techniques such as gas stripping and pervaporation require 5,220 and 3,295 kcal kg(-1) butanol, respectively.

  2. [Determination of tetrodotoxin in fermentation broth of distiller's yeast by ion chromatography].

    PubMed

    Shu, Jing; Li, Bailin; Ou, Jie

    2011-02-01

    A method was developed for the quantitative analysis of tetrodotoxin (TTX) in fermentation broth of distiller's yeast by ion chromatography. After extraction with acetonitrile solution (containing 0.1% phosphoric acid) and purification with an ion-exchange column, the tetrodotoxin was separated by ion chromatography and detected by a ultraviolet-visible (UV-VIS) absorbance detector. The experimental results showed that the tetrodotoxin had a good linearity (r2 = 0.997) in the range of 10 - 100 mg/L and the detection limit (3 of signal-to-noise ratio) was 1.0 mg/L. The average recoveries were between 90% - 103% with a relative standard deviation lower than 4.9%. The analysis of real samples verified the reliability of this method and demonstrated that the ion chromatography can be used for the quantification detection of the tetrodotoxin. The degradation experiment results suggested that distiller's yeast had a remarkable effect on the tetrodotoxin degradation.

  3. A novel process for recovery and refining of L-lactic acid from fermentation broth.

    PubMed

    Chen, Lijun; Zeng, Aiwu; Dong, Haibo; Li, Qi; Niu, Congcong

    2012-05-01

    This paper introduces a novel process for recovery and refining of L-lactic acid from a fermentation broth. The use of a solvent extraction step, in the novel approach, has significant impacts on the following centrifugal short-path distillation conditions (operating pressure, evaporator temperature and feed flow rate). As the conditions were varied, the l-lactic acid purity and yield in the distillate were monitored. For the purpose of comparison, a series of experiments were also carried out using the existing purification process. The results showed that both of the two processes can obtain l-lactic acid with a high purity around 91.3%, while the yield obtained using the novel process reached 61.73%, which was about 20.43% higher than that using the existing process. Additionally, multiple-pass distillation observed special attention by improving the yield up to 74.63%. Copyright © 2012 Elsevier Ltd. All rights reserved.

  4. Lethal paralytic shellfish poisoning from consumption of green mussel broth, Western Samar, Philippines, August 2013

    PubMed Central

    Ramos, Ruth Alma; de los Reyes, Vikki Carr; Sucaldito, Ma Nemia; Tayag, Enrique

    2015-01-01

    Background In July 2013, the Philippines’ Event-Based Surveillance & Response Unit received a paralytic shellfish poisoning (PSP) report from Tarangnan, Western Samar. A team from the Department of Health conducted an outbreak investigation to identify the implicated source and risk factors in coastal villages known for green mussel production and exportation. Methods A case was defined as a previously well individual from Tarangan, Western Samar who developed gastrointestinal symptoms and any motor and/or sensory symptoms after consumption of shellfish from 29 June to 4 July 2013 in the absence of any known cause. The team reviewed medical records, conducted active case finding and a case-control study. Relatives of cases who died were interviewed. Sera and urine specimens, green mussel and seawater samples were tested for saxitoxin levels using high performance liquid chromatography. Results Thirty-one cases and two deaths were identified. Consumption of > 1 cup of green mussel broth was associated with being a case. Seawater sample was positive for Pyrodinium bahamense var. compressum and green mussel samples were positive for saxitoxin. Inspection revealed villagers practice open defecation and improper garbage disposal. Conclusion This PSP outbreak was caused by the consumption of the green mussel broth contaminated by saxitoxin. As a result of this outbreak, dinoflagellate and saxitoxin surveillance was established, and since the outbreak, there have been no harmful algal blooms event or PSP case reported since. A “Save Cambatutay Bay” movement, focusing on proper waste disposal practice and clean-up drives has been mobilized. PMID:26306212

  5. Fermentation broth components influence droplet coalescence and hinder advanced biofuel recovery during fermentation.

    PubMed

    Heeres, Arjan S; Schroën, Karin; Heijnen, Joseph J; van der Wielen, Luuk A M; Cuellar, Maria C

    2015-08-01

    Developments in synthetic biology enabled the microbial production of long chain hydrocarbons, which can be used as advanced biofuels in aviation or transportation. Currently, these fuels are not economically competitive due to their production costs. The current process offers room for improvement: by utilizing lignocellulosic feedstock, increasing microbial yields, and using cheaper process technology. Gravity separation is an example of the latter, for which droplet growth by coalescence is crucial. The aim of this study was to study the effect of fermentation broth components on droplet coalescence. Droplet coalescence was measured using two setups: a microfluidic chip and regular laboratory scale stirred vessel (2 L). Some fermentation broth components had a large impact on droplet coalescence. Especially components present in hydrolysed cellulosic biomass and mannoproteins from the yeast cell wall retard coalescence. To achieve a technically feasible gravity separation that can be integrated with the fermentation, the negative effects of these components on coalescence should be minimized. This could be achieved by redesign of the fermentation medium or adjusting the fermentation conditions, aiming to minimize the release of surface active components by the microorganisms. This way, another step can be made towards economically feasible advanced biofuel production. © 2015 The Authors. Biotechnology Journal published by Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim. This is an open access article under the terms of the Creative Commons Attribution-Non-Commercial-NoDerivs Licence, which permits use and distribution in any medium, provided the original work is properly cited, the use is non-commercial and no modifications or adaptations are made.

  6. Wild-Type MIC Distributions and Epidemiological Cutoff Values for Posaconazole and Voriconazole and Candida spp. as Determined by 24-Hour CLSI Broth Microdilution▿

    PubMed Central

    Pfaller, M. A.; Boyken, L.; Hollis, R. J.; Kroeger, J.; Messer, S. A.; Tendolkar, S.; Diekema, D. J.

    2011-01-01

    We tested 16,191 strains of Candida against posaconazole and voriconazole, using the CLSI M27-A3 broth microdilution (BMD) method (24-h incubation), in order to define wild-type (WT) populations and epidemiological cutoff values (ECVs). From 2001 to 2009, 8,619 isolates of Candida albicans, 2,415 isolates of C. glabrata, 2,278 isolates of C. parapsilosis, 1,895 isolates of C. tropicalis, 508 isolates of C. krusei, 205 isolates of C. lusitaniae, 177 isolates of C. guilliermondii, and 93 isolates of C. kefyr were obtained from over 100 centers worldwide. The modal MICs (μg/ml) for posaconazole and voriconazole, respectively, were as follows: for C. albicans, 0.016 and 0.007; for C. glabrata, 0.5 and 0.06; for C. parapsilosis, 0.06 and 0.007; for C. tropicalis, 0.03 and 0.015; for C. krusei, 0.25 and 0.12; for C. lusitaniae, 0.03 and 0.007; for C. guilliermondii, 0.12 and 0.03; and for C. kefyr, 0.06 and 0.007. The ECVs (μg/ml [% of isolates that had MICs equal to or less than the ECV]) for posaconazole and voriconazole, respectively, were as follows: 0.06 (98.5) and 0.03 (98.9) for C. albicans, 2 (96.2) and 0.5 (90.4%) for C. glabrata, 0.25 (99.3) and 0.12 (97.9) for C. parapsilosis, 0.12 (97.6) and 0.06 (97.2) for C. tropicalis, 0.5 (99.8) and 0.5 (99.4) for C. krusei, 0.12 (95.6) and 0.03 (96.6) for C. lusitaniae, 0.5 (98.9) and 0.25 (98.3) for C. guilliermondii, and 0.25 (100.0) and 0.015 (100.0) for C. kefyr. In the absence of clinical breakpoints (CBPs) for posaconazole, these WT distributions and ECVs will be useful in surveillance for emergence of reduced susceptibility to posaconazole among Candida spp. Whereas a CBP for susceptibility of ≤1 μg/ml has been established for voriconazole and all species of Candida, it is notable that ECVs for this agent range from 10- to >100-fold lower than the CBP, depending on the species of Candida. The CBP is inadequate in detecting the emergence of voriconazole resistance among most Candida species encountered

  7. Removal of heavy metals from polluted soil using the citric acid fermentation broth: a promising washing agent.

    PubMed

    Zhang, Hongjiao; Gao, Yuntao; Xiong, Huabin

    2017-04-01

    The citric acid fermentation broth was prepared and it was employed to washing remediation of heavy metal-polluted soil. A well-defined washing effect was obtained, the removal percentages using citric acid fermentation broth are that 48.2% for Pb, 30.6% for Cu, 43.7% for Cr, and 58.4% for Cd and higher than that using citric acid solution. The kinetics of heavy metals desorption can be described by the double constant equation and Elovich equation and is a heterogeneous diffusion process. The speciation analysis shows that the citric acid fermentation broth can effectively reduce bioavailability and environmental risk of heavy metals. Spectroscopy characteristics analysis suggests that the washing method has only a small effect on the mineral composition and does not destroy the framework of soil system. Therefore, the citric acid fermentation broth is a promising washing agent and possesses a potential practical application value in the field of remediation of soils with a good washing performance.

  8. Comparison of broth enhancement to direct plating for screening of rectal cultures for ciprofloxacin-resistant Escherichia coli.

    PubMed

    Liss, Michael A; Nakamura, Kristen K; Peterson, Ellena M

    2013-01-01

    A transrectal prostate biopsy is the most common procedure used to establish the diagnosis of prostate cancer. Prior to biopsy, patients are commonly given ciprofloxacin for prophylaxis. However, a complication of the procedure is infection with ciprofloxacin-resistant organisms, in particular resistant Escherichia coli. In order to identify patients carrying ciprofloxacin-resistant E. coli, so as to tailor their antibiotic prophylaxis, rectal swabs are screened using selective broth and/or solid medium. In our evaluation, we compared broth enrichment and direct plating techniques by using brain heart infusion broth and MacConkey agar containing 1 μg/ml or 10 μg/ml of ciprofloxacin. Of the 100 patients included in the study, 20 were colonized with ciprofloxacin-resistant organisms, 19 of which were E. coli. There was no significant difference (P > 0.1) between the culture methods or the ciprofloxacin concentrations in the medium when identifying patients with ciprofloxacin-resistant E. coli; however, broth enrichment using 1 μg/ml ciprofloxacin was the most sensitive at 100%, but it was the least specific. Direct plating of rectal swabs onto MacConkey agar containing 10 μg/ml of ciprofloxacin was 100% specific and missed only 1 positive specimen, with a sensitivity of 94.7%; this method was the most cost-effective. Therefore, direct plating of rectal swabs onto selective medium proved to be a sensitive and cost-effective approach in identifying patients colonized with ciprofloxacin-resistant E. coli.

  9. Pre-treatment step with Leuconostoc mesenteroides or L. pseudomesenteroides strains removes furfural from Zymomonas mobilis ethanolic fermentation broth

    USDA-ARS?s Scientific Manuscript database

    Furfural (furan-2-carboxaldehyde), formed during dilute acid hydrolysis of biomass, is an inhibitor of growth and ethanol production by Zymomonas mobilis. The present study used a biological pre-treatment to reduce that amount of furfural in a model biofuel fermentation broth. The pre-treatment in...

  10. [Determination of E. coli with MUG (Fluorocult)-lauryl sulfate broth for the testing of microbial contamination in drugs].

    PubMed

    Huang, H; Oberkötter, E; Blume, H

    1994-06-01

    A test method for the determination of Escherichia coli in plant materials with the MUG (Fluorocult)-lauryl sulfate broth is described. It was found that more than 75% of the commonly used vegetable drugs exhibit fluorescence quench effects to different degrees when determining E. coli with the MUG-lauryl sulfate broth. Therefore a simple combination of two procedures was evaluated in order to avoid the matrix interferences: in a first step the drug sample was diluted eight times in a proportion of 1:10 with MUG-lauryl sulfate broth in eight separate tubes from 1 g down to 10(-7) g/tube (1st test series) and the resulting samples were incubated for 40 h at 36 degrees C. Subsequently, the tubes were tested for fluorescence. If the first tube of this series was found without fluorescence, in a second step a subsequent series of MUG-lauryl sulfate broth (2nd test series) was inoculated with 0.5 ml of the incubated culture from each of the first three tubes of series 1 and incubated again for 24 h at 36 degrees C. The results were evaluated from gas production, fluorescence as well as indole formation. Thus, the method allowed a simple and reproducible enumeration of E. coli for the test on microbial contamination in medicinal plant materials. The method was successfully applied to samples of 38 vegetable drugs for quantitative determination of E. coli (8 samples were found being contaminated with E. coli).

  11. [Simultaneous determination of organic acids and saccharides in lactic acid fermentation broth from biomass using high performance liquid chromatography].

    PubMed

    Ma, Rui; Ouyang, Jia; Li, Xin; Lian, Zhina; Cai, Cong

    2012-01-01

    Abstract: A high performance liquid chromatographic method for the simultaneous determination of organic acids and saccharides in lactic acid fermentation broth from biomass was developed. A Bio-Rad Aminex HPX-87H column was used at 55 degrees C. The mobile phase was 5 mmol/L sulfuric acid solution at a flow rate of 0.6 mL/min. The samples were detected by a refractive index detector (RID). The results showed that six organic acids and three saccharides in fermentation broth were completely separated and determined in 17 min. The linear correlation coefficients were above 0.999 8 in the range of 0.15-5.19 g/L. Under the optimized conditions, the recoveries of the organic acids and saccharides in Rhizopus oryzae fermentation broth at two spiked levels were in the range of 96.91%-103.11% with the relative standard deviations (RSDs, n = 6) of 0.81%-4.61%. This method is fast and accurate for the quantitative analysis of the organic acids and saccharides in microbial fermentation broths.

  12. Pre-treatment step with Leuconostoc mesenteroides or L. pseudomesenteroides strains removes furfural from Zymomonas mobilis ethanolic fermentation broth.

    PubMed

    Hunter, William J; Manter, Daniel K

    2014-10-01

    Furfural is an inhibitor of growth and ethanol production by Zymomonas mobilis. This study used a naturally occurring (not GMO) biological pre-treatment to reduce that amount of furfural in a model fermentation broth. Pre-treatment involved inoculating and incubating the fermentation broth with strains of Leuconostoc mesenteroides or Leuconostoc pseudomesenteroides. The Leuconostoc strains converted furfural to furfuryl alcohol without consuming large amounts of dextrose in the process. Coupling this pre-treatment to ethanolic fermentation reduced furfural in the broth and improved growth, dextrose uptake and ethanol formation. Pre-treatment permitted ethanol formation in the presence of 5.2 g L(-1) furfural, which was otherwise inhibitive. The pre-treatment and presence of the Leuconostoc strains in the fermentation broth did not interfere with Z. mobilis ethanolic fermentation or the amounts of ethanol produced. The method suggests a possible technique for reducing the effect that furfural has on the production of ethanol for use as a biofuel. Published by Elsevier Ltd.

  13. Dihydroberkleasmin A: a new eremophilane sesquiterpenoid from the fermentation broth of the plant endophytic fungus Pestalotiopsis photiniae.

    PubMed

    Yang, Xiao-Long; Zhang, Su; Zhu, Hua-Jie; Luo, Du-Qiang

    2011-02-23

    Dihydroberkleasmin A (1), a new ester-substituted sesquiterpenoid related to the eremophilane class, together with the known compound berkleasmin C (2), were isolated from the fermentation broth of the plant endophytic fungus Pestalotiopsis photiniae. The structure of dihydroberkleasmin A (1) was elucidated by extensive spectroscopic analysis. The stereochemistry was assigned by comparison of the NMR spectroscopic data with those of berkleasmin A.

  14. Supplementing chicken broth with monosodium glutamate reduces energy intake from high fat and sweet snacks in middle-aged healthy women.

    PubMed

    Imada, Toshifumi; Hao, Susan Shuzhen; Torii, Kunio; Kimura, Eiichiro

    2014-08-01

    Monosodium L-glutamate (MSG) and inosine monophosphate-5 (IMP) are flavor enhancers for umami taste. However, their effects on appetite and food intake are not well-researched. The objective of the current study was to test their additions in a broth preload on subsequent appetite ratings, energy intake and food choice. Eighty-six healthy middle-aged women with normal body weight received three preload conditions on 3 test days 1 week apart - a low-energy chicken flavor broth (200 ml) as the control preload, and broths with added MSG alone (0.5 g/100 ml, MSG broth) or in combination with IMP (0.05 g/100 ml) (MSG+ broth) served as the experimental conditions. Fifteen minutes after preload administration subjects were provided an ad libitum testing meal which consisted of 16 snacks varying in taste and fat content. MSG and MSG+ enhanced savory taste and broth properties of liking and pleasantness. In comparison with control, the MSG preload resulted in less consumption of total energy, as well as energy from sweet and high-fat snacks. Furthermore, MSG broth preload reduced added sugar intake. These findings were not observed after MSG+ preload. Appetite ratings were not different across the three preloads. Results suggest a potential role of MSG addition to a low-energy broth preload in subsequent energy intake and food choice. This trial was registered at clinicaltrials.gov as NCT01761045.

  15. Thermal inactivation and sublethal injury kinetics of Salmonella enterica and Listeria monocytogenes in broth versus agar surface.

    PubMed

    Wang, Xiang; Devlieghere, Frank; Geeraerd, Annemie; Uyttendaele, Mieke

    2017-02-21

    The objective of the present study was to compare the thermal inactivation and sublethal injury kinetics of Salmonella enterica and Listeria monocytogenes in broth (suspended cells) and on solid surface (agar-seeded cells). A 3-strain cocktail of S. enterica or L. monocytogenes inoculated in broth or on agar was subjected to heating in a water bath at various set temperatures (55.0, 57.5 and 60.0°C for S. enterica and 60.0, 62.5 and 65°C for L. monocytogenes). The occurrence of sublethally injured cells was determined by comparing enumerations on nonselective (TSAYE) and selective (XLD or ALOA) media. Results showed that the inactivation curves obtained from selective media were log-linear, and significant shoulders (p<0.05) were observed on some of the inactivation curves from TSAYE media. The D-values derived from the total population were higher than those from the uninjured cells. Generally, cells on agar surface exhibited higher heat resistance than those in broth. For S. enterica, cell injury increased with the exposure time, no difference was observed when treated at temperatures from 55.0 to 60.0°C, while for L. monocytogenes, cell injury increased significantly with heating time and treatment temperature (from 60.0 to 65°C). Moreover, the degree of sublethal injury affected by thermal treatment in broth or on agar surface depended upon the target microorganism. Higher proportions of injured S. enterica cells were observed for treatment in broth than on agar surface, while the opposite was found for L. monocytogenes. The provided information may be used to assess the efficacy of thermal treatment processes on surfaces for inactivation of S. enterica and L. monocytogenes, and it provides insight into the sublethally injured survival state of S. enterica and L. monocytogenes treated in liquid or on solid food.

  16. Hydrolysis of dilute acid pretreated mixed hardwood and purified microcrystalline cellulose by cell-free broth from Clostridium thermocellum

    SciTech Connect

    Lynd, L.R.; Grethlein, H.E.

    1987-01-01

    The cellulase activity in cell-free broths from Clostridium thermocellum is examined on both dilute-acid-pretreated mixed hardwood (90% maple, 10% birch) and Avicel. Experiments were conducted in vitro in order to distinguish properties of the cellulase from properties of the organism and to evaluate the effectiveness of C. thermocellum cellulase in the hydrolysis of a naturally occurring, lignin-containing substrate. The results obtained establish that essentially quantitative hydrolysis of cellulose from pretreated mixed hardwood is possible using this enzyme system. Pretreatment with 1% H/sub 2/SO/sub 4/ and a 9-s residence time at 220, 210, 200, and 180/sup 0/C allowed yields after enzymatic hydrolysis (percentage of glucan solubilized/glucan potentially solubilized) of 97.8, 86.1, 82.0, and 34.6%, respectively. Enzymatic hydrolysis of mixed hardwood with no pretreatment resulted in a yield of 10.1%. Hydrolysis yields of greater than 95% were obtained from 0.6 g/l mixed hardwood pretreated at 220/sup 0/C in 7 hours at broth strengths of 60 and 80% (v/v) and in approximately 48 hours with 33% broth. Hydrolysis of pretreated mixed hardwood is compared to hydrolysis of Avicel. The initial rate of Avicel hydrolysis saturates with respect to enzyme, whereas the initial rate of hydrolysis of pretreated wood is proportional to the amount of enzyme present. Initial hydrolysis rates for pretreated wood and Avicel at 0.6 g/l are greater for wood at low broth dilutions (1.25:1 to 5:1) by up to 2.7-fold and greater for Avicel at high broth dilutions (5:1 to 50:1) by up to 4.3-fold. Maximum rates of hydrolysis are achieved at less than 2 g substrate/liter for both pretreated wood and Avicel).

  17. Natural antibiotic susceptibility of Proteus spp., with special reference to P. mirabilis and P. penneri strains.

    PubMed

    Stock, I

    2003-02-01

    The natural susceptibility of 102 Proteus mirabilis and 35 Proteus penneri strains to 71 antibiotics was examined. Minimum inhibitory concentrations (MICs) were determined by applying a microdilution procedure in IsoSensitest broth (for all strains) and cation-adjusted Mueller Hinton broth (for some strains). P. mirabilis and P. penneri were naturally resistant to penicillin G, oxacillin, all tested macrolides, lincosamides, streptogramins, glycopeptides, rifampicin and fusidic acid. Both species were uniformly, naturally sensitive to all tested aminoglycosides, acylureidopenicillins, some cephalosporins, carbapenems, aztreonam, quinolones, sulfamethoxazole and co-trimoxazole. Species-specific differences in natural susceptibility affecting clinical assessment criteria were seen with tetracyclines, several beta-lactams, chloramphenicol and nitrufurantoin. P. mirabilis was naturally resistant to all tested tetracyclines, and was naturally sensitive to all beta-lactams, except penicillin G and oxacillin. Strains of P. penneri were naturally sensitive or of intermediate susceptibility to tetracyclines, and naturally resistant to amoxicillin (but sensitive or of intermediate susceptibility to aminopenicillins in the presence of beta-lactamase inhibitors) and some cephalosporins (i.e. cefaclor, cefazoline, loracarbef, cefuroxime, cefotiam, and cefdinir). P. penneri was less susceptible than P. mirabilis to chloramphenicol; P. mirabilis was less susceptible than P. penneri to nitrofurantoin. Major medium-dependent influences on the MICs were seen with fosfomycin. The present study describes a database concerning the natural antibiotic susceptibility of P. mirabilis and P. penneri strains to a range of antibiotics, which can be applied to validate forthcoming antibiotic susceptibility tests of these bacteria. It was shown that ten of fifteen amoxicillin-sensitive P. mirabilis strains produced beta-lactamases at a low level, supporting the thesis of the presence of a

  18. Effects of antimicrobial components of essential oils on growth of Bacillus cereus INRA L2104 in and the sensory qualities of carrot broth.

    PubMed

    Valero, M; Giner, M J

    2006-01-15

    The possible use of antimicrobials from seven plant essential oils as food preservatives was studied by examining their effects on the growth kinetics of activated Bacillus cereus INRA L2104 spores inoculated into tyndallized carrot broth. The effects of various concentrations of borneol, carvacrol, cinnamaldehyde, eugenol, menthol, thymol, and vanillin were determined. Five microliters of cinnamaldehyde, 15 microl of carvacrol, or 30 mg of thymol per 100 ml of inoculated carrot broth completely inhibited bacterial growth for more than 60 days at 16 degrees C. Lower concentrations of the three antimicrobials prolonged the lag phase and reduced both the exponential growth rate and the final population densities of cultures. The study of the sensory characteristics of the supplemented broths suggested that low concentration of cinnamaldehyde enhanced the taste of carrot broth, and that it did not have any adverse effect on the taste and smell of carrot broth at concentrations less than 6 microl 100 ml(-1).

  19. Comparison of the Vitek 2 yeast susceptibility system with CLSI microdilution for antifungal susceptibility testing of fluconazole and voriconazole against Candida spp., using new clinical breakpoints and epidemiological cutoff values.

    PubMed

    Pfaller, Michael A; Diekema, Daniel J; Procop, Gary W; Rinaldi, Michael G

    2013-09-01

    A commercially available, fully automated yeast susceptibility test system (Vitek 2; bioMérieux, Marcy d'Etoile, France) was compared in 3 different laboratories with the Clinical and Laboratory Standards Institute (CLSI) reference microdilution (BMD) method by testing 2 quality control strains, 10 reproducibility strains, and 425 isolates of Candida spp. against fluconazole and voriconazole. Reference CLSI BMD MIC endpoints and Vitek 2 MIC endpoints were read after 24 hours and 9.1-27.1 hours incubation, respectively. Excellent essential agreement (within 2 dilutions) between the reference and Vitek 2 MICs was observed for fluconazole (97.9%) and voriconazole (96.7%). Categorical agreement (CA) between the 2 methods was assessed using the new species-specific clinical breakpoints (CBPs): susceptible (S) ≤2 μg/mL, susceptible dose-dependent (SDD) 4 μg/mL, and resistant (R) ≥8 μg/mL for fluconazole and Candida albicans, Candida tropicalis, and Candida parapsilosis and ≤32 μg/mL (SDD), ≥64 μg/mL (R) for Candida glabrata; S ≤0.12 μg/mL, SDD 0.25-0.5 μg/mL, R ≥1 μg/mL for voriconazole and C. albicans, C. tropicalis, and C. parapsilosis, and ≤0.5 μg/mL (S), 1 μg/mL (SDD), ≥2 μg/mL (R) for Candida krusei. The epidemiological cutoff value (ECV) of 0.5 μg/mL for voriconazole and C. glabrata was used to differentiate wild-type (WT; MIC ≤ ECV) from non-WT (MIC > ECV) strains of this species. Due to the lack of CBPs for the less common species, the ECVs for fluconazole and voriconazole, respectively, were used for Candida lusitaniae (2 μg/mL and 0.03 μg/mL), Candida dubliniensis (0.5 μg/mL and 0.03 μg/mL), Candida guilliermondii (8 μg/mL and 0.25 μg/mL), and Candida pelliculosa (4 μg/mL and 0.25 μg/mL) to categorize isolates of these species as WT and non-WT. CA between the 2 methods was 96.8% for fluconazole and 96.5% for voriconazole with less than 1% very major errors and 1.3-3.0% major errors. The Vitek 2 yeast susceptibility system

  20. Modeling of the Competitive Growth of Listeria monocytogenes and Lactococcus lactis in Vegetable Broth

    PubMed Central

    Breidt, Frederick; Fleming, Henry P.

    1998-01-01

    Current mathematical models used by food microbiologists do not address the issue of competitive growth in mixed cultures of bacteria. We developed a mathematical model which consists of a system of nonlinear differential equations describing the growth of competing bacterial cell cultures. In this model, bacterial cell growth is limited by the accumulation of protonated lactic acid and decreasing pH. In our experimental system, pure and mixed cultures of Lactococcus lactis and Listeria monocytogenes were grown in a vegetable broth medium. Predictions of the model indicate that pH is the primary factor that limits the growth of L. monocytogenes in competition with a strain of L. lactis which does not produce the bacteriocin nisin. The model also predicts the values of parameters that affect the growth and death of the competing populations. Further development of this model will incorporate the effects of additional inhibitors, such as bacteriocins, and may aid in the selection of lactic acid bacterium cultures for use in competitive inhibition of pathogens in minimally processed foods. PMID:9726854

  1. Validation of a HILIC Method for the Analysis of Ergothioneine in Fermentation Broth.

    PubMed

    Liu, Qi; Zhang, Weiya; Wang, Hongyu; Li, Yunhua; Liu, Wei; Wang, Qunjie; Liu, Dongze; Chen, Ning; Jiang, Wenxia

    2016-07-01

    A hydrophilic interaction liquid chromatography method has been established for the quantification of ergothioneine (EGT) in fermentation broth. Chromatographic separation was conducted on a Venusil hydrophilic interaction liquid chromatography (HILIC) column (250 × 4.6 mm, 5 µm) at an elution rate of 1.0 mL/min with an isocratic mobile phase consisting of acetonitrile/20 mmol/L ammonium acetate solution (85 : 15, v/v) adjusted to pH 6.0 with acetic acid. Analytes were detected at 254 nm using a UV-VIS detector. The injection volume was 10 µL, and the column temperature was 40°C. The limits of detection and limits of quantification were 63 and 21 µg/L, respectively. Excellent linearity [correlation coefficient (R(2)) = 0.9999] was achieved for EGT quantification in the range of 5-400 mg/L. The relative standard deviations of repeatability, intermediate precision and stability were 1.47, 1.03 and 1.66%, respectively, and EGT recoveries were within 99.2-100.8%. The chromatographic peak corresponding to EGT in the HILIC spectrum was confirmed using ESI-MS. In general, the method developed here is simple, reliable, accurate, and stable and may be useful for routine analyses in EGT biosynthesis research. © The Author 2016. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

  2. Recovery of acids from anaerobic acidification broth by liquid-liquid extraction.

    PubMed

    Alkaya, Emrah; Kaptan, Serkan; Ozkan, Leyla; Uludag-Demirer, Sibel; Demirer, Göksel N

    2009-11-01

    In this study, anaerobic acidification of sugar beet processing wastes and subsequent liquid-liquid extraction of produced fermentation metabolites were investigated. The aim of extraction experiments was to asses the influence of pH and extractant (trioctylphosphine oxide (TOPO) in kerosene) concentrations on the recovery of volatile fatty acids (VFAs) from fermentation broth. The effect of TOPO in kerosene concentration was as crucial as the effect of pH on the recovery of VFAs via extraction. Consequently, pH 2.5 was determined as optimum. At this pH, percent recoveries of VFAs were changed from 43% to 98%, depending on the type of the acid extracted (acetic, butyric, propionic and valeric acids) and the concentration of TOPO in kerosene (5-20%). As the concentration of TOPO in kerosene was increased, efficiency of extraction was increased. As a result, highest VFA recoveries (61-98%) were observed at 20% TOPO in kerosene with distribution ratio values ranging between 1.54 and 40.79. At pH 2.5, the increase in TOPO concentration directly increased the chemical oxygen demand (COD) removal efficiencies, as it does for total VFA recovery. Up to 72% COD removals were achieved, at 20% TOPO in kerosene at pH 2.5, while the removal efficiencies remained between 19% and 22% at pH 5.5.

  3. Lactic acid recovery from cheese whey fermentation broth using combined ultrafiltration and nanofiltration membranes.

    PubMed

    Li, Yebo; Shahbazi, Abolghasem

    2006-01-01

    The separation of lactic acid from lactose in the ultrafiltration permeate of cheese whey broth was studied using a cross-flow nanofiltration membrane unit. Experiments to test lactic acid recovery were conducted at three levels of pressure (1.4, 2.1, and 2.8 MPa), two levels of initial lactic acid concentration (18.6 and 27 g/L), and two types of nanofiltration membranes (DS-5DK and DS-5HL). Higher pressure caused significantly higher permeate flux and higher lactose and lactic acid retention (p < 0.0001). Higher initial lactic acid concentrations also caused significantly higher permeate flux, but significantly lower lactose and lactic acid retention (p < 0.0001). The two tested membranes demonstrated significant differences on the permeate flux and lactose and lactic acid retention. Membrane DS-5DK was found to retain 100% of lactose at an initial lactic acid concentration of 18.6 g/L for all the tested pressures, and had a retention level of 99.5% of lactose at initial lactic acid concentration of 27 g/L when the pressure reached 2.8 MPa. For all the tests when lactose retention reached 99-100%, as much as 64% of the lactic acid could be recovered in the permeate.

  4. Performance of serum-free broth media for growth of Renibacterium salmoninarum

    USGS Publications Warehouse

    Starliper, C.E.; Schill, W.B.; Mathias, J.

    1998-01-01

    Growth of Renibacterium salmoninarum was compared in 14 different broth media; 13 serum-free, and 1 that contained newborn calf serum, KDM2+M. Supplementation with 1% v/v R. salmoninarum MCO4M metabolite was evaluated for 6 of the media that do not utilize it as part of their ingredients. Viable cells were enumerated on Days 10, 20, and 30 post inoculation to evaluate performance. The experiment was repeated 3 times using high, low, and medium (trials 1 to 3, respectively) cell concentrations as inoculum. In general there was no optimal medium and all performed well. The choice of which to employ depends on the ease of preparation and presence of certain ingredients that might affect subsequent assays. In trials 2 and 3, the pH was estimated using test papers at the same time as cells were counted. Maximum pH increase occurred with KDM2+M and those media containing charcoal. For most media, a simple pH determination could be used as a means to check that growth has occurred in a culture, particularly if charcoal was added directly to the media and a visual inspection could not be made to detect growth.

  5. 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

  6. Ergosterols from the Culture Broth of Marine Streptomyces anandii H41-59

    PubMed Central

    Zhang, Yang-Mei; Li, Hong-Yu; Hu, Chen; Sheng, Hui-Fan; Zhang, Ying; Lin, Bi-Run; Zhou, Guang-Xiong

    2016-01-01

    An actinomycete strain, H41-59, isolated from sea sediment in a mangrove district, was identified as Streptomyces anandii on the basis of 16S rDNA gene sequence analysis as well as the investigation of its morphological, physiological and biochemical characteristics. Three new ergosterols, ananstreps A–C (1–3), along with ten known ones (4–13), were isolated from the culture broth of this strain. The gross structures of these new compounds were elucidated on the basis of extensive analysis of spectroscopic data, including HR-ESI-MS, and NMR. The cytotoxicities of these isolates against human breast adenocarcinoma cell line MCF-7, human glioblastoma cell line SF-268, and human lung cancer cell line NCI-H460 and their antibacterial activities in inhibiting the growth of Candida albicans and some other pathogenic microorganisms were tested. Compounds 3–8, 10 and 11 displayed cytotoxicity with IC50 values in a range from 13.0 to 27.8 μg/mL. However, all the tested compounds showed no activity on C. albicans and other bacteria at the test concentration of 1 mg/mL with the paper disc diffusion method. PMID:27153073

  7. Compounds in a particular production lot of tryptic soy broth inhibit Staphylococcus aureus cell growth.

    PubMed

    Ishii, Masaki; Matsumoto, Yasuhiko; Sekimizu, Kazuhisa

    2015-06-01

    Staphylococcus aureus Newman strain and several methicillin-resistant S. aureus (MRSA) clinical isolates were grown on agar plates prepared with conventional lots of tryptic soy broth (TSB). Cell growth of these strains was inhibited on agar plates containing TSB of a particular product lot (lot A), whereas the cell growth of S. aureus RN4220 strain and several other MRSA clinical isolates was not inhibited. The cell growth of a strain of S. epidermidis was also inhibited on agar plates containing TSB of lot A, whereas the cell growth of Bacillus subtilis, Lactococcus lactis, Klebsiella pneumonia, Salmonella enterica, Serratia marcescens, Pseudomonas aeruginosa, and Escherichia coli was not inhibited. Although cell growth of the Newman strain was inhibited on agar plates containing TSB of lot A that was autoclaved in stainless steel or glass containers, cell growth inhibition was not observed when the medium was autoclaved in polypropylene containers. Compounds that inhibited the cell growth of the Newman strain were extracted from a polypropylene tube that was preincubated with liquid medium prepared from TSB of lot A. These findings suggest that polypropylene-binding compounds in TSB of lot A inhibited the cell growth of S. aureus Newman strain, some MRSA clinical isolates, and S. epidermidis.

  8. Profiling histidine dipeptides in plasma and urine after ingesting beef, chicken or chicken broth in humans.

    PubMed

    Yeum, Kyung-Jin; Orioli, Marica; Regazzoni, Luca; Carini, Marina; Rasmussen, Helen; Russell, Robert M; Aldini, Giancarlo

    2010-03-01

    The in vitro metabolic stability of histidine-dipeptides (HD), carnosine (CAR) and anserine (ANS), in human serum, and their absorption kinetics after ingesting pure carnosine or HD rich foods in humans have been investigated. Healthy women (n = 4) went through four phases of taking one dose of either 450 mg of pure carnosine, 150 g beef (B), 150 g chicken (C), or chicken broth (CB) from 150 g chicken with a >2-week washout period between each phase. Blood samples were collected at 0, 30, 60, 100, 180, 240, and 300 min, and urine samples before and after (up to 7 h) ingesting pure carnosine or food. Both plasma and urine samples were analyzed for HD concentrations using a sensitive and selective LC-ESI-MS/MS method. CAR was undetectable in plasma after ingesting pure carnosine, B, C or CB. By contrast, plasma ANS concentration was significantly increased (P < 0.05) after ingesting C or CB, respectively. Urinary concentrations of both CAR and ANS were 13- to 14-fold increased after ingesting B, and 14.8- and 243-fold after CB ingestion, respectively. Thus, dietary HD, which are rapidly hydrolyzed by carnosinase in plasma, and excreted in urine, may act as reactive carbonyl species sequestering agents.

  9. Commercial Lysogeny Broth culture media and oxidative stress: a cautious tale.

    PubMed

    Ezraty, Benjamin; Henry, Camille; Hérisse, Marion; Denamur, Erick; Barras, Frédéric

    2014-09-01

    Lysogeny Broth (LB), most often misnamed Luria-Bertani medium, ranks among the most commonly used growth media in microbiology. Surprisingly, we observed that oxidative levels vary with the commercial origin of the LB ready to use powder. Indeed, growth on solid media of Escherichia coli and Salmonella derivatives lacking antioxidative stress defenses, such as oxyR mutant devoid of the H2O2-sensing transcriptional activator or Hpx(-) strains lacking catalases and peroxidases, exhibit different phenotypes on LB-Sigma or LB-Difco. Using gene fusion and exogenously added catalase, we found that LB-Sigma contains higher levels of H2O2 than LB-Difco. Also we observed differences in population counts of 82 clinical and environmental isolates of E. coli, depending on the LB used. Further investigations revealed a significant influence of the commercial origin of agar as well. Besides being a warning to the wide population of LB users, our observations provide researchers in the oxidative stress field with a tool to appreciate the severity of mutations in antioxidative stress defenses. Copyright © 2014 Elsevier Inc. All rights reserved.

  10. Biodegradation of nitrobenzene in a lysogeny broth medium by a novel halophilic bacterium Bacillus licheniformis.

    PubMed

    Li, Tian; Deng, Xinping; Wang, Jinjun; Chen, Yucheng; He, Lin; Sun, Yuchuan; Song, Caixia; Zhou, Zhifeng

    2014-12-15

    The Bacillus licheniformis strain YX2, a novel nitrobenzene-degrading halophilic bacterium, was isolated from active sludge obtained from a pesticide factory. Strain YX2 can withstand highly acidic and alkaline conditions and high temperatures. Degradation of nitrobenzene (200mgL(-1)) by YX2 exceeded 70% after 72h in lysogeny broth medium (pH 4-9). Under optimal degradation conditions (33°C, pH 7 in LB medium) YX2 degraded 50, 100, 200, and 600mgL(-1) nitrobenzene within 36, 36, 72, and 156h, respectively. Even in the presence of benzene, phenol or aniline, strain YX2 efficiently degraded nitrobenzene. Furthermore, strain YX2 completely degraded 600mgL(-1) nitrobenzene in 7% NaCl (w/w). Thus, our data show that strain YX2 may have promise for removing nitrobenzene from complex wastewaters with high salinity and variable pH. Copyright © 2014 Elsevier Ltd. All rights reserved.

  11. The broth in my brother's brothel: morpho-orthographic segmentation in visual word recognition.

    PubMed

    Rastle, Kathleen; Davis, Matthew H; New, Boris

    2004-12-01

    Much research suggests that words comprising more than one morpheme are represented in a "decomposed" manner in the visual word recognition system. In the research presented here, we investigate what information is used to segment a word into its morphemic constituents and, in particular, whether semantic information plays a role in that segmentation. Participants made visual lexical decisions to stem targets preceded by masked primes sharing (1) a semantically transparent morphological relationship with the target (e.g., cleaner-CLEAN), (2) an apparent morphological relationship but no semantic relationship with the target (e.g., corner-CORN), and (3) a nonmorphological form relationship with the target (e.g., brothel-BROTH). Results showed significant and equivalent masked priming effects in cases in which primes and targets appeared to be morphologically related, and priming in these conditions could be distinguished from nonmorphological form priming. We argue that these findings suggest a level of representation at which apparently complex words are decomposed on the basis of their morpho-orthographic properties. Implications of these findings for computational models of reading are discussed.

  12. Purification and characterization of an endoxylanase from the culture broth of Bacillus cereus BSA1.

    PubMed

    Mandal, A; Kar, S; Das Mohapatra, P K; Maity, C; Pati, B R; Mondal, K C

    2011-01-01

    An extracellular xylanase from the fermented broth of Bacillus cereus BSA1 was purified and characterized. The enzyme was purified to 3.43 fold through ammonium sulphate precipitation, DEAE-cellulose chromatography and followed by gel filtration through Sephadex G-100 column. The molecular mass of the purified xylanse was about 33 kDa. The enzyme was an endoxylanase as it initially degraded xylan to xylooligomers. The purified enzyme showed optimum activity at 55 degrees C and at pH 7.0 and remained reasonably stable in a wide range ofpH (5.0-8.0) and temperature (40-65 degrees C). The Km and Vmax values were found to be 8.2 mg/ml and 181.8 micromol/(min mg), respectively. The enzyme had no apparent requirement ofcofactors, and its activity was strongly inhibited by Cu++, Hg++. It was also a salt tolerant enzyme and stable upto 2.5 M of NaCl and retained its 85% activity at 3.0 M. For stability and substrate binding, the enzyme needed hydrophobic interaction that revealed when most surfactants inhihited xylanase activity. Since the enzyme was active over wide range ofpH, temperature and remained active in higher salt concentration, it could find potential uses in biobleaching process in paper industries.

  13. An improved HPLC-DAD method for clavulanic acid quantification in fermentation broths of Streptomyces clavuligerus.

    PubMed

    Ramirez-Malule, Howard; Junne, Stefan; López, Carlos; Zapata, Julian; Sáez, Alex; Neubauer, Peter; Rios-Estepa, Rigoberto

    2016-02-20

    Clavulanic acid (CA) is an important secondary metabolite commercially produced by cultivation of Streptomyces clavuligerus (Sc). It is a potent inhibitor of bacterial β-lactamases. In this work, a specific and improved high performance liquid chromatography (HPLC) method, using a C-18 reversed phase column, diode array detector and gradient elution for CA quantification in fermentation broths of Sc, was developed and successfully validated. Samples were imidazole-derivatized for the purpose of creating a stable chromophore (clavulanate-imidazole). The calibration curve was linear over a typical range of CA concentration between 0.2 and 400mg/L. The detection and quantification limits were 0.01 and 0.02mg/L, respectively. The precision of the method was evaluated for CA spiked into production media and a recovery of 103.8%, on average, was obtained. The clavulanate-imidazole complex was not stable when the samples were not cooled during the analysis. The recovery rate was 39.3% on average. This assay was successfully tested for CA quantification in samples from Sc fermentation, using both, a chemically defined and a complex medium.

  14. Oil and air dispersion in a simulated fermentation broth as a function of mycelial morphology.

    PubMed

    Lucatero, Savidra; Larralde-Corona, Claudia Patricia; Corkidi, Gabriel; Galindo, Enrique

    2003-01-01

    The culture conditions of a multiphase fermentation involving morphologically complex mycelia were simulated in order to investigate the influence of mycelial morphology (Trichoderma harzianum) on castor oil and air dispersion. Measurements of oil drops and air bubbles were obtained using an image analysis system coupled to a mixing tank. Complex interactions of the phases involved could be clearly observed. The Sauter diameter and the size distributions of drops and bubbles were affected by the morphological type of biomass (pellets or dispersed mycelia) added to the system. Larger oil drop sizes were obtained with dispersed mycelia than with pellets, as a result of the high apparent viscosity of the broth, which caused a drop in the power drawn, reducing oil drop break-up. Unexpectedly, bubble sizes observed with dispersed mycelia were smaller than with pellets, a phenomenon which can be explained by the segregation occurring at high biomass concentrations with the dispersed mycelia. Very complex oil drops were produced, containing air bubbles and a high number of structures likely consisting of small water droplets. Bubble location was influenced by biomass morphology. The percentage (in volume) of oil-trapped bubbles increased (from 32 to 80%) as dispersed mycelia concentration increased. A practically constant (32%) percentage of oil-trapped bubbles was observed with pelleted morphology at all biomass concentrations. The results evidenced the high complexity of phases interactions and the importance of mycelial morphology in such processes.

  15. Radiometric method for testing susceptibility of mycobacteria to pyrazinamide in 7H12 broth.

    PubMed Central

    Heifets, L B; Iseman, M D

    1985-01-01

    The test of susceptibility to pyrazinamide requires an acid environment (pH less than or equal to 5.5). This, however, is not favorable to the growth of Mycobacterium tuberculosis, especially in solid agar media. To obviate this difficulty, we developed a testing method with 7H12 broth medium and based on radiometric readings of the growth. The radiometric method employed in this study (BACTEC system) provides an opportunity to detect the dynamics of growth by daily recording of the growth index, which reflects the metabolic activity of the multiplying bacteria. In our technique, M. tuberculosis isolates were initially cultivated at pH 6.8. After logarithmic growth had begun, phosphoric acid solution was added to obtain pH 5.5. When pyrazinamide was added simultaneously with the acid, the growth index of susceptible cultures decreased, whereas it continued to increase in pH 5.5 control vials and in tests with pyrazinamide-resistant strains. PMID:3972987

  16. Analysis of mixtures of fatty acids and fatty alcohols in fermentation broth.

    PubMed

    Liu, Yilan; Chen, Ting; Yang, Maohua; Wang, Caixia; Huo, Weiyan; Yan, Daojiang; Chen, Jinjin; Zhou, Jiemin; Xing, Jianmin

    2014-01-03

    Microbial production of fatty acids and fatty alcohols has attracted increasing concerns because of energy crisis and environmental impact of fossil fuels. Therefore, simple and efficient methods for the extraction and quantification of these compounds become necessary. In this study, a high-performance liquid chromatography-refractive index detection (HPLC-RID) method was developed for the simultaneous quantification of fatty acids and fatty alcohols in these samples. The optimum chromatographic conditions are C18 column eluted with methanol:water:acetic acid (90:9.9:0.1, v/v/v); column temperature, 26°C; flow rate, 1.0mL/min. Calibration curves of all selected analytes showed good linearity (r(2)≥0.9989). The intra-day and inter-day relative standard deviations (RSDs) of the 10 compounds were less than 4.46% and 5.38%, respectively, which indicated that the method had good repeatability and precision. Besides, a method for simultaneous extraction of fatty acids and fatty alcohols from fermentation broth was optimized by orthogonal design. The optimal extraction conditions were as follows: solvent, ethyl acetate; solvent to sample ratio, 0.5:1; rotation speed, 2min at 260rpm; extraction temperature, 10°C. This study provides simple and fast methods to simultaneously extract and quantify fatty acids and fatty alcohols for the first time. It will be useful for the study of microbial production of these products. Copyright © 2013 Elsevier B.V. All rights reserved.

  17. Ion-exclusion chromatography determination of organic acid in uridine 5'-monophosphate fermentation broth.

    PubMed

    Niu, Huanqing; Chen, Yong; Xie, Jingjing; Chen, Xiaochun; Bai, Jianxin; Wu, Jinglan; Liu, Dong; Ying, Hanjie

    2012-09-01

    Simultaneous determination of organic acids using ion-exclusion liquid chromatography and ultraviolet detection is described. The chromatographic conditions are optimized when an Aminex HPX-87H column (300 × 7.8 mm) is employed, with a solution of 3 mmol/L sulfuric acid as eluent, a flow rate of 0.4 mL/min and a column temperature of 60°C. Eight organic acids (including orotic acid, α-ketoglutaric acid, citric acid, pyruvic acid, malic acid, succinic acid, lactic acid and acetic acid) and one nucleotide are successfully quantified. The calibration curves for these analytes are linear, with correlation coefficients exceeding 0.999. The average recovery of organic acids is in the range of 97.6% ∼ 103.1%, and the relative standard deviation is in the range of 0.037% ∼ 0.38%. The method is subsequently applied to obtain organic acid profiles of uridine 5'-monophosphate culture broth fermented from orotic acid by Saccharomyces cerevisiae. These data demonstrate the quantitative accuracy for nucleotide fermentation mixtures, and suggest that the method may also be applicable to other biological samples.

  18. Bactericidal activity of soymilk fermentation broth by in vitro and animal models.

    PubMed

    Chin, Yi-Ping; Tsui, Ko-Chung; Chen, Mei-Chieh; Wang, Cheng-Yi; Yang, Chin-Yuh; Lin, Yuh-Ling

    2012-06-01

    Soybean fermentation broth (SFB) exhibits potent antibacterial activity against different species of bacteria in in vitro assays and animal models. Four isoflavone compounds-daidzin, genistin, genistein, and daidzein-of SFB were analyzed and quantified by high-performance liquid chromatography. In the in vitro test, daidzin and daidzein had more potent antibacterial activity than genistin. The minimum inhibition concentration values for these bacteria of SFB ranged from 1.25% to 5%, and the minimum bactericidal concentration values of strains ranged from 2.5% to 10%, depending on the species or strain. Vancomycin-resistant Entercoccus faecalis (VRE) strains were also tested for susceptibility to SFB in two species of animal model: the Sprague-Dawley rat and the BALB/c mouse. SFB-fed Sprague-Dawley rats showed excellent elimination efficiency against VRE, close to 99% compared with the phosphate-buffered saline-fed control group. In the BALB/c mouse model, SFB antibacterial activity was 65-80% against VRE compared with the control. In conclusion, SFB contains natural antibacterial substances such as daidzin, genistin, and daidzein that inhibit bacterial growth.

  19. Purification of chondroitin precursor from Escherichia coli K4 fermentation broth using membrane processing.

    PubMed

    Schiraldi, Chiara; Carcarino, Immacolata Loredana; Alfano, Alberto; Restaino, Odile Francesca; Panariello, Andrea; De Rosa, Mario

    2011-04-01

    Recently the possibility of producing the capsular polysaccharide K4, a fructosylated chondroitin, in fed-batch experiments was assessed. In the present study, a novel downstream process to obtain chondroitin from Escherichia coli K4 fermentation broth was developed. The process is simple, scalable and economical. In particular, downstream procedures were optimized with a particular aim of purifying a product suitable for further chemical modifications, in an attempt to develop a biotechnological platform for chondroitin sulfate production. During process development, membrane devices (ultrafiltration/diafiltration) were exploited, selecting the right cassette cut-offs for different phases of purification. The operational conditions (cross-flow rate and transmembrane pressure) used for the process were determined on an ÄKTA cross-flow instrument (GE Healthcare, USA), a lab-scale automatic tangential flow filtration system. In addition, parameters such as selectivity and throughput were calculated based on the analytical quantification of K4 and defructosylated K4, as well as the major contaminants. The complete downstream procedure yielded about 75% chondroitin with a purity higher than 90%.

  20. Isolation of α-arbutin from Xanthomonas CGMCC 1243 fermentation broth by macroporous resin adsorption chromatography.

    PubMed

    Liu, Chunqiao; Zhang, Peng; Liu, Luo; Xu, Tao; Tan, Tianwei; Wang, Fang; Deng, Li

    2013-04-15

    α-Arbutin is a glycosylated hydroquinone which has inhibitory function against tyrosinase. In this work, a one-step isolation of α-arbutin from Xanthomonas CGMCC 1243 fermentation broth by macroporous resin adsorption chromatography was investigated. The research results indicated that S-8 resin offered the best adsorption and desorption capacities for α-arbutin than others and its equilibrium adsorption data were well-fitted to the Freundlich isotherm. In order to optimize the operating parameters for separating α-arbutin, dynamic adsorption and desorption tests on S-8 column chromatography were carried out. Under optimized conditions (adsorption volume of 7 bed volume (BV), mobile phase of 25% (v/v) ethanol solution and elution volume of 3 BV), the purity and recovery of α-arbutin were 97.3% (w/w) and 90.9% (w/w), respectively. The product was identified as α-arbutin by (13)C NMR and (1)H NMR analysis. Moreover, we scaled up S-8 column from laboratory test (10 cm × 2 cm ID) to large scale (500 cm × 100 cm ID) without diminishing α-arbutin yield. In conclusion, the results in this work provide a one-step and cost-effective method for large-scale production of α-arbutin. Copyright © 2013. Published by Elsevier B.V.

  1. Urokinase Separation from Cell Culture Broth of a Human Kidney Cell Line

    PubMed Central

    Bansal, Vibha; Roychoudhury, Pradip K.; Kumar, Ashok

    2007-01-01

    A single step ion-exchange chromatography on a sulfo-propyl (SP)- Sepharose column was performed to separate both the high molecular weight (HMW)- and low molecular weight (LMW)- forms of enzymatically active urokinase type plasminogen activator from human kidney (HT1080) cell culture media. The level of urokinase secreted by the cell line reached to about 145 Plough units/ml culture broth within 48 h of cultivation. The conditioned cell culture media was applied directly to the column without any prior concentration steps. Polyacrylamide gel electrophoresis of the column eluates in the presence of sodium dodecyl sulphate showed that the cell line secretes three forms of two-chain high molecular weight (HMW) urokinase of molecular weights (Mr) 64,000, 60,900 and 55,000. In addition, two low molecular weight (LMW) forms of Mr 22,000 and 20,000; proteolytic cleavage products of HMW, were also found. The HMW and LMW forms had intrinsic plasminogen dependent proteolytic activity as judged by zymographic analysis. The specific activity of the pooled peak fractions increased (approximately 93-fold) to values as high as 1481 Plough units/ mg protein. Both HMW as well as LMW forms were obtained in significantly high yields. PMID:17200693

  2. Library Reference Services.

    ERIC Educational Resources Information Center

    Miller, Constance; And Others

    1985-01-01

    Seven articles on library reference services highlight reference obsolescence in academic libraries, major studies of unobtrusive reference tests, methods for evaluating reference desk performance, reference interview evaluation, problems of reference desk control, online searching by end users, and reference collection development in…

  3. Resuscitation of acid-injured Salmonella in enrichment broth, in apple juice and on the surfaces of fresh-cut cucumber and apple.

    PubMed

    Liao, C-H; Fett, W F

    2005-01-01

    To investigate the resuscitation of acid-injured Salmonella enterica in selected enrichment broths, in apple juice and on cut surfaces of apple and cucumber slices. Following exposure to 2.4% acetic acid for 7 min, S. enterica (serovars Mbandaka, Chester and Newport) cells were used to inoculate enrichment broths, phosphate-buffered saline (PBS), apple juice and fruit slices. Injured Salmonella cells resuscitated and regained the ability to form colonies on selective agar (Xylose-Lysine-Tergitol 4) if they were incubated in lactose broth (LB), universal pre-enrichment broth (UPB) or buffered peptone water (BPW), but not in tetrathionate broth, PBS or apple juice. The resuscitation occurred at a significantly (P > 0.05) faster rate in UPB than in LB or BPW. The resuscitation also occurred on the surfaces of fresh-cut cucumber at 20 degrees C, but not at 4 degrees C. Acid-injured Salmonella cells resuscitated in nonselective enrichment broths at different rates, but not in selective enrichment broth, apple juice, PBS or on fresh-cut apple. Pre-enrichment of food samples in UPB prior to selective enrichment is recommended. Injured Salmonella cells have the ability to resuscitate on fresh-cut surfaces of cucumber when stored at abusive temperatures.

  4. Reference Frames and Relativity.

    ERIC Educational Resources Information Center

    Swartz, Clifford

    1989-01-01

    Stresses the importance of a reference frame in mechanics. Shows the Galilean transformation in terms of relativity theory. Discusses accelerated reference frames and noninertial reference frames. Provides examples of reference frames with diagrams. (YP)

  5. Reference Frames and Relativity.

    ERIC Educational Resources Information Center

    Swartz, Clifford

    1989-01-01

    Stresses the importance of a reference frame in mechanics. Shows the Galilean transformation in terms of relativity theory. Discusses accelerated reference frames and noninertial reference frames. Provides examples of reference frames with diagrams. (YP)

  6. Determination of Ideal Broth Formulations Needed to Prepare Hydrous Cerium Oxide Microspheres via the Internal Gelation Process

    SciTech Connect

    Collins, Jack Lee; Chi, Anthony

    2009-02-01

    A simple test tube methodology was used to determine optimum process parameters for preparing hydrous cerium oxide microspheres via the internal gelation process.1 Broth formulations of cerium ammonium nitrate [(NH4)2Ce(NO3)6], hexamethylenetetramine, and urea were found that can be used to prepare hydrous cerium oxide gel spheres in the temperature range of 60 to 90 C. A few gel-forming runs were made in which microspheres were prepared with some of these formulations to be able to equate the test-tube gelation times to actual gelation times. These preparations confirmed that the test-tube methodology is reliable for determining the ideal broth formulations.

  7. Early Recovery of Salmonella from Food Using a 6-Hour Non-selective Pre-enrichment and Reformulation of Tetrathionate Broth

    PubMed Central

    Daquigan, Ninalynn; Grim, Christopher J.; White, James R.; Hanes, Darcy E.; Jarvis, Karen G.

    2016-01-01

    Culture based methods are commonly employed to detect pathogens in food and environmental samples. These methods are time consuming and complex, requiring multiple non-selective and selective enrichment broths, and usually take at least 1 week to recover and identify pathogens. Improving pathogen detection in foods is a primary goal for regulatory agencies and industry. Salmonella detection in food relies on a series of culture steps in broth formulations optimized to resuscitate Salmonella and reduce the abundance of competitive bacteria. Examples of non-selective pre-enrichment broths used to isolate Salmonella from food include Lactose, Universal Pre-enrichment, BPW, and Trypticase Soy broths. Tetrathionate (TT) and Rappaport–Vassiliadis (RV) broths are employed after a 24-h non-selective enrichment to select for Salmonella and hamper the growth of competitive bacteria. In this study, we tested a new formulation of TT broth that lacks brilliant green dye and has lower levels of TT . We employed this TT broth formulation in conjunction with a 6-h non-selective pre-enrichment period and determined that Salmonella recovery was possible one day earlier than standard food culture methods. We tested the shortened culture method in different non-selective enrichment broths, enumerated Salmonella in the non-selective enrichments, and used 16S rRNA gene sequencing to determine the proportional abundances of Salmonella in the TT and RV selective enrichments. Together these data revealed that a 6-h non-selective pre-enrichment reduces the levels of competitive bacteria inoculated into the selective TT and RV broths, enabling the recovery of Salmonella 1 day earlier than standard culture enrichment methods. PMID:28082968

  8. Extraction of natural red colorants from the fermented broth of Penicillium purpurogenum using aqueous two-phase polymer systems.

    PubMed

    Santos-Ebinuma, Valéria Carvalho; Lopes, André Moreni; Pessoa, Adalberto; Teixeira, Maria Francisca Simas

    2015-01-01

    Safety concerns related to the increasing and widespread application of synthetic coloring agents have increased the demand for natural colorants. Fungi have been employed in the production of novel and safer colorants. In order to obtain the colorants from fermented broth, suitable extraction systems must be developed. Aqueous two-phase polymer systems (ATPPS) offer a favorable chemical environment and provide a promising alternative for extracting and solubilizing these molecules. The aim of this study was to investigate the partitioning of red colorants from the fermented broth of Penicillium purpurogenum using an ATPPS composed of poly(ethylene glycol) (PEG) and sodium polyacrylate (NaPA). Red colorants partitioned preferentially to the top (PEG-rich phase). In systems composed of PEG 6,000 g/mol/NaPA 8,000 g/mol, optimum colorant partition coefficient (KC ) was obtained in the presence of NaCl 0.1 M (KC  = 10.30) while the PEG 10,000 g/mol/NaPA 8,000 g/mol system in the presence of Na2 SO4 0.5 M showed the highest KC (14.78). For both polymers, the mass balance (%MB) and yield in the PEG phase (%ηTOP ) were close to 100 and 79%, respectively. The protein selectivity in all conditions evaluated ranged from 2.0-3.0, which shows a suitable separation of the red colorants and proteins present in the fermented broth. The results suggest that the partitioning of the red colorants is dependent on both the PEG molecular size and salt type. Furthermore, the results obtained support the potential application of ATPPS as the first step of a purification process to recover colorants from fermented broth of microorganisms. © 2015 American Institute of Chemical Engineers.

  9. [Simultaneous determination of meso-erythritol and L-erythrulose in fermentation broth using high performance liquid chromatography].

    PubMed

    Ge, Chiyu; Zhang, Junli; Chen, Jianhua

    2012-08-01

    A high performance liquid chromatographic (HPLC) method has been developed for the simultaneous determination of meso-erythritol and L-erythrulose in fermentation broth. The chromatographic conditions were as follows: Lichrospher 5-NH2 column (250 mm x 4.6 mm) with the temperature of 30 degrees C, acetonitrile-water (90: 10, v/v) as mobile phase with the flow rate of 1.0 mL/min. meso-Erythritol was detected by refractive index (RI) detector at 35 degrees C and L-erythrulose was detected by ultraviolet (UV) detector at 277 nm at room temperature. The linear range for meso-erythritol was 1.00 - 100.00 g/L with a correlation coefficient of 0.998 5. The limit of detection (LOD) and the limit of quantification (LOQ) for meso-erythritol were 0.10 g/L and 0.45 g/L, respectively. The linear range for L-erythrulose was 1.00 - 100.00 g/L with a correlation coefficient of 0.995 8. The LOD and LOQ for L-erythrulose were 0.50 g/L and 0.87 g/L, respectively. The relative standard deviations (RSDs) of intraday and interday for meso-erythritol were less than 3.28% and 5.30%, respectively. The intraday and interday RSDs for L-erythrulose were less than 2.16% and 2.25%, respectively. The recoveries of meso-erythritol and L-erythrulose in fermentation broth were greater than 99%. The samples from fermentation broth were detected at different time points. The measurement by the novel HPLC method was not affected by the other components in the fermentation broth. Furthermore, the HPLC method can be used for the determination of the substrate meso-erythritol and the product L-erythrulose simultaneously.

  10. Isolation of Shiga Toxin-Producing Escherichia coli from Ground Beef Using Multiple Combinations of Enrichment Broths and Selective Agars.

    PubMed

    Brusa, Victoria; Piñeyro, Pablo E; Galli, Lucía; Linares, Luciano H; Ortega, Emanuel E; Padola, Nora L; Leotta, Gerardo A

    2016-03-01

    Shiga toxin-producing Escherichia coli (STEC) are foodborne pathogens, and beef cattle are recognized as the principal reservoir. The aims of this study were (1) to identify the most sensitive combination of selective enrichment broths and agars for STEC isolation in artificially inoculated ground beef samples, and (2) to evaluate the most efficient combination(s) of methods for naturally contaminated ground beef samples. A total of 192 ground beef samples were artificially inoculated with STEC and non-stx bacterial strains. A combination of four enrichment broths and three agars were evaluated for sensitivity, specificity, and positive predictive value for STEC isolation from experimentally inoculated samples. Enrichments with either modified tryptic soy broth (mTSB) containing 8 mg/L novobiocin (mTSB-8) or modified Escherichia coli (mEC) broth followed by isolation in MacConkey agar were the most sensitive combinations for STEC isolation of artificially inoculated samples. Independently, both enrichments media followed by isolation in MacConkey were used to evaluate ground beef samples from 43 retail stores, yielding 65.1% and 58.1% stx-positive samples by RT-PCR, respectively. No difference was observed in the isolate proportions between these two methods (8/25 [32%] and 8/28 [28.6%]). Identical serotypes and stx genotypes were observed in STEC strains isolated from the same samples by either method. In this study, no single enrichment protocol was sufficient to detect all STEC in artificially inoculated samples and had considerable variation in detection ability with naturally contaminated samples. Moreover, none of the single or combinations of multiple isolation agars used were capable of identifying all STEC serogroups in either artificially inoculated or naturally occurring STEC-contaminated ground beef. Therefore, it may be prudent to conclude that there is no single method or combination of isolation methods capable of identifying all STEC serogroups.

  11. Evaluation of disk approximation and single-well broth tests for detection of inducible clindamycin resistance in Streptococcus pneumoniae.

    PubMed

    Jorgensen, James H; McElmeel, M Leticia; Fulcher, Letitia C; McGee, Lesley; Glennen, Anita

    2011-09-01

    This study evaluated an agar disk diffusion D-zone test and an erythromycin-clindamycin (ERY + CLI) single-well broth test for inducible CLI resistance in Streptococcus pneumoniae. The standard CLSI disk approximation test and a single-well combination test incorporating 1 plus 0.5 μg/ml ERY + CLI detected >96% of isolates containing the ermB determinant.

  12. A novel Method for the selective recovery and purification of gamma-polyglutamic acid from Bacillus licheniformis fermentation broth.

    PubMed

    Manocha, Bhavik; Margaritis, Argyrios

    2010-01-01

    Microbially produced gamma-polyglutamic acid (gamma-PGA) is a commercially important biopolymer with many applications in biopharmaceutical, food, cosmetic and waste-water treatment industries. Owing to its increasing demand in various industries, production of gamma-PGA is well documented in the literature, however very few methods have been reported for its recovery. In this paper, we report a novel method for the selective recovery and purification of gamma-PGA from cell-free fermentation broth of Bacillus licheniformis. The cell-free fermentation broth was treated with divalent copper ions, resulting in the precipitation of gamma-PGA, which was collected as a pellet by centrifugation. The pellet was resolubilized and dialyzed against de-ionized water to obtain the purified gamma-PGA biopolymer. The efficiency and selectivity of gamma-PGA recovery was compared with ethanol precipitation method. We found that 85% of the original gamma-PGA content in the broth was recovered by copper sulfate-induced precipitation, compared to 82% recovery by ethanol precipitation method. Since ethanol is a commonly used solvent for protein precipitation, the purity of gamma-PGA precipitate was analyzed by measuring proteins that co-precipitated with gamma-PGA. Of the total proteins present in the broth, 48% proteins were found to be co-precipitated with gamma-PGA by ethanol precipitation, whereas in copper sulfate-induced precipitation, only 3% of proteins were detected in the final purified gamma-PGA, suggesting that copper sulfate-induced precipitation offers better selectivity than ethanol precipitation method. Total metal content analysis of the purified gamma-PGA revealed the undetectable amount of copper ions, whereas other metal ions detected were in low concentration range. The purified gamma-PGA was characterized using infrared spectroscopy. Copyright 2010 American Institute of Chemical Engineers

  13. A new compound from liquid fermentation broth of Armillaria mellea and the determination of its absolute configuration.

    PubMed

    Wang, Yun-Chao; Zhang, Yu-Wei; Zheng, Li-Hua; Bao, Yong-Li; Wu, Yin; Yu, Chun-Lei; Sun, Lu-Guo; Zhang, Yu; Huang, Yan-Xin; Sun, Ying; Li, Yu-Xin

    2013-01-01

    A new 2,5-diketopiperazine, (R)-2-(2-(furan-2-yl)-oxoethyl)-octahydropyrrolo[1,2-a]pyrazine-1,4-dione, and seven known compounds were isolated from the ethyl acetate extract of liquid fermentation broth of Armillaria mellea. The structures of the isolated compounds were established from NMR and HR-MS data. The absolute configuration of the new compound was established by comparing the experimental electronic circular dichroism (ECD) spectrum with the calculated ECD data.

  14. Susceptibility Testing of Fluconazole by the NCCLS Broth Macrodilution Method, E-Test, and Disk Diffusion for Application in the Routine Laboratory

    PubMed Central

    Vandenbossche, Inge; Vaneechoutte, Mario; Vandevenne, Marleen; De Baere, Thierry; Verschraegen, Gerda

    2002-01-01

    Antifungal susceptibility testing may be an important aid in the treatment of patients with life-threatening yeast infections. In order to establish the suitability of different susceptibility test methods for fluconazole with yeasts, the Rosco tablet and the E-test were compared with the gold standard NCCLS broth macrodilution method for 106 yeast strains. These included 102 clinical isolates of Candida spp., including Candida glabrata (n = 30), Candida albicans (n = 20), Candida tropicalis (n = 13), Candida parapsilosis (n = 10), Candida krusei (n = 8), plus Cryptococcus neoformans (n = 3), Saccharomyces cerevisiae (n = 2), and 16 strains belonging to other Candida spp. Four American Type Culture Collection strains of Candida were included as quality controls. The NCCLS method was found to be too complex and labor-intensive for routine testing. The E-test is an accurate alternative, but experience in determining MICs and careful attention to procedural details are critically important. The Rosco tablet showed the best agreement with the NCCLS reference method, especially when newly established breakpoints of R ≤ 10 mm and S ≥ 21 mm were used. PMID:11880416

  15. Isolation and characterization of exopolysaccharide with immunomodulatory activity from fermentation broth of Morchella conica

    PubMed Central

    2013-01-01

    Background and the purpose of this study Mushroom polysaccharides have traditionally been used for the prevention and treatment of a multitude of disorders like infectious illnesses, cancers and various autoimmune diseases. In vitro and in vivo studies suggest that certain polysaccharides affect immune system function. Morchella conica (M. conica) is a species of rare edible mushroom whose multiple medicinal functions have been proven. Thus, the objective of this study is to isolate and characterize of exopolysaccharide from submerged mycelial culture of M. conica, and to evaluate its immunomodulatory activity. Methods A water-soluble Morchella conica Polysaccharides (MCP) were extracted and isolated from the fermentation broth of M. conica through a combination of DEAE-cellulose and Sephacryl S-300 HR chromatograph. NMR and IR spectroscopy has played a developing role in identification of polysaccharide with different structure and composition from fungal and plant sources, as well as complex glycosaminoglycans of animal origin. Thus, NMR and IR spectroscopy were used to analyze the chemical structure and composition of the isolated polysaccharide. Moreover, the polysaccharide was tested for its immunomodulatory activity at different concentrations using in vitro model. Results The results showed that MCP may significantly modulate nitric oxide production in macrophages, and promote splenocytes proliferation. Analysis from HPLC, infrared spectra and nuclear magnetic resonance spectroscopy showed that MCP was a homogeneous mannan with an average molecular weight of approximately 81.2 kDa. The glycosidic bond links is →6)-α-D-Man p-(1→. Conclusion The results suggested that the extracted MCP may modulate nitric oxide production in macrophages and promote splenocytes proliferation, and it may act as a potent immunomodulatory agent. PMID:23351529

  16. RP-HPLC determination of recombinant human interferon omega in the Pichia pastoris fermentation broth.

    PubMed

    Liu, Hong; Pan, Hong-Chun; Peng, Li; Cai, Shao-Xi

    2005-07-15

    A rapid and valid reversed-phase high performance liquid chromatography (RP-HPLC) method for determination of recombinant human interferon omega (rhIFNomega) in the yeast Pichia pastoris fermentation broth was developed. The method is based on the hydrophobicity of rhIFNomega followed by RP-HPLC separation with UV detection. The chromatography analysis was performed on EC 250/4 NUCLEOSIL 300-5 C18 (250 mm x 4 mm i.d., 300 A, with a particle size of 5 microm) column. The compositions of the mobile phase A and B were 999:1 (v/v) water/TFA and 999:1 (v/v) acetonitrile/TFA at a flow rate of 1.0 ml min(-1). Detection was done by spectrophotometry at 280 nm and the column temperature was 30+/-1 degrees C. Calibration curve was linear (r=0.9986, n=7) in the range of 0.074-0.555 mg ml(-1) for rhIFNomega and the regression equation was y=2.02 x 10(6)x-1.27 x 10(5). Limit of detection for rhIFNomega was 0.053 mg ml(-1). The values of R.S.D. (%) of intra-day and inter-day precision were <5.65 and <5.68 (n=6), respectively. The R.S.D. (%) values and the average recovery rate of recovery experiment were <1.23 (n=3) and 97.97%.

  17. Indirect methods for characterization of carbon dioxide levels in fermentation broth.

    PubMed

    Frick, R; Junker, B

    1999-01-01

    Various factors which influence dissolved carbon dioxide levels were indirectly evaluated in pilot scale and laboratory studies. For pilot scale studies, off-gas carbon dioxide (percentage in exit air) was measured using a mass spectrometer and then its potential impact on dissolved carbon dioxide concentrations qualitatively examined. Greater volumetric air flowrates reduced off-gas carbon dioxide levels more effectively at lower airflow ranges and thus lowered expected dissolved carbon dioxide levels through gas stripping. Lower broth pH values decreased off-gas carbon dioxide levels but increased expected dissolved carbon dioxide levels due to the pH-dependence of the gas/liquid carbon dioxide equilibrium. While back-pressure increases had an insignificant effect on off-gas carbon dioxide levels, they directly affected expected dissolved carbon dioxide levels according to Henry's law. Laboratory studies, conducted using both uninoculated and inoculated fermentation media, quantified the response of the media to pH changes with bicarbonate addition, specifically its buffering capacity. This effect then was related qualitatively to expected dissolved carbon dioxide levels. Higher dissolved carbon dioxide levels, as demonstrated by reduced pH changes with bicarbonate addition, thus would be expected for salt solutions of increased ionic strength and higher protein content media. In addition, pH changes with greater bicarbonate additions declined for fermentation samples taken over the course of a one week cultivation, most likely due to the higher protein content associated with biomass growth. The presence of weak acids/bases initially in the media or formed as metabolic by products, as well as the concentration of buffering ions such as phosphate, also were believed to be important contributing elements to the buffering capacity of the solution.

  18. [Determination of enantiomeric purity for lactic acid in fermentation broth by Rhizopus oryzae with high performance liquid chromatography].

    PubMed

    Bai, D M; Zhao, X M; Hu, Z D

    2001-01-01

    A procedure for the resolution of DL-lactic acid and the determination of D-isomer ratio in L-lactic acid fermentation broth by Rhizopus oryzae is described. The effects of pH of mobile phase and concentration of chiral mobile phase additives on resolution of DL-lactic acid were investigated. The optical isomers of lactic acid were resolved by RP-HPLC with 2,3,6-tri-O-beta-cyclodextrin(TM-beta-CD) as a chiral mobile phase additive, and C18 column as stationary phase, and detected at wavelength 210 nm. The results showed that a correction factor should be introduced into the equation for calculation of the percentage of D-lactic acid, because the UV absorption of D-lactic acid and L-lactic acid might not be the same when TM-beta-CD was present. Quantitation was achieved with external standard method, the average recovery was 100.4%, and the relative standard deviation was 0.82%. This method can be used for the determination of the percentage of D-isomer in L-lactic acid fermentation broth by Rhizopus oryzae, and it is simple, rapid and accurate. The results showed that the mass fraction of D-isomer in the fermented broth increased during the period of storage.

  19. Radiation sensitivity of poliovirus, a model for norovirus, inoculated in oyster ( Crassostrea gigas) and culture broth under different conditions

    NASA Astrophysics Data System (ADS)

    Jung, Pil-Mun; Park, Jae Seok; Park, Jin-Gyu; Park, Jae-Nam; Han, In-Jun; Song, Beom-Seok; Choi, Jong-il; Kim, Jae-Hun; Byun, Myung-Woo; Baek, Min; Chung, Young-Jin; Lee, Ju-Woon

    2009-07-01

    Poliovirus is a recognized surrogate for norovirus, pathogen in water and food, due to the structural and genetic similarity. Although radiation sensitivity of poliovirus in water or media had been reported, there has been no research in food model such as shellfish. In this study, oyster ( Crassostrea gigas) was incubated in artificial seawater contaminated with poliovirus, and thus radiation sensitivity of poliovirus was determined in inoculated oyster. The effects of ionizing radiation on the sensitivity of poliovirus were also evaluated under different conditions such as pH (4-7) and salt concentration (1-15%) in culture broth, and temperature during irradiation. The D10 value of poliovirus in PBS buffer, virus culture broth and oyster was determined to 0.46, 2.84 and 2.94 kGy, respectively. The initial plaque forming unit (PFU) of poliovirus in culture broth was slightly decreased as the decrease of pH and the increase of salt concentration, but radiation sensitivity was not affected by pH and salt contents. However, radiation resistance of poliovirus was increased at frozen state. These results provide the basic information for the inactivation of pathogenic virus in foods by using irradiation.

  20. Functionalized magnetic mesoporous silica nanoparticles: fabrication, laccase adsorption performance and direct laccase capture from Trametes versicolor fermentation broth.

    PubMed

    Wang, Feng; Huang, Wei; Guo, Chen; Liu, Chun-Zhao

    2012-12-01

    A simple and highly efficient protocol using magnetic mesoporous silica nanoparticles (MMSNPs) with metal affinity ligands was developed to directly capture laccase from Trametes versicolor fermentation broth. The Cu(2+)-chelated magnetic mesoporous silica nanoparticles (MMSNPs-Cu(2+)) with pore sizes ranging from 3.6 to 27.1 nm exhibited size selectivity on laccase capture from the fermentation broth, and the MMSNPs-Cu(2+) with an average pore size of 14.5 nm provided 60.6-fold purification of laccase and 114.6% recovery yield of enzyme activity. Both size selectivity of the MMSNPs and affinity of the chelated metal ion resulted in high laccase capture efficiency from the fermentation broth. The most efficient MMSNPs-Cu(2+) demonstrated no significant loss in laccase capture effectiveness following 10 reuse cycles. This simple and efficient strategy has the potential to be used for the robust and inexpensive preparation of purified laccase at the industrial scale. Copyright © 2012 Elsevier Ltd. All rights reserved.

  1. Evaluation of a matrix-assisted laser desorption ionization-time of flight mass spectrometry assisted, selective broth method to screen for vancomycin-resistant enterococci in patients at high risk

    PubMed Central

    Huang, Tsi-Shu; Lee, Susan Shin-Jung; Lee, Chia-Chien; Chen, Chiu-Yen; Chen, Fang-Chen; Chen, Bao-Chen; Sy, Cheng Len

    2017-01-01

    Background Bile esculin azide with vancomycin (BEAV) medium is a sensitive, but slightly less specific method for vancomycin-resistant enterococci (VRE) screening. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) is a rapid method for identification of clinical pathogens. This study aimed to assess the performance of a novel combination screening test for VRE, using BEAV broth combined with MALDI-TOF MS. Materials and methods Clinical specimens were collected from patients at risk of VRE carriage, and tested by the novel combination method, using selective BEAV broth culture method followed by MALDI-TOF MS identification (SBEAVM). The reference method used for comparison was the ChromID VRE agar method. Results A total of 135 specimens were collected from 78 patients, and 63 specimens tested positive for VRE positive using the ChromID VRE method (positive rate 46.7%). The sensitivity, specificity, positive predictive value, and negative predictive value of SBEAVM method after an incubation period of 28 hours were 93.7%, 90.3%, 89.4%, and 94.2%, respectively. The SBEAVM method when compared to the ChromID VRE method had a shorter turnaround time (29 vs. 48–72 hours) and lower laboratory cost ($2.11 vs. $3.23 per test). Conclusions This study demonstrates that SBEAVM is a rapid, inexpensive, and accurate method for use in VRE screening. PMID:28609453

  2. Reference Service Policy Statement.

    ERIC Educational Resources Information Center

    Young, William F.

    This reference service policy manual provides general guidelines to encourage reference service of the highest possible quality and to insure uniform practice. The policy refers only to reference service in the University Libraries and is intended for use in conjunction with other policies and procedures issued by the Reference Services Division.…

  3. Reference Service Policy Statement.

    ERIC Educational Resources Information Center

    Young, William F.

    This reference service policy manual provides general guidelines to encourage reference service of the highest possible quality and to insure uniform practice. The policy refers only to reference service in the University Libraries and is intended for use in conjunction with other policies and procedures issued by the Reference Services Division.…

  4. Use of blood-free enrichment broth in the development of a rapid protocol to detect Campylobacter in twenty-five grams of chicken meat.

    PubMed

    Hayashi, Masahiro; Kubota-Hayashi, Sayoko; Natori, Tatsuya; Mizuno, Takuya; Miyata, Machiko; Yoshida, Shigeru; Zhang, Jiwei; Kawamoto, Keiko; Ohkusu, Kiyofumi; Makino, Souichi; Ezaki, Takayuki

    2013-04-15

    A Food Pathogen Enrichment (FPE) broth, which supports the growth of Campylobacter without lysed blood and CO2, was developed. The FPE broth supports the growth of Campylobacter to the same degree as Bolton and Preston broths. Using the FPE broth, we developed a novel rapid protocol to detect small numbers of Campylobacter in 25g of food. The sensitivity of FPE enrichment and PCR to detect Campylobacter spp. from spiked chicken meat was determined. The detection sensitivities for non-stressed C. jejuni and C. coli from fresh meat ranged from 5.8 to 1.1×10(1)CFU per 25g of chicken meat, and those for freeze-stressed C. jejuni and C. coli from frozen meat ranged from 9.9×10(1) to 2.0×10(2)CFU. The FPE broth enrichment culture (24h) of chicken meat, followed by PCR, resulted in a significantly higher detection score (80% positive) than conventional Bolton enrichment and subsequent colony isolation using mCCDA agar plates (18% positive). Differences between our new protocol and the Bolton enrichment method were due to the overgrowth of many resistant bacteria, especially extended-spectrum beta-lactamase-producing bacteria in the Bolton enrichment broth.

  5. Identification of Gram-Negative Bacteria and Genetic Resistance Determinants from Positive Blood Culture Broths by Use of the Verigene Gram-Negative Blood Culture Multiplex Microarray-Based Molecular Assay

    PubMed Central

    Ledeboer, Nathan A.; Lopansri, Bert K.; Dhiman, Neelam; Cavagnolo, Robert; Carroll, Karen C.; Granato, Paul; Thomson, Richard; Butler-Wu, Susan M.; Berger, Heather; Samuel, Linoj; Pancholi, Preeti; Swyers, Lettie; Hansen, Glen T.; Tran, Nam K.; Polage, Christopher R.; Thomson, Kenneth S.; Hanson, Nancy D.; Winegar, Richard

    2015-01-01

    Bloodstream infection is a serious condition associated with significant morbidity and mortality. The outcome of these infections can be positively affected by the early implementation of effective antibiotic therapy based on the identification of the infecting organism and genetic markers associated with antibiotic resistance. In this study, we evaluated the microarray-based Verigene Gram-negative blood culture (BC-GN) assay in the identification of 8 genus or species targets and 6 genetic resistance determinants in positive blood culture broths. A total of 1,847 blood cultures containing Gram-negative organisms were tested using the BC-GN assay. This comprised 729 prospective fresh, 781 prospective or retrospective frozen, and 337 simulated cultures representing 7 types of aerobic culture media. The results were compared to those with standard bacterial culture and biochemical identification with nucleic acid sequence confirmation of the resistance determinants. Among monomicrobial cultures, the positive percent agreement (PPA) of the BC-GN assay with the reference method was as follows; Escherichia coli, 100%; Klebsiella pneumoniae, 92.9%; Klebsiella oxytoca, 95.5%; Enterobacter spp., 99.3%; Pseudomonas aeruginosa, 98.9%; Proteus spp., 100%; Acinetobacter spp., 98.4%; and Citrobacter spp., 100%. All organism identification targets demonstrated >99.5% negative percent agreement (NPA) with the reference method. Of note, 25/26 cultures containing K. pneumoniae that were reported as not detected by the BC-GN assay were subsequently identified as Klebsiella variicola. The PPA for identification of resistance determinants was as follows; blaCTX-M, 98.9%; blaKPC, 100%; blaNDM, 96.2%; blaOXA, 94.3%; blaVIM, 100%; and blaIMP, 100%. All resistance determinant targets demonstrated >99.9% NPA. Among polymicrobial specimens, the BC-GN assay correctly identified at least one organism in 95.4% of the broths and correctly identified all organisms present in 54.5% of the broths

  6. Identification of Gram-Negative Bacteria and Genetic Resistance Determinants from Positive Blood Culture Broths by Use of the Verigene Gram-Negative Blood Culture Multiplex Microarray-Based Molecular Assay.

    PubMed

    Ledeboer, Nathan A; Lopansri, Bert K; Dhiman, Neelam; Cavagnolo, Robert; Carroll, Karen C; Granato, Paul; Thomson, Richard; Butler-Wu, Susan M; Berger, Heather; Samuel, Linoj; Pancholi, Preeti; Swyers, Lettie; Hansen, Glen T; Tran, Nam K; Polage, Christopher R; Thomson, Kenneth S; Hanson, Nancy D; Winegar, Richard; Buchan, Blake W

    2015-08-01

    Bloodstream infection is a serious condition associated with significant morbidity and mortality. The outcome of these infections can be positively affected by the early implementation of effective antibiotic therapy based on the identification of the infecting organism and genetic markers associated with antibiotic resistance. In this study, we evaluated the microarray-based Verigene Gram-negative blood culture (BC-GN) assay in the identification of 8 genus or species targets and 6 genetic resistance determinants in positive blood culture broths. A total of 1,847 blood cultures containing Gram-negative organisms were tested using the BC-GN assay. This comprised 729 prospective fresh, 781 prospective or retrospective frozen, and 337 simulated cultures representing 7 types of aerobic culture media. The results were compared to those with standard bacterial culture and biochemical identification with nucleic acid sequence confirmation of the resistance determinants. Among monomicrobial cultures, the positive percent agreement (PPA) of the BC-GN assay with the reference method was as follows; Escherichia coli, 100%; Klebsiella pneumoniae, 92.9%; Klebsiella oxytoca, 95.5%; Enterobacter spp., 99.3%; Pseudomonas aeruginosa, 98.9%; Proteus spp., 100%; Acinetobacter spp., 98.4%; and Citrobacter spp., 100%. All organism identification targets demonstrated >99.5% negative percent agreement (NPA) with the reference method. Of note, 25/26 cultures containing K. pneumoniae that were reported as not detected by the BC-GN assay were subsequently identified as Klebsiella variicola. The PPA for identification of resistance determinants was as follows; blaCTX-M, 98.9%; blaKPC, 100%; blaNDM, 96.2%; blaOXA, 94.3%; blaVIM, 100%; and blaIMP, 100%. All resistance determinant targets demonstrated >99.9% NPA. Among polymicrobial specimens, the BC-GN assay correctly identified at least one organism in 95.4% of the broths and correctly identified all organisms present in 54.5% of the broths

  7. Model-based design of a pilot-scale simulated moving bed for purification of citric acid from fermentation broth.

    PubMed

    Wu, Jinglan; Peng, Qijun; Arlt, Wolfgang; Minceva, Mirjana

    2009-12-11

    One of the conventional processes used for the recovery of citric acid from its fermentation broth is environmentally harmful and cost intensive. In this work an innovative benign process, which comprises simulated moving bed (SMB) technology and use of a tailor-made tertiary poly(4-vinylpyridine) (PVP) resin as a stationary phase is proposed. This paper focuses on a model-based design of the operation conditions for an existing pilot-scale SMB plant. The SMB unit is modeled on the basis of experimentally determined hydrodynamics, thermodynamics and mass transfer characteristics in a single chromatographic column. Three mathematical models are applied and validated for the prediction of the experimentally attained breakthrough and elution profiles of citric acid and the main impurity component (glucose). The transport dispersive model was selected for the SMB simulation and design studies, since it gives a satisfactory prediction of the elution profiles within acceptable computational time. The equivalent true moving bed (TMB) and SMB models give a good prediction of the experimentally attained SMB separation performances, obtained with a real clarified and concentrated fermentation broth as a feed mixture. The SMB separation requirements are set to at least 99.8% citric acid purity and 90% citric acid recovery in the extract stream. The complete regeneration in sections 1 and 4 is unnecessary. Therefore the net flow rates in all four SMB sections have been considered in the unit design. The influences of the operating conditions (the flow rate in each section, switching time and unit configuration) on the SMB performances were investigated systematically. The resulting SMB design provides 99.8% citric acid purity and 97.2% citric acid recovery in the extract. In addition the citric acid concentration in the extract is a half of its concentration in the pretreated fermentation broth (feed).

  8. A new cyclopeptide with antifungal activity from the co-culture broth of two marine mangrove fungi.

    PubMed

    Li, Chunyuan; Wang, Jinhua; Luo, Cuiping; Ding, Weijia; Cox, Daniel G

    2014-01-01

    A new cyclic tetrapeptide, cyclo-(L-leucyl-trans-4-hydroxy-L-prolyl-D-leucyl-trans-4-hydroxy-L-proline) (1), was isolated from the co-culture broth of two mangrove fungi Phomopsis sp. K38 and Alternaria sp. E33. The structure of 1 was determined by analysis of spectroscopic data and Marfey's analytic method. Primary bioassay demonstrated that compound 1 exhibited moderate to high inhibitory activity against four crop-threatening fungi including Gaeumannomyces graminis, Rhizoctonia cerealis, Helminthosporium sativum and Fusarium graminearum as compared with triadimefon.

  9. Reach for Reference. Four Recent Reference Books

    ERIC Educational Resources Information Center

    Safford, Barbara Ripp

    2004-01-01

    This article provides descriptions of four new science and technology encyclopedias that are appropriate for inclusion in upper elementary and/or middle school reference collections. "The Macmillan Encyclopedia of Weather" (Stern, Macmillan Reference/Gale), a one-volume encyclopedia for upper elementary and middle level students, is a…

  10. Reach for Reference. Four Recent Reference Books

    ERIC Educational Resources Information Center

    Safford, Barbara Ripp

    2004-01-01

    This article provides descriptions of four new science and technology encyclopedias that are appropriate for inclusion in upper elementary and/or middle school reference collections. "The Macmillan Encyclopedia of Weather" (Stern, Macmillan Reference/Gale), a one-volume encyclopedia for upper elementary and middle level students, is a…

  11. Fundamentals of Reference

    ERIC Educational Resources Information Center

    Mulac, Carolyn M.

    2012-01-01

    The all-in-one "Reference reference" you've been waiting for, this invaluable book offers a concise introduction to reference sources and services for a variety of readers, from library staff members who are asked to work in the reference department to managers and others who wish to familiarize themselves with this important area of…

  12. Live, Digital Reference.

    ERIC Educational Resources Information Center

    Kenney, Brian

    2002-01-01

    Discusses digital reference services, also known as virtual reference, chat reference, or online reference, based on a round table discussion at the 2002 American Library Association annual conference in Atlanta. Topics include numbers and marketing; sustainability; competition and models; evaluation methods; outsourcing; staffing and training;…

  13. Statistical Reference Datasets

    National Institute of Standards and Technology Data Gateway

    Statistical Reference Datasets (Web, free access)   The Statistical Reference Datasets is also supported by the Standard Reference Data Program. The purpose of this project is to improve the accuracy of statistical software by providing reference datasets with certified computational results that enable the objective evaluation of statistical software.

  14. Live, Digital Reference.

    ERIC Educational Resources Information Center

    Kenney, Brian

    2002-01-01

    Discusses digital reference services, also known as virtual reference, chat reference, or online reference, based on a round table discussion at the 2002 American Library Association annual conference in Atlanta. Topics include numbers and marketing; sustainability; competition and models; evaluation methods; outsourcing; staffing and training;…

  15. Direct blood culturing on solid medium outperforms an automated continuously monitored broth-based blood culture system in terms of time to identification and susceptibility testing

    PubMed Central

    Idelevich, E.A.; Grünastel, B.; Peters, G.; Becker, K.

    2015-01-01

    Pathogen identification and antimicrobial susceptibility testing (AST) should be available as soon as possible for patients with bloodstream infections. We investigated whether a lysis-centrifugation (LC) blood culture (BC) method, combined with matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) identification and Vitek 2 AST, provides a time advantage in comparison with the currently used automated broth-based BC system. Seven bacterial reference strains were added each to 10 mL human blood in final concentrations of 100, 10 and 1 CFU/mL. Inoculated blood was added to the Isolator 10 tube and centrifuged at 3000 g for 30 min, then 1.5 mL sediment was distributed onto five 150-mm agar plates. Growth was observed hourly and microcolonies were subjected to MALDI-TOF MS and Vitek 2 as soon as possible. For comparison, seeded blood was introduced into an aerobic BC bottle and incubated in the BACTEC 9240 automated BC system. For all species/concentration combinations except one, successful identification and Vitek 2 inoculation were achieved even before growth detection by BACTEC. The fastest identification and inoculation for AST were achieved with Escherichia coli in concentrations of 100 CFU/mL and 10 CFU/mL (after 7 h each, while BACTEC flagged respective samples positive after 9.5 h and 10 h). Use of the LC-BC method allows skipping of incubation in automated BC systems and, used in combination with rapid diagnostics from microcolonies, provides a considerable advantage in time to result. This suggests that the usefulness of direct BC on solid medium should be re-evaluated in the era of rapid microbiology. PMID:26909155

  16. Direct blood culturing on solid medium outperforms an automated continuously monitored broth-based blood culture system in terms of time to identification and susceptibility testing.

    PubMed

    Idelevich, E A; Grünastel, B; Peters, G; Becker, K

    2016-03-01

    Pathogen identification and antimicrobial susceptibility testing (AST) should be available as soon as possible for patients with bloodstream infections. We investigated whether a lysis-centrifugation (LC) blood culture (BC) method, combined with matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) identification and Vitek 2 AST, provides a time advantage in comparison with the currently used automated broth-based BC system. Seven bacterial reference strains were added each to 10 mL human blood in final concentrations of 100, 10 and 1 CFU/mL. Inoculated blood was added to the Isolator 10 tube and centrifuged at 3000 g for 30 min, then 1.5 mL sediment was distributed onto five 150-mm agar plates. Growth was observed hourly and microcolonies were subjected to MALDI-TOF MS and Vitek 2 as soon as possible. For comparison, seeded blood was introduced into an aerobic BC bottle and incubated in the BACTEC 9240 automated BC system. For all species/concentration combinations except one, successful identification and Vitek 2 inoculation were achieved even before growth detection by BACTEC. The fastest identification and inoculation for AST were achieved with Escherichia coli in concentrations of 100 CFU/mL and 10 CFU/mL (after 7 h each, while BACTEC flagged respective samples positive after 9.5 h and 10 h). Use of the LC-BC method allows skipping of incubation in automated BC systems and, used in combination with rapid diagnostics from microcolonies, provides a considerable advantage in time to result. This suggests that the usefulness of direct BC on solid medium should be re-evaluated in the era of rapid microbiology.

  17. Isolation of natural red colorants from fermented broth using ionic liquid-based aqueous two-phase systems.

    PubMed

    Ventura, Sónia P M; Santos-Ebinuma, Valéria C; Pereira, Jorge F B; Teixeira, Maria F S; Pessoa, Adalberto; Coutinho, João A P

    2013-05-01

    There is a growing demand for natural colorants. This is prompting the search for new alternative and "benign" separation systems allowing higher recoveries, extraction yields, and selectivities. This work investigates the use of aqueous two-phase systems (ATPS) based on ionic liquids as extraction processes for the recovery of red colorants from the fermented broth of Penicillium purpurogenum DPUA 1275. Several ATPS based on quaternary ammonium and imidazolium were studied in this work aiming at separating the red colorants produced from the remaining colorants and contaminant proteins present in the fermented broth. The results suggest that the red colorants can be isolated by an appropriate manipulation of some of the process conditions, such as the use of quaternary ammonium with short alkyl chains, alkaline media, and short tie-line lengths (extraction point systems with lower concentrations of ionic liquid). These conditions allow large partition coefficients for the red colorants (K red = 24.4 ± 2.3), high protein removal (60.7 ± 2.8 %) and selectivity parameters (S red/prot = 10.05).

  18. A platform technology of recovery of lactic acid from a fermentation broth of novel substrate Zizyphus oenophlia.

    PubMed

    Bishai, Moumita; De, Swarnalok; Adhikari, Basudam; Banerjee, Rintu

    2015-08-01

    Lactic acid, a biologically derived compound, exists ubiquitously in nature. Its existence ranges from human being to microorganisms. Having paramount industrial significance, lactic acid should be highly pure, devoid of any contaminants. Hence, development of minimum steps of platform technologies to purify it needs urgent attention. The article proposed a novel and simple process for separation of lactic acid from a potential substrate Zizyphus oenophlia, based on ion exchange chromatography. The process herein involves two steps of purification; firstly a weak anion exchange resin was used to separate lactic acid from other anions present in the broth. This was followed by use of strong cation exchanger which washes out the target molecule (lactic acid) while trapped other cations present in the solution. The selected ion exchangers were Amberlite IRA 96 and Amberlite IR 120. Amberlite IRA 96 retained the lactic acid from the broth while washing away other anions. Maximum binding capacity of the resin was found to 210.46 mg lactic acid/g bead. After the simple two-step purification process, the purity of lactic acid improves up to 99.17 % with a recovery yield of 98.9 %. Upon characterization, formation of only levo rotatory form of lactic acid confirms its easy metabolism by the human system, thus triggering its application towards biomaterial sector.

  19. [Determination of sugars, organic acids and alcohols in microbial consortium fermentation broth from cellulose using high performance liquid chromatography].

    PubMed

    Jiang, Yan; Fan, Guifang; Du, Ran; Li, Peipei; Jiang, Li

    2015-08-01

    A high performance liquid chromatographic method was established for the determination of metabolites (sugars, organic acids and alcohols) in microbial consortium fermentation broth from cellulose. Sulfate was first added in the samples to precipitate calcium ions in microbial consortium culture medium and lower the pH of the solution to avoid the dissociation of organic acids, then the filtrates were effectively separated using high performance liquid chromatography. Cellobiose, glucose, ethanol, butanol, glycerol, acetic acid and butyric acid were quantitatively analyzed. The detection limits were in the range of 0.10-2.00 mg/L. The linear correlation coefficients were greater than 0.999 6 in the range of 0.020 to 1.000 g/L. The recoveries were in the range of 85.41%-115.60% with the relative standard deviations of 0.22% -4.62% (n = 6). This method is accurate for the quantitative analysis of the alcohols, organic acids and saccharides in microbial consortium fermentation broth from cellulose.

  20. Multi-probe real-time PCR identification of four common Candida species in blood culture broth.

    PubMed

    Foongladda, Suporn; Mongkol, Nanthanida; Petlum, Pornphan; Chayakulkeeree, Methee

    2014-06-01

    We developed a single-tube real-time polymerase chain reaction (PCR) assay with multiple hybridization probes for detecting Candida albicans, C. tropicalis, C. glabrata, and C. parapsilosis. Primers were designed to amplify 18S rRNA gene of the genus Candida, and DNA probes were designed to hybridize two areas of the amplicons. The amplification curves and specific melting peaks of the probes hybridized with PCR product were used for definite species identifications. The reaction specificity was 100 % when evaluating the assay using DNA samples from 21 isolates of fungal and bacterial species. The assay was further evaluated in 129 fungal blood culture broth samples which were culture positive for fungus. Of the 129 samples, 119 were positively identified as: C. albicans (39), C. tropicalis (30), C. parapsilosis (23), C. glabrata (20), Candida spp. (5), and two samples containing mixed C. glabrata/C. albicans and C. glabrata/C. tropicalis. The five Candida spp. were identified by sequencing analysis as C. krusei, C. dubliniensis, C. aquaetextoris, and two isolates of C. athensensis. Of the ten samples which showed negative PCR results, six were Cryptococcus neoformans, and the others were Trichosporon sp., Rhodotorula sp., Fusarium sp., and Penicillium marneffei. Our findings show that the assay was highly effective in identifying the four medically important Candida species. The results can be available within 3 h after positivity of a blood culture broth sample.

  1. Comprehension of viscous morphology--evaluation of fractal and conventional parameters for rheological characterization of Aspergillus niger culture broth.

    PubMed

    Wucherpfennig, Thomas; Lakowitz, Antonia; Krull, Rainer

    2013-01-20

    The filamentous fungus Aspergillus niger is a widely used host in industrial processes from food, chemical to pharmaceutical industry. The most prominent feature of this filamentous microorganism in submerged cultivation is its complex morphology which comprises dense spherical pellets as well as viscous elongated filaments. Depending on culture conditions, the exhibited morphology has tremendous effect on the overall process, making a precise understanding of fungal growth and morphology indispensable. Morphology, however, is only industrially relevant as long as it can be linked to important cultivation characteristics of filamentous microorganisms such as culture broth flow behavior. In the present study, different conventional and fractal morphological parameters gained from automatic image analysis were tested for their eligibility to predict culture broth rheology from morphologic appearance. The introduced biomass independent rheological parameters K(BDW) and n(BDW) obtained by power law relationship were successfully estimated from morphology related fractal and conventional parameters. For improved characterization of morphologic appearance of filamentous fungi newly introduced fractal quotient and lacunarity were compared to conventional particle shape parameters in form of the earlier established Morphology number (MN). Copyright © 2012 Elsevier B.V. All rights reserved.

  2. Effect of Sex on Flavor-related and Functional Compounds in Freeze-dried Broth Made from Korean Native Chicken

    PubMed Central

    Jayasena, Dinesh D.; Jung, Samooel; Alahakoon, Amali U.; Nam, Ki Chang

    2014-01-01

    Studies on the flavour characteristics of meat-based broth, quantification of flavour-related and functional compounds, and factors affecting the availability of such compounds are minimal. The present study was designed to determine the effects of sex on flavor-related and functional compounds in freeze-dried broth (FDB) made from Korean native chickens (KNC). Male and female KNC from a commercial strain (WoorimatdagTM) were reared under similar commercial conditions. FDB was separately prepared using male and female birds aged 100 d (six birds of each sex) and analyzed for nucleotide, free amino acid, betaine, carnitine, carnosine, anserine, and creatine contents, and fatty acid composition. The levels of betaine, carnitine and creatine in FDB were not significantly different between the two sexes (p>0.05) in KNC. Carnosine and anserine were not detected in FDB samples. However, FDB from female chickens had significantly higher inosine-5-monophosphate and arachidonic acid contents than did FDB from male chickens. FDB prepared with male KNC contained higher levels of inosine, linoleic acid, glycine, alanine, lysine, and serine (p<0.05). However, glutamic acid, oleic acid, and DHA were present in comparable amounts (p>0.05) in FDB made from male and female KNC. Our findings suggest that the sex of KNC has significant effect on the contents of flavor-related compounds, but not functional compounds. PMID:26761282

  3. [Effects of salting, cut type, and initial simmering temperature on protein and fat contents of meat broths: I. Beef].

    PubMed

    Gotera-Prado, Z; Quintero, J B; Huerta-Leidenz, N; Prado Gotera, Z

    1997-12-01

    A 2 x 2 x 4 factorial design was used to study variation of protein and fat contents in beef broths as affected by cut type (flank, shank), salt treatments (addition of salt to the medium, no salt), and initial temperatures of simmering (25, 70, 75, and 100 degrees C). Flank portions yielded slightly more protein (0.29 g/100 mL) and had three-fold less fat (0.39 g/mL) than those of shank (0.25 and 1.12 g/mL, respectively) (P < 0.05). No linear relationship of temperature and amount of extractable components was observed, but it was clear that the greatest protein extraction was accomplished when meat was immersed in cooking water at boiling point (P < 0.05). In general, salting of water reduced fat content of beef broths. However, a significant Salting x Cut type interaction showed this effect was only present in shanks (P < 0.05). Conversely, the reducing effect (P < 0.05) of salting on amount of protein extracted from flank was not observed in shanks. Based on these data, we conclude that larger amounts of protein and less fat could be transferred from meat pieces to the medium by immersing beef in salted water at the boiling point.

  4. Determination of MICs of streptomycin for resistant Salmonella isolates in swine and poultry using a micro-broth dilution system.

    PubMed

    Edrington, Thomas S; Harvey, Roger B; Farrington, Leigh A; Nisbet, David J

    2002-03-01

    The MICs of streptomycin for Salmonella isolates from swine and poultry were determined by a micro-broth dilution technique. The Salmonella isolates were recovered from the lymph nodes and cecal contents of market-age swine and from the cecal contents of poultry at the time of slaughter and were found by disk diffusion to be resistant to 10 microg of streptomycin. MIC testing was carried out with the Sensititre susceptibility system for streptomycin, which uses a microwell concentration gradient of 16 to 800 microg/ml. Results indicated that >80% of the swine isolates had MICs of < or = 64 microg/ml, while 51% of poultry isolates exhibited MICs of > or = 128 microg/ml. The highest MICs observed in swine and poultry were 256 and 800 microg/ml, respectively. Replicate tests performed on 12 of the isolates chosen at random indicated a 100% correlation between runs. Advantages of this system include easily read results and precoated wells. Disadvantages include the cost and the inability to test concentrations of streptomycin other than those in the wells. We found this micro-broth dilution commercial test kit to provide a relatively quick and easy testing procedure for the determination of streptomycin resistance in Salmonella.

  5. Functional magnetic mesoporous nanoparticles for efficient purification of laccase from fermentation broth in magnetically stabilized fluidized bed.

    PubMed

    Wang, Feng; Guo, Chen; Liu, Chun-Zhao

    2013-12-01

    A magnetically stabilized fluidized bed (MSFB) with the Cu(2+)-chelated magnetic mesoporous silica nanoparticles (MMSNPs-Cu(2+)) was established to purify laccase directly from the fermentation broth of Trametes versicolor. The MMSNPs-Cu(2+) particles in the MSFB maintained a stable bed expansion of two to threefold at a flow rate of 120-180 cm/h. At the optimal magnetic field intensity of 120 Gs, both the maximal Bodenstein number and the smallest axial dispersion coefficient were achieved, which resulted in a stable fluidization stage. The dynamic binding capacity of laccase in the MSFB decreased from 192.5 to144.3 mg/g when the flow velocity through the bed increased from 44.2 to 69.8 cm/h. The MSFB with MMSNPs-Cu(2+) achieved efficient laccase purification from the fermentation broth with 62.4-fold purification of laccase and 108.9 % activity yield. These results provided an excellent platform for the application of these magnetic mesoporous nanoparticles integrated with the MSFB in developing novel protein purification process.

  6. Assessment of tolerance induction by Origanum vulgare L. essential oil or carvacrol in Pseudomonas aeruginosa cultivated in a meat-based broth and in a meat model.

    PubMed

    da Silva Luz, Isabelle; Gomes-Neto, Nelson Justino; Magnani, Marciane; de Souza, Evandro Leite

    2015-12-01

    This study assessed the efficacy of Origanum vulgare L. essential oil (OVEO) and carvacrol in inhibiting the growth of Pseudomonas aeruginosa ATCC 9027, as well as the development of direct tolerance and cross-tolerance when this bacterium was challenged with sublethal amounts of these substances in a meat-based broth and in a meat model. OVEO and carvacrol at their minimum inhibitory concentrations (MICs), 1/2 MIC and 1/4 MIC decreased the viable cell counts of P. aeruginosa in meat-based broth. Direct tolerance or cross-tolerance was not induced after exposure of the assayed bacterial strain to sublethal amounts of OVEO or carvacrol in meat-based broth and in an artificially contaminated ground beef. Bacterial cells progressively subcultured in meat-based broth with increasing amounts of the tested substances survived up to the MIC of OVEO and to 1/2 MIC of carvacrol. The results reveal a lack of induction of tolerance in P. aeruginosa by exposure to OVEO or carvacrol in meat-based broth and in a meat model.

  7. Genetics Home Reference

    MedlinePlus

    Skip Navigation Bar Home Current Issue Past Issues Genetics Home Reference Past Issues / Spring 2007 Table of ... of this page please turn Javascript on. The Genetics Home Reference (GHR) Web site — ghr.nlm.nih. ...

  8. The Test of Reference.

    ERIC Educational Resources Information Center

    Childers, Thomas

    1980-01-01

    Reports the results of an unobtrusive study, from a user's viewpoint, of reference services available in the Suffolk Cooperative Library System. The study raises questions of policy centering around user expectations of library reference services. (RAA)

  9. The Floating Reference Librarian

    ERIC Educational Resources Information Center

    Hernon, Peter; Pastine, Maureen

    1972-01-01

    The floating librarian'' is one who interprets and adjusts the formal library structure to meet legitimate needs. This is one of the ways the academic reference librarian can gain greater acceptance with students and faculty. (9 references) (Author/NH)

  10. Academic Reference Service

    ERIC Educational Resources Information Center

    Lopez, Manuel D.

    1973-01-01

    Cost benefit accounting is not being used in academic reference libraries today but administrators are beginning to require quantitative evaluation of services provided. Several systems are described and evaluated. (35 references) (DH)

  11. Herbal reference standards.

    PubMed

    Schwarz, Michael; Klier, Bernhard; Sievers, Hartwig

    2009-06-01

    This review describes the current definitions and regulatory requirements that apply to reference standards that are used to analyse herbal products. It also describes and discusses the current use of reference substances and reference extracts in the European and United States pharmacopoeias.

  12. Academic Library Reference Services.

    ERIC Educational Resources Information Center

    Batt, Fred

    This examination of the philosophy and objectives of academic library reference services provides an overview of the major reference approaches to fulfilling the following primary objectives of reference services: (1) providing accurate answers to patrons' questions and/or helping patrons find sources to pursue their research needs; (2) building…

  13. Academic Library Reference Services.

    ERIC Educational Resources Information Center

    Batt, Fred

    This examination of the philosophy and objectives of academic library reference services provides an overview of the major reference approaches to fulfilling the following primary objectives of reference services: (1) providing accurate answers to patrons' questions and/or helping patrons find sources to pursue their research needs; (2) building…

  14. Assessment of Reference Services.

    ERIC Educational Resources Information Center

    Von Seggern, Marilyn

    1987-01-01

    This annotated bibliography of materials dealing with the evaluation of library reference services is arranged by category including literature success, quality, and accuracy of answers; cost and task analysis; interviews and communication; classification of reference questions; reference collections; staff availability; use and nonuse of…

  15. Effectiveness of the antimicrobial removal device, BACTEC 16B medium, and thiol broth in neutralizing antibacterial activities of imipenem, norfloxacin, and related agents.

    PubMed

    Weinberg, E; Shungu, D L; Gadebusch, H H

    1984-02-01

    The Antimicrobial Removal Device (ARD), BACTEC 16B medium, and Thiol broth were evaluated for their effectiveness in reducing the activity of imipenem (IPM), cefoxitin, moxalactam, and ceftazidime in blood samples. In addition, the capability of the ARD and Thiol broth to bind norfloxacin and the ARD to bind oxolinic and nalidixic acids in urine samples was investigated. At the highest concentrations of the drugs tested (32 micrograms/ml for the four beta-lactams and 256 micrograms/ml for the three quinolinecarboxylic acids), there was at least a 95% reduction in the in vitro activity of each of the antibacterial agents for treated versus untreated samples. Of the compounds tested in the ARD system, the organic acids were more completely removed than were the beta-lactams. The Thiol broth was more effective than the ARD and the BACTEC 16B medium in inactivating imipenem, but it had no effect on the antibacterial activity of norfloxacin.

  16. Effectiveness of the antimicrobial removal device, BACTEC 16B medium, and thiol broth in neutralizing antibacterial activities of imipenem, norfloxacin, and related agents.

    PubMed Central

    Weinberg, E; Shungu, D L; Gadebusch, H H

    1984-01-01

    The Antimicrobial Removal Device (ARD), BACTEC 16B medium, and Thiol broth were evaluated for their effectiveness in reducing the activity of imipenem (IPM), cefoxitin, moxalactam, and ceftazidime in blood samples. In addition, the capability of the ARD and Thiol broth to bind norfloxacin and the ARD to bind oxolinic and nalidixic acids in urine samples was investigated. At the highest concentrations of the drugs tested (32 micrograms/ml for the four beta-lactams and 256 micrograms/ml for the three quinolinecarboxylic acids), there was at least a 95% reduction in the in vitro activity of each of the antibacterial agents for treated versus untreated samples. Of the compounds tested in the ARD system, the organic acids were more completely removed than were the beta-lactams. The Thiol broth was more effective than the ARD and the BACTEC 16B medium in inactivating imipenem, but it had no effect on the antibacterial activity of norfloxacin. PMID:6230369

  17. Preparative isolation and purification of anti-tumor agent ansamitocin P-3 from fermentation broth of Actinosynnema pretiosum using high-performance counter-current chromatography.

    PubMed

    Yao, Yuqin; Cheng, Zhihui; Ye, Haoyu; Xie, Yongmei; He, Jing; Tang, Minghai; Shen, Tao; Wang, Jiangman; Zhou, Yan; Lu, Zejun; Luo, Feng; Chen, Lijuan; Yu, Luoting; Yang, Jin-Liang; Peng, Aihua; Wei, Yuquan

    2010-05-01

    Ansamitocin P-3 is a potent anti-tumor maytansinoid found in Actinosynnema pretiosum. However, due to the complexity of the fermentation broth of Actinomycete, how to effectively separate ansamitocin P-3 is still a challenge. In this study, both analytical and preparative high-performance counter-current chromatography were successfully used to separate and purify ansamitocin P-3 from fermentation broth. A total of 28.8 mg ansamitocin P-3 with purity of 98.4% was separated from 160 mg crude sample of fermentation broth in less than 80 min with the two-phase solvent system of hexane-ethyl acetate-methanol-water (0.6:1:0.6:1, v/v/v/v). The purity and structural identification were determined by HPLC, (1)H NMR, (13)C NMR and mass spectroscopy.

  18. Determination of amino acids in cell culture and fermentation broth media using anion-exchange chromatography with integrated pulsed amperometric detection.

    PubMed

    Hanko, Valoran P; Rohrer, Jeffrey S

    2004-01-01

    Cell culture and fermentation broth media are used in the manufacture of biotherapeutics and many other biological materials. Characterizing the amino acid composition in cell culture and fermentation broth media is important because deficiencies in these nutrients can reduce desired yields or alter final product quality. Anion-exchange (AE) chromatography using sodium hydroxide (NaOH) and sodium acetate gradients, coupled with integrated pulsed amperometric detection (IPAD), determines amino acids without sample derivatization. AE-IPAD also detects carbohydrates, glycols, and sugar alcohols. The presence of these compounds, often at high concentrations in cell culture and fermentation broth media, can complicate amino acid determinations. To determine whether these samples can be analyzed without sample preparation, we studied the effects of altering and extending the initial NaOH eluent concentration on the retention of 42 different carbohydrates and related compounds, 30 amino acids and related compounds, and 3 additional compounds. We found that carbohydrate retention is impacted in a manner different from that of amino acid retention by a change in [NaOH]. We used this selectivity difference to design amino acid determinations of diluted cell culture and fermentation broth media, including Bacto yeast extract-peptone-dextrose (yeast culture medium) broth, Luria-Bertani (bacterial culture medium) broth, and minimal essential medium and serum-free protein-free hybridoma medium (mammalian cell culture media). These media were selected as representatives for both prokaryotic and eukaryotic culture systems capable of challenging the analytical technique presented in this paper. Glucose up to 10mM (0.2%, w/w) did not interfere with the chromatography, or decrease recovery greater than 20%, for the common amino acids arginine, lysine, alanine, threonine, glycine, valine, serine, proline, isoleucine, leucine, methionine, histidine, phenylalanine, glutamate, aspartate

  19. References for marine science

    NASA Astrophysics Data System (ADS)

    1990-06-01

    Standard and Reference Materials for Marine Science, National Oceanic and Atmospheric Administration Technical Memo OMA-51 (2nd edition, 434 pp.), by A. Y. Cantillo, is now available. This compilation of reference materials was prepared at the request of the Group of Experts on Standards and Reference Materials and was printed by NOAA. GESREM is sponsored by the International Atomic Energy Agency, the Intergovernmental Oceanographic Commission, and the United Nations Program.Reference materials are included on ashes, gases, instrument performance materials, oils, physical properties, rocks, sediments, sludges, tissues and waters. For each reference material, source, description and preparation, analyses and values, cost, references, and comments are given. Indices are included for elements, isotopes and organic compounds. Cross references to Chemical Abstracts Service registry numbers and alternate names and chemical structures of organic compounds are also provided.

  20. Evaluation of colistin and nalidixic acid in Todd-Hewitt broth for selective isolation of group B streptocci.

    PubMed

    Fenton, L J; Harper, M H

    1979-02-01

    Todd-Hewitt broth (THB) containing colistin and nalidixic acid was compared with four other media with respect to efficacy of isolation of group B streptocci. It was effective as plain THB, THB with gentamicin and nalidixic acid and blood, and THB with colistin, nalidixic acid, and blood. THB with gentamicin and nalidixic acid, but without blood, was inhibitory to group B streptocci. The value of THB with colistin and nalidixic acid lies in its ability to successfully inhibit gram-negative organisms and still promote group B streptococcal growth without the addition of blood. This greatly reduces the time and expense of media preparation and permits early determination of bacterial growth, so that other means of rapidly identifying group B streptocci can be applied at the earliest possible time.

  1. Evaluation of colistin and nalidixic acid in Todd-Hewitt broth for selective isolation of group B streptocci.

    PubMed Central

    Fenton, L J; Harper, M H

    1979-01-01

    Todd-Hewitt broth (THB) containing colistin and nalidixic acid was compared with four other media with respect to efficacy of isolation of group B streptocci. It was effective as plain THB, THB with gentamicin and nalidixic acid and blood, and THB with colistin, nalidixic acid, and blood. THB with gentamicin and nalidixic acid, but without blood, was inhibitory to group B streptocci. The value of THB with colistin and nalidixic acid lies in its ability to successfully inhibit gram-negative organisms and still promote group B streptococcal growth without the addition of blood. This greatly reduces the time and expense of media preparation and permits early determination of bacterial growth, so that other means of rapidly identifying group B streptocci can be applied at the earliest possible time. PMID:372220

  2. Purification of nattokinase by reverse micelles extraction from fermentation broth: effect of temperature and phase volume ratio.

    PubMed

    Liu, Jun-Guo; Xing, Jian-Min; Chang, Tian-Shi; Liu, Hui-Zhou

    2006-03-01

    Nattokinase is a novel fibrinolytic enzyme that is considered to be a promising agent for thrombosis therapy. In this study, reverse micelles extraction was applied to purify and concentrate nattokinase from fermentation broth. The effects of temperature and phase volume ratio used for the forward and backward extraction on the extraction process were examined. The optimal temperature for forward and backward extraction were 25 degrees C and 35 degrees C respectively. Nattokinase became more thermosensitive during reverse micelles extraction. And it could be enriched in the stripping phase eight times during backward extraction. It was found that nattokinase could be purified by AOT reverse micelles with up to 80% activity recovery and with a purification factor of 3.9.

  3. Oxidative production of xylonic acid using xylose in distillation stillage of cellulosic ethanol fermentation broth by Gluconobacter oxydans.

    PubMed

    Zhang, Hongsen; Han, Xushen; Wei, Chengxiang; Bao, Jie

    2017-01-01

    An oxidative production process of xylonic acid using xylose in distillation stillage of cellulosic ethanol fermentation broth was designed, experimentally investigated, and evaluated. Dry dilute acid pretreated and biodetoxified corn stover was simultaneously saccharified and fermented into 59.80g/L of ethanol (no xylose utilization). 65.39g/L of xylose was obtained in the distillation stillage without any concentrating step after ethanol was distillated. Then the xylose was completely converted into 66.42g/L of xylonic acid by Gluconobacter oxydans. The rigorous Aspen Plus modeling shows that the wastewater generation and energy consumption was significantly reduced comparing to the previous xylonic acid production process using xylose in pretreatment liquid. This study provided a practical process option for xylonic acid production from lignocellulose feedstock with significant reduction of wastewater and energy consumption.

  4. Impact of changes in broth composition on Chlorella vulgaris cultivation in a membrane photobioreactor (MPBR) with permeate recycle.

    PubMed

    Discart, V; Bilad, M R; Marbelia, L; Vankelecom, I F J

    2014-01-01

    A membrane photobioreactor (MPBR) is a proven and very useful concept in which microalgae can be simultaneously cultivated and pre-harvested. However, the behavior with respect to accumulation of algogenic organic matter, including transparent exopolymeric particles (TEPs), counter ions and unassimilated nutrients due to the recycling of the medium is still unclear, even though the understanding of this behavior is essential for the optimization of microalgae processing. Therefore, the dynamics of these compounds, especially TEPs, during coupled cultivation and harvesting of Chlorella vulgaris in an MPBR with permeate recycle are addressed in this study. Results show that TEPs are secreted during algae cell growth, and that their presence is thus inevitable. In the system with permeate recycle, substances such as counter ions and unassimilated nutrients get accumulated in the system. This was proven to limit the algae growth, together with the occurrence of bioflocculation due to an increasing broth pH.

  5. Effects of LB broth, naphthalene concentration, and acetone on the naphthalene degradation activities by Pseudomonas putida G7.

    PubMed

    Chang, Su-Yun; Liu, Xue-Gong; Ren, Bi-Qiong; Liu, Bo; Zhang, Kai; Zhang, Honggui; Wan, Yao

    2015-01-01

    Luria-Bertani broth and acetone were usually used in naphthalene degradation experiments as nutrient and solvent. However, their effect on the degradation was seldom mentioned. In this work, we investigated the effect of LB, naphthalene concentration, and acetone on the degradation of naphthalene by Pseudomonas putida G7, which is useful for the degradation of naphthalene on future field remediation. By adding LB, the naphthalene degradation efficiencies and naphthalene dioxygenase were both decreased by 98%, while the catechol dioxygenase was decreased by 90%. Degradation of naphthalene was also inhibited when naphthalene concentration was 56 ppm and higher, which was accompanied with the accumulation of orange-colored metabolism products. However, acetone can stimulate the degradation of naphthalene, and the stimulation was more obvious when naphthalene concentration was lower than 2000 ppm. By assaying the enzyme activities of naphthalene dioxygenase and catechol dioxygenase, it was thought that the degradation efficiency was depending on the more sensitive enzymes on the complicated conditions.

  6. Solid phase extraction of lactic acid from fermentation broth by anion-exchangeable silica confined ionic liquids.

    PubMed

    Bi, Wentao; Zhou, Jun; Row, Kyung Ho

    2011-01-15

    Three anion-exchangeable, silica-confined ionic liquids were synthesized for solid phase extraction of lactic acid from fermentation broth, followed by high-performance liquid chromatography coupled to ultraviolet detection. By comparing the adsorption isotherms of lactic acid on different silica-confined ionic liquids, interactions between the lactic acid and sorbents were investigated. The adsorbed amounts were then fitted into different adsorption isotherm equations; finally, the Langmuir equation was selected. Then the imidazolium silica with the highest adsorption capacity of lactic acid was packed into a cartridge for solid phase extraction. The loading volume of the cartridge was optimized by the Langmuir equation and geometry. After washing with distilled water and eluting with 0.25 mol L(-1) of an HCl solution, the lactic acid was separated from interference with a recovery yield of 91.9%. Furthermore, this kind of anion-exchangeable material exhibited potential for industrial applications and separation of other anionic bioactive compounds.

  7. [Gene expression differences of toxigenic and nontoxigenic Vibrio cholerae strains in mannitol fermentation medium and Luria-Bertani broth].

    PubMed

    Zhang, Hongzhi; Pang, Bo; Zhang, Li; Kan, Biao

    2009-06-01

    To analyze gene expression differences of toxigenic and nontoxigenic strains of El Tor Vibrio cholerae growing separately in mannitol fermentation medium and LB (Luria-Bertani) broth. Total RNA was extracted from the mannitol slow-fermenting strain N16961 (toxigenic) and the mannitol fast-fermenting strain 93097 (nontoxigenic) at 1 h of fermentation. The large scale gene expression profiles were detected and compared with high throughout microarray. By comparing the strains growing in different cultures, we found 142 differentially expressed genes in N16961 and 418 genes in 93097. Most of these genes were grouped into six functional classes. They were mainly related to transport and binding, energy metabolism, protein biosynthesis, and protein fate. The expression levels of genes in N16961 and 93097 were affected by culture conditions, which can serve as basis for further studying the mechanism of metabolism of mannitol.

  8. Pyrroloquinoline quinone from Gluconobacter oxydans fermentation broth enhances superoxide anion-scavenging capacity of Cu/Zn-SOD.

    PubMed

    Ma, Ke; Cui, Jun-Zhu; Ye, Jian-Bin; Hu, Xian-Mei; Ma, Ge-Li; Yang, Xue-Peng

    2017-09-01

    A bioassay-guided fractionation of extract from Gluconobacter oxydans fermentation broth afforded Compound 1, which was identified as pyrroloquinoline quinone (PQQ) by spectroscopic methods. PQQ has been shown to enhance the superoxide anion-scavenging capacity significantly for Cu/Zn-SOD. To illustrate the mechanism, the interaction between PQQ and Cu/Zn-SOD was investigated. The multiple binding sites involving hydrogen bonds and van der Waals force between PQQ and Cu/Zn-SOD were revealed by isothermal titration calorimetry. The α-helix content was increased in the Cu/Zn-SOD structure with the addition of PQQ into the solution through ultraviolet (UV) spectroscopy. These results indicated that PQQ could change the conformation of Cu/Zn-SOD through interaction, which could enhance its superoxide anion-scavenging capacity. Therefore, PQQ is a potential natural antioxidant. Copyright © 2017 Elsevier Ltd. All rights reserved.

  9. Antibiotic purification from fermentation broths by counter-current chromatography: analysis of product purity and yield trade-offs.

    PubMed

    Booth, A J; Ngiam, S H; Lye, G J

    2004-12-01

    Counter-current chromatography (CCC) is a low pressure, liquid-liquid chromatographic technique which has proven to be a powerful purification tool for the high-resolution fractionation of a variety of active pharmaceutical compounds. The successful integration of CCC into either existing or new manufacturing processes requires the predictable purification of target compounds from crude, fermentation-derived, feed streams. This work examines the feasibility of CCC for the purification of fermentation-derived erythromycin A (EA) from its structurally and chemically similar analogues. At the laboratory scale, the effect of feed pre-treatment using either clarified, forward extracted (butyl acetate) or back extracted broth on EA separation was investigated. This defined the degree of impurity removal required, i.e. back extracted broth, to ensure a reproducible elution profile of EA during CCC. Optimisation and scale-up of the separation studied the effects of mobile phase flow (2-40 ml.min(-1)) and solute loading (0.1-10 g) on the attainable EA purity and yield. The results in all cases demonstrated a high attainable EA purity (>97% w/w) with throughputs up to 0.33 kg.day(-1). Secondly, a predictive scale-up model was applied demonstrating, that from knowledge of the solute distribution ratio of EA (K(EA)) at the laboratory scale, the EA elution time at the pilot scale could be predicted to within 3-10%, depending upon the solute injection volume. In addition, this study has evaluated a "fractionation diagram" approach to visually determine the effects of key operational variables on separation performance. This resulted in accurate fraction cut-point determination for a required degree of product purity and yield. Overall, the results show CCC to be a predictable and scaleable separation technique capable of handling real feed streams.

  10. A simple one pot purification of bacterial amylase from fermented broth based on affinity toward starch-functionalized magnetic nanoparticle.

    PubMed

    Paul, Tanima; Chatterjee, Saptarshi; Bandyopadhyay, Arghya; Chattopadhyay, Dwiptirtha; Basu, Semanti; Sarkar, Keka

    2015-08-18

    Surface-functionalized adsorbant particles in combination with magnetic separation techniques have received considerable attention in recent years. Selective manipulation on such magnetic nanoparticles permits separation with high affinity in the presence of other suspended solids. Amylase is used extensively in food and allied industries. Purification of amylase from bacterial sources is a matter of concern because most of the industrial need for amylase is met by microbial sources. Here we report a simple, cost-effective, one-pot purification technique for bacterial amylase directly from fermented broth of Bacillus megaterium utilizing starch-coated superparamagnetic iron oxide nanoparticles (SPION). SPION was prepared by co-precipitation method and then functionalized by starch coating. The synthesized nanoparticles were characterized by transmission electron microscopy (TEM), a superconducting quantum interference device (SQUID, zeta potential, and ultraviolet-visible (UV-vis) and Fourier-transform infrared (FTIR) spectroscopy. The starch-coated nanoparticles efficiently purified amylase from bacterial fermented broth with 93.22% recovery and 12.57-fold purification. Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) revealed that the molecular mass of the purified amylase was 67 kD, and native gel showed the retention of amylase activity even after purification. Optimum pH and temperature of the purified amylase were 7 and 50°C, respectively, and it was stable over a range of 20°C to 50°C. Hence, an improved one-pot bacterial amylase purification method was developed using starch-coated SPION.

  11. The combination of lactate and diacetate synergistically reduces cold growth in brain heart infusion broth across Listeria monocytogenes lineages.

    PubMed

    Stasiewicz, Matthew J; Wiedmann, Martin; Bergholz, Teresa M

    2010-04-01

    Combinations of organic acids are often used in ready-to-eat foods to control the growth of Listeria monocytogenes during refrigerated storage. The purpose of this study was to quantitatively assess synergy between two organic acid growth inhibitors under conditions similar to those present in cold-smoked salmon, and to assess the effect of evolutionary lineage on response to those growth inhibitors. Thirteen strains of L. monocytogenes, representing lineages I and II, were grown at 7 degrees C in broth at pH 6.1 and 4.65% water-phase NaCl, which was supplemented with 2% potassium lactate, 0.14% sodium diacetate, or the combination of both at the same levels. Our data suggest that lineages adapt similarly to these inhibitors, as the only significant growth parameter difference between lineages was a minor effect (+/- 0.16 day, P = 0.0499) on lag phase (lambda). For all strains, lactate significantly extended lambda, from 2.6 +/- 0.4 to 3.8 +/- 0.5 days (P < 0.001), and lowered the maximum growth rate (mu(max)) from 0.54 +/- 0.06 to 0.49 +/- 0.04 log(CFU/ml)/day (P < 0.001), compared with the control. Diacetate was ineffective alone, but in combination with lactate, synergistically increased lambda to 6.6 +/- 1.6 days (P < 0.001) and decreased mu(max) to 0.34 +/- 0.05 log(CFU/ml)/day (P < 0.001). Monte Carlo simulations provided further evidence for synergy between diacetate and lactate by predicting signficantly slower growth to nominal endpoints for the combination of inhibitors. This study shows potassium lactate and sodium diacetate have significant synergistic effects on both lambda and mu(max) of L. monocytogenes at refrigeration temperature in broth, and justifies combining these inhibitors, at effective levels, in food product formulations.

  12. Effect of organic acids and temperature on survival of Shigella flexneri in broth at pH 4.

    PubMed

    Zaika, Laura L

    2002-09-01

    The survival of bacterial pathogens in acidified foods depends not only on the hydrogen ion concentration, but also on the type of acid and the storage temperature. Shigella flexneri is a foodborne pathogen that is acid tolerant. The survival of S. flexneri 5348 in brain heart infusion broth supplemented with 0.04 M acetic, citric, lactic, malic, or tartaric acid and adjusted to pH 4 with HCI or NaOH was studied. The control medium was brain heart infusion broth adjusted to pH 4 with HCI. Stationary-phase cells were inoculated into media at initial populations of 6 to 7 log10 CFU/ml and incubated at 4, 19, 28, and 37 degrees C. A two-phase linear inactivation model was applied to plate count data to derive lag times (tL) and slopes of the curves, from which D-values and time required for a 4-log10 decrease in population (T4D) were calculated. In all cases, survival increased with decreasing temperature. For each acid, tL, the D-value, and T4D increased with decreasing temperature. All acids inhibited S. flexneri to some extent but to differing degrees as follows: lactic acid, acetic acid > citric acid, malic acid, tartaric acid > HCl. The T4D values for the control medium and for media containing acetic, citric, lactic, malic, and tartaric acids were 64, 47, 50, 34, 58, and 52 h, respectively, at 37 degrees C and 2,607, 1,498, 1,905, 1,346, 1,726, and 2,134 h, respectively, at 4 degrees C. The results of this study indicate that organic acids may aid in the inactivation of Shigella. However, these data also suggest that foods stored at or below room temperature containing low levels (< 1%) of acids could cause illness if contaminated with Shigella.

  13. Synthetic growth reference charts.

    PubMed

    Hermanussen, Michael; Stec, Karol; Aßmann, Christian; Meigen, Christof; Van Buuren, Stef

    2016-01-01

    To reanalyze the between-population variance in height, weight, and body mass index (BMI), and to provide a globally applicable technique for generating synthetic growth reference charts. Using a baseline set of 196 female and 197 male growth studies published since 1831, common factors of height, weight, and BMI are extracted via Principal Components separately for height, weight, and BMI. Combining information from single growth studies and the common factors using in principle a Bayesian rationale allows for provision of completed reference charts. The suggested approach can be used for generating synthetic growth reference charts with LMS values for height, weight, and BMI, from birth to maturity, from any limited set of height and weight measurements of a given population. Generating synthetic growth reference charts by incorporating information from a large set of reference growth studies seems suitable for populations with no autochthonous references at hand yet. © 2015 Wiley Periodicals, Inc.

  14. High frequency reference electrode

    DOEpatents

    Kronberg, James W.

    1994-01-01

    A high frequency reference electrode for electrochemical experiments comprises a mercury-calomel or silver-silver chloride reference electrode with a layer of platinum around it and a layer of a chemically and electrically resistant material such as TEFLON around the platinum covering all but a small ring or "halo" at the tip of the reference electrode, adjacent to the active portion of the reference electrode. The voltage output of the platinum layer, which serves as a redox electrode, and that of the reference electrode are coupled by a capacitor or a set of capacitors and the coupled output transmitted to a standard laboratory potentiostat. The platinum may be applied by thermal decomposition to the surface of the reference electrode. The electrode provides superior high-frequency response over conventional electrodes.

  15. High frequency reference electrode

    DOEpatents

    Kronberg, J.W.

    1994-05-31

    A high frequency reference electrode for electrochemical experiments comprises a mercury-calomel or silver-silver chloride reference electrode with a layer of platinum around it and a layer of a chemically and electrically resistant material such as TEFLON around the platinum covering all but a small ring or halo' at the tip of the reference electrode, adjacent to the active portion of the reference electrode. The voltage output of the platinum layer, which serves as a redox electrode, and that of the reference electrode are coupled by a capacitor or a set of capacitors and the coupled output transmitted to a standard laboratory potentiostat. The platinum may be applied by thermal decomposition to the surface of the reference electrode. The electrode provides superior high-frequency response over conventional electrodes. 4 figs.

  16. Preparing the references.

    PubMed

    Peh, W C G; Ng, K H

    2009-07-01

    In a scientific paper, the references serve to provide background information and allow the researcher to compare and contrast the work of others in relation to his own study. Authors are responsible for the accuracy of all references cited. The references quoted should be easily accessible and retrievable by anyone wishing to obtain further information. There is a strong preference for citing journal articles listed in PubMed. The two major reference format systems are the Vancouver and Harvard systems, with increasing preference for the Vancouver system. Authors should adhere exactly to the instructions to authors of the target journal.

  17. Evaluation of universal pre-enrichment broth for isolation of Salmonella spp., Escherichia coli O157:H7, and Listeria monocytogenes from dairy farm environmental samples.

    PubMed

    Nam, H M; Murinda, S E; Nguyen, L T; Oliver, S P

    2004-01-01

    Use of universal pre-enrichment broth (UPB) as a primary enrichment medium for detection of Salmonella spp., Escherichia coli O157:H7, and Listeria monocytogenes from dairy farm environmental samples was evaluated. There were no differences in bacterial growth between UPB and selective primary enrichment broths for each pathogen inoculated individually or in combination at 10(1) and 10(2) colony forming units/mL. In addition, no differences were observed when UPB and selective primary enrichment broths were compared for detection efficiency of pathogens in artificially contaminated raw milk and fecal samples. Listeria enrichment broth (LEB) was compared with UPB to support growth of L. monocytogenes from naturally contaminated environmental samples. Listeria monocytogenes was isolated from seven of 30 samples enriched in UPB and six of 30 samples enriched in LEB. Dairy farm environmental samples were examined for recovery of the three pathogens using UPB. Subsequent isolation was achieved using selective secondary enrichment of each pathogen. Listeria monocytogenes, Salmonella spp., and E. coli O157:H7 were isolated in 13.4% (30 of 224), 8.9% (20 of 224), and 2.2% (five of 224) of samples, respectively. Isolation rates of the three pathogens were somewhat higher than in previous reports. Overall, UPB supported growth of test pathogens to detectable levels within 24 h. Our results demonstrate that UPB has potential for routine use in isolation of foodborne pathogens from diverse environmental samples.

  18. Effect of sample preparation and bacterial concentration on Salmonella enterica detection in poultry meat using culture methods and PCR assaying of preenrichment broths.

    PubMed

    Kanki, Masashi; Sakata, Junko; Taguchi, Masumi; Kumeda, Yuko; Ishibashi, Masanori; Kawai, Takao; Kawatsu, Kentaro; Yamasaki, Wataru; Inoue, Kiyoshi; Miyahara, Michiko

    2009-02-01

    We evaluated the sensitivity of a PCR assay in the detection of Salmonella enterica at the broth preenrichment step of poultry meat. A total of 162 retail poultry meat samples, which were prepared by manual massaging, stomacher or no homogenization were compared for Salmonella recovery. Using these homogenization methods, the PCR assay at the broth preenrichment step detected Salmonella in, respectively, 48.9%, 62.2% and 50.0% of meat and giblet samples detected as Salmonella-positive using the culture method. In ground chicken, however, Salmonella was detected in 21.7% of samples treated by stomacher homogenization, compared to 40.7% and 48% of untreated and hand-massaged samples, respectively. These results suggest that stomaching of ground chicken causes excessive effusion of food constituents, which affects PCR results. Using the most probable number (MPN) technique, Salmonella was detected at under 1.0 CFU/g in 12 ground chicken samples and under 10(3)CFU/ml of broth in seven of the 12 broth-enriched samples, which considered the minimum concentration detectable by PCR assay. These results show that Salmonella detection using routine PCR assays is difficult in poultry meat, and in particular ground chicken, due to low amounts of Salmonella and the presence of inhibitors.

  19. Determination of gibberellins in fermentation broth produced by Fusarium verticilliodes MTCC 156 by high-performance liquid chromatography tandem mass spectrometry.

    PubMed

    Sharma, Rohit; Iyer, Jitesh P; Chakraborti, Asit K; Banerjee, Uttam Chand

    2004-02-01

    A method for the detection of gibberellins produced by Fusarium verticilliodes is described using HPLCMS/MS (HPLC tandem MS). A Hypersil (5 microm) octadecylsilane column with methanol/water as eluent in the ratio 3:1 at a flow rate of 0.5 ml/min was used. In the HPLCMS, GA(3) (gibberellic acid; m / z 346.3) eluted at retention time tr=3.08 min, with the corresponding mass chromatogram having peaks at m / z 346.7 and 328.8 corresponding to the M+ and M+-H2O ions respectively. The ethyl acetate extract from the broth, subjected to HPLCMS analysis under similar conditions, showed a constituent with tr=2.13 min, the mass chromatogram of which exhibited peaks at m / z 348.9 and 331.9 corresponding to the MH+ and MH+-H2O ions respectively. Comparison of the MS and MS/MS results (direct infusion) of an authentic sample of GA3 and the ethyl acetate extract from the broth revealed the formation of reduced GA3 in the broth. The present study, utilizing HPLCMS/MS, describes an improved methodology for the unambiguous determination and estimation of gibberellins from fermentation broth.

  20. Growth Kinetics of Listeria monocytogenes in Broth and Beef Frankfurters– Determination of Lag Phase Duration and Exponential Growth Rate under Isothermal Conditions

    USDA-ARS?s Scientific Manuscript database

    The objective of this research was to develop a new kinetic model to describe the isothermal growth of microorganisms. The new model was tested with Listeria monocytogenes in broth and frankfurters, and compared with two commonly used models - Baranyi and modified Gompertz models. Bias factor (BF)...

  1. Suppression of pecan and peach pathogens using metabolites or broths of from symbiotic bacteria obtained from the guts of entomopathogenic nematodes

    USDA-ARS?s Scientific Manuscript database

    Concentrated metabolites from the bacteria Xenorhabdus spp. and Photorhabdus spp. have previously been shown to suppress growth of peach and pecan pathogens in vitro, and reduce disease on detached leaves or terminals. The objectives of this study were 1) determine if bacterial broths (in addition t...

  2. Effect of fermentation time on antioxidative activities of Ganoderma lucidum broth using leguminous plants as part of the liquid fermentation medium.

    PubMed

    Chien, Yung Liang; Ho, Chi Tang; Chiang, Been Huang; Hwang, Lucy Sun

    2011-06-15

    Oxidative damage plays an important role in the pathology of human diseases. Ganoderma lucidum, a medicinal fungus, has been used for thousands of years in traditional Oriental medicine. It is reported to have antioxidant functions such as inhibition of lipid peroxidation. The objective of the present study was to investigate the effect of fermentation time on the antioxidative activities of G. lucidum broth filtrate using leguminous plants as part of the liquid fermentation medium. Inhibition of Cu(2+)-induced oxidation of human low-density lipoprotein (LDL), DPPH radical-scavenging activity, total phenolic compounds, isoflavones and protocatechuic acid were measured to evaluate the antioxidant activity of G. lucidum fermentation broth filtrate. Our results showed that black soybean and Astragalus membranaceus improved the antioxidant activity of the G. lucidum fermentation broth filtrate. Protocatechuic acid was identified by LC-MS as the antioxidant compounds whose relative potency of inhibiting LDL oxidation to Trolox is 1.55. Protocatechuic acid showed positive correlation with the antioxidant activity of the fermentation broth filtrate while isoflavones did not contribute to antioxidant activity. Copyright © 2010 Elsevier Ltd. All rights reserved.

  3. Marketing Reference Services.

    ERIC Educational Resources Information Center

    Norman, O. Gene

    1995-01-01

    Relates the marketing concept to library reference services. Highlights include a review of the literature and an overview of marketing, including research, the marketing mix, strategic plan, marketing plan, and marketing audit. Marketing principles are applied to reference services through the marketing mix elements of product, price, place, and…

  4. An Online Reference System.

    ERIC Educational Resources Information Center

    Chisman, Janet; Treat, William

    1984-01-01

    Describes a computer aid developed to assist in academic library reference service using the DataPhase Circulation System, an automated system that features full cataloging records in database and permits local programing. Access points (subject, type of reference work, course) and database structure and user screens are highlighted. (EJS)

  5. Marketing Reference Services.

    ERIC Educational Resources Information Center

    Norman, O. Gene

    1995-01-01

    Relates the marketing concept to library reference services. Highlights include a review of the literature and an overview of marketing, including research, the marketing mix, strategic plan, marketing plan, and marketing audit. Marketing principles are applied to reference services through the marketing mix elements of product, price, place, and…

  6. Reference Point Heterogeneity

    PubMed Central

    Terzi, Ayse; Koedijk, Kees; Noussair, Charles N.; Pownall, Rachel

    2016-01-01

    It is well-established that, when confronted with a decision to be taken under risk, individuals use reference payoff levels as important inputs. The purpose of this paper is to study which reference points characterize decisions in a setting in which there are several plausible reference levels of payoff. We report an experiment, in which we investigate which of four potential reference points: (1) a population average payoff level, (2) the announced expected payoff of peers in a similar decision situation, (3) a historical average level of earnings that others have received in the same task, and (4) an announced anticipated individual payoff level, best describes decisions in a decontextualized risky decision making task. We find heterogeneity among individuals in the reference points they employ. The population average payoff level is the modal reference point, followed by experimenter's stated expectation of a participant's individual earnings, followed in turn by the average earnings of other participants in previous sessions of the same experiment. A sizeable share of individuals show multiple reference points simultaneously. The reference point that best fits the choices of the individual is not affected by a shock to her income. PMID:27672374

  7. Creating a Reference Toolbox.

    ERIC Educational Resources Information Center

    Scott, Jane

    1997-01-01

    To help students understand that references are tools used to locate specific information, one librarian has her third-grade students create their own reference toolboxes as she introduces dictionaries, atlases, encyclopedias, and thesauri. Presents a lesson plan to introduce print and nonprint thesauri to third and fourth graders and includes a…

  8. Art Reference, SCLS 1972.

    ERIC Educational Resources Information Center

    Lukac, Milan

    To help librarians answer patrons' questions about art works, especially paintings, the procedures followed by the reference division of the Suffolk Cooperative Library System are outlined, and a list of reference materials is suggested. Topics covered include biographical information about artists, identification of paintings, location of…

  9. Rethinking Virtual Reference

    ERIC Educational Resources Information Center

    Tenopir, Carol

    2004-01-01

    Virtual reference services seem a natural extension of libraries digital collections and the emphasis on access to the library anytime, anywhere. If patrons use the library from home, it makes sense to provide them with person-to-person online reference. The Library of Congress (LC), OCLC, and several large library systems have developed and…

  10. China Connections Reference Book.

    ERIC Educational Resources Information Center

    Kalat, Marie B.; Hoermann, Elizabeth F.

    This reference book focuses on six aspects of the geography of the People's Republic of China. They are: territory, governing units, population and land use, waterways, land forms, and climates. Designed as a primary reference, the book explains how the Chinese people and their lifestyles are affected by China's geography. Special components…

  11. Reference Point Heterogeneity.

    PubMed

    Terzi, Ayse; Koedijk, Kees; Noussair, Charles N; Pownall, Rachel

    2016-01-01

    It is well-established that, when confronted with a decision to be taken under risk, individuals use reference payoff levels as important inputs. The purpose of this paper is to study which reference points characterize decisions in a setting in which there are several plausible reference levels of payoff. We report an experiment, in which we investigate which of four potential reference points: (1) a population average payoff level, (2) the announced expected payoff of peers in a similar decision situation, (3) a historical average level of earnings that others have received in the same task, and (4) an announced anticipated individual payoff level, best describes decisions in a decontextualized risky decision making task. We find heterogeneity among individuals in the reference points they employ. The population average payoff level is the modal reference point, followed by experimenter's stated expectation of a participant's individual earnings, followed in turn by the average earnings of other participants in previous sessions of the same experiment. A sizeable share of individuals show multiple reference points simultaneously. The reference point that best fits the choices of the individual is not affected by a shock to her income.

  12. Library Reference Service.

    ERIC Educational Resources Information Center

    Schippleck, Suzanne

    The Inglewood, California, public library provides a manual on reference service. The theory, purpose, and objectives of reference are noted, and goals and activities are described in terms of budget, personnel, resources, and services. A chapter on organization covers service structure, information services, relationships with other library…

  13. Evaluation of the Xpert vanA/vanB Assay Using Enriched Inoculated Broths for Direct Detection of vanB Vancomycin-Resistant Enterococci

    PubMed Central

    Arends, J. P.; Kampinga, G. A.; Ahmad, H. M.; Dijkhuizen, B.; van Barneveld, P.; Rossen, J. W. A.; Friedrich, A. W.

    2014-01-01

    Rapid and accurate detection of VRE (vancomycin-resistant enterococci) is required for adequate antimicrobial treatment and infection prevention measures. Previous studies using PCR for the detection of VRE, including Cepheid's Xpert vanA/vanB assay, reported accurate detection of vanA VRE; however, many false-positive results were found for vanB VRE. This is mainly due to nonenterococcal vanB genes, which can be found in the gut flora. Our goal was to optimize the rapid and accurate detection of vanB VRE and to improve the positive predictive value (PPV) by limiting false-positive results. We evaluated the use of the Xpert vanA/vanB assay on rectal swabs and on enriched inoculated broths for the detection of vanB VRE. By adjusting the cycle threshold (CT) cutoff value to ≤25 for positivity by PCR on enriched broths, the sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) were 96.9%, 100%, 100%, and 99.5% for vanB VRE, respectively. As shown in this study, CT values of ≤25 acquired from enriched broths can be considered true positive. For broths with CT values between 25 and 30, we recommend confirming the results by culture. CT values of >30 appeared to be true negative. In conclusion, this study shows that the Cepheid's Xpert vanA/vanB assay performed on enriched inoculated broths with an adjusted cutoff CT value is a useful and rapid tool for the detection of vanB VRE. PMID:25297325

  14. Identification of bacteria in blood culture broths using matrix-assisted laser desorption-ionization Sepsityper™ and time of flight mass spectrometry.

    PubMed

    Kok, Jen; Thomas, Lee C; Olma, Thomas; Chen, Sharon C A; Iredell, Jonathan R

    2011-01-01

    Matrix-assisted laser desorption ionization time of flight mass spectrometry (MALDI-TOF MS) is a novel method for the direct identification of bacteria from blood culture broths. We evaluate for the first time, the performance of the MALDI Sepsityper™ Kit and MS for the identification of bacteria compared to standard phenotypic methods using the manufacturer's specified bacterial identification criteria (spectral scores ≥1.700-1.999 and ≥2.000 indicated identification to genus and species level, respectively). Five hundred and seven positive blood culture broths were prospectively examined, of which 379 (74.8%; 358 monomicrobial, 21 polymicrobial) were identified by MALDI-TOF MS; 195 (100%) and 132 (67.7%) of 195 gram-positive; and 163 (100%) and 149 (91.4%) of 163 gram-negative organisms from monomicrobial blood cultures were correctly identified to genus and species level, respectively. Spectral scores <1.700 (no identification) were obtained in 128/507 (25.2%) positive blood culture broths, including 31.6% and 32.3% of gram-positive and polymicrobial blood cultures, respectively. Significantly more gram-negative organisms were identified compared to gram-positive organisms at species level (p<0.0001). Five blood cultures were misidentified, but at species level only; including four monomicrobial blood cultures with Streptococcus oralis/mitis that were misidentified as Streptococcus pneumoniae. Positive predictive values for the direct identification of both gram-positive and gram-negative bacteria from monomicrobial blood culture broths to genus level were 100%. A diagnostic algorithm for positive blood culture broths that incorporates gram staining and MALDI-TOF MS should identify the majority of pathogens, particularly to genus level.

  15. The subduction reference framework

    NASA Astrophysics Data System (ADS)

    Seton, M.; Müller, D.; Gurnis, M.; Flament, N.; Whittaker, J.

    2010-12-01

    Plate tectonic reconstructions are essential for determining the spatial and temporal context for geological and geophysical data and help distinguish competing models for regional plate kinematic histories and the relationships between tectonic features and events. Plate reconstructions, a series of relative plate motions anchored to an absolute reference frame via a plate circuit, can act as surface boundary constraints for mantle convection models, allowing us to link surface processes to the deep earth. One of the limitations in global plate motion models has been to accurately determine the positions of plates through time. Traditionally, this has been based on either palaeomagnetic or hotspot reference frames, however both these methodologies have some shortcomings. Palaeomagnetic reference frames can determine latitudes but not longitudes, with additional inaccuracies due to true polar wander. Hotspot reference frames can only be confidently tied back to about 130 Ma and there is evidence that mantle plumes have moved relative to each other. New “hybrid” reference frames are emerging, which consist of fixed or moving hotspot reference frames merged with true polar wander (TPW) corrected palaeomagnetic reference frames. We have devised a methodology to link plate reconstructions to mantle convection back to Pangaea breakup time to converge on a solution that correctly aligns slab material in the mantle to the locations of subduction zones in the past. We aim to construct a “Subduction Reference Frame” for plate motions since 200 Ma by iteratively matching forward geodynamic models with tomographically imaged slabs in the mantle. Our forward models involve coupling global plate kinematics, the thermal structure of the oceanic lithosphere and slab assimilation to a spherical mantle convection code, CitcomS. Preliminary results have been obtained for a plate motion model using a moving hotspot reference frame to 100 Ma and a TPW corrected reference frame

  16. EPA QUICK REFERENCE GUIDES

    EPA Science Inventory

    EPA Quick Reference Guides are compilations of information on chemical and biological terrorist agents. The information is presented in consistent format and includes agent characteristics, release scenarios, health and safety data, real-time field detection, effect levels, samp...

  17. Value of Information References

    DOE Data Explorer

    Morency, Christina

    2014-12-12

    This file contains a list of relevant references on value of information (VOI) in RIS format. VOI provides a quantitative analysis to evaluate the outcome of the combined technologies (seismology, hydrology, geodesy) used to monitor Brady's Geothermal Field.

  18. Genetics Home Reference

    MedlinePlus

    ... MENU Toggle navigation Home Page Search Share: Email Facebook Twitter Home Health Conditions Genes Chromosomes & mtDNA Resources Help Me Understand Genetics Genetics Home Reference provides consumer-friendly information about the effects of genetic variation ...

  19. Selecting a reference object.

    PubMed

    Miller, Jared E; Carlson, Laura A; Hill, Patrick L

    2011-07-01

    One way to describe the location of an object is to relate it to another object. Often there are many nearby objects, each of which could serve as a candidate to be the reference object. A common theoretical assumption is that features that make a given object salient relative to the candidate set are instrumental in determining which is selected. The current research tests this assumption, assessing the relative importance of spatial, perceptual, and functional-interactive features. Three experiments demonstrated that spatial features have the strongest influence on reference object selection, with the perceptual feature of color playing no significant role. Functional-interactive features were shown to be spatially dependent, having an influence only when the spatial configuration enabled an interaction between the located object and the reference object. These findings challenge the common perspective that salience in and of itself dictates reference object selection and argue for a reliance on spatial features.

  20. Civil engineering reference guide

    SciTech Connect

    Merritt, F.S.

    1986-01-01

    The civil engineering reference guide contains the following: Structural theory. Structural steel design. Concrete design and construction. Wood design and construction. Bridge engineering. Geotechnical engineering. Water engineering. Environmental engineering. Surveying.

  1. EPA QUICK REFERENCE GUIDES

    EPA Science Inventory

    EPA Quick Reference Guides are compilations of information on chemical and biological terrorist agents. The information is presented in consistent format and includes agent characteristics, release scenarios, health and safety data, real-time field detection, effect levels, samp...

  2. Genetics Home Reference: sialidosis

    MedlinePlus

    ... features. Sialidosis type I, also referred to as cherry-red spot myoclonus syndrome, is the less severe ... or night blindness. An eye abnormality called a cherry-red spot, which can be identified with an ...

  3. Enterprise Reference Library

    NASA Technical Reports Server (NTRS)

    Bickham, Grandin; Saile, Lynn; Havelka, Jacque; Fitts, Mary

    2011-01-01

    Introduction: Johnson Space Center (JSC) offers two extensive libraries that contain journals, research literature and electronic resources. Searching capabilities are available to those individuals residing onsite or through a librarian s search. Many individuals have rich collections of references, but no mechanisms to share reference libraries across researchers, projects, or directorates exist. Likewise, information regarding which references are provided to which individuals is not available, resulting in duplicate requests, redundant labor costs and associated copying fees. In addition, this tends to limit collaboration between colleagues and promotes the establishment of individual, unshared silos of information The Integrated Medical Model (IMM) team has utilized a centralized reference management tool during the development, test, and operational phases of this project. The Enterprise Reference Library project expands the capabilities developed for IMM to address the above issues and enhance collaboration across JSC. Method: After significant market analysis for a multi-user reference management tool, no available commercial tool was found to meet this need, so a software program was built around a commercial tool, Reference Manager 12 by The Thomson Corporation. A use case approach guided the requirements development phase. The premise of the design is that individuals use their own reference management software and export to SharePoint when their library is incorporated into the Enterprise Reference Library. This results in a searchable user-specific library application. An accompanying share folder will warehouse the electronic full-text articles, which allows the global user community to access full -text articles. Discussion: An enterprise reference library solution can provide a multidisciplinary collection of full text articles. This approach improves efficiency in obtaining and storing reference material while greatly reducing labor, purchasing and

  4. Precision displacement reference system

    DOEpatents

    Bieg, Lothar F.; Dubois, Robert R.; Strother, Jerry D.

    2000-02-22

    A precision displacement reference system is described, which enables real time accountability over the applied displacement feedback system to precision machine tools, positioning mechanisms, motion devices, and related operations. As independent measurements of tool location is taken by a displacement feedback system, a rotating reference disk compares feedback counts with performed motion. These measurements are compared to characterize and analyze real time mechanical and control performance during operation.

  5. Membrane reference electrode

    DOEpatents

    Redey, L.; Bloom, I.D.

    1988-01-21

    A reference electrode utilizes a small thin, flat membrane of a highly conductive glass placed on a small diameter insulator tube having a reference material inside in contact with an internal voltage lead. When the sensor is placed in a non-aqueous ionic electrolytic solution, the concentration difference across the glass membrane generates a low voltage signal in precise relationship to the concentration of the species to be measured, with high spatial resolution. 2 figs.

  6. USGS reference materials

    USGS Publications Warehouse

    ,

    1995-01-01

    Every year in the United States, millions of measurements are made on the chemical composition of items that affect us on a daily basis. Determining the accuracy of these measurements is based on the analysis of appropriate reference materials whose composition was previously determined through rigorous testing. Today, reference materials help us evaluate the composition of the food we eat, medicine we use, soil we grow our crops in, and hundreds of other products that affect our everyday lives.

  7. Membrane reference electrode

    DOEpatents

    Redey, Laszlo; Bloom, Ira D.

    1989-01-01

    A reference electrode utilizes a small thin, flat membrane of a highly conductive glass placed on a small diameter insulator tube having a reference material inside in contact with an internal voltage lead. When the sensor is placed in a non-aqueous ionic electrolytic solution, the concentration difference across the glass membrane generates a low voltage signal in precise relationship to the concentration of the species to be measured with high spatial resolution.

  8. Comparison of different options for harvest of a therapeutic protein product from high cell density yeast fermentation broth.

    PubMed

    Wang, Alice; Lewus, Rachael; Rathore, Anurag S

    2006-05-05

    Recovery of therapeutic protein from high cell density yeast fermentations at commercial scale is a challenging task. In this study, we investigate and compare three different harvest approaches, namely centrifugation followed by depth filtration, centrifugation followed by filter-aid enhanced depth filtration, and microfiltration. This is achieved by presenting a case study involving recovery of a therapeutic protein from Pichia pastoris fermentation broth. The focus of this study is on performance of the depth filtration and the microfiltration steps. The experimental data has been fitted to the conventional models for cake filtration to evaluate specific cake resistance and cake compressibility. In the case of microfiltration, the experimental data agrees well with flux predicted by shear induced diffusion model. It is shown that, under optimal conditions, all three options can deliver the desired product recovery ( >80%), harvest time ( <15 h including sequential concentration/diafiltration step), and clarification ( <6 NTU). However, the three options differ in terms of process development time required, capital cost, consumable cost, ease of scale-ability and process robustness. It is recommended that these be kept under consideration when making a final decision on a harvesting approach.

  9. Metabolic Labeling to Characterize the Overall Composition of Francisella Lipid A and Lipopolysaccharide Grown in Broth and in Human Phagocytes

    PubMed Central

    Barker, Jason H.; Kaufman, Justin W.; Zhang, DeSheng; Weiss, Jerrold P.

    2013-01-01

    SUMMARY A hallmark of Francisella tularensis, a highly virulent Gram-negative bacterium, is its unusual lipopolysaccharide (LPS). Detailed analyses of Francisella LPS have revealed both structural heterogeneity and characteristics that may contribute to innate immune evasion. However, none of the methods yet employed has been sufficient to determine the overall LPS composition of Francisella. We now demonstrate that metabolic labeling of Francisellae with [14C]acetate, combined with fractionation of [14C]acetate-labeled lipids by ethanol precipitation rather than hot phenol-water extraction, permits a more sensitive and quantitative appraisal of overall compositional heterogeneity in lipid A and LPS. We show that the majority of lipid A of different Francisellae strains grown in diverse bacteriologic media and within human phagocytes accumulated as very hydrophobic species, including free lipid A, with < 10% of the lipid A molecules substituted with O-antigen polysaccharides. The spectrum of lipid A and LPS species varied in both a medium-dependent and strain-dependent fashion, and intracellular growth in THP-1 cells yielded lipid A species that were not present in the same bacteria grown in brain heart infusion broth. In summary, metabolic labeling with [14C]acetate greatly facilitates assessment of the effect of genotypic and/or environmental variables on the synthesis and net accumulation of lipid A and LPS by Francisella, including during growth within the cytosol of infected host cells. PMID:23729477

  10. A refrigeration temperature of 4 degrees C does not prevent static growth of Yersinia pestis in heart infusion broth.

    PubMed

    Torosian, Stephen D; Regan, Patrick M; Doran, Tara; Taylor, Michael A; Margolin, Aaron

    2009-09-01

    Multiple barriers such as inspections, testing, and proper storage conditions are used to minimize the risk of contaminated food. Knowledge of which barriers, such as refrigeration, are effective in preventing pathogen growth and persistence, can help direct the focus of efforts during food sampling. In this study, the doubling times were evaluated for 10 strains of Yersinia pestis of different genetic background cultured in heart infusion broth (HIB) kept at 4 degrees C +/- 1 degrees C under static conditions. Nine out of the 10 strains were able to grow at 4 degrees C +/- 1 degrees C. Apparent doubling times for 7 of the strains ranged from 41 to 50 h. Strain Harbin and strain D1 had apparent doubling times of 65 and 35 h, respectively, and strain O19 Ca-6 did not grow at all. Analysis of variance showed that the averaged growth data (colony forming units per mL) between strains that grew were not significantly different. The data presented here demonstrate that refrigeration alone is not an effective barrier to prevent static growth of Y. pestis in HIB. These findings provide the preliminary impetus to investigate Y. pestis growth in a variety of food matrices that may provide a similar environment as HIB.

  11. Attenuation of TNF-α secretion by L-proline-based cyclic dipeptides produced by culture broth of Pseudomonas aeruginosa.

    PubMed

    Khan, Rukaiyya; Basha, Ameer; Goverdhanam, Ragavendra; Rao, Poorna Chandra; Tanemura, Yuhei; Fujimoto, Yoshinori; Begum, Ahil Sajeli

    2015-12-15

    To identify small molecule inhibitors of TNF-α, bioassay- and LC-MS-guided chemical investigation on EtOAc extract of Pseudomonas aeruginosa ABS-36 culture broth (EEPA) was performed, which yielded four proline-based cyclic dipeptides, cyclo(Gly-l-Pro) (1), cyclo(l-Pro-l-Phe) (2), cyclo(trans-4-hydroxy-l-Pro-l-Phe) (3) and cyclo(trans-4-hydroxy-l-Pro-l-Leu) (4). Compounds 1 and 3 exhibited potent inhibition of TNF-α release with IC50 values of 4.5 and 14.2μg/mL, respectively, while EEPA showed IC50 of 38.8μg/mL under lipopolysaccharide treated RAW 264.7 cell ELISA assay. Also, marked attenuation of mRNA-expression of TNF-α was shown by all compounds. In vivo testing in rats of EEPA and chemically synthesized 4 validated significant TNF-α reduction with 51% (500mg/kg) and 79% (50mg/kg), respectively. In addition, all compounds exhibited significant diminution of IL-1β and IL-6 mRNA-expression levels and NO production. All samples displayed only weak toxicity to lipopolysaccharide-induced RAW 264.7 cells.

  12. Effects of the origins of Botrytis cinerea on earthy aromas from grape broth media further inoculated with Penicillium expansum.

    PubMed

    Morales-Valle, H; Silva, L C; Paterson, R R M; Venâncio, A; Lima, N

    2011-08-01

    Earthy "off" aromas from wine and grape juice are highly detrimental to the production of quality grape products. These volatile compounds are produced on grapes by Botrytis cinerea, Penicillium expansum and/or a combination of P. expansum and B. cinerea strains. B. cinerea strains were isolated from different (a) vineyards in Spain and Portugal, (b) grape varieties (c) bunches (i.e., sound and botrytized) and (d) positions in the botrytized bunch (i.e., interior or exterior). A novel Headspace-Phase Microextraction (SPME) followed by Gas Chromatrography/Mass Spectrometry (GC-MS) dedicated to analyze geosmin, methylisoborneol (MIB), 1-octen-3-ol, fenchone and fenchol in grape broth medium was used. Approximately 50% of the B. cinerea strains induced detectable geosmin. One strain accumulated significant amounts of anisoles, demonstrating that this contamination might already occur in the vineyard. Strains from the interior of Cainho grape bunches induced more geosmin and hence it may be possible to reduce this volatile in wine by avoiding using these grapes in case of B. cinerea attack. Copyright © 2011 Elsevier Ltd. All rights reserved.

  13. Two-stage pervaporation process for effective in situ removal acetone-butanol-ethanol from fermentation broth.

    PubMed

    Cai, Di; Hu, Song; Miao, Qi; Chen, Changjing; Chen, Huidong; Zhang, Changwei; Li, Ping; Qin, Peiyong; Tan, Tianwei

    2017-01-01

    Two-stage pervaporation for ABE recovery from fermentation broth was studied to reduce the energy cost. The permeate after the first stage in situ pervaporation system was further used as the feedstock in the second stage of pervaporation unit using the same PDMS/PVDF membrane. A total 782.5g/L of ABE (304.56g/L of acetone, 451.98g/L of butanol and 25.97g/L of ethanol) was achieved in the second stage permeate, while the overall acetone, butanol and ethanol separation factors were: 70.7-89.73, 70.48-84.74 and 9.05-13.58, respectively. Furthermore, the theoretical evaporation energy requirement for ABE separation in the consolidate fermentation, which containing two-stage pervaporation and the following distillation process, was estimated less than ∼13.2MJ/kg-butanol. The required evaporation energy was only 36.7% of the energy content of butanol. The novel two-stage pervaporation process was effective in increasing ABE production and reducing energy consumption of the solvents separation system. Copyright © 2016 Elsevier Ltd. All rights reserved.

  14. Hemolytic and proteolytic activities of Aeromonas hydrophila and Aeromonas veronii biovar sobria in broth and salmon extract at different temperatures.

    PubMed

    González-Rodríguez, María-Nieves; Santos, Jesús A; Otero, Andrés; García-López, Maria-Luisa

    2004-02-01

    Expression of hemolytic and proteolytic activities throughout the growth cycle was investigated with two enterotoxic aeromonad strains assigned to the species Aeromonas hydrophila and Aeromonas veronii biovar sobria. Although growth kinetic data were dependent on strain, temperature, and substrate, maximum populations attained were higher than 9 log CFU/ml in aerated tryptone soya broth plus yeast extract (TSBYE) and salmon extract within the range 4 to 28 degrees C. For both strains in TSBYE, variable amounts of hemolytic activity were first detected at any temperature when aeromonad counts were over 9 log CFU/ml. Afterwards, this activity increased up to similar levels (109 to 112 hemolytic units per ml) without a significant increase in populations. Salmon extract supported hemolysin synthesis at 28 but not 4 degrees C. Proteolytic activity of the A. hydrophila strain was only expressed in salmon extract at 28 degrees C, whereas A. veronii biovar sobria did at 28 degrees C in both substrates and at 10 degrees C in TSBYE.

  15. The use of Macronet resins to recover gamma-decalactone produced by Rhodotorula aurantiaca from the culture broth.

    PubMed

    Alchihab, Mohamed; Aldric, Jean-Marc; Aguedo, Mario; Destain, Jacqueline; Wathelet, Jean-Paul; Thonart, Philippe

    2010-02-01

    During the biotransformation of castor oil into gamma-decalactone, R. aurantiaca produced both the lactone form and its precursor (4-hydroxydecanoic acid). After six days of culture, a maximum yield of gamma-decalactone of 6.5 g/l was obtained. The parameters of gamma-decalactone adsorption on three Macronet resins (MN-202, MN-102 and MN-100) were investigated in water. Adsorption isotherms of gamma-decalactone for the three Macronet resins were linear. The trapping of gamma-decalactone produced by R. aurantiaca on these resins was then carried out. gamma-Decalactone was effectively retained by all the studied Macronet resins. The resin MN-202 trapped gamma-decalactone more efficiently than MN-102 and MN-100. The percentages of gamma-decalactone adsorbed on the resins MN-202, MN-102 and MN-100 were, respectively, 85, 75 and 81%, whereas around 70% of the adsorbed gamma-decalactone was then desorbed. We propose an industrial process that uses Macronet resins to extract gamma-decalactone from culture broth of R. aurantiaca.

  16. A Laccase with HIV-1 Reverse Transcriptase Inhibitory Activity from the Broth of Mycelial Culture of the Mushroom Lentinus tigrinus

    PubMed Central

    Xu, LiJing; Wang, HeXiang; Ng, TziBun

    2012-01-01

    A 59 kDa laccase with inhibitory activity against HIV-1 reverse transcriptase (IC50 = 2.4 μM) was isolated from the broth of mycelial culture of the mushroom Lentinus tigrinus. The isolation procedure involved ion exchange chromatography on DEAE-cellulose and CM-cellulose, and gel filtration by fast protein liquid chromatography on Superdex 75. The laccase was adsorbed on both types of ion exchangers. About 95-fold purification was achieved with a 25.9% yield of the enzyme. The procedure resulted in a specific enzyme activity of 76.6 U/mg. Its N-terminal amino acid sequence was GIPDLHDLTV, which showed little similarity to other mushroom laccase and other Lentinus tigrinus strain laccase. Its characteristics were different from previously reported laccase of other Lentinus tigrinus strain. Maximal laccase activity was observed at a pH of 4 and at a temperature of 60°C, respectively. This study yielded the information about the potentially exploitable activities of Lentinus tigrinus laccase. PMID:22536022

  17. Analgesic and anti-inflammatory effects of the dry matter of culture broth of Termitomyces albuminosus and its extracts.

    PubMed

    Lu, Yi-Yu; Ao, Zong-Hua; Lu, Zhen-Ming; Xu, Hong-Yu; Zhang, Xiao-Mei; Dou, Wen-Fang; Xu, Zheng-Hong

    2008-12-08

    The objectives of this study were to investigate the analgesic and anti-inflammatory effects of the dry matter of culture broth (DMCB) of Termitomyces albuminosus in submerged culture and its crude saponin extract (CSE) and crude polysaccharide extract (CPE). The analgesic effects of DMCB, CSE and CPE were evaluated with models of acetic acid-induced writhing response and formalin test in mouse. The anti-inflammatory effects of DMCB, CSE and CPE were evaluated by using models of xylene-induced mouse ear swelling and carrageen-induced mouse paw edema. The DMCB, CSE and CPE significantly decreased the acetic acid-induced writhing response and the licking time on the late phase in the formalin test. Treatment of DMCB (1000mg/kg), CSE (200mg/kg) or CPE (200mg/kg) inhibited the mouse ear swelling by 61.8%, 79.0% and 81.6%, respectively. In the carrageen-induced mouse paw edema test, the group treated with indomethacin showed the strongest inhibition of edema formation by 77.8% in the third hour after carrageenan administration, while DMCB (1000mg/kg), CSE (200mg/kg) and CPE (200mg/kg) showed 48.4%, 55.6% and 40.5%, respectively. The results suggested that DMCB of Termitomyces albuminosus possessed the analgesic and anti-inflammatory activities. Saponins and polysaccharides were proposed to be the major active constituents of Termitomyces albuminosus in submerged culture.

  18. Biofilm formation by Staphylococcus aureus from food contact surfaces in a meat-based broth and sensitivity to sanitizers.

    PubMed

    de Souza, Evandro Leite; Meira, Quênia Gramile Silva; de Medeiros Barbosa, Isabella; Athayde, Ana Júlia Alves Aguiar; da Conceição, Maria Lúcia; de Siqueira Júnior, José Pinto

    2014-01-01

    This study assessed the capacity of adhesion, the detachment kinetic and the biofilm formation by Staphylococcus aureus isolated from food services on stainless steel and polypropylene surfaces (2 × 2 cm) when cultivated in a meat-based broth at 28 and 7 °C. It was also to study the efficacy of the sanitizers sodium hypochlorite (250 mg/L) and peracetic acid (30 mg/L) in inactivating the bacterial cells in the preformed biofilm. S. aureus strains adhered in high numbers regardless the assayed surface kind and incubation temperature over 72 h. Cells detachment of surfaces revealed high persistence over the incubation period. Number of cells needed for biofilm formation was noted at all experimental systems already after 3 days. Peracetic acid and sodium hypochlorite were not efficient in completely removing the cells of S. aureus adhered on polypropylene and stainless steel surfaces. From these results, the assayed strains revealed high capacity to adhere and form biofilm on polypropylene and stainless steel surfaces under different growth conditions. Moreover, the cells in biofilm matrix were resistant for total removal when submitted to the exposure to sanitizers.

  19. Biofilm formation by Staphylococcus aureus from food contact surfaces in a meat-based broth and sensitivity to sanitizers

    PubMed Central

    de Souza, Evandro Leite; Meira, Quênia Gramile Silva; de Medeiros Barbosa, Isabella; Athayde, Ana Júlia Alves Aguiar; da Conceição, Maria Lúcia; de Siqueira Júnior, José Pinto

    2014-01-01

    This study assessed the capacity of adhesion, the detachment kinetic and the biofilm formation by Staphylococcus aureus isolated from food services on stainless steel and polypropylene surfaces (2 × 2 cm) when cultivated in a meat-based broth at 28 and 7 °C. It was also to study the efficacy of the sanitizers sodium hypochlorite (250 mg/L) and peracetic acid (30 mg/L) in inactivating the bacterial cells in the preformed biofilm. S. aureus strains adhered in high numbers regardless the assayed surface kind and incubation temperature over 72 h. Cells detachment of surfaces revealed high persistence over the incubation period. Number of cells needed for biofilm formation was noted at all experimental systems already after 3 days. Peracetic acid and sodium hypochlorite were not efficient in completely removing the cells of S. aureus adhered on polypropylene and stainless steel surfaces. From these results, the assayed strains revealed high capacity to adhere and form biofilm on polypropylene and stainless steel surfaces under different growth conditions. Moreover, the cells in biofilm matrix were resistant for total removal when submitted to the exposure to sanitizers. PMID:24948915

  20. Evolution of culture broth rheological properties during propagation of the entomopathogenic nematode Steinernema carpocapsae, in submerged monoxenic culture.

    PubMed

    Chavarría-Hernández, Norberto; Rodríguez-Hernández, Adriana Inés; Pérez-Guevara, Fermín; de la Torre, Mayra

    2003-01-01

    This article presents the evolution of culture broth rheological properties during monoxenic cultures of Steinernema carpocapsae in cylindrical bottles agitated orbitally. Rheological properties were evaluated in simple-shear flow conditions and were well-modeled by the Ostwald-de Waele model. Rheological properties varied from slightly dilatant, n = 1.2 (-), to moderately pseudoplastic flow behavior, n = 0.6 (-). Nematode concentrations increased from 750 +/- 190 to 130 900 +/- 6900 nematodes/mL, and the apparent viscosity (eta(a)) evolved from 4.5 +/- 0.7 to 46.6 +/- 3.2 mPa.s during the fermentations. Rheological behavior did not appear to be strongly influenced by nematode number and/or its stage of development; however, the release of substances from the decomposition of nematode cadavers appeared to be of great importance. Among the different developmental stages of the nematodes, only juveniles of the first stage (J1) were highly susceptible to the shearing conditions tested (shear stress, tau(r)()(theta), from 0.9 to 3.5 Pa during periods of 80-100 min), resulting in the viability loss of 85% of J1 nematodes.

  1. Preparative separation of six antimycin A components from antimycin fermentation broth by high-speed counter-current chromatography.

    PubMed

    Wang, Jiaoyan; Wen, Yaoming; Chen, Xiuming; Lin, Yuhan; Zhou, Jingming; Xie, Ying; Wang, Huaiwang; Jiang, Hong; Zheng, Wei

    2010-09-03

    A method of using high-speed counter-current chromatography (HSCCC) was established for preparative isolation and purification of antimycin A components from antimycin fermentation broth. Six antimycin A components were successfully purified for the first time by HSCCC with a two-phase solvent system composed of n-hexane-ethyl acetate-methanol-water (5:2:4:1, by volume). Total of 20mg antimycin A(4)(a or b), 25mg antimycin A(3)(a or b), 21mg antimycin A(8)(a or b), 34mg antimycin A(2)(a or b), 26mg antimycin A(1)(a or b) and 34mg antimycin A(1)(a or b) with the purities of 93.2, 98.6, 96.2, 94.1, 94.9 and 96.7%, respectively, determined by high-performance liquid chromatography (HPLC), were yielded from 200mg crude sample only in one HSCCC run. 2010 Elsevier B.V. All rights reserved.

  2. HPLC-fluorescence detection method for determination of key intermediates of the lincomycin biosynthesis in fermentation broth.

    PubMed

    Kameník, Zdenek; Kopecký, Jan; Marecková, Markéta; Ulanová, Dana; Novotná, Jitka; Pospísil, Stanislav; Olsovská, Jana

    2009-03-01

    The biosynthetic pathway of the clinically important antibiotic lincomycin is not known in details. The precise knowledge of the lincomycin biosynthesis is a prerequisite for generation of improved derivatives by means of combinatorial genetics. Methods allowing determination of the key intermediates are very important tools of the pathway investigation. Two new high-performance liquid chromatography methods with fluorescence detection for determination of lincomycin precursors in fermentation broth of Streptomyces lincolnensis and its lincomycin nonproducing mutants were developed. The first one enables simultaneous analysis of methylthiolincosamide (MTL) and N-demethyllincomycin (NDL), whereas the second one is suitable for 4-propyl-L-proline (PPL) assay. Both methods are based on the pre-column derivatization: MTL and NDL with 4-chloro-7-nitrobenzofurazan; PPL with o-phthaldialdehyde. The methods were validated with lower limit of quantification values of 2.50, 3.75, and 3.75 microg ml(-1) for MTL, NDL, and PPL, respectively. The inter- and intra-day accuracies and precisions were all within 12%. Stability of oxidized and derivatized analytes was investigated.

  3. Broth and agar hop-gradient plates used to evaluate the beer-spoilage potential of Lactobacillus and Pediococcus isolates.

    PubMed

    Haakensen, M; Schubert, A; Ziola, B

    2009-03-15

    Identification of the beer-spoilage Lactobacillus and Pediococcus bacteria has largely taken two approaches; identification of spoilage-associated genes or identification of specific species of bacteria regardless of ability to grow in beer. The problem with these two approaches is that they are either overly inclusive (i.e., detect all bacteria of a given species regardless of spoilage potential) or overly selective (i.e., rely upon individual, putative spoilage-associated genes). Our goal was to design a method to assess the ability of Lactobacillus and Pediococcus to spoil beer that is independent of speciation or genetic background. In searching for a method by which to differentiate between beer-spoilage bacteria and bacteria that cannot grow in beer, we explored the ability of lactobacilli and pediococci isolates to grow in the presence of varying concentrations of hop-compounds and ethanol in broth medium versus on agar medium. The best method for differentiating between bacteria that can grow in beer and bacteria that do not pose a threat as beer-spoilage organisms was found to be a hop-gradient agar plate containing ethanol. This hop-gradient agar plate technique provides a rapid and simple solution to the dilemma of assessing the ability of Lactobacillus and Pediococcus isolates to grow in beer, and provides new insights into the different strategies used by these bacteria to survive under the stringent conditions of beer.

  4. Reference Man anatomical model

    SciTech Connect

    Cristy, M.

    1994-10-01

    The 70-kg Standard Man or Reference Man has been used in physiological models since at least the 1920s to represent adult males. It came into use in radiation protection in the late 1940s and was developed extensively during the 1950s and used by the International Commission on Radiological Protection (ICRP) in its Publication 2 in 1959. The current Reference Man for Purposes of Radiation Protection is a monumental book published in 1975 by the ICRP as ICRP Publication 23. It has a wealth of information useful for radiation dosimetry, including anatomical and physiological data, gross and elemental composition of the body and organs and tissues of the body. The anatomical data includes specified reference values for an adult male and an adult female. Other reference values are primarily for the adult male. The anatomical data include much data on fetuses and children, although reference values are not established. There is an ICRP task group currently working on revising selected parts of the Reference Man document.

  5. Validation study to demonstrate the equivalence of a minor modification (TECRA ULTIMA protocol) to AOAC Method 998.09 (TECRA Salmonella Visual Immunoassay) with the cultural reference method.

    PubMed

    Briggs, Julie; Dailianis, Angela; Hughes, Denise; Garthwaite, Ian

    2004-01-01

    The TECRA Salmonella Visual Immunoassay (VIA) using Rappaport-Vassiliadis RV[R10] as a single selective enrichment broth has Final Action approval (AOAC Method 998.09). TECRA has recently developed a protocol (TECRA ULTIMA), which involves the addition of a new additive to a 1 mL aliquot of the RV[R10] broth, prior to the heat-killing step, thereby allowing the RV[R10] broth to be tested directly in the kit and thus eliminating the need for the 2 h post-enrichment in M broth. An in-house validation study was conducted to compare the modified AOAC Method 998.09 to the reference culture method. Three foods were used in the study: Naturally contaminated raw ground poultry at high (10-50 cells/25 g), and low (1-5 cells/25 g) levels; and milk powder and peanut butter, artificially inoculated at low and high levels with Salmonella bovismorbificans and S. enterica Mbandaka, respectively. Twenty test portions were analyzed for each level with 10 uninoculated control samples per food. Overall, no significant differences (p <0.05) were observed when the proportion of positive test portions for the modified VIA were compared with that for the reference method. This minor modification, which employs the additive (provided in the TECRA ULTIMA SALMONELLA Test Kit) to permit the direct analysis of RV[R10] broth has demonstrated the utility of the TECRA ULTIMA SALMONELLA protocol. It is recommended that the minor modification to Method 998.09 be approved First Action as an additional option within the method.

  6. Every Reference Librarian a Reviewer.

    ERIC Educational Resources Information Center

    Rettig, James

    1987-01-01

    Discusses the need for reference librarians to learn reference materials well enough to be able to review materials and correctly refer patrons to information sources. The review process is outlined and some evaluation of current reference services is provided. (CLB)

  7. Improved liquid-chromatography tandem mass spectrometry method for the determination of the bioactive dipeptides, carnosine and anserine: application to analysis in chicken broth.

    PubMed

    Macià, Alba; Motilva, Maria-José; Romero, Maria-Paz; Labrador, Agustín; Domínguez, Alba; Peiro, Lluís

    2012-05-15

    An improved method, based on ultra-performance liquid chromatography (UPLC) coupled to tandem mass spectrometry (MS/MS), has been developed to determine the bioactive dipeptides carnosine (CAR) and anserine (ANS) in chicken broth. These analytes are hydrophilic (polar) and in order to improve their retention, the chromatographic mode used was hydrophilic interaction chromatography (HILIC) (1.7 μm particle size). In order to remove the salt before the chromatographic analysis of the chicken broth (0.8%, w/w), an exhaustive sample pre-treatment strategy was necessary since the salt is not volatilized and could block the ionization source and lead to signal suppression. The chicken broth was firstly centrifuged to remove the fat and chicken proteins, and then was pretreated by off-line solid-phase extraction (SPE), using traditional cartridges, or off-line μElution plate (μSPE), using microplates, and the results were compared. Due to the high polar character of the dipeptides studied and the sample matrix, these compounds were not retained in the sorbent hydrophilic-lipophilic balanced (HLB) and were eluted in the load step, whereas the salt was retained in the sorbent. This fact was observed by the addition of silver nitrate in the chicken broth extract, where before the SPE or μSPE a white precipitate (silver chloride) was formed and after the SPE or μSPE this precipitate was not observed. By using these sample pre-treatment strategies, the extraction recoveries were higher than 80%, and the matrix effect was lower than 12%. Once the improved method was developed, the quality parameters of the method were studied. The LODs and LOQs of the CAR and ANS were lower than 6 and 1.8 μg/l, respectively. Then, the method was applied to analyse a commercial chicken broth. This improved method allowed determining CAR and ANS between 6 and 10mg dipeptide/l chicken broth in 10 min (sample pre-treatment and chromatographic analysis). Therefore, the proposed improved method

  8. In Vitro Activity of Ceftazidime-Avibactam against Contemporary Pseudomonas aeruginosa Isolates from U.S. Medical Centers by Census Region, 2014.

    PubMed

    Huband, Michael D; Castanheira, Mariana; Flamm, Robert K; Farrell, David J; Jones, Ronald N; Sader, Helio S

    2016-04-01

    Thein vitroantibacterial activities of ceftazidime-avibactam and comparator agents were evaluated using reference broth microdilution methods against 1,743Pseudomonas aeruginosaisolates collected in 2014 from 69 U.S. medical centers, representing each of the nine census regions. Ceftazidime-avibactam demonstrated potent activity againstP. aeruginosa, including many isolates not susceptible to ceftazidime, meropenem, and piperacillin-tazobactam. In each of the nine census regions, ceftazidime-avibactam demonstrated the highest percentage of susceptible isolates.

  9. Setting reference targets

    SciTech Connect

    Ruland, R.E.

    1997-04-01

    Reference Targets are used to represent virtual quantities like the magnetic axis of a magnet or the definition of a coordinate system. To explain the function of reference targets in the sequence of the alignment process, this paper will first briefly discuss the geometry of the trajectory design space and of the surveying space, then continue with an overview of a typical alignment process. This is followed by a discussion on magnet fiducialization. While the magnetic measurement methods to determine the magnetic centerline are only listed (they will be discussed in detail in a subsequent talk), emphasis is given to the optical/mechanical methods and to the task of transferring the centerline position to reference targets.

  10. [Determination of lactic acid in fermentation broth of Rhizopus oryzae by reversed-phase high performance liquid chromatography (RP-HPLC)].

    PubMed

    Bai, D M; Ban, R; Zhao, X M; Hu, Z D

    2000-11-01

    A method for determining lactic acid in fermentation broth of Rhizopus oryzae by RP-HPLC is described. The operating conditions were Wakosil-II 5 C18 RS column(4.6 mm i.d. x 150 mm, 5 microns) at room temperature, 0.01 mol/L phosphoric acid solution (pH 2.5) as mobile phase with a flow rate of 1.0 mL/min and UV detection at 210 nm. The retention time of lactic acid was 3.820 min. This method is simple, rapid and accurate. The results will not be affected by other components in the broth. The relative standard deviation was 0.22% (n = 5), and the recovery was over 99%.

  11. Maximizing neonatal early onset group B streptococcal disease prevention with universal culture screening at 35 to 37 weeks gestation: a comparison of GBS detection rates between LIM broth and CNA culture media.

    PubMed

    Orsello, Christopher; Dommermuth, Ronald

    2003-06-01

    Group B streptococcal (GBS) disease is the most common cause of early onset neonatal sepsis. The Centers for Disease Control (CDC) recommends performing recto-vaginal cultures on pregnant woman to detect GBS, followed by treatment of women with positive cultures. Our facility adopted selective culture screening in 1997 using a colistin-naladixic acid (CNA) plate media instead of the more expensive LIM broth media. CNA plate cultures cost one third that of LIM broth and allow for final results in 24 hours, versus 48-72 hours with LIM broth. We hypothesized that CNA media saves time, money, and detects GBS as effectively as LIM broth. This study determined which media is superior at detecting recto-vaginal GBS. This was a case-control study involving 152 consecutive pregnant patients at 35-37 weeks from August 1 to October 1, 2001, at Naval Hospital Bremerton, Wash. We obtained two recto-vaginal swabs from each patient. One was cultured in LIM broth and the other on CNA medium. We then compared differences in the rates of positive cultures with LIM broth versus CNA medium using chi-square statistics and calculation of odds ratios (OR). LIM broth detected GBS in 35 of 145 (24.19%) women versus 21 of 145 (14.5%) using CNA. CNA failed to detect GBS in 15 cases in which LIM broth succeeded (OR=1.88; 95% CI=1.03-3.4). LIM broth is superior at detecting maternal GBS colonization and is recommended over CNA plate to maximize prevention of early onset neonatal GBS disease.

  12. Aluminum reference electrode

    DOEpatents

    Sadoway, D.R.

    1988-08-16

    A stable reference electrode is described for use in monitoring and controlling the process of electrolytic reduction of a metal. In the case of Hall cell reduction of aluminum, the reference electrode comprises a pool of molten aluminum and a solution of molten cryolite, Na[sub 3]AlF[sub 6], wherein the electrical connection to the molten aluminum does not contact the highly corrosive molten salt solution. This is accomplished by altering the density of either the aluminum (decreasing the density) or the electrolyte (increasing the density) so that the aluminum floats on top of the molten salt solution. 1 fig.

  13. Aluminum reference electrode

    DOEpatents

    Sadoway, Donald R.

    1988-01-01

    A stable reference electrode for use in monitoring and controlling the process of electrolytic reduction of a metal. In the case of Hall cell reduction of aluminum, the reference electrode comprises a pool of molten aluminum and a solution of molten cryolite, Na.sub.3 AlF.sub.6, wherein the electrical connection to the molten aluminum does not contact the highly corrosive molten salt solution. This is accomplished by altering the density of either the aluminum (decreasing the density) or the electrolyte (increasing the density) so that the aluminum floats on top of the molten salt solution.

  14. IERS Reference System.

    NASA Astrophysics Data System (ADS)

    Yokoyama, K.

    Present circumstances related to IERS activities are described from various points of view. The NASA Dynamics of Solid Earth (DOSE) program and the IERS intensive campaign proposed by J. Dickey of JPL are particularly interesting. It is important to implement international cooperation to establish a fundamental radio reference frame by carrying out global solution based on all geodetic observations, past and future. A precession and nutation model may be determined observationally with an accuracy of 0.2 - 0.3 mas in a few years. Then it will become possible to establish the radio reference frame with this accuracy.

  15. NASCAP programmer's reference manual

    NASA Technical Reports Server (NTRS)

    Mandell, M. J.; Stannard, P. R.; Katz, I.

    1993-01-01

    The NASA Charging Analyzer Program (NASCAP) is a computer program designed to model the electrostatic charging of complicated three-dimensional objects, both in a test tank and at geosynchronous altitudes. This document is a programmer's reference manual and user's guide. It is designed as a reference to experienced users of the code, as well as an introduction to its use for beginners. All of the many capabilities of NASCAP are covered in detail, together with examples of their use. These include the definition of objects, plasma environments, potential calculations, particle emission and detection simulations, and charging analysis.

  16. Multifunctional reference electrode

    DOEpatents

    Redey, L.; Vissers, D.R.

    1981-12-30

    A multifunctional, low mass reference electrode of a nickel tube, thermocouple means inside the nickel tube electrically insulated therefrom for measuring the temperature thereof, a housing surrounding the nickel tube, an electrolyte having a fixed sulfide ion activity between the housing and the outer surface of the nickel tube forming the nickel/nickel sulfide/sulfide half-cell are described. An ion diffusion barrier is associated with the housing in contact with the electrolyte. Also disclosed is a cell using the reference electrode to measure characteristics of a working electrode.

  17. Multifunctional reference electrode

    DOEpatents

    Redey, Laszlo; Vissers, Donald R.

    1983-01-01

    A multifunctional, low mass reference electrode of a nickel tube, thermocouple means inside the nickel tube electrically insulated therefrom for measuring the temperature thereof, a housing surrounding the nickel tube, an electrolyte having a fixed sulfide ion activity between the housing and the outer surface of the nickel tube forming the nickel/nickel sulfide/sulfide half-cell. An ion diffusion barrier is associated with the housing in contact with the electrolyte. Also disclosed is a cell using the reference electrode to measure characteristics of a working electrode.

  18. Determination of carbohydrates, sugar alcohols, and glycols in cell cultures and fermentation broths using high-performance anion-exchange chromatography with pulsed amperometric detection.

    PubMed

    Hanko, V P; Rohrer, J S

    2000-08-01

    Cell cultures and fermentation broths are complex mixtures of organic and inorganic compounds. Many of these compounds are synthesized or metabolized by microorganisms, and their concentrations can impact the yields of desired products. Carbohydrates serve as carbon sources for many microorganisms, while sugar alcohols (alditols), glycols (glycerol), and alcohols (methanol and ethanol) are metabolic products. We used high-performance anion-exchange chromatography with pulsed amperometric detection (HPAE-PAD) to simultaneously analyze for carbohydrates, alditols, and glycerol in growing yeast (Saccharomyces cerevisiae) cultures and their final fermentation broths. Both cultures were grown on complex undefined media, aliquots centrifuged to remove particulates, and the supernatants diluted and directly injected for analysis. Pulsed amperometry allowed a direct detection of the carbohydrates, alditols, and glycols present in the cultures and fermentation broths with very little interference from other matrix components. The broad linear range of three to four orders of magnitude allowed samples to be analyzed without multiple dilutions. Peak area RSDs were 2-7% for 2, 3-butanediol, ethanol, glycerol, erythritol, rhamnose, arabitol, sorbitol, galactitol, mannitol, arabinose, glucose, galactose, lactose, ribose, raffinose, and maltose spiked into a heat-inactivated yeast culture broth supernatant that was analyzed repetitively for 48 h. This method is useful for directly monitoring culture changes during fermentation. The carbohydrates in yeast cultures were monitored over 1 day. A yeast culture with medium consisting primarily of glucose and trace levels of trehalose and arabinose showed a drop in sugar concentration over time and an increase in glycerol. Yeast growing on a modified culture medium consisting of multiple carbohydrates and alditols showed preference for specific carbon sources and showed the ability to regulate pathways leading to catalysis of

  19. Tuberculosis Diagnosis and Multidrug Resistance Testing by Direct Sputum Culture in Selective Broth without Decontamination or Centrifugation ▿ †

    PubMed Central

    Grandjean, Louis; Martin, Laura; Gilman, Robert H.; Valencia, Teresa; Herrera, Beatriz; Quino, Willi; Ramos, Eric; Rivero, Maribel; Montoya, Rosario; Escombe, A. Roderick; Coleman, David; Mitchison, Denis; Evans, Carlton A.

    2008-01-01

    Tuberculosis culture usually requires sputum decontamination and centrifugation to prevent cultures from being overgrown by contaminating bacteria and fungi. However, decontamination destroys many tuberculous bacilli, and centrifugation often is not possible in resource-poor settings. We therefore assessed the performance of Mycobacterium tuberculosis culture with unprocessed samples plated directly by using tuberculosis-selective media and compared this procedure to conventional culture using centrifuge decontamination. Quadruplicate aliquots of strain H37RV were cultured in 7H9 broth with and without selective antimicrobials and after centrifuge decontamination. The subsequent comparison was made with 715 sputum samples. Split paired sputum samples were cultured conventionally with centrifuge decontamination and by direct culture in tuberculosis-selective media containing antibiotics. Centrifuge decontamination reduced tuberculosis H37RV colonies by 78% (P < 0.001), whereas direct culture in tuberculosis-selective media had no inhibitory effect. Similarly, in sputum cultures that were not overgrown by contaminants, conventional culture yielded fewer tuberculosis colonies than direct culture (P < 0.001). However, the sensitivity of conventional culture was greater than that of direct culture, because samples were less affected by contamination. Thus, of the 340 sputum samples that were tuberculosis culture positive, conventional culture detected 97%, whereas direct culture detected 81% (P < 0.001). Conventional and direct cultures both took a median of 8.0 days to diagnose tuberculosis (P = 0.8). In those direct cultures that detected drug resistance or susceptibility, there was a 97% agreement with the results of conventional culture (Kappa agreement statistic, 0.84; P < 0.001). Direct culture is a simple, low-technology, and rapid technique for diagnosing tuberculosis and determining drug susceptibility. Compared to that of conventional culture, direct culture

  20. Tuberculosis diagnosis and multidrug resistance testing by direct sputum culture in selective broth without decontamination or centrifugation.

    PubMed

    Grandjean, Louis; Martin, Laura; Gilman, Robert H; Valencia, Teresa; Herrera, Beatriz; Quino, Willi; Ramos, Eric; Rivero, Maribel; Montoya, Rosario; Escombe, A Roderick; Coleman, David; Mitchison, Denis; Evans, Carlton A

    2008-07-01

    Tuberculosis culture usually requires sputum decontamination and centrifugation to prevent cultures from being overgrown by contaminating bacteria and fungi. However, decontamination destroys many tuberculous bacilli, and centrifugation often is not possible in resource-poor settings. We therefore assessed the performance of Mycobacterium tuberculosis culture with unprocessed samples plated directly by using tuberculosis-selective media and compared this procedure to conventional culture using centrifuge decontamination. Quadruplicate aliquots of strain H37RV were cultured in 7H9 broth with and without selective antimicrobials and after centrifuge decontamination. The subsequent comparison was made with 715 sputum samples. Split paired sputum samples were cultured conventionally with centrifuge decontamination and by direct culture in tuberculosis-selective media containing antibiotics. Centrifuge decontamination reduced tuberculosis H37RV colonies by 78% (P < 0.001), whereas direct culture in tuberculosis-selective media had no inhibitory effect. Similarly, in sputum cultures that were not overgrown by contaminants, conventional culture yielded fewer tuberculosis colonies than direct culture (P < 0.001). However, the sensitivity of conventional culture was greater than that of direct culture, because samples were less affected by contamination. Thus, of the 340 sputum samples that were tuberculosis culture positive, conventional culture detected 97%, whereas direct culture detected 81% (P < 0.001). Conventional and direct cultures both took a median of 8.0 days to diagnose tuberculosis (P = 0.8). In those direct cultures that detected drug resistance or susceptibility, there was a 97% agreement with the results of conventional culture (Kappa agreement statistic, 0.84; P < 0.001). Direct culture is a simple, low-technology, and rapid technique for diagnosing tuberculosis and determining drug susceptibility. Compared to that of conventional culture, direct culture