fluconazole  (Millipore)


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  • 99
    Name:
    Fluconazole
    Description:
    Fluconazole is used to treat adult neutropenic patients with invasive candidiasis IC
    Catalog Number:
    f8929
    Price:
    None
    Applications:
    Fluconazole has been used to assess minimum inhibitory concentration (MIC)-fungal assay. It has also been used as a positive control in antifungal assay of plants.
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    Structured Review

    Millipore fluconazole
    Fluconazole
    Fluconazole is used to treat adult neutropenic patients with invasive candidiasis IC
    https://www.bioz.com/result/fluconazole/product/Millipore
    Average 99 stars, based on 36 article reviews
    Price from $9.99 to $1999.99
    fluconazole - by Bioz Stars, 2020-09
    99/100 stars

    Images

    1) Product Images from "Selection and evaluation of appropriate reference genes for RT-qPCR based expression analysis in Candida tropicalis following azole treatment"

    Article Title: Selection and evaluation of appropriate reference genes for RT-qPCR based expression analysis in Candida tropicalis following azole treatment

    Journal: Scientific Reports

    doi: 10.1038/s41598-020-58744-7

    Violin plot representing the distribution of the CT values obtained for 10 candidate reference genes form 60 samples (30 fluconazole treated and 30 untreated control). Violin plot representing minimum value to maximum value with probability density of the data.
    Figure Legend Snippet: Violin plot representing the distribution of the CT values obtained for 10 candidate reference genes form 60 samples (30 fluconazole treated and 30 untreated control). Violin plot representing minimum value to maximum value with probability density of the data.

    Techniques Used:

    2) Product Images from "Fluconazole Alters the Polysaccharide Capsule of Cryptococcus gattii and Leads to Distinct Behaviors in Murine Cryptococcosis"

    Article Title: Fluconazole Alters the Polysaccharide Capsule of Cryptococcus gattii and Leads to Distinct Behaviors in Murine Cryptococcosis

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0112669

    Reduced susceptibility to fluconazole leads to very low CAP59 and CAP64 gene expression and affects physical properties of the GXM capsule. Microscopic visualization with India ink stain of L27/01 ( A ) and L27/01 F strains ( B ). Capsule size of L27/01 and L27/01 F strains ( C ). SEM analysis of L27/01 ( D ) and L27/01 F ( E ) strains. Zeta potential of capsular and secreted PS of cells ( F ). Cell suspensions were analyzed with an AXIOPLAN (Carl Zeiss) fluorescence microscope. Images were processed using ImageJ. Staining of L27/01 ( G ) and L27/01 F ( H ) strains with mAb 18B7 showing differences in epitope presentation in the PS capsule between strains. Ratio of gene expression of CAP59 , CAP64 and UXS1 to actin ( I ). Size distribution of PS fibers from capsular ( J and K ) and exo-PS samples ( L and M ) of L27/01 and L27/01 F strains, respectively. Bar = 10 µm (A) and (D). Bar = 5 µm (B), (E), (G) and (H). *P
    Figure Legend Snippet: Reduced susceptibility to fluconazole leads to very low CAP59 and CAP64 gene expression and affects physical properties of the GXM capsule. Microscopic visualization with India ink stain of L27/01 ( A ) and L27/01 F strains ( B ). Capsule size of L27/01 and L27/01 F strains ( C ). SEM analysis of L27/01 ( D ) and L27/01 F ( E ) strains. Zeta potential of capsular and secreted PS of cells ( F ). Cell suspensions were analyzed with an AXIOPLAN (Carl Zeiss) fluorescence microscope. Images were processed using ImageJ. Staining of L27/01 ( G ) and L27/01 F ( H ) strains with mAb 18B7 showing differences in epitope presentation in the PS capsule between strains. Ratio of gene expression of CAP59 , CAP64 and UXS1 to actin ( I ). Size distribution of PS fibers from capsular ( J and K ) and exo-PS samples ( L and M ) of L27/01 and L27/01 F strains, respectively. Bar = 10 µm (A) and (D). Bar = 5 µm (B), (E), (G) and (H). *P

    Techniques Used: Expressing, Staining, Fluorescence, Microscopy

    Reduced susceptibility to fluconazole impairs C. gattii migration to the CNS. Lungs ( A ), brain ( B ), and Bronchoalveolar lavage fluid (BALF) ( C ) were removed (n = 8), homogenized, diluted and plated onto Sabouraud dextrose agar for measurement of fungal burden 0.5, 1, 7, 15, and 30 days post-infection with 10 6 cells of L27/01 or L27/01 F strains. ND: non-detected. Black bars refer to L27/01 and grey bars refer to L27/01 F . *Data represent the mean of two independent experiments in triplicate. P
    Figure Legend Snippet: Reduced susceptibility to fluconazole impairs C. gattii migration to the CNS. Lungs ( A ), brain ( B ), and Bronchoalveolar lavage fluid (BALF) ( C ) were removed (n = 8), homogenized, diluted and plated onto Sabouraud dextrose agar for measurement of fungal burden 0.5, 1, 7, 15, and 30 days post-infection with 10 6 cells of L27/01 or L27/01 F strains. ND: non-detected. Black bars refer to L27/01 and grey bars refer to L27/01 F . *Data represent the mean of two independent experiments in triplicate. P

    Techniques Used: Migration, Infection

    L27/01 developed reduced susceptibility to fluconazole with overexpression of the MDR1 gene, but the parental strain is more virulent than L27/01 F in mice. Pulsed field gel electrophoresis (PFGE) patterns of L27/01 F and L27/01 and strains on 1% PFGE certified agarose (lanes 2 and 3, respectively). Lane 1 show PFGE Size Marker (SM), 0.225–2.2 Mb S. cerevisiae chromosomal DNA Saccharomyces cerevisiae used as size standard ( A ). MDR1 gene expression in L27/01 and L27/01 F strains ( B ). Ergosterol levels of L27/01 and L27/01 F strains ( C ). Urease and Laccase activities of L27/01 and L27/01 F strains: one enzymatic unit was calculated per 10 5 and 10 8 cells of C. gattii for urease and laccase activity, respectively, after 24 hours ( D ). Eight mice per group were inoculated by the intratracheal route with 10 6 cells of L27/01 or L27/01 F . Animals administered with PBS represent the non-infected (control) group. Animals were monitored daily. Survival curve ( E ). Weight variation (Score) ( F ). Each 1 (one) gram received or lost corresponds to 10 points added or subtracted, respectively. *The vertical dotted line indicates a significant difference from the appointed day. CFU/g of lungs ( G ) from animals infected with 1×10 6 cells of L27/01 or L27/01 F strains treated i.p. with fluconazole at 10 mg/kg/day (n = 6). NI: not infected group. NT: non treated group. Black bars refer to L27/01 (▪) and grey bars refer to L27/01 F (▴). Data represent the mean of three independent experiments in triplicate. *P
    Figure Legend Snippet: L27/01 developed reduced susceptibility to fluconazole with overexpression of the MDR1 gene, but the parental strain is more virulent than L27/01 F in mice. Pulsed field gel electrophoresis (PFGE) patterns of L27/01 F and L27/01 and strains on 1% PFGE certified agarose (lanes 2 and 3, respectively). Lane 1 show PFGE Size Marker (SM), 0.225–2.2 Mb S. cerevisiae chromosomal DNA Saccharomyces cerevisiae used as size standard ( A ). MDR1 gene expression in L27/01 and L27/01 F strains ( B ). Ergosterol levels of L27/01 and L27/01 F strains ( C ). Urease and Laccase activities of L27/01 and L27/01 F strains: one enzymatic unit was calculated per 10 5 and 10 8 cells of C. gattii for urease and laccase activity, respectively, after 24 hours ( D ). Eight mice per group were inoculated by the intratracheal route with 10 6 cells of L27/01 or L27/01 F . Animals administered with PBS represent the non-infected (control) group. Animals were monitored daily. Survival curve ( E ). Weight variation (Score) ( F ). Each 1 (one) gram received or lost corresponds to 10 points added or subtracted, respectively. *The vertical dotted line indicates a significant difference from the appointed day. CFU/g of lungs ( G ) from animals infected with 1×10 6 cells of L27/01 or L27/01 F strains treated i.p. with fluconazole at 10 mg/kg/day (n = 6). NI: not infected group. NT: non treated group. Black bars refer to L27/01 (▪) and grey bars refer to L27/01 F (▴). Data represent the mean of three independent experiments in triplicate. *P

    Techniques Used: Over Expression, Mouse Assay, Pulsed-Field Gel, Electrophoresis, Marker, Expressing, Activity Assay, Infection

    Synopsis of the methodology. Fluconazole-resistant strain selection ( A–F ). After determining the MIC of fluconazole on SDA, an average of five colonies obtained from the highest fluconazole concentration were selected for culture on SDA plates supplemented with this drug. L27/01 ( A ) was the strain able to grow at the highest fluconazole concentration ( B ) and was chosen for culture on SDA plates supplemented with this drug. This strain was cultured in solid medium with increasing concentrations ( B–C ) of fluconazole until growth ceased at 100 µg/mL ( D ). To verify the maintenance of resistance to fluconazole and cross-resistance between this drug and amphotericin B, the selected strain was cultured in SDA without drug every 48 h 170 times ( E ), and the MIC test was performed by microdilution every 5 subcultures ( F ). Colonies grown at 95 µg/mL were maintained in this concentration, and the strain was named “L27/01 F ” ( G ). L27/01 strain grew in the absence of drug ( H ). The genetic similarity between L27/01 and L27/01 F strains was evaluated by randomly amplified polymorphic DNA (RAPD)-PCR, PFGE and ITS Sequencing. CAP59, CAP64, ARF-1, ERG11, UXS-1 levels by real-time PCR were evaluated. Lipid evaluation was performed to compare the ergosterol content of L27/01 and L27/01 F cell membranes. Also, Urease and Laccase activities of L27/01 and L27/01 F strains were determined ( I ). Evaluation of murine cryptococcosis after inoculation with L27/01 or L27/01 F strain: survival curve ( J ). Investigation of whether fluconazole affects the polysaccharide (PS) capsule ( K ). The phagocytosis assay was performed to assess the influence of PS capsules from L27/01 and L27/01 F strains on phagocytosis and intracellular proliferation rate (IPR) in murine peritoneal macrophages from C57BL/6 mice ( L ). Cryptococcal cell dissemination, immune response and behavioral alterations ( M ). SDA: Sabouraud Dextrose Agar. MIC: Minimum inhibitory concentration. PFGE: Pulsed field gel electrophoresis. CFU: Colony forming units. BALF: bronchoalveolar lavage fluid. MPO: Myeloperoxidase activity. i.t.: intratracheal infection. PS: polysaccharide.
    Figure Legend Snippet: Synopsis of the methodology. Fluconazole-resistant strain selection ( A–F ). After determining the MIC of fluconazole on SDA, an average of five colonies obtained from the highest fluconazole concentration were selected for culture on SDA plates supplemented with this drug. L27/01 ( A ) was the strain able to grow at the highest fluconazole concentration ( B ) and was chosen for culture on SDA plates supplemented with this drug. This strain was cultured in solid medium with increasing concentrations ( B–C ) of fluconazole until growth ceased at 100 µg/mL ( D ). To verify the maintenance of resistance to fluconazole and cross-resistance between this drug and amphotericin B, the selected strain was cultured in SDA without drug every 48 h 170 times ( E ), and the MIC test was performed by microdilution every 5 subcultures ( F ). Colonies grown at 95 µg/mL were maintained in this concentration, and the strain was named “L27/01 F ” ( G ). L27/01 strain grew in the absence of drug ( H ). The genetic similarity between L27/01 and L27/01 F strains was evaluated by randomly amplified polymorphic DNA (RAPD)-PCR, PFGE and ITS Sequencing. CAP59, CAP64, ARF-1, ERG11, UXS-1 levels by real-time PCR were evaluated. Lipid evaluation was performed to compare the ergosterol content of L27/01 and L27/01 F cell membranes. Also, Urease and Laccase activities of L27/01 and L27/01 F strains were determined ( I ). Evaluation of murine cryptococcosis after inoculation with L27/01 or L27/01 F strain: survival curve ( J ). Investigation of whether fluconazole affects the polysaccharide (PS) capsule ( K ). The phagocytosis assay was performed to assess the influence of PS capsules from L27/01 and L27/01 F strains on phagocytosis and intracellular proliferation rate (IPR) in murine peritoneal macrophages from C57BL/6 mice ( L ). Cryptococcal cell dissemination, immune response and behavioral alterations ( M ). SDA: Sabouraud Dextrose Agar. MIC: Minimum inhibitory concentration. PFGE: Pulsed field gel electrophoresis. CFU: Colony forming units. BALF: bronchoalveolar lavage fluid. MPO: Myeloperoxidase activity. i.t.: intratracheal infection. PS: polysaccharide.

    Techniques Used: Selection, Concentration Assay, Cell Culture, Amplification, Polymerase Chain Reaction, Sequencing, Real-time Polymerase Chain Reaction, Phagocytosis Assay, Mouse Assay, Pulsed-Field Gel, Electrophoresis, Activity Assay, Infection

    3) Product Images from "Potent Synergy between Spirocyclic Pyrrolidinoindolinones and Fluconazole against Candida albicans"

    Article Title: Potent Synergy between Spirocyclic Pyrrolidinoindolinones and Fluconazole against Candida albicans

    Journal: ChemMedChem

    doi: 10.1002/cmdc.201500271

    Fluconazole synergizer CID 6584729 and diastereomer 1 .
    Figure Legend Snippet: Fluconazole synergizer CID 6584729 and diastereomer 1 .

    Techniques Used:

    4) Product Images from "Intestinal Candida parapsilosis isolates from Rett syndrome subjects bear potential virulent traits and capacity to persist within the host"

    Article Title: Intestinal Candida parapsilosis isolates from Rett syndrome subjects bear potential virulent traits and capacity to persist within the host

    Journal: BMC Gastroenterology

    doi: 10.1186/s12876-018-0785-z

    a ) Fluconazole and b ) itraconazole resistance as measured by MIC values in C. albicans and C. parapsilosis isolates from HC and RTT subjects. MIC values are reported as means ± standard errors. Exact p-values are reported and considered significant if
    Figure Legend Snippet: a ) Fluconazole and b ) itraconazole resistance as measured by MIC values in C. albicans and C. parapsilosis isolates from HC and RTT subjects. MIC values are reported as means ± standard errors. Exact p-values are reported and considered significant if

    Techniques Used:

    5) Product Images from "Resorbable Beads Provide Extended Release of Antifungal Medication: In Vitro and In Vivo Analyses"

    Article Title: Resorbable Beads Provide Extended Release of Antifungal Medication: In Vitro and In Vivo Analyses

    Journal: Pharmaceutics

    doi: 10.3390/pharmaceutics11110550

    Cell viability of the fluconazole-incorporated beads (* p
    Figure Legend Snippet: Cell viability of the fluconazole-incorporated beads (* p

    Techniques Used:

    Photographs of the fabricated drug-loaded beads. The white cylindrical beads are fluconazole-impregnated Poly( d , l -lactide- co -glycolide) (PLGA) beads. The yellow cylindrical beads are amphotericin B-impregnated PLGA beads.
    Figure Legend Snippet: Photographs of the fabricated drug-loaded beads. The white cylindrical beads are fluconazole-impregnated Poly( d , l -lactide- co -glycolide) (PLGA) beads. The yellow cylindrical beads are amphotericin B-impregnated PLGA beads.

    Techniques Used:

    In vitro release curves of fluconazole from drug-loaded beads with various polymer:drug ratios. ( A ). The accumulated release of beads of different sizes with 4:1 polymer:drug ratios. ( B ). The accumulated release of beads of different sizes with 6:1 polymer:drug ratios.
    Figure Legend Snippet: In vitro release curves of fluconazole from drug-loaded beads with various polymer:drug ratios. ( A ). The accumulated release of beads of different sizes with 4:1 polymer:drug ratios. ( B ). The accumulated release of beads of different sizes with 6:1 polymer:drug ratios.

    Techniques Used: In Vitro

    Fourier-transform infrared spectra of ( A ) pure Poly( d , l -lactide- co -glycolide) (PLGA) and amphotericin B/PLGA beads ( B ) pure Poly( d , l -lactide- co -glycolide) (PLGA) and fluconazole/PLGA beads.
    Figure Legend Snippet: Fourier-transform infrared spectra of ( A ) pure Poly( d , l -lactide- co -glycolide) (PLGA) and amphotericin B/PLGA beads ( B ) pure Poly( d , l -lactide- co -glycolide) (PLGA) and fluconazole/PLGA beads.

    Techniques Used:

    Images of the surgical procedure. ( A ) The right femoral site was depilated and sterilized. ( B ) a bone cavity was formed (5.0 × 10.0 mm 2 ); ( C ) a polymethylmethacrylate spacer was placed into the bone cavity; ( D ) after 2 weeks, fluconazole-impregnated Poly( d , l -lactide- co -glycolide) cylindrical beads (5.0 × 6.0 mm 2 ) were placed into the right femoral bone cavity.
    Figure Legend Snippet: Images of the surgical procedure. ( A ) The right femoral site was depilated and sterilized. ( B ) a bone cavity was formed (5.0 × 10.0 mm 2 ); ( C ) a polymethylmethacrylate spacer was placed into the bone cavity; ( D ) after 2 weeks, fluconazole-impregnated Poly( d , l -lactide- co -glycolide) cylindrical beads (5.0 × 6.0 mm 2 ) were placed into the right femoral bone cavity.

    Techniques Used:

    In vivo release curves of fluconazole from the drug-loaded beads.
    Figure Legend Snippet: In vivo release curves of fluconazole from the drug-loaded beads.

    Techniques Used: In Vivo

    Effect of molding temperature on ( A ) amphotericin B and ( B ) fluconazole stability determined by Fourier-transform infrared spectroscopy.
    Figure Legend Snippet: Effect of molding temperature on ( A ) amphotericin B and ( B ) fluconazole stability determined by Fourier-transform infrared spectroscopy.

    Techniques Used: Spectroscopy

    6) Product Images from "Sir2 regulates stability of repetitive domains differentially in the human fungal pathogen Candida albicans"

    Article Title: Sir2 regulates stability of repetitive domains differentially in the human fungal pathogen Candida albicans

    Journal: Nucleic Acids Research

    doi: 10.1093/nar/gkw594

    Instability of TLO genes triggered by stress conditions masks the action of Sir2 . ( A ) Schematic of TLOα10-URA3 + . ( B ) TLOα10-URA3 + fluctuation analysis in WT and sir2 Δ/Δ cells without (−) and with (+) fluconazole treatment. p -values = 4.846 × 10 −09 and 0.6591, respectively. ( C ) Fluctuation analysis for LOH Rates in TLOα10-URA3 + in WT and sir2 Δ/Δ cells without (−) and with (+) H 2 O 2 treatment. p -values = 4.846 × 10 −09 and 0.0001475, respectively. ( D ) TLOα10-URA3 + in WT and sir2 Δ/Δ cells fluctuation analysis at 30°C or 39°C. p -values = 4.846 × 10 −09 and 3.065 × 10 −05 , respectively. ( E ) Schematic of TLOα10 Δ TRE-URA3 + . ( F ) TLOα10 Δ TRE-URA3 + fluctuation analysis without (−) and with (+) fluconazole treatment. p -values = 3.065 × 10 −05 .
    Figure Legend Snippet: Instability of TLO genes triggered by stress conditions masks the action of Sir2 . ( A ) Schematic of TLOα10-URA3 + . ( B ) TLOα10-URA3 + fluctuation analysis in WT and sir2 Δ/Δ cells without (−) and with (+) fluconazole treatment. p -values = 4.846 × 10 −09 and 0.6591, respectively. ( C ) Fluctuation analysis for LOH Rates in TLOα10-URA3 + in WT and sir2 Δ/Δ cells without (−) and with (+) H 2 O 2 treatment. p -values = 4.846 × 10 −09 and 0.0001475, respectively. ( D ) TLOα10-URA3 + in WT and sir2 Δ/Δ cells fluctuation analysis at 30°C or 39°C. p -values = 4.846 × 10 −09 and 3.065 × 10 −05 , respectively. ( E ) Schematic of TLOα10 Δ TRE-URA3 + . ( F ) TLOα10 Δ TRE-URA3 + fluctuation analysis without (−) and with (+) fluconazole treatment. p -values = 3.065 × 10 −05 .

    Techniques Used:

    Regulation of genome plasticity in C. albicans . ( A ) Schematic model to represent recombination control at the rDNA locus in WT, sir2Δ/Δ and csm1Δ/Δ cells. ( B ) Schematic model to represent recombination control at subtelomeric regions in WT and sir2Δ/Δ cells. ( C ) Schematic model to represent recombination rates in control or under stress conditions (30/39°C, fluconazole and H 2 O 2 treatment) in WT, sir2Δ/Δ cells.
    Figure Legend Snippet: Regulation of genome plasticity in C. albicans . ( A ) Schematic model to represent recombination control at the rDNA locus in WT, sir2Δ/Δ and csm1Δ/Δ cells. ( B ) Schematic model to represent recombination control at subtelomeric regions in WT and sir2Δ/Δ cells. ( C ) Schematic model to represent recombination rates in control or under stress conditions (30/39°C, fluconazole and H 2 O 2 treatment) in WT, sir2Δ/Δ cells.

    Techniques Used:

    Stress conditions increase LOH associated with all genomic loci tested. ( A ) Schematic of TLOα10-URA3 + , TLOγ16-URA3 + , URA3 + and rDNA-URA3 + . ( B ) TLOα10-URA3 + , TLOγ16-URA3 + , URA3 + and rDNA-URA3 + fluctuation analysis without (−) and with (+) fluconazole treatment. The calculated p -values in the presence and absence of fluconazole for each strain are respectively 2.035 × 10 −07 , 1.125 × 10 −05 , 3.697 × 10 −07 and 2.035 × 10 −07 .
    Figure Legend Snippet: Stress conditions increase LOH associated with all genomic loci tested. ( A ) Schematic of TLOα10-URA3 + , TLOγ16-URA3 + , URA3 + and rDNA-URA3 + . ( B ) TLOα10-URA3 + , TLOγ16-URA3 + , URA3 + and rDNA-URA3 + fluctuation analysis without (−) and with (+) fluconazole treatment. The calculated p -values in the presence and absence of fluconazole for each strain are respectively 2.035 × 10 −07 , 1.125 × 10 −05 , 3.697 × 10 −07 and 2.035 × 10 −07 .

    Techniques Used:

    7) Product Images from "In vitro evaluation of the activity of an essential oil from Pistacia vera L. variety Bronte hull against Candida sp."

    Article Title: In vitro evaluation of the activity of an essential oil from Pistacia vera L. variety Bronte hull against Candida sp.

    Journal: BMC Complementary and Alternative Medicine

    doi: 10.1186/s12906-018-2425-0

    Structural formula of fluconazole ( a ), voriconazole ( b ) and caspofungin ( c )
    Figure Legend Snippet: Structural formula of fluconazole ( a ), voriconazole ( b ) and caspofungin ( c )

    Techniques Used:

    8) Product Images from "Novel Cell-Killing Mechanisms of Hydroxyurea and the Implication toward Combination Therapy for the Treatment of Fungal Infections"

    Article Title: Novel Cell-Killing Mechanisms of Hydroxyurea and the Implication toward Combination Therapy for the Treatment of Fungal Infections

    Journal: Antimicrobial Agents and Chemotherapy

    doi: 10.1128/AAC.00734-17

    Time course analysis of the synergistic cytotoxic effect of HU in combination with SMP, terbinafine, itraconazole, clotrimazole, ketoconazole, or fluconazole in S. pombe . Logarithmically growing wild-type S. pombe was inoculated in YE6S medium on 96-well plates at 3,000 cells/well, and then 10 mM HU and 21 μM SMP (A), 5 mM HU and 0.091 μM terbinafine (B), 10 mM HU and 0.021 μM itraconazole (C), 10 mM HU and 0.043 μM clotrimazole (D), 5 mM HU and 9.4 μM ketoconazole (E), or 10 mM HU and 22.8 μM fluconazole (F) were added to the cell cultures either alone or in combinations. The same amounts of carriers were added for the control. The plates were incubated at 30°C and scanned at the indicated time points to monitor cell survival. All data points are the averages of the readings from three separate wells.
    Figure Legend Snippet: Time course analysis of the synergistic cytotoxic effect of HU in combination with SMP, terbinafine, itraconazole, clotrimazole, ketoconazole, or fluconazole in S. pombe . Logarithmically growing wild-type S. pombe was inoculated in YE6S medium on 96-well plates at 3,000 cells/well, and then 10 mM HU and 21 μM SMP (A), 5 mM HU and 0.091 μM terbinafine (B), 10 mM HU and 0.021 μM itraconazole (C), 10 mM HU and 0.043 μM clotrimazole (D), 5 mM HU and 9.4 μM ketoconazole (E), or 10 mM HU and 22.8 μM fluconazole (F) were added to the cell cultures either alone or in combinations. The same amounts of carriers were added for the control. The plates were incubated at 30°C and scanned at the indicated time points to monitor cell survival. All data points are the averages of the readings from three separate wells.

    Techniques Used: Incubation

    9) Product Images from "Genetic Analysis of NDT80 Family Transcription Factors in Candida albicans Using New CRISPR-Cas9 Approaches"

    Article Title: Genetic Analysis of NDT80 Family Transcription Factors in Candida albicans Using New CRISPR-Cas9 Approaches

    Journal: mSphere

    doi: 10.1128/mSphere.00545-18

    Sensitivity of mutants to antifungal drugs. The sensitivity to amphotericin B and fluconazole was determined by a disk diffusion assay in which cells were spread onto the surface of an RPMI 1640 medium plate and then filter discs containing 25 µg of the drugs were placed on the surface of the plate. The ron1 Δ rep1 Δ ndt80 Δ triple mutant was slightly more sensitive to amphotericin B (AMB) than the wild-type SC5314 strain but was not more sensitive to fluconazole (FLC). None of the mutants showed significant differences in the zone of 50% growth inhibition (RAD 50 ) in fluconazole compared to the wild-type strain. However, the deletion mutants lacking NDT80 (mutants ndt80 Δ, rep1 Δ ndt80 Δ, ron1 Δ ndt80 Δ, and ron1 Δ rep1 Δ ndt80 Δ) did not show the trailing growth around the fluconazole disks that was seen for the other strains. The plates were incubated at 30°C for 48 h and then photographed. Image analysis software was used to measure the zone of growth inhibition to determine the average radius that corresponded to a 50% growth reduction (RAD 50 ). Double asterisks show statistically significant differences ( P
    Figure Legend Snippet: Sensitivity of mutants to antifungal drugs. The sensitivity to amphotericin B and fluconazole was determined by a disk diffusion assay in which cells were spread onto the surface of an RPMI 1640 medium plate and then filter discs containing 25 µg of the drugs were placed on the surface of the plate. The ron1 Δ rep1 Δ ndt80 Δ triple mutant was slightly more sensitive to amphotericin B (AMB) than the wild-type SC5314 strain but was not more sensitive to fluconazole (FLC). None of the mutants showed significant differences in the zone of 50% growth inhibition (RAD 50 ) in fluconazole compared to the wild-type strain. However, the deletion mutants lacking NDT80 (mutants ndt80 Δ, rep1 Δ ndt80 Δ, ron1 Δ ndt80 Δ, and ron1 Δ rep1 Δ ndt80 Δ) did not show the trailing growth around the fluconazole disks that was seen for the other strains. The plates were incubated at 30°C for 48 h and then photographed. Image analysis software was used to measure the zone of growth inhibition to determine the average radius that corresponded to a 50% growth reduction (RAD 50 ). Double asterisks show statistically significant differences ( P

    Techniques Used: Diffusion-based Assay, Mutagenesis, Inhibition, Incubation, Software

    10) Product Images from "Candida albicansVPS4 contributes differentially to epithelial and mucosal pathogenesis"

    Article Title: Candida albicansVPS4 contributes differentially to epithelial and mucosal pathogenesis

    Journal: Virulence

    doi: 10.4161/21505594.2014.956648

    The C. albicans vps4 Δ mutant is more sensitive to cell wall stress and challenge with antifungals. C. albicans DAY185, vps4 Δ, and VPS4 reintegrant strains were grown on agar plates containing cell-wall perturbing agents or antifungal drugs. Strains are indicated on the left. Cell densities decrease from left to right (1 × 10 7 , 2 × 10 6 , 4 × 10 5 , and 8 × 10 4 cells per mL). Normal growth on YPD medium is shown on the top right. YPD medium containing cell wall perturbing agents including 0.02% SDS, 200 μg/mL Congo Red, and 50 μg/mL Calcofluor White, as well as medium containing the antifungal drugs caspofungin (0.1 μg/mL) and fluconazole (4 μg/mL), are shown.
    Figure Legend Snippet: The C. albicans vps4 Δ mutant is more sensitive to cell wall stress and challenge with antifungals. C. albicans DAY185, vps4 Δ, and VPS4 reintegrant strains were grown on agar plates containing cell-wall perturbing agents or antifungal drugs. Strains are indicated on the left. Cell densities decrease from left to right (1 × 10 7 , 2 × 10 6 , 4 × 10 5 , and 8 × 10 4 cells per mL). Normal growth on YPD medium is shown on the top right. YPD medium containing cell wall perturbing agents including 0.02% SDS, 200 μg/mL Congo Red, and 50 μg/mL Calcofluor White, as well as medium containing the antifungal drugs caspofungin (0.1 μg/mL) and fluconazole (4 μg/mL), are shown.

    Techniques Used: Mutagenesis

    11) Product Images from "Characterization of a novel antibiofilm effect of nitric oxide-releasing aspirin (NCX-4040) on Candida albicans isolates from denture stomatitis patients"

    Article Title: Characterization of a novel antibiofilm effect of nitric oxide-releasing aspirin (NCX-4040) on Candida albicans isolates from denture stomatitis patients

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0176755

    NO-ASA increases the antifungal effect of fluconazole in resistant strains. Bars represent mean ± SD of inhibition zone diameters. Discs contained 25 μg fluconazole, alone or in combination with 25 μg nitric oxide-releasing aspirin or 25 μg ASA. Controls containing only vehicle (DMSO), aspirin (ASA), or NCX-4040 NO-ASA showed no inhibition zone diameters (not shown). *p
    Figure Legend Snippet: NO-ASA increases the antifungal effect of fluconazole in resistant strains. Bars represent mean ± SD of inhibition zone diameters. Discs contained 25 μg fluconazole, alone or in combination with 25 μg nitric oxide-releasing aspirin or 25 μg ASA. Controls containing only vehicle (DMSO), aspirin (ASA), or NCX-4040 NO-ASA showed no inhibition zone diameters (not shown). *p

    Techniques Used: Inhibition

    NO-ASA inhibits C . albicans planktonic morphogenesis. Upper Panel : light microscopy photographs showing representative filamentous cells (hyphae or pseudohyphae) for the 17p strain after 3 h or 12 h of incubation with DMSO (A, E) ; 800 μM fluconazole (B, F) or 500 μM aspirin (C, G). Cells treated with 500 μM NO-ASA (D, H) showed mainly budding yeast cells and scarce filamentous cells (arrow in H). Lower panel : Bars represent mean ± SD of percentage of filamentous cells as compared to untreated controls for each strain assayed after 3 hours of incubation (untreated control bars were omitted for clarity). ****p
    Figure Legend Snippet: NO-ASA inhibits C . albicans planktonic morphogenesis. Upper Panel : light microscopy photographs showing representative filamentous cells (hyphae or pseudohyphae) for the 17p strain after 3 h or 12 h of incubation with DMSO (A, E) ; 800 μM fluconazole (B, F) or 500 μM aspirin (C, G). Cells treated with 500 μM NO-ASA (D, H) showed mainly budding yeast cells and scarce filamentous cells (arrow in H). Lower panel : Bars represent mean ± SD of percentage of filamentous cells as compared to untreated controls for each strain assayed after 3 hours of incubation (untreated control bars were omitted for clarity). ****p

    Techniques Used: Light Microscopy, Incubation

    NO-ASA inhibits the adhesion of C . albicans to abiotic surfaces. Bars represent mean ± SD of percentage of cells adhered to the bottom of wells as compared controls for each strain assayed. Due fluconazole 800 μM did not affect adhesion as compared to untreated strains it was used as control for the analysis. **p
    Figure Legend Snippet: NO-ASA inhibits the adhesion of C . albicans to abiotic surfaces. Bars represent mean ± SD of percentage of cells adhered to the bottom of wells as compared controls for each strain assayed. Due fluconazole 800 μM did not affect adhesion as compared to untreated strains it was used as control for the analysis. **p

    Techniques Used:

    12) Product Images from "Unexpected effects of azole transporter inhibitors on antifungal susceptibility in Candida glabrata and other pathogenic Candida species"

    Article Title: Unexpected effects of azole transporter inhibitors on antifungal susceptibility in Candida glabrata and other pathogenic Candida species

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0180990

    (A) The effect of clorgyline on fluconazole susceptibility in the C . glabrata CBS138 strain. Checkerboard susceptibility assay was performed as described in the Materials and Methods section. Percentages of cell growth relative to drug-free control are expressed as means ± SE. Clorgyline concentration: white bar, 0 μg/ml; light grey bar, 40 μg/ml; dark grey bar, 80 μg/ml; and black bar, 160 μg/ml. (B) Spot dilution assay. Logarithmic-phase cells were adjusted to 2 × 10 7 cells/ml, and 5 μl of serial 10-fold dilutions were then spotted onto SC plates containing FLCZ, clorgyline, or both. Plates were incubated at 30°C for 48 h. Final concentrations: clorgyline, 80 μg/ml; and FLCZ, 64 μg/ml.
    Figure Legend Snippet: (A) The effect of clorgyline on fluconazole susceptibility in the C . glabrata CBS138 strain. Checkerboard susceptibility assay was performed as described in the Materials and Methods section. Percentages of cell growth relative to drug-free control are expressed as means ± SE. Clorgyline concentration: white bar, 0 μg/ml; light grey bar, 40 μg/ml; dark grey bar, 80 μg/ml; and black bar, 160 μg/ml. (B) Spot dilution assay. Logarithmic-phase cells were adjusted to 2 × 10 7 cells/ml, and 5 μl of serial 10-fold dilutions were then spotted onto SC plates containing FLCZ, clorgyline, or both. Plates were incubated at 30°C for 48 h. Final concentrations: clorgyline, 80 μg/ml; and FLCZ, 64 μg/ml.

    Techniques Used: Drug Susceptibility Assay, Concentration Assay, Dilution Assay, Incubation

    13) Product Images from "Interspecies Outer Membrane Vesicles (OMVs) Modulate the Sensitivity of Pathogenic Bacteria and Pathogenic Yeasts to Cationic Peptides and Serum Complement"

    Article Title: Interspecies Outer Membrane Vesicles (OMVs) Modulate the Sensitivity of Pathogenic Bacteria and Pathogenic Yeasts to Cationic Peptides and Serum Complement

    Journal: International Journal of Molecular Sciences

    doi: 10.3390/ijms20225577

    OMVs protect yeast against synergistic fungicidal activity of polymyxin B in combination with fluconazole: ( a ) Synergistic activities determined by checkerboard assay; ( b ) time-kill assay; ( c ) 24 h growth-curve kinetics for C. albicans incubated with drug alone and drug in combinations with or without OMVs. The kinetics were measured using Varioskan™ LUX reader with measurements at 1 h intervals. The data for experiments (a,b) show means ± SD from two independent experiments carried out in triplicate; the data for experiment (c) show mean values from two independent repetitions carried out in duplicate; for better readability, SDs were not included.
    Figure Legend Snippet: OMVs protect yeast against synergistic fungicidal activity of polymyxin B in combination with fluconazole: ( a ) Synergistic activities determined by checkerboard assay; ( b ) time-kill assay; ( c ) 24 h growth-curve kinetics for C. albicans incubated with drug alone and drug in combinations with or without OMVs. The kinetics were measured using Varioskan™ LUX reader with measurements at 1 h intervals. The data for experiments (a,b) show means ± SD from two independent experiments carried out in triplicate; the data for experiment (c) show mean values from two independent repetitions carried out in duplicate; for better readability, SDs were not included.

    Techniques Used: Activity Assay, Time-Kill Assay, Incubation

    14) Product Images from "In Vitro and In Vivo Antifungal Activity of Lichochalcone-A against Candida albicans Biofilms"

    Article Title: In Vitro and In Vivo Antifungal Activity of Lichochalcone-A against Candida albicans Biofilms

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0157188

    Co-culture fluorescence microscopy stained with calcofluor white stain (Blue: C . albicans ) and Viability/Cytotoxicity Assay Kit for Animal Live Dead Cells (Green: live fibroblast cells; Red: dead fibroblast cells). A. Vehicle control (ethanol 1%); B. Positive control (fluconazole 32.2 μM); and C. Lichochalcone-A at 62.5 μM. Lichochalcone-A displayed low candida (blue fluorescence) growth with even distribution of live fibroblast cells (green fluorescence). Scale bars are set in μm.
    Figure Legend Snippet: Co-culture fluorescence microscopy stained with calcofluor white stain (Blue: C . albicans ) and Viability/Cytotoxicity Assay Kit for Animal Live Dead Cells (Green: live fibroblast cells; Red: dead fibroblast cells). A. Vehicle control (ethanol 1%); B. Positive control (fluconazole 32.2 μM); and C. Lichochalcone-A at 62.5 μM. Lichochalcone-A displayed low candida (blue fluorescence) growth with even distribution of live fibroblast cells (green fluorescence). Scale bars are set in μm.

    Techniques Used: Co-Culture Assay, Fluorescence, Microscopy, Staining, Cytotoxicity Assay, Positive Control

    Pro-inflammatory cytokines expression of IL-1α and IL-1β, and anti-inflammatory cytokine expression of IL-10 in C . albicans treated with lichochalcone-A (62.5 μM and 150 μM), fluconazole (32 μM) (positive control), and 1% ethanol (vehicle control). Significant decrease in IL-1α and IL-1β (*p
    Figure Legend Snippet: Pro-inflammatory cytokines expression of IL-1α and IL-1β, and anti-inflammatory cytokine expression of IL-10 in C . albicans treated with lichochalcone-A (62.5 μM and 150 μM), fluconazole (32 μM) (positive control), and 1% ethanol (vehicle control). Significant decrease in IL-1α and IL-1β (*p

    Techniques Used: Expressing, Positive Control

    Time Kill of C . albicans (MYA 2876) inoculum (10³ CFU/ml) tested against lichochalcone-A (at 10x MIC, and 20x MIC), fluconazole 32 μM, (positive control), 1% ethanol (vehicle control), and medium with inoculum only (negative control), plot expressed as average values for log10 of the numbers of CFU/milliliter versus time (hrs.).
    Figure Legend Snippet: Time Kill of C . albicans (MYA 2876) inoculum (10³ CFU/ml) tested against lichochalcone-A (at 10x MIC, and 20x MIC), fluconazole 32 μM, (positive control), 1% ethanol (vehicle control), and medium with inoculum only (negative control), plot expressed as average values for log10 of the numbers of CFU/milliliter versus time (hrs.).

    Techniques Used: Positive Control, Negative Control

    Fungal viability of C . albicans biofilm expressed in CFU/ml/ grams of dry weight after treatment with lichochalcone-A. The antifungal activity of lichochalcone-A (625 μM; 10x MIC) against C . albicans MYA 2876 biofilms was compared to the vehicle control group (1% ethanol) and positive control group (fluconazole 320 μM; 10x MIC) The standard deviations of each sample are shown in the graph, and all the mean differences between the control groups and test (lichochalcone-A at 625 μM) were statistically significant (*p
    Figure Legend Snippet: Fungal viability of C . albicans biofilm expressed in CFU/ml/ grams of dry weight after treatment with lichochalcone-A. The antifungal activity of lichochalcone-A (625 μM; 10x MIC) against C . albicans MYA 2876 biofilms was compared to the vehicle control group (1% ethanol) and positive control group (fluconazole 320 μM; 10x MIC) The standard deviations of each sample are shown in the graph, and all the mean differences between the control groups and test (lichochalcone-A at 625 μM) were statistically significant (*p

    Techniques Used: Activity Assay, Positive Control

    15) Product Images from "Biological exploration of a novel 1,2,4-triazole-indole hybrid molecule as antifungal agent"

    Article Title: Biological exploration of a novel 1,2,4-triazole-indole hybrid molecule as antifungal agent

    Journal: Journal of Enzyme Inhibition and Medicinal Chemistry

    doi: 10.1080/14756366.2019.1705292

    Survival curve of mice after treatment with fluconazole (3 x 5 mg/kg per os , □) or 8 g (2 x 30 mg/kg ip, Δ; 3 x 20 mg/kg ip, ▲). Control group (●). ** p
    Figure Legend Snippet: Survival curve of mice after treatment with fluconazole (3 x 5 mg/kg per os , □) or 8 g (2 x 30 mg/kg ip, Δ; 3 x 20 mg/kg ip, ▲). Control group (●). ** p

    Techniques Used: Mouse Assay

    16) Product Images from "Novel Antifungal Drug Discovery Based on Targeting Pathways Regulating the Fungus-Conserved Upc2 Transcription Factor"

    Article Title: Novel Antifungal Drug Discovery Based on Targeting Pathways Regulating the Fungus-Conserved Upc2 Transcription Factor

    Journal: Antimicrobial Agents and Chemotherapy

    doi: 10.1128/AAC.01677-13

    Cells expressing an UPC2-LacZ promoter fusion show a dose-dependent increase in β-galactosidase activity in response to increasing concentrations of fluconazole (in micrograms per milliliter). Cells were diluted 1:20 into the final assay volume
    Figure Legend Snippet: Cells expressing an UPC2-LacZ promoter fusion show a dose-dependent increase in β-galactosidase activity in response to increasing concentrations of fluconazole (in micrograms per milliliter). Cells were diluted 1:20 into the final assay volume

    Techniques Used: Expressing, Activity Assay

    Compounds 1, 2, and 3 decrease Upc2 SRE binding in the presence of fluconazole. Wild-type S. cerevisiae cells were grown to exponential phase in the absence and presence of fluconazole and each compound for 6 h at 30°C. ScUpc2 was immunoprecipitated
    Figure Legend Snippet: Compounds 1, 2, and 3 decrease Upc2 SRE binding in the presence of fluconazole. Wild-type S. cerevisiae cells were grown to exponential phase in the absence and presence of fluconazole and each compound for 6 h at 30°C. ScUpc2 was immunoprecipitated

    Techniques Used: Binding Assay, Immunoprecipitation

    Compounds 1, 2, and 3 inhibit fluconazole-induced Upc2-dependent activity. Cells were grown to exponential phase in the absence (−) and presence (+) of fluconazole (FLC) and each compound (compound 1, 2, or 3) for 6 h at 30°C. Total RNA
    Figure Legend Snippet: Compounds 1, 2, and 3 inhibit fluconazole-induced Upc2-dependent activity. Cells were grown to exponential phase in the absence (−) and presence (+) of fluconazole (FLC) and each compound (compound 1, 2, or 3) for 6 h at 30°C. Total RNA

    Techniques Used: Activity Assay

    Compounds 1, 2, and 3 decrease Upc2 protein expression in the presence of fluconazole. Wild-type S. cerevisiae cells were grown to exponential phase in the absence and presence of fluconazole (FLC) and each compound (compound 1, 2, or 3) for 6 h at 30°C
    Figure Legend Snippet: Compounds 1, 2, and 3 decrease Upc2 protein expression in the presence of fluconazole. Wild-type S. cerevisiae cells were grown to exponential phase in the absence and presence of fluconazole (FLC) and each compound (compound 1, 2, or 3) for 6 h at 30°C

    Techniques Used: Expressing

    Compounds 1, 2, and 3 inhibit fluconazole-induced CgUpc2-dependent activity. Cells were grown to exponential phase in the absence (−) and presence (+) of fluconazole and each compound (compound 1, 2, or 3) for 6 h at 30°C. Total RNA was
    Figure Legend Snippet: Compounds 1, 2, and 3 inhibit fluconazole-induced CgUpc2-dependent activity. Cells were grown to exponential phase in the absence (−) and presence (+) of fluconazole and each compound (compound 1, 2, or 3) for 6 h at 30°C. Total RNA was

    Techniques Used: Activity Assay

    17) Product Images from "Sensitive Assay for Antifungal Activity of Glucan Synthase Inhibitors That Uses Germ Tube Formation in Candida albicans as an End Point"

    Article Title: Sensitive Assay for Antifungal Activity of Glucan Synthase Inhibitors That Uses Germ Tube Formation in Candida albicans as an End Point

    Journal: Antimicrobial Agents and Chemotherapy

    doi: 10.1128/AAC.47.10.3305-3310.2003

    IC 50 s for 288 compounds in the germ tube formation assay. The values for reference compounds (amphotericin B, caspofungin, cilofungin, fluconazole, ketoconazole, and voriconazole) are indicated by the arrows. The remaining compounds are from a novel class of glucan synthase inhibitors.
    Figure Legend Snippet: IC 50 s for 288 compounds in the germ tube formation assay. The values for reference compounds (amphotericin B, caspofungin, cilofungin, fluconazole, ketoconazole, and voriconazole) are indicated by the arrows. The remaining compounds are from a novel class of glucan synthase inhibitors.

    Techniques Used: Tube Formation Assay

    18) Product Images from "Functional analysis of Paracoccidioides brasiliensis 14-3-3 adhesin expressed in Saccharomyces cerevisiae"

    Article Title: Functional analysis of Paracoccidioides brasiliensis 14-3-3 adhesin expressed in Saccharomyces cerevisiae

    Journal: BMC Microbiology

    doi: 10.1186/s12866-015-0586-2

    Evaluation of complementation. A spot assay was used to evaluate susceptibility to fluconazole 35 μM in wt ( a ), Δbmh1 ( b ) and Δbmh2 ( c ) S. cerevisiae transformants. There was a decreased sensitivity of transformants pYES-14-3-3 and pYES-BMH1 compared with the empty vector transformant (pYES). As a growth control, the transformants were also spotted in SD-URA without fluconazole
    Figure Legend Snippet: Evaluation of complementation. A spot assay was used to evaluate susceptibility to fluconazole 35 μM in wt ( a ), Δbmh1 ( b ) and Δbmh2 ( c ) S. cerevisiae transformants. There was a decreased sensitivity of transformants pYES-14-3-3 and pYES-BMH1 compared with the empty vector transformant (pYES). As a growth control, the transformants were also spotted in SD-URA without fluconazole

    Techniques Used: Spot Test, Plasmid Preparation

    19) Product Images from "Antifungal Potential of Host Defense Peptide Mimetics in a Mouse Model of Disseminated Candidiasis"

    Article Title: Antifungal Potential of Host Defense Peptide Mimetics in a Mouse Model of Disseminated Candidiasis

    Journal: Journal of Fungi

    doi: 10.3390/jof4010030

    Dose-response of Compounds 3 and 6 in vivo. Female CD.1 mice ( n = 5 per group) were infected IV with 3.5 × 10 4 cfu/mouse before being treated 2 h later with a single SC injection of increasing concentrations of Compound 3 or 6 for 24 h. As a positive control, female Swiss CD.1 mice were also gavaged at 2 h with 20 mg/kg fluconazole for 24 h. Data are presented as log reduction in kidney burden (±SEM) from the mean cfu of untreated mice ( n = 5). The dose-dependent reduction in kidney burden was significant ( p
    Figure Legend Snippet: Dose-response of Compounds 3 and 6 in vivo. Female CD.1 mice ( n = 5 per group) were infected IV with 3.5 × 10 4 cfu/mouse before being treated 2 h later with a single SC injection of increasing concentrations of Compound 3 or 6 for 24 h. As a positive control, female Swiss CD.1 mice were also gavaged at 2 h with 20 mg/kg fluconazole for 24 h. Data are presented as log reduction in kidney burden (±SEM) from the mean cfu of untreated mice ( n = 5). The dose-dependent reduction in kidney burden was significant ( p

    Techniques Used: In Vivo, Mouse Assay, Infection, Injection, Positive Control

    Comparison of HDP mimetic efficacy at 50% MTD. Mean log 10 reduction of colony forming units (cfu) of Candida albicans following SC treatment with different HDP mimetics at 50% of the maximum tolerated dose (MTD) 2 h following IV infection with 3.5 × 10 4 cfu/mouse for 24 h. For comparison, female Swiss CD.1 mice were gavaged 2 h after infection with 20 mg/kg fluconazole for 24 h, which caused a 3-log reduction in cfu. Groups were compared to a control group dosed with kleptose and infected with 3 × 10 4 cfu C. albicans per mouse.
    Figure Legend Snippet: Comparison of HDP mimetic efficacy at 50% MTD. Mean log 10 reduction of colony forming units (cfu) of Candida albicans following SC treatment with different HDP mimetics at 50% of the maximum tolerated dose (MTD) 2 h following IV infection with 3.5 × 10 4 cfu/mouse for 24 h. For comparison, female Swiss CD.1 mice were gavaged 2 h after infection with 20 mg/kg fluconazole for 24 h, which caused a 3-log reduction in cfu. Groups were compared to a control group dosed with kleptose and infected with 3 × 10 4 cfu C. albicans per mouse.

    Techniques Used: Infection, Mouse Assay

    24-h treatment of C. albicans disseminated candidiasis with Compound 6 in single and split doses compared with fluconazole. Mean log 10 reduction of colony forming units (cfu) of Candida albicans following SC treatment with Compound 6 at 50% of the maximum tolerated dose (MTD), 40 mg/kg, 2 h following IV infection with 3.5 × 10 4 cfu/mouse for 24 h. Split doses of 20 mg/kg were given twice at 2 and 4 h and 2 and 6 h after infection. Groups were compared to a control group dosed with kleptose and infected with 3 × 10 4 cfu C. albicans per mouse. For comparison, female Swiss CD.1 mice were gavaged 2 h after infection with 20 mg/kg fluconazole for 24 h.
    Figure Legend Snippet: 24-h treatment of C. albicans disseminated candidiasis with Compound 6 in single and split doses compared with fluconazole. Mean log 10 reduction of colony forming units (cfu) of Candida albicans following SC treatment with Compound 6 at 50% of the maximum tolerated dose (MTD), 40 mg/kg, 2 h following IV infection with 3.5 × 10 4 cfu/mouse for 24 h. Split doses of 20 mg/kg were given twice at 2 and 4 h and 2 and 6 h after infection. Groups were compared to a control group dosed with kleptose and infected with 3 × 10 4 cfu C. albicans per mouse. For comparison, female Swiss CD.1 mice were gavaged 2 h after infection with 20 mg/kg fluconazole for 24 h.

    Techniques Used: Infection, Mouse Assay

    20) Product Images from "Effect of Acetic Acid and Lactic Acid at Low pH in Growth and Azole Resistance of Candida albicans and Candida glabrata"

    Article Title: Effect of Acetic Acid and Lactic Acid at Low pH in Growth and Azole Resistance of Candida albicans and Candida glabrata

    Journal: Frontiers in Microbiology

    doi: 10.3389/fmicb.2018.03265

    Heat map representing the inhibitory effect of lactic acid and clotrimazole, fluconazole, miconazole, and tioconazole against C. albicans, C. glabrata BG2 and C. glabrata CBS138 in MM medium (at pH 4). To assess an eventual synergistic effect of lactic acid with the different azoles in determining growth inhibition of Candida spp., growth of the different strains in MM medium (at pH4) in the presence of different concentrations of acetic acid and of the different azoles was assessed based on the OD600nm after 24 h of incubation in the presence of the acids, as detailed in Materials and Methods. Synergism between lactic acid and an azole was only considered when growth inhibition achieved in the presence of the acid and of the azole was, at least, 50% higher the inhibition observed in the presence of the azole alone or of the acid alone. Conditions where the acid and an azole were found to synergize are represented by an arrow. Details on the results obtained in those conditions where azoles and lactic acid were found to synergize are provided in Supplementary Figure S7 .
    Figure Legend Snippet: Heat map representing the inhibitory effect of lactic acid and clotrimazole, fluconazole, miconazole, and tioconazole against C. albicans, C. glabrata BG2 and C. glabrata CBS138 in MM medium (at pH 4). To assess an eventual synergistic effect of lactic acid with the different azoles in determining growth inhibition of Candida spp., growth of the different strains in MM medium (at pH4) in the presence of different concentrations of acetic acid and of the different azoles was assessed based on the OD600nm after 24 h of incubation in the presence of the acids, as detailed in Materials and Methods. Synergism between lactic acid and an azole was only considered when growth inhibition achieved in the presence of the acid and of the azole was, at least, 50% higher the inhibition observed in the presence of the azole alone or of the acid alone. Conditions where the acid and an azole were found to synergize are represented by an arrow. Details on the results obtained in those conditions where azoles and lactic acid were found to synergize are provided in Supplementary Figure S7 .

    Techniques Used: Inhibition, Incubation

    Heat map representing the inhibitory effect of acetic acid and clotrimazole, fluconazole, miconazole, and tioconazole against C. albicans, C. glabrata BG2 and C. glabrata CBS138 in MM medium (at pH 4). To assess an eventual synergistic effect of acetic acids with the different azoles in determining growth inhibition of Candida spp., growth of the different strains in MM medium (at pH4) in the presence of different concentrations of acetic acid and of the different azoles was assessed based on the OD600nm after 24 h of incubation in the presence of the acids, as detailed in Materials and Methods. Synergism between acetic acid and an azole was only considered when growth inhibition achieved in the presence of the acid and of the azole was, at least, 50% higher the inhibition observed in the presence of the azole alone or of the acid alone. Conditions where the acid and an azole were found to synergize are represented by an arrow. Details on the results obtained in those conditions where azoles and acetic acid were found to synergize are provided in Supplementary Figure S7 ( * p -value below 0.05; ** p -value below 0.01; *** p -value below 0.001; **** p -value below 0.0001).
    Figure Legend Snippet: Heat map representing the inhibitory effect of acetic acid and clotrimazole, fluconazole, miconazole, and tioconazole against C. albicans, C. glabrata BG2 and C. glabrata CBS138 in MM medium (at pH 4). To assess an eventual synergistic effect of acetic acids with the different azoles in determining growth inhibition of Candida spp., growth of the different strains in MM medium (at pH4) in the presence of different concentrations of acetic acid and of the different azoles was assessed based on the OD600nm after 24 h of incubation in the presence of the acids, as detailed in Materials and Methods. Synergism between acetic acid and an azole was only considered when growth inhibition achieved in the presence of the acid and of the azole was, at least, 50% higher the inhibition observed in the presence of the azole alone or of the acid alone. Conditions where the acid and an azole were found to synergize are represented by an arrow. Details on the results obtained in those conditions where azoles and acetic acid were found to synergize are provided in Supplementary Figure S7 ( * p -value below 0.05; ** p -value below 0.01; *** p -value below 0.001; **** p -value below 0.0001).

    Techniques Used: Inhibition, Incubation

    Susceptibility of C. glabrata (A) and C. albicans (B) strains to acetic acid, to fluconazole or to a combination of fluconazole and acetic acid. This picture depicts the final OD600nm of cultures of the different strains used in our study after 24 h of cultivation in RPMI medium supplemented with acetic acid (40 or 60 mM, at pH 4) or in this same medium supplemented with fluconazole (64 mg/L) or with fluconazole and acetic acid ( * p -value below 0.05; ** p -value below 0.01; *** p -value below 0.001; **** p -value below 0.0001).
    Figure Legend Snippet: Susceptibility of C. glabrata (A) and C. albicans (B) strains to acetic acid, to fluconazole or to a combination of fluconazole and acetic acid. This picture depicts the final OD600nm of cultures of the different strains used in our study after 24 h of cultivation in RPMI medium supplemented with acetic acid (40 or 60 mM, at pH 4) or in this same medium supplemented with fluconazole (64 mg/L) or with fluconazole and acetic acid ( * p -value below 0.05; ** p -value below 0.01; *** p -value below 0.001; **** p -value below 0.0001).

    Techniques Used:

    21) Product Images from "Synergistic antifungal effect of cyclized chalcone derivatives and fluconazole against Candida albicans"

    Article Title: Synergistic antifungal effect of cyclized chalcone derivatives and fluconazole against Candida albicans

    Journal: MedChemComm

    doi: 10.1039/c7md00440k

    Representative isobolograms for the synergistic interactions in nine different ratios of 6B with fluconazole (A) against fluconazole susceptible C. albicans 0079gr and 5B with fluconazole against fluconazole resistant C. albicans 167–2.
    Figure Legend Snippet: Representative isobolograms for the synergistic interactions in nine different ratios of 6B with fluconazole (A) against fluconazole susceptible C. albicans 0079gr and 5B with fluconazole against fluconazole resistant C. albicans 167–2.

    Techniques Used:

    Energy dependent R6G efflux in fluconazole susceptible C. albicans ATCC90028 cells (open symbols) and in fluconazole-resistant C. albicans A71 (filled symbols), incubated either with R6G (10 μM) alone (squares) or with R6G plus mono- and bis-chalcone derivatives. All mono-analogues are represented by circles while all bis-analogues are represented by triangles.
    Figure Legend Snippet: Energy dependent R6G efflux in fluconazole susceptible C. albicans ATCC90028 cells (open symbols) and in fluconazole-resistant C. albicans A71 (filled symbols), incubated either with R6G (10 μM) alone (squares) or with R6G plus mono- and bis-chalcone derivatives. All mono-analogues are represented by circles while all bis-analogues are represented by triangles.

    Techniques Used: Incubation

    22) Product Images from "Fluconazole Alters the Polysaccharide Capsule of Cryptococcus gattii and Leads to Distinct Behaviors in Murine Cryptococcosis"

    Article Title: Fluconazole Alters the Polysaccharide Capsule of Cryptococcus gattii and Leads to Distinct Behaviors in Murine Cryptococcosis

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0112669

    Reduced susceptibility to fluconazole leads to very low CAP59 and CAP64 gene expression and affects physical properties of the GXM capsule. Microscopic visualization with India ink stain of L27/01 ( A ) and L27/01 F strains ( B ). Capsule size of L27/01 and L27/01 F strains ( C ). SEM analysis of L27/01 ( D ) and L27/01 F ( E ) strains. Zeta potential of capsular and secreted PS of cells ( F ). Cell suspensions were analyzed with an AXIOPLAN (Carl Zeiss) fluorescence microscope. Images were processed using ImageJ. Staining of L27/01 ( G ) and L27/01 F ( H ) strains with mAb 18B7 showing differences in epitope presentation in the PS capsule between strains. Ratio of gene expression of CAP59 , CAP64 and UXS1 to actin ( I ). Size distribution of PS fibers from capsular ( J and K ) and exo-PS samples ( L and M ) of L27/01 and L27/01 F strains, respectively. Bar = 10 µm (A) and (D). Bar = 5 µm (B), (E), (G) and (H). *P
    Figure Legend Snippet: Reduced susceptibility to fluconazole leads to very low CAP59 and CAP64 gene expression and affects physical properties of the GXM capsule. Microscopic visualization with India ink stain of L27/01 ( A ) and L27/01 F strains ( B ). Capsule size of L27/01 and L27/01 F strains ( C ). SEM analysis of L27/01 ( D ) and L27/01 F ( E ) strains. Zeta potential of capsular and secreted PS of cells ( F ). Cell suspensions were analyzed with an AXIOPLAN (Carl Zeiss) fluorescence microscope. Images were processed using ImageJ. Staining of L27/01 ( G ) and L27/01 F ( H ) strains with mAb 18B7 showing differences in epitope presentation in the PS capsule between strains. Ratio of gene expression of CAP59 , CAP64 and UXS1 to actin ( I ). Size distribution of PS fibers from capsular ( J and K ) and exo-PS samples ( L and M ) of L27/01 and L27/01 F strains, respectively. Bar = 10 µm (A) and (D). Bar = 5 µm (B), (E), (G) and (H). *P

    Techniques Used: Expressing, Staining, Fluorescence, Microscopy

    Reduced susceptibility to fluconazole impairs C. gattii migration to the CNS. Lungs ( A ), brain ( B ), and Bronchoalveolar lavage fluid (BALF) ( C ) were removed (n = 8), homogenized, diluted and plated onto Sabouraud dextrose agar for measurement of fungal burden 0.5, 1, 7, 15, and 30 days post-infection with 10 6 cells of L27/01 or L27/01 F strains. ND: non-detected. Black bars refer to L27/01 and grey bars refer to L27/01 F . *Data represent the mean of two independent experiments in triplicate. P
    Figure Legend Snippet: Reduced susceptibility to fluconazole impairs C. gattii migration to the CNS. Lungs ( A ), brain ( B ), and Bronchoalveolar lavage fluid (BALF) ( C ) were removed (n = 8), homogenized, diluted and plated onto Sabouraud dextrose agar for measurement of fungal burden 0.5, 1, 7, 15, and 30 days post-infection with 10 6 cells of L27/01 or L27/01 F strains. ND: non-detected. Black bars refer to L27/01 and grey bars refer to L27/01 F . *Data represent the mean of two independent experiments in triplicate. P

    Techniques Used: Migration, Infection

    L27/01 developed reduced susceptibility to fluconazole with overexpression of the MDR1 gene, but the parental strain is more virulent than L27/01 F in mice. Pulsed field gel electrophoresis (PFGE) patterns of L27/01 F and L27/01 and strains on 1% PFGE certified agarose (lanes 2 and 3, respectively). Lane 1 show PFGE Size Marker (SM), 0.225–2.2 Mb S. cerevisiae chromosomal DNA Saccharomyces cerevisiae used as size standard ( A ). MDR1 gene expression in L27/01 and L27/01 F strains ( B ). Ergosterol levels of L27/01 and L27/01 F strains ( C ). Urease and Laccase activities of L27/01 and L27/01 F strains: one enzymatic unit was calculated per 10 5 and 10 8 cells of C. gattii for urease and laccase activity, respectively, after 24 hours ( D ). Eight mice per group were inoculated by the intratracheal route with 10 6 cells of L27/01 or L27/01 F . Animals administered with PBS represent the non-infected (control) group. Animals were monitored daily. Survival curve ( E ). Weight variation (Score) ( F ). Each 1 (one) gram received or lost corresponds to 10 points added or subtracted, respectively. *The vertical dotted line indicates a significant difference from the appointed day. CFU/g of lungs ( G ) from animals infected with 1×10 6 cells of L27/01 or L27/01 F strains treated i.p. with fluconazole at 10 mg/kg/day (n = 6). NI: not infected group. NT: non treated group. Black bars refer to L27/01 (▪) and grey bars refer to L27/01 F (▴). Data represent the mean of three independent experiments in triplicate. *P
    Figure Legend Snippet: L27/01 developed reduced susceptibility to fluconazole with overexpression of the MDR1 gene, but the parental strain is more virulent than L27/01 F in mice. Pulsed field gel electrophoresis (PFGE) patterns of L27/01 F and L27/01 and strains on 1% PFGE certified agarose (lanes 2 and 3, respectively). Lane 1 show PFGE Size Marker (SM), 0.225–2.2 Mb S. cerevisiae chromosomal DNA Saccharomyces cerevisiae used as size standard ( A ). MDR1 gene expression in L27/01 and L27/01 F strains ( B ). Ergosterol levels of L27/01 and L27/01 F strains ( C ). Urease and Laccase activities of L27/01 and L27/01 F strains: one enzymatic unit was calculated per 10 5 and 10 8 cells of C. gattii for urease and laccase activity, respectively, after 24 hours ( D ). Eight mice per group were inoculated by the intratracheal route with 10 6 cells of L27/01 or L27/01 F . Animals administered with PBS represent the non-infected (control) group. Animals were monitored daily. Survival curve ( E ). Weight variation (Score) ( F ). Each 1 (one) gram received or lost corresponds to 10 points added or subtracted, respectively. *The vertical dotted line indicates a significant difference from the appointed day. CFU/g of lungs ( G ) from animals infected with 1×10 6 cells of L27/01 or L27/01 F strains treated i.p. with fluconazole at 10 mg/kg/day (n = 6). NI: not infected group. NT: non treated group. Black bars refer to L27/01 (▪) and grey bars refer to L27/01 F (▴). Data represent the mean of three independent experiments in triplicate. *P

    Techniques Used: Over Expression, Mouse Assay, Pulsed-Field Gel, Electrophoresis, Marker, Expressing, Activity Assay, Infection

    Synopsis of the methodology. Fluconazole-resistant strain selection ( A–F ). After determining the MIC of fluconazole on SDA, an average of five colonies obtained from the highest fluconazole concentration were selected for culture on SDA plates supplemented with this drug. L27/01 ( A ) was the strain able to grow at the highest fluconazole concentration ( B ) and was chosen for culture on SDA plates supplemented with this drug. This strain was cultured in solid medium with increasing concentrations ( B–C ) of fluconazole until growth ceased at 100 µg/mL ( D ). To verify the maintenance of resistance to fluconazole and cross-resistance between this drug and amphotericin B, the selected strain was cultured in SDA without drug every 48 h 170 times ( E ), and the MIC test was performed by microdilution every 5 subcultures ( F ). Colonies grown at 95 µg/mL were maintained in this concentration, and the strain was named “L27/01 F ” ( G ). L27/01 strain grew in the absence of drug ( H ). The genetic similarity between L27/01 and L27/01 F strains was evaluated by randomly amplified polymorphic DNA (RAPD)-PCR, PFGE and ITS Sequencing. CAP59, CAP64, ARF-1, ERG11, UXS-1 levels by real-time PCR were evaluated. Lipid evaluation was performed to compare the ergosterol content of L27/01 and L27/01 F cell membranes. Also, Urease and Laccase activities of L27/01 and L27/01 F strains were determined ( I ). Evaluation of murine cryptococcosis after inoculation with L27/01 or L27/01 F strain: survival curve ( J ). Investigation of whether fluconazole affects the polysaccharide (PS) capsule ( K ). The phagocytosis assay was performed to assess the influence of PS capsules from L27/01 and L27/01 F strains on phagocytosis and intracellular proliferation rate (IPR) in murine peritoneal macrophages from C57BL/6 mice ( L ). Cryptococcal cell dissemination, immune response and behavioral alterations ( M ). SDA: Sabouraud Dextrose Agar. MIC: Minimum inhibitory concentration. PFGE: Pulsed field gel electrophoresis. CFU: Colony forming units. BALF: bronchoalveolar lavage fluid. MPO: Myeloperoxidase activity. i.t.: intratracheal infection. PS: polysaccharide.
    Figure Legend Snippet: Synopsis of the methodology. Fluconazole-resistant strain selection ( A–F ). After determining the MIC of fluconazole on SDA, an average of five colonies obtained from the highest fluconazole concentration were selected for culture on SDA plates supplemented with this drug. L27/01 ( A ) was the strain able to grow at the highest fluconazole concentration ( B ) and was chosen for culture on SDA plates supplemented with this drug. This strain was cultured in solid medium with increasing concentrations ( B–C ) of fluconazole until growth ceased at 100 µg/mL ( D ). To verify the maintenance of resistance to fluconazole and cross-resistance between this drug and amphotericin B, the selected strain was cultured in SDA without drug every 48 h 170 times ( E ), and the MIC test was performed by microdilution every 5 subcultures ( F ). Colonies grown at 95 µg/mL were maintained in this concentration, and the strain was named “L27/01 F ” ( G ). L27/01 strain grew in the absence of drug ( H ). The genetic similarity between L27/01 and L27/01 F strains was evaluated by randomly amplified polymorphic DNA (RAPD)-PCR, PFGE and ITS Sequencing. CAP59, CAP64, ARF-1, ERG11, UXS-1 levels by real-time PCR were evaluated. Lipid evaluation was performed to compare the ergosterol content of L27/01 and L27/01 F cell membranes. Also, Urease and Laccase activities of L27/01 and L27/01 F strains were determined ( I ). Evaluation of murine cryptococcosis after inoculation with L27/01 or L27/01 F strain: survival curve ( J ). Investigation of whether fluconazole affects the polysaccharide (PS) capsule ( K ). The phagocytosis assay was performed to assess the influence of PS capsules from L27/01 and L27/01 F strains on phagocytosis and intracellular proliferation rate (IPR) in murine peritoneal macrophages from C57BL/6 mice ( L ). Cryptococcal cell dissemination, immune response and behavioral alterations ( M ). SDA: Sabouraud Dextrose Agar. MIC: Minimum inhibitory concentration. PFGE: Pulsed field gel electrophoresis. CFU: Colony forming units. BALF: bronchoalveolar lavage fluid. MPO: Myeloperoxidase activity. i.t.: intratracheal infection. PS: polysaccharide.

    Techniques Used: Selection, Concentration Assay, Cell Culture, Amplification, Polymerase Chain Reaction, Sequencing, Real-time Polymerase Chain Reaction, Phagocytosis Assay, Mouse Assay, Pulsed-Field Gel, Electrophoresis, Activity Assay, Infection

    23) Product Images from "Competitive Fitness of Fluconazole-Resistant Clinical Candida albicans Strains"

    Article Title: Competitive Fitness of Fluconazole-Resistant Clinical Candida albicans Strains

    Journal: Antimicrobial Agents and Chemotherapy

    doi: 10.1128/AAC.00584-17

    Competitive fitness of fluconazole-susceptible clinical isolates F2 and 5833 (light gray bars) and genetically engineered derivatives carrying the MRR1 GOF mutations from matched resistant isolates F5 and 6692, respectively (dark gray bars). Two independently generated RFP -labeled derivatives of isolates F2 and 5833 were each mixed with two independently constructed corresponding mutants; i.e., four coculture experiments were performed for each comparison. Shown are the relative proportions of the strains in the inoculum (day 0) and after two rounds of coculture (day 1 and day 2) at 30°C in YPD medium. Results are the means and standard deviations from the four coculture experiments. Significant differences in the proportions of the strains with MRR1 GOF mutations at day 1 and day 2 compared to day 0 are indicated by asterisks ( P
    Figure Legend Snippet: Competitive fitness of fluconazole-susceptible clinical isolates F2 and 5833 (light gray bars) and genetically engineered derivatives carrying the MRR1 GOF mutations from matched resistant isolates F5 and 6692, respectively (dark gray bars). Two independently generated RFP -labeled derivatives of isolates F2 and 5833 were each mixed with two independently constructed corresponding mutants; i.e., four coculture experiments were performed for each comparison. Shown are the relative proportions of the strains in the inoculum (day 0) and after two rounds of coculture (day 1 and day 2) at 30°C in YPD medium. Results are the means and standard deviations from the four coculture experiments. Significant differences in the proportions of the strains with MRR1 GOF mutations at day 1 and day 2 compared to day 0 are indicated by asterisks ( P

    Techniques Used: Generated, Labeling, Construct

    Competitive fitness of fluconazole-susceptible isolates F2, B3, 5833, and TW1 (light gray bars) and matched fluconazole-resistant isolates F5, B4, 6692, and TW17 (dark gray bars), respectively, in a mouse model of gastrointestinal colonization. In each case, the susceptible isolate was mixed with two independently generated RFP -labeled derivatives of the matched resistant isolate and the resistant isolate was mixed with two independently generated RFP -labeled derivatives of the matched susceptible isolate. For all comparisons, each of the four mixtures was used to infect 3 mice (4 mice were infected with the 5833/6692ADH1R1A and TW1/TW17ADH1R1A pairs); i.e., the competitive fitness of each isolate pair was tested in 12 mice (13 mice for 5833/6692 and TW1/TW17). Shown are the relative proportions of the strains in the inoculum (day 0) and in samples recovered at the indicated times from the feces of the animals. Results are the means and standard deviations from the 12 (or 13) coinfection experiments. Significant differences in the proportions of resistant isolates after passage through the gastrointestinal tract compared to the inoculum are indicated by asterisks ( P
    Figure Legend Snippet: Competitive fitness of fluconazole-susceptible isolates F2, B3, 5833, and TW1 (light gray bars) and matched fluconazole-resistant isolates F5, B4, 6692, and TW17 (dark gray bars), respectively, in a mouse model of gastrointestinal colonization. In each case, the susceptible isolate was mixed with two independently generated RFP -labeled derivatives of the matched resistant isolate and the resistant isolate was mixed with two independently generated RFP -labeled derivatives of the matched susceptible isolate. For all comparisons, each of the four mixtures was used to infect 3 mice (4 mice were infected with the 5833/6692ADH1R1A and TW1/TW17ADH1R1A pairs); i.e., the competitive fitness of each isolate pair was tested in 12 mice (13 mice for 5833/6692 and TW1/TW17). Shown are the relative proportions of the strains in the inoculum (day 0) and in samples recovered at the indicated times from the feces of the animals. Results are the means and standard deviations from the 12 (or 13) coinfection experiments. Significant differences in the proportions of resistant isolates after passage through the gastrointestinal tract compared to the inoculum are indicated by asterisks ( P

    Techniques Used: Generated, Labeling, Mouse Assay, Infection

    Competitive fitness of matched pairs of fluconazole-resistant isolates (dark gray bars) and fluconazole-susceptible isolates (light gray bars) from the same patients. In each case, the susceptible isolate was mixed with two independently generated RFP -labeled derivatives of the matched resistant isolate and the resistant isolate was mixed with two independently generated RFP -labeled derivatives of the matched susceptible isolate. Shown are the relative proportions of the two strains in the inoculum (day 0) and after two rounds of coculture (day 1 and day 2) at 30°C in YPD medium. Results are the means and standard deviations for the four cocultures. For isolate pair F2/F5, each combination was tested four times; i.e., results are from 16 cocultures. Significant differences in the proportions of the resistant isolates at day 1 and day 2 compared to day 0 are indicated by asterisks ( P
    Figure Legend Snippet: Competitive fitness of matched pairs of fluconazole-resistant isolates (dark gray bars) and fluconazole-susceptible isolates (light gray bars) from the same patients. In each case, the susceptible isolate was mixed with two independently generated RFP -labeled derivatives of the matched resistant isolate and the resistant isolate was mixed with two independently generated RFP -labeled derivatives of the matched susceptible isolate. Shown are the relative proportions of the two strains in the inoculum (day 0) and after two rounds of coculture (day 1 and day 2) at 30°C in YPD medium. Results are the means and standard deviations for the four cocultures. For isolate pair F2/F5, each combination was tested four times; i.e., results are from 16 cocultures. Significant differences in the proportions of the resistant isolates at day 1 and day 2 compared to day 0 are indicated by asterisks ( P

    Techniques Used: Generated, Labeling

    24) Product Images from "Acylhydrazones as Antifungal Agents Targeting the Synthesis of Fungal Sphingolipids"

    Article Title: Acylhydrazones as Antifungal Agents Targeting the Synthesis of Fungal Sphingolipids

    Journal: Antimicrobial Agents and Chemotherapy

    doi: 10.1128/AAC.00156-18

    Effect of 20 mg/kg/day oral administration against cryptococcosis, candidiasis, and aspergillosis. (A) Survival of mice infected intranasally with 5 × 10 5 C. neoformans cells and treated with fluconazole, BHBM, or D13. *, D13 versus no drug, P value of 0.0018; #, D3 versus BHBM, P value of 0.0057. (B) Survival of mice infected intravenously with 10 4 C. albicans cells and treated with fluconazole, BHBM, or D13. *, D13 versus no drug, P value of 0.0004; #, BHBM versus no drug, P value of 0.015. (C) Survival of mice infected intranasally with 2 × 10 4 conidia of A. fumigatus and treated with voriconazole, BHBM, or D13. *, D13 versus no drug, P value of
    Figure Legend Snippet: Effect of 20 mg/kg/day oral administration against cryptococcosis, candidiasis, and aspergillosis. (A) Survival of mice infected intranasally with 5 × 10 5 C. neoformans cells and treated with fluconazole, BHBM, or D13. *, D13 versus no drug, P value of 0.0018; #, D3 versus BHBM, P value of 0.0057. (B) Survival of mice infected intravenously with 10 4 C. albicans cells and treated with fluconazole, BHBM, or D13. *, D13 versus no drug, P value of 0.0004; #, BHBM versus no drug, P value of 0.015. (C) Survival of mice infected intranasally with 2 × 10 4 conidia of A. fumigatus and treated with voriconazole, BHBM, or D13. *, D13 versus no drug, P value of

    Techniques Used: Mouse Assay, Infection

    25) Product Images from "NSG2 (ORF19.273) Encoding Protein Controls Sensitivity of Candida albicans to Azoles through Regulating the Synthesis of C14-Methylated Sterols"

    Article Title: NSG2 (ORF19.273) Encoding Protein Controls Sensitivity of Candida albicans to Azoles through Regulating the Synthesis of C14-Methylated Sterols

    Journal: Frontiers in Microbiology

    doi: 10.3389/fmicb.2018.00218

    Determination of sterols in NSG2 -deficient strains by gas chromatography–mass spectrometry (GC–MS). (A) Sterol synthesis pathway in C. albicans . The red box is on behalf of accumulated sterols in nsg2 ). (B) Sterol compositions of SN152, nsg2 Δ/Δ and nsg2 Δ/Δ+NSG2 which were cultured in YPD medium for 16 h and extracted by NaOH agents. (C) Sterol compositions of SN152, nsg2 Δ/Δ and nsg2 Δ/Δ+NSG2 which were treated with 8 μg/ml of fluconazole for 6 h. The percentage of each sterol is calculated by all detected sterols in each sample. ∗ p
    Figure Legend Snippet: Determination of sterols in NSG2 -deficient strains by gas chromatography–mass spectrometry (GC–MS). (A) Sterol synthesis pathway in C. albicans . The red box is on behalf of accumulated sterols in nsg2 ). (B) Sterol compositions of SN152, nsg2 Δ/Δ and nsg2 Δ/Δ+NSG2 which were cultured in YPD medium for 16 h and extracted by NaOH agents. (C) Sterol compositions of SN152, nsg2 Δ/Δ and nsg2 Δ/Δ+NSG2 which were treated with 8 μg/ml of fluconazole for 6 h. The percentage of each sterol is calculated by all detected sterols in each sample. ∗ p

    Techniques Used: Gas Chromatography, Mass Spectrometry, Gas Chromatography-Mass Spectrometry, Cell Culture

    NSG2 -deficient strains were sensitive to fluconazole in vivo . (A) Survival curves of BALB/C mice infected by SN152, nsg2 Δ/Δ or nsg2 Δ/Δ+NSG2 strains. A total of 3 × 10 5 CFU cells were injected to female BALB/C (6–8 weeks) through tail vein and observed until all mice dead. (B) Kidney CFU assay in mice with systemic candidiasis after infected with 3 × 10 5 CFU after 2 days. (C) Survival curves of BALB/C mice infected by SN152, nsg2 Δ/Δ or nsg2 Δ/Δ+NSG2 and treated with 2 mg/kg fluconazole for 1 week, ∗∗ p
    Figure Legend Snippet: NSG2 -deficient strains were sensitive to fluconazole in vivo . (A) Survival curves of BALB/C mice infected by SN152, nsg2 Δ/Δ or nsg2 Δ/Δ+NSG2 strains. A total of 3 × 10 5 CFU cells were injected to female BALB/C (6–8 weeks) through tail vein and observed until all mice dead. (B) Kidney CFU assay in mice with systemic candidiasis after infected with 3 × 10 5 CFU after 2 days. (C) Survival curves of BALB/C mice infected by SN152, nsg2 Δ/Δ or nsg2 Δ/Δ+NSG2 and treated with 2 mg/kg fluconazole for 1 week, ∗∗ p

    Techniques Used: In Vivo, Mouse Assay, Infection, Injection, Colony-forming Unit Assay

    The integrity and functions of cell membrane by NSG2 disruption. (A) Ultra-structure images of Candida albicans SN152, nsg2 Δ/Δ and nsg2 Δ/Δ+NSG2 strains in the presence or absence of 8 μg/ml of fluconazloe for 16 h. Solid arrow indicated the slight damage in the membrane of nsg2 Δ/Δ and the dashed arrow indicated the extensive solubilization of the cytoplasmic membrane in the presence of 8 μg/ml of fluconazole. The magnification was indicated at the bottom of each picture. (B) Representative flow cytometric profiles and graph showing the proportions of PI + -gated cells in SN152, nsg2 Δ/Δ and nsg2 Δ/Δ+NSG2 treated with fluconazoles or not. Cells were treated with fluconazoles for 8 h and stained by 2 μM PI for 50 min. ∗∗ p
    Figure Legend Snippet: The integrity and functions of cell membrane by NSG2 disruption. (A) Ultra-structure images of Candida albicans SN152, nsg2 Δ/Δ and nsg2 Δ/Δ+NSG2 strains in the presence or absence of 8 μg/ml of fluconazloe for 16 h. Solid arrow indicated the slight damage in the membrane of nsg2 Δ/Δ and the dashed arrow indicated the extensive solubilization of the cytoplasmic membrane in the presence of 8 μg/ml of fluconazole. The magnification was indicated at the bottom of each picture. (B) Representative flow cytometric profiles and graph showing the proportions of PI + -gated cells in SN152, nsg2 Δ/Δ and nsg2 Δ/Δ+NSG2 treated with fluconazoles or not. Cells were treated with fluconazoles for 8 h and stained by 2 μM PI for 50 min. ∗∗ p

    Techniques Used: Flow Cytometry, Staining

    26) Product Images from "A Systematic Screen Reveals a Diverse Collection of Medications That Induce Antifungal Resistance in Candida Species"

    Article Title: A Systematic Screen Reveals a Diverse Collection of Medications That Induce Antifungal Resistance in Candida Species

    Journal: Antimicrobial Agents and Chemotherapy

    doi: 10.1128/AAC.00054-19

    Several antagonistic compounds elevate Candida albicans fluconazole resistance at submicromolar concentrations. Checkerboard assays were performed in RPMI medium with C. albicans (strain SC5314) across a range of fluconazole doses and of (A) aripiprazole, (B) etofenamate, (C) mometasone, (D) penbutolol, (E) pinaverium, (F) prenylamine, and (G) thiethylperazine. After 24 h of incubation at 35°C, the growth in each well was quantified as OD 600 , normalized to the untreated control well, and data are presented as heat maps. Data represent averages of results from technical duplicates from a single experiment representative of two biological replicates.
    Figure Legend Snippet: Several antagonistic compounds elevate Candida albicans fluconazole resistance at submicromolar concentrations. Checkerboard assays were performed in RPMI medium with C. albicans (strain SC5314) across a range of fluconazole doses and of (A) aripiprazole, (B) etofenamate, (C) mometasone, (D) penbutolol, (E) pinaverium, (F) prenylamine, and (G) thiethylperazine. After 24 h of incubation at 35°C, the growth in each well was quantified as OD 600 , normalized to the untreated control well, and data are presented as heat maps. Data represent averages of results from technical duplicates from a single experiment representative of two biological replicates.

    Techniques Used: Incubation

    There is significant overlap in the azole antagonistic compounds from the C. albicans , C. parapsilosis , and C. tropicalis screens. The number of common and distinct compounds identified as hits from the fluconazole screens with these three species at 1, 25, and 1.6 μM, respectively, is illustrated in the Venn diagram.
    Figure Legend Snippet: There is significant overlap in the azole antagonistic compounds from the C. albicans , C. parapsilosis , and C. tropicalis screens. The number of common and distinct compounds identified as hits from the fluconazole screens with these three species at 1, 25, and 1.6 μM, respectively, is illustrated in the Venn diagram.

    Techniques Used:

    A variety of approved medications substantially diminish Candida albicans susceptibility to fluconazole. C. albicans (strain SC5314) susceptibility to fluconazole was tested in the absence (DMSO vehicle alone) or presence of 5 μM of the indicated compounds in RPMI medium according to the CLSI broth microdilution protocol. Growth was quantified as OD 600 after 24 h of incubation at 35°C, and data are presented as percent growth relative to the drug-free control. Data represent averages of results from technical duplicates from a single experiment representative of two biological replicates.
    Figure Legend Snippet: A variety of approved medications substantially diminish Candida albicans susceptibility to fluconazole. C. albicans (strain SC5314) susceptibility to fluconazole was tested in the absence (DMSO vehicle alone) or presence of 5 μM of the indicated compounds in RPMI medium according to the CLSI broth microdilution protocol. Growth was quantified as OD 600 after 24 h of incubation at 35°C, and data are presented as percent growth relative to the drug-free control. Data represent averages of results from technical duplicates from a single experiment representative of two biological replicates.

    Techniques Used: Incubation

    Aripiprazole induces transcription of the CDR1 and CDR2 genes in C. albicans . SC5314 was grown in YPD medium buffered with MOPS and adjusted to pH 7 in the presence of the indicated concentrations of drugs or of DMSO vehicle alone before RNA was isolated, and the relative abundance of the CDR1 , CDR2 , and MDR1 transcripts were determined by qRT-PCR analysis. The relative abundances of these transcripts in each sample were normalized to that of the ACT1 transcript and expressed relative to the level seen with the DMSO control (fold expression). Flu, fluconazole; Ari, aripiprazole; Eto, etofenamate. Mean and standard deviations of results from three separate experiments, each performed with three technical replicates, are shown.
    Figure Legend Snippet: Aripiprazole induces transcription of the CDR1 and CDR2 genes in C. albicans . SC5314 was grown in YPD medium buffered with MOPS and adjusted to pH 7 in the presence of the indicated concentrations of drugs or of DMSO vehicle alone before RNA was isolated, and the relative abundance of the CDR1 , CDR2 , and MDR1 transcripts were determined by qRT-PCR analysis. The relative abundances of these transcripts in each sample were normalized to that of the ACT1 transcript and expressed relative to the level seen with the DMSO control (fold expression). Flu, fluconazole; Ari, aripiprazole; Eto, etofenamate. Mean and standard deviations of results from three separate experiments, each performed with three technical replicates, are shown.

    Techniques Used: Isolation, Quantitative RT-PCR, Expressing

    Drug-induced and genetically encoded mechanisms can act in an additive manner to confer azole resistance in Candida albicans . The susceptibility of wild-type C. albicans (SC5314) (A), of strains harboring activated (gain of function—GOF) alleles of MRR1 (B), TAC1 (D), or UPC2 (F), or of isogenic control strains (C, E, and G) to fluconazole was tested in the absence (DMSO vehicle alone) or presence of 5 μM of the indicated compounds in RPMI medium according to the CLSI broth microdilution protocol. Growth was quantified as OD 600 after 24 h of incubation at 35°C, expressed as a percentage of growth in the drug-free control, and presented as heat maps. Data represent averages of results from technical duplicates from a single experiment representative of two biological replicates.
    Figure Legend Snippet: Drug-induced and genetically encoded mechanisms can act in an additive manner to confer azole resistance in Candida albicans . The susceptibility of wild-type C. albicans (SC5314) (A), of strains harboring activated (gain of function—GOF) alleles of MRR1 (B), TAC1 (D), or UPC2 (F), or of isogenic control strains (C, E, and G) to fluconazole was tested in the absence (DMSO vehicle alone) or presence of 5 μM of the indicated compounds in RPMI medium according to the CLSI broth microdilution protocol. Growth was quantified as OD 600 after 24 h of incubation at 35°C, expressed as a percentage of growth in the drug-free control, and presented as heat maps. Data represent averages of results from technical duplicates from a single experiment representative of two biological replicates.

    Techniques Used: Activated Clotting Time Assay, Incubation

    27) Product Images from "Candida glabrata susceptibility to antifungals and phagocytosis is modulated by acetate"

    Article Title: Candida glabrata susceptibility to antifungals and phagocytosis is modulated by acetate

    Journal: Frontiers in Microbiology

    doi: 10.3389/fmicb.2015.00919

    Effect of acetic acid and fluconazole in the expression of ADY2a, ADY2b, FPS1, FPS2 , and ATO3 in C. glabrata ATCC 2001 planktonic (A) and biofilm cells (B) . Cells and biofilms were grown in two different conditions: RPMI pH 5.0 and RPMI 0.5% acetic acid at pH 5.0. Legend displays fluconazole concentrations used. Gene expression was calculated using ΔCT method and normalized to PGK1 gene. Error bars represent SD. ∗ , ∗∗ , and ∗∗∗ means that results are statistically significant ( p
    Figure Legend Snippet: Effect of acetic acid and fluconazole in the expression of ADY2a, ADY2b, FPS1, FPS2 , and ATO3 in C. glabrata ATCC 2001 planktonic (A) and biofilm cells (B) . Cells and biofilms were grown in two different conditions: RPMI pH 5.0 and RPMI 0.5% acetic acid at pH 5.0. Legend displays fluconazole concentrations used. Gene expression was calculated using ΔCT method and normalized to PGK1 gene. Error bars represent SD. ∗ , ∗∗ , and ∗∗∗ means that results are statistically significant ( p

    Techniques Used: Expressing

    Effect of fluconazole in Candida glabrata ATCC2001 pre-formed biofilms. Graph (A) represents the percentage of biomass reduction that was quantified by crystal violet staining and (B) the percentage of colony-forming units (CFUs) reduction. Two conditions were tested: RPMI pH 5.0 and RPMI 0.5% acetic acid at pH 5.0. Error bars represent SD. ∗ and ∗∗∗ means that results are statistically significant ( p
    Figure Legend Snippet: Effect of fluconazole in Candida glabrata ATCC2001 pre-formed biofilms. Graph (A) represents the percentage of biomass reduction that was quantified by crystal violet staining and (B) the percentage of colony-forming units (CFUs) reduction. Two conditions were tested: RPMI pH 5.0 and RPMI 0.5% acetic acid at pH 5.0. Error bars represent SD. ∗ and ∗∗∗ means that results are statistically significant ( p

    Techniques Used: Staining

    28) Product Images from "Identification of Genes Upregulated by the Transcription Factor Bcr1 That Are Involved in Impermeability, Impenetrability, and Drug Resistance of Candida albicansa/? Biofilms"

    Article Title: Identification of Genes Upregulated by the Transcription Factor Bcr1 That Are Involved in Impermeability, Impenetrability, and Drug Resistance of Candida albicansa/? Biofilms

    Journal: Eukaryotic Cell

    doi: 10.1128/EC.00071-13

    Bcr1 plays a fundamental role in conferring upon a/ α biofilms the traits of impermeability, impenetrability, and fluconazole resistance, deemed important in commensalism and pathogenicity. (A to D) Sypro Ruby impermeability (staining). (E to H)
    Figure Legend Snippet: Bcr1 plays a fundamental role in conferring upon a/ α biofilms the traits of impermeability, impenetrability, and fluconazole resistance, deemed important in commensalism and pathogenicity. (A to D) Sypro Ruby impermeability (staining). (E to H)

    Techniques Used: Staining

    Effects of deleting Bcr1-upregulated genes in a/ α biofilms upon impermeability, impenetrability, and fluconazole resistance. (A to H) Impermeability of Sypro Ruby. (I to P) Impenetrability of DiI-stained human polymorphonuclear leukocytes. (Q)
    Figure Legend Snippet: Effects of deleting Bcr1-upregulated genes in a/ α biofilms upon impermeability, impenetrability, and fluconazole resistance. (A to H) Impermeability of Sypro Ruby. (I to P) Impenetrability of DiI-stained human polymorphonuclear leukocytes. (Q)

    Techniques Used: Staining

    Effects of overexpressing Bcr1-upregulated genes in white a/a biofilms upon impermeability, impenetrability, and fluconazole resistance. (A to G) Sypro Ruby impermeability. (H to N) Impenetrability of DiI-stained human polymorphonuclear leukocytes. (O)
    Figure Legend Snippet: Effects of overexpressing Bcr1-upregulated genes in white a/a biofilms upon impermeability, impenetrability, and fluconazole resistance. (A to G) Sypro Ruby impermeability. (H to N) Impenetrability of DiI-stained human polymorphonuclear leukocytes. (O)

    Techniques Used: Staining

    29) Product Images from "Triclosan Demonstrates Synergic Effect with Amphotericin B and Fluconazole and Induces Apoptosis-Like Cell Death in Cryptococcus neoformans"

    Article Title: Triclosan Demonstrates Synergic Effect with Amphotericin B and Fluconazole and Induces Apoptosis-Like Cell Death in Cryptococcus neoformans

    Journal: Frontiers in Microbiology

    doi: 10.3389/fmicb.2016.00360

    The effect of triclosan with amphotericin B and fluconazole. (A,B) Agar disk diffusion assay for triclosan (TCS) in combination with (A) amphotericin B (AMB), or (B) fluconazole (FLC) in C. neoformans H99-containing agar plates. 25 μg of each drug was applied on a disk. (C,D) Agar disk diffusion assay for TCS (1.56, 3.125, and 6.25 μg/ml) in combination with at subinhibitory concentrations (6.25 μg/ml) of (C) AMB and (D) FLU. Diameters of the inhibition zone were measured after 48 h of incubation. Asterisk ∗ indicates partial inhibition. Data shown are representative pictures of three independent experiments. (E,F) Graphs show the inhibition diameter (mm) of C. neoformans plate after triclosan treatment in combination with AMB or FLU. X-axis shows Log concentration of triclosan. Data shown are mean ± SD. ∗ P
    Figure Legend Snippet: The effect of triclosan with amphotericin B and fluconazole. (A,B) Agar disk diffusion assay for triclosan (TCS) in combination with (A) amphotericin B (AMB), or (B) fluconazole (FLC) in C. neoformans H99-containing agar plates. 25 μg of each drug was applied on a disk. (C,D) Agar disk diffusion assay for TCS (1.56, 3.125, and 6.25 μg/ml) in combination with at subinhibitory concentrations (6.25 μg/ml) of (C) AMB and (D) FLU. Diameters of the inhibition zone were measured after 48 h of incubation. Asterisk ∗ indicates partial inhibition. Data shown are representative pictures of three independent experiments. (E,F) Graphs show the inhibition diameter (mm) of C. neoformans plate after triclosan treatment in combination with AMB or FLU. X-axis shows Log concentration of triclosan. Data shown are mean ± SD. ∗ P

    Techniques Used: Diffusion-based Assay, Inhibition, Incubation, Concentration Assay

    30) Product Images from "Characterization of Biofilm Formation and the Role of BCR1 in Clinical Isolates of Candida parapsilosis"

    Article Title: Characterization of Biofilm Formation and the Role of BCR1 in Clinical Isolates of Candida parapsilosis

    Journal: Eukaryotic Cell

    doi: 10.1128/EC.00181-13

    Antifungal susceptibility of C. parapsilosis biofilms. Biofilms formed on SE for 48 h were exposed to amphotericin B, fluconazole, and caspofungin for 48 h at 37°C. The metabolic activity of biofilm cells treated with antimycotics was determined
    Figure Legend Snippet: Antifungal susceptibility of C. parapsilosis biofilms. Biofilms formed on SE for 48 h were exposed to amphotericin B, fluconazole, and caspofungin for 48 h at 37°C. The metabolic activity of biofilm cells treated with antimycotics was determined

    Techniques Used: Activity Assay

    31) Product Images from "Changes in the composition of intestinal fungi and their role in mice with dextran sulfate sodium-induced colitis"

    Article Title: Changes in the composition of intestinal fungi and their role in mice with dextran sulfate sodium-induced colitis

    Journal: Scientific Reports

    doi: 10.1038/srep10416

    Fluconazole (anti-fungal, AF) treatment exacerbates acute DSS-colitis in mice. Six groups of mice were treated as shown in Table 2. (A) Histology scores and (B) colon lengths were measured on day 23 after sacrifice. (C) Weights were measured on days 14–23, and the percent weight change was calculated. The values are expressed as mean ± SEM. (D) Hematoxylin and eosin staining of representative cross-sections of murine distal colon (HE, ×200). Colons of normal, antibiotic cocktail (AB), and AF mice have normal appearances. AB + DSS mice exhibit slight inflammation with a low level of lymphocyte infiltration. DSS mice display mucosal and submucosal inflammation, bowel wall thickening, and a moderate level of lymphocyte infiltration and regeneration with crypt depletion. AF + DSS mice show transmural inflammation, extensive lymphocyte infiltration, and loss of the entire crypt and epithelium. (n = 8–12/group). * P
    Figure Legend Snippet: Fluconazole (anti-fungal, AF) treatment exacerbates acute DSS-colitis in mice. Six groups of mice were treated as shown in Table 2. (A) Histology scores and (B) colon lengths were measured on day 23 after sacrifice. (C) Weights were measured on days 14–23, and the percent weight change was calculated. The values are expressed as mean ± SEM. (D) Hematoxylin and eosin staining of representative cross-sections of murine distal colon (HE, ×200). Colons of normal, antibiotic cocktail (AB), and AF mice have normal appearances. AB + DSS mice exhibit slight inflammation with a low level of lymphocyte infiltration. DSS mice display mucosal and submucosal inflammation, bowel wall thickening, and a moderate level of lymphocyte infiltration and regeneration with crypt depletion. AF + DSS mice show transmural inflammation, extensive lymphocyte infiltration, and loss of the entire crypt and epithelium. (n = 8–12/group). * P

    Techniques Used: Mouse Assay, Staining

    Change in mucosal bacterial compositions by antibiotic cocktail (AB) and fluconazole (AF) treatment. (A) The 16s rDNA levels in the colonic mucosa of AB mice were significantly decreased relative to normal and AF mice. (B) The bacterial Shannon index was significantly increased in the mucosa of AF mice relative to normal and AB mice. (C) Partial least-squares discriminant analysis (PLS-DA) scores plot based on the relative abundance of bacterial OTUs (97% similarity level) in the colonic mucosa of normal, AB, and AF mice. * P
    Figure Legend Snippet: Change in mucosal bacterial compositions by antibiotic cocktail (AB) and fluconazole (AF) treatment. (A) The 16s rDNA levels in the colonic mucosa of AB mice were significantly decreased relative to normal and AF mice. (B) The bacterial Shannon index was significantly increased in the mucosa of AF mice relative to normal and AB mice. (C) Partial least-squares discriminant analysis (PLS-DA) scores plot based on the relative abundance of bacterial OTUs (97% similarity level) in the colonic mucosa of normal, AB, and AF mice. * P

    Techniques Used: Mouse Assay

    32) Product Images from "Synthetic organotelluride compounds induce the reversal of Pdr5p mediated fluconazole resistance in Saccharomyces cerevisiae"

    Article Title: Synthetic organotelluride compounds induce the reversal of Pdr5p mediated fluconazole resistance in Saccharomyces cerevisiae

    Journal: BMC Microbiology

    doi: 10.1186/s12866-014-0201-y

    Evaluation of the reversion of the fluconazole resistance by the organotellurides. (A) AD124567 strain of S. cerevisiae : Fluconazole (−): yeast cell growth on YPD solid in absence of fluconazole. Fluconazole (+): yeast cell growth on YPD solid medium in presence of fluconazole at 120 μg/mL. Medium containing FK506 10 μM + fluconazole 120 μg/mL was used as positive control. (B) Resistant Candida albicans strain (clinical isolate): Fluconazole (−): yeast cell growth on Sabouraud solid medium in absence of fluconazole. Fluconazole (+): yeast cell growth on Sabouraud solid in presence of fluconazole at 64 μg/mL.
    Figure Legend Snippet: Evaluation of the reversion of the fluconazole resistance by the organotellurides. (A) AD124567 strain of S. cerevisiae : Fluconazole (−): yeast cell growth on YPD solid in absence of fluconazole. Fluconazole (+): yeast cell growth on YPD solid medium in presence of fluconazole at 120 μg/mL. Medium containing FK506 10 μM + fluconazole 120 μg/mL was used as positive control. (B) Resistant Candida albicans strain (clinical isolate): Fluconazole (−): yeast cell growth on Sabouraud solid medium in absence of fluconazole. Fluconazole (+): yeast cell growth on Sabouraud solid in presence of fluconazole at 64 μg/mL.

    Techniques Used: Positive Control

    33) Product Images from "Altered Susceptibility of Candida glabrata Bloodstream Isolates to Triazoles at Clinically Relevant pH Values: Comparison of the NCCLS M27-A2, Sensititre YeastOne, and Etest Methods"

    Article Title: Altered Susceptibility of Candida glabrata Bloodstream Isolates to Triazoles at Clinically Relevant pH Values: Comparison of the NCCLS M27-A2, Sensititre YeastOne, and Etest Methods

    Journal: Antimicrobial Agents and Chemotherapy

    doi: 10.1128/AAC.48.11.4441-4443.2004

    Optical density (OD) of microwells at 48 h of fluconazole exposure at the specified pH values against C. glabrata (isolate R).
    Figure Legend Snippet: Optical density (OD) of microwells at 48 h of fluconazole exposure at the specified pH values against C. glabrata (isolate R).

    Techniques Used:

    34) Product Images from "Antifungal Activity of Naphthoquinoidal Compounds In Vitro against Fluconazole-Resistant Strains of Different Candida Species: A Special Emphasis on Mechanisms of Action on Candida tropicalis"

    Article Title: Antifungal Activity of Naphthoquinoidal Compounds In Vitro against Fluconazole-Resistant Strains of Different Candida Species: A Special Emphasis on Mechanisms of Action on Candida tropicalis

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0093698

    An analysis of changes in cell size/granularity (FSC×SSC) to evaluate the effects of fluconazole (64 µg/mL), Ampho (4 µg/mL) and naphthofuranquinone (NFQ) compounds 1, 2 and 3 at concentrations of 32 µg/mL (a); 64 µg/mL (b) and 128 µg/mL (c) on isolates of FLC-resistant C. tropicalis after a 24 h incubation. * p
    Figure Legend Snippet: An analysis of changes in cell size/granularity (FSC×SSC) to evaluate the effects of fluconazole (64 µg/mL), Ampho (4 µg/mL) and naphthofuranquinone (NFQ) compounds 1, 2 and 3 at concentrations of 32 µg/mL (a); 64 µg/mL (b) and 128 µg/mL (c) on isolates of FLC-resistant C. tropicalis after a 24 h incubation. * p

    Techniques Used: Incubation

    Effects of fluconazole (64 µg/mL), Ampho (4 µg/mL) and naphthofuranquinone (NFQ) compounds 1, 2 and 3 at concentrations of 32 µg/mL (a); 64 µg/mL (b) and 128 µg/mL (c) on membrane integrity (as determined by a PI exclusion test) on isolates of FLC-resistant C. tropicalis after 24 h incubation. The population of cells in each lower right quadrant corresponds to the percentage of cells with damaged membranes (PI positive). * p
    Figure Legend Snippet: Effects of fluconazole (64 µg/mL), Ampho (4 µg/mL) and naphthofuranquinone (NFQ) compounds 1, 2 and 3 at concentrations of 32 µg/mL (a); 64 µg/mL (b) and 128 µg/mL (c) on membrane integrity (as determined by a PI exclusion test) on isolates of FLC-resistant C. tropicalis after 24 h incubation. The population of cells in each lower right quadrant corresponds to the percentage of cells with damaged membranes (PI positive). * p

    Techniques Used: Incubation

    An evaluation of ROS formation in fluconazole-sensitive (A) and fluconazole-resistant (B) C. tropicalis isolates after treatment with naphthofuranquinone (NFQ) compounds 1, 2 and 3 using the concentrations 32 µg/mL (a); 64 µg/mL (b) and 128 µg/mL (c). The percentage of ROS formation in the fluconazole-sensitive and fluconazole-resistant C. tropicalis isolates was evaluated for 24 hours. * P
    Figure Legend Snippet: An evaluation of ROS formation in fluconazole-sensitive (A) and fluconazole-resistant (B) C. tropicalis isolates after treatment with naphthofuranquinone (NFQ) compounds 1, 2 and 3 using the concentrations 32 µg/mL (a); 64 µg/mL (b) and 128 µg/mL (c). The percentage of ROS formation in the fluconazole-sensitive and fluconazole-resistant C. tropicalis isolates was evaluated for 24 hours. * P

    Techniques Used:

    An assessment of the mitochondrial membrane potential (Δψm) of fluconazole-resistant C. tropicalis strains. The cells were labeled with Rh123 (50 nM). The graph shows strains incubated for 24 hours with RPMI-1640 (control), with FLC (64 µg/mL) and Ampho (4 µg/mL) and with naphthofuranquinone (NFQ) compounds 1, 2 and 3 at concentrations of 32 µg/mL (a); 64 µg/mL (b) and 128 µg/mL (c) * P
    Figure Legend Snippet: An assessment of the mitochondrial membrane potential (Δψm) of fluconazole-resistant C. tropicalis strains. The cells were labeled with Rh123 (50 nM). The graph shows strains incubated for 24 hours with RPMI-1640 (control), with FLC (64 µg/mL) and Ampho (4 µg/mL) and with naphthofuranquinone (NFQ) compounds 1, 2 and 3 at concentrations of 32 µg/mL (a); 64 µg/mL (b) and 128 µg/mL (c) * P

    Techniques Used: Labeling, Incubation

    35) Product Images from "Commensal fungi recapitulate the protective benefits of intestinal bacteria"

    Article Title: Commensal fungi recapitulate the protective benefits of intestinal bacteria

    Journal: Cell host & microbe

    doi: 10.1016/j.chom.2017.10.013

    Persistent fungal intestinal colonization is required for maintaining their protective benefits (A) Fungal colony forming units (CFUs) in the feces with or without supplementing fluconazole (FLUC) to specific-pathogen free mice previously administered C. albicans (CA) and maintained on drinking water supplemented with an antibiotic cocktail (ABX) containing ampicillin, gentamicin, metronidazole, neomycin, and vancomycin. (B) Percent survival after DSS supplementation in the drinking water (for six days) for mice described in panel (A). (C) Percent survival after influenza A PR8-OVA (Flu A) intranasal infection (6 × 10 4 PFUs) for mice described in panel (A). * P
    Figure Legend Snippet: Persistent fungal intestinal colonization is required for maintaining their protective benefits (A) Fungal colony forming units (CFUs) in the feces with or without supplementing fluconazole (FLUC) to specific-pathogen free mice previously administered C. albicans (CA) and maintained on drinking water supplemented with an antibiotic cocktail (ABX) containing ampicillin, gentamicin, metronidazole, neomycin, and vancomycin. (B) Percent survival after DSS supplementation in the drinking water (for six days) for mice described in panel (A). (C) Percent survival after influenza A PR8-OVA (Flu A) intranasal infection (6 × 10 4 PFUs) for mice described in panel (A). * P

    Techniques Used: Mouse Assay, Infection

    36) Product Images from "Antifungal Activity and Synergism with Azoles of Polish Propolis"

    Article Title: Antifungal Activity and Synergism with Azoles of Polish Propolis

    Journal: Pathogens

    doi: 10.3390/pathogens7020056

    Effect of fluconazole (FLU) in the range of concentration 512–128 μg/mL, caspofungin (CAS) 8–2 μg/mL and amphotericin B (AmB) 8–2 μg/mL on eradication of biofilm formed by ( A ) C. albicans , ( B ) C. glabrata and ( C ) C. krusei clinical isolates from PVC and silicone catheters.
    Figure Legend Snippet: Effect of fluconazole (FLU) in the range of concentration 512–128 μg/mL, caspofungin (CAS) 8–2 μg/mL and amphotericin B (AmB) 8–2 μg/mL on eradication of biofilm formed by ( A ) C. albicans , ( B ) C. glabrata and ( C ) C. krusei clinical isolates from PVC and silicone catheters.

    Techniques Used: Concentration Assay

    37) Product Images from "Dynamic Interaction between Fluconazole and Amphotericin B against Cryptococcus gattii"

    Article Title: Dynamic Interaction between Fluconazole and Amphotericin B against Cryptococcus gattii

    Journal: Antimicrobial Agents and Chemotherapy

    doi: 10.1128/AAC.06098-11

    Time-kill curves of fluconazole and amphotericin B alone or in combination against 14 C. gattii strains. (A) Time-kill curve performed with fluconazole at 1× MIC (●), 2× the MIC (■), and 4× the MIC (▲). (B) Time-kill curve performed with amphotericin B (MIC). (C) Time-kill curve performed with antifungal combinations: FLC (1.0 μg/ml) + AMB (MIC) (♦), FLC (4.0 μg/ml) + AMB (MIC) (■), and FLC (16.0 μg/ml) + AMB (MIC) (▲). AMB, amphotericin B; FLC, fluconazole.
    Figure Legend Snippet: Time-kill curves of fluconazole and amphotericin B alone or in combination against 14 C. gattii strains. (A) Time-kill curve performed with fluconazole at 1× MIC (●), 2× the MIC (■), and 4× the MIC (▲). (B) Time-kill curve performed with amphotericin B (MIC). (C) Time-kill curve performed with antifungal combinations: FLC (1.0 μg/ml) + AMB (MIC) (♦), FLC (4.0 μg/ml) + AMB (MIC) (■), and FLC (16.0 μg/ml) + AMB (MIC) (▲). AMB, amphotericin B; FLC, fluconazole.

    Techniques Used:

    Schematic representation of the reduction in ergosterol content after treatment with fluconazole and its influence on amphotericin B activity. (A) Without fluconazole, the action of amphotericin B depends on the constitutive levels of ergosterol of each strain. (B) Fluconazole at 2 to 4 μg/ml leads to an increasing loss of the ergosterol content and starts to impair the action of amphotericin B but with no visually interference on the fungus killing. (C) Fluconazole at ≥4 μg/ml leads to lower ergosterol levels, and fewer sites for amphotericin B remain, probably leading to the antagonistic interaction.
    Figure Legend Snippet: Schematic representation of the reduction in ergosterol content after treatment with fluconazole and its influence on amphotericin B activity. (A) Without fluconazole, the action of amphotericin B depends on the constitutive levels of ergosterol of each strain. (B) Fluconazole at 2 to 4 μg/ml leads to an increasing loss of the ergosterol content and starts to impair the action of amphotericin B but with no visually interference on the fungus killing. (C) Fluconazole at ≥4 μg/ml leads to lower ergosterol levels, and fewer sites for amphotericin B remain, probably leading to the antagonistic interaction.

    Techniques Used: Activity Assay

    38) Product Images from "Disruption of Microbial Biofilms by an Extracellular Protein Isolated from Epibiotic Tropical Marine Strain of Bacillus licheniformis"

    Article Title: Disruption of Microbial Biofilms by an Extracellular Protein Isolated from Epibiotic Tropical Marine Strain of Bacillus licheniformis

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0064501

    Inhibition of C. albicans biofilms after co-incubation with (a) B. licheniformis BL-DZ1 protein (b) fluconazole. Disruption of pre-formed biofilms by (c) B. licheniformis BL-DZ1 protein (d) fluconazole [* = MIC value].
    Figure Legend Snippet: Inhibition of C. albicans biofilms after co-incubation with (a) B. licheniformis BL-DZ1 protein (b) fluconazole. Disruption of pre-formed biofilms by (c) B. licheniformis BL-DZ1 protein (d) fluconazole [* = MIC value].

    Techniques Used: Inhibition, Incubation

    CLSM analysis of C. albicans biofilms (a) control. After co-incubation with (b) protein BL-DZ1 (c) fluconazole. Disruption of pre-formed biofilms (d) control, after treatment with (e) protein BL-DZ1 (f) fluconazole. Bar indicates 20 µm scale.
    Figure Legend Snippet: CLSM analysis of C. albicans biofilms (a) control. After co-incubation with (b) protein BL-DZ1 (c) fluconazole. Disruption of pre-formed biofilms (d) control, after treatment with (e) protein BL-DZ1 (f) fluconazole. Bar indicates 20 µm scale.

    Techniques Used: Confocal Laser Scanning Microscopy, Incubation

    39) Product Images from "Quantitation of ergosterol content and gene expression profile of ERG11 gene in fluconazole-resistant Candida albicans"

    Article Title: Quantitation of ergosterol content and gene expression profile of ERG11 gene in fluconazole-resistant Candida albicans

    Journal: Current Medical Mycology

    doi: 10.29252/cmm.3.1.13

    Expression analysis of ERG11 gene in fluconazole-susceptible (A) and -resistant (B) C. albicans treated with different concentrations of fluconazole based on MIC (2 × MIC, 1 × MIC, ½ × MIC, and ¼ × MIC) by semi-quantitative RT-PCR. M: 100 bp DNA Ladder, A1: Actin with 2 × MIC concentration of fluconazole, E1: ERG11 with 2 × MIC concentration of fluconazole, C1: Internal control without M-MuLV reverse transcriptase, A2: Actin with 1 × MIC concentration of fluconazole, E2: ERG11 with 1 × MIC concentration of fluconazole, C2: Internal control without M-MuLV reverse transcriptase, A3: Actin with ½ × MIC concentration of fluconazole, E3: ERG11 with ½ × MIC concentration of fluconazole, C3: Internal control without M-MuLV reverse transcriptase, A4: Actin with ¼ × MIC concentration of fluconazole, E4: ERG11 with ¼ × MIC concentration of fluconazole, C4: Internal control without M-MuLV reverse transcriptase, A5: Actin without fluconazole (untreated control), E5: ERG11 without fluconazole (untreated control), C5: Internal control without M-MuLV reverse transcriptase, Co: Control negative for PCR.
    Figure Legend Snippet: Expression analysis of ERG11 gene in fluconazole-susceptible (A) and -resistant (B) C. albicans treated with different concentrations of fluconazole based on MIC (2 × MIC, 1 × MIC, ½ × MIC, and ¼ × MIC) by semi-quantitative RT-PCR. M: 100 bp DNA Ladder, A1: Actin with 2 × MIC concentration of fluconazole, E1: ERG11 with 2 × MIC concentration of fluconazole, C1: Internal control without M-MuLV reverse transcriptase, A2: Actin with 1 × MIC concentration of fluconazole, E2: ERG11 with 1 × MIC concentration of fluconazole, C2: Internal control without M-MuLV reverse transcriptase, A3: Actin with ½ × MIC concentration of fluconazole, E3: ERG11 with ½ × MIC concentration of fluconazole, C3: Internal control without M-MuLV reverse transcriptase, A4: Actin with ¼ × MIC concentration of fluconazole, E4: ERG11 with ¼ × MIC concentration of fluconazole, C4: Internal control without M-MuLV reverse transcriptase, A5: Actin without fluconazole (untreated control), E5: ERG11 without fluconazole (untreated control), C5: Internal control without M-MuLV reverse transcriptase, Co: Control negative for PCR.

    Techniques Used: Expressing, Quantitative RT-PCR, Concentration Assay, Polymerase Chain Reaction

    Percent ergosterol levels of fluconazole-susceptible and -resistant C. albicans after 24 h of treatment with different concentrations of fluconazole based on MIC (2 × MIC, 1 × MIC, ½ × MIC, and ¼ × MIC). Data are means with standard error from three independent experiments in triplicate assays. Statistically significant differences between the treatment isolates and controls are indicated with an asterisk ( P ≤ 0.05 ).
    Figure Legend Snippet: Percent ergosterol levels of fluconazole-susceptible and -resistant C. albicans after 24 h of treatment with different concentrations of fluconazole based on MIC (2 × MIC, 1 × MIC, ½ × MIC, and ¼ × MIC). Data are means with standard error from three independent experiments in triplicate assays. Statistically significant differences between the treatment isolates and controls are indicated with an asterisk ( P ≤ 0.05 ).

    Techniques Used:

    Relative quantitation of ERG11 gene expression (normalized to house-keeping gene, actin) in fluconazole-susceptible and -resistant C. albicans treated with different concentrations of fluconazole based on minimum inhibitory concentration (2 × MIC, 1 × MIC, ½ × MIC, and ¼ × MIC). Data are means of fold changes with standard deviation from three independent experiments amplified in triplicate.
    Figure Legend Snippet: Relative quantitation of ERG11 gene expression (normalized to house-keeping gene, actin) in fluconazole-susceptible and -resistant C. albicans treated with different concentrations of fluconazole based on minimum inhibitory concentration (2 × MIC, 1 × MIC, ½ × MIC, and ¼ × MIC). Data are means of fold changes with standard deviation from three independent experiments amplified in triplicate.

    Techniques Used: Quantitation Assay, Expressing, Concentration Assay, Standard Deviation, Amplification

    40) Product Images from "Biguanides enhance antifungal activity against Candida glabrata"

    Article Title: Biguanides enhance antifungal activity against Candida glabrata

    Journal: Virulence

    doi: 10.1080/21505594.2018.1475798

    Metformin enhances antifungal activity against drug resistant C. glabrata isolates. Fluconazole-resistant clinical C. glabrata isolates were incubated with 3.9 mg/mL of metformin and voriconazole (A) or fluconazole (B). The MIC 50 to each antifungal agent was determine with and without metformin. (C) Echinocandin and fluconazole double-resistant strain FR/MR was incubated with metformin and increasing amounts of micafungin. Micafungin MIC 50 was determined for strain FR/MR with and without metformin. * denotes p ≤ 0.05, ** p ≤ 0.01. Data represent a minimum of 3 independent experiments.
    Figure Legend Snippet: Metformin enhances antifungal activity against drug resistant C. glabrata isolates. Fluconazole-resistant clinical C. glabrata isolates were incubated with 3.9 mg/mL of metformin and voriconazole (A) or fluconazole (B). The MIC 50 to each antifungal agent was determine with and without metformin. (C) Echinocandin and fluconazole double-resistant strain FR/MR was incubated with metformin and increasing amounts of micafungin. Micafungin MIC 50 was determined for strain FR/MR with and without metformin. * denotes p ≤ 0.05, ** p ≤ 0.01. Data represent a minimum of 3 independent experiments.

    Techniques Used: Activity Assay, Incubation

    Combination therapy of metformin with common antifungal agents reduces C. glabrata activity. (A) Wild type C. glabrata treated with 6.5 mg/mL metformin in combination with voriconazole, fluconazole, amphotericin or micafungin in the presence of Prestoblue for 18hrs at 30°C. AU represent Prestoblue fluorescence unit. Data points were fitted to a four-parameter logistic curve. (B) MIC 50 were determined from four-parameter logistic curves comparing control antifungal drug with metformin combination. (C) C. glabrata was treated with voriconazole, metformin, or the combination for 18hrs, resuspended and plated onto YPD agar plates for 24-48hrs at 30°C. Data are plotted as fold population change compared to initial inoculum. * denotes p ≤ 0.05, ** p ≤ 0.01. Data represent 3 independent experiments.
    Figure Legend Snippet: Combination therapy of metformin with common antifungal agents reduces C. glabrata activity. (A) Wild type C. glabrata treated with 6.5 mg/mL metformin in combination with voriconazole, fluconazole, amphotericin or micafungin in the presence of Prestoblue for 18hrs at 30°C. AU represent Prestoblue fluorescence unit. Data points were fitted to a four-parameter logistic curve. (B) MIC 50 were determined from four-parameter logistic curves comparing control antifungal drug with metformin combination. (C) C. glabrata was treated with voriconazole, metformin, or the combination for 18hrs, resuspended and plated onto YPD agar plates for 24-48hrs at 30°C. Data are plotted as fold population change compared to initial inoculum. * denotes p ≤ 0.05, ** p ≤ 0.01. Data represent 3 independent experiments.

    Techniques Used: Activity Assay, Fluorescence

    Related Articles

    RNA Extraction:

    Article Title: Selection and evaluation of appropriate reference genes for RT-qPCR based expression analysis in Candida tropicalis following azole treatment
    Article Snippet: .. YPD broth in the presence (sub-inhibitory concentrations) and absence of fluconazole (Sigma-Aldrich, Germany) were supplemented with the freshly grown cells of C. tropicalis at a concentration of 1 × 106 cells/mL and incubated up to 7 hours for RNA extraction . .. Extraction of RNA and cDNA synthesis Total RNA from C. tropicalis isolates was extracted at the logarithmic phase using TRIzol reagent (Invitrogen, USA) as per the manufacturer’s protocol.

    Nuclear Magnetic Resonance:

    Article Title: ALARM NMR for HTS triage and chemical probe validation
    Article Snippet: .. ALARM NMR protein (from Support Protocol 1 ) SnakeSkin™ dialysis membrane, 10 kDa MWCO (ThermoFisher, Cat # 68305) Regeneration dialysis buffer (see recipe) Nitrogen (N2 ) gas Bradford reagent or microvolume spectrophotometer (e.g., ThermoFisher NanoDrop™) Reaction buffers (see recipes) 1.7 mm SampleJet NMR tubes (Bruker, Cat # Z106462) 384-well nonsterile, nonbinding surface, non-tissue culture treated microplate (e.g., Corning # 3640) Plate seals (e.g., Corning # 07-200-683) DMSO CPM (Sigma-Aldrich, Cat # C1484) positive reactive control compound 2-Chloro-1,4-naphthoquinone (Santa Cruz Biotechnology, Cat # sc-209130) positive reactive control compound Fluconazole (Sigma-Aldrich, Cat # PHR1160) negative reactive control compound Additional test compounds (as 10 mM DMSO stocks) D2 O (Cambridge Isotope Labs) Heating block Refrigerator with N2 line Stir plate with stir bar Tabletop centrifuge (e.g., Beckman-Coulter Allegra X-12) Oven incubator NMR spectrometer (recommended 400 MHz or higher fields; e.g., Bruker Avance 700 MHz spectrometer) Room temperature probe or CryoProbe™ (preferred) NMR autosampler (preferred) NMR analysis software (e.g., Bruker TopSpin®) Note: additional required reagents are listed under recipes. ..

    Spectrophotometry:

    Article Title: ALARM NMR for HTS triage and chemical probe validation
    Article Snippet: .. ALARM NMR protein (from Support Protocol 1 ) SnakeSkin™ dialysis membrane, 10 kDa MWCO (ThermoFisher, Cat # 68305) Regeneration dialysis buffer (see recipe) Nitrogen (N2 ) gas Bradford reagent or microvolume spectrophotometer (e.g., ThermoFisher NanoDrop™) Reaction buffers (see recipes) 1.7 mm SampleJet NMR tubes (Bruker, Cat # Z106462) 384-well nonsterile, nonbinding surface, non-tissue culture treated microplate (e.g., Corning # 3640) Plate seals (e.g., Corning # 07-200-683) DMSO CPM (Sigma-Aldrich, Cat # C1484) positive reactive control compound 2-Chloro-1,4-naphthoquinone (Santa Cruz Biotechnology, Cat # sc-209130) positive reactive control compound Fluconazole (Sigma-Aldrich, Cat # PHR1160) negative reactive control compound Additional test compounds (as 10 mM DMSO stocks) D2 O (Cambridge Isotope Labs) Heating block Refrigerator with N2 line Stir plate with stir bar Tabletop centrifuge (e.g., Beckman-Coulter Allegra X-12) Oven incubator NMR spectrometer (recommended 400 MHz or higher fields; e.g., Bruker Avance 700 MHz spectrometer) Room temperature probe or CryoProbe™ (preferred) NMR autosampler (preferred) NMR analysis software (e.g., Bruker TopSpin®) Note: additional required reagents are listed under recipes. ..

    Concentration Assay:

    Article Title: Selection and evaluation of appropriate reference genes for RT-qPCR based expression analysis in Candida tropicalis following azole treatment
    Article Snippet: .. YPD broth in the presence (sub-inhibitory concentrations) and absence of fluconazole (Sigma-Aldrich, Germany) were supplemented with the freshly grown cells of C. tropicalis at a concentration of 1 × 106 cells/mL and incubated up to 7 hours for RNA extraction . .. Extraction of RNA and cDNA synthesis Total RNA from C. tropicalis isolates was extracted at the logarithmic phase using TRIzol reagent (Invitrogen, USA) as per the manufacturer’s protocol.

    Article Title: Sir2 regulates stability of repetitive domains differentially in the human fungal pathogen Candida albicans
    Article Snippet: .. When indicated, media were supplemented with 5-fluorotic acid (5-FOA, Melford) at a concentration of 1 mg/ml, Nourseothricin (clonNAT, Melford) at a concentration of 100 μg/ml, Fluconazole (Sigma) at a concentration of 1 μg/ml or 0.5 mM H2 O2 . ..

    Article Title: Intestinal Candida parapsilosis isolates from Rett syndrome subjects bear potential virulent traits and capacity to persist within the host
    Article Snippet: .. Antifungal susceptibility testing All fungal isolates were tested for susceptibility to fluconazole, itraconazole and 5-flucytosine (Sigma-Aldrich) by Minimum Inibitory Concentration (MIC) assays according to the European Committee on Antimicrobial Susceptibility Testing (EUCAST) recommendations [ , ]. ..

    Incubation:

    Article Title: Selection and evaluation of appropriate reference genes for RT-qPCR based expression analysis in Candida tropicalis following azole treatment
    Article Snippet: .. YPD broth in the presence (sub-inhibitory concentrations) and absence of fluconazole (Sigma-Aldrich, Germany) were supplemented with the freshly grown cells of C. tropicalis at a concentration of 1 × 106 cells/mL and incubated up to 7 hours for RNA extraction . .. Extraction of RNA and cDNA synthesis Total RNA from C. tropicalis isolates was extracted at the logarithmic phase using TRIzol reagent (Invitrogen, USA) as per the manufacturer’s protocol.

    other:

    Article Title: Resorbable Beads Provide Extended Release of Antifungal Medication: In Vitro and In Vivo Analyses
    Article Snippet: Fabrication of Poly(d ,l -lactide-co-glycolide) PLGA/Amphotericin B and PLGA/Fluconazole Beads All materials utilized in this study, including PLGA (50:50), amphotericin B, and fluconazole, were acquired from Sigma-Aldrich (St. Louis, MO, USA).

    Article Title: Unexpected effects of azole transporter inhibitors on antifungal susceptibility in Candida glabrata and other pathogenic Candida species
    Article Snippet: Fluconazole (Sigma Aldrich) was dissolved in dimethyl sulfoxide (DMSO), and stored at 4°C.

    Article Title: Fluconazole Alters the Polysaccharide Capsule of Cryptococcus gattii and Leads to Distinct Behaviors in Murine Cryptococcosis
    Article Snippet: The MICs for fluconazole and amphotericin B (Sigma-Aldrich) were also determined by the microdilution method , – .

    Blocking Assay:

    Article Title: ALARM NMR for HTS triage and chemical probe validation
    Article Snippet: .. ALARM NMR protein (from Support Protocol 1 ) SnakeSkin™ dialysis membrane, 10 kDa MWCO (ThermoFisher, Cat # 68305) Regeneration dialysis buffer (see recipe) Nitrogen (N2 ) gas Bradford reagent or microvolume spectrophotometer (e.g., ThermoFisher NanoDrop™) Reaction buffers (see recipes) 1.7 mm SampleJet NMR tubes (Bruker, Cat # Z106462) 384-well nonsterile, nonbinding surface, non-tissue culture treated microplate (e.g., Corning # 3640) Plate seals (e.g., Corning # 07-200-683) DMSO CPM (Sigma-Aldrich, Cat # C1484) positive reactive control compound 2-Chloro-1,4-naphthoquinone (Santa Cruz Biotechnology, Cat # sc-209130) positive reactive control compound Fluconazole (Sigma-Aldrich, Cat # PHR1160) negative reactive control compound Additional test compounds (as 10 mM DMSO stocks) D2 O (Cambridge Isotope Labs) Heating block Refrigerator with N2 line Stir plate with stir bar Tabletop centrifuge (e.g., Beckman-Coulter Allegra X-12) Oven incubator NMR spectrometer (recommended 400 MHz or higher fields; e.g., Bruker Avance 700 MHz spectrometer) Room temperature probe or CryoProbe™ (preferred) NMR autosampler (preferred) NMR analysis software (e.g., Bruker TopSpin®) Note: additional required reagents are listed under recipes. ..

    Software:

    Article Title: ALARM NMR for HTS triage and chemical probe validation
    Article Snippet: .. ALARM NMR protein (from Support Protocol 1 ) SnakeSkin™ dialysis membrane, 10 kDa MWCO (ThermoFisher, Cat # 68305) Regeneration dialysis buffer (see recipe) Nitrogen (N2 ) gas Bradford reagent or microvolume spectrophotometer (e.g., ThermoFisher NanoDrop™) Reaction buffers (see recipes) 1.7 mm SampleJet NMR tubes (Bruker, Cat # Z106462) 384-well nonsterile, nonbinding surface, non-tissue culture treated microplate (e.g., Corning # 3640) Plate seals (e.g., Corning # 07-200-683) DMSO CPM (Sigma-Aldrich, Cat # C1484) positive reactive control compound 2-Chloro-1,4-naphthoquinone (Santa Cruz Biotechnology, Cat # sc-209130) positive reactive control compound Fluconazole (Sigma-Aldrich, Cat # PHR1160) negative reactive control compound Additional test compounds (as 10 mM DMSO stocks) D2 O (Cambridge Isotope Labs) Heating block Refrigerator with N2 line Stir plate with stir bar Tabletop centrifuge (e.g., Beckman-Coulter Allegra X-12) Oven incubator NMR spectrometer (recommended 400 MHz or higher fields; e.g., Bruker Avance 700 MHz spectrometer) Room temperature probe or CryoProbe™ (preferred) NMR autosampler (preferred) NMR analysis software (e.g., Bruker TopSpin®) Note: additional required reagents are listed under recipes. ..

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  • 99
    Millipore fluconazole
    Effects of E1210 and other antifungals on the adherence of C. albicans IFM49971 cells to polystyrene surfaces. C. albicans cells were incubated in the presence of each compound for 1 h in MOPS–RPMI 1640, and cells that adhered to polystyrene surfaces were stained using crystal violet. Each data point represents the mean ± SEM from three independent plates assayed in duplicate. Symbols: ●, E1210; ○, <t>fluconazole;</t> △, micafungin; □, amphotericin B.
    Fluconazole, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 36 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    93
    Millipore zona pellucida
    Oocyte yield, quality and developmental potential. (a) Oocyte yield was determined by mating age-matched Fyn-null or wild-type (WT) female mice with vasectomised WT males. Oocytes were collected the morning on which vaginal plugs were detected. A separate group of female mice was stimulated with pregnant mare’s serum gonadotropin and human chorionic gonadotrophin before mating. (b) Oocyte size and the thickness of the zona <t>pellucida</t> was measured from images made by Hoffman interference microscopy through the equator of oocytes. (c) The maturation status of oocytes recovered from WT and Fyn-null female mice was determined by confocal fluorescence microscopy following fixation and labelling with ethidium homodimer-2 (EthD-2) to reveal chromosome status. The developmental competence of WT and Fyn-null zygotes produced by mating with fertile WT males was determined by recovering oocytes and zygotes the morning on which vaginal plugs were detected and culturing them to allow development under identical conditions. Oocytes that failed to progress to the two-cell stage by 41 h after mating were fixed, stained with EthD-2 and examined by confocal fluorescence microscopy to determine meiotic and fertilisation status. Immature oocytes were excluded from calculation of the percentage rates of fertilisation and development. Significantly different values are indicated by *, **, ***, or **** (P
    Zona Pellucida, supplied by Millipore, used in various techniques. Bioz Stars score: 93/100, based on 46 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/zona pellucida/product/Millipore
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    Image Search Results


    Effects of E1210 and other antifungals on the adherence of C. albicans IFM49971 cells to polystyrene surfaces. C. albicans cells were incubated in the presence of each compound for 1 h in MOPS–RPMI 1640, and cells that adhered to polystyrene surfaces were stained using crystal violet. Each data point represents the mean ± SEM from three independent plates assayed in duplicate. Symbols: ●, E1210; ○, fluconazole; △, micafungin; □, amphotericin B.

    Journal: Antimicrobial Agents and Chemotherapy

    Article Title: E1210, a New Broad-Spectrum Antifungal, Suppresses Candida albicans Hyphal Growth through Inhibition of Glycosylphosphatidylinositol Biosynthesis

    doi: 10.1128/AAC.00731-11

    Figure Lengend Snippet: Effects of E1210 and other antifungals on the adherence of C. albicans IFM49971 cells to polystyrene surfaces. C. albicans cells were incubated in the presence of each compound for 1 h in MOPS–RPMI 1640, and cells that adhered to polystyrene surfaces were stained using crystal violet. Each data point represents the mean ± SEM from three independent plates assayed in duplicate. Symbols: ●, E1210; ○, fluconazole; △, micafungin; □, amphotericin B.

    Article Snippet: Since Fu et al. ( ) showed that the Als1p expression on the surface of C. albicans cells was induced after incubation for 1 h in MOPS–RPMI 1640, C. albicans cells were incubated for 1 h at 35°C in MOPS–RPMI 1640 that contained serial fourfold dilutions of E1210, micafungin, or fluconazole on a filter plate (MultiScreen HTS 96-well filtration plate; Millipore Corp., Billerica, MA).

    Techniques: Incubation, Staining

    Effects of E1210 and other antifungals on the density of C. albicans IFM49971 biofilms after incubation for 24 h. The biofilm density was quantified using safranin staining. Each data point represents the mean ± SEM from experiments on three independent plates assayed in duplicate. Symbols: ●, E1210; ○, fluconazole; △, micafungin; □, amphotericin B.

    Journal: Antimicrobial Agents and Chemotherapy

    Article Title: E1210, a New Broad-Spectrum Antifungal, Suppresses Candida albicans Hyphal Growth through Inhibition of Glycosylphosphatidylinositol Biosynthesis

    doi: 10.1128/AAC.00731-11

    Figure Lengend Snippet: Effects of E1210 and other antifungals on the density of C. albicans IFM49971 biofilms after incubation for 24 h. The biofilm density was quantified using safranin staining. Each data point represents the mean ± SEM from experiments on three independent plates assayed in duplicate. Symbols: ●, E1210; ○, fluconazole; △, micafungin; □, amphotericin B.

    Article Snippet: Since Fu et al. ( ) showed that the Als1p expression on the surface of C. albicans cells was induced after incubation for 1 h in MOPS–RPMI 1640, C. albicans cells were incubated for 1 h at 35°C in MOPS–RPMI 1640 that contained serial fourfold dilutions of E1210, micafungin, or fluconazole on a filter plate (MultiScreen HTS 96-well filtration plate; Millipore Corp., Billerica, MA).

    Techniques: Incubation, Staining

    Autoradiograph of TLC separation of GPI intermediates produced from UDP-[ 14 C]GlcNAc in the presence of E1210. The membranes of the S. cerevisiae GWT1 -disrupted cells overexpressing C. albicans GWT1 (A), A. fumigatus GWT1 (B), and human PIG-W (C) were assayed for the inositol acylation of GPI in the presence of E1210, fluconazole (FLCZ), amphotericin B (AMPH), and micafungin (MCFG). (D) PI-PLC treatment of radiolabeled lipids produced by membranes of S. cerevisiae overexpressing CaGWT1 and of S. cerevisiae overexpressing ScGWT1 . These images were trimmed from representative autoradiographs of the TLC plates derived from three independent experiments.

    Journal: Antimicrobial Agents and Chemotherapy

    Article Title: E1210, a New Broad-Spectrum Antifungal, Suppresses Candida albicans Hyphal Growth through Inhibition of Glycosylphosphatidylinositol Biosynthesis

    doi: 10.1128/AAC.00731-11

    Figure Lengend Snippet: Autoradiograph of TLC separation of GPI intermediates produced from UDP-[ 14 C]GlcNAc in the presence of E1210. The membranes of the S. cerevisiae GWT1 -disrupted cells overexpressing C. albicans GWT1 (A), A. fumigatus GWT1 (B), and human PIG-W (C) were assayed for the inositol acylation of GPI in the presence of E1210, fluconazole (FLCZ), amphotericin B (AMPH), and micafungin (MCFG). (D) PI-PLC treatment of radiolabeled lipids produced by membranes of S. cerevisiae overexpressing CaGWT1 and of S. cerevisiae overexpressing ScGWT1 . These images were trimmed from representative autoradiographs of the TLC plates derived from three independent experiments.

    Article Snippet: Since Fu et al. ( ) showed that the Als1p expression on the surface of C. albicans cells was induced after incubation for 1 h in MOPS–RPMI 1640, C. albicans cells were incubated for 1 h at 35°C in MOPS–RPMI 1640 that contained serial fourfold dilutions of E1210, micafungin, or fluconazole on a filter plate (MultiScreen HTS 96-well filtration plate; Millipore Corp., Billerica, MA).

    Techniques: Autoradiography, Thin Layer Chromatography, Produced, Planar Chromatography, Derivative Assay

    Microscopic images of C. albicans cells after treatment with E1210 and other antifungals for 4 h. Cells were left untreated or were treated with 0.5×MIC, 1×MIC, and the highest concentration tested of each compound for 4 h. The magnification of all images is ×200. Abbreviations: FLCZ, fluconazole; MCFG, micafungin; AMPH, amphotericin B.

    Journal: Antimicrobial Agents and Chemotherapy

    Article Title: E1210, a New Broad-Spectrum Antifungal, Suppresses Candida albicans Hyphal Growth through Inhibition of Glycosylphosphatidylinositol Biosynthesis

    doi: 10.1128/AAC.00731-11

    Figure Lengend Snippet: Microscopic images of C. albicans cells after treatment with E1210 and other antifungals for 4 h. Cells were left untreated or were treated with 0.5×MIC, 1×MIC, and the highest concentration tested of each compound for 4 h. The magnification of all images is ×200. Abbreviations: FLCZ, fluconazole; MCFG, micafungin; AMPH, amphotericin B.

    Article Snippet: Since Fu et al. ( ) showed that the Als1p expression on the surface of C. albicans cells was induced after incubation for 1 h in MOPS–RPMI 1640, C. albicans cells were incubated for 1 h at 35°C in MOPS–RPMI 1640 that contained serial fourfold dilutions of E1210, micafungin, or fluconazole on a filter plate (MultiScreen HTS 96-well filtration plate; Millipore Corp., Billerica, MA).

    Techniques: Concentration Assay

    Effects of E1210 and other antifungals on germ tube formation in C. albicans IFM49971 cells. C. albicans cells were incubated in the presence of each compound for 4 h in MOPS–RPMI 1640, and cells that converted from yeast form to the hyphal form and adhered to polystyrene surfaces were stained with crystal violet. Each data point represents the mean ± SEM from three independent plates assayed in duplicate. Symbols: ●, E1210; ○, fluconazole; △, micafungin; □, amphotericin B.

    Journal: Antimicrobial Agents and Chemotherapy

    Article Title: E1210, a New Broad-Spectrum Antifungal, Suppresses Candida albicans Hyphal Growth through Inhibition of Glycosylphosphatidylinositol Biosynthesis

    doi: 10.1128/AAC.00731-11

    Figure Lengend Snippet: Effects of E1210 and other antifungals on germ tube formation in C. albicans IFM49971 cells. C. albicans cells were incubated in the presence of each compound for 4 h in MOPS–RPMI 1640, and cells that converted from yeast form to the hyphal form and adhered to polystyrene surfaces were stained with crystal violet. Each data point represents the mean ± SEM from three independent plates assayed in duplicate. Symbols: ●, E1210; ○, fluconazole; △, micafungin; □, amphotericin B.

    Article Snippet: Since Fu et al. ( ) showed that the Als1p expression on the surface of C. albicans cells was induced after incubation for 1 h in MOPS–RPMI 1640, C. albicans cells were incubated for 1 h at 35°C in MOPS–RPMI 1640 that contained serial fourfold dilutions of E1210, micafungin, or fluconazole on a filter plate (MultiScreen HTS 96-well filtration plate; Millipore Corp., Billerica, MA).

    Techniques: Incubation, Staining

    Oxidative activity of biofilm treatment with Fluconazole (64 µg/mL) and Ethanolic Extract of Licania rigid Leaves (EEFLr) (2048 µg/mL) formed by clinical Candida yeasts. Data represent the mean and standard deviation of absorbance during biofilm production and treatment, compared with the control (biofilm formation without treatment). For the analysis, Tukey’s multiple comparisons test was performed for all means obtained at 5% significance level. The symbols “*” and “****” indicate significant differences between treatments and the control with ( p ≤ 0.05) and ( p ≤ 0.0001), respectively.

    Journal: Antibiotics

    Article Title: Evaluation of the Antifungal Activity of the Licania Rigida Leaf Ethanolic Extract against Biofilms Formed by Candida Sp. Isolates in Acrylic Resin Discs

    doi: 10.3390/antibiotics8040250

    Figure Lengend Snippet: Oxidative activity of biofilm treatment with Fluconazole (64 µg/mL) and Ethanolic Extract of Licania rigid Leaves (EEFLr) (2048 µg/mL) formed by clinical Candida yeasts. Data represent the mean and standard deviation of absorbance during biofilm production and treatment, compared with the control (biofilm formation without treatment). For the analysis, Tukey’s multiple comparisons test was performed for all means obtained at 5% significance level. The symbols “*” and “****” indicate significant differences between treatments and the control with ( p ≤ 0.05) and ( p ≤ 0.0001), respectively.

    Article Snippet: Six hours after Fluconazole and extract addition, the biofilms were quantified using the tetrazolium salt reduction assay, whereby 20 μL, in the 5 μL to 1mL of PBS buffer ratio, sterilized by membrane filtration (0.22 μL pores, Millipore, St. Louis, MO 63103, USA) was added to each well of the microtiter plate, including control wells.

    Techniques: Activity Assay, Standard Deviation

    Oocyte yield, quality and developmental potential. (a) Oocyte yield was determined by mating age-matched Fyn-null or wild-type (WT) female mice with vasectomised WT males. Oocytes were collected the morning on which vaginal plugs were detected. A separate group of female mice was stimulated with pregnant mare’s serum gonadotropin and human chorionic gonadotrophin before mating. (b) Oocyte size and the thickness of the zona pellucida was measured from images made by Hoffman interference microscopy through the equator of oocytes. (c) The maturation status of oocytes recovered from WT and Fyn-null female mice was determined by confocal fluorescence microscopy following fixation and labelling with ethidium homodimer-2 (EthD-2) to reveal chromosome status. The developmental competence of WT and Fyn-null zygotes produced by mating with fertile WT males was determined by recovering oocytes and zygotes the morning on which vaginal plugs were detected and culturing them to allow development under identical conditions. Oocytes that failed to progress to the two-cell stage by 41 h after mating were fixed, stained with EthD-2 and examined by confocal fluorescence microscopy to determine meiotic and fertilisation status. Immature oocytes were excluded from calculation of the percentage rates of fertilisation and development. Significantly different values are indicated by *, **, ***, or **** (P

    Journal: Reproduction, fertility, and development

    Article Title: Role of Fyn kinase in oocyte developmental potential

    doi: 10.1071/RD09311

    Figure Lengend Snippet: Oocyte yield, quality and developmental potential. (a) Oocyte yield was determined by mating age-matched Fyn-null or wild-type (WT) female mice with vasectomised WT males. Oocytes were collected the morning on which vaginal plugs were detected. A separate group of female mice was stimulated with pregnant mare’s serum gonadotropin and human chorionic gonadotrophin before mating. (b) Oocyte size and the thickness of the zona pellucida was measured from images made by Hoffman interference microscopy through the equator of oocytes. (c) The maturation status of oocytes recovered from WT and Fyn-null female mice was determined by confocal fluorescence microscopy following fixation and labelling with ethidium homodimer-2 (EthD-2) to reveal chromosome status. The developmental competence of WT and Fyn-null zygotes produced by mating with fertile WT males was determined by recovering oocytes and zygotes the morning on which vaginal plugs were detected and culturing them to allow development under identical conditions. Oocytes that failed to progress to the two-cell stage by 41 h after mating were fixed, stained with EthD-2 and examined by confocal fluorescence microscopy to determine meiotic and fertilisation status. Immature oocytes were excluded from calculation of the percentage rates of fertilisation and development. Significantly different values are indicated by *, **, ***, or **** (P

    Article Snippet: Cumulus-free MII oocytes were treated in acid Tyrode’s solution (pH 2.5) to remove the zona pellucida, followed by incubation in potassium simplex optimized medium with amino acids (KSOMAA ) (Millipore Corporation, Phillipsburg, NJ, USA) containing 1 mg mL−1 BSA (embryo tested; Sigma-Aldrich) and 1 mM fura-2 AM (Invitrogen) premixed with 0.02% pluronic F-127 (Invitrogen) for 30 min at 378C.

    Techniques: Mouse Assay, Microscopy, Fluorescence, Ethidium Homodimer Assay, Produced, Staining

    Morphology of Fyn-null oocytes. Oocytes recovered from wild-type (WT) B6;129SF2/J and Fyn-null B6;129S7-Fyn tm1Sor/J (Fyn −/− ) mice were examined by Hoffman modulation contrast. The Fyn-null oocytes contained dense, irregular inclusions. The diameter of the two groups of oocytes did not differ significantly (P = 0.563); however, the thickness of the zona pellucida of Fyn-null oocytes was significantly less than that of WT oocytes (P

    Journal: Reproduction, fertility, and development

    Article Title: Role of Fyn kinase in oocyte developmental potential

    doi: 10.1071/RD09311

    Figure Lengend Snippet: Morphology of Fyn-null oocytes. Oocytes recovered from wild-type (WT) B6;129SF2/J and Fyn-null B6;129S7-Fyn tm1Sor/J (Fyn −/− ) mice were examined by Hoffman modulation contrast. The Fyn-null oocytes contained dense, irregular inclusions. The diameter of the two groups of oocytes did not differ significantly (P = 0.563); however, the thickness of the zona pellucida of Fyn-null oocytes was significantly less than that of WT oocytes (P

    Article Snippet: Cumulus-free MII oocytes were treated in acid Tyrode’s solution (pH 2.5) to remove the zona pellucida, followed by incubation in potassium simplex optimized medium with amino acids (KSOMAA ) (Millipore Corporation, Phillipsburg, NJ, USA) containing 1 mg mL−1 BSA (embryo tested; Sigma-Aldrich) and 1 mM fura-2 AM (Invitrogen) premixed with 0.02% pluronic F-127 (Invitrogen) for 30 min at 378C.

    Techniques: Mouse Assay

    Calcium oscillatory patterns typical of fertilised Fyn-null oocytes. Cumulus- and zona pellucida-free oocytes from nine Fyn-null and five wild-type (WT) female mice were loaded with fura-2, bound to polylysine-coated coverslips and fertilised with WT spermatozoa in vitro so that fertilisation-induced calcium oscillations could be recorded. (a) The typical pattern of calcium oscillations following fertilisation of WT (dashed line) and Fyn-null (solid line) oocytes. (b) The average duration and amplitude of the first calcium oscillation, as well as the total peak number during the first hour after insemination. The total integrated calcium input signal during the first hour after insemination is presented on the far right. The y-axis values show the mean ± s.d.; n values represent the number of recordings analysed from each group. Groups that differ significantly (P

    Journal: Reproduction, fertility, and development

    Article Title: Role of Fyn kinase in oocyte developmental potential

    doi: 10.1071/RD09311

    Figure Lengend Snippet: Calcium oscillatory patterns typical of fertilised Fyn-null oocytes. Cumulus- and zona pellucida-free oocytes from nine Fyn-null and five wild-type (WT) female mice were loaded with fura-2, bound to polylysine-coated coverslips and fertilised with WT spermatozoa in vitro so that fertilisation-induced calcium oscillations could be recorded. (a) The typical pattern of calcium oscillations following fertilisation of WT (dashed line) and Fyn-null (solid line) oocytes. (b) The average duration and amplitude of the first calcium oscillation, as well as the total peak number during the first hour after insemination. The total integrated calcium input signal during the first hour after insemination is presented on the far right. The y-axis values show the mean ± s.d.; n values represent the number of recordings analysed from each group. Groups that differ significantly (P

    Article Snippet: Cumulus-free MII oocytes were treated in acid Tyrode’s solution (pH 2.5) to remove the zona pellucida, followed by incubation in potassium simplex optimized medium with amino acids (KSOMAA ) (Millipore Corporation, Phillipsburg, NJ, USA) containing 1 mg mL−1 BSA (embryo tested; Sigma-Aldrich) and 1 mM fura-2 AM (Invitrogen) premixed with 0.02% pluronic F-127 (Invitrogen) for 30 min at 378C.

    Techniques: Mouse Assay, In Vitro

    Morula-stage embryos re-engineered through diploid aggregation. Flowchart of chimeric offspring derivation from random incorporation of embryonic stem cells into early stage embryos achieved through imposed diploid aggregation. Yellow box : Initial embryos are generated from timed pregnant KCNJ11 gene-deficient donors previously superovulated through hormonal activation using sequential i.p. injections of PMSG and HCG. Knockout embryos are harvested at 2.5 d.p.c. to collect at the morula stage. Zona pellucida is removed using acid Tyrode's solution to prepare aggregation competent early embryos. Blue box : Simultaneously, embryonic stem cells are grown for two passages to produce low-density cultures that when digested are able to generate clumps of 8–15 pluripotent stem cells. Green box : Synchronized pseudopregnant surrogates are produced by appropriate selection of females in oestrus, and mated with vasectomized studs. Bottom : Mosaic morula generated after aggregation of KNCJ11 gene-deficient embryos with wild-type embryonic stem cells. Following intrauterine surgical transfer, surrogate females support normal embryonic development and give birth to chimeric offspring.

    Journal: Philosophical Transactions of the Royal Society B: Biological Sciences

    Article Title: KCNJ11 knockout morula re-engineered by stem cell diploid aggregation

    doi: 10.1098/rstb.2008.0179

    Figure Lengend Snippet: Morula-stage embryos re-engineered through diploid aggregation. Flowchart of chimeric offspring derivation from random incorporation of embryonic stem cells into early stage embryos achieved through imposed diploid aggregation. Yellow box : Initial embryos are generated from timed pregnant KCNJ11 gene-deficient donors previously superovulated through hormonal activation using sequential i.p. injections of PMSG and HCG. Knockout embryos are harvested at 2.5 d.p.c. to collect at the morula stage. Zona pellucida is removed using acid Tyrode's solution to prepare aggregation competent early embryos. Blue box : Simultaneously, embryonic stem cells are grown for two passages to produce low-density cultures that when digested are able to generate clumps of 8–15 pluripotent stem cells. Green box : Synchronized pseudopregnant surrogates are produced by appropriate selection of females in oestrus, and mated with vasectomized studs. Bottom : Mosaic morula generated after aggregation of KNCJ11 gene-deficient embryos with wild-type embryonic stem cells. Following intrauterine surgical transfer, surrogate females support normal embryonic development and give birth to chimeric offspring.

    Article Snippet: Embryos, in groups of 20–30, were transferred with as little M2 medium as possible into the acid Tyrode solution, and continuously irrigated to keep neighbouring embryos separated until zona pelucida dissolved within 30–40 s. Once stripped of their zona pellucida, denuded morulae were washed in five drops of M2 followed by five drops of EmbryoMax KSOM (Millipore, Billerica, MA), preparing them for subsequent in vitro manipulation.

    Techniques: Generated, Activation Assay, Knock-Out, Produced, Selection

    KCNJ11 gene-deficient embryos aggregated with embryonic stem cells. ( a ) Stage-specific embryos were harvested from KCNJ11 gene deficient donors. ( b ) Zona pellucida was removed with acid Tyrode washes to produce aggregation competent embryos. ( c ) Embryonic stem cells are prepared in 8–15 cell clumps. ( d ) Diploid aggregation was coerced between two morula-stage embryos and embryonic stem cell clumps.

    Journal: Philosophical Transactions of the Royal Society B: Biological Sciences

    Article Title: KCNJ11 knockout morula re-engineered by stem cell diploid aggregation

    doi: 10.1098/rstb.2008.0179

    Figure Lengend Snippet: KCNJ11 gene-deficient embryos aggregated with embryonic stem cells. ( a ) Stage-specific embryos were harvested from KCNJ11 gene deficient donors. ( b ) Zona pellucida was removed with acid Tyrode washes to produce aggregation competent embryos. ( c ) Embryonic stem cells are prepared in 8–15 cell clumps. ( d ) Diploid aggregation was coerced between two morula-stage embryos and embryonic stem cell clumps.

    Article Snippet: Embryos, in groups of 20–30, were transferred with as little M2 medium as possible into the acid Tyrode solution, and continuously irrigated to keep neighbouring embryos separated until zona pelucida dissolved within 30–40 s. Once stripped of their zona pellucida, denuded morulae were washed in five drops of M2 followed by five drops of EmbryoMax KSOM (Millipore, Billerica, MA), preparing them for subsequent in vitro manipulation.

    Techniques: