Layer plate CAS assay for the quantitation of siderophore production and determination of exudation patterns for fungi

Andrews, Megan Y.; Santelli, Cara M.; Duckworth, Owen W.

doi:10.1016/j.mimet.2015.12.012

Cited by 21 publications

(24 citation statements)

References 11 publications

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“…Liquid CAS assay. We prepared 10ϫ CAS assay reagent as described previously (85). One part 10ϫ CAS reagent was combined with 9 parts siderophore-containing liquid, and the mixture was incubated at 37°C for 6 h. Mixtures were measured with a plate reader (Opsys MR; DYNEX Technologies) and compared to RPMI medium -1ϫ CAS reagent as reference values.…”

Section: Methodsmentioning

confidence: 99%

Studies of Pseudomonas aeruginosa Mutants Indicate Pyoverdine as the Central Factor in Inhibition of Aspergillus fumigatus Biofilm

et al. 2018

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and are common opportunistic bacterial and fungal pathogens, respectively. They often coexist in airways of immunocompromised patients and individuals with cystic fibrosis, where they form biofilms and cause acute and chronic illnesses. Hence, the interactions between them have long been of interest and it is known that can inhibit We have approached the definition of the inhibitory molecules by studying 24 mutants with various virulence genes deleted for the ability to inhibit biofilms. The ability of cells or their extracellular products produced during planktonic or biofilm growth to affect biofilm metabolism or planktonic growth was studied in agar and liquid assays using conidia or hyphae. Four mutants, the, ,, and mutants, were shown to be defective in various assays. This suggested the siderophore pyoverdine as the key inhibitory molecule, although additional quorum sensing-regulated factors likely contribute to the deficiency of the latter two mutants. Studies of pure pyoverdine substantiated these conclusions and included the restoration of inhibition by the pyoverdine deletion mutants. A correlation between the concentration of pyoverdine produced and antifungal activity was also observed in clinical isolates derived from lungs of cystic fibrosis patients. The key inhibitory mechanism of pyoverdine was chelation of iron and denial of iron to Interactions between human pathogens found in the same body locale are of vast interest. These interactions could result in exacerbation or amelioration of diseases. The bacterium affects the growth of the fungus Both pathogens form biofilms that are resistant to therapeutic drugs and host immunity. and biofilms are found , e.g., in the lungs of cystic fibrosis patients. Studying 24 mutants, we identified pyoverdine as the major anti- compound produced by Pyoverdine captures iron from the environment, thus depriving of a nutrient essential for its growth and metabolism. We show how microbes of different kingdoms compete for essential resources. Iron deprivation could be a therapeutic approach to the control of pathogen growth.

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Section: Methodsmentioning

confidence: 99%

Studies of Pseudomonas aeruginosa Mutants Indicate Pyoverdine as the Central Factor in Inhibition of Aspergillus fumigatus Biofilm

et al. 2018

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“…In addition, we provide detailed protocols for making CAS assay layer plates, the digital analysis protocol for determining area of color change, and discuss growth media that do and do not work with the layer plate method. The results of these data, their interpretation, and further discussion can be found in Andrews et al, 2016 [1] .…”

mentioning

confidence: 76%

“… The data presented in this note details the method used to quantify siderophore equivalents exuded by fungi on agar-solidified growth plates [1] . The visual data included illustrate the use of the CAS layer plate method to compare siderophore exudation patterns between different fungi, or between the same fungi on different media.…”

Section: Value Of the Datamentioning

confidence: 99%

“…The data presented in this note details the method used to quantify siderophore equivalents exuded by fungi on agar-solidified growth plates [1] .…”

Section: Value Of the Datamentioning

confidence: 99%

“…This data in brief article provides the image data files supporting the method developed in [1] , and provides clear examples and greater detail to aid in using the method. The image data included in Fig.…”

Section: Datamentioning

confidence: 99%

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Digital image quantification of siderophores on agar plates

2016

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This article presents visual image data and detailed methodology for the use of a new method for quantifying the exudation of siderophores during fungal growth. The data include images showing time series for calibration, fungal exudation, and negative controls, as well as replication accuracy information. In addition, we provide detailed protocols for making CAS assay layer plates, the digital analysis protocol for determining area of color change, and discuss growth media that do and do not work with the layer plate method. The results of these data, their interpretation, and further discussion can be found in Andrews et al., 2016 [1].

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