I'm isolating bacterial from soil on Nutrient Agar media but after 4-5 days I see fungal growth start in the plates. I read about use of Nystatin to prevent fungal growth. How much quantity or concentration of Nystatin should I use for 250 ml of culture media.
Nystatin is an antifungal agent commonly used in microbiology to inhibit the growth of fungi in culture media. The appropriate concentration of nystatin to use in a Nutrient Agar medium depends on the specific application and the susceptibility of the fungi you intend to inhabit.
Generally, nystatin is added to agar media at concentrations ranging from 50 to 100 units per milliliter (U/mL) or 50 to 100 micrograms per milliliter (μg/mL). However, the optimal concentration may vary depending on factors such as the type of fungi present in the sample and the desired level of inhibition. For Nutrient Agar medium, a typical concentration of nystatin is around 50 U/mL or 50 μg/mL. This concentration is often effective at inhibiting the growth of most common fungi without significantly affecting the growth of bacteria or other microorganisms that may be present in the sample. It's important to note that nystatin is sensitive to heat and light, so it should be added to the medium after autoclaving and cooling to avoid degradation. Ultimately, it may be necessary to perform a series of experiments using different concentrations of nystatin to determine the optimal concentration for your specific application and experimental conditions.
Kais Khudhair al Hadrawi Thank you for the answer.
On adding Nystatin powder to the autoclaved and cooled media, it did not dissolve well but instead formed clumps. Is it added directly or as a solution to the media.
I add 50mg/L of Nystatin as a methanol solution (or rather suspension). The 50mg does not dissolve completely in 1mL of methanol but the suspension is homogenic so if you add this to warm agar (with stirring) then you get a somewhat even dispersal. Hope that helps...
Jakub Grzesiak Thank you for the answer. will definitely try this method.
To prevent fungal contamination in bacterial cultures on Nutrient Agar (NA) medium, adding an antifungal agent like Nystatin can be very effective. Here's a detailed guide on how to use Nystatin for this purpose:
Nystatin Concentration for Nutrient Agar
Typical Concentration:
The usual concentration of Nystatin to inhibit fungal growth without affecting bacterial growth is 50-100 units per milliliter (U/mL) of medium.
Steps to Prepare Nystatin-Infused Nutrient Agar
Important Considerations
Summary
To prevent fungal growth in Nutrient Agar medium for isolating bacteria from soil, use Nystatin at a concentration of 50-100 U/mL. For 250 mL of Nutrient Agar, add 25 mL of a 1,000 U/mL Nystatin stock solution to achieve a final concentration of 100 U/mL. Ensure the medium is cooled to 45-50°C before adding Nystatin and mix thoroughly before pouring plates. This approach will help you maintain a fungal-free environment for your bacterial cultures.
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