Potato Dextrose Agar (PDA) – Principle, Composition, Preparation, Uses

What is Potato Dextrose Agar (PDA)?

• Potato Dextrose Agar (PDA) is a commonly used microbiological growth medium that serves as a foundation for cultivating and identifying yeasts and molds. It is prepared by infusing potatoes and incorporating dextrose, which provides a source of energy for fungal growth. PDA is widely endorsed by authoritative bodies such as the American Public Health Association (APHA) and the Food and Drug Administration (FDA) for conducting plate counts of yeasts and molds in the analysis of various food and dairy products.
• PDA is considered a versatile medium suitable for a wide range of applications. It can be supplemented with different additives to enhance its selectivity or inhibit the growth of unwanted bacteria. For instance, selective agents like Chloramphenicol, Tartaric Acid, and Chlortetracycline can be added to prevent bacterial growth while allowing the growth of desired yeasts and molds. This modification enables PDA to be utilized as a selective medium for specific purposes, such as isolating and cultivating clinically significant yeast and molds.
• In addition to its role in food and dairy microbiology, PDA finds utility in maintaining stock cultures of certain dermatophytes. Dermatophytes are fungi that cause infections of the skin, hair, and nails, and PDA provides a suitable environment for their preservation.
• One of the advantageous properties of PDA is its ability to stimulate sporulation and pigmentation of fungi. This feature aids in the cultivation and differentiation of pathogenic and non-pathogenic fungi, contributing to their identification and characterization. By observing the colony morphology and pigmentation patterns on PDA, microbiologists can gather valuable information about the fungal species under investigation.
• Specific variations of PDA with additional components are recommended for different microbial examinations. PDA with tartaric acid (TA) is suggested for the microbial analysis of food and dairy products. The inclusion of chlortetracycline is recommended for enumerating yeast and mold in cosmetic samples. Additionally, PDA supplemented with chloramphenicol is suitable for the selective cultivation of fungi in mixed samples.
• Overall, Potato Dextrose Agar (PDA) serves as a valuable tool in the field of microbiology for the cultivation, identification, and enumeration of yeasts, molds, and dermatophytes. Its versatility, ease of preparation, and ability to support fungal growth make it a widely used medium in various laboratory settings.

Principle of Potato Dextrose Agar (PDA)

The principle of Potato Dextrose Agar (PDA) lies in its composition and the interactions between its components. PDA is designed to provide a favorable environment for the growth and proliferation of fungi, particularly yeasts and molds.

The carbohydrate source in PDA is dextrose, which serves as a growth stimulant for fungal organisms. It provides a readily available energy source that supports their metabolic activities and promotes their multiplication. The potato infusion in PDA acts as a nutrient base, supplying additional essential nutrients necessary for the luxuriant growth of most fungi.

To solidify the agar medium, agar, a polysaccharide derived from seaweed, is added. Agar allows the PDA to take on a gel-like consistency, providing a solid surface for the fungi to grow and form colonies.

In some formulations of PDA, a specified amount of sterile tartaric acid (10%) may be incorporated to lower the pH of the medium to approximately 3.5. This acidic pH is intentionally adjusted to inhibit the growth of bacteria, creating a selective environment that favors the growth of fungi over bacterial contaminants. By inhibiting bacterial growth, PDA helps to ensure that the resulting colonies on the medium primarily consist of fungi.

It is important to note that once the tartaric acid has been added to the PDA, the medium should not be reheated. Heating the acidified medium can lead to the hydrolysis of agar, rendering it unable to solidify properly.

Overall, the principle of Potato Dextrose Agar (PDA) is to provide a nutrient-rich medium with dextrose as a growth stimulant, potato infusion as a nutrient base, and agar as a solidifying agent. The addition of tartaric acid helps to create a selective environment by inhibiting bacterial growth, allowing for the cultivation and enumeration of fungi in various microbiological applications.

Potato Dextrose Agar Composition 

The composition of Potato Dextrose Agar (PDA) can vary depending on specific formulations and desired applications. Here are the compositions of different variations of PDA mentioned in the provided information:

1. Standard Potato Dextrose Agar:
   • Dextrose: 20 g/L
   • Potato extract: 4 g/L
   • Agar: 15 g/L
2. Potato Dextrose Agar with Chloramphenicol:
   • Dextrose: 20 g/L
   • Potato extract: 4 g/L
   • Agar: 15 g/L
   • Chloramphenicol: 25 mg/L (pH-5.6 +/- 0.2 at 25°C)
3. Potato Dextrose Agar with Chlortetracycline:
   • Dextrose: 20 g/L
   • Potato extract: 4 g/L
   • Agar: 15 g/L
   • Chlortetracycline: 40 mg/L
4. Potato Dextrose Agar with tartaric acid:
   • Dextrose: 20 g/L
   • Potato extract: 4 g/L
   • Agar: 15 g/L
   • Tartaric acid: 1.4 g/L (pH-3.5 +/- 0.3 at 25°C)
5. Potato Dextrose Agar Salt Agar:
   • Dextrose: 20 g/L
   • Potato extract: 4 g/L
   • Agar: 15 g/L
   • Sodium Chloride: 75.0 g/L

Ingredients	Standard PDA	PDA with Chloramphenicol	PDA with Chlortetracycline	PDA with Tartaric Acid	PDA Salt Agar
Dextrose	20 g/L	20 g/L	20 g/L	20 g/L	20 g/L
Potato extract	4 g/L	4 g/L	4 g/L	4 g/L	4 g/L
Agar	15 g/L	15 g/L	15 g/L	15 g/L	15 g/L
Chloramphenicol	–	25 mg/L	–	–	–
Chlortetracycline	–	–	40 mg/L	–	–
Tartaric acid	–	–	–	1.4 g/L	–
Sodium Chloride	–	–	–	–	75.0 g/L
pH (at 25°C)	–	5.6 +/- 0.2	–	3.5 +/- 0.3	–

Please note that “-” indicates the absence of a particular ingredient in that variation of PDA.

Preparation Procedure of Potato Dextrose Agar

The preparation of Potato Dextrose Agar (PDA) involves a straightforward process. Here are the steps based on the provided information:

1. Suspend 39 grams of dehydrated PDA media obtained from a commercial supplier in 1000 ml of distilled water.
2. Heat the mixture to boiling while stirring to ensure complete dissolution of the medium.
3. Autoclave the PDA at 15 lbs pressure (121°C) for 15 minutes to sterilize the medium and eliminate any potential contaminants.
4. After autoclaving, mix the PDA well to ensure homogeneity before dispensing it into sterile Petri dishes or tubes.
5. If a specific pH of 3.5 is required for certain applications, the medium should be acidified using sterile 10% tartaric acid. Add approximately 1 ml of tartaric acid to every 100 ml of sterile and cooled PDA medium. It’s essential not to heat the medium after adding the acid.
6. To process a specimen, streak it onto the PDA medium using a sterile inoculating loop. This step helps to obtain isolated colonies for further analysis and identification.
7. Invert the Petri dishes (agar side up) to prevent condensation from dripping onto the colonies. Incubate the plates at a temperature range of 25-30°C, providing increased humidity to promote fungal growth.
8. Check the plates weekly for the presence of fungal growth. Cultures should be incubated for 4-6 weeks before being reported as negative, ensuring sufficient time for fungal organisms to develop and become detectable.

By following these steps, one can properly prepare Potato Dextrose Agar for various microbiological applications, facilitating the growth and observation of fungi in a laboratory setting.

Preparation of Potato Dextrose Agar from commercial Powder

1. Add 39 grams of commercially powered water to 1 liter of pure water.
2. Boil and mix until it dissolves completely.
3. Sterilize media using autoclaving at 121oC for 15 mins.
4. Dispense aseptically into sterilized Petri dishes.

Result of Potato Dextrose Agar

The interpretation of results on Potato Dextrose Agar (PDA) involves examining the characteristics of the colonies that develop on the medium. Here’s how the results can be interpreted based on the provided information:

1. Yeasts: Yeasts typically appear as creamy to white colonies on PDA. These colonies are usually smooth, moist, and have a slightly raised or domed shape. Yeasts are single-celled fungi, and their growth on PDA often displays a smooth and uniform texture.
2. Molds: Molds, on the other hand, form filamentous colonies with a more diverse range of colors compared to yeasts. The colonies of molds can exhibit various hues, including green, blue, gray, black, or even other vibrant colors. The texture of mold colonies is often fuzzy or velvety due to the presence of hyphae, which are long branching filaments characteristic of molds.

It’s important to note that the specific characteristics of colonies on PDA can vary depending on the fungal species being examined. Therefore, additional tests or microscopic examination may be required to identify and differentiate specific fungal isolates accurately.

By observing the colony morphology, color, and texture on Potato Dextrose Agar, microbiologists can make preliminary assessments about the types of fungi present in the sample. This information aids in the identification and classification of yeast and mold isolates in various research, clinical, or quality control settings.

Colony morphology of some fungi on Potato Dextrose Agar

Fungi	Colony Characteristics	Surface color	Reverse color	Zonation	Sporulation
A.candidus	Velvety thick	Creamish white	Slightly creamish	Radially furrowed on the reverse	Moderate
A.niger	Velvety	White with typical black spores	Yellow	Heavily furrowed on the reverse	Heavy
A.sulphureus	Velvety	Dirty white with yellow spores at the center	Orange to chocolate color	Slightly radially furrowed	Moderate
A. versicolor	Floccose	White to orange-cream with green spores at the center	Bright orange	Heavily wrinkled on reverse	Moderate
Penicillium corylophilum	Velvety	Dark green	Colorless to Creamish	With shallow centre and radially furrowed raised margin	Moderate
P. expansum	Velvety	Dark green with clear exudates and distinct sterile white margin	Yellow	Radially furrowed	Heavy
Penicillium spp	Powdery	Olivaceous green with sterile white margin	Orange to red, wrinkled	Radially furrowed	Heavy
Fusarium oxysporum	Floccose	Magenta pink	Magenta-red turning violet	With concentric zones of dark and light reddish coloration	Poor

Quality Control

Quality control of Potato Dextrose Agar (PDA) is crucial to ensure its effectiveness and reliability for the cultivation and identification of yeasts and molds. The provided information highlights the quality control measures performed for PDA, including control strains, incubation conditions, expected results, and reference to the batch-related Certificate of Analysis. Here’s an overview:

1. Control Strains and Incubation:
   • Geotrichum candidum 1240 (DSM #): Incubated for 5 days at 28 °C with the expected result of good to very good growth.
   • Penicillium commune 10428: Incubated for 5 days at 28 °C with the expected result of medium to good growth.
   • Trichophyton ajelloi 28454: Incubated for 5 days at 28 °C with the expected result of medium to good growth.
   • Candida albicans 10231, Saccharomyces cerevisiae 9763, Rhodotorula mucilaginosa 70403 (DSM #): Incubated for ≤ 5 days at 20-25 °C with the expected result of recovery ≥ 70%.
   • Aspergillus brasiliensis (formerly A. niger) 16404: Incubated for ≤ 5 days at 20-25 °C with the expected result of recovery ≥ 50%.
2. Quality Control (Spiral Plating Method):
   • The control strains mentioned above are inoculated onto PDA using the spiral plating method.
   • The inoculum size ranges from 10-100 colony-forming units (CFU).
   • Incubation is carried out for ≤ 5 days at 20-25 °C.
   • The expected result for Candida albicans, Saccharomyces cerevisiae, and Rhodotorula mucilaginosa is a recovery of ≥ 70%.
   • The expected result for Aspergillus brasiliensis is a recovery of ≥ 50%.

The actual batch-related Certificate of Analysis provides specific quality control data for each batch of PDA, including the results obtained for the mentioned control strains. This document serves as a reference to assess the quality and performance of the PDA batch.

By implementing robust quality control measures, laboratories can ensure the reliability and consistency of PDA for the cultivation and identification of yeasts and molds in microbiological testing and research.

Uses of Potato Dextrose Agar

Potato Dextrose Agar (PDA) finds a range of uses in microbiology laboratories. Here are some common applications of PDA based on the provided information:

• Detection of Yeasts and Molds in Dairy Products and Prepared Foods: PDA is widely employed for the detection and enumeration of yeasts and molds in dairy products and various prepared foods. Its nutrient-rich composition and ability to support the growth of these microorganisms make it a suitable medium for assessing the microbial quality of food samples.
• Cultivation of Yeasts and Molds from Clinical Specimens: PDA can be utilized for the cultivation and isolation of yeasts and molds from clinical specimens, aiding in the identification and diagnosis of fungal infections in patients. It provides a favorable environment for the growth and observation of pathogenic and non-pathogenic fungi obtained from clinical samples.
• Microbial Examination of Food and Dairy Products: PDA supplemented with tartaric acid (TA) is specifically recommended for the microbial examination of food and dairy products. The addition of tartaric acid helps to lower the pH of the medium, inhibiting the growth of bacteria and creating a selective environment that favors the growth of yeasts and molds.
• Microbial Enumeration of Yeast and Mold from Cosmetics: PDA supplemented with chlortetracycline is recommended for the microbial enumeration of yeast and mold from cosmetic products. The inclusion of chlortetracycline as a selective agent helps to inhibit the growth of bacteria while allowing the growth of specific fungi, aiding in the assessment of cosmetic product quality and safety.
• Selective Cultivation of Fungi from Mixed Samples: PDA supplemented with chloramphenicol is recommended for the selective cultivation of fungi from mixed samples. Chloramphenicol acts as a selective agent, inhibiting the growth of bacteria and providing a favorable environment for the growth of fungi. This selective cultivation approach helps in the isolation and identification of fungal species present in mixed samples.

Limitations of Potato Dextrose Agar

While Potato Dextrose Agar (PDA) is a valuable medium for the cultivation and preliminary identification of yeasts and molds, it does have some limitations. Here is a specific limitation based on the provided information:

1. Identification Limitation: PDA provides a suitable environment for the growth and development of yeasts and molds, allowing for their initial observation and characterization based on colony morphology. However, to achieve complete and accurate identification of the fungal isolates, further testing is necessary. Biochemical, immunological, molecular, or mass spectrometry testing methods are recommended to be performed on colonies obtained from pure culture. These additional tests provide more specific and reliable information about the fungal species present.

It’s important to note that while PDA can support the growth of a wide range of fungi, it may not be the ideal medium for every type of fungus. Some fastidious or specialized fungi may require specific growth media tailored to their nutritional requirements.

In summary, while PDA is a valuable tool for the cultivation and preliminary identification of yeasts and molds, its limitations highlight the need for complementary testing methods to achieve complete and accurate identification of fungal isolates.

FAQ

References

• https://microbeonline.com/potato-dextrose-agar-pda-principle-composition-colony-characteristics/
• https://www.mycrobe.org/blog/2018/7/6/potato-dextrose-agar-pda
• https://sharebiology.com/potato-dextrose-agar-pda/#gs.gi6eed
• https://microbenotes.com/potato-dextrose-agar-pda/
• https://www.sigmaaldrich.com/deepweb/assets/sigmaaldrich/product/documents/418/488/tn1348en-mk.pdf
• https://www.appliedfooddiagnostics.com/products/micro-media-pd
• https://hardydiagnostics.com/w60
• https://www.fda.gov/food/laboratory-methods-food/bam-media-m127-potato-dextrose-agar
• https://himedialabs.com/TD/M096.pdf