Incubating plates at different temperatures, such as starting at 20 to 25 degrees Celsius and
then increasing to 30 to 35 degrees Celsius.
As optimum incubation conditions for yeasts & molds and bacteria is 22.5 ±2.5°C for 3 days
(72 hours) and 32.5±2.5°C for 2 days (48 hours) respectively, these must be incubated at
these incubation conditions for their proper growth. Now the question is that how to incubate
the single plate at two different temperatures?
We can incubate the same plate first at 32.5 ±2.5°C for 2 days and then at 22.5±2.5°C for
remaining 3 days. This is given in USP General Chapter <1116> Microbiological Control and
Monitoring of Aseptic Processing Environments, that was revised in 2012 and states that if
the lower temperature is selected first to incubate the plates then the growth of gram-positive
cocci may be affected.
These are associated with human- generated contamination So their recovery is important.
Incubation first at 32.5 ±2.5°C shall also help to grow the slow-growing microorganisms.
- 𝐈𝐧𝐢𝐭𝐢𝐚𝐥 𝐈𝐧𝐜𝐮𝐛𝐚𝐭𝐢𝐨𝐧 (𝟮𝟬-𝟮𝟱°𝗖)
This temperature range is often referred to as the “room temperature” incubation.
It is used to promote the growth of certain microorganisms, particularly those that thrive at lower temperatures or are more sensitive to higher temperatures.
Incubating at 20-25°C allows for the recovery of a broader range of organisms and can helpdistinguish between different species based on their growth characteristics. - 𝐒𝐮𝐛𝐬𝐞𝐪𝐮𝐞𝐧𝐭 𝐈𝐧𝐜𝐮𝐛𝐚𝐭𝐢𝐨𝐧 (𝟯𝟬-𝟯𝟱°𝗖)
After the initial incubation, plates are moved to a higher temperature range, typically 30 to 35 degrees Celsius. This temperature is closer to the optimal growth temperature for many common bacteria.
Increasing the temperature can help in the selective growth of specific bacterial species while inhibiting the growth of others. It is a way to encourage the growth of target organisms while discouraging unwanted contaminants. - 𝐒𝐞𝐥𝐞𝐜𝐭𝐢𝐯𝐞 𝐆𝐫𝐨𝐰𝐭𝐡
The two-step incubation process can be used as a selective method.
For example, somemicroorganisms may be psychrophiles (cold-loving), and they will primarily grow during the initial incubation phase, while mesophiles (organisms that grow best at moderate temperatures) will thrive during the subsequent higher-temperature phase. - 𝐈𝐝𝐞𝐧𝐭𝐢𝐟𝐢𝐜𝐚𝐭𝐢𝐨𝐧
Different species of microorganisms may have distinct growth characteristics at various temperatures. By using this two-step approach, microbiologists can obtain valuable information about the identity of the microorganisms present in a sample based on their growth behavior.