Why do agar plates need to be dried

The incubator fans pull more moisture out of the growth promotions plates simply as there is more moisture still available within the agar than from the environmental monitoring plates. As such, you will see higher humidity traces in the growth promotion plate incubator.

It is necessary to control for this effect, as it could lead to condensation formation in your incubators which in turn can increase the risk of contamination.

Additives — neutralisers are known to contribute to the formation of cracks through the middle of the media as desiccation occurs. The addition of neutralisers forms chemical reactions within the media, reducing its strength, leading to sheering of the media and crack formation. Storage time and conditions — extended storage of a plate, despite being wrapped in protective layers, at higher temperatures before use, allows increased time for evaporation to take place, especially if the wrapping utilised is optimised for gamma irradiation and not moisture control.

It is essential to understand the material qualities of the wrapping used to protect plates. As alluded to above, certain materials, such as those of nylon make up, are optimised for irradiation to increase the passage of dosage and have higher permeability properties, thus allowing moisture out of the packaging. I advise that during media qualification studies, you consider shelf-life storage. Observe the media for the formation of growth and or condensation within the packaging.

Your observations may alert you to an issue with the packaging. Broad temperature range changes — rapid and extreme temperature changes in storage and in-use can lead to excess water loss from media. For example, the moving of the plates between refrigerated storage and ambient cleanroom environments. While it is acceptable to utilise refrigerated storage media in your environmental monitoring programme with appropriate controls to bring the media to ambient conditions before use, give consideration to whether this may be leading to desiccation issues.

The concentration of agar in the plate — for solid media agar that requires irradiation to ensure sterility, suppliers can add additional agar to the media to better 'fix' the water into the medium.

My findings show that irradiation can lead to water loss and that desiccation after that can be an unavoidable issue. Increased agar can lead to increased sheering forces when moisture is lost and thus may be the cause of crack formation in your media.

Consistency and variability of the supplier's manufacturing process — minor changes or variability in manufacturing processes of media can lead to changes in media performance at a site. These variations include a change of supplier for base agar formulations. If you observe a step-change in the number of desiccated plates, check with your manufacturer for modifications to the manufacturing process.

Consider even like for like changes in your assessment of possible root causes for desiccation. Another area for consideration is the transport of media to your site. Has the supplier experienced any issues in its logistics management of your supply of plates, are the plates stored for longer than previously seen, for example, is there a new carrier transporting your media to site.

Particle bounce — particles bouncing off the surface of media can increase the evaporation effects across the surface of an agar plate.

Particle bounce is more likely to be an issue in higher particulate environments and as such is not considered a problem for low bioburden cleanroom facility or aseptic manufacturing site. This issue is noted here to inform the reader that it should be a consideration if your site is likely to experience this type of environmental challenge.

High doses of irradiation — high doses of irradiation can lead to chemical alteration oxidation of the media, which in turn leads to desiccation issues. A dose of 10 kGy is considered safe for the sterilisation of media and suppliers across industry use between 10 — 20 kGy. Doses above 15 kGy are a potential source of investigation should you see desiccation in your solid media.

Failure to validate exposure times — not all media are created equal or will be suitable to use on your site. When purchasing new media, validate your exposure times to show that the media is capable of supporting growth post-exposure.

The Redipor range of irradiated agar comes in a number of fill volumes to meet the needs of your working environment: 18ml is the standard fill volume for 90mm petri dishes 27ml is our deep filled equivalent.

This is the most popular for high air flow environments 32ml is our extra deep fill and also the maximum we can fill to. These are for environments where dehydration is a real problem Assessing operating environment In regards to how the agar shrinks, ideally it will shrink downwards before it shrinks inwards. Selecting media formulation We are aware of issues where the agar cracks down the middle as it shrinks.

Sandle T. Settle plate exposure under unidirectional airflow and the effect of weight loss upon microbial growth.

The pharmaceutical and cleanroom industry's pocket guide to prepared culture media. Sign Up for our Blog. We promise that we won't SPAM you. Make sure you keep the color code to hand though. Now you should have no issues making agar plates that are perfect every time. Originally published July 5, Reviewed and updated February Has this helped you? Then please share with your network. Hi pouring in plates should be under laminar hood?

I have to autoclave the plates before pouring? Hi, I made NGM agar with the powder and water, added the dextrose, autoclaved it, then plated it. I found out today that I missed out on 3 ingredients. Can I save the current media by heating the plates and returning the liquid to the jar I made it in, and add the required ingredients and autoclave again? Facebook Twitter LinkedIn More. As expected, the weight loss of agar in Petri dish increased with increasing drying time.

As the drying time increased, MCF generally decreased Figure 3. For E. This means that the software can differentiate the patterns of colonies grown on agar that was air-dried for a minimum of 10 min as a unique scatter pattern. Similarly, E.

O and O at 10, 30 and 60 min of drying time, respectively. This means each serovar was affected differently by short drying time of agar plates Figure 3. We were also intrigued to observe the effect of extreme air-drying conditions abusive or mal-handling practices on the scatter pattern. Same analysis was conducted using different air-drying parameters during preparation of the Petri dishes, where the SMAC agar plates were subjected to extreme drying conditions.

There was a substantial loss of surface water layer and weight of agar plate during extreme drying condition Table 3. Results of this study suggest that scatter patterns of serovar O26 were least affected while serovar O and O were severely affected after growth on plates that were subjected to extreme drying conditions as calculated by MCF values Figure 4.

However, in the case of E. In case of E. However, the high. Table 2. Surface water layer and relative weight loss of the SMAC agar depending on the short drying time min after solidification. Table 3. Surface water layer and relative weight loss of the SMAC agar under extreme drying condition.

The variations in the findings of short and extreme drying conditions must be related to loss of agar media water content, which was found to be reduced by 2. Such variation in colony profile will impart immense difference in BARDOT scatter patterns based on physical theoretical variations of light forward scattering as mentioned in our previous studies [18,27]. This may be the reason for the irreproducible and incongruent scatter patterns under extreme drying condition, when compared to short drying condition.

It was also observed that storage of agar plates in sealed plastic bag at room temperature has insignificant effect on the weight loss of agar plates. Here, it can be concluded that the storage of the Petri dishes in sealed plastic bag at room temperature for more than 5 days seems to have an impact on scatter pattern of serovar O26, whereas as it did not affect the pattern of serovar, O and O on plates when stored for 24 days at room temperature in sealed plastic bag Figure 5.

For up to 24 days of storage of agar plate in sealed plastic bag at room temperature, the surface water layer on the agar was not significantly different from the one of a fresh agar plate prepared at day one data not presented. Figure 4. Effect of weight loss of agar on the MCF value for scatter patterns of colonies of E. SMAC agar plates were air-dried for 3, 5, 7 and 9 h after solidification with plate lid open in a laminar flow biosafety cabinet.

There was a substantial loss of surface water layer and weight of agar plate during extreme drying condition see Table 3. Figure 5. MCF values for scatter patterns of colonies of three E. Finally, E. For the two serovars O and O26 , there must be additional factors that may cause changes during this storage period. However, for E. It does not appear that the surface water layer affected the BARDOT scatter images, because for those days, there was no significant differences on the surface water layer when compared with the others days data not shown.

Results obtained in this study revealed some limitations, which need further investigation: 1 this study was conducted using the three serovars of only one bacterial species grown on single selective medium; 2 weight loss and surface water layer experiments were extremely sensitive and dependent on relative humidity of the laboratory environment, which is influenced by seasonal variations throughout the year; 3 lack of Sensitive instrument and isolated, controlled environment for the experiments.

This study has to be expanded to include growth of different bacteria on selective or nonselective media. Application of sensible instrument to measure weight loss, surface water layer and other environmental parameter under isolated system will definitely reduce the level of variations in the experimental setups and measurement.

Extreme drying conditions for SMAC agar plate did not result in consistent scatter pattern and it must be emphasized that such extreme conditions of drying is not usual conditions of media preparation in the routine microbiological laboratory setup. Such extreme conditions were inducted in our study to analyze the level of sensitivity and robustness of the BARDOT system.

In future, it would be beneficial to monitor the effect of other experimental variables such as oxygen and nutrient availability on the optical scatter pattern of bacteria, which can also be quantified by determining MCF. Furthermore, the data from this study will help generate a numerical growth model of bacteria under altered environmental conditions. A quantitative MCF value also can be used as one of the parameters to observe the effect of various nutrients, chemicals or antibiotics in media formulation.

Figure 6. Diagram representing the cross-validation matrix and representative scatter patterns of the three serovars of E.

SMAC plates were subjected to air-drying up to 10 min after solidification and then stored in cold room for up to 30 days. In cross-validation matrix values presented in diagonal rows represent the level of similarity within the submitted batch and the values in anti-diagonal rows represent the level of similarity defined as MCF value in this study between the two batches of scatter images.

Lower MCF values Figure 8. This finding is extremely important. Figure 7. MCF values for scatter patterns of colonies of E. Figure 8.