Learning Objectives

1. Explain the purpose of viable air sampling
2. Discuss how conclusions can be made from the results of viable air sampling

Happy Friday!

This weeks 483 we're going to focus on viable air sampling, which is one part of an environmental sampling plan. The information gained from viable air sampling can be very telling of the cleanliness and control you have over your cleanroom.

In this particular 483 viable air sampling was performed by a third party testing company for the purposes of re-certifying the cleanroom. Here's what the observation said:

So if you're unfamiliar with viable air sampling, it's the type of sampling where you use a piece of equipment to draw in a known volume of air and impact the air on a plate of growth media to see if there are microorganisms in the air.

This 483 states that a third party contractor had done the viable air sampling inside their ISO 5 engineering control or hood and in their ISO 7 buffer room. What's interesting here is that the 483 shows the results from not one certification of their cleanroom but the inspector reported a second certification six months later as well. What's most concerning is the April 2019 results where a total of 19 CFU or colony forming units were found. However, six months later, while there was still microorganisms found in the air in the same location, it was much less with only one found. Let's continue to just focus on the ISO 5 location as this is the most critical part of the cleanroom as this is where the compounding is performed.

In either case, April and October, an investigation should have been done in the cleanroom to figure out what the underlying issue is. I encourage you to go back and listen to the podcast episode on Investigations and CAPAs for learning how to properly investigate and document microbial excursions.

In this 483, because it was a third party contractor, they also reported the type of organisms present. This can be extremely helpful in finding out the root cause of the contamination. In April, 19 CFU were found and they were:

• Bacillus
• Coagulase negative Staphylococcus
• Micrococcus

There's actually a chart within USP <797> that tells us when we should take action if there's greater than 1 CFU found to be in our ISO class 5 area. So the fact that in April there were 19 found, an investigation absolutely should have been performed.

Looking up where these particular microorganisms come from would be the first step in an investigation. Bacillus are gram positive bacteria that are usually able to form spores, which are highly resistant when they're in their spore form. Bacillus can be found in soil or can be of animal origin.

Coagulase negative staphylococcus and micrococcus are typically normal flora found on humans. This information gives us two possible places to look for ingress routes for these organisms: materials and people or operators inside the cleanroom.

Remember, these were found in their ISO class 5 engineering control which is supposed to be extremely clean and free of microorganisms at any given time. So this could also suggest that maybe there's something wrong with their engineering control as well as it should be actively pushing microorganisms out of the hood.

In an investigation, we would possibly look at:

• Gowning procedures for operators
• Cleaning procedures: what agents are being used, the frequency of cleaning
• Operator technique while inside the cleanroom
• Our process for bringing materials into the cleanroom
• The engineering control itself: is it working properly, is there enough airflow, is there anything wrong with the HEPA filters

Six months later when the recertification was done again, there was only 1 CFU of bacillus found in the ISO 5 hood. This suggests that the firm did possibly investigate the cause of the contamination and made some improvements. However, there was no documentation found to definitively say that was done. I will say it might've been helpful if there was also results from surface sampling in the hood as this might give us further indication of what was causing the contamination.

Without sharing the entire 483, there are other key issues found that suggest other possibilities. For instance observation 2 notes that cleaning is performed in the ISO 5 area with non-sterile cleaning agents and wipes. Observation 6 discusses how there's a loose light fixture inside the ISO 5 laminar airflow hood. Observation 8 points out that prior to entering the cleanroom some materials were not disinfected. In observation 9 it's discovered that there's signs of rust and chipped paint inside the ISO 5 laminar airflow hood.

Needless to say, there's a lot of issues that we can look at and remediate to fix the underlying contamination. USP <797> says:

Data collected in response to corrective actions must be reviewed to confirm that the actions taken have been effective. The corrective action plan must be dependent on the cfu count and the microorganism recovered. Some examples of corrective action include process or facility improvements, personnel training, cleaning and disinfecting, or HEPA filter replacement and/or repair. The extent of the investigation should be consistent with the deviation and should include an evaluation of trends. The corrective action plan must be documented.

USP <797> Data evaluation and action levels

The point I'm trying to make here is that there's an abundance of information given for us to find and remediate the root cause of this particular contamination. The abundance of CFU found in the April 2019 recertification should have prompted the pharmacy to fix at minimum:

• Cleaning, materials agents and frequencies should be examined, possibly no longer using non-sterile wipes to clean and disinfect the hood
• Operator's gowning should be examined to see if people are gowning properly and if the gowning itself is sufficient (is there skin exposed while inside the laminar airflow hood)
• The visible rust and other issues with the ISO 5 area should have been remediated

Another point to be made is that viable air sampling is only performed every six months during recertification of the cleanroom. Perhaps after the first results, where there was 19 CFU found in the ISO 5 area, that should've also prompted more frequent viable air sampling. No follow up was performed until six months later. It's hard to make the case of any trend taking place when the data points are six months apart from each other. Follow up viable air sampling most definitely should've been performed to see if any of the improvements actually did remediate the issue. Also, sterile operations didn't cease when there was 19 CFU found, which calls into question any of the batches that