Question: Are the chapters <72> and <73> applicable and what is the impact on the validation procedure?
Answer: From a compendial standpoint, a USP General Chapter numbered below 1000 is applicable or compendially required if it is referenced in a monograph, another applicable General Chapter, or General Notices. USP <72> and <73> will become official in August 2025 but are not applicable until they are referenced in a monograph, in another applicable General Chapter, or in the General Notices. Therefore, <72> and <73> are considered alternative methods and must meet the requirements in the General Notices, 6.30 Alternative and Harmonized Methods and Procedures. Thus, for the methods applying <72> and <73>, a primary validation and a verification of the method is required prior to performing testing.Primary validation should demonstrate that the RMM system is adequate in detecting the intended target and must characterize the principle of detection. It may be carried out by the vendor of the technology or by the end user. It is the responsibility of the user to review the supplier's primary validation package.
The user must verify that the selected method is appropriate for use in testing specific products or materials. A verification must include specific experiments to confirm that the method is suitable for its intended purpose under the conditions of use for the material, drug substance, and/or drug product. The method suitability procedure described in <72> and <73> can be used for method verification as it was designed with more stringent requirements. The test strains selected for the suitability testing should be relevant to the product and manufacturing process and supported by a suitable justification.
Regulatory authorities may require supplemental data in addition to the primary validation and suitability as per <72> and <73> prior to acceptance. Users are encouraged to consult the relevant regulatory authority to discuss submission requirements.
Question: For the method suitability test is the inclusion of slow growing microorganisms and/or local isolates mandatory?
Answer: Inclusion of slow growing microorganisms and/or local isolates may be considered if relevant for the process or product risk. It is not an obligation to include slow growing microorganisms or local isolates if appropriately justified.
For instance, if the rapid microbiological method is used as a rapid screening test with a detection of less than 48h and most of the typical contaminants are fast growing microorganisms there might be no need to include a slow-growing microorganisms.
Question: Would short life products also consider cryopreserved cell therapy products? What falls in the scope of short life products?
Answer: Short-life products are defined in <1071> as products with a short-shelf life or with a short time between manufacturing and administration and for which a sterility test requiring a 14-day incubation is not suitable. These products may include compounded sterile preparations (CSPs), nuclear medicine products, and Advanced Therapy Medicinal Products (ATMPs). For example, many radiopharmaceuticals would be considered short-life products due to the short half-life of radiotracers, while CSPs and autologous cell therapies would be considered short-life products due to their short beyond-use dating. Cryopreserved products may fall under the definition of short-life products if they have to be administered to patients within a short time period.
Question: How does one determine the generation time of a slow-growing microorganism?
Answer: The procedure can be found in general Microbiology books. Calculations are detailed in <72> & <73>.
Question: If my inoculum control count of one test strain is above 10 CFU do I fail the method suitability test?
Answer: Considering the variability of the microbial inoculum and assuming it follows a Poisson distribution, it is possible that a low sample size targeting not more than 10 CFU may have an inoculum control that exceeds 10 CFU. A scientific justification would be required if the inoculum control final mean value exceeds 10 CFU and is maintained for the suitability test results. For instance, even if individual inoculum replicate counts on the plate might exceed 10 CFU, the mean value may still lie within 10 CFU. Introducing a higher number of replicates (triplicates instead of duplicates) and targeting the inoculum size to slightly less than 10 CFU mean value would reduce the risk of exceeding 10 CFU.
Mean value assuming Poisson distribution (P) 0 counts (P) exceeding 10 CFU in one replicate (P) exceeding 10 CFU in two replicates (P) exceeding 10 CFU in three replicates 8 <0.1% 18.4% 3.4% 0.6% 6 0.2%
4.3%
0.18%
<0.1%
4 1.8%
0.8%
<0.1%
<0.1%
Question: what type of nutrient media may be used for detecting micro aerophilic microorganisms, may gradient-layered facultative media such as fluid thioglycollate be utilized?
Answer: Different media formulations are provided by manufacturers. There is no preferred media, but microorganism recovery must be demonstrated.
Question: How to determine a cut-off level for the ATP direct inoculation method?
Answer: Either use the method provided by the vendor, or determine a statistically based cut off level.
Question: Do I need to use my own facility isolates or can I use a representative organism (i.e. same species) from a culture collection?
Answer: Preferably use facility isolates, but if this is not feasible then use a culture collection of the same species.
