Combined Interpretations of the 2003, 2009, and 2016 Standards that apply to Volume 1 of the 2016 TNI Standard
MODULE 5: MICROBIOLOGY TECHNICAL REQUIREMENTS
Question: Are microorganisms considered standards? Does the lab need to assign an expiration date on them? The reference cultures the lab receives from ATCC does not have expiration dates. The lab is following the protocol for microorganism listed in Appendix D3.7.
TNI Response: The ATCC reference cultures are "reference materials" and must adhere to the requirements of 126.96.36.199 (2003). These requirements include the requirement of having an expiration date whether supplied by the vendor or assigned by the laboratory. This requirement was removed in 2009 and remains as such. The SIR is obsolete.
Section: 188.8.131.52 Ongoing Demonstration of Capability Requirements
Question: For ongoing DOC e.g. for HPC SimPlate, the lab performs a blind PT sample or a Quality Control sample with results meeting the manufacturer's acceptance criteria. However, we would like to be able to use section 184.108.40.206.e) "A documented process of reviewing QC samples performed by an analyst, or groups of analysts, relative to the QC requirements of the method, laboratory SOP, client specifications, and/or this Standard. This review can be used to identify patterns for individuals or groups of analysts and determine if corrective action or retraining is necessary."
SM/TNI requires repeat counts performed monthly with criteria of 5% RPD for a single analyst or 10% for more than one analyst counting. Can this process be applicable or acceptable to meet section 220.127.116.11.e and be applied for continuing DOC for other analyst who did not actually perform the PT or QCS? If not, please expand on exactly what this section mean with a clear example.
TNI Response: Your example would apply to 18.104.22.168.e if the analyst using 22.214.171.124 only read out positive results for that method. Please note that 126.96.36.199 does not include determination of non-positive environmental or QC samples, and therefore a DOC following 188.8.131.52.e using 184.108.40.206 would not prove an analyst competent in determining non-positive environmental or QC sample results or competent at performing any other parts of the method. Therefore 220.127.116.11 cannot be used solely to meet the requirements for an ongoing DOC.
18.104.22.168.e refers to those instances where a lab may use other approaches to an on-going DOC, such as lab generated blind samples, replicate analysis, in-batch positives and negatives or other reasonable approaches. If the laboratory has a documented procedure for analyzing samples using the method, utilizing associated techniques (ex. aseptic technique) and meeting QC requirements, on a defined basis in a similar manner for all analysts (such as monthly positive and negative QC samples for each analyst), which could be used to determine patterns/trends, it could be used as documentation for an on-going DOC.
Section: 22.214.171.124 Quality and Sterility of Standards, Reagents, Materials, and Media
Question: How are other AB's interpreting this standard: "A sterility blank shall be performed on each batch of dilution water prepared in the laboratory and on each batch of pre-prepared, ready-to-use dilution water with non-selective growth media." Are you requiring a sterility check with non-selective growth media for any dilution water (i.e. Deionized, distilled, etc.) or just those dilution waters whose preparation is described in microbiological procedures, like phosphate buffer water or peptone water?
TNI Response: If sterility is required for use (whether in-house or purchased), then sterility must be checked prior to use. If the water is being used to prepare media or buffered water, then a sterility check is not needed until after the reagent has been prepared. A sterility check must be performed on any dilution water. Clarified in the 2016 Standard by adding in 126.96.36.199.a) "All materials and supplies that would be needed to process…."
Question: Can the standard be interpreted to mean a sterility blank is not required every 10 samples, as implied in the last sentence of the section, if the funnels are single use? It is assumed that the point of running the blanks every 10 samples is to show proper rinsing technique for multi-use funnels. Because the final sentence in this section begins with "In addition," and is displaced from the sentence that says, "For pre-sterilized single use funnels a sterility check shall be performed on one funnel per lot," it seems to add a requirement to filtration series using all types of funnels, but I do not believe that was the intent.
TNI Response: It was the intent that single use funnels, provided they are only used once, would not require any additional sterility check beyond once per lot as written in the Standard. However, if the method being followed has more stringent requirements, those requirements must be followed. The basic issue with this SIR is that until the 2016 standard made clear that sterility checks and method blanks are different things required for different reasons the language was confusing. Section 188.8.131.52 is about sterility checks, but this question is about method blanks.
Question: The lab conducts SM9020a testing on potable water only. We purchase single use sterile water from IDEXX who also manufactures the Colilert media. It is their recommendation to use sterile water in blanks and dilutions. This is the only use our lab has for sterile water as everything else is prepackaged ready to use. The sterile water comes in sealed 100ml aliquots with reagent water criteria certification from IDEXX. The water is purchased in lots and the lot is verified upon receipt for pH, conductance and sterility. It is stored in a refrigerator at 4C. The lab is small and only conducts 20 tests per month. Is the in-house QA and manufacture certification of the sterile water sufficient for the use of the water as a lot per TNI standards - or should it be conform to pre-dispensed buffer water standards?
TNI Response: The requirement is to check the purchased water once per lot and, in addition, once per month when using a bottle that lasts longer than one month. If the water is to be used for preparation of media, reagents or as a diluent, which will put it in contact with microorganisms, then the requirements of verification as stated in 184.108.40.206(c) are needed along with the verification of sterility. As this is something that is somewhat static and the source is not changing over time as an In-house DI system may be an initial check that mirrors the DI water requirements is reasonable with some relief regarding frequency as noted in 220.127.116.11.d) v. Refer to 18.104.22.168 e).
Question: V1M5: 22.214.171.124.b.i states, "All media shall be tested for performance (e.g., for selectivity, sensitivity, sterility, growth promotion, and growth inhibition). These tests shall be performed at a minimum with first use."
Is the intention to require that the 'minimum' testing MUST be performed during/with first use, or is 'before first use' acceptable?
TNI Response: Performance testing is required to be performed before the first use, or with the first use. The laboratory may determine the approach they prefer.
Section: 126.96.36.199 Method Blanks
Question: Can you please clarify what is meant by "methods that specify colony counts such as membrane filter or plated media." Does this mean all methods that are enumeration? Or just the membrane filter and plated media methods?
TNI Response: Comparability of responses found in environmental microbiological testing is an important laboratory tool for ensuring comparability between multiple analysts or parallel counts determined in a laboratory with only a single analyst. The language in the 2009 Standard relates to the colony counts determined with membrane filter testing and plated media.
Section: 188.8.131.52 Variability/Reproducibility of Quanti-Tray
Question: My question is regarding the Test Variability/Reproducibility section in the new 2016 standard. The verbiage essentially states to perform for methods that specify counts (i.e., cfu/100 mL or MPN/100mL)..." or other methods which specify a quantitative result." My question is, was the intent to hold labs accountable for performing this for IDEXX Quanti-tray as well? It is an MPN method; however, I did not feel the intent was to perform for this method but rather other MPN methods (e.g., multiple tube fermentation). Furthermore, if a laboratory is not obtaining any positive samples over the course of a month using this method, would they still be required to perform one? Standard methods instructs to perform this for plate counting methods, so I wanted to make sure the intent as to what methods to hold labs accountable for.
TNI Response: The standard is clear on the inclusion MPN methods and this does not exclude Quanti-tray. Furthermore, the STD states 'duplicate counts shall be performed monthly'. It would be up to the lab to determine how they obtain the positive sample with which to perform the counts.
Section: 184.108.40.206 Selectivity
Question: In Standard Methods there is a requirement to do at least one positive sample verification monthly, in some cases 10% etc. What if you do not have any positive samples, should you do a positive verification anyway? I see that for DW (source) even if you do not have any positive samples, a positive verification check is to be done quarterly. How about the other type of waters? For example to check any false negatives? Frequency?
TNI Response: Questions involving specific methods should be directed to the writer of the method. The requirement for positive controls exists so that the lab can demonstrate that IT isn't the reason there are no positives, i.e., it isn't doing something that causes no growth.
Section: 220.127.116.11 Constant and Consistent Test Conditions
Question: What does "autoclave maintenance... shall include a pressure check" mean? Does that specifically mean that the pressure relief valve shall be checked and documented? If would be extremely helpful if these sections could be written more specifically.
TNI Response: Since PV=nRT, checking the temperature and assuring no leaks (so that V is constant) is sufficient to meet the requirements of the standard. Another suggestion: A note could be made in a logbook indicating that seals were checked and no cracks were found. The 2016 standard added in the PV= nRT language and called it sufficient.
Section: 1.7.5 Sample Handling
Question: Regarding microbiological samples and especially the potentially reduced frequency of checks for absence of chlorine residual for potable water sources (including source water): What are the definitions of "source" and "source water" in this context?
TNI Response: "Source water" is defined by EPA as "Untreated water from streams, rivers, lakes, or underground aquifers, which is used to supply private and public drinking water supplies"
Question: Regarding microbiological samples and especially the potentially reduced frequency of checks for absence of chlorine residual for potable water sources (including source water): What constitutes a unique "source"? Some considerations are:
a. A water body (stream or lake) as a source vs. discrete segments (i.e., random or recurring sampling locations) of the water body as unique sources of source water.
b. Individual wells producing source water.
c. A common aquifer with multiple wells producing source water from that aquifer.
d. A water distribution system as a source vs. individual sampling locations specified in the sampling plan for the distribution system as unique sources.
It seems like the intent of this section was to perform checks to verify removal of chlorine residual from samples from all new potable water sources and random checks for routine samples submitted by long-term customers whose samples satisfy all four conditions in 1.7.5.b.i to iv.
TNI Response: It is the intent of the standard to verify removal of chlorine residual from all samples that may have been chlorinated. This check is made on sample containers received by the laboratory, regardless of how they were collected. A definition of source was added in 1.3.1 of the 2016 standard and section 18.104.22.168 was extensively revised.
Question: Regarding microbiological samples and especially the potentially reduced frequency of checks for absence of chlorine residual for potable water sources (including source water): What constitutes sufficient documentation or objective evidence that "the laboratory can 'show' that the received sample container are from their laboratory?" For example, IDEXX containers provided by various laboratories to customers are indistinguishable, unless a laboratory marks the containers to show they came from a specific laboratory in a large nationwide chain of laboratories.
TNI Response: This question is a business decision as to how a lab would like to prove the bottle came from their laboratory and therefore not a question of clarity with regard to the standard.