The spore test: weekly biological monitoring, and exactly what CDC says to do when one comes back positive

What it says#

Three monitors run on a sterilizer, and CDC is clear that they answer different questions and that none of them substitutes for another.

Mechanical monitoring means checking the sterilizer gauges, computer displays or printouts and documenting the sterilization pressure, temperature and exposure time in the sterilization records. Because those parameters can be observed during the cycle, CDC says this "might be the first indication of a problem." The instruction attached to it is blunt: do not use instrument packages if the mechanical monitoring indicates that the sterilizer did not reach the required temperature, time or pressure.

Chemical monitoring uses sensitive chemicals that change color when exposed to high temperatures, or to combinations of time and temperature. A chemical indicator goes inside every package to verify that the sterilizing agent penetrated the package and reached the instruments; if the internal indicator is not visible from outside, an external indicator is used as well. Inspect it immediately when removing packages, and if the color change did not occur, do not use the instruments. CDC distinguishes a single-parameter internal indicator, which reports one variable, from a multiparameter internal indicator designed to react to two or more, which gives a more reliable indication that sterilization conditions have been met.

Biological monitoring, the spore test, "assesses the sterilization process directly by killing known highly resistant microorganisms." CDC's reasoning is that because the spores used in biological indicators are more resistant than the common microbial contaminants found on patient-care equipment, a negative spore test indicates that other potential microorganisms in the load have been killed. The organism differs by method: the 2008 guideline says Bacillus atrophaeus spores monitor dry heat and ethylene oxide, and Geobacillus stearothermophilus spores monitor steam.

The frequency is at least weekly, per sterilizer, with a matching control from the same lot number. The 2003 guideline adds what the control is for: it is not processed through the sterilizer, is incubated alongside the test indicator, and must show growth, which is how you know the spores were alive to begin with. Follow the manufacturer's directions for where in the chamber the indicator goes. A spore test is also used for every load containing an implantable device, and CDC says ideally implantable items are not used until they test negative. Mail-in monitoring services are explicitly fine; the 2003 guideline notes that studies found mail delays have no substantial effect on final test results.

Then the part that matters most, which the dental page states as a numbered path and which is reproduced here in CDC's order and words. If the mechanical and chemical indicators suggest the sterilizer is functioning properly, "a single positive spore test result probably does not indicate sterilizer malfunction." First, remove the sterilizer from service and review the sterilization operating procedures to determine potential reasons for the failed test. Recall and quarantine any implantable items and do not use them until they are shown to be sterile. Items other than implantable items do not necessarily need to be recalled. Repeat the spore test immediately using the same cycle that produced the positive spore test. If the result of the repeat spore test is negative and operating procedures were correct, then the sterilizer can be returned to service. If the repeat spore test result is positive, remove the sterilizer from service and do not use until it has been inspected or repaired and re-challenged with spore tests in three consecutive fully loaded chamber sterilization cycles. And: "When possible, items from suspect loads dating back to the last negative spore test should be recalled, rewrapped, and re-sterilized."

CDC points from there to Table 12 of the 2008 guideline, a suggested protocol for managing a positive biological indicator in a steam sterilizer. Table 12 adds the checks to run while the sterilizer is down: confirm the sterilizer was used correctly, verifying the time and temperature settings; check for electrical irregularities or changes in the steam supply from the standard of at least 97% steam with less than 3% moisture; confirm the correct biological indicator was used and correctly interpreted; and if problems persist, request an inspection, discuss the abnormalities with the manufacturer, or repeat the test with a different manufacturer's indicator.

Who it applies to#

Every practice that owns a sterilizer, and every sterilizer separately. The at-least-weekly clock is per machine, not per office, which is the detail that trips up a practice that bought a second autoclave and kept running one test.

CDC addresses the obvious dodge directly. Asked whether a spore test is needed for a sterilizer that is not used full time, CDC answers that it "does not provide a separate recommendation for sterilizers that are used on a part-time basis" and repeats the at-least-weekly expectation with a matching control. There is no exemption for a slow schedule, a satellite office or a backup unit that only runs on Fridays.

What biological monitoring cannot reach is liquid chemical sterilization. CDC states that the sterilization process with liquid chemical sterilants cannot be verified with biological indicators, which is part of why heat-sensitive semicritical items are discouraged in favor of heat-tolerant or disposable alternatives. The classification that decides which items those are is covered in this directory's entry spaulding-classification-dental-instruments, and the cycle parameters your machine has to hit are in cdc-sterilization-cycle-parameters-dental.

What changed#

The substance is stable. One phrase is not, and an office should know which version it is following. The 2003 guideline says a sterilizer that fails a repeat test "should remain out of service until it has been inspected, repaired, and rechallenged with BI tests in three consecutive empty chamber sterilization cycles." CDC's dental sterilization monitoring page, updated May 15, 2024, says the same rechallenge happens "in three consecutive fully loaded chamber sterilization cycles." Empty in one document, fully loaded in the other. The current dental page is the newer statement, so that is the one to follow, and this entry reports both rather than picking a favorite.

The 2003 guideline also describes a more conservative approach it attributes to another source, in which any positive spore test is assumed to be a sterilizer malfunction and everything processed back to the last negative indicator is considered nonsterile, retrieved and reprocessed or quarantined. CDC says the margin of safety in steam sterilization makes that conservative, and that no published study documents disease transmission through a nonretrieved surgical instrument after a steam cycle with a positive biological indicator. But it also says the conservative approach "should always be used for sterilization methods other than steam," naming dry heat, unsaturated chemical vapor, ethylene oxide and hydrogen peroxide gas plasma. If you run a dry heat or chemical vapor unit, that is your rule.

Where the source is#

CDC's Best Practices for Sterilization Monitoring in Dental Settings, updated May 15, 2024, is a short free page on cdc.gov and is the link under the title; it carries the frequency, the three monitors and the failure path in the order given above. The 2003 dental guideline holds the longer narrative, the control indicator explanation and the non-steam conservative approach; it is free as an MMWR PDF and as a web page. Table 12 of the 2008 Guideline for Disinfection and Sterilization in Healthcare Facilities gives the suggested management protocol and is free in the same PDF. The 2016 Summary of Infection Prevention Practices repeats the weekly expectation in its key recommendations. All are public domain. During this check cdc.gov refused automated requests carrying a browser user agent and answered plain ones; the documents themselves are open to anyone.