I have problems with the new autoclave. We checked the sterilization process and realized that the biological indicator (B. stearothermophilus) grew after the sterilization.
The technician put the vial of the microorganism inside a vessel with the cap loosley tighten. After sterilization at 121 ºC for 15 min they put the vial in a oven at 60 ºC. Next day we checked the vial and there was growth in it.
I have repeated the process autoclaving at 121 ºC for 20 minutes but once again the bioindicator grew after "sterilization". When I put the vial in a beaker without any cap the sterilization of the microorganism worked. So, I thought the problem was that the temperatura inside of the vessel is not enough. I chechek the temperature with a sounding line and the temperature inside the vessel was 118-120 ºC.
However, they reply that they always have validated the autoclave in the same way without any problem. The technical support suggest to increase the temperature of the autclave to 124 ºC but the normative specifies 121 ºC
Why this problem now?
Dear Dr. Alba Blánquez ,
Geobacillus stearothermophilus strains have been reported in the past to be occasionally resistant to even the standard moist heat sterilization protocols and a positive bioindicator normally results from either the not-so-normal heat resistance by the thermophilic firmicutes or due to human errors as well as faulty equipments. There have been reports of the G. stearothermophilus strain to have a kill temperature as high as 132 degree Celsius and the kill time being over 17 minutes. In your case, overlooking the possible human errors, the spores may very well be extra-resistant. The most effective sterilization of such spores take place within the first half of the exposure to the sterilization cycle and failing to achieve it could have resulted in the positive response by the BI. The sounding line provided you the temperature inside the vial but is the actual pressure being raised inside the vial at 15 psi within the first 8 minutes? The loosely capped vial could very well be a reason behind the positive BI since it is expressively mentioned in various relevant literatures and protocols that the maintenance of the dryness inside the vial/ or avoidance of moisture build is prerequisite to negative BI response since moisture somehow prevents the core alpha glucosidase inactivation at the core of the spores.
You can try to position the vials differently in order to find out any dead spots inside the autoclave where the steam isn't properly circulating, as an added measure.
I hope this has helped you in your quest.
With regards,
Dr. Abhishek Mukherjee
Another form of autoclave indicator makes use of biological indicators, These are stadndardized populations of resistant bacterial spores such as Geobacillus sterothermophilus .....The bacterial spore coat test is used to determine if the sterilization cycle parameters were sufficient to kill the test microorganisms. The problems include * Not determining that the physical monitors were correct for the load * Not identifying that the incorrect cycle was run for the load contents * Using the wrong B1 or test pack/PCD for the load * Using the wrong class 5 integrating indicator or B 1 result *Using the wrong internal chemical indicator * Incorrect reading of internal chemical indicator * Incorrect storage od any chemical or biological indicators.* Inproper cycle for the load contents * Use of improper cycle for the load contents * Improper packaging materials or packaging technique. * Sterilizer loading * Poor steam quality and quantity wet steam * Superheated steam. For more details consult https://multimedia.3m.com
If the problem is heat transfer rate then it should be responsive to time
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