Environmental testing positive in zone 2
Hello everyone,
I was presented with the following scenario and would like some input:
Environmental testing of zone 2 in chocolate production showed presumptive Listeria on the outside of dosing equipment and presumptive Salmonella around molds (tested by swabs). Each pallet produced was sampled, taking pieces of the product from each box, for a total of 10% of the boxes in each pallet randomly sampled. The pooled samples (per pallet) were tested for Listeria (in 25g) and Salmonella by PCR (in 375g), all came back negative.
Would you release this production? What additional testing would you do?
Thank you!
Presumptive positive is not always real positive AFAIK: I've experienced that several times, and it always appeared that re-testing of the same sample by different method showed negative. I don't know what methods your lab's using, but my lab uses some kind of sensitive testing machine, which sometimes reads product DNAs as Listeria ones (rough and approximate explanation from the lab guys)
Presumptive positive is not always real positive AFAIK: I've experienced that several times, and it always appeared that re-testing of the same sample by different method showed negative. I don't know what methods your lab's using, but my lab uses some kind of sensitive testing machine, which sometimes reads product DNAs as Listeria ones (rough and approximate explanation from the lab guys)
Yes, the DNA testing was the one used for Salmonella.
The thing that is bothering my customer is that if there was indeed a spot contamination (not the entire production), how would he be sure with sampling?
I would release the products as multiple representative samples showed clean. However, I would wait till other forum members' opinion - the more, the better:) RE: additional test - did you try using different method? Your decision would also depend on a batch volume: if it's relatively small and not that costly, you might consider disposing it rather than being worried...
I would release the products as multiple representative samples showed clean. However, I would wait till other forum members' opinion - the more, the better:) RE: additional test - did you try using different method? Your decision would also depend on a batch volume: if it's relatively small and not that costly, you might consider disposing it rather than being worried...
The methods were different. The environmental testing was done by swabbing and the sample testing was done by PCR or ELFA.
The product is quite costly. It was produced around 8,000 lbs.
I would NOT release the product-------see the maple leaf food white paper
A) did you follow AQL when selecting the samples? 10% does not sound like enough to me (agree with your customer) how large is a batch? is that 10% by piece or by weight
B) presumptive for SPECIES or generic?
C) testing for anything is particularly challenging in chocolate as the microbes tend to "bind" in the cocoa butter and can become almost undetectable
I would NOT release the product-------see the maple leaf food white paper
A) did you follow AQL when selecting the samples? 10% does not sound like enough to me (agree with your customer) how large is a batch? is that 10% by piece or by weight
B) presumptive for SPECIES or generic?
C) testing for anything is particularly challenging in chocolate as the microbes tend to "bind" in the cocoa butter and can become almost undetectable
Do you have a link for this white paper?
A) Each pallet has 60 cases (each with 30lbs). Out of that, 6 cases were sampled per pallet. Around 1 lb. of sample per box was taken.
B) Presumptive for Listeria spp. and Salmonella enterica
Some additional reading
Some chocolate manufacturers have implemented a precautionary pasteurization step for all of the raw ingredients, such as chocolate liquor, coming into the chocolate production plant (Cordier, 1994). However, others appear to be moving in the opposite direction in response to the consumer's increasing interest in 'superfoods', such as 'raw' chocolate, which is characterized by its low-temperature thermal processing (Nieburg, 2019). In the absence of a roasting step, the ability of Salmonella to survive the subsequent conching and tempering steps in chocolate-making i) presents a public health concern and ii) suggests that other bacterial pathogens with increased thermotolerance in low aw environments such as L. monocytogenes may also be able to survive the process (Sumner et al., 1991; Tsai et al., 2019; van Asselt and Zwietering, 2006).
https://atrium.lib.u...ed=y&sequence=3
https://www.scienced...362028X22103844
We used Salmonella detection in dark chocolate as a model to test the impact of different enrichment times (minimum and maximum validated times) and procedures on detection of low levels of difficult-to-detect Salmonella strains, for three PCR kits that were AOAC International Performance Tested Method certified for detection of Salmonella spp. in dark chocolate. Initial inclusivity studies with pure cultures showed that all three kits detected 70 of 70 Salmonella spp. strains at 1 log above the theoretical limit of detection, with some strains yielding later cycle threshold values or having variable detection among technical replicates, indicating reduced assay performance for these strains. Based on these data, we selected a S. enterica subsp. enterica serovar Poona strain as well as three non-subsp. enterica strains to test the ability of the three kits to detect Salmonella in dark chocolate inoculated at low levels (0.06 to 1.18 most probable number per 25 g). With primary enrichment in skim milk at 35°C, detection frequency for all assays did not significantly differ from the reference method for both the minimum and maximum validated enrichment times. However, a pilot study that used primary enrichment in buffered peptone water at 42°C yielded significantly fewer positive samples (13 of 80) than were obtained with the U.S. Food and Drug Administration Bacteriological Analytical Manual method using enrichment in skim milk at 35°C (40 of 80 positive samples); strains representing subsp. houtenae and salamae were detected in significantly fewer chocolate samples than enrichment with skim milk. Our data indicate that continued efforts to simplify rapid pathogen detection kits may reduce kit performance in a way that can only be detected with stringent evaluation protocols that are designed to identify kit failure modes.
Hi rdquality,
I would be rejecting the product, Chocolate is notorious for being a risk even with very low levels of salmonella contamination and your sampling may not pick it up. Ideally you should be shutting down the line for a deep clean and carrying out extra environmental sampling.
To add to the useful info posted by Scampi, there is something to be learned from the Cadbury salmonella outbreak and their interpretation of results.
Unanswered questions in Cadbury salmonella case
“Cadbury’s method of analysing and interpreting the lab results was also outdated. They were using a statistical method that relies on an even spread of the contaminant, but that is not the way it works. Chocolate is not homogenous. You could have salmonella in three bars and none in several thousand. You can’t measure the risk by averaging out the infection across all the bars.”
Also see Lesson Two: Being extra cautious can pay off
In short the site that continued production caused 500 people to be hospitalised.
Kind regards,
Tony
Some additional reading
Some chocolate manufacturers have implemented a precautionary pasteurization step for all of the raw ingredients, such as chocolate liquor, coming into the chocolate production plant (Cordier, 1994). However, others appear to be moving in the opposite direction in response to the consumer's increasing interest in 'superfoods', such as 'raw' chocolate, which is characterized by its low-temperature thermal processing (Nieburg, 2019). In the absence of a roasting step, the ability of Salmonella to survive the subsequent conching and tempering steps in chocolate-making i) presents a public health concern and ii) suggests that other bacterial pathogens with increased thermotolerance in low aw environments such as L. monocytogenes may also be able to survive the process (Sumner et al., 1991; Tsai et al., 2019; van Asselt and Zwietering, 2006).
https://atrium.lib.u...ed=y&sequence=3
https://www.scienced...362028X22103844
We used Salmonella detection in dark chocolate as a model to test the impact of different enrichment times (minimum and maximum validated times) and procedures on detection of low levels of difficult-to-detect Salmonella strains, for three PCR kits that were AOAC International Performance Tested Method certified for detection of Salmonella spp. in dark chocolate. Initial inclusivity studies with pure cultures showed that all three kits detected 70 of 70 Salmonella spp. strains at 1 log above the theoretical limit of detection, with some strains yielding later cycle threshold values or having variable detection among technical replicates, indicating reduced assay performance for these strains. Based on these data, we selected a S. enterica subsp. enterica serovar Poona strain as well as three non-subsp. enterica strains to test the ability of the three kits to detect Salmonella in dark chocolate inoculated at low levels (0.06 to 1.18 most probable number per 25 g). With primary enrichment in skim milk at 35°C, detection frequency for all assays did not significantly differ from the reference method for both the minimum and maximum validated enrichment times. However, a pilot study that used primary enrichment in buffered peptone water at 42°C yielded significantly fewer positive samples (13 of 80) than were obtained with the U.S. Food and Drug Administration Bacteriological Analytical Manual method using enrichment in skim milk at 35°C (40 of 80 positive samples); strains representing subsp. houtenae and salamae were detected in significantly fewer chocolate samples than enrichment with skim milk. Our data indicate that continued efforts to simplify rapid pathogen detection kits may reduce kit performance in a way that can only be detected with stringent evaluation protocols that are designed to identify kit failure modes.
Hi rdquality,
I would be rejecting the product, Chocolate is notorious for being a risk even with very low levels of salmonella contamination and your sampling may not pick it up. Ideally you should be shutting down the line for a deep clean and carrying out extra environmental sampling.
To add to the useful info posted by Scampi, there is something to be learned from the Cadbury salmonella outbreak and their interpretation of results.
Unanswered questions in Cadbury salmonella case
“Cadbury’s method of analysing and interpreting the lab results was also outdated. They were using a statistical method that relies on an even spread of the contaminant, but that is not the way it works. Chocolate is not homogenous. You could have salmonella in three bars and none in several thousand. You can’t measure the risk by averaging out the infection across all the bars.”
Also see Lesson Two: Being extra cautious can pay off
In short the site that continued production caused 500 people to be hospitalised.
Kind regards,
Tony
But would you reject the entire production even if the environmental testing was presumptive positive, without actual confirmation by another method?
I would also toss it, rerun SOP's, test again, then run and release.
Presumptive? So is the lab retesting or still testing and providing you with a definitive answer?
If they alerted you of presumptive positive, they would then have to confirm it with you. If you are just at presumptive positive stage, I would have all of product on Hold until you receive the final result. If it comes back as positive for sure, I would toss.
In my past, when we were alerted of presumptive positive, the line would be shut down, additional swabbing in a certain pattern around where the presumptive was would be done. Then we would have sanitation clean and re-swab again. You at the very least need a mapping of where this probably came from even if it ends up being negative. We called it a mini swab-a-thon. Not sure if that helps. I have been out of food for 3 years now, so I would tend to go with what the other fine people who work in food on here said.
Presumptive? So is the lab retesting or still testing and providing you with a definitive answer?
If they alerted you of presumptive positive, they would then have to confirm it with you. If you are just at presumptive positive stage, I would have all of product on Hold until you receive the final result. If it comes back as positive for sure, I would toss.
In my past, when we were alerted of presumptive positive, the line would be shut down, additional swabbing in a certain pattern around where the presumptive was would be done. Then we would have sanitation clean and re-swab again. You at the very least need a mapping of where this probably came from even if it ends up being negative. We called it a mini swab-a-thon. Not sure if that helps. I have been out of food for 3 years now, so I would tend to go with what the other fine people who work in food on here said.
The environmental testing was done by swabbing - these methods give a presumptive result. However, it can take up to 48 hours to get the results back. By the time the results were available, the line was thoroughly cleaned and sanitized and additional swabbing of the area came back negative. All results of the product itself came back negative.
Ah, I would rather err on the side of caution and discard as at this point you have no idea if that production would have been affected or not. Another concern is you now have no idea of the potential cause. You have a presumptive positive, there is no way in my mind you could justify releasing product. You release and people get sick, you have that presumptive positive and the weight of the law would come after you I would think. Sorry. Not a fun position to be in at all.
The lab should have (automatically) confirmed presumptive results. Afaik just involves additional analytical work..
Negative sampling/resampling can never prove non-presence of a pathogen in a lot..
And the converse for a confirmed positive result.
But would you reject the entire production even if the environmental testing was presumptive positive, without actual confirmation by another method?
YES---that is how this process is supposed to work----------you should never default to "the product is fine" when enviro tests get a presumptive this is particularly true in products like chocolate
See the previous post about the challenges that chocolate has and this is WHY chocolate recalls tend to be very large--------everything since the last clean swabs should theoretically be recalled (unless of course your company has amazing insurance that somehow covers them for negligence--which in my mind is what this would be)
Based on what our external lab has told us, presumptive positive EM results indicate that the area sampled has Listeria cells which are either dead or alive.
When we receive a PP result, the next step is either Culture Confirmation (are the cells living or dead) or DNA sequencing (determines the species).
We treat all PP the same without moving forward to either step since it doesn't matter to us whether the cells are living, dead, or from a non-pathogenic species. We treat every PP as if we had found living L. mono cells in the sampled area and treat it accordingly based on the zone.
Based on what our external lab has told us, presumptive positive EM results indicate that the area sampled has Listeria cells which are either dead or alive.
When we receive a PP result, the next step is either Culture Confirmation (are the cells living or dead) or DNA sequencing (determines the species).
We treat all PP the same without moving forward to either step since it doesn't matter to us whether the cells are living, dead, or from a non-pathogenic species. We treat every PP as if we had found living L. mono cells in the sampled area and treat it accordingly based on the zone.
Hi GQS,
Yr (conservative) follow-up action is up-to-you however from a purely microbiological POV, the action could be considered "illogical".
Presumptive Positive Procedures are naturally attractive when the results are always negative. If otherwise, there are possibilities for "dissension".