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Is there a rapid method for quantifying pathogenic bacteria?

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#1 Heffer03

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Posted 27 October 2015 - 07:43 PM

Hello everyone!
 
I have a question, I need some help with it. We're planning to change the plating method to a rapid method, such as PCR, lateral flow, ELISA, etc, in order to analyse pathogenic bacteria, Listeria and Salmonella. Since we provide service to meat/poultry business. This would happen in a short term in our lab.
The thing is that this client, has a new requirement: they are asking for a method that would quantify Salmonella and Listeria. I've read that the qPCR would be a choice. But since, there's an enrichment step the quantification it's not possible.
Is there (in this world  :smile: ) a rapid method that would quantify pathogenic bacteria?
 
Thank you

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#2 Charles.C

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Posted 28 October 2015 - 02:19 AM

 

Hello everyone!
 
I have a question, I need some help with it. We're planning to change the plating method to a rapid method, such as PCR, lateral flow, ELISA, etc, in order to analyse pathogenic bacteria, Listeria and Salmonella. Since we provide service to meat/poultry business. This would happen in a short term in our lab.
The thing is that this client, has a new requirement: they are asking for a method that would quantify Salmonella and Listeria. I've read that the qPCR would be a choice. But since, there's an enrichment step the quantification it's not possible.
Is there (in this world  :smile: ) a rapid method that would quantify pathogenic bacteria?
 
Thank you

 

 

Hi Heffer,

 

Thanks for yr query and Welcome to the Forum ! :welcome:

 

The options may depend on yr specific requirements for speed, sensitivity, etc.

 

From Google, one possible solution  -

 

http://www.sigmaaldr...rapid-test.html


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Charles.C


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#3 sbarzee

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Posted 28 October 2015 - 02:32 AM

Why do you have to have an enrichment step?


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#4 Charles.C

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Posted 28 October 2015 - 05:22 AM

Why do you have to have an enrichment step?

 

 

To increase the concentation of target species.

 

A question of detection capability.


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Charles.C


#5 FishNChips

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Posted 05 November 2015 - 12:20 AM

Hello,

  I'm a long-time lurker, first-time poster.  As a microbiology lab. tech with experience using qPCR and immunoassays for Salmonella, I think I can offer some insight here. 

 

Like Charles C said, it's a question of limit of detection.  If you look at methods at the FDA BAM (http://www.fda.gov/F...s/ucm070149.htm) you ideally are detecting one viable cell per 25 g of material.  That limit is not going to be achievable by qPCR, immunoassay, or similar methods without enrichment - so there's no way to dispense with the enrichment step and maintain that L.O.D.   As the original post alluded to, there is no way to establish a correlation between the number of cells post-enrichment and pre-enrichment, so by doing the enrichment the method is no longer quantitative. 

 

If you had something that is highly contaminated (say, hundreds of cells per gram), yes you can do a direct DNA extraction and run qPCR.  I suppose this would be useful as an in-process control but it wouldn't meet criteria for finished-products.  


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#6 Charles.C

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Posted 05 November 2015 - 04:55 AM

Hi FishNChips,

 

Thks for the nice Post.

 

Please don't stop at One. :smile:


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Charles.C


#7 Heffer03

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Posted 05 November 2015 - 02:45 PM

Thank you both, for answering .

 

Is now clear the limitation of any rapid method with quantifying pathogens. 
 
I'm thinking that the best way to approach the situation would be: To run a qPCR, immunoassay or any rapid method in order to obtain the faster Negative results. Any presumptive Positive has to confirm with the reference method (since we're using a rapid method) and simultaneously we can perform a MPN analysis to find out the estimated pathogen population. 
It isn't the greatest option, if you think about the optimization of the lab workload. But that's the only one that it comes to mind...Do you have any other idea?  :helpplease: 

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#8 Charles.C

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Posted 05 November 2015 - 04:24 PM

Hi Heffer,

 

Note that the rapid methods in my earlier post are potentially quantitative and accredited. The point is whether their detection capability is low enough. As per post 5 maybe not ( I only use classical (cheap) micro. myself).

 

If not, only option I know is to add an enrichment step within a MPN based procedure. The time acceptability will then depend on yr definition of "rapid".

Obviously the MPN workload compared to direct is more if >= 3x3 tubes although could be potentially decreased somewhat via a non-routine array, eg 5-tube set (eg as used in water analysis). It all depends on sensitivity again.


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Charles.C


#9 3esa

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Posted 05 November 2015 - 05:42 PM

Why are you quantifying pathogenic bacteria to begin with?  Sorry I might have missed that.

 

Usually, anything with Salmonella or Listeria positive is already an adulterated product, regardless of numbers.


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#10 Charles.C

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Posted 05 November 2015 - 05:49 PM

Why are you quantifying pathogenic bacteria to begin with?  Sorry I might have missed that.

 

Usually, anything with Salmonella or Listeria positive is already an adulterated product, regardless of numbers.

 

Hi 3esa,

 

USA is not always globally copied.

 

See the parallel thread -

 

http://www.ifsqn.com...ria-spp-in-rte/


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Charles.C






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