Researchers propose a ‘probiotic ID card’ based on DNA and flow cytometry
The new research was the work of investigators associated with several universities and research institutes in Parma and Milan as well as the APC Microbiome Institute in Cork. The research was published in the journal Frontiers in microbiology.
Advent of “probiogenomics”
The researchers used the term “probiogenomics” to describe their approach which produces what they called a “probiotic identity card” (PIC). According to the authors, this involved “The use of FC assays to determine the absolute abundance and viability of probiotic microorganisms in a given product/sample. In addition, gene-targeted metagenomic analyzes involving the 16S rRNA gene and ITS profiling have been replaced by shotgun metagenomics, allowing the classification of probiotics to the species level using a methodology of single sequencing.
To test the suitability of the approach, the researchers purchased twelve commercially available probiotic supplements in powder form. The supplements, which were not identified by brand, claimed from one to nine different species on the labels. The species claimed were a mixture of common probiotic strains, primarily from the Bifidobacteriumand Lactobacillusgenera, and included a spore-forming bacterium and a yeast species. The amounts listed ranged from one billion to 70 billion CFU (colony forming units).
Only half of the products generally met the label claim
Applying their methodology, the researchers found that six of the 12 products met the label claims in terms of the species present. Three of the products lacked at least one species claimed on the label, and two of the products contained other unidentified microbial species.
Additionally, the researchers claimed that three products were mislabeled as containing germs from the L.paracaseitaxon, instead of L. caseitaxon claimed on the label.
“This is a well-known problem since the strains belonging to L.casei and L.paracasei are phenotypically and genotypically closely related,”they said.
In terms of CFU counts, five of the six products that accurately indicated the microbial species present also accurately indicated CFU amounts. The sixth product, which contained multiple strains, gave only one CFU figure for all strains and was therefore not included in this part of the analysis. Another product gave an accurate number of CFUs, but was also contaminated with an undeclared strain. Tests on the last five products revealed serious discrepancies.
“The PIC pipeline described in this work improves on the previously proposed genetic identity map (Lugli et al., 2019), removing redundant sequencing experiments, such as 16S rRNA gene and ITS profiling, and allowing for the accurate enumeration of viable cells of each probiotic strain present in the probiotic supplement,”concluded the researchers.
“Using the PIC approach, five out of 12 probiotic supplements, or >40%, reveal inconsistencies in the formulations compared to what was declared, raising concerns about the current protocols applied by the internal quality controls of the producer of probiotic supplements”,they added.
API: a promising approach
The International Probiotics Association (IPA) said the approach detailed by the researchers could be a valuable addition to probiotic supplement testing, while acknowledging the questions raised by the high failure rate seen in this first application of the technique. .
“What is new in their approach is the use of molecular and not culture-based techniques. This should provide more reliable and accurate information. This is a step up from standard culture on MRS agar,”IPA said in a statement posted to NutraIngredients-USA.
“The good news in their findings is that half of the products tested contain the correct total numbers, the correct species, and those species at the correct numbers. The bad news is of course the other half,” says IPA.
IPA noted that some details were missing from the researchers’ analysis, such as how close the products were to expiration dates when tested. Another possible issue concerns the chain of custody; where and when were the supplements purchased, and could some of them have been mishandled along the way?
The IPA also mentioned that some manufacturing constraints of the probiotics were not mentioned in the document, such as the different stability and shelf life profiles of the strains in a multi-strain product, which could result in different amounts. of surpluses.
Complementary tool in search of quality
Overall, although the trade group viewed the document as a positive addition to the continued quest for quality probiotics.
“If there is anything to be desired in the analysis, it would be the testing of several batches of the same product or even the same batch but from different points of sale. To better understand if the poor quality was an unfortunate event or if it is more structural. Again, information on the best before date would be important,”said the IPA.
“While there are regulations in GMP for quality guidelines, the article further strengthens the case for the continued development of harmonization standards for quality products and enumeration technologies within our industry. . An initiative that the IPA has taken to heart and is currently working diligently towards,”concludes the group.
Source: Frontiers in microbiology
January 21, 2022 | https://doi.org/10.3389/fmicb.2021.790881
The probiotic ID card: a new “probiogenomics” approach to studying probiotic supplements
Authors: Lugli GA, et al.
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