Characterization of antibiotic-resistance traits in Akkermansia muciniphila strains of human origin.

Akkermansia muciniphila, a commensal bacterium commonly found in healthy gut microbiota, is widely considered a next-generation beneficial bacterium candidate to improve metabolic and inflammatory disorders. Recently the EFSA's Panel on Nutrition, Novel food, and Food Allergens has declared that pasteurized A. muciniphila DSM 22959T (also MucT, ATCC BAA-835) can be considered safe as a novel food, opening the door to its commercialization as a food supplement. Despite its recognized health benefits, there is still little information regarding the antimicrobial susceptibility of this species and reference cut-off values to distinguish strains with intrinsic or acquired resistance from susceptible strains. In this study, we combined a genomic approach with the evaluation of the antibiotic susceptibility in five human A. muciniphila isolates. Genomic mining for antimicrobial resistance genes and MICs determinations revealed that only one strain harboring tetW gene showed resistance to tetracycline, whereas all A. muciniphila strains showed low sensitivity to ciprofloxacin and aminoglycosides with no genotypic correlation. Although all strains harbor the gene adeF, encoding for a subunit of the resistance-nodulation-cell division efflux pump system, potentially involved in ciprofloxacin resistance, the susceptibility towards ciprofloxacin determined in presence of efflux pump inhibitors was not affected. Overall, our outcomes revealed the importance to extend the antibiotic susceptibility test to a larger number of new isolates of A. muciniphila to better assess the safety aspects of this species.

Antimicrobial activity of resveratrol-derived monomers and dimers against foodborne pathogens.

Plant polyphenolic compounds are considered a promising source for new antibacterial agents. In this study, we evaluated the antimicrobial activity of a collection of resveratrol-derived monomers and dimers screened as single molecules against a panel of nine foodborne pathogens. The results demonstrated that two monomers (i.e., pterostilbene 2 and (E)-3-hydroxy-4',5-dimethoxystilbene 9) and three dimers (i.e., δ-viniferin 10, viniferifuran 14 and dehydro-δ-viniferin 15) were endowed with significant antibacterial activity against gram-positive bacteria. The exposure of gram-positive foodborne pathogens to 100 µg/mL of 2, 9 and 15 induced severe cell membrane damage, resulting in the disruption of the phospholipid bilayer. The most promising dimeric compound, dehydro-δ-viniferin 15, was tested against Listeria monocytogenes, resulting in a loss of cultivability, viability and cell membrane potential. TEM analysis revealed grave morphological modifications on the cell membrane and leakage of intracellular content, confirming that the cell membrane was the principal biological target of the tested derivative.

Development of omics-based protocols for the microbiological characterization of multi-strain formulations marketed as probiotics: the case of VSL#3.

The growing commercial interest in multi-strain formulations marketed as probiotics has not been accompanied by an equal increase in the evaluation of quality levels of these biotechnological products. The multi-strain product VSL#3 was used as a model to setup a microbiological characterization that could be extended to other formulations with high complexity. Shotgun metagenomics by deep Illumina sequencing was applied to DNA isolated from the commercial VSL#3 product to confirm strains identity safety and composition. Single-cell analysis was used to evaluate the cell viability, and ß-galactosidase and urease activity have been used as marker to monitor the reproducibility of the production process. Similarly, these lots were characterized in detail by a metaproteomics approach for which a robust protein extraction protocol was combined with advanced mass spectrometry. The results identified over 1600 protein groups belonging to all strains present in the VSL#3 formulation. Of interest, only 3.2 % proteins showed significant differences mainly related to small variations in strain abundance. The protocols developed in this study addressed several quality criteria that are relevant for marketed multi-strain products and these represent the first efforts to define the quality of complex probiotic formulations such as VSL#3.