Functional screening of antibiotic resistance genes from a representative metagenomic library of food fermenting microbiota.
Chiara DevirgiliisPaola ZinnoMariarita StirpeSimona BarileGiuditta PerozziPublished in: BioMed research international (2014)
Lactic acid bacteria (LAB) represent the predominant microbiota in fermented foods. Foodborne LAB have received increasing attention as potential reservoir of antibiotic resistance (AR) determinants, which may be horizontally transferred to opportunistic pathogens. We have previously reported isolation of AR LAB from the raw ingredients of a fermented cheese, while AR genes could be detected in the final, marketed product only by PCR amplification, thus pointing at the need for more sensitive microbial isolation techniques. We turned therefore to construction of a metagenomic library containing microbial DNA extracted directly from the food matrix. To maximize yield and purity and to ensure that genomic complexity of the library was representative of the original bacterial population, we defined a suitable protocol for total DNA extraction from cheese which can also be applied to other lipid-rich foods. Functional library screening on different antibiotics allowed recovery of ampicillin and kanamycin resistant clones originating from Streptococcus salivarius subsp. thermophilus and Lactobacillus helveticus genomes. We report molecular characterization of the cloned inserts, which were fully sequenced and shown to confer AR phenotype to recipient bacteria. We also show that metagenomics can be applied to food microbiota to identify underrepresented species carrying specific genes of interest.
Keyphrases
- lactic acid
- antibiotic resistance genes
- microbial community
- human health
- wastewater treatment
- circulating tumor
- nucleic acid
- single molecule
- cell free
- genome wide
- anaerobic digestion
- randomized controlled trial
- cross sectional
- bioinformatics analysis
- risk assessment
- genome wide identification
- climate change
- copy number
- working memory
- pseudomonas aeruginosa
- gene expression
- antimicrobial resistance
- real time pcr