Microbiota Analysis for the Optimization of Campylobacter Isolation From Chicken Carcasses Using Selective Media

Abstract

Since contaminated poultry meat is the major source of transmitting Campylobacter jejuni to humans, the isolation of Campylobacter from poultry carcasses is frequently performed in many countries as a baseline survey to ensure food safety. However, existing isolation methods have technical limitations in isolating this fastidious bacterium, such as a growth competition with indigenous bacteria in food samples. In this study, we compared the differences in microbiota compositions between Bolton and Preston selective media, two most common selective media to isolate Campylobacter , and investigated how different microbiota compositions resulting from different enrichment methods may affect isolation frequencies. A next-generation sequencing (NGS) analysis of 16S rRNA demonstrated that Bolton and Preston-selective enrichments generated different microbiota structures that shared only 31.57% of Operating Taxonomic Unit (OTU) types. Particularly, Escherichia was highly prevalent in Bolton selective media, and the enrichment cultures that increase Escherichia negatively affected the efficacy of Campylobacte r isolation. Furthermore, the combination of the selective media made a significant difference in the isolation frequency. The Bolton broth and Preston agar combination exhibited the highest (60.0%) frequencies of Campylobacter isolation, whereas the Bolton broth and Bolton agar combination showed the lowest (2.5%). These results show that each selective medium generates a unique microbiota structure and that the sequence of combining the selective media also critically affects the isolation frequency by altering microbiota compositions. In this study, we demonstrated how a microbiota analysis using NGS can be utilized to optimize a protocol for bacterial isolation from food samples.

Bacteriophage-Mediated Modulation of Bacterial Competition during Selective Enrichment of Campylobacter

“…Successful bacterial isolation relies on how effectively selective culture media can eliminate bacterial competitors while facilitating the growth of the target bacteria. Our previous studies demonstrated that each Campylobacter -selective medium makes substantial changes in the microbiome of enrichment cultures because of the selective pressure generated by antimicrobial supplements ( 11 , 12 ). Proteobacteria and Firmicutes are the major phyla after the enrichment of chicken carcasses with BB, whereas Proteobacteria and Fusobacteria were dominant after the enrichment with PB ( 11 ).…”

“…Our previous studies demonstrated that each Campylobacter -selective medium makes substantial changes in the microbiome of enrichment cultures because of the selective pressure generated by antimicrobial supplements ( 11 , 12 ). Proteobacteria and Firmicutes are the major phyla after the enrichment of chicken carcasses with BB, whereas Proteobacteria and Fusobacteria were dominant after the enrichment with PB ( 11 ). Commonly, antibiotic-resistant E. coli , such as ESBL-producing E. coli , is predominantly enriched in both BB and PB ( 11 ).…”

“…Proteobacteria and Firmicutes are the major phyla after the enrichment of chicken carcasses with BB, whereas Proteobacteria and Fusobacteria were dominant after the enrichment with PB ( 11 ). Commonly, antibiotic-resistant E. coli , such as ESBL-producing E. coli , is predominantly enriched in both BB and PB ( 11 ). Consistently, it has been reported that ESBL-producing E. coli prevalent on chicken carcasses significantly compromises the efficacy of Campylobacter isolation ( 7 , 11 ), suggesting the need for improving the current isolation methods.…”

“…Commonly, antibiotic-resistant E. coli , such as ESBL-producing E. coli , is predominantly enriched in both BB and PB ( 11 ). Consistently, it has been reported that ESBL-producing E. coli prevalent on chicken carcasses significantly compromises the efficacy of Campylobacter isolation ( 7 , 11 ), suggesting the need for improving the current isolation methods. However, ESBL genes in E. coli are generally encoded on plasmids harboring a cluster of antibiotic resistance genes ( 24 , 25 ).…”

“…ESBL-producing E. coli are highly prevalent in chicken and interrupts Campylobacter isolation using current protocols ( 7 , 8 ), leading to a revision of the international standard for the detection and enumeration of Campylobacter ( 9 , 10 ). Our previous analysis of microbiome changes during the enrichment of Campylobacter in selective media demonstrated that ESBL-producing E. coli are more abundantly enriched than Campylobacter ( 11 , 12 ). These studies suggested that inhibiting ESBL-producing E. coli at the enrichment step is the key to the success in Campylobacter isolation.…”