Adaptation of an in-house PCR for the detection of Helicobacter pylori and the mutations associated with macrolide resistance into ready-to-use PCR microwell strips.

BACKGROUND AND OBJECTIVES: The present study describes the successful adaptation of an in-house Polymerase Chain Reaction (PCR) for Helicobacter pylori detection coupled with the main mutations associated with resistance to clarithromycin in ready-to-use PCR microwell strips. MATERIALS AND METHODS: These microwell strips can be used on LightCycler® 480, and are delivered with nine microliters of the reaction mixture dispensed into 8-well microwell strips. An extraction control PCR targeting the ß-globin household gene is amplified in the same run as H pylori detection. RESULTS AND CONCLUSION: These microwell strips can be stored at -20°C for 1 year and left at room temperature and in the light for up to 4 h with no impact on the PCR results. Microwell strips can also undergo a thaw and refreeze cycle without impacting the PCR results. These PCR microwell strips are available for purchase from Eurogentec.

Emergence of Erythromycin Resistance Methyltransferases in Campylobacter coli Strains in France.

Antimicrobial resistance in campylobacters has been described worldwide. The emergence of multiresistant isolates, particularly among Campylobacter coli isolates, is concerning. New resistance mechanisms appear frequently, and DNA-sequence-based methods such as whole-genome sequencing (WGS) have become useful tools to monitor their emergence. The genomes of 51 multiresistant French Campylobacter sp. clinical strains from 2018 to 2019 were analyzed to identify associated resistance mechanisms. Analyses of erythromycin-resistant strains revealed 23S rRNA mutations among most of them and two different methyltransferases in 4 strains: Erm(B) and a novel methyltransferase, named Erm(N) here. The erm(B) gene was found in multidrug-resistant genomic islands, whereas erm(N) was inserted within CRISPR arrays of the CRISPR-cas9 operon. Moreover, using PCR screening in erythromycin-resistant strains from our collection, we show that erm(N) was already present in 3 French clinical strains 2 years before its first report in 2018 in Quebec, Canada. Bacterial transformations confirmed that the insertion of erm(N) into a CRISPR-cas9 operon can confer macrolide resistance. Campylobacter species are easily able to adapt to their environment and acquire new resistance mechanisms, and the emergence of methyltransferases in campylobacters in France is a matter of concern in the coming years.