Genetic and Transcriptomic Variations for Amoxicillin Resistance in Helicobacter pylori under Cryopreservation

Area of Science:

• Microbiology
• Molecular Biology
• Drug Resistance

Background:

• Amoxicillin resistance in Helicobacter pylori (H. pylori) is a growing concern for treating gastric infections.
• Clinical practices in China often involve freezing gastric specimens, which may affect H. pylori drug susceptibility testing.
• A decrease in amoxicillin resistance after freezing has been observed, potentially leading to inaccurate susceptibility results.

Purpose of the Study:

• To investigate the underlying reasons for the observed decrease in amoxicillin resistance in H. pylori following cryopreservation.
• To identify genetic and transcriptomic changes associated with altered amoxicillin resistance after freezing.

Main Methods:

• A high-level amoxicillin-resistant H. pylori clone (NX24r) was generated via drug pressure screening.
• Cryopreservation of the resistant clone at -80 °C for 3 months.
• Analysis of mutations in key resistance genes (PBP1, HefC) and whole-transcriptome sequencing before and after cryopreservation.

Main Results:

• Cryopreservation significantly reduced the minimum inhibitory concentration (MIC) of amoxicillin for the NX24r clone.
• Mutations in PBP1 and HefC, initially associated with high resistance, were not detected after cryopreservation.
• Transcriptome analysis revealed down-regulation of genes involved in plasma membrane biosynthesis and transport (e.g., lepB, secD) in the cryopreserved clone.

Conclusions:

• The decrease in amoxicillin resistance after cryopreservation is likely not due to loss of initial resistance-conferring mutations.
• Down-regulation of specific plasma membrane-related genes following freezing may contribute to the reduced amoxicillin resistance.
• Findings highlight the need to consider the impact of cryopreservation on H. pylori drug susceptibility testing.