Genetic requirements and transcriptomics of Helicobacter pylori biofilm formation on abiotic and biotic surfaces

Skander Hathroubi,Shuai Hu,Karen M. Ottemann

doi:10.1038/s41522-020-00167-3

Abstract

Biofilm growth is a widespread mechanism that protects bacteria against harsh environments, antimicrobials, and immune responses. These types of conditions challenge chronic colonizers such as Helicobacter pylori but it is not fully understood how H. pylori biofilm growth is defined and its impact on H. pylori survival. To provide insights into H. pylori biofilm growth properties, we characterized biofilm formation on abiotic and biotic surfaces, identified genes required for biofilm formation, and defined the biofilm-associated gene expression of the laboratory model H. pylori strain G27. We report that H. pylori G27 forms biofilms with a high biomass and complex flagella-filled 3D structures on both plastic and gastric epithelial cells. Using a screen for biofilm-defective mutants and transcriptomics, we discovered that biofilm cells demonstrated lower transcripts for TCA cycle enzymes but higher ones for flagellar formation, two type four secretion systems, hydrogenase, and acetone metabolism. We confirmed that biofilm formation requires flagella, hydrogenase, and acetone metabolism on both abiotic and biotic surfaces. Altogether, these data suggest that H. pylori is capable of adjusting its phenotype when grown as biofilm, changing its metabolism, and re-shaping flagella, typically locomotion organelles, into adhesive structures.

Highlights

Helicobacter pylori is a significant human pathogen that is difficult to cure
H. pylori G27 forms biofilm on AGS gastric epithelial cells We examined biofilm formation on gastric epithelial cells as a course analysis suggested that biofilm mass was detectable at 24 h, and steadily increased to a maximum at 3 days of growth
We used noncentrifuged cells because there was not a significant difference between Fla+ and Fla- variants in early attachment, and we reasoned this approach would allow us to assess whether flagella were important for biotic surface biofilm formation. Both mutants colonized AGS cells at lower numbers than wild-type strains, and the flagellated Fla+ motB mutant exhibited a significantly higher colonization than the aflagellated Fla− fliA mutant (Fig. 4B). These results show that H. pylori employs flagella for adherence and biofilm formation on AGS cells, similar to what has been reported for abiotic surfaces

Summary

INTRODUCTION

Helicobacter pylori is a significant human pathogen that is difficult to cure. H. pylori infects more than half of the world’s population, making it one of the most common bacterial infections[1]. The standard of care is a 2-week course of a combination of a proton pump inhibitor and two-three antibiotics (clarithromycin, metronidazole, amoxicillin, or tetracycline), but this treatment leaves ~25% of people uncured[4,5] It is not yet fully understood why cure is so difficult, because H. pylori strains are not typically multidrug resistant. Recent studies have suggested that biofilm-grown H. pylori are tolerant to the commonly used antibiotics clarithromycin, amoxicillin, and metronidazole, and express high levels of putative antibiotic efflux pumps[20,21,22]. These findings suggest that biofilm formation could be a contributor to H. pylori persistence and the difficulty in curing this infection. We characterize H. pylori G27’s biofilm formation on abiotic and biotic surfaces, and perform genomic and transcriptomic approaches to unravel genes associated with the biofilm mode of growth in this strong biofilm-forming H. pylori

RESULTS

DISCUSSION

Findings

METHODS