Novel Drug-like HsrA Inhibitors Exhibit Potent Narrow-Spectrum Antimicrobial Activities against Helicobacter pylori

<i>Helicobacter pylori</i> infection constitutes a silent pandemic of global concern. In the last decades, the alarming increase in multidrug resistance evolved by this pathogen has led to a marked drop in the eradication rates of traditional therapies worldwide. By using a high-throughput screening strategy, in combination with in vitro DNA binding assays and antibacterial activity testing, we identified a battery of novel drug-like HsrA inhibitors with MIC values ranging from 0.031 to 4 mg/L against several antibiotic-resistant strains of <i>H. pylori</i>, and minor effects against both Gram-negative and Gram-positive species of human microbiota. The most potent anti-<i>H. pylori</i> candidate demonstrated a high therapeutic index, an additive effect in combination with metronidazole and clarithromycin as well as a strong antimicrobial action against <i>Campylobacter jejuni</i>, another clinically relevant pathogen of phylum <i>Campylobacterota</i>. Transcriptomic analysis suggests that the in vivo inhibition of HsrA triggers lethal global disturbances in <i>H. pylori</i> physiology including the arrest of protein biosynthesis, malfunction of respiratory chain, detriment in ATP generation, and oxidative stress. The novel drug-like HsrA inhibitors described here constitute valuable candidates to a new family of narrow-spectrum antibiotics that allow overcoming the current resistome, protecting from dysbiosis, and increasing therapeutic options for novel personalized treatments against <i>H. pylori</i>.