Abstract
Abstract This project exploits SAMMY, a cutting-edge technology enabling the sequential isolation of chromatin fractions based on DNA accessibility, to explore bacterial dormancy in the major human pathogen Helicobacter pylori (HP). Dormant coccoid forms of HP contribute to persistence, antibiotic treatment failure, and constitute a major threat to public health worldwide. SAMMY will be applied for the first time to a bacterial system to generate an OMICS-level map of nucleoid conformations under various growth and stress conditions. We will combine whole transcriptome analysis and investigate the role of crucial transcription factors of HP in chromatin organization and coccoid formation. This innovative approach will uncover key structural and regulatory features of the HP nucleoid, providing new insights into dormancy mechanisms and persistence, and paving the way for novel antimicrobial strategies.
Results achieved
The NORA project successfully achieved its main objective of adapting and validating the SAMMY-seq technology for the study of bacterial chromatin organization in Helicobacter pylori, providing the first reproducible framework for investigating nucleoid accessibility and structural dynamics in bacterial cells. The project established optimized experimental and bioinformatic workflows that can be readily transferred to other bacterial species, significantly expanding the applicability of this innovative sequencing-based approach. Using SAMMY-seq, the project generated the first genome-wide characterization of nucleoid accessibility across different physiological states of H. pylori, including exponential growth, stationary phase, coccoid forms, and biofilms. These analyses revealed extensive chromatin reorganization during the transition to dormancy and identified genomic regions whose accessibility changes are associated with stress adaptation, genome plasticity, and pathogenicity-related functions. A major outcome of the project was the production of the first comprehensive transcriptomic profile of H. pylori coccoid cells. Integration of transcriptomic and nucleoid accessibility datasets demonstrated that dormant coccoid cells retain a substantial and structured RNA repertoire rather than representing merely degraded bacterial remnants. This finding provides new insights into the biology of bacterial persistence and supports the concept that coccoid cells remain physiologically organized and potentially capable of resuscitation. The project further identified and characterized novel molecular determinants potentially involved in dormancy regulation. In particular, transcriptomic and functional analyses highlighted the IsoA4/AapA4 toxin–antitoxin system as a candidate regulator of morphological transition and growth control. In parallel, significant progress was achieved in the development of conditional genetic tools for studying the essential transcriptional regulator HP1043, strengthening evidence for its role in maintaining the vegetative state and regulating genes associated with the transition toward dormancy. Beyond the biological discoveries, NORA delivered important resources for the scientific community. The SAMMY-seq analysis workflow was adapted to bacterial genomes, implemented as an open-source Nextflow pipeline, and publicly released through the nf-core framework, enabling reproducible and scalable analyses of nucleoid accessibility data. Project results were disseminated through national and international conferences, contributed to doctoral training activities, and laid the foundations for future studies aimed at comparing chromatin organization and dormancy mechanisms across different bacterial species. Overall, the project demonstrated the feasibility of applying SAMMY-seq to bacteria, generated unprecedented datasets on H. pylori dormancy, identified new candidate regulators of the coccoid transition, and established methodological and computational tools that will support future research on bacterial persistence and antimicrobial target discovery.Dettagli del progetto
Responsabile scientifico: Davide Roncarati
Strutture Unibo coinvolte:
Dipartimento di Farmacia e Biotecnologie
Coordinatore:
CNR - Consiglio Nazionale delle Ricerche(Italy)
Contributo totale Unibo: Euro (EUR) 92.179,00
Durata del progetto in mesi: 24
Data di inizio
28/09/2023
Data di fine:
28/02/2026
