Gnotobiology: from 19th-century global foundations to 21st-century omics — six decades of Czech contribution to microbiome research

Gnotobiology, from the Greek gnotos (meaning 'known') and bios (meaning 'life'), is a research discipline that uses organisms with a defined microbiological status to study the interaction between hosts and microbes. This review traces six decades of Czech gnotobiology, beginning with the launch of a dedicated gnotobiology programme at Nový Hrádek in 1962 by Jaroslav Šterzl, whose visionary aims anticipated by decades the current recognition of the microbiota as a central determinant of immune and broader physiological function. The site - originally established in 1953 as the Biological Station - was thereby transformed into one of only four gnotobiological laboratories worldwide at that time and the first in Central and Eastern Europe. The facility pioneered the rearing of germ-free piglets, rats, rabbits, and mice, establishing the experimental foundation for the laboratory's work on immune ontogeny, mucosal immunity and tolerance, and microbiota-host interactions in immune development and regulation. This review discusses the key discoveries made using these models. Among them, work at the Institute of Microbiology (Prague and Nový Hrádek) demonstrated that germ-free animals have underdeveloped lymphoid tissue and impaired adaptive immunity. The review also describes the subsequent development of gnotobiotic models of human metabolic, immune-mediated, neoplastic, and neuropsychiatric diseases. The completion of the Human Genome Project in 2001 and the emergence of microbial metagenomics in the early 2000s sparked renewed interest in host-microbe interactions and led to a rediscovery of gnotobiotic approaches as essential tools for establishing causation in microbiome research. We examine how integrating these approaches with high-throughput sequencing, metabolomics, and other omics technologies has shifted the focus from cataloguing the microbiome to mechanistically dissecting host-microbe interactions. Finally, we outline future directions, including humanized gnotobiotic models, microbiota-based therapeutics, and the convergence of gnotobiology with personalized medicine and synthetic biology.