High-Resolution Genomic Profiling of Campylobacter jejuni Across Human, Livestock, and Yellow-Legged Gull Reservoirs in Croatia Reveals Lineage Segregation and Shared Environmental Genotypes.

Campylobacter jejuni remains a leading cause of foodborne gastroenteritis worldwide, yet the epidemiological role of wild birds as reservoirs remains poorly understood. This study investigated the genetic diversity, transmission dynamics, and virulome architecture of 396 C. jejuni isolates collected in Croatia (2021-2025) from human, yellow-legged gull, broiler, bovine, and turkey sources. Whole-genome sequencing revealed high genomic diversity (SI>0.88), encompassing 124 sequence types across 25 clonal complexes. While classical MLST clustered 17.01% (n = 25/147, 95% CI: 11.83-23.82%) of gull isolates with human-associated lineages, high-resolution cg/wgMLST refined this overlap, segregating the majority of yellow-legged gull strains into a distinct, host-adapted ST-1275 clade (51.02%, n = 75/147). Furthermore, cg/wgMLST identified shared multi-host genotypes, such as CT6518 (ST-353) and CT3006 (ST-443), across wild gulls, domestic livestock, and clinical human cases. Virulome screening revealed a conserved pathogenic core alongside a lineage-specific periphery; the Guillain-Barré-syndrome-associated wlaN gene showed strong restriction to the dominant ST-21 generalist lineage, whereas the Type IV Secretion System was exceptionally rare (<1.5%). These findings demonstrate that, while most yellow-legged gull isolates constitute host-adapted lineages, gulls also harbor overlapping genotypes shared with livestock and clinical human isolates (6.80%, 95% CI: 3.69-12.18%), compatible with shared ecological niches and common contamination sources rather than direct bird-to-human transmission. This highlights their role as environmental sentinels and potential bridge reservoirs within One Health transmission interfaces. This study provides an indispensable baseline for integrated One Health biosecurity and surveillance strategies that bring together poultry production, ecosystem management, and public health infrastructure.