Whole-Genome Sequencing and Comparative Genomic Analysis of Citrobacter farmeri and Enterobacter cloacae from Unhatched Green Turtle Eggs

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Bacteria associated with unhatched sea turtle eggs remain poorly characterized at the genomic level. This study provides genome-scale characterization of bacterial isolates recovered from unhatched green sea turtle (Chelonia mydas) eggs at Akyatan Beach—a critical nesting site in the Eastern Mediterranean. Sampling 30 nests during the nesting season, we isolated bacteria from infected eggshells and dead embryos. Following Matrix-Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry(MALDI-TOF MS) identification and 16S rRNA validation, we performed whole-genome sequencing (WGS) on Citrobacter farmeri and Enterobacter cloacae, two opportunistic pathogens of significant clinical and ecological concern. High-quality draft genomes revealed remarkable metabolic versatility, particularly within carbon and nitrogen pathways. Most notably, we identified extensive resistomes including resistance to β-lactams, fluoroquinolones, and aminoglycosides, alongside virulence factors for adhesion and iron acquisition. ANI analysis confirmed high genomic similarity to clinical reference strains, comparative genomic analysis revealed a substantial accessory gene pool, suggesting potential genomic flexibility between the two isolates. These findings provide the first genome-scale insight into these pathogens in C. mydas nests, and highlighting their genomic potential for opportunistic pathogenicity. Our results advocate for integrating genomic microbial surveillance into nesting beach management through a ‘One Health’ lens.