From onset to healing: temporal dynamics of microbial communities pinpoint Midichloria-like organism's key role in the development of the fish skin disease red mark syndrome.

BACKGROUND: Red mark syndrome (RMS) is an infectious disease affecting rainbow trout (Oncorhynchus mykiss), especially at market size, forcing farmers to downgrade the product with heavy economic repercussions. The causative agent of RMS has not been established according to Koch's postulates, since possible candidates have not been isolated and propagated in vitro. While the 16S rRNA gene of a Midichloria-like organism (MLO) is consistently detected in active skin lesions, the role of other bacteria in the disease has not been excluded. In this work, we provide a temporal perspective to elucidate the relationships between the bacteriome in rainbow trout skin and water, during the development and resolution of clinical disease in naive fish infected by cohabitation with RMS-affected fish in the same tank. RESULTS: We quantified the MLO by qPCR in skin and, for the first time, in water using environmental DNA, showing that its quantity in both sample types corresponds with disease progression. Using 16S rRNA gene profiling, we provide further evidence that the MLO is likely the primary pathogen triggering RMS, as it was the only significantly enriched taxon in active lesions. Furthermore, the skin microbiome of affected fish reverted to a control-like state during healing. RMS also caused changes in the overall skin microbiome at peak pathology: the relative abundance of "Ca. Branchiomonas" and an unclassified gammaproteobacterium increased in apparently healthy skin areas of RMS-affected fish while remaining low in controls, suggesting they may be opportunistic bacteria implicated in skin dysbiosis. CONCLUSIONS: Our findings on the temporal dynamics of the skin microbiome during disease progression and recovery strengthen the evidence that Midichloria-like organism is the primary pathogen of RMS. The observed shifts in other bacterial taxa suggest possible secondary roles in disease-associated skin dysbiosis. Finally, detection of MLO in environmental DNA from water provides new insights into RMS transmission and potential monitoring strategies.