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Extensive use of antibiotics has been the primary treatment for the Salmonid Rickettsial Septicemia, a salmonid disease caused by the bacterium  Piscirickettsia salmonis . Occurrence of antibiotic resistance has been explored in various  P. salmonis  isolates using different assays; however,  P. salmonis  is a nutritionally demanding intracellular facultative pathogen; thus, assessing its antibiotic susceptibility with standardized and validated protocols is essential. In this work, we studied the pathogen response to antibiotics using a genomic, a transcriptomic, and a phenotypic approach. A new defined medium (CMMAB) was developed based on a metabolic model of  P. salmonis . CMMAB was formulated to increase bacterial growth in nutrient-limited conditions and to be suitable for performing antibiotic susceptibility tests. Antibiotic resistance was evaluated based on a comprehensive search of antibiotic resistance genes (ARGs) from  P. salmonis  genomes. Minimum inhibitory concentration assays were conducted to test the pathogen susceptibility to antibiotics from drug categories with predicted ARGs. In all tested  P. salmonis  strains, resistance to erythromycin, ampicillin, penicillin G, streptomycin, spectinomycin, polymyxin B, ceftazidime, and trimethoprim was medium-dependent, showing resistance to higher antibiotic concentrations in the CMMAB medium. The mechanism for antibiotic resistance to ampicillin in the defined medium was further explored and was proven to be associated to a decrease in the bacterial central metabolism, including the TCA cycle, the pentose-phosphate pathway, energy production, and nucleotide metabolism, and it was not associated with decreased growth rate of the bacterium or with the expression of any predicted ARG. Our results suggest that nutrient scarcity plays a role in the bacterial antibiotic resistance, protecting against the detrimental effects of antibiotics, and thus, we propose that  P. salmonis  exhibits a metabolic resistance to ampicillin when growing in a nutrient-limited medium.

Nutrient Scarcity in a New Defined Medium Reveals Metabolic Resistance to Antibiotics in the Fish Pathogen Piscirickettsia salmonis