Infectious diseases, such as lactococcosis caused by Lactococcus garvieae, are portrayed as critical limiting factors in aquaculture. The antimicrobial properties of Lactic Acid Bacteria (LAB), mainly the production of organic acids and bacteriocins (e. g., the lanthionine containing nisins A and Z; NisA and NisZ, respectively), led to propose LAB as probiotics to be used as an alternative and/or complementary strategy to vaccination and chemotherapy in aquaculture. L. lactis RBT18, isolated from cultured rainbow trout, exerts a strong direct and extracellular antimicrobial activity against L. garvieae and other ichthyopathogens, being the latter heat-resistant (100ºC, 10 min), and thus suggesting the involvement of a thermostable antimicrobial compound (i. e., bacteriocin). Cross-immunity tests using the agar-well-diffusion test (ADT) and PCR assays suggested that NisA/Z is the bacteriocin responsible for the extracellular antimicrobial activity exerted by L. lactis RBT18. To demonstrate this hypothesis, the bacteriocin was purified to homogeneity by two multi-chromatographic procedures. MALDI TOF-MS analyses of purified samples after the last reverse-phase chromatography step identified the presence of NisZ (3,330 Da), and its oxidized form (3,346 Da), derived from the oxidation of a lanthionine ring. The oxidized NisZ showed a diminished antimicrobial activity which would increase the chances of bacterial pathogens to evade its antimicrobial activity. Further experiments are necessary to assess the in vitro and in vivo safety and efficiency of L. lactis RBT18 as probiotic in aquaculture, but also to optimize the environmental conditions to reduce bacteriocin oxidation and thus bacterial pathogen resistance.