Objectives To determine the effect of sea salt on the resistance ofTrichoderma harzianumLZDX-32-08 to hygromycin B and speculate the possible mechanisms involved via transcriptome analysis. Results Sea salt addition in media to simulate marine environment significantly increased the tolerance of marine-derived fungusTrichoderma harzianumLZDX-32-08 to hygromycin B from 40 to 500 mu g/ml. Meanwhile, sea salt addition also elicited the hygromycin B resistance of 5 other marine or terrestrial fungi. Transcriptomic analyses ofT. harzianumcultivated on PDA, PDA supplemented with sea salt and PDA with both sea salt and hygromycin B revealed that genes coding for P-type ATPases, multidrug resistance related transporters and acetyltransferases were up-regulated, while genes coding for Ca2+/H(+)antiporter and 1,3-glucosidase were down-regulated, indicating probable increased efflux and inactivation of hygromycin B as well as enhanced biofilm formation, which could jointly contribute to the drug resistance. Conclusions Marine environment or high ion concentration in the environment could be an importance inducer for antifungal resistance. Possible mechanisms and related key genes were proposed for understanding the molecular basis and overcoming this resistance.