The western mosquitofish (Gambusia affinis (G. affinis)) is often the only fish that survives in various selenium-contaminated aquatic environments. However, mechanisms of its survival in such environments remain seldom explored. In this study, the acute toxicity of selenomethionine (Se-Met) in a variety of species (four oviparous and four ovoviviparous species) was evaluated, followed by the chronic effects of Se-Met on G. affinis and on zebrafish (Danio rerio (D. rerio)), focusing on Se accumulation in the gonads and offspring, reproductive biology, and offspring development. Our results showed that the LC50-96h of Se-Met was the largest in G. affinis among the eight fish, confirming its elevated resistance to Se. Meanwhile, the transcripts of genes related to selenoproteins and sulfoproteins showed opposite responses between G. affinis and D. rerio to Se-Met exposure, along with the altered levels of total selenoproteins in oviparous but not in ovoviviparous fish. The chronic exposures showed that Se levels in the gonads, effects on gonadal development and embryonic and early larval development, and Se efflux in the embryos and larvae were apparently different between G. affinis and D. rerio. Finally, differentially expressed genes (particularly antioxidant and inflammation) were mostly stimulated in G. affinis but inhibited in D. rerio. This study has demonstrated that the Se efflux capacity of the offspring and genes related to Se metabolism and antioxidant physiology can help partially explain the survival of G. affinis in Se-contaminated ecosystems.