The amphibian Xenopus laevis is to date the only species outside mammals where an MHC class I-like restricted innate-like T cell subset (iVa6-Ja1.43) reminiscent of iNKT cells has been identified and functionally characterized. Here, we report the identification of a distinct iT cell subset (Va45-Ja1.14) requiring a different MHC class I-like molecule (XNC4) for its development and function. We used two complementary reverse genetic approaches by transgenesis: RNAi to either impair either XNC4 or the Va45-Ja1.14 rearrangement and CRISPR-Cas9 to disrupt the Ja1.14 gene segment, to demonstrate that this innate-like immune surveillance system is critical for host resistance to mycobacteria infection. Both XNC4 deficiency that prevents iVa45-Ja1.14 T cell development, and direct disruption of the iVa45-Ja1.14 rearrangement dramatically impair tadpole immune response and resistance to M. marinum infection. In contrast, iVa45-Ja1.14 deficient tadpoles remain competent against ranavirus infection, unlike iVa6 iT cell deficient tadpoles that are highly susceptible to ranavirus infection. These data suggest that amphibians, which are evolutionary separated from mammals for more than 350 million years, have independently diversified a prominent and convergent MHC-like restricted innate-like T cell system.