Ontogeny of the organized nasopharynx-associated lymphoid tissue in rainbow trout

Understanding the ontogeny of teleost mucosa-associated lymphoid tissues (MALT) is critical for determining the earliest timepoint for effective mucosal vaccination of young fish. Here, we describe the developmental sequence that leads to the formation of an organized MALT structure in rainbow trout, the organized nasopharynx-associated lymphoid tissue (O-NALT). Control rainbow trout were sampled between 340 and 1860 degree days (DD) and routine histology and immunofluorescence staining were used to determine cellular changes in immune cells in the nasal cavity as well as O-NALT formation. We identified that O-NALT is first seeded by CD8α⁺ T cells at 700 DD followed by IgM⁺ B cells and cd4-2b⁺ cells at 1000 DD. Histomorphologically, trout O-NALT is fully formed at 1400 DD. Whole body gene expression analyses uncovered waves of igmh, cd4-2b, and cd8a expression that recapitulate the cellular seeding sequence of O-NALT by specific lymphocyte subsets. Our results indicate that 1) O-NALT formation results from a specific sequence of lymphocyte subset colonization pioneered by CD8α⁺ T cells and 2) the presence of the full O-NALT structure at 1400 DD may mark this timepoint as the earliest developmental stage at which mucosal vaccines can induce long lasting, specific immune responses.