A Tessellated Lymphoid Network Provides Whole-Body Antigen Surveillance in Zebrafish

T cells survey the host for cognate antigen in mammals by trafficking to and searching within lymph nodes distributed throughout the body. Non-mammalian jawed vertebrates lack lymph nodes but still maintain highly diverse repertoires of T cells. How these T cells find antigen in hosts devoid of lymph nodes remains wholly unclear. In this study, we exploit the in vivo imaging amenability of the zebrafish to investigate T cell organization, trafficking, and antigen surveillance in an animal completely lacking lymph nodes. We find that T cells organize into a previously undescribed whole-body scale-associated pattern that we have named the tessellated lymphoid network (TLN). We find that the TLN harbors significantly more T cells than the gills, gut, kidney, and spleen, as well as populations of antigen-presenting cells, suggesting a central role in adaptive immune responses in fish. Within the TLN, T cells perform a highly directional ballistic streaming mode of motility, allowing them to traffic through the host in a coordinated ventral-to-dorsal loop. Local infection results in a dramatic shift in T cell motility, from rapid streaming to a slower random walk in regions proximal to the infection. Here, T cells sequentially interact with and ultimately form stable contacts with antigen-presenting cells. Finally, T cells within the TLN of infected fish show transcriptional changes consistent with TCR signaling and subsequent T cell activation. The TLN thus provides a mechanism for T cells to both traffic through the host and, in the context of an infection, effectively scan for and detect antigen, mirroring the function of lymph nodes in mammals. Adult zebrafish either without infection or infected by L. monocytogenes through puncture wounding were euthanized and descaled to collect associated cells at 4 days post-infection. Libraries were constructed from these cells using the Chromium Single Cell Gene Expression Solution 3’ v2 (10x Genomics) and sequenced using NovaSeq6000 (Illumina) with read lengths of 29-bp + 90-bp (Read1 + Read2).