Growth of postembryonic larval intestine. The larval intestine continues as a straight tube for the 2 weeks following embryogenesis (A). The main change in appearance of the intestine between 6 and 19 dpf is doubling or tripling of the intestinal diameter. During the third week (20 to 26 dpf), a loop develops by first bending the intestine laterally and then rotating the bend dorsally (B). The loop continues to grow anteriorly and appears to draw in a portion of the posterior intestine into the loop (C). The intestine continues to grow in length during the fourth week (27 to 33 dpf) but not at the same rate as in the third week. One of the changes is a narrowing of the anterior intestinal diameter (D). Scale bar = 100 μm

Intestinal fold appearance. Intestinal folds viewed from the lumen side are defined at 19 dpf but appear shallow (A). Folds appear deeper by 26 dpf (B). Histological sections of 19 dpf folds (C) appear less developed than folds of 26 dpf intestines (D). Images from the lumen side and the histological images are from the anterior most portion of the mid region of the intestine as diagramed in the green lateral view of 19 dpf (E) and ventral view of 26 dpf (G); anterior is to the left). Cartoons in E to H also diagram regions recorded in Table 1. Anterior, pink; green, mid; red, posterior (E,G). The 19 dpf (F) and 26 dpf (H) diagram regions of folds recorded: black line represents the fold and dots represent nuclei. Green to green nuclei is width of base; green to blue (at tip of fold‐ top of diagram) is height of fold. Scale bar = 200 μm in A,B; 20 μm in C,D. Red line base of mesenchyme/limit of intestine; asterisk, lumen in C and D.

Position of proliferating cells between 6 and 19 dpf. Epithelial cells are labeled with a pulse of Edu (1 hr) at 7 dpf (A, blue arrow) and 14 dpf (D, blue arrow) followed by a chase to 19 dpf (red arrow). All labeled cells start in the interfold base (B, 7dpf and E, 14 dpf) and remain at this position following the chase period (C, 7 to 19 dpf and F, 14 to 19 dpf). Arrows indicate Edu‐labeled cells (B, C, E, and F); Scale bar = 10 μm; n = 5.

A–J: Epithelial cells migrate up the folds during the third and the fourth week postembryogenesis. Individuals are injected with Edu (green arrow) either at 20 (A) or 27 (E) dpf. Samples are taken at 1 hr to determine initial cell position (first blue arrow, A and E) and then at 2‐day intervals to the end of the week (three additional blue arrows, A and E). With a pulse of Edu at either 20 dpf (A) or 27 dpf (D) epithelial cells are labeled only at the fold base (arrows). Following a chase for 2 days to either 22 (C) or 29 (G) dpf, progeny reach the mid‐fold (arrows) and reach the tip by 4 days at either24 (B) or 31 (E) dpf. In each of the histological panels, the dashed line is drawn between the epithelial and mesenchymal layers. During these 2 weeks, mesenchymal cells are also labeled (examples at 24 dpf‐C and 31‐F; mesenchymal cells indicated with arrowheads); n = 4.

A–I: Development of apoptotic cells. In cross sections of the Tg(βactin:SecAV‐YFP) pu17 line, no apoptotic cells are observed during the third postembryonic week (example 23 dpf A to C). A small numbers of apoptotic cells are first observed at the tip of the fold at 33 dpf (arrow in E and merge with DAPI nuclear stain in F). Apoptotic cells become more prominent at the fold tip by 2 months postembryogenesis (arrows in H and merge with DAPI nuclear stain in I).

Secretory cells associate with clusters of proliferating intestinal epithelial cells. A pulse of Edu at 96 hpf followed by a chase to 120 hpf labels a group of proliferating cells (A) which is present at the fold base (B). Secretory cells (anti‐2F11) are interspersed within proliferating epithelial cells. Labeling with Edu on three different days during the first week postembryogenesis (C) (6–12 dpf) or the second week postembryogenesis (E) (13–19 dpf) labels clusters of proliferating epithelial cells. Secretory cells develop within groups of proliferating cells at the fold base (D) (6–12 dpf) and (F) (13–19 dpf) in a similar manner and number as observed at the end of embryogenesis. Some fold base epithelial cells begin differentiating into secretory cells (arrowhead A, C, E, and F) as they co‐label with 2F11 and Edu. Pan‐secretory 2F11, red; Edu, green; DAPI, blue; asterisk, intestinal lumen. Anterior is to the left and dorsal to the top in A, C, and E. Scale bar = 10 μm.

Secretory cells interdigitate between proliferating epithelial cells between 20 and 33 dpf. Labeling with Edu on three different days during the third week postembryogenesis labels multiple cells at the interfold base (A). Secretory cells now interdigitate between many of the proliferating epithelial cells. At 33 dpf, a single pulse of Edu labels multiple cells at the interfold base (B). Similar to 26 dpf, the interfold base now has many secretory cells and proliferating epithelial cells interdigitated. Numerous interdigitating secretory cells are observed between proliferating epithelial cells in the adult intestine following a single Edu pulse (C). The edge of the fold facing the lumen is outlined with the yellow dashed line. Red, pan‐secretory 2F11; green, Edu; blue, DAPI; white dashed line outer edge of intestine, asterisk, intestinal lumen. Scale bar = 10 μm.