46 Nutritional Programming (NP) has been studied as a means of mitigating the negative effects of 47 dietary plant protein (PP), but the optimal timing and mechanism behind NP are still unknown.48 The objectives of this study were: 1) To determine whether zebrafish (Danio rerio) can be 49 programmed to soybean meal (SBM) through early feeding and broodstock exposure to 50 improve SBM utilization; 2) To determine if NP in zebrafish affects expression of genes 51 associated with intestinal nutrient uptake; 3) To determine if early stage NP and/or broodstock 52 affects gene expression associated with intestinal inflammation or any morphological changes 53 in the intestinal tract that might improve dietary SBM utilization. Two broodstocks were used to 54 form the six experimental groups. One broodstock group received fishmeal (FM) diet (FMBS), 55 while the other was fed ("programmed with") SBM diet (PPBS). The first ((+) Control) and the 56 second group ((-) Control) received FM and SBM diet for the entire study, respectively, and 57 were progeny of FMBS. The last four groups consisted of a non-programmed (FMBS-X-PP and 58 PPBS-X-PP) and a programmed group (FMBS-NP-PP and PPBS-NP-PP) from each of the 59 broodstocks. The programming occurred through feeding with SBM diet during 13-23 dph. The 60 non-control groups underwent a PP-Challenge, receiving SBM diet during 36-60 dph. During the 61 PP-Challenge, both PPBS groups experienced significantly lower weight gains than the (+) 62 Control group. NP in early life stages significantly increased the expression of PepT1 in PPBS-NP-63 PP, compared to PPBS-X-PP. NP also tended to increase the expression of fabp2 in the 64 programmed vs. non-programmed groups of both broodstocks. The highest distal villus length-65 to-width ratio was observed in the dual-programmed group, suggesting an increase in surface 3 66 area for nutrient absorption within the intestine. The results of this study suggest that NP 67 during early life stages may increase intestinal absorption of nutrients from PP-based feeds.68 Introduction
69Plant protein-(PP) based diets in aquaculture are advancing but, are not yet fully 70 suitable replacements for fishmeal (FM) for many species due to their adverse effects on 71 growth and health. Soybean meal (SBM) is the most commonly used protein source in plant-72 based diets but, has been shown to reduce fish growth at high dietary inclusion levels (above 73 25%) and to cause inflammation in the intestines of many fish species due to presence of anti-74 nutritional factors [1]. The reduced growth performance of fish fed SBM-based diets has been 75 studied in both carnivorous and omnivorous species [2-4]. For example, rainbow trout 76 (Oncorhynchus mykiss) fed a PP-based diet showed a 17% decrease in growth and a significant 77 reduction in feeding efficiency [2]. Wang et al. [4] concluded that, in orange-spotted grouper 78 (Epinephelus coioides), increasing levels of dietary soybean meal led to reduced growth 79 performance and increased intestinal damage. Simil...