Increased antibiotic free production has increased emphasis on poultry gut health leading to evaluation of dietary alternatives such as nucleotides to improve production. Therefore, yeast derived nucleotides were evaluated for effects on growth performance, gastrointestinal ecology and immune system short- (trial 1) and long-term (trial 2) experiments. Nucleotides improved growth performance in trial 1 but not in trial 2. Nucleotides improved cationic transporter 1 expression and jejunal histomorphology in Eimeria challenged birds in trial 1 but negatively affected non-challenged birds in trial 1 and no effect in trial 2. Nucleotides increased bursal weights in trial 2, but not the spleen and thymus. Nucleotides effect on gastrointestinal microbiota community was independent of Eimeria, but interactive on ceca short chain fatty acid concentration and pH. Overall, nucleotides attenuated some negative effects of Eimeria but also resulted in negative effects. Supplementation of nucleotides may be most beneficial for animals encountering enteric stress conditions.