The combined effects of probiotics (Lactobacillus acidophilus, Lactobacillus casei, Streptococcus faecium, and Saccharomyces cerevisiae) and organic acids (sorbic and citric acid) on intestinal morphology and expression of immunerelated genes were investigated. One-day-old chicks were randomly assigned to 1 of 3 groups: birds not receiving probiotic or organic acids (control; T1), or birds receiving an oral combination (1 g/L in water) of 108 CFU/g of each of the aforementioned probiotics and organic acids (1% sorbic acid and 0.2% citric acid) for 7 (T2) or 14 d (T3). Each group was divided into 5 replicate pens of 20 birds each, and 5 birds from each group (1 from each pen) were killed on d 11 and 22. Intestinal sections were collected for histological assessment, and reverse transcriptase-PCR analysis was used to assess defensin and cathelicidins expression. Quantitative real-time PCR was used to assess toll-like receptors (TLR) and cytokine expression. Duodenal villus height was greater in T2 and T3 at d 11 (P ≤ 0.036) and 22 (P ≤ 0.015) compared with T1. At d 11, duodenal goblet cell/unit area was less in T3, whereas it was greater in T2 compared with T1 in the jejunum (P = 0.009). Ileal goblet cell/unit area was greater in T3 at d 22 compared with T1 (P < 0.001). Avian beta-defensin-3 was expressed in all tissues except the bursa of T3 birds at d 11, and TLR-2 was down regulated in the cecal tonsil of birds in T2 and T3 at d 11 compared with T1 (P = 0.020 and 0.003, respectively). Expression of IL-12p35 in the ileum at d 11 was down regulated in T2 and T3 compared with T1 (P = 0.030 and 0.012, respectively). Reduced expression of INF-γ was observed in the ileum in T3 compared with T1 at d 11 (P = 0.047). Ileal IL-6 and IL-10 and cecal tonsil interferon-gamma (INF-γ) expressions were greater T2 at d 22 (P ≤ 0.047) than T1. In conclusion, supplementation of combined probiotics and organic acids resulted in inconsistent gut morphology associated responses, and avian beta-defensins and cathelicidins expression were not associated with combined probiotics and organic acids supplementation. Birds supplemented with combined probiotics and organic acids for 7 d showing similar responses in TLR-2, IL-12p35, and IFN-γ compared with those supplemented for 14 d indicates that shorter periods of supplementation might be enough to elicit beneficial responses. © 2012 American Society of Animal Science. All rights reserved.