Two in vivo and in vitro studies were conducted to investigate the effects of supplementation of a natural polyherbal mixture (PHM) manufactured from selected herbs in broiler chickens challenged with coccidiosis. For the in vitro trial, E. tenella and E. maxima sporozoites were used to test how PHM affected sporozoites viability at 24, 48, and 72 h. The treatments were as follows: negative control with phosphate buffered saline (NC-PBS), solvent control containing 1 % dimethyl sulfoxide (DMSO), salinomycin at 12 mg/kg with 1 % DMSO in PBS (SAL), and 500 mg/kg PHM in PBS (PHM). For the in vivo trial, a total of 288 0-day-old male Cobb 500 were randomly distributed into 3 treatments with 8 replicates, and study lasted for 28 days. Treatments were as follows: non-challenge control with a basal diet (NC), Eimeria spp. challenge with a basal diet (CC), and Eimeria spp. challenge with a basal diet containing 500 mg/kg of the PHM (PHM). Chickens in challenged groups were inoculated with 62,500 oocyst of E. acervulina, 12,500 oocyst of E. maxima and E. tenella on 14 days. In vitro results showed that PHM increased (P < 0.001) a Eimeria sporozoite reduction percentage. As for in vivo results, the PHM group had similar body weight gain, feed intake, and feed efficiency compared to the NC group. The use of PHM reduced fecal oocyst counts of E. tenella and E. maxima from 6 to 9 days post inoculation (DPI; P < 0.05). Moreover, PHM supplementation decreased duodenum and ceca lesion scores (P < 0.001). The PHM group also had reduced expression levels of claudin 1 (CLDN-1), interleukin 1 beta (IL-1β) and tumor necrosis factor alpha (TNF-α) in the jejunum compared to the CC group (P < 0.01). In conclusion, the supplementation with 500 mg/kg of polyherbal mixture both in vitro and in vivo reduced the viability of E. tenella and E. maxima sporozoites, and this could explain that PHM effectively mitigated negative effects caused by the challenge with Eimeria spp., suggesting that it could be a dietary strategy to improve performance and gut health in broilers under coccidiosis.

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