The Toll-like receptor 4 (TLR4) pathway involves in the pathogen recognition and defense against infection in mammals. Considering that avian and mammalian TLR are differentially mediated, the action of a natural product on avian TLR4 pathway was unclear. High, medium and low doses of Astragalus polysaccharide (APS), were treated the chicken at 7-days-old age by gavage. The sIgA level in the intestinal fluid, the expression of chTLR4 mRNA/protein in bursa of Fabricius as well as the expression of downstream molecules of chTLR4 (chMyD88, chTRIF, chNF-κB, chIRF3, chIFN-β and chTNF-α) were measured on alternate days. The content of sIgA and the chTLR4 mRNA expression/protein level were increased in non-dose-dependent manner after APS supplement. Also, the expressions of a subset of MyD88-independent pathway genes were more than MyD88-independent, in particular with low doses of APS supplement for 7 days. These suggest that administration of APS activates chTLR4 pathway in bursa of Fabricius in MyD88-independent pathway. Meanwhile, low dose of APS shows better performance regarding the activation of chTLR4 and regulation of MyD88-independent pathway.