Introduction:BuShen HuoXue (BSHX) decoction is commonly used in the clinical treatment of premature ovarian failure because it can increase estradiol level and decrease follicle-stimulating hormone level. In this study, we determined the potential therapeutic effects of BSHX decoctionviaanti-stress pathway and the underlying mechanism by using the nematodeCaenorhabditis elegansas an assay system.Methods:Bisphenol A (BPA, 175 μg/mL) was used to establish a fertility-defectiveC. elegansmodel. Nematodes were cultivated according to standard methods. Brood size, DTC, the number of apoptotic cells and oocytes were used to evaluate the fertility of nematodes. Nematodes were cultivated at 35°C as heat stress. RNA isolation and RT-qPCR were used to detect the mRNA expression level of genes. Intestinal ROS and intestinal permeability were used to evaluate the function of intestinal barrier. BSHX decoction was extracted with water and analyzed by LC/Q-TOF.Results and Discussion:In BPA-treated N2 nematodes, 62.5 mg/mL BSHX decoction significantly improved the brood size and the oocytes quality at different developmental stages. BSHX decoction improved resistance to heat stress through thehsf-1-mediated heat-shock signaling pathway. Further analysis showed that the decoction significantly improved the transcriptional levels ofhsf-1downstream target genes, such ashsp-16.1,hsp-16.2,hsp-16.41, andhsp-16.48. Other thanhsp-16.2expression in the gonad, the decoction also affected intestinalhsp-16.2expression and significantly reversed the adverse effects induced by BPA. Moreover, the decoction ameliorated intestinal ROS and permeability. Thus, BSHX decoction can improve fertility by increasing intestinal barrier function viahsp-16.2-mediated heat-shock signaling pathway inC. elegans. These findings reveal the underlying regulatory mechanisms ofhsp-16.2-mediated heat resistance against fertility defect.