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Background In the era of increased antibiotic resistance and ever stricter control on antibiotic use, it is urgent to develop green, safe and non-residue alternatives to antibiotics applied to the poultry industry. To this end, we supplied the potential Lactobacillus Plantarum (L. Plantarum) fermented Astragalus in the diet of laying hens, with a final addition of 3‰. Its effects have been assessed on laying performance, egg quality, antioxidant and immunological status and intestinal microbiota, and are compared to the control group, to the Astragalus group containing 3‰ unfermented Astragalus, and to the L. Plantarum group containing 2% L. Plantarum (1 × 108 CFU/mL). Results During the second half of the experimental period (15 to 28 days), the egg production rate was significantly higher in the fermented Astragalus group than that in the other groups, with the fermented Astragalus group having the lowest feed conversion ratio. No significant difference (P > 0.05) was observed among treatments on egg quality. Fermented Astragalus-treated hens exhibited significantly increased catalase (CAT), glutathione peroxidase (GSH-Px), superoxide dismutase (SOD) and total antioxidant capacity (T-AOC) in serum, and reduced malondialdehyde (MDA) in serum. Furthermore, fermented Astragalus supplementation resulted in a significant increase in ileal microbiota abundance relative to control. Conclusions Feeding laying hens with L. Plantarum fermented Astragalus has beneficial effects on production, antioxidant potential, immunity and ileal microbiota. L. Plantarum fermented Astragalus is expected to be a novel feed additive used in poultry production.