Medicinal plants and probiotics affect the productive performance of avian species through comparable biological mechanisms, including modifications to the intestinal microbiome, alterations in histomorphology and effects on the immune system and blood metabolites. In light of this, the current study was designed and conducted to compare the efficacy of Zataria multiflora, Salvia officinalis and probiotic microorganisms on some physiological parameters and the performance of laying hens. A total of 225 white leghorn hens aged 42 weeks were randomly assigned into 9 treatments with 5 replicates of 10 birds each. Experimental treatments consisted of a control group, 0.2% and 0.4% Z. multiflora, 0.2% and 0.4% S. officinalis, 0.2% and 0.4% the combination of Z. multiflora and S. officinalis and 0.005% and 0.01% probiotic microorganisms. Our findings showed that dietary inclusion of 0.01% probiotic, 0.4% Z. multiflora and 0.4% Z. multiflora + S. officinalis treatments decreased the intestinal Escherichia coli (p < 0.01) and Salmonella (p = 0.06) enumerations and increased the lamina propria lymphoid follicle diameter (p < 0.01). Feeding 0.4% Z. multiflora, 0.4% Z. multiflora + S. officinalis could enhance the villus height and crypt depth (p < 0.05). Probiotic supplementation decreased crypt depth (p = 0.019) and increased villus height to crypt depth ratio (p < 0.05). Dietary supplementation of 0.4% Z. multiflora and 0.01% probiotic treatments significantly showed the highest antibody titre against the Newcastle disease virus vaccine and the lowest heterophil-to-lymphocyte ratio (p < 0.05). Over the whole trial period, feeding 0.01% probiotic and 0.4% Z. multiflora treatments could improve the egg production percentage and feed conversion ratio (p < 0.05). In conclusion, the present findings indicate that the dietary administration of 0.4% Z. multiflora and 0.01% probiotic treatments could improve the productive performance of laying hens by strengthening both mucosal and systemic immune functions, as well as improving health indicators related to the intestinal absorptive area.
