Objective Nephrolithiasis is a common urological disease. This study aims to evaluate the preventive and therapeutic effects of hydro-alcoholic extract of Aerva lanata (L.) roots (HAEAL) on ethylene glycol-induced nephrolithiasis in rats. Methods Fifty grams of shade-dried coarsely powdered Aerva lanata (L.) root was successively extracted with organic solvents in increasing order of polarity [petroleum ether (60 −80 °C), chloroform, and ethanol] using a Soxhlet apparatus, and then concentrated. Physical tests including nature, color, odor, and texture were performed on the herbal suspension. In vitro nephrolithiasis assessment was performed by nucleation assay, aggregation assay, and crystal growth assay. Thirty adult male Wistar albino rats were randomly divided into five groups (six rats in each group). Group 1: negative control group without induction or treatment till day 28. Group 2: positive control group receiving a daily oral solution of 0.75% ethylene glycol till day 14, and mixed with distilled water till day 28. Group 3: standard group receiving a daily oral solution of 0.75% ethylene glycol till day 14 and Cystone (750 mg/kg) from day 15 to day 28. Group 4: low dose HAEAL group receiving a daily oral solution of 0.75% ethylene glycol till day 14, and 400 mg/kg HAEAL from day 15 to day 28 (1 mL per day). Group 5: high dose HAEAL group receiving a daily oral solution of 0.75% ethylene glycol till day 14, and 800 mg/kg HAEAL from day 15 to day 28 (1 mL per day). Urine (urine volume, pH value, appearance, odor, and turbidity) examination and serum test were performed. On day 29, the kidneys were dissected, and histopathology examination was performed to determine the degree of tubular injury. Results The suspension showed stability and aroma with no turbidity at room temperature. The suspension did not show changes in color and odor until day 3, indicating that the preparation was stable for 72 h. Body weight decreased in the positive control group indicating stone formation and changes in water intake. Both standard and HAEAL treatments restored the body weight to normal levels after treatment, indicating the beneficial effects of the treatment. Histopathological examination revealed no significant findings in the negative control group, whereas the positive control group showed inflammation in the kidney parenchyma. Compared with positive control group, there was increase in urine volume and excretion of urinary constituents such as calcium and oxalate (P < 0.01) as well as improved clearance rate (P < 0.05) in HAEAL treatment groups, in addition, the urine pH value of HAEAL groups was increased. Conclusion HAEAL reduced nephrolithiasis formation and had a diuretic effect, which could be used to promote the expulsion of stones. Further studies are needed to enhance the stability of the suspension for the production of better pharmaceutical formulations.