Fecal samples were collected from patients admitted to The Eighth Affiliated Hospital of Sun Yat-sen University from April 2021 to December 2024, and a total of 131 cases tested positive for Blastocystis hominis infections using PCR assay and 50 cases tested negative were randomly divided into the experimental group (92 cases tested positive, 35 cases tested negative) and validation group (39 cases tested positive, 15 cases tested negative) using a random number table. Each fecal sample of experimental group was prepared into suspensions, and 50, 100, and 150 μl of suspensions were sampled for preparation of thin smears. Compared to conventional Wright-Giemsa staining, which involves an additional drying step, they dried at 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, and 80 ℃. The smears were stained with Wright-Giemsa stains for 5 to 10 minutes and then observed under an optical microscope. The diagnostic performance of the modified Wright-Giemsa staining method was evaluated using PCR amplification and genotyping results with B. hominis-specific primers BhRDr/RD5 as a reference standard. The validation group used both the modified Wright-Giemsa staining method and PCR method for simultaneous detection of B. hominis. The sensitivity, specificity, Youden index, predictive values (positive/negative), kappa coefficient, and concordance rate were as reference standards to evaluate the diagnostic performance of the modified method. The optimal fecal smear volume was 50 μl, and the most suitable drying temperature was (70 ± 5) ℃. PCR amplification produced a specific band at approximately 600 bp, with ST3 as the predominant genotype, followed by ST7, ST1, ST6, and ST2. The diagnostic sensitivity, specificity, kappa coefficient and concordance rate of the modified Wright-Giemsa staining method were 98.9% (95% CI: 94.0% to 100.0%), 97.1% (95% CI: 85.3% to 99.9%), 0.961 (95% CI: 0.906 to 1.000), and 98.4% (125/127) in the experimental group, showing high consistency with PCR assay. McNemar’s test was used to compare the differences in detection results between the two methods in the validation group, and the differences were not statistically significant (P > 0.05). The diagnostic sensitivity, specificity, positive predictive value, negative predictive value, Youden index, kappa coefficient and concordance rate of the modified Wright-Giemsa staining method were 97.4% (95% CI: 86.2% to 99.9%), 86.7% (95% CI: 59.5% to 98.3%), 95.0% (95% CI: 83.8% to 99.5%), 92.9% (95% CI: 69.8% to 99.8%), 0.841 (95% CI: 0.662 to 1.000), 0.858 (95% CI: 0.703 to 1.000), and 94.4% (51/54), respectively. The modified Wright-Giemsa staining method established in this study is simple to operate, yields stable results, and shows high consistency with PCR assay, which is suitable for rapid screening of B. hominis in grassroots medical institutions and field epidemiological investigations.