Abstract: Objective To evaluate the applicability of multiple-locus variable-number tandem repeat analysis (MLVA) for molecular typing of Stenotrophomonas maltophilia. Methods Twelve tandem repeat loci in the total genome of S.maltophilia were analyzed with PCR and run in 2% standard agarose gel. Gels were visualized under UV light. Gel images were managed by using BioNumerics software package. Band size estimates were converted to a number of units within a character dataset with BioNumerics software. The copy numbers of twelve tandem repeat loci of 106 S.maltophilia strains was analyzed with BioNumerics (Version 4.0, Applied Maths BVBA, Belium). Results Totally 106 S. maltophilia isolates were genotyped with MLVA assay. Thirty-four MLVA genotypes were detected based on the similarity coefficient of 100%. The isolates from patients in a outbreak or from same region showed identical genotypes and could be divided into 11 groups based on the similarity coefficient of 45%. A number of MLVA clusters appeared to be partially limited to those from same geographic areas and sources. Conclusion MLVA assay is confirmed as a test which are rapid, simple, comparable, reproducible and with high discriminatory power. It can be applied in the research of molecular epidemiology of S. maltophilia.