The purpose of this study was to analyze the metallo-beta-lactamases (MBLs) genotype andoprDmutations of the beta-lactam antibiotic-resistantPseudomonas aeruginosa(PA) strains isolated from southern China. We collected 110 strains of beta-lactam antibiotic-resistant PA from 2 hospitals during January 2016-December 2017 from Dongguan, South China. MBLs were detected, amplified, and typed using EDTA disc synergy test, PCR, and Sanger gene sequencing. The mutations and expression levels ofoprDwere detected using Sanger gene sequencing and qPCR. A total of 16.36% (18/110) beta-lactam antibiotic-resistant PA strains produced MBLs, and the main genotypes of MBLs were IMP-25, VIM-2, and SIM-2. Sanger gene sequencing results showed that 107 of the 110 strains harbored mutations inoprDsequence, while 3 strains were negative foroprDamplification (2.73%). Among the 107 strains with positive amplification (97.27%), the rate of intentional mutations (including deletions, insertions, and premature stop codons) was 93.46% (100/107) and that of no disrupted mutation was 6.54% (7/107). qPCR analysis confirmed that the expression level of the OprD protein in the 7 strains of no disrupted mutation was significantly reduced. Among the beta-lactam antibiotic-resistant PA strains in southern China, 16.36% were positive for MBLs. The loss rate ofoprDwas 2.73%, and almost all PA strains showedoprDamplification variation or transcription downregulation. Thus, impairedoprDexpression and MBLs production may be some of the mechanisms of beta-lactam antibiotic-resistance of PA strains in southern China.