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Antimicrobial Agents and Chemotherapy, January 2006, p. 204-209, Vol. 50, No. 1
0066-4804/06/$08.00+0     doi:10.1128/AAC.50.1.204-209.2006
Copyright © 2006, American Society for Microbiology. All Rights Reserved.

Simultaneous Detection of Nine Antibiotic Resistance-Related Genes in Streptococcus agalactiae Using Multiplex PCR and Reverse Line Blot Hybridization Assay

Xianyu Zeng,^1,2 Fanrong Kong,¹ Hui Wang,^1,2 Archie Darbar,^1, and Gwendolyn L. Gilbert^1*

Centre for Infectious Diseases and Microbiology, Institute of Clinical Pathology and Medical Research, Westmead, New South Wales, Australia,¹ Department of Dermatology, Wuhan First Hospital, Wuhan, Hubei Province, People's Republic of China²

Received 24 July 2005/ Returned for modification 28 August 2005/ Accepted 24 October 2005

Streptococcus agalactiae (group B streptococcus [GBS]) is the leading cause of neonatal and maternal sepsis. Penicillin is recommended for intrapartum prophylaxis, but erythromycin or clindamycin is used for penicillin-allergic carriers. Antibiotic resistance (AR) has increased recently and needs to be monitored. We have developed a multiplex PCR-based reverse line blot (mPCR/RLB) hybridization assay to detect, simultaneously, seven genes encoding AR—erm(A/TR), erm(B), mef(A/E), tet(M), tet(O), aphA-3, and aad-6—and two AR-related genes, int-Tn and mreA. We tested 512 GBS isolates from Asia and Australasia and compared mPCR/RLB with antibiotic susceptibility phenotype or single-gene PCR. Phenotypic resistance to tetracycline was identified in 450 (88%) isolates, of which 442 had tet(M) (93%) and/or tet(O) (6%). Of 67 (13%) erythromycin-resistant isolates, 18 were susceptible to clindamycin, i.e., had the M phenotype, encoded by mef(A/E); 39 had constitutive (cMLS_B) and 10 inducible clindamycin resistance, and of these, 34 contained erm(B) and 12 erm(A/TR). Of four additional isolates with mef(A/E), three contained erm(B) with cMLS_B and one was erythromycin susceptible. Of 61 (12%) clindamycin-resistant isolates, 20 were susceptible to erythromycin and two had intermediate resistance. Based on sequencing, 21 of 22 isolates with mef had mef(E), and 8 of 353 with int-Tn had an atypical sequence. Several AR genes, erm(B), tet(O), aphA-3, aad-6, and mef(A/E), were significantly more common among Asian than Australasian isolates, and there were significant differences in distribution of AR genes between GBS serotypes. Our mPCR/RLB assay is simple, rapid, and suitable for surveillance of antibiotic resistance in GBS.

Supplemental material for this article may be found at http://aac.asm.org/.

Present address: Department of Microbiology, Royal North Shore Hospital, St. Leonards, NSW, Australia.

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