Wei-Wen Hung1, Yen-Hsu Chen2, Sung-Pin Tseng3, Ya-Ting Jao4, Lee-Jene Teng5, Wei-Chun Hung6. 1. Division of Endocrinology and Metabolism, Department of Internal Medicine, Kaohsiung Medical University Hospital, Kaohsiung, Taiwan. 2. Graduate Institute of Medicine, Kaohsiung Medical University, Kaohsiung, Taiwan; Division of Infectious Diseases, Department of Internal Medicine, Kaohsiung Medical University Hospital, Kaohsiung, Taiwan. 3. Department of Medical Laboratory Science and Biotechnology, College of Health Sciences, Kaohsiung Medical University, Kaohsiung, Taiwan. 4. Infection Control Center, Kaohsiung Medical University Hospital, Kaohsiung, Taiwan. 5. Department of Clinical Laboratory Sciences and Medical Biotechnology, National Taiwan University College of Medicine, Taipei, Taiwan. Electronic address: ljteng@ntu.edu.tw. 6. Department of Microbiology and Immunology, College of Medicine, Kaohsiung Medical University, Kaohsiung, Taiwan; Department of Medical Research, Kaohsiung Medical University Hospital, Kaohsiung, Taiwan. Electronic address: wchung@kmu.edu.tw.
Abstract
BACKGROUND/ PURPOSE: Accurate identification is important for effective treatment because Enterococcus species have talents to cope with various antibiotics either by intrinsic resistance or by acquisition of mobile genetic elements. The groEL gene is a permissive target in identification of bacteria. We aimed to develop simple assays based on groEL for identification of enterococci. RESULTS: We continued our previous work and determined groEL gene sequences of Enterococcus species isolated from clinical specimens. Phylogenetic analysis based on groEL revealed that each strain clustered well with their reference strains (bootstrap value 100%), in which Enterococcusfaecium and Enterococcusgallinarum could be split into two clades. The divergence of E. faecium was coincident with hospital-associated clade, known as clade A, and community-associated clade, known as clade B. A PCR-restriction fragment length polymorphism (PCR-RFLP) assay was therefore designed to differentiate the two E. faecium clades, based on the specific RsaI cutting sites present in the two clades. To differentiate 7 clinical relevant Enterococcus species, the multiplex PCR assay was designed to identify Enterococcusavium, Enterococcuscasseliflavus, Enterococcusfaecalis, E. faecium, E. gallinarum, Enterococcushirae and Enterococcusraffinosus. Specificity was tested with other Enterococcus species including Enterococcuscecorum, Enterococcusdurans and Enterococcusmundtii. None of these bacterial species generated products of similar size to those of the seven Enterococcus species. CONCLUSION: The simple PCR-RFLP and multiplex PCR assays on the basis of groEL gene provided an alternative way to identify Enterococcus species.
BACKGROUND/ PURPOSE: Accurate identification is important for effective treatment because Enterococcus species have talents to cope with various antibiotics either by intrinsic resistance or by acquisition of mobile genetic elements. The groEL gene is a permissive target in identification of bacteria. We aimed to develop simple assays based on groEL for identification of enterococci. RESULTS: We continued our previous work and determined groEL gene sequences of Enterococcus species isolated from clinical specimens. Phylogenetic analysis based on groEL revealed that each strain clustered well with their reference strains (bootstrap value 100%), in which Enterococcusfaecium and Enterococcusgallinarum could be split into two clades. The divergence of E. faecium was coincident with hospital-associated clade, known as clade A, and community-associated clade, known as clade B. A PCR-restriction fragment length polymorphism (PCR-RFLP) assay was therefore designed to differentiate the two E. faecium clades, based on the specific RsaI cutting sites present in the two clades. To differentiate 7 clinical relevant Enterococcus species, the multiplex PCR assay was designed to identify Enterococcusavium, Enterococcuscasseliflavus, Enterococcusfaecalis, E. faecium, E. gallinarum, Enterococcushirae and Enterococcusraffinosus. Specificity was tested with other Enterococcus species including Enterococcuscecorum, Enterococcusdurans and Enterococcusmundtii. None of these bacterial species generated products of similar size to those of the seven Enterococcus species. CONCLUSION: The simple PCR-RFLP and multiplex PCR assays on the basis of groEL gene provided an alternative way to identify Enterococcus species.
Authors: András Fodor; Birhan Addisie Abate; Péter Deák; László Fodor; Ervin Gyenge; Michael G Klein; Zsuzsanna Koncz; Josephat Muvevi; László Ötvös; Gyöngyi Székely; Dávid Vozik; László Makrai Journal: Pathogens Date: 2020-06-29