A Study of Ni-resistant bacteria isolated from gingival crevicular fluid on the patients wearing Ni-Cr alloy prosthesis (In terms of molecular biological aspects)

니켈-크롬 합금 보철물 주위 치은열구 내에서 발견된 니켈 내성 균주에 관한 분자생물학적 연구

As a material of metal-ceramic prosthesis, nickel as a form of Ni-Cr alloy has been used for many dental prostheses in many cases. However, several problems in use of the alloy have been revealed (ex : tissue stimulation, skin allergy, hypersensitivity, cytotoxicity and carcinogenecity). Little is known about nickel with respect to the relationship between Ni-prosthesis and gaining of Ni-resistance in oral microorganisms. The present study was undertaken to check wheather use of Ni-prosthesis leads to occurrence of Ni-resistant microorganisms. So this study may suggest the possible relationships between the oral microorganisms and nickel-resistance in oral environment. Bacteria were isolated from the gingival crevicular fluid on the pateints wearing Ni-Cr prosthesis. The isolated bacteria were tested for their Ni-resistance in nickel containing media at different concentration from 3mM to 110mM. E. coli HB101 was used as control. The Ni-resistant bacteria were isolated and biochemically identified. The Ni-resistant bacteria were tested several bio-chemical, molecular-biological tests. Performed tests were ; measuring the growth curve, antibiotic test, growth ability test in liquid media, isolation of the chromosome and plasmid, digestion of DNA by restriction enzyme, electrophoresis of chromosome and plasmid DNA, identification of Ni-resistant genes by the DNA hybridization. The results were as follows: 1) The bacteria isolated from gingival crevicular fluid on the patients wearing Ni-Cr alloy pros-thesis showed nickel-resistance. 2) The isolated microorganisms grew at nickel containing media of high concentrations (60mM-110mM). 3) Based on the biochemical tests, the isolated microorganisms were identified as Enterococcus faecalis(13 cases), Klebsiella pneumoniae(1 case) and Enterobacter gergeviae(1 case). 4) Enterococcus faecalis expressed not only nickel resistance but also the multi-drug resistance to several antibiotics ; chloramphenicol, kanamicin, streptomycin, lincomycin, clindamycin. However, all strain showed the sensitivity against the tetracycline. 5) DNA hybridization result suggest that there is no homology between the previousely known gene of nickel resistance in Klebsiella pneumoniae and chromosomal DNA of Enterococcus faecalis.