Abstract
An EcoRI fragment (1.2 kb) which is highly conserved among Leptospira interrogans isolated in Korea was cloned into pBluescript vector from L. interrogans serovar lai WH20. The EcoRI fragment was sequenced, and a pair of primers (LP1 and LP2) was designed for PCR assay. PCR amplification of target DNA obtained from cultured L. interrogans showed that 274 bp could be detected when as little as 100 fg of leptospiral genomic DNA was used in the reaction mixture. No amplification of DNA was detected from DNA of Leptospira biflexa serovars patoc and sau paulo, Borrelia burgdorferi, Staphylococcus aureus, Escherichia coli, and Salmonella typhimurium. Amplification of 274-bp target DNA could be detected in DNA samples purified from 500 microliters of blood collected from experimentally infected gerbils 2 days after infection, while antibodies to L. interrogans could be detected by the microscopic agglutination test 7 days after infection. The specificity and high sensitivity of the test provided valuable tools for the early diagnosis of leptospirosis.
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- Cole J. R., Jr, Sulzer C. R., Pursell A. R. Improved microtechnique for the leptospiral microscopic agglutination test. Appl Microbiol. 1973 Jun;25(6):976–980. doi: 10.1128/am.25.6.976-980.1973. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Le Febvre R. B., Foley J. W., Thiermann A. B. Rapid and simplified protocol for isolation and characterization of leptospiral chromosomal DNA for taxonomy and diagnosis. J Clin Microbiol. 1985 Oct;22(4):606–608. doi: 10.1128/jcm.22.4.606-608.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Millar B. D., Chappel R. J., Adler B. Detection of leptospires in biological fluids using DNA hybridisation. Vet Microbiol. 1987 Oct;15(1-2):71–78. doi: 10.1016/0378-1135(87)90131-3. [DOI] [PubMed] [Google Scholar]
- Mérien F., Amouriaux P., Perolat P., Baranton G., Saint Girons I. Polymerase chain reaction for detection of Leptospira spp. in clinical samples. J Clin Microbiol. 1992 Sep;30(9):2219–2224. doi: 10.1128/jcm.30.9.2219-2224.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Nielsen J. N., Armstrong C. H., Nielsen N. C. Relationship among selected Leptospira interrogans serogroups as determined by nucleic acid hybridization. J Clin Microbiol. 1989 Dec;27(12):2724–2729. doi: 10.1128/jcm.27.12.2724-2729.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Ralph D., McClelland M., Welsh J., Baranton G., Perolat P. Leptospira species categorized by arbitrarily primed polymerase chain reaction (PCR) and by mapped restriction polymorphisms in PCR-amplified rRNA genes. J Bacteriol. 1993 Feb;175(4):973–981. doi: 10.1128/jb.175.4.973-981.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Shinozaki F., Sada E., Tamai T., Kobayashi Y. Characterization of Leptospira strains HY-1, HY-2, and HY-10 isolated in Korea by means of monoclonal antibodies and restriction endonuclease DNA analysis. Am J Trop Med Hyg. 1992 Mar;46(3):342–349. doi: 10.4269/ajtmh.1992.46.342. [DOI] [PubMed] [Google Scholar]
- Terpstra W. J., Schoone G. J., ter Schegget J. Detection of leptospiral DNA by nucleic acid hybridisation with 32P- and biotin-labelled probes. J Med Microbiol. 1986 Aug;22(1):23–28. doi: 10.1099/00222615-22-1-23. [DOI] [PubMed] [Google Scholar]
- Zuerner R. L., Bolin C. A. Repetitive sequence element cloned from Leptospira interrogans serovar hardjo type hardjo-bovis provides a sensitive diagnostic probe for bovine leptospirosis. J Clin Microbiol. 1988 Dec;26(12):2495–2500. doi: 10.1128/jcm.26.12.2495-2500.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Zuerner R. L. Physical map of chromosomal and plasmid DNA comprising the genome of Leptospira interrogans. Nucleic Acids Res. 1991 Sep 25;19(18):4857–4860. doi: 10.1093/nar/19.18.4857. [DOI] [PMC free article] [PubMed] [Google Scholar]