Rickettsioses in South Korea, materials and methods.

To the Editor: We read with interest the article by Choi et al. (1), which describes the molecular detection of Rickettsia typhi and 4 spotted fever group rickettsiae by nested polymerase chain reaction (PCR) in the serum of febrile Korean patients. The value of the study, however, is limited by imprecision, inconsistencies, and the impossibility of verifying data. First, neither epidemiologic nor clinical data are provided for studied patients, although these are essential for interpreting PCR results. Second, multiplex nested PCR is hampered by a high risk of contamination (2). Alternatively, nested PCR techniques that use a closed assay or single-use primers without positive controls limit such a risk (3). In all cases, the use of negative controls is critical (2,3). In this study, negative controls are neither described in the Materials and Methods section nor shown on the gels. In addition, the authors used as positive controls 4 of the 5 Rickettsia species they detected. Therefore, apart from R. felis, which was not used as a positive control, positive products may result from cross-contamination. Finally, technically, the data are impossible to reproduce: 1) primer sets WJ77/80 and WJ79/83/78 cited in the legends of Figures 2 and 3 are neither described nor referenced in the text, 2) sequence of the RpCS.877p primer in Table 1 differs from that in the referenced article (4), 3) described sequences have not been deposited in GenBank, and 4) all rompB primers described in Table 1 exhibit 1-6 nucleotide mismatches with ompB sequences of at least 1 of the detected species. Based on these errors, the 7 cases of dual infections with R. conorii and R. typhi, which have never been reported before, are doubtful, and these data need to be confirmed.

Pierre-Edouard Fournier, * Jean-Marc Rolain, * and Didier Raoult *

* Universite de la Mediterranee, Marseille, France

References

(1.) Choi YJ, Jang WJ, Kim JH, Ryu JS, Lee SH, Park KH, et al. Spotted fever group and typhus group rickettsioses in humans, South Korea. Emerg Infect Dis. 2005;11:237-44.

(2.) Hayden RT. In vitro nucleic acid amplification techniques. In: Persing DH, Tenover FC, Versalovic J, Tang YW, Unger ER, Relman DA, et al., editors. Molecular microbiology. Washington: ASM Press; 2003. p. 43-69.

(3.) Fournier PE, Raoult D. Suicide PCR on skin biopsy specimens for diagnosis of rickettsioses. J Clin Microbiol. 2004;42: 3428-34.

(4.) Roux V, Rydkina E, Eremeeva M, Raoult D. Citrate synthase gene comparison, a new tool for phylogenetic analysis, and its application for the rickettsiae. Int J Syst Bacteriol. 1997;47:252-61.

Address for correspondence: Didier Raoult, CNRS UMR 6020, IFR 48, Faculte de Medecine, Universite de la Mediterranee, 27 Blvd Jean Moulin, 13385 Marseille CEDEX 5, France; fax: [phone]; email: didier. [e-mail]