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      Culture and identification of Borrelia spirochetes in human vaginal and seminal secretions

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          Abstract

          Background: Recent reports indicate that more than 300,000 cases of Lyme disease are diagnosed yearly in the USA. Preliminary clinical, epidemiological and immunological studies suggest that infection with the Lyme disease spirochete Borrelia burgdorferi (Bb) could be transferred from person to person via intimate human contact without a tick vector. Failure to detect viable Borrelia spirochetes in vaginal and seminal secretions would argue against this hypothesis.

          Methods: Patients with and without a history of Lyme disease were selected for the study after informed consent was obtained. Serological testing for Bb was performed on all subjects. Semen or vaginal secretions were inoculated into BSK-H medium and cultured for four weeks. Examination of genital cultures and culture concentrates for the presence of spirochetes was performed using light and darkfield microscopy, and spirochete concentrates were subjected to Dieterle silver staining, anti-Bb immunohistochemical staining, molecular hybridization and PCR analysis for further characterization. Immunohistochemical and molecular testing was performed in three independent laboratories in a blinded fashion. Positive and negative controls were included in all experiments.

          Results: Control subjects who were asymptomatic and seronegative for Bb had no detectable spirochetes in genital secretions by PCR analysis. In contrast, spirochetes were observed in cultures of genital secretions from 11 of 13 subjects diagnosed with Lyme disease, and motile spirochetes were detected in genital culture concentrates from 12 of 13 Lyme disease patients using light and darkfield microscopy. Morphological features of spirochetes were confirmed by Dieterle silver staining and immunohistochemical staining of culture concentrates. Molecular hybridization and PCR testing confirmed that the spirochetes isolated from semen and vaginal secretions were strains of Borrelia, and all cultures were negative for treponemal spirochetes. PCR sequencing of cultured spirochetes from three couples having unprotected sex indicated that two couples had identical strains of Bb sensu stricto in their semen and vaginal secretions, while the third couple had identical strains of B. hermsii detected in their genital secretions.

          Conclusions: The culture of viable Borrelia spirochetes in genital secretions suggests that Lyme disease could be transmitted by intimate contact from person to person. Further studies are needed to evaluate this hypothesis.

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          Most cited references68

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          Lyme disease-a tick-borne spirochetosis?

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            Lyme disease-a tick-borne spirochetosis?

            A treponema-like spirochete was detected in and isolated from adult Ixodes dammini, the incriminated tick vector of Lyme disease. Causally related to the spirochetes may be long-lasting cutaneous lesions that appeared on New Zealand White rabbits 10 to 12 weeks after infected ticks fed on them. Samples of serum from patients with Lyme disease were shown by indirect immunofluorescence to contain antibodies to this agent. It is suggested that the newly discovered spirochete is involved in the etiology of Lyme disease.
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              Biological basis for syphilis.

              Syphilis is a chronic sexually transmitted disease caused by Treponema pallidum subsp. pallidum. Clinical manifestations separate the disease into stages; late stages of disease are now uncommon compared to the preantibiotic era. T. pallidum has an unusually small genome and lacks genes that encode many metabolic functions and classical virulence factors. The organism is extremely sensitive to environmental conditions and has not been continuously cultivated in vitro. Nonetheless, T. pallidum is highly infectious and survives for decades in the untreated host. Early syphilis lesions result from the host's immune response to the treponemes. Bacterial clearance and resolution of early lesions results from a delayed hypersensitivity response, although some organisms escape to cause persistent infection. One factor contributing to T. pallidum's chronicity is the paucity of integral outer membrane proteins, rendering intact organisms virtually invisible to the immune system. Antigenic variation of TprK, a putative surface-exposed protein, is likely to contribute to immune evasion. T. pallidum remains exquisitely sensitive to penicillin, but macrolide resistance has recently been identified in a number of geographic regions. The development of a syphilis vaccine, thus far elusive, would have a significant positive impact on global health.
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                Author and article information

                Journal
                F1000Res
                F1000Res
                F1000Research
                F1000Research
                F1000Research (London, UK )
                2046-1402
                27 April 2015
                2014
                : 3
                : 309
                Affiliations
                [1 ]International Lyme and Associated Diseases Society, Bethesda, MD, 20827-1461, USA
                [2 ]Australian Biologics, Sydney, NSW 2000, Australia
                [3 ]Department of Biology and Environmental Science, University of New Haven, West Haven, CT, 06516, USA
                [1 ]Institute of Parasitology, Biology Centre Czech Academy of Sciences, České Budějovice, Czech Republic
                [1 ]Division of Bacteriology and Parasitology, Tulane National Primate Research Center, Tulane University Health Sciences, Covington, LA, USA
                [1 ]School of Life Sciences, University of Glasgow, Glasgow, UK
                [1 ]Division of Bacteriology and Parasitology, Tulane National Primate Research Center, Tulane University Health Sciences, Covington, LA, USA
                USMA, USA
                [1 ]School of Life Sciences, University of Glasgow, Glasgow, UK
                [1 ]Falmouth Hospital, Falmouth, MA, USA
                USMA, USA
                Author notes

                MJM recruited patients, performed the spirochete cultures and wrote the original manuscript. CB, KRF, AT and ES performed the IFA and PCR studies. JB, YW and AF performed the PCR studies. HAS and PJM provided patient samples and edited the manuscript. RBS recruited patients, coordinated all studies, revised the manuscript and edited it for publication. All authors approved the manuscript for publication.

                Competing interests: The authors have no competing interests to declare. Preliminary results of the study were presented at the Western Regional Meeting of the American Federation for Medical Research, Carmel, CA, on January 25, 2014, and published in abstract form ( J Invest Med 2014; 62: 280–1).

                Competing interests: No competing interests were disclosed.

                Competing interests: No competing interests were disclosed.

                Competing interests: No competing interests were disclosed.

                Competing interests: No competing interests were disclosed.

                Competing interests: None

                Competing interests: No competing interests were disclosed.

                Competing interests: No competing interests were disclosed.

                Competing interests: No competing interests were disclosed.

                Article
                10.12688/f1000research.5778.3
                5482345
                28690828
                6420cdc0-6bae-4643-89ce-f5f448d4a191
                Copyright: © 2015 Middelveen MJ et al.

                This is an open access article distributed under the terms of the Creative Commons Attribution Licence, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

                History
                : 22 April 2015
                Funding
                Supported in part by a grant to MJM from the Lindorf Family Foundation, Newark, OH. This work is dedicated to the memory of Dr. Willy Burgdorfer.
                The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.
                Categories
                Research Article
                Articles
                Applied Microbiology
                Bacterial Infections
                Clinical Immunology

                lyme borreliosis,chronic lyme disease,borrelia burgdorferi,spirochetes,sexual transmission.

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