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

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.

Presence of Babesia odocoilei and Borrelia burgdorferi Sensu Stricto in a Tick and Dual Parasitism of Amblyomma inornatum and Ixodes scapularis on a Bird in Canada

“…As a stealth pathogen, Bbsl slips by host defenses and sequesters in deep-seated tissue (i.e., ligaments, tendons, bone, eye, brain, muscle, glial and neuronal cells, synovium, and scar tissue) [68,69,70,71,72,73,74,75,76,77,78,79]. The persistence of Bbsl in chronic Lyme disease patients is well documented [4,80,81]. In the advanced stage, patients can have cognitive impairment that results in a gradually developing spectrum of neuropsychiatric symptoms [82,83].…”

Erosive Vulvovaginitis Associated With Borrelia burgdorferi Infection

“…Persistent Borrelia infection despite antibiotic therapy has recently been described, and survival of the spirochete in privileged sites such as the genital tract has been postulated. 7,10 B burgdorferi has been cultured from vaginal and seminal secretions of Lyme disease patients, and the fact that active infection was present in an erosive genital lesion suggests that sexual transmission of Lyme disease may be possible. Although previous studies in animal models suggest that sexual transmission of B burgdorferi may occur, further work is needed to examine this possibility.…”

“…Sections were stained with hematoxylin and eosin (H&E), Dieterle silver nitrate stain, and anti– B burgdorferi immunostain (Abcam), as previously described. 6,7 Vaginal and skin cultures of Borrelia were performed as previously described by inoculation of vaginal secretions or vulvar skin scrapings from the ulceration into Barbour-Stoner-Kelly H–complete medium with 6% rabbit serum (Sigma Aldrich, #B8291) and the following antibiotics: phosphomycin (0.02 mg/mL; Sigma Aldrich), rifampicin (0.05 mg/mL; Sigma Aldrich), and amphotericin B (2.5 µg/mL; Sigma Aldrich). 6,7 Polymerase chain reaction (PCR) amplification of Borrelia DNA and Sanger sequencing of the vaginal culture were performed at Australian Biologics Laboratory as described previously.…”

“…6,7 Vaginal and skin cultures of Borrelia were performed as previously described by inoculation of vaginal secretions or vulvar skin scrapings from the ulceration into Barbour-Stoner-Kelly H-complete medium with 6% rabbit serum (Sigma Aldrich, #B8291) and the following antibiotics: phosphomycin (0.02 mg/mL; Sigma Aldrich), rifampicin (0.05 mg/ mL; Sigma Aldrich), and amphotericin B (2.5 µg/mL; Sigma Aldrich). 6,7 Polymerase chain reaction (PCR) amplification of Borrelia DNA and Sanger sequencing of the vaginal culture were performed at Australian Biologics Laboratory as described previously. 6,7 Culture pellets were obtained by centrifugation, stabilized in AL buffer (Qiagen), and then forwarded to Australian Biologics for B burgdorferi detection and identification.…”

“…6,7 Polymerase chain reaction (PCR) amplification of Borrelia DNA and Sanger sequencing of the vaginal culture were performed at Australian Biologics Laboratory as described previously. 6,7 Culture pellets were obtained by centrifugation, stabilized in AL buffer (Qiagen), and then forwarded to Australian Biologics for B burgdorferi detection and identification. DNA was extracted using the QIAamp DNA Mini Kit (Qiagen).…”