Vector Interactions and Molecular Adaptations of Lyme Disease and Relapsing Fever Spirochetes Associated with Transmission by Ticks

Pathogenic spirochetes in the genus Borrelia are transmitted primarily by two families of ticks. The Lyme disease spirochete, Borrelia burgdorferi, is transmitted by the slow-feeding ixodid tick Ixodes scapularis, whereas the relapsing fever spirochete, B. hermsii, is transmitted by Ornithodoros hermsi, a fast-feeding argasid tick. Lyme disease spirochetes are generally restricted to the midgut in unfed I. scapularis. When nymphal ticks feed, the bacteria pass through the hemocoel to the salivary glands and are transmitted to a new host in the saliva after 2 days. Relapsing fever spirochetes infect the midgut in unfed O. hermsi but persist in other sites including the salivary glands. Thus, relapsing fever spirochetes are efficiently transmitted in saliva by these fast-feeding ticks within minutes of their attachment to a mammalian host. We describe how B. burgdorferi and B. hermsii change their outer surface during their alternating infections in ticks and mammals, which in turn suggests biological functions for a few surface-exposed lipoproteins

Genome size and organization in the blacklegged tick, \u3ci\u3eIxodes scapularis\u3c/i\u3e and the Southern cattle tick, \u3ci\u3eBoophilus microplus\u3c/i\u3e

Genome sizes and the organization of repetitive DNA were determined in the hard ticks Ixodes scapularis and Boophilus microplus using reassociation kinetics. The I. scapularis genome contains ~ 2.15 pg (2.1 × 10 3 Mbp) of DNA and consists of no foldback (FB), 27% highly repetitive (HR), 39% moderately repetitive (MR), and 34% unique DNA. The B. microplus genome contains 7.5 pg (7.1 × 10 3 Mbp) DNA, and consists of 0.82% FB, 31% HR, 38% MR, and 30% unique DNA. In both species, repetitive sequences occur in a mixture of long and short period interspersion but most (65–80%) of the DNA follows a pattern of short period interspersion. Genome size and organization in the three tick species so far examined are distinct from other arthropods in having a greater proportion of MR, a lower proportion of unique and HR DNA of very low sequence complexity

Genome size and organization in the blacklegged tick, \u3ci\u3eIxodes scapularis\u3c/i\u3e and the Southern cattle tick, \u3ci\u3eBoophilus microplus\u3c/i\u3e

Genome sizes and the organization of repetitive DNA were determined in the hard ticks Ixodes scapularis and Boophilus microplus using reassociation kinetics. The I. scapularis genome contains ~ 2.15 pg (2.1 × 10 3 Mbp) of DNA and consists of no foldback (FB), 27% highly repetitive (HR), 39% moderately repetitive (MR), and 34% unique DNA. The B. microplus genome contains 7.5 pg (7.1 × 10 3 Mbp) DNA, and consists of 0.82% FB, 31% HR, 38% MR, and 30% unique DNA. In both species, repetitive sequences occur in a mixture of long and short period interspersion but most (65–80%) of the DNA follows a pattern of short period interspersion. Genome size and organization in the three tick species so far examined are distinct from other arthropods in having a greater proportion of MR, a lower proportion of unique and HR DNA of very low sequence complexity

Vector Competence of Ixodes scapularis and Ixodes ricinus (Acari: Ixodidae) for Three Genospecies of Borrelia burgdorferi

The vector competence of 2 tick species, Ixodes ricinus (L.) and Ixodes scapularis Say, was determined and compared for 3 genospecies of Borrelia burgdorferi. The 3 genospecies of B. burgdorferi used in the following experiments were Borrelia burgdorferi sensu stricto (B-31 and B-31.D1 clone), Borrelia afzelii (strain Pgau.C3), and Borrelia garinii (strain VS286 and VSBP). Spirochetes from all 5 strains were inoculated intradermally into outbred mice; larval ticks of both species were subsequently fed on those mice and replete larvae were assayed for infection by culture in BSK-H media every 7 d for 4 wk. Infection frequencies in I. scapularis exposed to the 5 strains were as follows: B-31 (90%), B-31.D1 (83%), Pgau.C3 (87%), VS286 (10%), and VSBP (5%). The comparable infection frequencies for /. ricinus were B-31 (3%), B-31.D1 (3%), Pgau.C3 (90%), VS286 (5%), and VSBP (3%). Resultant nymphal /. scapularis successfully transmitted B-31, B-31.D1, Pgau.C3, and VS286 to outbred mice. /. ricinus nymphs transmitted Pgau.C3 and VS286. Both species failed to transmit strain VSB

Partial pathogen protection by tick-bite sensitization and epitope recognition in peptide-immunized HLA DR3 transgenic mice

Ticks are notorious vectors of disease for humans, and many species of ticks transmit multiple pathogens, sometimes in the same tick bite. Accordingly, a broad-spectrum vaccine that targets vector ticks and pathogen transmission at the tick/host interface, rather than multiple vaccines against every possible tickborne pathogen, could become an important tool for resolving an emerging public health crisis. The concept for such a tick protective vaccine comes from observations of an acquired tick resistance (ATR) that can develop in non-natural hosts of ticks following sensitization to tick salivary components. Mice are commonly used as models to study immune responses to human pathogens but normal mice are natural hosts for many species of ticks and fail to develop ATR. We evaluated HLA DR3 transgenic (tg) “humanized” mice as a potential model of ATR and assessed the possibility of using this animal model for tick protective vaccine discovery studies. Serial tick infestations with pathogen-free Ixodes scapularis ticks were used to tick-bite sensitize HLA DR3 tg mice. Sensitization resulted in a cytokine skew favoring a Th2 bias as well as partial (57%) protection to infection with Lyme disease spirochetes (Borrelia burgdorferi) following infected tick challenge when compared to tick naïve counterparts. I. scapularis salivary gland homogenate (SGH) and a group of immunoinformatic-predicted T cell epitopes identified from the I. scapularis salivary transcriptome were used separately to vaccinate HLA DR3 tg mice, and these mice also were assessed for both pathogen protection and epitope recognition. Reduced pathogen transmission along with a Th2 skew resulted from SGH vaccination, while no significant protection and a possible T regulatory bias was seen in epitope-vaccinated mice. This study provides the first proof-of-concept for using HLA DR tg “humanized” mice for studying the potential tick protective effects of immunoinformatic- or otherwise-derived tick salivary components as tickborne disease vaccines

Molecular characterization of a new isolate of Borrelia lusitaniae derived from Apodemus sylvaticus in Portugal

A total of 196 small mammals were collected in Portugal and tested for Borrelia burgdorferi sensu lato. Tissue samples were taken from each animal and cultured in Barbour-Stoenner-Kelly (BSK)-II medium. The single strain of spirochete isolated was confirmed as Borrelia lusitaniae by genetic analyses. This is the first report of B. lusitaniae isolated from Apodemus sylvaticus

Borrelia burgdorferi small lipoprotein Lp6.6 is a member of multiple protein complexes in the outer membrane and facilitates pathogen transmission from ticks to mice

Borrelia burgdorferi lipoprotein Lp6.6 is a differentially produced spirochete antigen. An assessment of lp6.6 expression covering representative stages of the infectious cycle of spirochetes demonstrates that the gene is solely expressed during pathogen persistence in ticks. Deletion of lp6.6 in infectious B. burgdorferi did not influence in vitro growth, or its ability to persist and induce inflammation in mice, migrate to larval or nymphal ticks or survive through the larval-nymphal molt. However, Lp6.6-deficient spirochetes displayed significant impairment in their ability to transmit from infected ticks to naïve mice, which was restored upon genetic complementation of the mutant with a wild-type copy of lp6.6, establishing that Lp6.6 plays a role in pathogen transmission from ticks to mammals. Lp6.6 is a subsurface, yet highly abundant, outer membrane antigen. Two-dimensional blue native/SDS-PAGE coupled with liquid chromatography-mass spectrometry (LC-MS/MS) analysis and protein cross-linking studies independently shows that Lp6.6 exists in multiple protein complexes in the outer membrane. We speculate that the function of Lp6.6 is connected to the physiological processes of these membrane complexes. Further characterization of differentially produced membrane antigens and associated protein complexes will likely aid in our understanding of the molecular details of B. burgdorferi persistence and transmission through a complex enzootic cycle

Evaluation of the preventive capacities of a topically applied azithromycin formulation against Lyme borreliosis in a murine model

Objectives: Systemic antibiotic treatment of Lyme borreliosis is effective during the early stages of the infection, while chronic manifestations of the disease may remain refractory and difficult to treat. This study was carried out in order to evaluate the potential of topically applied azithromycin to eliminate the spirochaetal organisms in the skin of the freshly bitten host and thereby prevent Lyme borreliosis. Methods: Laboratory mice were challenged with Borrelia burgdorferi sensu stricto by needle inoculation or via infected ticks as vectors. Then, an azithromycin-containing formulation was applied once daily to the sites of exposure for three consecutive days. In the case of needle inoculation, a 5% azithromycin formulation was applied starting 1 h, 3 days and 5 days after infection. In the case of tick exposure, 4%, 10% and 20% azithromycin formulations were applied, starting directly after the detachment of the engorged ticks. Subsequently, the infection status of the mice was determined. Results: Concentrations of azithromycin in murine skin were >3800-fold higher than the published minimal inhibitory concentration for B. burgdorferi as soon as 3 h after the first application. After needle inoculation, spirochaetes were not detectable in all infected mice after treatment, if the first application started 1 h or even after 3 days post-infection. Furthermore, no borrelial organisms were detected after topical treatment when ticks were used for spirochaete inoculation. Conclusions: Our data indicate that topical treatment with a formulation containing azithromycin is a promising approach to prevent Lyme borreliosis shortly after a tick bite