Spirochetes are gram-negative bacteria, slender and motile, with 3 to 30 uneven coils. They are coiled in a helical shape with one or more complete turns.^15,16 The Borrelia species are microaerophilic, fastidious bacteria. They, along with leptospira and treponema, belong to the eubacterial phylum of spirochetes.¹⁷ Lyme disease is caused by one of three spcies: B. burgdorferi, B. afzelii, or B. garinii, with B. burgdorferi causing disease in the United States, and the latter two causing most Lyme disease in Europe and Asia.¹⁸ The Borrelia species, like all other spirochetes, have a protoplasmic cylinder encased by a cell membrane, a flagellum, and finally an outer membrane that is loosely associated with the underlying structures.¹⁷ The entire outer membrane can move to one end of the cylinder, a phenomenon that may be important in cellular adherence called capping or patching.¹⁹ B. burgdorferi is the longest (20 to 30 μm) and narrowest (0.2 to 0.3 μm) of the Borrelia species and has the fewest flagella (7 to 11).²⁰

B. burgdorferi has a ratio of guanine to cytosine of approximately 1:3, and it is 31% to 59% DNA homologous with other borrelia.²¹ The genome of B. burgdorferi contains 1.5 mega-baes, consisting of a linear chromosome 950 kilobases in length with 9 linear and 12 circular plasmids.²² Much of its genome encodes lipoproteins, including the outer-surface membrane proteins A-F (Osp A-F), encoded on plasmid DNA, with only a relatively small part encoding proteins that promote biosynthetic activity, making the organism dependent on its host for its nutritional survival.²² It is hypothesized that the outer surface protein aid in the adaptation of the organism to many different environments, including the host and vector sites, thereby serving as important factors determining the pathogenicity of the organism.²³ The cell wall of B. burgdorferi contains a surface-exposed lipoprotein, vlsE, that has been shown to undergo antigenic variation early in the disease course.²⁴ The 41-kd antigen located on the flagellum is similar to flagellar antigens of other spirochetes²⁵ and the 58- or 60-kd antigen appears to be a heat-shock protein that is cross-reactive with an equivalent antigen (58 to 65 kd) in a wide range of bacteria.²⁶

Borrelia species grow best at 33°C in a complex liquid medium called Barbour-Stoenner-Kelly medium,²⁷ although B. burgdorferi can grow in colonies when this medium is solidified with 1.3% agarose.²⁸ Borrelia grow slowly compared with most bacteria. Each spirochete grows in length for 12 to 24 hours before dividing into two cells.²⁷ Although a primary isolate of this spirochete is easily obtained from ticks, it is difficult to obtain from patients.^10,11 After 10 to 15 passages, B. burgdorferi loses its pathogenicity in culture and the organisms are no longer infectious.²⁹

Borrelia burgdorferi is transmitted by certain Ixodes ticks^{7,30,31,32,33} and possibly by biting flies.^34,35,36 In the United States, Lyme disease is seen primarily in three regions: (1) Northeast; (2) Midwest; primarly in Minnesota and Wisconsin; and (3) Northwest, mainly in Northern California and Oregon. Ixodes ticks that are part of the Ixodes ricinus complex include Ixodes scapularis in the northeastern and midwestern United States,^7,30 Ixodes pacificus in the western United States,³¹ Ixodes ricinus in Europe,³² and Ixodes persulcatus in Asia.³³ A number of ixodid ticks are found in Australia; however, because the vector of infection has not yet been identified, there is controversy as to whether Lyme disease truly exists in Australia.^37,38

Ticks of the I. ricinus complex go through three stages during their 2-year life cycle: the larval, nymph, and adult stage. They feed once during each of the three stages; larval ticks take one blood meal in late summer, nymphs feed during the following spring and early summer, and adults feed during that autumn.³⁹ The preferred host for both the larval and the nymph stages of I. scapularis in the United States is the white-footed mouse, Peromyscus leucopus.³⁰ It is critical that the tick, in both of its immature stages, feed on the same host because the life cycle of the spirochete depends on horizontal transmission. B. burgdorferi is transmitted from infected nymphs to mice in early summer and from infected mice to larvae in late summer. The larvae then molt to become infected nymphs that begin the cycle in the following year.⁴⁰ The white-footed mice are tolerant to infection with B. burgdorferi and are capable of remaining spirochetemic throughout the summer without an inflammatory response.³⁰ The life cycle of the larval and nymph stages is as follows: The tick feeds on the infected mouse, the spirochetes remain in the midgut of the tick until the following year, and then they develop into nymphs. The tick attaches itself to another host, and the B. burgdorferi spirochetes migrate to the tick’s salivary glands and are injected with its saliva as it feeds.^41,42 The tick must remain attached for 24 hours or more before transmission occurs.⁴³

The preferred host for I. scapularis‘s adult stage is the white-tailed deer.⁴⁴ The deer are not involved in the life cycle of the spirochete but are probably critical to the survival of the ticks.⁴⁵ If the deer are withdrawn from the life cycle, the ticks may adapt and survive on other animal hosts.⁴⁵ Infected ticks may bite human during either of the three stages, but Lyme disease most commonly occurs in late spring or early summer, the height of nymph feeding. Transmission of the spirochete to humans occurs if the tick is attached to the person for at least 24 hours.⁴⁶ Clinical Lyme disease is not known to occur in wild animals, although it does occur in domestic animals, including dogs,⁴⁷ horses,^48,49,50 and cattle.^50,51 Lyme disease may occur even in urban areas where there are parks with trees. In Bridgeport, CT, 28% of deer were found to be infected with adult female ticks.⁵²

As noted earlier, the transmission of the B. burgdorferi spirochete form host to vector and finally to infecting a person involves an intricate life cycle. In order to maintain itself within this life cycle, the spirochete must be able to modify and adapt to different environments. As described earlier, various outer surface proteins (Osp) are able to undergo antigenic variation in order to help the organism adapt. When the spirochete is contained within the midgut of the tick, Osp A is expressed,⁵³ and when the tick feeds and the spirochete travels to the salivary gland and human host, Osp C is expressed more.⁵⁴

The clinical features of all stages of Lyme disease are described in detail later in this chapter. Once a person becomes infected with B. burgdorferi, the earliest manifestation is a skin reaction, when both an innate and adaptive immune response is initiated.¹⁸ The spirochete can then disseminate throughout the body, and there has been considerable progress in defining various factors that may aid in the dissemination and therefore infectiveness of B. burgdorferi. Osp C sequences are highly variable, and only four of the many groups of sequence are associated with disseminated disease.⁵⁵ Furthermore, spirochetes bind to a number of receptors on host cells.¹⁸ There is a spirochetal protein that binds to platelet-specific integrin α_IIbβ_3,⁵⁶ a glycosaminoglycan binding protein on the spirochete that binds endothelial heparin sulfate and dermatin sulfate,⁵⁷ spirochetal protein, BBK32, that binds fibronectin,⁵⁸ and spirochetal proteins DbpA and DbpB that binds to decorin, which is found to collagen.⁵⁹ Coleman et al. have demonstrated that plasminogen binding by the spirochete and spirochetal upregulation and activation of matrix metalloproteinases and urokinase-type plasminogen activator receptor (uPAR, CD87) are all important for dissemination and enhancement of sphirochetemia.^60,61,62

During infection with B. burgdorferi, the patient’s immune response produces T- and B- cell responses to a variety of spirochetal antigens, including Osp A and Osp B.^63,64,65 Immunoglobulin (Ig) M and IgG are produced as part of the immune response, and killing of spirochetes is mediated by borreliacidal antibody production⁶⁶ and the classical complent pathway.⁶⁷ As in chronic forms of Lyme disease, B. burgdorferi may survive for a long period of time. This may be mediated by downregulation of Osp C⁶⁸ and antigenic variation of vlsE.²⁴

The Syrian hamster^69,70 has been shown to be susceptible to infection by human and tick isolates of B. burgdorferi. At least 91% of Syrian hamsters had one or more culture-positive organs when infected with 108 spirochetes by intraperitoneal injection.⁶⁹ In this study, a spirochetemia was present for the first 6 days of infection. At 14 days postinfection, when spirochetemia could not be detected, spirochetes were isolated from the spleen, kidney, liver, testes, brain, and 45% of homogenized eyes. Spirochetes were isolated from the eyes and kidney of one animal 52 days postinfection, suggesting that these organisms may cause a persistent multiorgan infection. In another study, seven hamsters were inoculated intraperitoneally with B. burgdorferi and examined by both cultural and histologic techniques at 1 to 9 months postinfection.⁷⁰ The eyes of four of the seven hamsters were culture-positive for spirochetes 14 days after inoculation. Spirochetes were noted in the vitreous of two of the seven hamsters by histology. More spirochetes were found in the eye than in the liver, spleen, or kidneys. No pathologic tissue changes were noted in the globes, except for occasional mononuclear phagocytic cells in the vitreous. The only pathologic tissue changes noted in this study were a rare lymphocytic focus in the liver of one hamster and a chronic portal triaditis in another.

The lack of morphologic organ damage and severe inflammation in these experimentally infected hamsters contrasts with histopathologic alterations seen in infected human tissue.⁷¹ The spirochete has been demonstrated in the vitreous of a patient with a panophthalmitis.⁷² It is possible that the lack of a cellular response and an intermittent spirochetemia in small animals may be factors in the continuing presence and maintenance of the spirochete in its natural habitat.⁷⁰

Many other animal models of Lyme disease have been developed, including gerbil, guinea pig, dog, and rat. The animal models that have been studied the most, however, are the mouse, hamster, and monkey models.⁷³ Rhesus monkeys inoculated with B. burgdorferi have an illness similar to human Lyme disease. Pachner et al. have found elevated blood and spinal fluid levels of interleukin-6 in infected monkeys.^74,75

Animal models have been used particularly to study the pathogenesis of Lyme neuroborreliosis.⁷⁵ The adult Rhesus monkey has developed erythema migrans within a few weeks after intradermal injection with the N40Br strain of B. burgdorferi.⁷⁵ Anti-B. burgdorferi antibodies may develop within the first few weeks and in the third and fourth week after the injection. Spirochete DNA can be detected in the blood using polymerase chain reaction (PCR). Soon after, the animals develop spirochetal invasion of the central nervous system with 50 to 300 white cells and antibodies against B. burgdorferi in the spinal fluid. These studies support the concept that spirochetemia and invasion of the spinal fluid occur at an early stage in Lyme disease, even though they may be clinically asymptomatic. It also adds support to the theory that B. burgdorferi can cause latent infection in both central and peripheral nervous tissue.⁷⁶

HLA specificities encoded by certain class II, d-locus alleles of the major histocompatibility complex are associated with susceptibility to a number of diseases with autoimmune features. The class II molecules are expressed primarily on B cells and macrophages that bind and present antigen to T-helper cells, which then initiate an immune response against these antigens. The development of autoimmune responses can influence genetic variations in the structure of class II molecules by affecting the composition of the T-cell sequencing in the development of self-tolerance during thymic maturation or by determining the type and manner of antigen binding.

There is noted to be an association between HLA-DR2 and DR4 and certain manifestations of Lyme disease.^77,78,79 In a study of 32 patients with meningopolyneuritis,⁷⁷ 56% were HLA-DR2-positive and 46% were HLA-DR4-positive. In a study of 22 patients with acrodermatitis,⁷⁸ 52% had HLA-DR2.^80,81 A late neurologic manifestation of Lyme disease, borrelial encephalomyelitis,⁸² may resemble multiple sclerosis. Multiple sclerosis is also associated with an increased frequency of HLA-DR2.⁸³

Steere et al. demonstrated an association of chronic Lyme arthritis with HLA-DR4 and HLA-DR2 alleles.⁷⁹ Of 80 patients with Lyme arthritis, 57% of those with chronic arthritis were associated with this histocompatibility antigen complex. Only 23% of those with arthritis of moderate duration (6 to 11 months) and only 9% of those with arthritis of short duration (1 to 5 months) had this specificity. After the HLA-DR4-positive patients were excluded from each group, a secondary association was noted with HLA-DR2 in 75% of the remaining patients with chronic arthritis, 50% of those with arthritis of moderate duration, and only 20% of those with arthritis of short duration.

The presence of HLA-DR4 in patients with arthritis was associated with a lack of response to antibiotic therapy. The frequency of treatment failure was not significantly increased in patients with HLA-DR2 specificity even after the exclusion of HLA-DR4-positive patients from the analysis. Thus, it is possible that the molecular basis of susceptibility may not be the same for both specificities.

Kalish, Leong, and Steere later determined that there is an association between chronic Lyme arthritis unresponsive to antibiotic treatment and the combination of HLA-DR4 specificity and Osp A or Osp B reactivity.⁸⁴ Approximately 10% of patients with Lyme arthritis develop chronic arthritis even after appropriate antibiotic treatment, and most of these patients have HLA-DRB1*0401 or related alleles.⁸⁵ Borrelia burgdorferi DNA is not present on PCR of synovial fluid from these chronic Lyme arthritis patients, but was detectable prior to antibiotic initiation,^86,87 indicating that the arthritis is not caused by chronic infection, but rather some immune response. Kalish, et al. showed that the strength of IgG reativity with one or more epitopes on the c-terminal fragment of Osp A correlates with the duration of chronic Lyme arthritis.⁸⁸ The immunodominant epitope of Osp A presented by the DRB1*0401 molecule was predicted to be located at amino acid 165-173, and this region has a sequence homologous to only one human protein, human-lymphocyte-function-associated antigen 1 (hLFA-1).^89,91 Therefore, chronic inflammation in patients with chronic Lyme arthritis seems to be secondary to molecular mimcry between an immunodominant T-cell epitope of Osp A and the protein hLFA-1.^89,90,91 The immunogenetic susceptibility secondary to cross-reactivity between B. burgdoreri antigen Osp A and a human host antigen hLFA-1 is the only form of chronic inflammatory arthritis with a known molecular mimicry component.⁹⁰