Lyme disease (LD), which is caused by genospecies of the Borrelia burgdorferi sensu lato complex, is the most common vector-borne disease in the Northern hemisphere. Spirochetes are transmitted by Ixodes ticks and maintained in diverse vertebrate animal hosts. Following tick bite, spirochetes initially establish a localized infection in the skin. However, they may also disseminate hematogenously to several distal sites, including heart, joints, or the CNS. Because they need to survive in diverse microenvironments, from tick vector to mammalian hosts, spirochetes have developed multiple strategies to combat the numerous host defense mechanisms. One of these strategies includes the production of a number of complement-regulator acquiring surface proteins (CRASPs) which encompass CspA, CspZ, and OspE paralogs to blunt the complement pathway. These proteins are capable of preventing complement activation on the spirochete surface by binding to complement regulator Factor H. The genes encoding these CRASPs differ in their expression patterns during the tick-to-host infection cycle, implying that these proteins may exhibit different functions during infection. This review summarizes the recent published reports which investigated the roles that each of these molecules plays in conferring tick-borne transmission and dissemination in vertebrate hosts. These findings offer novel mechanistic insights into LD pathobiology and may facilitate the identification of new targets for preventive strategies against Lyme borreliosis.

The three genospeciesBorrelia burgdorferi,Borrelia garinii, andBorrelia afzelii, all causative agents of Lyme disease, differ in their susceptibilities to human complement-mediated lysis. We recently reported that serum resistance of borrelias correlates largely with their ability to bind the human complement regulators FHL-1/reconectin and factor H. To date, two complement regulator-acquiring-proteins (CRASP-1 and CRASP-2) have been identified in serum-resistantB.afzeliiisolates (P. Kraiczy, C. Skerka, M. Kirschfink, V. Brade, and P. F. Zipfel, Eur. J. Immunol. 31:1674–1684, 2001). Here, we present a comprehensive study of the CRASPs detectable in both serum-resistant and intermediate serum-sensitiveB. afzeliiandB. burgdorferiisolates. These CRASPs were designated according to the genospecies either as BaCRASPs, when derived fromB. afzelii, or as BbCRASPs, for proteins identified inB. burgdorferiisolates. Each borrelial isolate expresses distinct CRASPs that can be differentiated by their mobility and binding phenotypes. A detailed comparison reveals overlapping and even identical binding profiles for BaCRASP-1 (27.5 kDa), BbCRASP-1 (25.9 kDa), and BbCRASP-2 (23.2 kDa), which bind FHL-1/reconectin strongly and interact weakly with factor H. In contrast, twoB. afzeliiproteins (BaCRASP-4 [19.2 kDa] and BaCRASP-5 [22.5 kDa]) and threeB. burgdorferiproteins (BbCRASP-3 [19.8 kDa], BbCRASP-4 [18.5 kDa], and BbCRASP-5 [17.7 kDa]) bind factor H but not FHL-1/reconectin. Most CRASPs bind both human immune regulators at their C-terminal ends. Temperature-dependent up-regulation of CRASPs (BaCRASP-1, BaCRASP-2, and BaCRASP-5) is detected in low-passage borrelias cultured at 33 or 37°C compared with those cultured at 20°C. The characterization of the individual CRASPs on the molecular level is expected to identify new virulence factors and potential vaccine candidates.

Summary: Borrelia burgdorferi, the aetiological agent of Lyme disease, employs sophisticated means to survive in diverse mammalian hosts. Recent studies demonstrated that acquisition of complement regulators factor H and factor H‐like protein‐1 (FHL‐1) allows spirochetes to resist complement‐mediated killing. Serum‐resistant B. burgdorferi express up to five distinct complement regulator‐acquiring surface proteins (CRASPs) that bind factor H and/or FHL‐1. In this study we have identified and characterized one of those B. burgdorferi proteins, named BbCRASP‐2. BbCRASP‐2 is distinct from the four previously identified factor H/FHL‐1‐binding CRASPs of B. burgdorferi strains. The single copy of the gene encoding BbCRASP‐2, cspZ, is located on the linear plasmid lp28‐3. BbCRASP‐2 is highly divergent from the factor H/FHL‐1‐binding protein BbCRASP‐1 and from members of the factor H‐binding Erp (OspE/F‐related) protein family. Peptide mapping analysis revealed that the factor H/FHL‐1 binding site is discontinuous and it was found that C‐terminal truncations abrogate factor H and FHL‐1 binding. The predominant BbCRASP‐2 binding site of both host complement regulators was mapped to the short consensus repeat 7 (SCR 7). Factor H and FHL‐1 bound to BbCRASP‐2 maintain cofactor activity for factor I‐mediated C3b inactivation and accelerate the decay of the C3 convertase. Expression of BbCRASP‐2 in serum‐sensitive B. burgdorferi mutant B313 increased resistance to complement‐mediated lysis. The characterization of BbCRASP‐2 now provides a complete picture of the three diverse complement regulator‐binding protein families of B. burgdorferi yielding new insights into the pathogenesis of Lyme disease.

The causative agents of Lyme disease, Borrelia burgdorferi s.s., B. garinii, and B. afzelii, differ in their susceptibility to complement‐mediated lysis. This phenomenon apparently depends on the expression of proteins termed complement regulator‐acquiring surface proteins (CRASP) and their binding to the inhibitory plasma proteins factor H and FHL‐1. To characterize these bacterial proteins in more detail we have now isolated from a B. burgdorferi expression library a novel factor H‐binding protein. In accordance with our previous studies this protein was termed BbCRASP‐3 and represents a novel member of the polymorphic Erp (OspE/F‐related) protein family. On the basis of protease accessibility assays using intact spirochetes, BbCRASP‐3 isidentified as a surface‐exposed protein and binds the C‐terminal short consensus repeats of factor H. Applying deletion mutants of BbCRASP‐3, the factor H‐binding site was mapped to the nine‐amino‐acid motif LEVLKKNLK localized at the C‐terminal end of BbCRASP‐3. Factor H bound to BbCRASP‐3 maintains its cofactor activity in factor I‐mediated C3b inactivation. Binding of BbCRASP‐3 to factor H can be inhibited by heparin, a physiological ligand of the complement regulator factor H. Blocking of factor‐H‐binding by soluble BbCRASP‐3 leads to an increase of complement deposition on intermediate serum‐resistant strain ZS7. In conclusion, BbCRASP‐3 has been identified as a novel factor H‐binding protein on B. burgdorferi which by conferring complement resistance to the pathogen may contribute to its persistence in the mammalian host.