Lyme disease, caused by the spirochete Borrelia burgdorferi, is the most common vector‐borne disease in the United States and Europe. The spirochetes are transmitted from mammalian and avian reservoir hosts to humans via ticks. Following tick bites, spirochetes colonize the host skin and then disseminate haematogenously to various organs, a process that requires this pathogen to evade host complement, an innate immune defence system. CspZ, a spirochete surface protein, facilitates resistance to complement‐mediated killing in vitro by binding to the complement regulator, factor H (FH). Low expression levels of CspZ in spirochetes cultivated in vitro or during initiation of infection in vivo have been a major hurdle in delineating the role of this protein in pathogenesis. Here, we show that treatment of B. burgdorferi with human blood induces CspZ production and enhances resistance to complement. By contrast, a cspZ‐deficient mutant and a strain that expressed an FH‐nonbinding CspZ variant were impaired in their ability to cause bacteraemia and colonize tissues of mice or quail; virulence of these mutants was however restored in complement C3‐deficient mice. These novel findings suggest that FH binding to CspZ facilitates B. burgdorferi complement evasion in vivo and promotes systemic infection in vertebrate hosts.

Lyme disease presents a suitable model for the investigation of host tropism – a pathogen’s ability to colonize and survive in different host species – since its causative agent, the spirochete Borrelia burgdorferi (Bb) is transmitted by ticks and maintained in rodent and bird reservoir hosts. In order to survive in vertebrates and escape from killing by complement, a first-line host immune defense, Bb produces the outer surface protein CspZ that binds the complement inhibitor factor H (FH) to promote infection. Protein sequence conservation seems to be linked to FH-binding activity divergence, raising the hypothesis that even minor variation can confer host-specific, FH-binding-mediated infectivity. Our work shows that that this minor variation is located in a loop in the CspZ protein localized in the CspZ-FH binding interface. Our functional experiments prove that this loop promotes bird- or rodent-specific FH-binding activity and infectivity. Swapping loops between rodent- and bird-associated CspZ variants alters their capability to confer host-specific dissemination. We further investigated the evolutionary mechanisms driving the emergence of the CspZ loop-mediated, host-dependent complement evasion. This multifaceted work demonstrates how a single, short protein motif can significantly impact host tropism.

Modern infectious disease outbreaks often involve changes in host tropism, the preferential adaptation of pathogens to specific hosts. The Lyme disease-causing bacterium Borrelia burgdorferi ( Bb ) is an ideal model to investigate the molecular mechanisms of host tropism, because different variants of these tick-transmitted bacteria are distinctly maintained in rodents or bird reservoir hosts. To survive in hosts and escape complement-mediated immune clearance, Bb produces the outer surface protein CspZ that binds the complement inhibitor factor H (FH) to facilitate bacterial dissemination in vertebrates. Despite high sequence conservation, CspZ variants differ in human FH-binding ability. Together with the FH polymorphisms between vertebrate hosts, these findings suggest that minor sequence variation in this bacterial outer surface protein may confer dramatic differences in host-specific, FH-binding-mediated infectivity. We tested this hypothesis by determining the crystal structure of the CspZ–human FH complex, and identifying minor variation localized in the FH-binding interface yielding bird and rodent FH-specific binding activity that impacts infectivity. Swapping the divergent region in the FH-binding interface between rodent- and bird-associated CspZ variants alters the ability to promote rodent- and bird-specific early-onset dissemination. We further linked these loops and respective host-specific, complement-dependent phenotypes with distinct CspZ phylogenetic lineages, elucidating evolutionary mechanisms driving host tropism emergence. Our multidisciplinary work provides a novel molecular basis for how a single, short protein motif could greatly modulate pathogen host tropism.