SummaryBorrelia burgdo~feri, the spirochetal agent of Lyme disease, is transmitted by Ixodes ticks. A vaccine based on B. burgdo~feri outer surface protein (Osp) A protects mice from spirochete infection. Here we report on the expression of OspA on spirochetes inside engorging ticks and relate OspA expression to antispirochetal immunity. Spirochetes in the gut of unfed nymphal ticks were stained by an OspA antibody, whereas in feeding ticks, the majority of spirochetes in the gut and salivary glands did not stain with the antibody. Thus, OspA was not expressed on most spirochetes during transmission from the vector to the vertebrate host. To examine the mechanism of protection afforded by OspA antibody, mice were passively immunized with OspA antibody at different times relative to tick attachment. When OspA antibody was administered to mice before or at the time of tick attachment, spirochetal development events in the vector, such as growth and salivary gland invasion, were blocked and the mice were protected from B. burgdotferi infection. When OspA antibody was administered to mice 48 h after tick attachment, spirochetes persisted in the nymphs and the mice were not protected despite the presence of circulating antibodies in the host as well as in the tick blood meal. Thus, OspA immunity appears to be effective only during a narrow window time at the beginning of the blood meal when antibodies bind to OspA-expressing spirochetes in the tick gut and block transmission from the vector to the host.T he spirochetal agent of Lyme disease, Borrelia burgdooCeri is transmitted by Ixodes ticks. In unfed nymphal ticks, spirochetes are found in the gut (l). During tick feeding, the bacteria disseminate to the salivary glands and infect the vertebrate host (2-4). A vaccine based on B. burgdo~ri outer surface protein (Osp) A which protects mice from infection is currently being tested in human trials (5-7). Here we examine the expression of OspA on spirochetes within engorging nymphs and relate OspA expression to protection afforded by OspA antibody. Materials and MethodsMice. Pathogen-free C3H/HeN (C3H) mice were purchased from the National Institutes of Health (Bethesda, MD). Maintenance and Infection of Ticks. Ixodes dammini (also known as L scapularis)were derived from a colony in its second generation from field-collected adults, and were free of inherited infection. Larvae were allowed to engorge on CD-1 mice that had been infected 2 wk previously by the bites of three to five nymphal I. dammini containing the N40 strain of B. burgdotferi.Engorged larvae were collected, held in mesh-covered plaster of paris containing vials, and were allowed to molt at 95% relative humidity at 21~ The infected nymphs were held under the same conditions before their use in experiments, usually within 2 mo of molting. Antibody Staining of OspA-expressing Spirochetes in NymphalTicks. Guts and salivary glands were dissected out of nymphs and prepared for antibody staining as previously described (2). mAb CIII.78, which binds to a COOH-t...

Abstract. Parallel experiments in living cells and invitro were undertaken to characterize the mechanism by which misfolded and unassembled glycoproteins are retained in the ER. A thermoreversible folding mutant of vesicular stomatitis virus (VSV) G protein called ts045 was analyzed. At 39°C, newly synthesized G failed to fold correctly according to several criteria: intrachain disulfide bonds were incomplete; the B2 epitope was absent; and the protein was associated with immunoglobulin heavy chain binding protein (BiP), a heat shock-related, ER protein. When the temperature was lowered to 32°C, these properties were reversed, and the protein was transported to the cell surface.Upon the shift up from 32°C back to 39°C, G protein in the ER returned to the misfolded form and was retained, while the protein that had reached a pre-Golgi compartment or beyond was thermostable and remained transport competent. The misfolding reaction could be reconstituted in a cell free system using ts045 virus particles and protein extracts from microsomes. Taken together, the results showed that ER is unique among the organelles of the secretory pathway in containing specific factors capable of misfolding G protein at the nonpermissive temperature and thus participating in its retention.

Ixodes ticks are infected by Borrelia burgdorferi when larvae feed on spirochete-infected mice. We studied the acquisition of B. burgdorferi by larval ticks, characterized the production of outer surface protein A (OspA) by spirochetes entering larvae, and examined the effects of OspA antibodies on the establishment of B. burgdorferi infections in ticks. Most larvae were infected by spirochetes 24 to 48 h after placement on mice. OspA antibodies stained the first spirochetes observed in larvae, suggesting that OspA is synthesized early during the colonization of the vector. When OspA antibodies were administered to B. burgdorferi-infected mice and larvae were then placed on the animals, the severity of larval infection and the number of infected ticks (7 of 16) were decreased compared with that of controls (15 of 16). The inhibitory effects of OspA antibodies were observed with passive antibody transfer as well as active host-generated immunity. The lower larval infection rate observed in the presence of OspA antibodies was exacerbated after the larval molt since only 1 of 12 nymphs was infected, and none of the mice that were fed upon by these nymphs became infected with B. burgdorferi. Therefore, an OspA antibody response in mice altered the reservoir competence of the vertebrate host by inhibiting the movement of B. burgdorferi from the host to the vector.