OspC facilitates Borrelia burgdorferi invasion of Ixodes scapularis salivary glands
Utpal Pal, Xiaofeng Yang, Manchuan Chen, Linda K. Bockenstedt, John F. Anderson, Richard A. Flavell, Michael V. Norgard, Erol Fikrig
Utpal Pal, Xiaofeng Yang, Manchuan Chen, Linda K. Bockenstedt, John F. Anderson, Richard A. Flavell, Michael V. Norgard, Erol Fikrig
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Article Infectious disease

OspC facilitates Borrelia burgdorferi invasion of Ixodes scapularis salivary glands

Abstract

Outer surface protein C (OspC) is a differentially expressed major surface lipoprotein of Borrelia burgdorferi. ospC is swiftly upregulated when spirochetes leave the Ixodes scapularis tick gut, migrate to the salivary gland, and exit the arthropod vector. Here we show that OspC strongly binds to the tick salivary gland, suggesting a role for OspC in spirochete adherence to this tissue. In vivo studies using a murine model of Lyme borreliosis showed that while OspC F(ab)2 fragments did not influence either the viability of spirochetes or ospC gene expression, they did interfere with B. burgdorferi invasion of tick salivary glands. We then generated ospC knockout spirochetes in an infectious clone of B. burgdorferi and examined them within the vector. OspC-deficient or wild-type spirochetes persisted equally within the gut of unfed ticks and multiplied during the tick engorgement; however, unlike wild-type B. burgdorferi, the mutants were unable to invade salivary glands. Salivary gland colonization of OspC-deficient spirochetes was completely restored when this mutant was complemented in trans with a plasmid harboring the wild-type ospC gene. These studies conclusively demonstrate the importance of OspC in the invasion of tick salivary glands by B. burgdorferi, a critical step in the transmission of spirochetes from the arthropod vector to the mammalian host.

Authors

Utpal Pal, Xiaofeng Yang, Manchuan Chen, Linda K. Bockenstedt, John F. Anderson, Richard A. Flavell, Michael V. Norgard, Erol Fikrig

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OspC F(ab)2 fragments interfere with the invasion of B. burgdorferi to t...
OspC F(ab)2 fragments interfere with the invasion of B. burgdorferi to the I. scapularis salivary glands. (a) Distribution of B. burgdorferi within the I. scapularis salivary glands (top row) or gut (bottom row) 72 hours after the onset of feeding. B. burgdorferi–infested nymphal ticks were fed on mice that had been treated with F(ab)2 fragments prepared from either normal rabbit sera (NRS-Fab) or polyclonal OspC sera (OspC-Fab). The spirochetes (arrows) were stained with a FITC-labeled goat anti–B. burgdorferi (shown in green), and the nuclei of the gut epithelial cells were stained with propidium iodide (shown in red). Due to the lower abundance of spirochetes in the salivary gland than in gut, only a few spirochetes can be visualized through a single focal plane of the microscope. Images were recorded at ×400 magnification and are presented as a merged images for clarity (n = 3). (b) Detection of B. burgdorferi mRNA within engorged I. scapularis salivary glands or gut. Nymphs treated with NRS-Fab or OspC-Fab were analyzed by RT-PCR for the detection of viable B. burgdorferi at 48 or 96 hours after the onset of feeding. Equal amounts of total RNA from ticks that fed on F(ab)2-treated mice were converted to cDNA with reverse transcriptase, subjected to PCR with flaB primers, and analyzed on a 1.5% agarose gel. I. scapularis β-actin was used as a control to confirm equal loading of total RNA isolated from infected, fed ticks. An aliquot of prepared RNA from each group was subjected to RT-PCR in the absence of reverse transcriptase to confirm the absence of genomic DNA (data not shown).