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Background: Ixodes scapularis saliva enables the transmission of infectious agents to the mammalian host due to its immunomodulatory, anesthetic and anti-coagulant properties. However, how I. scapularis saliva influences host cytokine secretion in the presence of the obligate intracellular rickettsial pathogen Anaplasma phagocytophilum remains elusive., Methods: Bone marrow derived macrophages (BMDMs) were stimulated with pathogen associated molecular patterns (PAMPs) and A. phagocytophilum. Cytokine secretion was measured in the presence and absence of I. scapularis saliva. Human peripheral blood mononuclear cells (PBMCs) were also stimulated with Tumor Necrosis Factor (TNF)-α in the presence and absence of I. scapularis saliva and interleukin (IL)-8 was measured., Results: I. scapularis saliva inhibits inflammatory cytokine secretion by macrophages during stimulation of Toll-like (TLR) and Nod-like receptor (NLR) signaling pathways. The effect of I. scapularis saliva on immune cells is not restricted to murine macrophages because decreasing levels of interleukin (IL)-8 were observed after TNF-α stimulation of human peripheral blood mononuclear cells. I. scapularis saliva also mitigates pro-inflammatory cytokine response by murine macrophages during challenge with A. phagocytophilum., Conclusions: These findings suggest that I. scapularis may inhibit inflammatory cytokine secretion during rickettsial transmission at the vector-host interface.

The effects of repeated infestations with Dermacentor andersoni nymphs on the lymphocyte functions of BALB/c mice were investigated. The in-vitro proliferation responses to concanavalin-A or salivary-gland molecules, the production of cytokines, and the expression of two adhesion molecules-leucocyte function-associated antigen-1 (LFA-1) and very late activation-4 (VLA-4)-were all studied. In addition, the ability of salivary-gland extract or saliva from D. andersoni to modulate expression of lymphocyte adhesion molecules in vitro was determined. The proliferative responses of T-lymphocytes to concanavalin-A were significantly suppressed after first and second infestations, and significant increases in lymphocyte proliferation in the presence of tick salivary-gland antigen were observed in infested mice. After two infestations, production of interleukin-2 was significantly decreased but that of interferon-gamma remained unchanged. Production of interleukin-4 and interleukin-10 was significantly enhanced in infested mice after both the first and second infestations. Expression of LFA-1 and VLA-4 by lymphocytes from infested mice was suppressed. Furthermore, both a salivary-gland extract and the saliva of D. andersoni reduced the in-vitro expression of both of these adhesion molecules by lymphocytes from tick-naive mice.

Lyme borreliosis, caused by Borrelia burgdorferi, is transmitted by Ixodes scapularis in the eastern and midwestern United States and by Ixodes pacificus in the far-Western United States. Studies have shown that infestation with I. scapularis nymphs modulates host cytokine production; however, the influence of I. pacificus infestation on host cytokines remains uninvestigated. This study demonstrated how repeated infestations with pathogen-free I. scapularis or I. pacificus nymphs affects the production of the macrophage cytokines IL-1beta, IL-6 and tumour necrosis factor-alpha and the T lymphocyte cytokines IL-2, IL-4, IL-5, IL-6, IL-10 and IFN-gamma by C3H/HeN mice. Female mice were infested once or twice with pathogen-free I. scapularis or I. pacificus nymphs, with a 14-day tick-free period between exposures. After each infestation, tick biology parameters were assessed and macrophage and T lymphocyte cytokine production measured by antigen capture ELISA. Acquired resistance to tick feeding did not develop after infestation with either tick species. Differences in cytokine production were observed between infested and noninfested mice, and between mice infested with either I. scapularis or I. pacificus nymphs. Infestations polarized cytokine production towards a Th2 cytokine profile, with suppression of pro-inflammatory Th1 cytokines. This pattern of cytokine production is more pronounced for I. pacificus infested mice.

Schoeler, G. B., Manweiler, S. A., and Wikel, S. K. 1999. Ixodes scapularis: Effects of repeated infestations with pathogen-free nymphs on macrophage and T lymphocyte cytokine responses of BALB/c and C3H/HeN mice. Experimental Parasitology 92, 239-248. Ixodes scapularis is the principal vector in the United States of Borrelia burgdorferi, the causative agent of Lyme borreliosis, the human granulocytic ehrichiosis agent, and Babesia microti. Infestation with I. scapularis nymphs has previously been shown to modulate host T lymphocyte cytokine production. Tick-induced host immunomodulation is increasingly recognized as a contributing factor in successful transmission and/or establishment of tick-borne pathogens. This study was conducted to determine the effects of repeated infestations with pathogen-free I. scapularis nymphs on the production of the macrophage cytokines interleukin (IL)-1beta and tumor necrosis factor-alpha and the T lymphocyte cytokines IL-2, IL-4, IL-10, and interferon-gamma in both BALB/c and C3H/HeN mice. The pattern of T lymphocyte cytokine production was evaluated to determine if repeated tick infestation polarizes the immune response toward a Th-1 or Th-2 cytokine profile. Female BALB/c and C3H/HeN mice were infested one to four times with pathogen-free I. scapularis nymphs, with a 14-day tick-free period between each exposure. After each infestation, tick biology parameters were measured and macrophage and T lymphocyte cytokine production was assessed. Elaboration of T lymphocyte and macrophage cytokines was quantitated by antigen capture enzyme-linked immunosorbent assay. Acquired resistance to I. scapularis feeding was not developed by either mouse strain. Significant differences in cytokine production were observed between infested and noninfested mice, as well as between the two mouse strains, following tick infestation. Infestation of both strains with pathogen-free I. scapularis results in a polarization of the host immune response toward a Th-2, anti-inflammatory pattern, with a corresponding suppression of Th-1 responses., (Copyright 1999 Academic Press.)

Ixodid tick infestation induces host acquired resistance, which involves immunoglobulin cell-mediated and complement-dependent effector pathways. Ticks have developed countermeasures to modulate host antiarthropod responses. Ixodid-mediated host immunomodulation results in vitro in reduced responsiveness to T-lymphocyte mitogens for cells obtained from infested hosts and impaired antibody responses to a thymic dependent antigen. Salivary gland extracts from days 0-9 of engorgement from unmated, female Dermacentor andersoni Stiles suppressed lymphocyte proliferative responses (LPS) to the T-cell mitogen Con A up to 68.4%, whereas responsiveness to E. coli LPS was enhanced. Cytokines assessed in this study included interleukin-1, IL-1, and tumor necrosis factor (TNF) alpha produced by macrophages, and interleukin-2, IL-2, and gamma interferon (IFN-G) secreted by T-lymphocytes. Salivary gland extracts prepared from tissues obtained on days 0-5 of engorgement suppressed IL-1 elaboration from 89.8% on day 0 through 37.5% on day 6. Levels of TNF were reduced from 40.7 to 94.6% throughout the course of the study. Production of IL-2 was suppressed by 14.1-31.9%, and IFN-G was reduced by 8.7-57.0%. Reduced IL-1 levels during the early phases of feeding indicated reduced host ability to activate T-lymphocytes and provide costimulatory, differentiation, and development signals for B-cells. Both IL-1 and TNF are endogenous pyrogens and activate polymorphonuclear leukocytes. Activities of TNF and IFN-G include antiviral properties and induction of expression of class I and II major histocompatibility complex molecules, which are critical components in the recognition of antigen by T-lymphocytes. The autocrine role of IL-2 in proliferation of T-lymphocytes is central to the development of immune reactivity involving T-cell regulation or effector functions or both. Reductions in cytokine levels would suppress immune responses directed toward immunogens introduced into the host during the course of tick feeding. These results indicates that immunomodulation of the host during tick feeding facilitates engorgement and pathogen transmission.