Effect of Vaccinations With Recombinant Fusion Proteins on Ancylostoma Caninum Habitat Selection in the Canine Intestine

Abstract

Laboratory dogs were vaccinated subcutaneously with 3 different recombinant fusion proteins, each precipitated with alum or calcium phosphate. The vaccinated dogs were then challenged orally with 400 third-stage infective larvae (L3) of the canine hookworm, Ancylostoma caninum. The 3 A. caninum antigens selected were Ac-TMP, an adult-specific secreted tissue inhibitor of metalloproteases; Ac-AP, an adult-specific secreted factor Xa serine protease inhibitor anticoagulant; and Ac-ARR-1, a cathepsin D-like aspartic protease. Each of the 3 groups comprised 6 male beagles (8 +/- 1 wk of age). A fourth group comprised control dogs injected with alum. All of the dogs vaccinated with Ac-TMP or Ac-APR-1 exhibited a vigorous antigen-specific antibody response, whereas only a single dog vaccinated with Ac-AP developed an antibody response. Dogs with circulating antibody responses exhibited 4.5-18% reduction in the numbers of adult hookworms recovered from the small intestines at necropsy, relative to alum-injected dogs. In contrast, there was a concomitant increase in the number of adult hookworms recovered from the colon. The increase in colonic hookworms was as high as 500%, relative to alum-injected dogs. Female adult hookworms were more likely to migrate into the colon than were males. Anti-enzyme and anti-enzyme inhibitor antibodies correlated with an alteration in adult hookworm habitat selection in the canine gastroinntestinal tract.

Binding of Excreted and/or Secreted Products of Adult Hookworms to Human NK Cells inNecator americanus-Infected Individuals from Brazil

“…N. americanus was maintained in hamsters (32), and Ancylostoma caninum was maintained in experimental beagles (18). Adult worms were removed from the intestines of euthanized animals, washed in phosphate-buffered saline (PBS), and then cultured overnight in RPMI 1640 containing 100 U/ml penicillin G sodium, 100 g/ml streptomycin sulfate, and 0.25 g/ml amphotericin B (all reagents from SigmaAldrich, St. Louis, MO).…”

Ancylostoma caninum MTP-1, an Astacin-Like Metalloprotease Secreted by Infective Hookworm Larvae, Is Involved in Tissue Migration

“…A purpose-bred, parasite-naïve male beagle aged 8 weeks was purchased from Marshall Farms (North Rose, NY) and maintained in the George Washington University Animal Research Facility as previously described (12). The experiments were conducted through an approved protocol by the George Washington University Animal Care and Use Committee.…”

“…Before the first vaccination and after each subsequent one, a serum sample was obtained. Ac-MTP-1 (100 g) was formulated with the adjuvant AS03 as described elsewhere (16), and the dog was immunized intramuscularly three times beginning at age 62 days with Ac-MTP-1 and adjuvant as previously described (12). The dog was given boosters at 21-day intervals.…”

“…Blood was drawn once every 21 days, and serum was separated from cells by centrifugation. Enzyme-linked immunosorbent assays were performed as previously described (12). Recombinant Ac-MTP-1 was used to coat microtiter plates at a concentration of 5.0 g ml Ϫ1 .…”

Biochemical Characterization and Vaccine Potential of a Heme-Binding Glutathione Transferase from the Adult Hookworm Ancylostoma caninum

“…Inclusion criteria for enrollment included body weight equal to or exceeding 2.5 kg, hematocrit equal to or exceeding 28.0, and white blood cell count not exceeding 20 ϫ 10 6 /ml. All dogs in a trial were purchased from the same vendor (Harlan Farms) and maintained in the George Washington University (GWU) Animal Research Facility as previously described (20). An initial screening fecal examination was conducted shortly after the arrival of the dogs at the GWU Animal Research Facility to confirm the absence of canine hookworm infection.…”