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PubMed1996

Hookworm: developmental biology of the infectious process.

Hookworms cause severe anemia and malnutrition in developing countries of the tropics, with an estimated one billion people infected worldwide. An in vitro system that models the early events of infection has provided new information about the linkage between the infectious process and the parasite's developmental biology. The cloning and expression of Ancylostoma secreted protein, ASP 1 - a secreted molecule associated with these developmental processes - is an example of how this system allows us to dissect the infectious process at the molecular level.

Amino Acid Sequence↗

PubMed2001

Electron and light microscopy of neutrophil responses in mice vaccinated and challenged with third-stage infective hookworm (Ancylostoma caninum) larvae.

The role of neutrophils in mediating host inflammation was examined in mice vaccinated with living third-stage infective hookworm larvae (L3). Mice were vaccinated by oral immunization with 500 L3 (Ancylostoma caninum) once every 2 weeks for a total of three immunizations. The vaccinated mice were then challenged intraperitoneally with 2000 L3) 1 week after the final immunization. To stimulate peritoneal production of neutrophils, 2 ml of 2% glycogen were injected intraperitoneally at 16 h prior to the challenge infection. Neutrophils were found to comprise 85% of the peritoneal cell population. L3 from the challenge infection were collected and then examined at timed intervals by inverted light microscopy, scanning electron microscopy (SEM) and transmission electron microscopy (TEM). Greater than a fivefold increase in the total numbers of peritoneal cells was noted in the vaccinated mice as compared to unvaccinated mice. In the peritoneal cavity of vaccinated mice, the neutrophils adhered to the L3 within 2 h, and over 55% of the L3 were surround by clusters of neutrophils to form a sausage-like sheath 4 h later. At 24-72 h after challenge, almost all of the L3 recovered from the vaccinated mice were covered with thick clusters of cells. Both SEM and TEM demonstrated extensive ultrastructural damage to the L3. In contrast, the L3 recovered from the unvaccinated mice appeared to be unaffected by neutrophils. These studies suggest that neutrophils, like macrophages, can have an important role as effector cells in L3-vaccinated mice.

Ancylostoma↗

PubMed1985

The influence of temperature and osmotic stress on the development and eclosion of hookworm eggs.

Time lapse video micro recording techniques have been used to investigate the development and eclosion of Ancylostoma ceylanicum and A. tubaeforme eggs under a range of temperature and osmotic conditions. Over the range 15 to 35 degrees C increasing temperature resulted in decreased development time, the hyperbolic form of the curve being similar to that found by other workers. The adoption of a developmental parameter to mark the start of timing provided more reproducible results than previously reported. Osmotic stress in the range 0-600 mOsM/kg provided by a number of inorganic salts and organic sugars increased hatching time but did not greatly reduce the percentage that hatched. There was no correlation between the ionic composition of the solution and its effect on the eggs. Solutions of EDTA, carbonates and iodides markedly reduced hatching. Behavioural studies of the pre-hatched larvae suggested that the eggs were permeable and susceptible to osmotic and ionic changes early in their development and well before they were ready to hatch. No appreciable differences were found between the two species of hookworm investigated.

Ancylostoma↗

PubMed1996

The anti-haemostatic strategies of the human hookworm Necator americanus.

The human hookworm Necator americanus appears to have evolved a number of complementary strategies to overcome the host's haemostatic processes. These include the inhibition of blood coagulation, platelet aggregation and mediator release, and the secretion of fibrinogenolytic enzymes. These strategies presumably allow the parasite to establish the chronic infections so often documented in human populations.

Animals↗

PubMed1981

Efficacies of vaccines against canine hookworm disease.

Three types of vaccine (1) a sonicated extract of killed larvae, (2) 5-fluorouracil-treated larvae and (3) 40 krad. X-irradiated larvae have been evaluated. Pups aged 3-4 months, weighing 2 +/- 0.5 kg and free of natural hookworm infection were selected for experimental studies. The sonicated extract was given either in a single dose or as 2 doses at weekly intervals, whereas the 5-fluorouracil-treated and X-irradiated larvae were given in a single dose only. After completion of the vaccination schedule, the pups were challenged with filariform larvae and were sacrificed 3 weeks after challenge. Non-vaccinated pups and those vaccinated with a single dose of sonicated extract lost weight and had diarrhoea or dysentery after challenge. Furthermore, in non-vaccinated pups, haemoglobin levels fell and at autopsy the intestinal mucosa was found to be grossly congested and free blood was present in the lumen of the intestine. Such findings were not obtained in pups vaccinated with the other types of vaccines. The percentage of infective larvae which developed to the adult stage in non-vaccinated pups was 82.7 +/- 9.53 compared with 45.1 +/- 12.33, 30.1 +/- 10.05, 20.6 +/- 10.60 and 8.0 +/- 7.46%, respectively in pups vaccinated with a single dose of sonicated extract, double dose of sonicated extract, 5-fluorouracil-treated and 40 krad. X-irradiated larval vaccines. While the irradiated larval vaccine provided maximum protection against clinical ancylostomiasis, 2 doses of the sonicated larval vaccine could also afford almost equal protection to pups.

Ancylostoma↗

PubMed1986

Permeability changes in the egg-shell of hookworms during development and eclosion.

The development and hatching of Ancylostoma ceylanicum and Ancylostoma tubaeforme eggs in a range of potentially inimical solutions has been examined using time-lapse video micro-recording techniques. In all compounds tested the larvae developed to the active pretzel stage. The inception of activity resulted in an increase in shell permeability to water and small molecules. In anthelmintics of molecular weight (Mr) 400 development continued normally until about the time that control eggs hatched, when there was a marked decrease in larval activity and eclosion did not occur. The addition of a low Mr surface-active agent potentiated the effect of the anthelmintic and caused larval activity to be lost earlier. The calcium inhibitor lanthanum chloride affected larval development rather than eclosion. Large Mr compounds did not affect either development or hatching and the results suggest that the egg-shell may act as a form of sieve, allowing the essential oxygen to reach the developing larva while excluding large potentially dangerous molecules. No appreciable differences were found between the two species of hookworm examined.

Acridine Orange↗

PubMed1988

Experimental and field studies on the viability of eggs, and the infectivity of third-stage larvae of the human hookworm Necator americanus recovered from the creek waters of the Niger Delta, Nigeria, and their epidemiological significance.

In a combined field and laboratory study, the eggs and 3rd-stage infective larvae of the human hookworm Necator americanus were recovered from the creek waters of the Niger Delta, Nigeria. They were tested for viability and infectivity. Eggs recovered from fresh water creeks showed a viability or hatching index of between 17.5 and 23.7%. Eggs recovered from brackish water showed a viability index of 18.8-21.4% in 'normal' cultures, and a maximum index of 6.3% when cultured in brackish water. The differences in the viability indices of these eggs were not significant (P greater than 0.05). Infective 3rd-stage larvae were recovered from both fresh creek water (205-258 L3/litre of water), and brackish water (45 L3/litre of water). Larvae hatched from brackish water eggs that were cultured in the brackish water were 20% infective by 3 days, but lost all infective potential by 7 days post-hatching. Larvae from fresh water eggs, cultured in fresh water and 'normal' laboratory cultures reached 50% infectivity in 3-5 days, losing potential infectivity in 11-15 days post-hatching. The infectivity patterns of larvae recovered directly from fresh and brackish waters did not differ significantly (P greater than 0.05) from the infectivity patterns of larvae recovered from laboratory cultures of eggs from all the experimental sources. Although larvae recovered directly from fresh and brackish waters survived in these respective media under laboratory conditions, there was a significant difference (P less than 0.05) in their survival patterns. The survival and infectivity rates of field larvae were considerably reduced over time relative to control larvae.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1995

Ancylostoma caninum anticoagulant peptide: a hookworm-derived inhibitor of human coagulation factor Xa.

Human hookworm infection is a major cause of gastrointestinal blood loss and iron deficiency anemia, affecting up to one billion people in the developing world. These soil-transmitted helminths cause blood loss during attachment to the intestinal mucosa by lacerating capillaries and ingesting extravasated blood. We have isolated the major anticoagulant used by adult worms to facilitate feeding and exacerbate intestinal blood loss. This 8.7-kDa peptide, named the Ancylostoma caninum anticoagulant peptide (AcAP), was purified by using a combination of ion-exchange chromatography, gel-filtration chromatography, and reverse-phase HPLC. N-terminal sequencing of AcAP reveals no homology to any previously identified anticoagulant or protease inhibitor. Single-stage chromogenic assays reveal that AcAP is a highly potent and specific inhibitor of human coagulation, with an intrinsic K*i for the inhibition of free factor Xa of 323.5 pM. In plasma-based clotting time assays, AcAP was more effective at prolonging the prothrombin time than both recombinant hirudin and tick anticoagulant peptide. These data suggest that AcAP, a specific inhibitor of factor Xa, is one of the most potent naturally occurring anticoagulants described to date.

Amino Acid Sequence↗

PubMed1986

Response of laboratory-adapted human hookworm and other nematodes to ivermectin.

The activity of ivermectin was examined in Necator americanus, a human hookworm, adapted to the laboratory hamster. A dose of 30 mg kg-1 X 1 or 10 mg kg-1 X 2 was required for complete clearance of pre-adult N. americanus; however, hamsters carrying adult N. americanus were completely cured of infection by doses of 15 mg kg-1 or 7.5 mg kg-1 X 2. The doses of ivermectin required for complete elimination of N. americanus were much higher than those reported for other intestinal nematodes. The probable reasons for these higher doses are discussed. Tests were also carried out with other rodent parasites, namely Nematospiroides dubius, Strongyloides ratti and Trichuris muris. Doses as low as 0.3 mg kg-1 X 1 completely eliminated adult N. dubius from mice, whereas++ S. ratti needed a repeated dose (0.3 mg kg-1 X 2). None of the mice was cured of T. muris infection even at doses of 10 mg kg-1 X 2, although some degree of cure was apparent at the toxic dose. It thus appears that ivermectin is in no way superior in its activity against N. americanus and T. muris than the existing anthelmintics.

Animals↗

PubMed1983

Secretion of a proteolytic anticoagulant by Ancylostoma hookworms.

Hookworms of the genus Ancylostoma secrete an anticoagulant that both inhibits the clotting of human plasma and promotes fibrin clot dissolution. This anticoagulant activity is attributable to a 36,000 dalton proteolytic enzyme. The protease can degrade fibrinogen into five smaller polypeptides that intrinsically have anticoagulating properties, covert plasminogen to a mini-plasminogen-like molecule, and hydrolyze a synthetic peptide substrate with specificity for elastolytic enzymes. It is hypothesized that the parasite uses this enzyme to prevent blood clotting while feeding on villous capillaries.

Ancylostoma↗

PubMed1992

Isotypic variation in antibody responses in a community in Papua New Guinea to larval and adult antigens during infection, and following reinfection, with the hookworm Necator americanus.

The natural infection of a community with the hookworm Necator americanus induces a vigorous humoral response to both larval and adult parasite antigens. This response occurs in all five human antibody isotypes, and data are presented to show that, at the population level, isotypes respond differently, following chemotherapy and during reinfection, to changes in antigen stimulation. The differential response probably reflects the fact that the parasite, during the course of its life cycle, presents different amounts of antigens at different anatomical locations. It is suggested that IgG and IgM responses against adult excretory-secretory (ES) products most accurately reflect the efficacy of chemotherapy, and the load of resident adult infection, while IgG responses against larval somatic antigens reflect continuous exposure to infection. These hypotheses should now be tested, at the level of the individual, in a longitudinal manner using more closely spaced sampling intervals. This repetitive sampling, and the inclusion of a measure of the exposure of the population to infective stages, will allow more definitive conclusions to be made about the role of the immune response in controlling worm burdens.

Adolescent↗

PubMed2004

Impact of antihelminthic treatment on infection by Ascaris lumbricoides, Trichuris trichiura and hookworms in Covas, a rural community of Pernambuco, Brazil.

This work aims to evaluate the impact of drug treatment on infection by Ascaris lumbricoides (Al), Trichuris trichiura (Tt) and hookworms (Hook) in a rural community from the sugar-cane zone of Pernambuco, Brazil. Four parasitological surveys were carried out from March 2001 to March 2002. Individual diagnosis was based on eight slides (four by the Kato-Katz method and four by the Hoffman method) per survey. Infected subjects were assigned to two groups for treatment with either albendazole (n = 62) or mebendazole (n = 57). Prevalence of infection fell significantly (p < 0.05) one month after treatment: Al (from 47.7% to 6.6%); Tt (from 45.7% to 31.8%) and Hook (from 47.7% to 24.5%). One year after treatment, infections by Tt and Hook remained significantly below pre-control levels. A substantial decrease in single-infection cases and multiple infections was found. Egg-negative rate was significant for Al (94.0%), Hook (68.3%) but not for Tt (45.5%), and did not differ significantly between subjects treated with mebendazole or albendazole. Egg counts fell significantly in the individuals remaining positive for Tt. It is recommended that antihelminthic treatment should be selective and given at yearly intervals preferably with albendazole, due to its cost-effectiveness.

Adolescent↗

PubMed1966

Treatment of hookworm infection with 2, 6-Diiodo-4-Nitrophenol.

Single dose(1.0 cc) of 20 % 2, 6-diiodo-4-nitrophenol solution was injected to 82 hookworm infected cases and 79% of them became egg-negative within 2-3 weeks after the injection. Mild to severe local pain were experienced for several minutes in all cases. Two cases complained of paralysis of the arms of the same side. The tablet of the same formula was given orally to 21 cases for 1-4 days. The 2-4 day course with 360 mg/day were enough to eliminate the parasite, and showed no systemic side effects.

Journal Article↗

PubMed2000

Eotaxin is specifically cleaved by hookworm metalloproteases preventing its action in vitro and in vivo.

Eotaxin is a potent eosinophil chemoattractant that acts selectively through CCR3, which is expressed on eosinophils, basophils, mast cells, and Th2-type T cells. This arm of the immune system is believed to have evolved to control helminthic parasites. We hypothesized that helminths may employ mechanisms to inhibit eosinophil recruitment, to prolong worm survival in the host. We observed that the excretory/secretory products of the hookworm Necator americanus inhibited eosinophil recruitment in vivo in response to eotaxin, but not leukotriene B(4), a phenomenon that could be prevented by the addition of protease inhibitors. Using Western blotting, N. americanus supernatant was shown to cause rapid proteolysis of eotaxin, but not IL-8 or eotaxin-2. N. americanus homogenate was fractionated by gel filtration chromatography, and a FACS-based bioassay measured the ability of each fraction to inhibit the activity of a variety of chemokines. This resulted in two peaks of eotaxin-degrading activity, corresponding to approximately 15 and 50 kDa molecular mass. This activity was specific for eotaxin, as responses to other agonists tested were unaffected. Proteolysis of eotaxin was prevented by EDTA and phenanthroline, indicating that metalloprotease activity was involved. Production of enzymes inactivating eotaxin may be a strategy employed by helminths to prevent recruitment and activation of eosinophils at the site of infection. As such this represents a novel mechanism of regulation of chemokine function in vivo. The existence of CCR3 ligands other than eotaxin (e.g., eotaxin-2) may reflect the evolution of host counter measures to parasite defense systems.

Animals↗

PubMed2004

A secreted protein from the human hookworm necator americanus binds selectively to NK cells and induces IFN-gamma production.

Parasitic helminths induce chronic infections in their hosts although, with most human helminthiases, protective immunity gradually develops with age or exposure of the host. One exception is infection with the human hookworm, Necator americanus, where virtually no protection ensues over time. Such observations suggest these parasites have developed unique mechanisms to evade host immunity, leading us to investigate the role of the excretory/secretory (ES) products of adult N. americanus in manipulating host immune responses. Specifically, we found that a protein(s) from ES products of adult N. americanus bound selectively to mouse and human NK cells. Moreover, incubation of purified NK cells with N. americanus ES products stimulated the production of augmented (4- to 30-fold) levels of IFN-gamma. This augmentation was dependent on the presence of both IL-2 and IL-12 and was endotoxin-independent. This is the first report of a pathogen protein that binds exclusively to NK cells and the first report of a nematode-derived product that induces abundant levels of cytokines from NK cells. Such an interaction could provide a means of cross-regulating deleterious Th2 immune responses in the host, thereby contributing to the long-term survival of N. americanus.

Animals↗

PubMed1987

The clinical and immunologic responses of normal human volunteers to low dose hookworm (Necator americanus) infection.

Five normal human volunteers were exposed to approximately 50 infective larvae of Necator americanus and were observed for the development of clinical signs or symptoms and for changes in blood eosinophil levels, IgG antibody titers, total and parasite-specific IgE, and lymphocyte blastogenic responses for 6-10 weeks. Bronchoalveolar lavage was performed on four subjects prior to infection and at times when larval migration through the pulmonary tree was likely. Eggs were demonstrated in the stools of four volunteers who remained untreated for more than 6 weeks; one volunteer had to be treated at day 40 because of severe gastrointestinal symptoms. All others also complained of abdominal pain and flatulence between days 35-40. All volunteers developed marked blood eosinophilia which peaked between days 38-64 and ranged from 1,350-3,828 eosinophils/mm3. Small increases in total and parasite-specific IgE and IgG were noted in some volunteers. One volunteer showed a significant lymphocyte blastogenic response. With the exception of mucosal erythema, bronchoalveolar lavage results were unremarkable. Our data indicate that a single small inoculum of hookworm larvae is capable of producing significant transient gastrointestinal morbidity and marked blood eosinophilia but does not induce other prominent T cell- and B cell-dependent immune responses.

Adult↗

PubMed1994

Secretion of cysteine proteinase activity by the zoonotic hookworm Ancylostoma caninum.

The zoonotic hookworm, Ancylostoma caninum, probably induces human eosinophilic enteritis by inducing allergic responses to its secretions. This species is already known to secrete metalloproteinases, but in other parasites, cysteine proteinases are involved in pathogenesis. We studied somatic extracts of A. caninum adults and infective larvae and adult excretory/secretory (ES) antigens for cysteine proteinase activity using fluorogenic peptide substrates and by gelatin and fluorogenic substrate polyacrylamide gel electrophoresis. Proteolytic activity was observed against the cathepsins L and B-specific substrate Z-phe-arg-AMC, against the plasmin substrate Boc-val-leu-lys-AMC, and against gelatin. The Z-phe-arg-AMC-hydrolyzing activity in ES antigens and in adult extracts was enhanced up to 15-fold by the reducing agent dithiothreitol (DTT), but was totally blocked by specific inhibitors of cysteine proteinases, including the peptidyl diazomethyl ketone Z-phe-ala-CHN2,E-64, leupeptin, and N-ethylmaleimide. In a similar fashion, gelatinolytic activity in ES antigens detected using substrate gels was enhanced by the addition of reducing agents and inhibited by Z-phe-ala-CHN2 and E-64. The DTT-enhanced, Z-phe-arg-AMC-hydrolyzing activity in ES antigens was active over a wide pH range (pH 5-9). Similar cysteine proteinase activity to that detected in ES antigens was present in extracts of adult and infective larvae of A. caninum. Because the substrate Z-phe-arg-AMC was specifically hydrolyzed, and because this hydrolysis was totally blocked by cysteine proteinase-specific inhibitors, ES antigens and tissue extracts of A. caninum clearly possess cysteine proteinase activity.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗

PubMed1994

Pathogenicity of the hookworm, Ancylostoma pluridentatum, in a Florida panther (Felis concolor coryi) kitten.

We evaluated clinical signs and administered anthelmintic treatment to a wild-caught, captive Florida panther (Felis concolor coryi) kitten from Big Cypress National Preserve, Florida (USA) infected with the hookworm, Ancylostoma pluridentatum. Clinical signs observed included poor body condition, lethargy, and below normal red blood cell numbers, hemoglobin concentration, and packed cell volume, and elevated eosinophil numbers. In addition, a maximum of 936 Ancylostoma sp. eggs/g of feces were found on Day 11 of captivity. Following oral administration of 20 mg/kg pyrantel pamoate on Day 11, 26 A. pluridentatum were collected from the feces. Based on the resolution of clinical signs, cessation of egg shedding, and a return to normal hematologic values following anthelmintic treatment, we believe that infection with A. pluridentatum was the primary cause of the stressed conditions in the panther kitten.

Administration, Oral↗