Hookworms are some of the most widespread of the soil-transmitted helminths (STH) with an estimated 438.9 million people infected. Until relatively recently Ancylostoma ceylanicum was regarded as a rare cause of hookworm infection in humans, with little public health relevance. However, recent advances in molecular diagnostics have revealed a much higher prevalence of this zoonotic hookworm than previously thought, particularly in Asia. This study examined the prevalence of STH and A. ceylanicum in the municipalities of Palapag and Laoang in the Philippines utilizing real-time polymerase chain reaction (PCR) on stool samples previously collected as part of a cross-sectional survey of schistosomiasis japonica. Prevalence of hookworm in humans was high with 52.8% (n = 228/432) individuals positive for any hookworm, 34.5% (n = 149/432) infected with Necator americanus, and 29.6% (n = 128/432) with Ancylostoma spp; of these, 34 were PCR-positive for A. ceylanicum. Considering dogs, 12 (n = 33) were PCR-positive for A. ceylanicum. This is the first study to utilize molecular diagnostics to identify A. ceylanicum in the Philippines with both humans and dogs infected. Control and elimination of this zoonotic hookworm will require a multifaceted approach including chemotherapy of humans, identification of animal reservoirs, improvements in health infrastructure, and health education to help prevent infection.

Caenorhabditis elegans is a free-living nematode that resides in soil and typically feeds on bacteria. We postulate that haematophagic C. elegans could provide a model to evaluate vaccine responses to intestinal proteins from hematophagous nematode parasites, such as Necator americanus. Human erythrocytes, fluorescently labelled with tetramethylrhodamine succinimidyl ester, demonstrated a stable bright emission and facilitated visualization of feeding events with fluorescent microscopy. C. elegans were observed feeding on erythrocytes and were shown to rupture red blood cells upon capture to release and ingest their contents. In addition, C. elegans survived equally on a diet of erythrocytes. There was no statistically significant difference in survival when compared with a diet of Escherichia coli OP50. The enzymes responsible for the digestion and detoxification of haem and haemoglobin, which are key components of the hookworm vaccine, were found in the C. elegans intestine. These findings support our postulate that free-living nematodes could provide a model for the assessment of neutralizing antibodies to current and future hematophagous parasite vaccine candidates.

Despite the profound health implications of Necator americanus infection in humans, many aspects of its interaction with the host immune system are poorly understood. Here we investigated the early events at the interface of N. americanus larvae (L3) and human dendritic cells (DCs). Our data show that co-culturing DCs and the larvae trigger ex-sheathing of hookworms rapidly where a majority of DCs are sequestered onto the larval sheath allowing the ex-sheathed larvae to migrate away unchallenged. Intriguingly, DCs show negligible interaction with the ex-sheathed larvae, alluding to differences between the surface chemistry of the larva and its sheath. Furthermore, blocking of two key C-type lectin receptors on DC surface (i.e. DC-SIGN and mannose receptor) resulted in inhibition of ex-sheathing process and DC sequestration, highlighting the importance of C-type lectins on DCs in the induction of the ex-sheathing. Analyses of DC phenotype and cytokine profile after co-culture with the N. americanus larvae showed an immature phenotype as evidenced by the low expression of the maturation markers and cytokines. These data provide new insights into early events at the interface of human DCs and N. americanus larvae and could explain how L3 evade immune recognition upon initial interaction with DCs.

This study quantifies the rate and intensity of re-infection with human hookworm and Schistosoma mansoni infection 12 months following successful treatment, and investigates the influence of socio-economic, geographical and environmental factors. A longitudinal study of 642 individuals aged over 5 years was conducted in Minas Gerais State, Brazil from June 2004 to March 2006. Risk factors were assessed using interval censored regression for the rate and negative binomial regression for intensity. The crude rate and intensity of hookworm re-infection was 0·21 per year (95% confidence interval (CI) 0·15–0·29) and 70·9 epg (95% CI 47·2–106·6). For S. mansoni the rate was 0·06 per year (95% CI 0·03–0·10) and intensity 6·51 epg (95% CI 3·82–11·11). Rate and intensity of re-infection with hookworm were highest among males and positively associated with previous infection status, absence of a toilet and house structure. Rate and intensity of S. mansoni re-infection were associated with previous infection status as well as geographical, environmental and socio-economic factors. The implications of findings for the design of anti-helminth vaccine trials are discussed.

The relationships between the number of infective larvae (L3s) to which animals were exposed and the establishment of A. ceylanicum and N. americanus in hamsters were examined. There was no evidence of density-dependent constraints on the establishment of A. ceylanicum in the range 10–1000 larvae, but an experiment in which the range was extended to 1500 L3s gave a significant negative correlation between the magnitude of the infecting dose and percentage establishment. Even so the percentage reduction was relatively small, approximating to 0·89% per 100 larvae administered, and in practice density-dependent constraints on establishment are unlikely to affect experiments with this species in which much lower doses (<250 L3) are generally employed. The range of doses for N. americanus L3s was smaller (10–400). Of the four experiments reported, two gave a significant reduction of establishment with increasing dose and two did not. When the data was split into low doses (<100 L3s) and high doses (>100 L3s), falling establishment with increasing dose was only detected in the lower dose range. There was no difference in the establishment when doses of 100 L3s were compared with 250 or 400 L3s. On balance, it was concluded that density-dependent constraints on establishment of N. americanus in hamsters were not marked and would have little significant effect on experiments utilizing fewer than 200 L3s (approximately 7·6% reduction between 10 and 200 L3s). These results are discussed in relation to host regulation of hookworm burdens.

An attempt was made to identify the human hookworm involved in failed-treatment cases using abnormal hosts and scanning electron microscopy. Thirty-seven, 2 to 6 month old Chinese hamsters (Cricetulus griseus) from a closed, outbred, conventional colony, were each given between 20 and 120 filariform larvae per os. The larvae were cultured from faeces from mebendazole (Vermox®) 500 mg single-dose, failed-treatment cases living in the lowveld farming area of the Transvaal Province, South Africa. About 60 to 78 days after inoculation, the animals were killed and adult worms were removed from their small intestines. Eleven (30%) of the 37 hamsters harboured a total of 31 adult worms (19 males and 12 females), while 26 hamsters were refractory to infection. The greatest number of worms recovered from a single animal was six. A total of 27 worms (17 males and 10 females) were subjected to examination by scanning electron microscopy. Micrographs showed male and female worms to be morphologically all of the Necator americanus species, as identified by a pair of ventral and dorsal cutting plates, a dorsal tooth and the fused terminus of spicules in the male bursa. The transverse cuticular striations were distinct and smooth. Several points of interest arose from the results of this study and are discussed.

A simple method for the collection of third-stage larvae of Necator americanus has been described. This technique provides repeated recovery of very clean larvae from cultures in moderate numbers.

Laboratory bred DSN hamsters were exposed to varying doses of infective larvae of Ancylostoma ceylanicum (orally) or Necator americanus (percutaneously) and were autopsied at times which corresponded to a period immediately before cessation of growth of worms or soon afterwards. A total of 829 (404 male and 425 female) A. ceylanicum and 1582 (781 male and 801 female) N. americanus were measured. At worm burdens of fewer than 100, the length of A. ceylanicum appeared to increase with infection intensity and no evidence was found that growth was retarded under crowded conditions. In an experiment comparing directly low (mean worm burden = 22) and heavy infections (mean worm burden = 180) significant negative associations between both weight and width, and worm burden were detected, but length again increased with worm burden. In contrast, 5 experiments with N. americanus indicated negative relationships between measures of worm size (length, width, wet and dry weight) and worm burden. It was concluded that N. americanus is subject to regulation by density-dependent processes within the host while A. ceylanicum is not sensitive to the same degree.

Reinfection with hookworm (Necator americanus) following chemotherapy was studied over 2 years in a rural village in Madang Province, Papua New Guinea. The prevalence of hookworm infection had returned to pre-treatment levels after 2 years, and the geometric mean hookworm burden had returned to 58 % of the pre-treatment value. The rate of acquisition of adult worms was independent of host age, and was estimated as a geometric mean of 2·9–3·3 worms/host/year (arithmetic mean 7·9–8·9 worms/host/year). There was significant predisposition to hookworm infection; the strength of this predisposition did not vary significantly between age or sex classes.

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.

The proteolytic activities present in adult Necator americanus excretory–secretory products have been assessed using biologically relevant, naturally occurring substrates (haemoglobin and fibrinogen) and a number of synthetic fluorogenic and chromogenic substrates. One broad peak of activity was observed against haemoglobin in the pH range 5 to 7, with maximum activity at pH 6·6, while fibrinogenolytic activity was shown to be greater at pH 3·5. Inhibition studies against haemoglobin, fibrinogen and synthetic substrates using a battery of appropriate protease inhibitors indicated the presence of a mixture of aspartyl, cysteinyl and serine proteases. Metal ion (Ca2+, Zn2+ and Fe2+) stimulation was demonstrated, with stimulation by Zn2+ being the most marked. These results are discussed in the context of recent developments in the field of parasite proteolytic enzymes, where they have been suggested as targets for immuno- and chemotherapy.

Neonatal hamsters were exposed to varying doses of Necator americanus larvae and changes in the stability of the resulting worm burdens were monitored over a period of 25 weeks. No change in worm burdens was evident for the first 5 weeks of infection, irrespective of the infection intensity, but the more heavily infected groups subsequently lost worms in a density-dependent manner. Male and female hamsters lost comparable proportions of their established parasite burdens indicating that there was no host sex-linked difference in this respect. By week 15 infections had stabilized and the residual worm burdens, usually a maximum of 30 worms survived for a considerably longer period of time. Initially the percentage of male worms varied from 45% to 50% but as infection progressed male worms comprised a significantly increasing proportion of the total worm burden. By week 25 the percentage of male worms was usually in excess of 60%. The growth of infected animals was not severely affected by N. americanus, even when heavy worm burdens established initially, but a significant effect was detected particularly in week 5, prior to worm loss, when the adult worms would have been feeding on intestinal tissues and causing blood loss for a period of about 2 weeks. The most severe depression in the packed cell volume was also recorded in week 5, indicating that anaemia had been initiated in infected hamsters. Whilst, the regulation of parasite burdens in weeks 5–10 post-infection may have resulted from host immunity, the persistence of the residual worm burdens, the marked density-dependent anaemia and the subtle effect on host weight, all reflected well-documented aspects of chronic human necatoriasis.

The course of a primary Necator americanus infection was studied in the lungs and small intestines of syngeneic mice. Following percutaneous infection no difference in initial larval establishment in the lungs was found between male BALB/c, NIH or B10.G mice. However, significant differences in the subsequent kinetics of infection were demonstrated between the BALB/c and NIH strains. Lung worm burdens declined more slowly in NIH mice than in BALB/c strain. Surprisingly, however, a greater proportion of larvae remaining in the lungs of BALB/c mice, 9 days p.i., were trapped than in NIH mice. Nevertheless, establishment in the small intestines of the BALB/c strain was consistently greater than in NIH mice. Host immunosuppression resulted in increased larval retention in the lungs of both the BALB/c and NIH strains as well as in the small intestines of BALB/c mice. Treatment with hydrocortisone acetate did not increase intestinal worm burdens in NIH mice. The data presented suggest that, in this complex, dynamic model system, designation of ‘susceptible’ and ‘resistant’ strains is inappropriate. The factors underlying the observed strain differences in resistance to infection are discussed.

Baseline data from an immuno-epidemiological study of hookworm infection in a rural village in Madang Province, Papua New Guinea are reported. Necator americanus was found to be the commonest helminth infection, with a prevalence of near 100% and intensity of 40 worms per host in adults. Enterobius vermicularis, Ascaris lumbricoides and Trichuris trichiura were also present, at prevalences of 53, 10 and 3% respectively; Ancylostoma duodenale was absent. The frequency distribution of N. americanus was highly over-dispersed, and was well described by a negative binomial distribution with aggregation parameter, k, of 0·370. Intensity of infection was significantly related to host age, but did not differ between the sexes. Haemoglobin levels and haematocrit values were indicative of anaemia in the community, but were unrelated to hookworm infection. Levels of antibodies (IgG, IgA and 1gM combined) against adult Necator cuticular collagen and excretory-secretory (ES) products were determined.

The development and mortality of the eggs of Ancylostoma duodenale and Necator americanus in distilled water were monitored over a range of temperatures between 15 and 35°C. Egg demography was examined within the context of a fourparameter mathematical model of development and mortality. Over the range of temperatures studied, egg mortality (μ) was an increasing exponential function of temperature (T) measured in degrees Celsius. A single model adequately described the mortality of both species (ln [μ] = 0·041*T − 6·87). The minimum time (τ) to hatching was consistently less for A. duodenale (ln [1/T] = −(0·79+53·05*[l/T]) than N. americanus (ln [1/T] = −(0·99 + 53·05*[l/T])). The hatching rate [σ when t > τ) was an increasing function of time and temperature in both cases, but the precise functional relationship was species specific.

The feeding behaviour of parasitic 3rd-stage larvae (L3) of the hookworms Ancylostoma caninum, A. ceylanicum and Necator americanus was examined. Less than 11% of A. caninum L3 recovered from the small intestines of dogs infected orally were feeding at 4–48 h post-infection (p.i.). and none of the A. ceylanicum L3 recovered from the intestines of orally infected hamsters had resumed feeding. All L4 of both species recovered at 36 and 48 h p.i. had resumed feeding. On the other hand, approximately 16% of the A. ceylanicum L3 recovered from the skin of percutaneously infected hamsters at 18 h were feeding, and the percentage feeding increased to nearly 58% at 44 h p.i. Necator americanus L3 recovered from the skin of percutaneously infected neonatal hamsters resumed feeding at 6–12 h p.i. and reached 90–94% by 18 h. Feeding began to decline at 66 h, and reached 29% at 120 h p.i. This decrease was associated with the migration of larvae from the skin to the lungs. By 192 h p.i. over 95% of the larvae had reached the small intestine, and all had moulted to the L4. The results indicate that parasitic L3 resume feeding in the skin during percutaneous infections, and suggest that feeding by hookworm L3 correlates with the resumption of development.

This comprehensive review briefly describes the history and pharmacology of albendazole as an anthelminthic drug and presents detailed summaries of the efficacy and safety of albendazole's use as an anthelminthic in humans. Cure rates and % egg reduction rates are presented from studies published through March 1998 both for the recommended single dose of 400mg for hookworm (separately for Necator americanus and Ancylostoma duodenale when possible), Ascaris lumbricoides, Trichuris trichiura, and Enterobius vermicularis and, in separate tables, for doses other than a single dose of 400mg. Overall cure rates are also presented separately for studies involving only children 2–15 years. Similar tables are also provided for the recommended dose of 400mg per day for 3 days in Strongyloides stercoralis, Taenia spp. and Hymenolepis nana infections and separately for other dose regimens. The remarkable safety record involving more than several hundred million patient exposures over a 20 year period is also documented, both with data on adverse experiences occurring in clinical trials and with those in the published literature and/or spontaneously reported to the company. The incidence of side effects reported in the published literature is very low, with only gastrointestinal side effects occurring with an overall frequency of just >1%. Albendazole's unique broad-spectrum activity is exemplified in the overall cure rates calculated from studies employing the recommended doses for hookworm (78% in 68 studies: 92% for A. duodenale in 23 studies and 75% for N. americanus in 30 studies), A. lumbricoides (95% in 64 studies), T. trichiura (48% in 57 studies), E. vermicularis (98% in 27 studies), S. stercoralis (62% in 19 studies), H. nana (68% in 11 studies), and Taenia spp. (85% in 7 studies). The facts that albendazole is safe and easy to administer, both in treatment of individuals and in treatment of whole communities where it has been given by paramedical and nonmedical personnel, have enabled its use to improve general community health, including the improved nutrition and development of children.

Hookworms are gut-dwelling, blood-feeding nematodes that infect hundreds of millions of people, particularly in the tropics. As part of a program aiming to define novel drug targets and vaccine candidates for human parasitic nematodes, genes expressed in adults of the human hookworm Necator americanus were surveyed by the expressed sequence tag approach. In total 161 new hookworm genes were identified. For the majority of these, a function could be assigned by homology. The dataset includes proteases, protease inhibitors, a lipid binding protein, C-type lectins, an anti-bacterial factor, globins and other genes of interest from a drug or vaccine development viewpoint. Three different classes of small, secreted proteins were identified that may be involved in the host–parasite interaction, including potential potassium channel blocking peptides. One third of the genes were novel. These included highly expressed, secreted (glyco)proteins which may be part of the excretory–secretory products of these important pathogens. Of particular interest are a family of 9 genes with similarity to the immunomodulatory protein, neutrophil inhibitory factor, that may play a role in establishing an immunocompromised niche for this successful parasite.

The blood and broncho-alveolar leucocyte (BAL) responses, to a primary Necator americanus infection, were studied in male BALB/c and NIH mice. Following percutaneous infection, a significant blood leucocytosis occurred in both the BALB/c and NIH strains. The peak response occurred, in both strains, on day 10 p.i. and this was reflected in the eosinophil response which peaked at the same time-point. The change in blood eosinophil numbers, as a result of infection, was found to be the greatest recorded for any cell type. In BALB/c mice, however, infection elicited a significantly more intense total leucocyte, lymphocyte and eosinophil response than in NIH mice. In contrast, the BAL response was more intense in the NIH strain. Peak BAL responses were observed between days 12 and 16 p.i., in both strains, and the change in broncho-alveolar eosinophil numbers, as a result of infection, was found to be the greatest recorded for any cell type. The relationship between the observed leucocyte responses and resistance to the migrating larvae of a primary infection is discussed.