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PubMed2000

PCR assay for the specific amplification of Oesophagostomum bifurcum DNA from human faeces.

Oesophagostomiasis in humans due to infection with Oesophagostomum bifurcum (nodule worm) is of major human health significance in northern Togo and Ghana where the human hookworm, Necator americanus, also exists at high prevalence. Accurate diagnosis of O. bifurcum infection in humans is central to studying the epidemiology and controlling the parasite. To overcome limitations of current copro-diagnostic methods, we have developed an alternative, molecular approach. Utilising genetic markers in the second internal transcribed spacer (ITS-2) of ribosomal DNA, we have established a two-step, semi-nested PCR method for the specific amplification of minute amounts (fg) of O. bifurcum DNA from human faecal samples. Using a panel of 155 well-defined faecal and DNA samples, the assay achieved a sensitivity of 94.6% and a specificity of 100%. This PCR assay will be useful for the diagnosis of O. bifurcum infection and as a molecular tool for elucidating the epidemiology of human oesophagostomiasis.

Animals↗
PubMed1991

Identification of human hookworm in failed-treatment cases using Chinese hamsters (Cricetulus griseus) and scanning electron microscopy.

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.

Animals↗
PubMed1993

Observations on the morphology of adults and larval stages of Oesophagostomum sp. isolated from man in northern Togo and Ghana.

Infection with Oesophagostomum sp. appears to be extremely common in man in northern Togo and Ghana. Adult specimens were recovered from the intestinal lumen by treatment with pyrantel pamoate and the morphological characteristics of oesophagostomes of man could for the first time be compared with information available on the morphology of oesophagostomes of monkeys. The observations and measurements demonstrated that the species involved is Oesophagostomum bifurcum and that the eggs of this species cannot be differentiated from those of Necator americanus. Both infections occur simultaneously in the population involved. The L1 larvae, too, cannot be differentiated from hookworm L1 larvae. The L3 larvae, however, are characteristic. Diagnoses of human Oesophagostomum infections is based on the detection of these larvae in coprocultures. In the present paper, the eggs, the L1 and L3 larval stages and the adults, are carefully described and photos are given.

Animals↗
PubMed1996

Analysis of the prevalence of infection and associations between human gastrointestinal nematodes among different age classes living in the urban and suburban communities of Port Harcourt, Nigeria.

Fresh stool samples from 300 subjects presenting at out-patient clinics in two major hospitals in Port Harcourt were analysed for the prevalence and intensity of gastrointestinal (GI) nematode infections and 85.3% were found to carry at least one of the four species of GI nematodes recognized during the study. The most common species was Ascaris lumbricoides (54%), followed by Trichuris trichiura (43.7%) and Necator americanus (42.7%) and lastly Strongyloides stercoralis (33%). Peak prevalence for A. lumbricoides was among the < or = 9 years age cohorts, whilst those for the remaining species all coincided among the 15-19 years age cohort. The frequencies of single, double and triple-species infection categories, across the species and within most of the specific combinations were lower than the expected frequencies calculated from overall prevalence data for each species. Observed cases of non-infected individuals and those carrying all four species were higher than expected, suggesting that a greater than expected subset of the population was free from infection, particularly among subjects > or = 30 years old and another subset more prone to infection by all four species, predominantly among the age cohorts < or = 29 years. N. americanus and T. trichiura were more commonly encountered among multiple species infection combinations than expected, but the intensity of the former declined as the number of other concurrently infecting species increased. In contrast, the intensity of infection with S. stercoralis increased as the number of other species increased. Positive associations between A. lumbricoides and T. trichiura, N. americanus and S. stercoralis, and T. trichiura and N. americanus were identified in respect of greater than expected co-occurrence of these combinations and significant positive correlations between their respective intensities of infection. These were related to specific age cohorts which were identified as particularly prone to multiple infections.

Adolescent↗
PubMed1975

Mechanism of skin penetration by Ancylostoma tubaeforme larvae.

Skin penetration by infective Ancylostoma tubaeforme larvae has been investigated cinematographically and using in vitro techniques. The dermal tissue appears to cause little hinderance to larval migration but complete penetration through the skin from the dermal direction did not occur, although total penetration from the epidermal surface was frequently accomplished. No evidence could be found for enzymic secretions emanating from the worms under conditions that gave positive results from Necator americanus and Strongyloides fülleborni infective larvae. The results indicated that A. tubaeforme was able to penetrate without the use of enzymic secretions and an alternative, mechanical mechanism for penetration is advanced.

Adenosine Triphosphate↗
PubMed1985

Sodium azide preservation of faecal specimens for Kato analysis.

The modified Kato technique has the advantages of reproducibility, simplicity and economy: the disadvantage is that it cannot be used in conjunction with traditional faecal preservatives. Sodium azide has been evaluated as a preservative for human faeces for subsequent Kato analysis. More than 400 faecal samples (from normal and malnourished children, and from mixed-age participants in a field survey of the Turks and Caicos Islands) were each mixed with 2-5 mg of sodium azide powder and stored in 2 or 4 ml autoanalyser cups at ambient tropical temperature. At intervals up to 30 weeks, aliquots were prepared for Kato analysis. Trichuris trichiura, Ascaris lumbricoides and Necator americanus eggs were well preserved without degenerative or developmental changes in morphology. Quantitative analyses of 18 samples indicated that the mean egg count/sample did not change significantly after storage for 1, 2, 4, 8, 12 and 16 weeks in preservative. The use of azide preservative extends the applications of the Kato technique to field and clinical studies in which delays may occur between specimen collection and examination. The direct costs of azide preservation are substantially lower than for traditional methods and the preserved specimens are more compact and resistant to leakage.

Ascaris↗
PubMed1993

Reinfection with hookworm after chemotherapy in Papua New Guinea.

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.

Age Factors↗
PubMed1993

Hookworm infections and human iron metabolism.

Ancylostoma duodenale and Necator americanus are extremely common species of soil-transmitted helminth which flourish where poverty and malnutrition prevail. Hookworms contribute significantly to iron-deficiency anaemia, which remains one of the world's major nutritional problems, through the feeding activities of intestinal stages leading to chronic blood loss into the gut. In this article, a mathematical model is proposed to explain how human iron metabolism may respond to hookworm infection of varying intensity. The model draws attention to the importance of the regulation of stored iron levels in the process. The results from the model are presented for the effects of hookworm infection on the iron metabolism of a healthy adult male. Calculations are also presented in which the effects of hookworms on the iron metabolism of a non-pregnant woman are compared with those of a pregnant woman. Use of the model may help develop a better understanding of the pathology of hookworm disease.

Adolescent↗
PubMed1988

An investigation of hookworm infection and reinfection following mass anthelmintic treatment in the south Indian fishing community of Vairavankuppam.

Hookworm infections, as assessed by counting worms expelled following anthelmintic treatment and by egg output, were found to be of low prevalence and intensity in a South Indian fishing community. The initial overall prevalence of infection in the community was 43%, and the average burden was estimated at 2.2 hookworms per person. The age profiles of prevalence and intensity differed between males and females, with the latter harbouring significantly higher levels of infection. Children of both sexes under 10 years of age rarely harboured hookworms. Treatment with pyrantel pamoate was estimated to be 91% effective in clearing hookworm infections. Egg counts made on stools collected during an 11-month period of reinfection indicated that female patients became reinfected soon after treatment, while little hookworm egg excretion was observed in males during the observation period following treatment. Females acquired a significantly higher number of worms during the reinfection period compared with males, although the average burden in females reached only 28% of the initial, pre-treatment level. The hookworm population consisted of predominantly Necator americanus, and less than 10% of Ancylostoma duodenale. The parasites were highly aggregated within the host population with 10% of the community harbouring over 65% of the total hookworms. Low values of the negative binomial aggregation parameter, k, (indicating extreme over-dispersion) were recorded in groups stratified by age and sex. Highly significant positive correlations were observed between the initial (pre-treatment) and reinfection worm burdens of female (but not of male) patients. It is suggested that occupational practices related to walking through areas contaminated with hookworm larvae play an important role in generating the observed patterns of infection within this community.

Adult↗
PubMed2003

Hookworm cathepsin D aspartic proteases: contributing roles in the host-specific degradation of serum proteins and skin macromolecules.

Cathepsin D aspartic proteases of hookworms were recently implicated in the host-specific digestion of haemoglobin by adult parasites. Ac-APR-1 from the dog hookworm, Ancylostoma caninum and Na-APR-1 from the human hookworm, Necator americanus, were shown to be expressed in the infective larval stage (L3) as well as adult worms. We now show that both proteases degraded skin macromolecules and serum proteins, some of which were cleaved more readily from permissive definitive hosts as opposed to non-permissive hosts. Na-APR-1 degraded human collagens more efficiently than did Ac-APR-1, and Ac-APR-1 degraded canine serum albumin more efficiently than did Na-APR-1. On the other hand, both enzymes degraded human serum proteins (albumin and fibrinogen) with approximately equal efficiency under the conditions of our assays in vitro. Molecular models of these 2 orthologous, aspartic proteases showed that, despite having active site clefts with identical primary sequences, residues in the S3 pocket adopted different conformations, likely accounting for different substrate preferences reported previously. Antisera raised to both proteases partially inhibited (16-26%) migration of hookworm L3 through hamster skin in vitro, further implying a connective tissue invasive role for these enzymes in addition to digestion of serum and erythrocyte proteins for nutrition.

Ancylostoma↗
PubMed2006

Bacterial synthesis of PHA block copolymers.

Polyhydroxyalkanoates (PHA) containing block copolymers were synthesized in Cupriavidus necator using periodic substrate addition. Poly(3-hydroxybutyrate) (PHB) segments were formed during fructose utilization. Pulse feeds of pentanoic acid resulted in the synthesis of 3-hydroxyvalerate monomers, forming poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV) random copolymer. PHA synthesis was controlled using analysis of oxygen uptake and carbon evolution rates from the bioreactor off-gas. A combination of characterization techniques applied to the polymer batches strongly suggests the presence of block copolymers: (i) Thermodynamically stable polymer samples obtained by fractionation and analyzed by differential scanning calorimetry (DSC) and nuclear magnetic resonance spectroscopy (NMR) indicate that some fractions, representing approximately 30% of the total polymer sample, exhibit melting characteristics and nearest-neighbor statistics indicative of block copolymers, (ii) preliminary rheology experiments indicate additional mesophase transitions only found in block copolymer materials, (iii) dynamic mechanical analysis shows extension of the rubbery plateaus in block copolymer samples, and (iv) uniaxial extension tests result in differences in mechanical properties (modulus and elongation at failure) expected of similarly prepared block copolymer and single polymer type materials.

Cells, Cultured↗
PubMed1980

Homology between roundworm (Ascaris) and hookworm (N. americanus) antigens detected by human IgE antibodies.

Utilizing the techniques of radioallergosorbent test (RAST), inhibition of RAST and isoelectric focusing on polyacrylamide gels, antigenic cross-reactivity has been shown between extracts of Ascaris lumbricoides. Ascaris suum and Necator americanus (hookworm). Antigenic homology is extensive between the two Ascaris preparations. The antigens common to hookworm and Ascaris, while representing a minor component of Ascaris body fluid, are nonetheless potent stimulators of IgE antibody production in subjects infected only with hookworm larvae. These shared antigens appear to have a pI of less than 5.1. The hookworm specific antigens also have a pI of less than 5.1, whereas those specific to Ascaris have pI values in the ranges of 5.0 to 6.0 and 7.0 to 8.6. These latter antigens, i.e. those with pI between 7.0 and 8.6, apparently elicit an immune response only during an active infection with Ascaris.

Antigens↗
PubMed1999

A hookworm allergen which strongly resembles calreticulin.

Immmoglobulin E-rich plasma from patients from Papua New Guinea infected with Necator americanus has been used to probe an adult N. americanus cDNA library for the presence of hookworm allergens. Using this approach, one hookworm allergen has been identified as calreticulin, which was subsequently expressed in Escherichia coli. Little serological cross reactivity was seen between the recombinant calreticulins of this hookworm and its host. Prospective roles for hookworm calreticulin in the host-parasite relationship are discussed in depth.

Allergens↗
PubMed1978

Human Ancylostoma infections in Nigeria.

An estimate of the proportion of infections with Ancylostoma duodenale and Necator americanus by the examination of larvae collected from the stools of 220 persons infected with hookworm was made. Fifty-nine per cent of the infections were due to N. americanus only: A. duodenale always occurred in association with N. americanus. In 12% of the infections it is suggested that there were approximately three female A. duodenale to 95 female N. americanus. The highest numbers of A. duodenale in mixed infections (28 female A. duodenale with 45 female N. americanus) occurred in only 1% of the infections. Evidence is given that egg size can be important in the differentiation of the two species. A method of calculating the proportion of the two hookworms, and hence of other intestinal nematodes, in mixed infections in an individual as well as in the general population, is also described.

Adolescent↗
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↗
PubMed1998

Parasitic infections in villagers from three districts of the Brazilian Amazon.

A survey of intestinal parasites among 827 rural villagers in the Brazilian Amazon revealed that the prevalences of infection with Necator americanus, Trichuris trichiura and/or Ascaris lumbricoides were high. However, the intensities of infection with the two parasites for which there were enough data on intensity for further analysis, N. americanus and T. trichiura, were only low or moderate in each of the three districts studied. Possible reasons for the unusual (but not unique) absence of high intensities include moderately good sanitation, low population densities, the way in which work activities are located geographically, the use of indigenous anthelmintics and the geographical isolation of the communities which provided the subjects. Whether the light to moderate levels of infection observed require intervention is discussed.

Adolescent↗
PubMed2003

Hookworm aspartic protease, Na-APR-2, cleaves human hemoglobin and serum proteins in a host-specific fashion.

Hookworms are voracious blood-feeders. The cloning and functional expression of an aspartic protease, Na-APR-2, from the human hookworm Necator americanus are described here. Na-APR-2 is more similar to a family of nematode-specific, aspartic proteases than it is to cathepsin D or pepsin, and the term "nemepsins" for members of this family of nematode-specific hydrolases is proposed. Na-apr-2 mRNA was detected in blood-feeding, developmental stages only of N. americanus, and the protease was expressed in the intestinal lumen, amphids, and excretory glands. Recombinant Na-APR-2 cleaved human hemoglobin (Hb) and serum proteins almost twice as efficiently as the orthologous substrates from the nonpermissive dog host. Moreover, only 25% of the Na-APR-2 cleavage sites within human Hb were shared with those generated by the related N. americanus cathepsin D, Na-APR-1. Antiserum against Na-APR-2 inhibited migration of 50% of third-stage N. americanus larvae through skin, which suggests that aspartic proteases might be effective vaccines against human hookworm disease.

Amino Acid Sequence↗
PubMed2004

Immune responses in human necatoriasis: association between interleukin-5 responses and resistance to reinfection.

Cytokine and proliferative responses to Necator americanus infection were measured in a treatment-reinfection study of infected subjects from an area of Papua New Guinea where N. americanus is highly endemic. Before treatment, most subjects produced detectable interleukin (IL)-4 (97%), IL-5 (86%), and interferon (IFN)- gamma (64%) in response to adult N. americanus antigen. Pretreatment IFN- gamma responses were negatively associated with hookworm burden, decreasing by 18 pg/mL for each increase of 1000 eggs/gram (epg) (n=75; P<.01). Mean IFN- gamma responses increased significantly after anthelmintic treatment, from 166 to 322 pg/mL (n=42; P<.01). The intensity of reinfection was significantly negatively correlated with pretreatment IL-5 responses, decreasing by 551 epg for each 100 pg/mL increase in production of IL-5 (n=51; P<.01). These data indicate that there is a mixed cytokine response in necatoriasis, with worm burden-associated suppression of IFN- gamma responses to adult N. americanus antigen. Resistance to reinfection is associated with the parasite-specific IL-5 response.

Adolescent↗