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PubMed1990

An economical procedure for screening of hybridoma supernatants for surface reactive antibodies to filarial larvae.

Third- and fourth-stage larvae of Onchocerca volvulus can be attached to the lids of 96-well microtiter plates to facilitate the screening of hybridoma supernatants reactive with filarial larvae. Plates are coated with a solution consisting of 0.1% gelatin, and 0.01% chrome alum. Following coating of plate lids one larva per depression is placed on the lid. An immunofluorescence assay (IFA) was conducted in which there were two 45 minute incubations and eight separate washes by immersion in PBS. At the completion of the assay over 90% of the larvae remain attached. The background in IFA is minimal and the results are consistent with those obtained using a suspension IFA. This technique permits rapid identification of antibody producing hybridomas with small numbers of larvae.

Animals↗
PubMed1987

Onchocerciasis in Plateau State of Nigeria. II. The prevalence among residents around the Assob River area.

A study of the prevalence of human onchocerciasis in eight villages around the Assob River in Jos Plateau, Nigeria, was undertaken between October 1983 and March 1985 using the skin-snip method. A total of 1673 individuals were examined. One hundred and eighty five (11.06%) had microfilariae of Onchocerca volvulus in their skin biopsies, and of these infected people, five (2.7%) were blind. The highest prevalence rate recorded was in Ngwan Madaki, where 51 (26.3%) of the 194 people examined were infected. The lowest percentage prevalence was recorded in Dogon Fili with four (1.95%) of the 205 people examined being positive. The proportion of infected people increased with age; and onchocercal lesions were significantly more common in older than in younger persons (P less than 0.05). There was no significant difference between infection rates in males and females (P greater than 0.05), although the overall percentage infection and the numbers of onchocercal lesions were higher in males than in females.

Adolescent↗
PubMed1990

Circulating immune complex-associated parasite antigens in human onchocerciasis.

This study identified and characterized parasite antigens in sera from humans infected with Onchocerca volvulus. Immune complexes were precipitated from human sera with polyethylene glycol and analyzed by immunoblot with rabbit antibodies to O. volvulus. A 23-kDa parasite antigen was detected in sera from 17 of 23 Nigerian onchocerciasis patients and 5 of 10 endemic controls. Other parasite antigens with apparent molecular masses of 62, 66, and 70 kDa were less consistently observed. These antigens were not present in Nigerian or US nonendemic control sera, in sera from patients with various other parasitic infections, or in sera from US patients with autoimmune diseases. Biochemical studies indicated that these antigens are nonglycosylated acidic proteins that do not contain phosphorylcholine. These antigens may be useful as targets for improved diagnostic tests for onchocerciasis based on parasite antigen detection.

Adult↗
PubMed1982

IgE antibodies are more species-specific than IgG antibodies in human onchocerciasis and lymphatic filariasis.

To explore the relative species specificities of the IgE and IgG antibody responses to helminth infections in man, we studied four pools of sera from patients infected with Wuchereria bancrofti, Brugia malayi, Onchocerca volvulus or Ascaris lumbricoides and ten individual sera from patients with onchocerciasis. IgE antibodies were detected by radioallergosorbent test (RAST) analysis and IgG antibodies by a Staphylococcus protein A radioimmunoassay (Staph A-RIA). Analysis of the binding curves with four different immunosorbents (prepared from antigens of B. malayi, O. volvulus, Dipetalonema viteae and A. lumbricoides) in the RAST and the binding curves with these same four antigens in the Staph A-RIA confirmed the relative species specificities for both the IgE and IgG antibody responses. Then determination of these antibody levels after specific absorption of the sera with both homologous and heterologous antigens showed that in all instances there was significantly less cross-reactivity with heterologous parasite antigens (i.e. higher species specificity) in the IgE antibody response to filarial infection than in the corresponding IgG antibody response. Such findings imply that efforts toward developing techniques for specific immunodiagnosis of filarial infections are likely to be particularly successful if focused on the IgE antibody response of exposed individuals.

Antigens↗
PubMed1995

Irreversible effects of ivermectin on adult parasites in onchocerciasis patients in the Onchocerciasis Control Programme in West Africa.

Ivermectin is an effective drug for the treatment of human onchocerciasis, a disease caused by the parasitic filarial nematode Onchocerca volvulus. When humans are treated, the microfilariae normally found in the skin are rapidly and very nearly completely eliminated. Nonetheless, after a delay, microfilariae gradually reappear in the skin. This study is concerned with the causes of this delay. Hypotheses are tested by comparing the results of model calculations with skin microfilaria counts collected from 114 patients during a trial of five annual treatments in the focus area of Asubende, Ghana. The results obtained strongly suggest that annual treatment with ivermectin causes an irreversible decline in microfilariae production of approximately 30%/treatment. This result has important implications for public health strategies designed to eliminate onchocerciasis as a significant health hazard.

Adult↗
PubMed2002

Sensitive and specific serodiagnosis of onchocerciasis with recombinant hybrid proteins.

Onchocerciasis remains a major health hazard in many tropical countries. However, the existing tools for diagnosis of the disease have limitations, particularly regarding the detection of low level or early infections. To design an optimized reagent, we exploited the high antibody reactivity of patient sera against the Onchocerca volvulus proteins Ov20 and Ov33, which have been described as highly sensitive and specific immunodiagnostic reagents for producing hybrid proteins. The construct OvH2 was composed of Ov20 fused to Ov33, while OvH3 consisted of the C-terminus of Ov20 linked to Ov33. When these constructs were tested with sera from patients with onchocerciasis and control sera, OvH2 showed a sensitivity of 98.5% and a specificity of 97.7% and OvH3 showed a sensitivity of 98.5% and a specificity of 95.35%. All non-responders were from Ecuador. These results surpass those of existing single recombinant antigens, suggesting that our hybrid proteins combined the sensitivity of the two parent proteins. Tests based on OvH2 should prove suitable for monitoring onchocerciasis control programs and individual diagnosis.

Animals↗
PubMed1979

Inflammatory reactions in onchocerciasis: a report on current knowledge and recommendations for further study.

This report concerns the host's reactions to the presence of the parasite both in the course of the natural disease and during drug treatment. The various stages of Onchocerca volvulus are discussed in terms of the type of tissue reaction seen. The discussion then turns to basic hypotheses concerning the etiology of these reactions, emphasis being placed on the fact that while pathological changes are considerable in some locations there is a remarkable lack of reaction in others. Some of the mechanisms possibly involved in this apparent absence of host response are discussed, including anti-complement factors, poor antigenicity, acquisition of host antigen, immune tolerance, and blocking antibodies. In any study of the inflammatory response it is recommended that critical evaluations be made of histological material, haematological studies, the definition of the antigenic nature of O. volvulus, characterization of immunological reactivity of patients, and the definition of the migratory pathways of the parasite.The marked host reactions seen following chemotherapy, especially those related to the interaction of the drug diethylcarbamazine with microfilariae, are discussed at some length. The etiology of these reactions is considered and recommendations are made for the experimental elucidation of the mechanisms involved. Emphasis is placed on the necessity for detailed sequential histopathological and immunopathological studies in the definition of the tissue lesions found in onchocerciasis. Characterization of these lesions will assist greatly the approach to control of the adverse reactions seen during treatment.The use of anti-inflammatory agents in clinical trials is discussed and comments are made concerning the most suitable clinical situations for testing drugs and the types of drug that should be tested.

Anti-Inflammatory Agents↗
PubMed1987

Isolation of an antigenic fraction for diagnosis of onchocerciasis.

A surface enriched fraction was prepared from adults of Onchocerca volvulus by brief extraction of entire worms with detergent. This was then gel filtered to yield a low molecular weight fraction which functioned specifically in ELISA analysis. An identical result was also obtained when the related cattle parasite, O. gibsoni, was similarly fractionated and tested. The low molecular weight fraction contained at least four antigenic components when examined by coprecipitation and immunoblotting studies. One ml of packed worms yielded sufficient low molecular weight antigen to examine about 2,000 human sera by the ELISA procedure, and the test was sensitive at human serum dilutions down to 1/400. A preliminary study with individual sera from Onchocerciasis endemic and non-endemic areas of Southern Mexico yielded 0/24 false positives, 3/24 false negatives and a significant ELISA value in 21/24 sera from proven cases of Onchocerciasis.

Animals↗
PubMed1988

The in vitro determination of the visual resolution in onchocercal nodules of bovine and human origin by a portable ophthalmologic ultrasound scanner.

A portable ophthalmologic real-time ultrasound linear scanner (Ophthascan) was used to determine the optic resolution of excised onchocercal nodules of Onchocerca volvulus and O. gibsoni origin. Fresh, thawed and fixed nodules were used. Following macroscopic and histologic examination it became apparent that the Ophthascan could clearly distinguish between echogenic worm coils and non-echogenic areas of necrosis. The fibrotic nodular capsule was more echogenic than the worm coils inside the nodule. The optic resolution did not permit to distinguish the internal organ structures within a worm segment. This in vitro determination of the visual resolution of an opthalmologic real-time ultrasound linear scanner is in accordance with in vivo results of an ongoing trial with a new macrofilaricide. Thus sonography can be used to visualise the interior of superficial onchocercal nodules in vivo and to follow their evolution with or without chemotherapy. The non-invasive diagnostic and sequential assessment of onchocercal nodules by sonography will be a valuable contribution in onchocerciasis, especially for the assessment of macrofilaricides.

Animals↗
PubMed1982

Early formation of the nodule in Guatemalan onchocerciasis.

Thirty-eight nodules containing adults of Onchocerca volvulus were removed from 36 patients who had no detectable microfilariae in skin snips. Worms were digested from nodules in collagenase solution, maintained alive in vitro, and the number, sex, and state of fecundity were recorded. A total of 48 female and 8 male worms were recovered; 39 females were in the nodules without the presence of a male. Eleven females (22.9%) had microfilariae in utero or produced microfilariae in vitro; seven of these were found together with males in the nodules while four were not. No nodules were found around male worms unless a female was also present. These observations indicate that the nodule forms only around female worms and that mating probably occurs before or early during nodule formation. Furthermore, the production of microfilariae by the the female is not essential for nodule formation since many nodules contained non-fecund, living females.

Animals↗
PubMed1978

Variation in delayed hypersensitivity in onchocerciasis.

Soluble antigen preparations were made from Onchocerca volvulus adults recovered from subcutaneous nodules, and from Necator americanus third-stage larvae. Intradermal skin tests were carried out on a total of 100 individuals clinically classified as having either the generalized form of onchocerciasis (86 cases, or the localized sowda form (14 cases). 91 of the people studied produced immediate reactions to the onchocercal antigen, but only those with sowda showed delayed reactions, though one person with generalized disease showed a doubtful delayed reaction. Reactions to N. americanus antigen, both immediate and delayed, were seen in both forms of the disease. The lack of delayed skin reaction in the generalized form of onchocerciasis is discussed, and a comparison is made with other diseases. Preliminary evidence for the existence of a true spectrum of clinical and immunological forms of onchocerciasis is suggested by one case in this study.

Adolescent↗
PubMedDate not supplied

[Onchocerciasis chemotherapy. II. Evaluation of two therapeutic schemes on microfilarial density, utilising the association diethylcarbamazine and levamisole (author's transl)].

As the study of the association diethylcarbamazine-levamisole gave encouraging results on the dermal microfilarial density of Onchocerca volvulus (Leuckart, 1893), the authors followed it up in order to determine the optimum regimen for mass treatment. Two therapeutic schemes have been tested: A--Initial treatment: 14 days with the dayly dosis of respectively 200 mg of diethylcarbamazine and 120 mg of levamisole progressively reached in four days. After one year: a single dayly dosis of respectively 200 mg and 60 mg for five days. B--Initial treatment: 7 days with the same dosis as above. After one year: dayly dosis of respectively 200 mg and 60 mg for seven days. It appears that scheme A may be considered as the best baseline to achieve the optimum regimen for the mass treatment of onchocerciasis. The aim of such a treatment is to decrease the dermal microfilarial density to a level compatible with the patient good condition.

Adolescent↗
PubMed2001

Influence of intercodon and base frequencies on codon usage in filarial parasites.

Base frequency, codon usage, and intercodon identity were analyzed in five filarial parasite species representing five Onchocercidae genera. Wucheria bancrofti, Brugia malayi, Onchocerca volvulus, Acanthocheilonema viteae, and Dirofilaria immitis gene sequences were downloaded from NCBI, and analysis was performed using locally designed computer programs and other freely available applications. A clear sequence bias was observed among the nematode species examined. At the nucleotide level, AT basepairs were present in gene sequences at higher frequencies than GC. In addition, codons ending in A or T were used proportionately more than those with G or C in the third-codon position. In addition, the amino acids used most often corresponded to codons ending in AT basepairs. Intercodon base proportion was biased in that A was found most often at N4, second only to T in certain specific cases. Since all of these sequence biases were observed in a relatively consistent fashion among all of the organisms studied, we conclude that sequence bias is a genetic characteristic, which is associated with multiple filarial genera.

Animals↗
PubMed1997

The entry of ivermectin and suramin into Onchocerca ochengi nodules.

No currently available drug, which is safe for mass treatment, effectively kills adults of Onchocerca volvulus, the causal agent of onchocerciasis in humans, or of O. ochengi, a cattle parasite used as a model of O. volvulus. Since adults of both of these filarial nematodes are found in well developed nodules, the lack of efficacy of these drugs may be a result of their poor penetration into the nodules. To check if this was the problem, the distributions of the microfilaricide, ivermectin, and the partial macrofilaricide, suramin, in plasma, skin, nodule capsules and nodule contents were determined in cattle naturally infected with O. ochengi in Cameroon. The cattle were treated with either a single, subcutaneous injection of 500 micrograms ivermectin/kg, or with intravenous injections of [14C]-labelled suramin, each of 10 mg/kg, given one a day for 6 days. Concentrations of ivermectin and suramin in various tissues were then assayed by high-pressure liquid chromatography and scintillation counting, respectively. On day 7 post-treatment (pt), suramin concentrations were consistently highest in the nodule, contents and capsule wall (11.0 and 8.9 nCi/g, respectively) and significantly less in skin and plasma (1.2 and 1.4 nCi/g, respectively; P < 0.05). The distribution of ivermectin on day 7 pt was similar, with the highest concentrations in the capsule wall, nodule contents and plasma (58.4 ng/g, 43 ng/g and 48.6 ng/ml, respectively; P > 0.05) and the concentration in the skin (6.4 ng/g) significantly lower than those in the capsule or plasma (P < 0.05). High intra-nodular concentrations of both drugs were maintained for 5-7 days at least and those of ivermectin would be expected to kill nematodes other than filariae. It is apparent that failure of ivermectin and suramin to kill adult Onchocerca spp. is not because the drugs penetrate nodules inadequately.

Animals↗
PubMed1980

The chemotherapy of onchocerciasis IV. Further trials with metrifonate.

Thirty-four healthy adult males, moderately to heavily infected with Onchocerca volvulus, were treated daily with metrifonate at 10 mg/kg body weight for either three or six days. The patients' reactions and the effects on the microfilariae were measured on a 'single-blind' basis. Muscarinic effects of acetylcholine were prominent despite the use of belladonna alkaloids, and nicotinic effects--leading to a proximal muscle weakness--occurred in one patient after the fifth dose. A syndrome of polyarthritis, fever and elevated erythrocyte sedimentation rate occurred in 11 patients, in two of whom microfilariae were demonstrated in the joint fluid. The six-day regime destroyed 83% of the microfilarial load and the three-day regime destroyed 72%, but the difference was not statistically significant (as assessed one week after the completion of treatment). It seems unlikely that metrifonate will replace DEC as the 'reference' microfilaricide and attempts to improve its efficacy by employing higher dosage must employ an intermittent dose regime.

Acetylcholine↗
PubMed1990

Large scale ivermectin distribution and its epidemiological consequences.

Community trials were started to address questions concerning the safety of ivermectin during large scale treatment, its potential for transmission control, its effect in preventing ocular onchocercal disease, its acceptability and the organization of large scale treatment. A summary is presented of the major, latest results on the short-term epidemiological impact of large scale ivermectin treatment, as observed in eight community trials undertaken in the Onchocerciasis Control Programme in West Africa (OCP). Ivermectin treatment resulted in a 96%-99% reduction in the mean load of microfilariae (mf) in the skin in treated patients. The subsequent mf-repopulation of the skin was faster than in the clinical trials and after 12 months the mean loads had returned to more than 40% of the pre-treatment load. Ocular mf loads were also greatly reduced and a post-treatment regression of early lesions of the anterior segment of the eye was observed. The transmission of Onchocerca volvulus was reduced by some 60% during the first year after treatment in one trial but no additional reduction was observed after the second treatment round. These results, and other recent research findings, have been used to quantify an epidemiological model for the transmission and control of onchocerciasis. Preliminary results of computer simulations of the predicted long-term epidemiological impact of large scale ivermectin treatment indicate that ivermectin treatment may play a very important role in disease control but that it is unlikely to become a practical tool for transmission control in endemic foci. Ivermectin treatment appears to be the most appropriate method for control of recrudescence of infection in an area where the parasite reservoir has been virtually eliminated by vector control, such as in the core area of the OCP.

Africa, Western↗
PubMed1988

Autoantibody induced by experimental Onchocerca infection. Effect of different routes of administration of microfilariae and of treatment with diethylcarbamazine citrate and ivermectin.

Hartley guinea pigs were injected with microfilariae (Mf) of Onchocerca lienalis as a model for acute inflammatory responses to Mf in human Onchocerca volvulus infection. IgG autoantibody reactive with a 3 M KCl extract of guinea pig cornea was detected by ELISA in the serum of guinea pigs injected with O. lienalis Mf three or more times sub-conjunctivally, or two or more times subcutaneously. Administration of the microfilaricides diethylcarbamazine citrate and ivermectin did not alter the proportion of animals expressing autoantibody or the mean autoantibody titer. The severity of acute corneal inflammatory reactions to Mf was similar in animals with and without circulating autoantibody. Although autoantibody responses did not correlate with acute corneal inflammatory reactions to dead Mf, the ability of Mf to induce formation of an antibody reactive with a component of autologous cornea suggests that autoimmune mechanisms might participate in chronic onchocercal lesions in the cornea, eg, sclerosing keratitis.

Animals↗
PubMed1994

Transient changes in cytokine profiles following ivermectin treatment of onchocerciasis.

Cytokine production by peripheral blood mononuclear cells after antigen or mitogen stimulation was assessed before and after semiannual ivermectin treatment of 27 patients with onchocerciasis. Before treatment, Onchocerca volvulus antigen (OvA) elicited interleukin (IL)-5 production but inhibited production of IL-10, granulocyte-macrophage colony-stimulating factor (GM-CSF), and tumor necrosis factor-alpha. Six months after the first dose of ivermectin, there were increases in the IL-2, IL-4, IL-5, and interferon-gamma responses to mitogen and in the GM-CSF and IL-10 responses to OvA. By 24 months (after four ivermectin doses), OvA-induced GM-CSF production and mitogen-induced IL-2 and IL-10 production remained elevated above pretreatment levels, whereas that of other cytokines returned to or below pretreatment levels. These transient changes in cytokine response profiles of patients with onchocerciasis following ivermectin treatment likely reflect changes in antigen load.

Adult↗