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PubMed1987

In vitro drug screening in isolated male Onchocerca gibsoni using motility suppression.

A primary in vitro screen was developed to screen for drug activity against isolated Onchocerca gibsoni. The assay estimates variation in motility through the use of a motility meter. Of the seven compounds tested in the screen; ivermectin, CGP 6140, CGP20376, Mel W and furapyrimidone gave MI50 concentrations (the concentration at which the motility was reduced to 50% of the control value at 72 hours) below 10(-4) M, whereas suramin gave variable results depending on the varying susceptibility of individual worms and levamisole at 10(-4) M had no significant effect on the worms. The effects of these drugs were not reversible as removal of the worms into drug-free medium caused no increase in motility. Thus the reduction in motility is regarded as indicating significant metabolic damage. The results compared favourably with reported in vivo tertiary screens for activity against Onchocerca species. This is a quantitative, inexpensive and reproducible method for assessing the effectiveness of drugs against Onchocerca and could be included into the primary screens for activity against filarial worms.

Animals↗
PubMed1986

Onchocerca spp: frequency in Thoroughbreds at necropsy in Kentucky.

Examination of the ligamentum nuchae for Onchocerca spp was completed in 523 Thoroughbreds (1 to 29 years old) at necropsy in Kentucky during a 13-month period (1984 to 1985). Onchocerca spp were found in 306 (59%) of the horses. Frequencies of this parasite were 3%, 17%, 40%, 43%, and 42% in 1-, 2-, 3-, 4-, and 5-year-old horses, respectively. For 6- to 29-year-old horses, frequency varied from 71% to 100%, except for 26-year-old horses (50%); 87% of the 304 horses examined in the 6- to 29-year-old age groups was infected. The suspensory ligaments and flexor tendons from fetlocks (thoracic limb only) of 100 Thoroughbreds (3 to 24 years old) were examined at necropsy during a 4-month period (1984) for Onchocerca spp. Infected horses were not found.

Animals↗
PubMed1981

A diagnostic skin test for Onchocerca volvulus infection.

Onchocerca supernatant (OS) was prepared by a technique permitting live microfilariae to migrate from nodule tissue through agar gel into sterile Hanks balanced salt/Penicillin-Streptomycin solution where they metabolized. The OS, after dialysis, was passed through Seitz viral filter and either concentrated or lyophilized. Using rabbit antiserum in immunodiffusion and immunoelectrophoresis tests, microfilariae proteins and also human protein were detected in out OS. No common antigens were found between this and somatic extracts of Loa loa, O. gutturosa, O. volvulus, L. carinii, D. immittis and A. lumbricoides. 125I labelled OS was purified by passage through protein A column and then through immunosorbent column of horse anti-human serum linked to CNB-activated sepharose 4B. Autoradiography, after sodium dodecyl sulphate polyacylamide slab gel eletrophoresis of purified OS, showed 10 protein bands in the molecular range 10,000 to 125,000. Skin prick tests with OS, shown not to be contaminated with Hepatitis B antigens, elicited immediate hypersensitivity reaction. Using our criteria, positive reactions were seen in 81% of proven onchocerca cases and only occasionally in Loasis 4.5%, ascaridiasis 13.5% or healthy controls 2.4%. The poor skin reactivity to OS in loasis was not due to immunosuppression as these patients, when also infested with ascaris, reacted just as well as onchocerca patients with ascaris to skin prick test using somatic extracts of ascaris.

Adolescent↗
PubMed1981

The transmission of Onchocerca tarsicola (Filarioidea: Onchocercidae) by Odagmia ornata and Prosimulium nigripes (Diptera: Simuliidae).

Onchocerca tarsicola is a subcutaneous filariid of the red deer living on the tendons of the radial-carpal and tibial-tarsal joints. Its microfilariae concentrate mainly in the external ears. The extrinsic cycle of this parasite was investigated. In a hyperendemic area wild-caught Odagmia ornata were dissected, as well as samples which had fed on a tame deer and been kept in the laboratory after the infecting blood-meal. About 3 to 5% of the flies were infected with larval stages of Onchocerca tarsicola. More than 40% of Odagmia ornata induced to feed on the ears of the deer became infected and about 30% of the flies which survived more than 18 days contained third-stage larvae. The larval development in the vector is described. Prosimulium nigripes appears to be a second natural vector of Onchocerca tarsicola. Observations concerning the potential vectors of O. flexuosa, O. tubingensis and Cutifilaria wenki living in red deer are discussed.

Animals↗
PubMed1996

Identification of a common filarial larva in Simulium damnosum s.l. (type D, Duke, 1967) as Onchocerca ramachandrini from the wart hog.

Filarial larvae resembling Type D (Duke, 1967), which are common in the Simulium damnosum s.i. vectors of human onchocerciasis ("riverblindness") in several parts of West Africa, were dissected from wild-caught flies in north Cameroon and examined morphologically. This was done in order to establish a possible synonymy with infective larvae (L3) of 2 recently discovered Onchocerca species of wart hogs (Onchocerca ramachandrini Bain, Wahl, and Renz, 1993 and Onchocerca sp. Wahl and Bain, 1995), which had been found to resemble Type D. After dissection of approximately 1,700 S. damnosum s.1., 13 Type D-like larvae were recovered from 12 infected flies. Their morphology corresponded to O. ramachandrini.

Animals↗
PubMed1995

Investigation of Culicoides (Diptera: Ceratopogonidae) in relation to the transmission of bovine Onchocerca and other filariae in central Kyushu, Japan.

In Kyushu, Japan, where a human case of zoonotic onchocerciasis was found, a survey was conducted to know if the bovine Onchocerca species of this region, O. gutturosa, O. lienalis and O. sp., which develop in local Simulium spp., were also transmitted by Culicoides. A total of 17,006 out of 42,582 females of eight Culicoides species captured by light-traps from May to November in 1989 and 1990 at two cattle sheds, one in Oita and one in Kumamoto, were dissected and examine for Onchocerca infection. Overall results showed that none of the species had filarial infections except 4 of 946 C. arakawae, an ornithophilic species, collected in Oita which harboured filarial larvae with a short tail, belonging to Lemdaninae. Unsheathed microfilariae indistinguishable from O. lienalis or O. gutturosa were found in the midgut of only one blood-fed C. matsuzawai collected in Oita. It is concluded that Culicoides species would not be vectors of these three bovine Onchocerca species. Additional collections of Culicoides in a residential area of Oita showed that several filarial species, Lemdaninae and Splendidofilarinae, probably from birds, are transmitted by C. arakawae.

Animals↗
PubMed1977

A comparison of individual dissection and mass separation for recovery of Onchocerca larvae from vector black flies.

The effectiveness of a mass separation technique, previously used for the extraction of larvae of lymphatic-dwelling filarial worms from batches of vector mosquitoes, was tested as a means of recovering infective-stage larvae of Onchocerca volvulus from Simulium ochraceum in Guatemala. Blood-engorged flies, collected from 10 infected human attractants, were maintained for 9 days to allow ingested microfilariae to develop to the infective stage. The numbers of Onchocerca larvae recovered after groups of these flies were crushed and washed into tissue culture fluid in Baermann funnels was compared with the numbers obtained by individual dissections of flies fed on the same subjects. The mass separation procedure gave a mean recovery rate of 0.03 larva/fly and detected larvae only in flies which had fed on those subjects with the highest microfilarial skin densities. Dissections yielded 0.50 larva/fly (a 16.7-fold increase) and detected larvae in flies collected from all test subjects. The explanation for the ineffectiveness of the mass separation technique may lie in the observed sluggishness of infective-stage Onchocerca larvae and a consequent inability to free themselves from the fly fragments in the Baermann funnel.

Animals↗
PubMed1997

PCR and DNA hybridization indicate the absence of animal filariae from vectors of Onchocerca volvulus in Uganda.

In order to identify Onchocerca volvulus larvae from vectors, DNA of filaria larvae from dissected blackflies was isolated, and a 150-bp long tandemly repeated DNA sequence (0-150), which occurs in many Onchocerca species, was amplified using polymerase chain reaction (PCR). Subsequently, the PCR product was blotted onto a nylon membrane and hybridized with DNA probes specific for O. volvulus or Onchocerca ochengi. Filaria larvae from 395 infected Simulium neavei were examined and 259 samples produced detectable PCR products. Among these samples, 239 (92%) reacted with an O. volvulus-specific oligonucleotide. A sample of 69 PCR products was tested using an O. ochengi DNA probe, but all failed to hybridize. Filaria larvae from 64 infected Simulium damnosum, presumably of the cytotypes "Nyamagasani" and "Nkusi" were studied and 0-150 was amplified from 38 samples. From these samples, 35 (92%) hybridized specifically with an O. volvulus probe but none with the O. ochengi-specific DNA sequence. Nonamplified samples were obtained mainly from blackflies that contained only 1 or 2 filaria larvae, and therefore, an insufficient DNA extraction was assumed. It can be concluded that few, if any, filaria species of animal origin were transmitted by S. neavei and S. damnosum s.l. in Kabarole and Kasese districts in Uganda.

Animals↗
PubMed1988

Chemotherapy of Onchocerca lienalis microfilariae in mice: a model for the evaluation of novel compounds for the treatment of onchocerciasis.

The model of Onchocerca lienalis microfilariae (mf) injected into inbred CBA/Ca mice was studied for its usefulness as an additional primary/secondary drug screen for onchocerciasis. Invermectin, DEC, suramin, flubendazole, mebendazole, levamisole, Mel W, furapyrimidone, metrifonate, amoscanate and the new Ciba-Geigy compounds CGP 6140, CGP 20'376 and CGI 17658 all significantly reduced levels of mf at a dose of 5 X 100 mg/kg or less. An early dosing protocol, on days 3-7 after infection, was found to be generally more effective than dosing on days 11-15, followed by necropsy on day 18. In some cases there were important differences in levels of drug activity depending on whether the drug was administered by the subcutaneous or oral route, indicating that new compounds should be tested via both routes. Ivermectin was by far the most active compound examined, virtually clearing mf from the skin at a dose of 5 X 0.0063 mg/kg and producing a significant mf reduction (63.5%) at 5 X 0.0008 mg/kg following subcutaneous administration. In comparison, DEC was much less active, producing a 32.4% mf reduction at 5 X 25 mg/kg ranging up to a maximum of 72% reduction at 5 X 100 mg/kg. CGI 17658 was the most active compound examined next to ivermectin, almost 100% effective against skin mf at a dose of 5 X 6.25 mg/kg via the oral route while being less effective via subcutaneous administration (65% reduction). The lowest effective dose examined was 5 X 3.13 mg/kg (per os) which reduced mf levels by 64%. CGP 20'376 was also very active, resulting in a 46% (subcutaneous) and 62% (per os) reduction at a dose of 5 X 6.25 mg/kg. This mouse model has clearly identified all the known microfilaricides examined and also, to a lesser extent, those compounds considered to be principally macrofilaricides. We believe it has value as an additional drug screen for onchocerciasis, which will enable the evaluation of novel compounds against skin-dwelling Onchocerca mf at the primary/secondary level, providing complementary information to new in vitro screens using adult Onchocerca.

Administration, Oral↗
PubMed2001

Subconjunctival zoonotic Onchocerca in an Albanian man.

A case of subconjunctival infection with a zoonotic species of Onchocerca is described, in a 16-year-old Albanian man who had immigrated to Greece. This is the first report of human infection with Onchocerca in this tissue location and only the eighth report of zoonotic Onchocerca in man.

Adolescent↗
PubMed1994

[Subcutaneous nodules and cutaneous lesions caused by different Onchocerca in African cattle].

Subcutaneous nodules and skin biopsy specimens obtained from the umbilical area of 6 onchocercal Bos indicus were collected in Cameroon and subjected to a histological examination. The nodules containing Onchocerca ochengi and Onchocerca dukei showed the same structure as Onchocerca volvulus nodules in man; they consisted of inflammatory pseudo-cysts often containing a female filaria. These pseudo-cysts were classified as "young", "active" and "old" depending on the inflammatory cellular component of their walls. The nodules were surrounded by a connective tissue capsule criss-crossed by vessels containing sections of microfilariae and morula in the vascular lumen. The skin, infected with microfilariae of these two species, of O. gutturosa and O. armillata, showed various dermatitis lesions with fibrosis, similar to those observed in patients with onchocerciasis. In most cases, inflammatory infiltrates surrounded the lymphatic capillaries along the blood vessels, giving rise to lymphangitis. The presence of microfilariae inside the granulomatous tissue demonstrates that microfilariae were deposited directly into the cystic lumen. Only the microfilariae which escaped the inflammatory reaction were able to penetrate the lymphatic vessels. They could then migrate into the lymphatic system and reach the superficial layer of the dermis. Bovine nodular onchocercomata represent an interesting model for human onchocerciasis.

Animals↗
PubMed1998

The peroxidoxin 2 protein of the human parasite Onchocerca volvulus: recombinant expression, immunolocalization, and demonstration of homologous molecules in other species.

The peroxidoxin protein of the filarial parasite Onchocerca volvulus (OvPXN-2) belongs to a group of highly conserved antioxidant molecules. For a more detailed characterization of this protein and for determination of its expression pattern the OvPXN-2 protein was recombinantly expressed as a His-tagged protein. Under reducing conditions the recombinant protein had an apparent molecular mass of 28 kDa. Considering the size of the His-tag and the FLAG epitope introduced to the recombinant protein, this size is in agreement with that of the native protein identified in O. volvulus extract. Antiserum raised against the recombinant protein was used for immunolocalization. In O. volvulus the antigen is predominantly expressed in the hypodermis and particularly the lateral and median chords show high levels of expression. The protein is also expressed strongly in the hypodermis of infective larvae and more weakly in microfilariae. Related cross-reacting proteins were detected in several Onchocerca species and other filariae. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis in combination with Western blotting revealed proteins with almost identical mobility in extracts prepared from O. ochengi, O. gibsoni, and Dirofilaria immitis.

Animals↗
PubMed1994

Identification and characterisation of a novel repetitive antigen from Onchocerca spp.

A novel repetitive antigen from the cattle parasite Onchocerca gibsoni was shown to be recognised by sera from humans infected with Onchocerca volvulus, Wuchereria bancroftii or Brugia malayi. The O. gibsoni protein was produced in a recombinant form, and antibodies raised to this protein used to screen cDNA libraries for O. volvulus. A series of clones were isolated which encoded repetitive regions very similar to those in O. gibsoni, but interspersed between these were longer repeating units which we have not so far found in O. gibsoni. The repetitive antigen was shown to be of high molecular weight and present only in the insoluble (membrane) fraction of O. gibsoni microfilariae. Immunofluorescence techniques demonstrated that the antigen was associated both with muscle and with specific membrane layers, including a peripheral layer which corresponds to either the outer hypodermis or an inner region of the cuticle in adult female O. gibsoni. In many respects, the proteins encoded by the O. gibsoni and O. volvulus cDNA clones resembled repetitive antigens from several distantly related eukaryotic parasites, and a possible common role in immune evasion is discussed.

Amino Acid Sequence↗
PubMed2002

Immunity to Onchocerca spp. in animal hosts.

This review summarizes research using Onchocerca spp. in chimpanzees, cattle and mice to gain insight into the protective immune response to the filarial worm Onchocerca volvulus in humans. In addition, Acanthocheilonema viteae has been evaluated as a surrogate filarial worm for studying immunity to the infection. Immune mechanisms controlling these infections are described and initial success using recombinant antigen vaccines in these models is reviewed.

Animals↗
PubMed1994

Induction of the prophenoloxidase-activating system of Simulium (Diptera: Simuliidae) following Onchocerca (Nematoda: Filarioidea) infection.

Trials were carried out to study the humoral immune response of blackflies to filariae following infection using the intrathoracic injection technique. An induced 66 kDa protein was abundant in the haemolymph of the European species Simulium ornatum following infection with bovine Onchocerca lienalis. This protein was apparently at higher concentrations in the haemolymph of sham-inoculated flies, i.e. flies that received sterile medium without the parasites. A molecule of the same size was also observed in the haemolymph of infected S. damnosum s.l. following infection with human O. volvulus or bovine O. ochengi. However, the level of this protein was lower in blackflies injected with microfilariae of bovine O. dukei. Unlike O. volvulus and O. ochengi this species is not transmitted by S. damnosum s.l. under natural conditions. No such reaction was observed if the African blackflies had received a sham inoculation. Feeding experiments with wild-caught nulliparous S. damnosum sl. on Onchocerca-infected cattle supported the results of the injection trials. The 66 kDa protein could only be found in the haemolymph of specimens infected via a blood meal. This 66 kDa molecule was identified as phenoloxidase. It appeared in the haemolymph due to the activation of the prophenoloxidase system following the filarial infection and we hypothesize that it may be sequestered by the parasites, as part of a non-self recognition system.

Animals↗
PubMed1996

Protective immunity induced by vaccination with Onchocerca volvulus tropomyosin in rodents.

A cDNA clone of Onchocerca volvulus, designated MOv14, and encoding 136 amino-acid residues from the C-terminus of O. volvulus tropomyosin, was evaluated as a protective immunogen in two complimentary rodent models of onchocerciasis. Vaccination of BALB/c mice with the recombinant fusion of MOv14 coupled to Maltose-Binding Protein (MBP) induced significant reductions (48-62%) in the recovery of Onchocerca lienalis microfilariae from the skin, compared to control groups immunized with MBP alone. The predominant antibody response generated to MOv14 by vaccination was of IgG1. Following a similar vaccination protocol in Mongolian jirds, two independent experiments demonstrated that 16 weeks after infection with Acanthocheilonema viteae there was a 46% reduction in the recovery of adult worms in vaccinated animals compared to control groups. Antibodies generated by vaccination recognized a product released during culture of A. viteae infective larvae which migrated at a distinct molecular mass from native tropomyosin from somatic tissues.

Animals↗
PubMed2006

Molecular systematics of five Onchocerca species (Nematoda: Filarioidea) including the human parasite, O. volvulus, suggest sympatric speciation.

The genus Onchocerca (Nematoda: Filarioidea) consists of parasites of ungulate mammals with the exception of O. volvulus, which is a human parasite. The relationship between O. volvulus, O. ochengi and O. gibsoni remains unresolved. Based on morphology of the microfilariae and infective larvae, vector transmission and geographical distribution, O. ochengi and O. volvulus have been placed as sister species. Nevertheless, the cuticle morphology and chromosomal data (O. volvulus and O. gibsoni have n=4 while O. ochengi is n=5) suggest that O. gibsoni could be more closely related to O. volvulus than O. ochengi. Sequences from the 12S rRNA, 16S rRNA and ND5 mitochondrial genes have been used to reconstruct the phylogeny of five Onchocerca species including O. volvulus. Analyses with maximum likelihood and maximum parsimony showed that O. ochengi is the sister species of O. volvulus, in accordance with the classification based on morphology and geographical location. The separate specific status of the species O. gutturosa and O. lienalis was supported, although their phylogenetic relationship was not well resolved. The analyses indicated that the basal species was O. gibsoni, a South-East Asian and Australasian species, but this result was not statistically significant. The possible involvement of sympatric speciation in the evolution of this group of parasites is discussed.

Animals↗
PubMed1995

Onchocerca volvulus: a comparative study of in vitro neutrophil killing of microfilariae and humoral responses in infected and endemic normals.

In vitro neutrophil-mediated microfilarial killing, humoral responses to crude adult worm antigens (OVAg) and to four recombinant Onchocerca antigens (GST-OV7, GST-OV103, GST-OV3.6, and MBP-OV16), as well as Western blot analysis to stage-specific Onchocerca proteins were studied in individuals from Bassa County, Liberia, infected with onchocerciasis and in endemic normals, defined as individuals without microfilaridermia. Both groups exhibited high levels of specific serum-dependent killing of microfilariae but could not be differentiated on the basis of these results. However, infected individuals had a significantly higher frequency of antibody response to OVAg (P = 0.0001) except GST-OV103. Based on the pattern of response to the different antigens, 17 categories of specific recognition were observed. Nine of these categories were common to both the infected individuals and the endemic normals, 2 were unique to the infected individuals, and 6 were unique to the endemic normals. Among the endemic normals, we identified a subcategory of individuals who had nondetectable levels of total IgG to OVAg by ELISA and had significantly lower IgG4 responses to OVAg. These same individuals demonstrated evidence of past infection, based on serum antibodies detectable by Western blot analysis to male and female adult worms and skin microfilariae, and had a positive response to two or less of the recombinant antigens. We believe that this subcategory may represent individuals with a unique status of immunity.

Adolescent↗