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At least 19 recordsLinked to original sources
PubMed1995

Mitochondrial alleles of Simulium damnosum sensu lato infected with Onchocerca volvulus.

Onchocerca volvulus infected Simulium damnosum s.l. were analysed by directed heteroduplex analysis. Of 73 infected flies, 68 produced heteroduplex products identical to those previously identified. All 6 major sibling species, except S. leonense, were present in this group. In the 5 remaining flies, 2 new heteroduplex patterns were noted. Molecular phylogenetic analysis of these samples suggested that they belonged to the S. squamosum/S. yahense subcomplex. The ability to reliably genotype adult flies will permit studies of the vectorial capacity of the sibling species of S. damnosum s.l. for the blinding and non-blinding strains of O. volvulus.

Alleles↗
PubMed1997

In vitro production and characterization of excretory/secretory products of Onchocerca volvulus.

Onchocerca volvulus excretory/secretory products [ESP] free of host contaminants are often needed for immunologic and biochemical studies. Since prolonged in vitro survival of O. volvulus in culture requires the presence of human serum, as a supplement, it was necessary to investigate other culture medium supplements in which pure ESP could be generated. Thus heat-inactivated normal rabbit serum, fetal calf serum and the synthetic serum substitute Ultroser-G were tested for their abilities to sustain O. volvulus adult females and nodular microfilariae in vitro and their abilities to promote the synthesis and release of ESP. Using [35S]-methionine as the radioactive precursor, O. volvulus nodular microfilariae and adult females were shown to actively synthesize and release excretory/secretory proteins with increasing efficiency in human serum, rabbit serum, Ultroser-G and fetal calf serum. Analysis of the ESP by SDS-PAGE revealed that at least 30 polypeptides in the 7-174 kDa range were continuously released. About 14 of these components were immunogenic to the human host as shown by immunoprecipitation. Prominent antigenic polypeptides were those with relative molecular weights of 13, 16, 33, 68 and 170. Rabbit serum, fetal calf serum and Ultroser-G could therefore, conveniently replace human serum in cultures of O. volvulus nodular microfilariae and adult females.

Animals↗
PubMed2000

The genomes of Onchocerca volvulus.

Onchocerca volvulus, the filarial parasite that causes onchocerciasis or river blindness, contains three distinct genomes. These include the nuclear genome, the mitochondrial genome and the genome of an intracellular endosymbiont of the genus Wolbachia. The nuclear genome is roughly 1.5x10(8) bp in size, and is arranged on four chromosome pairs. Analysis of expressed sequence tags from different life-cycle stages has resulted in the identification of transcripts from roughly 4000 O. volvulus genes. Several of these transcripts are highly abundant, including those encoding collagen and cuticular proteins. Analysis of several gene sequences from O. volvulus suggests that the nuclear genes of O. volvulus are relatively compact and are interrupted relatively frequently by small introns. The intron-exon boundaries of these genes generally follow the GU-AG rule characteristic of the splice donor and acceptors of other vertebrate organisms. The nuclear genome also contains at least one repeated sequence family of a 150 bp repeat which is arranged in tandem arrays and appears subject to concerted evolution. The mitochondrial genome of O. volvulus is remarkably compact, only 13747 bp in size. Consistent with the small size of the genome, four gene pairs overlap, eight contain no intergenic regions and the remaining gene pairs are separated by small intergenic domains ranging from 1 to 46 bp. The protein-coding genes of the O. volvulus mitochondrial genome exhibit a striking codon bias, with 15/20 amino acids having a single codon preference greater than 70%. Intraspecific variation in both the nuclear and mitochondrial genomes appears to be quite limited, consistent with the hypothesis that O. volvulus has suffered a genetic bottleneck in the recent past.

Animals↗
PubMed1994

A novel type of glutathione S-transferase in Onchocerca volvulus.

Onchocerca volvulus is a pathogenic human filarial parasite which, like other helminth parasites, is capable of evading the host's immune responses by a variety of defense mechanisms which are likely to include the detoxification and repair mechanisms of the enzyme glutathione S-transferase (GST). In this study, we show that one of the previously described GSTs from O. volvulus appears to possess the characteristics of a secreted enzyme. When the complete O. volvulus GST1 (OvGST1) sequence presented here is compared with those of other GSTs, 50 additional residues at the N terminus are observed, the first 25 showing characteristics of a signal peptide. This is consistent with the N-terminal sequence data on the native mature enzyme which begins at amino acid 26, based on the deduced protein sequence from the cDNA. The native protein, without the signal peptide sequence, possesses a 24-amino-acid extension not present in other GSTs. The deduced amino acid sequence of the OvGST1 cDNA clone was shown to possess four potential N-glycosylation sites. Digestion of O. volvulus homogenate with endoglycosidase, followed by detection of OvGST1 with specific antibody, indicated that the enzyme possesses at least two N-linked oligosaccharide chains. Gel filtration of the Escherichia coli-produced recombinant OvGST1 showed that it is enzymatically active as a nonglycosylated dimer. OvGST1 is found in the media surrounding adult worms maintained in culture, indicating that, in vitro, this enzyme is released from the worm. The strongest immunostaining for OvGST1 was observed in the outer cellular covering of the adult worm body, the syncytial hypodermis, especially in the interchordal hypodermis, where the peripheral membrane forms a series of lamellae which run into the outer zone of the hypodermal cytoplasm.

Amino Acid Sequence↗
PubMed2001

Structural analysis and antibody response to the extracellular glutathione S-transferases from Onchocerca volvulus.

Onchocerca volvulus is a human pathogenic filarial parasite which, like other parasitic nematodes, is capable of surviving in an immunologically competent host by employing a variety of immune evasion strategies and defense mechanisms including the detoxification and repair mechanisms of the glutathione S-transferases (GSTs). In this study we analyzed the glycosylation pattern and the immunological properties of extracellular O. volvulus GST1a and -1b (OvGST1a and -1b). The enzymes differ in only 10 amino acids, and both are glycoproteins that have cleavable signal peptides and unusual N-terminal extensions. These characteristics have not been described for other GSTs so far. Mass spectrometry analyses indicate that both enzymes carry high-mannose type oligosaccharides on at least four glycosylation sites. Glycosylation sites 1 to 3 of OvGST1a (OvGST1b sites 2 to 4) are occupied by truncated N-glycans (Man(2)GlcNAc2 to Man(5)GlcNAc(2)), and N glycosylation site 4 of OvGST1a (OvGST1b site 5) carries Man(5)GlcNAc2 to Man(9)GlcNAc(2). To analyze the capacity of these secretory GSTs to stimulate host immune responses, we studied the antibody responses of onchocerciasis patients against the native affinity-purified OvGST1a and -1b. By enzyme-linked immunosorbent assay we showed that OvGST1a and -1b are immunodominant antigens, with less than 7% nonresponder patients. A direct comparison of the antibody responses to the glycosylated and deglycosylated forms demonstrates the high immunogenicity of the N-glycans. Analyses of the antibody responses to the unusual N-terminal extension show an enhanced recognition of this portion by patients as opposed to recognition of the recombinant protein without extension.

Amino Acid Sequence↗
PubMed1992

Effects of three-month doses of ivermectin on adult Onchocerca volvulus.

Onchocerca volvulus worms in nodules from Guatemalan patients treated with four, eight, or 11 single doses of ivermectin (150 micrograms/kg of body weight) that were given once every three months were examined by routine histologic techniques and compared with worms in control nodules from untreated persons living in the same location over the same time periods. All treated nodules were removed four months after the last dose of ivermectin, i.e., 13, 25, or 34 months after the start of the trial. At the 25th and 34th months, i.e., after the eighth or eleventh doses of ivermectin, there were excess mortalities in female worms of 25.5% and 32.6%, respectively, over and above the levels in controls. Furthermore, the proportions of live females still producing scanty embryos up to the gastrula stage were only 7.7% and 18.2%, and no females were producing microfilariae. Ivermectin given at 3-month intervals also reduced significantly the mean numbers of live male worms in nodules, as well as the proportions of inseminated females. This regimen was effective in preventing embryogenesis to the microfilarial stage while, at the same time, it caused a slow but steady attrition of the adult worms.

Adolescent↗
PubMed1990

On the reproductive activity of the female Onchocerca volvulus.

Onchocerca volvulus worms, extracted from nodules by collagenase digestion, stained with haematoxylin and cleared in glycerol, were unravelled for longitudinal examination and later embedded in brain blocks for study of serial transverse sections. A classification system for female worms is proposed, based on the reproductive status of 446 worms from Guatemala, 94 from Liberia and 125 from Mali. They were categorized into fecund, inseminated specimens; uninseminated, but potentially fertile specimens, shedding ova destined to degenerate; worms changing from the uninseminated to the inseminated state and vice versa, which were few in number; old worms, with degenerate ovaries, whose genital tracts were either empty or had disappeared; and moribund or dead worms, characterized by loss of turgor, collapse and degeneration, calcification, or invasion by polymorphic, basophilic cells. Potentially fertile worms shed oocytes continuously and, when they were inseminated, embryonic development ensured. No evidence was found of a periodic cycle of reproduction. Inseminated worms were found in nodules without a male worm, and uninseminated worms in nodules harbouring male worms. Measurements are recorded of portions of the female reproductive tract and of the length of uterus occupied by the various embryonic stages in fully fecund worms. A significant difference in the length of the body behind the first and second ovaries was observed as between worms from West African savanna (Mali) and forest (Liberia). Limited observations were also made on meiosis in the oocyte, penetration of the oocyte by sperm, formation of the ovum, syngamy and zygote formation.

Animals↗
PubMed1986

Identification of antigens of Onchocerca volvulus and Onchocerca gibsoni for diagnostic use.

Adults of Onchocerca volvulus and Onchocerca gibsoni were identically fractionated into a surface-enriched fraction, a phosphate buffered saline (PBS) extract and a PBS insoluble-detergent soluble fraction. Glycoproteins were prepared from these extracts and all fractions were examined by the Western blot technique using sera from individuals infected with a variety of filarial and non-filarial nematode worms. Using antisera to O. volvulus, a number of antigens were demonstrated in all of the extracts, with some antigens of each extract being unique. Many antigens were glycoproteins, and a high cross-reactivity was observed between O. volvulus and O. gibsoni. The different fractions of both species were also analysed using a panel of different sera in order to identify Onchocerca-specific antigens. The studies revealed that the lower molecular weight antigens showed greater Onchocerca specificity in all of the extracts examined. The surface-enriched fraction, however, clearly contained less widely cross-reacting components than the somatic and glycoprotein fractions. Finally, using surface labelling and coprecipitation techniques, O. gibsoni was shown to possess a 20 kDa Onchocerca-specific antigen, previously described for O. volvulus. The findings indicate a number of Onchocerca-specific antigens which may have potential in diagnosis of human onchocerciasis. They also show that the related bovine parasite O. gibsoni, may be an alternative source of material.

Animals↗
PubMed1998

[A case of cross infection by Onchocerca volvulus and Onchocerca ochengi in Simulium damnosum S.L].

During a routine entomological survey conducted within the framework of the Program to Control Onchocerciasis in West Africa, a female simulium forest fly was found to be contaminated by 13 Onchocerca volvulus larvae and 7 Onchocerca ochengi larvae. The two Onchocerca species were identified using specific DNA probes. We speculate that cross infection could be related either to behavioral factors, e.g. interruption of blood meals on two different hosts, or developmental factors, e.g. asynchronous development of parasites of the same species or specific differences in the duration of parasite cycles. Further study will be needed to determine the incidence and scope of cross infection in areas where accurate assessment of the impact of vector control on transmission of onchocerciasis in man is required.

Africa, Western↗
PubMed1996

In vitro cultivation and development of Onchocerca volvulus and Onchocerca lienalis microfilariae.

Onchocerca volvulus and O. lienalis skin-derived microfilariae (mf) were cultured in vitro; parasite viability was assessed at intervals by measuring their ability to develop in Simulium ornatum. In the presence of monkey kidney feeder cells, both species retained full viability when cultured for up to 5 hr before intrathoracic injection into Simulium. In the absence of feeder cells and in contrast to O. lienalis, O. volvulus mf rapidly lost their viability. In further trials (including feeder cells), O. volvulus mf retained full viability for 14 days, while O. lienalis mf retained full viability for a least 19 days but with a proportion able to develop to third-stage larvae (L3) after 70 days in culture. In experiments using this system to culture O. volvulus mf (ex utero) derived from adult female worms but with the addition of reduced glutathione and/or 20-hydroxyecdysone, parasites were consistently more active over a 70-day period than those cultured without these additives. None of the mf cultured without additives were able to develop to L3 in Simulium when tested for up to 51 days in culture, while a proportion of mf incubated with reduced glutathione and/or 20-hydroxyecdysone produced small but significant numbers of L3 after 28, 43, and 51 days, representing the first time that uterine mf have been cultured to a form infective for the vector.

Animals↗
PubMed1991

Comparison of the effects of a single dose and of four six-monthly doses of ivermectin on adult Onchocerca volvulus.

Adult Onchocerca volvulus worms, extracted from nodules of Guatemalans by collagenase digestion, were examined whole and by histological techniques. One group of persons received a single 150 micrograms/kg dose of ivermectin; two other groups (one with older and one with younger nodules) received four similar doses of ivermectin at 6-month intervals. For each group, there were comparable untreated controls. All nodules were removed six months after the last dose. After a single dose, the only significant difference from the controls was in the decreased proportion of female worms producing live microfilariae. After four doses, there were significant increases in the proportions of moribund/dead female worms and of live uninseminated females, when compared with the corresponding controls. There were also fewer male worms present, but this difference was not significant. Six months after the conclusion of the 4-dose regimen, the proportion of female worms producing live microfilariae was significantly lower than in the groups that had received a single dose.

Animals↗
PubMed1996

Biochemical analysis, gene structure and localization of the 24 kDa glutathione S-transferase from Onchocerca volvulus.

Survival of Onchocerca volvulus, a pathogenic human filarial parasite, is likely to depend upon the detoxification activities of the glutathione S-transferases (GSTs). The 24 kDa O. volvulus GST, OvGST2, was expressed in a bacterial system and the recombinant protein was purified to homogeneity by affinity chromatography. Specific activities of the recombinant OvGST2 (rOvGST2) with a variety of substrates, and in the presence of inhibitors, were determined. With the universal substrate 1-chloro-2,4-dinitrobenzene, the specific activity of rOvGST2 was 2130 nmol min-1 mg-1. The rOvGST2 showed relatively limited selenium-independent glutathione peroxidase activity, but secondary products of lipid peroxidation, namely members of the trans,trans-alka-2,4-dienal,trans-alk-2-enal and 4-hydroxyalk-2-enal series, were conjugated to glutathione via OvGST2 dependent activity. The gene encoding the OvGST2 was isolated and the nucleotide sequence determined. The ovgst2 gene was found to possess seven exons with six intervening sequences, with all except one having consensus splice-site junctions. This intron/exon organisation of the ovgst2 gene is almost identical with those described for the mammalian Pi class GST genes, consistent with the protein structural evidence that the OvGST2 is related to the Pi class GSTs. Southern blot analysis with total parasite genomic DNA indicated a single copy gene, with a restriction pattern consistent with that of the isolated gene. The tissue distribution of the OvGST2 was examined in O. volvulus by immunohistochemistry and was shown to be distinct from that of the OvGST1. The OvGST2 was located throughout the syncytial hypodermis of male and female adult worms, as well as in the uterine epithelium. Microfilariae, and infective third stage larvae of O. volvulus, isolated from Simulium neavei, were immunopositive for OvGST2.

Amino Acid Sequence↗
PubMed1995

Immunity to onchocerciasis: putative immune persons produce a Th1-like response to Onchocerca volvulus.

Immunity to Onchocerca volvulus (Ov) infection is suggested by the presence of putatively immune (PI) subjects in a region of Ecuador in which Ov is endemic. PI subjects were identified by traditional diagnostic methods combined with a polymerase chain reaction-based assay for Ov DNA in skin snips. Responses of peripheral blood mononuclear cells (PBMC) from the PI group (n = 16) were compared with those of persons with active infection (microfiladermic [MF] subjects; n = 51). PBMC of PI subjects proliferated significantly more to Ov antigen (OvAg; P < .009) than did PBMC of MF persons but less to streptolysin-O (P < .001). Cytokine analysis of PBMC culture supernatants revealed that PI subjects (n = 11) produced significantly more interferon-gamma to OvAg than did those in the MF group (n = 18; P = .018), less interleukin (IL)-5 to nonparasite antigen (P = .003) and mitogen (P = .012), and less IL-10 spontaneously (P = .016). Thus, immunity to Ov may in part be mediated by an antigen-specific Th1-type response.

Adolescent↗
PubMed1986

Detection of protein kinase substrates in extracts of Onchocerca volvulus.

Extracts of Onchocerca volvulus were phosphorylated in the presence of (gamma 32P)ATP and Mg2+ by endogenous protein kinase activity and exogenous rabbit muscle catalytic sub-unit of the adenosine 3'5' monophosphate dependent protein kinase (E.C. 2.7.1.37). Sodium dodecylsulfate polyacrylamide gel electrophoretic analysis of the 32P-labelled extracts revealed at least seven (32P)-phosphoproteins with apparent Mr of 92,000; 86,000; 40,000; 27,000; 26,000; 23,000 and 17,000. The phosphorylation of the components with apparent Mr of 23,000 and 17,000 was catalysed by both endogenous and exogenous protein kinases, whereas the other components required exogenous protein kinase for their phosphorylation. The endogenous protein kinase activity was inhibited by suramin and the heat-stable protein inhibitor of the adenosine 3'5' monophosphate dependent protein kinase. The (32P)phosphoproteins identified in this investigation are probably candidate regulatory molecules in O. volvulus; though their physiological functions remain to be determined.

Animals↗
PubMed1988

Isolation and in vitro translation of Onchocerca volvulus mRNA.

Adult Onchocerca volvulus recovered for excised nodules by dissection or treatment with collagenase have been used as a source of RNA for in vitro translation experiments. RNA was purified using either the hot phenol/SDS procedure or the guanidine isothiocyanate protocol. Immunoprecipitation experiments performed on in vitro products demonstrate a marked heterogeneity in responses by individed human infection sera. Further immunoprecipitation experiments demonstrate cross reactivity between O. volvulus and other filarial nematodes.

Animals↗
PubMed1986

Antigenic similarity of Onchocerca volvulus to other helminths examined by monoclonal antibodies against O. volvulus.

Antigens in Onchocerca volvulus were characterized using anti-O. vovulus monoclonal antibodies (McAb). A total of 17 McAb which were not species-specific to O. volvulus was examined for the antigenic similarity of O. volvulus with 12 parasitic helminths using indirect immunofluorescence. The range of reactivity of these McAb was wider in nematodes in contrast to species of cestoda or trematoda. The antigenic similarity within helminths examined was generally in agreement with their taxonomic relationships but was not shown in the lower taxonomic hierarchy. The helminth which holds the antigens most in common to O. volvulus was not the filarial parasites but Ascaris suum, an intestinal nematoda. The least antigenic similarity to O. volvulus was found in Schistosoma mansoni and S. japonicum, trematoda, among helminths examined. On the other hand, the existence of a common antigen shared with all the 13 helminths examined was demonstrated by a IgM McAb.

Antibodies, Monoclonal↗