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[
Acta Trop,
1978]
Soluble antigen was prepared from adult worms and microfilariae of Onchocerca volvulus. In 27 patients clinically characterized as having generalized type (17) and localized reaction type (10) onchocerciasis, the response to this antigen was assessed in vivo by skin testing and in vitro by leucocyte migration inhibition assay. Three varieties of skin hypersensitivity were observed: Immediate, Arthus-type and delayed or type IV hypersensitivity. The first 2 occurred in all patients, but type IV hypersensitivity was noted only in 10, eight of whom had localized reactive type onchocerciasis, whilst the other 2 had mild generalized lesions which had for the most part healed with treatment. The leucocyte migration inhibition assay showed a spectrum of responsiveness with significant inhibition in the presence of antigen in 8 patients with localized lesions and in 2 others with mild generalized disease which had largely healed.--The specificity of the skin reaction and leucocyte migration inhibition to the antigen used was confirmed by the results obtained when a control group of 18 subjects, with no evidence of previous filarial infection, was similarly tested. 16 of them showed no skin hypersensitivity and 2 had a doubtful immediate reaction. No significant inhibition of leucocyte migration in the presence of antigen was noted in this group.
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[
Acta Trop,
1981]
A method for the selective recovery of living microfilariae from Onchocerca volvulus nodules is described. The microfilariae migrate through solidified agar gel into overlayering Hanks balanced salt solution (HBSS). The highest recovery rates of the worms were obtained with 0.3 and 0.4% agar. Optimal conditions for in vitro cultures of the larvae in HBSS were established; pH range 7.0-7.5, glucose concentration 2-5 mg/ml for long term cultures, osmolality 200-309 mOsmoles/l, temperature 4 degrees C for prolonged cultures and 24-28 degrees C for overall best yield of excretory/secretory products (ESP). Subculturing of the larvae reduced contamination of ESP with human serum protein to minimal amounts after 9 recultures done within 96 h.
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[
Trop Med Parasitol,
1985]
The effects of Onchocerca volvulus infection on immunoglobulin levels have been studied in subjects from three localities with different onchocerciasis prevalence rates. Infestation by O. volvulus provoked a 200-500% rise in total serum IgE levels. Immunoglobulins G, M. and A were increased less dramatically. To study further the IgE response, radioallergosorbent test (RAST) systems were developed using total extract of O. volvulus and Onchocerca supernatant (excretory-secretory) antigens. Both antigen preparations reacted with more than 80% of the onchocerciasis sera from the hyperendemic village, but the reactions were not proportionate to skin microfilarial density. There were also positive reactions with sera of some individuals whose skin biopsies showed no microfilariae, indicating prior exposure to the parasite. European control sera tested under similar conditions reacted only slightly in the RAST system. The developed RAST systems may thus be used for determining prior exposure to O. volvulus.
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[
Bull Soc Pathol Exot,
1999]
Onchocerciasis, also known as "river blindness", presents a plenum of clinical manifestations which vary from one individual to another, and from one area to another. This large spectrum of clinical manifestations of the disease is an indication of the complexity of the pathogenesis of onchocerciasis and suggests that many interacting factors might influence the clinical features of the disease. The present study has focused on the heterogenicity of the host immune response as a plausible explanation for differences in clinical manifestations of the infection. Host genetic factors, namely HLA genes, might play an important role in determining the nature of the immune response mounted against the parasite Onchocerca volvulus, and thus the development of different manifestations of the infection. Genetic diversity of onchocerciasis was assessed in different endemic foci in Cameroon. In order to investigate the possibility that the Major Histocompatibility Complex (MHC) genes might be associated with the different clinical types of onchocerciasis, 146 subjects living in three endemic areas of Cameroon were studied. They were classified in four groups: A (asymptomatic subjects), P (putatively immune subjects) L (patients with localised disease) and G (patients with generalised disease). The four groups differed in the distribution of HLA class II alleles as determined by Direct Heteroduplex Analysis. On the one hand, allele HLA-DQA1*0501 appeared to be associated with protection against severe onchocerciasis; on the other, allele HLA-DQB1*0201 might play an important role in the severe form of the disease.
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Ngu JL, Lando G, Barr PJ, Nkenfou C, Morgan G, Tume CB, Seigel J, Bathurst I, Asonganyi T, McKerrow JL, Sun E
[
Am J Trop Med Hyg,
1997]
A yeast (Saccharomyces cerevisiae) expression system has been adapted to produce reagent quantities of a major Onchocerca antigen, Ov33. Using a pool of monoclonal antibodies produced against third-stage larvae, a cDNA library constructed from adult O. volvulus worms was screened. Twenty-seven cDNAs were isolated, two of which had sequence homology to Ov33, a putative aspartyl protease inhibitor, which is the immunodominant antigen of O. volvulus. These cDNAs were expressed at high levels intracellularly or through the secretory pathway of S. cerevisiae. Localization studies using antisera produced against purified recombinant protein demonstrated that Ov33 is a very abundant parasite protein present in the hypodermis, muscle, and uterus of female worms, as well as in embryonic microfilariae. The soluble recombinant protein secreted by yeast (C71) demonstrated inhibitory activity against the aspartyl protease pepsin. Antibodies to the recombinant protein-mediated leukocyte adherence to and killing of skin microfilariae. The sensitivity of a diagnostic test using recombinant Ov33 was evaluated using sera from 441 patients. The mean sensitivities for the two recombinant constructs, C27 and C71, were 82.2% and 85.4%, respectively. The combined sensitivity using both recombinant proteins was 94%.
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Shimono K, Honda N, Hasegawa T, Takahashi M, Hashimoto N, Sudo Y, Hayashi S, Mizutani K, Miyauchi S, Yamamoto M, Takagi S, Yamashita K, Tsukamoto T, Murata T
[
J Biol Chem,
2016]
Thermophilic rhodopsin (TR) is a photoreceptor protein with an extremely high thermal stability and the first characterized light-driven electrogenic proton pump derived from the extreme thermophile Thermus thermophilus JL-18. In this study, we confirmed its high thermal stability compared with other microbial rhodopsins and also report the potential availability of TR for optogenetics as a light-induced neural silencer. The x-ray crystal structure of TR revealed that its overall structure is quite similar to that of xanthorhodopsin, including the presence of a putative binding site for a carotenoid antenna; but several distinct structural characteristics of TR, including a decreased surface charge and a larger number of hydrophobic residues and aromatic-aromatic interactions, were also clarified. Based on the crystal structure, the structural changes of TR upon thermal stimulation were investigated by molecular dynamics simulations. The simulations revealed the presence of a thermally induced structural substate in which an increase of hydrophobic interactions in the extracellular domain, the movement of extracellular domains, the formation of a hydrogen bond, and the tilting of transmembrane helices were observed. From the computational and mutational analysis, we propose that an extracellular LPGG motif between helices F and G plays an important role in the thermal stability, acting as a "thermal sensor." These findings will be valuable for understanding retinal proteins with regard to high protein stability and high optogenetic performance.