Hamburg, Universiẗat, Diss., 2002.

Submitted by Angelo Silva (asilva@icict.fiocruz.br) on 2016-07-13T18:31:00Z No. of bitstreams: 2 license.txt: 1748 bytes, checksum: 8a4605be74aa9ea9d79846c1fba20a33 (MD5) 71824.pdf: 2172041 bytes, checksum: 706d176d456c37e0c405bbdd3df5101a (MD5) / Approved for entry into archive by Anderson Silva (avargas@icict.fiocruz.br) on 2016-07-21T19:07:35Z (GMT) No. of bitstreams: 2 71824.pdf: 2172041 bytes, checksum: 706d176d456c37e0c405bbdd3df5101a (MD5) license.txt: 1748 bytes, checksum: 8a4605be74aa9ea9d79846c1fba20a33 (MD5) / Made available in DSpace on 2016-07-21T19:07:35Z (GMT). No. of bitstreams: 2 71824.pdf: 2172041 bytes, checksum: 706d176d456c37e0c405bbdd3df5101a (MD5) license.txt: 1748 bytes, checksum: 8a4605be74aa9ea9d79846c1fba20a33 (MD5) Previous issue date: 2015 / Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Rio de Janeiro, RJ, Brasil / No Brasil, a área endêmica para oncocercose está restrita a Amazônia, em Terra Indígena Yanomami. O Programa Brasileiro de Eliminação da Oncocercose (PBEO) adota como principal estratégia de controle o tratamento em massa dessa população com Ivermectina, estando em consonância com o Programa para Eliminação da Oncocercose nas Américas (OEPA) e os outros cinco países endêmicos na América Latina \2013 México, Guatemala, Equador, Colômbia e Venezuela. De acordo com estratégias de monitoramento definidas pela Organização Mundial da Saúde (OMS), foi realizado esse estudo visando à disponibilização de ferramentas para avaliação do estado epidemiológico da oncocercose baseadas em análises de parâmetros entomológicos, fornecendo subsídios para medidas e as ações efetivas de controle. Foram realizadas, de 2009 a 2011, avaliações entomológicas, moleculares e epidemiológicas nos três polos base sentinelas para o PBEO \2013 Xitei (subpolo Ketaa e Watatase), Balawau (subpolo Maxapapi e Wanapiu) e Toototobi (subpolo Xiroxiropiu) para o monitoramento do impacto das estratégias de controle do PBEO/OEPA na transmissão da oncocercose no Brasil. Para análise dos parâmetros entomológicos foram realizadas capturas sistemáticas mensais de simulídeos, de 4-8 dias consecutivos de 7-18h Para análise molecular, grupos de até 50 fêmeas foram organizados de acordo com a espécie e outros parâmetros, separados em cabeças e corpos e cada grupo foi testado para a presença de DNA do parasito por PCR-ELISA. Do total de 74.397 simulídeos capturados (54% S. guianense; 40% S. incrustatum; 6% S. oyapockense), 16.971 foram coletados em Xitei/Watatase, 22.910 em Xitei/Ketaa, 2.301 em Balawaú/Maxapapi, 10.986 em Balawaú/Wanapiu e 21.229 de Toototobi. Do total de 1559 pools examinados (821 de Xitei, 302 de Balawaú e 436 de Toototobi), a prevalência estimada de S. guianense infectado foi de 1,0/2.000 simulídeos (95% LSIC \2013 2,2) no polo base Toototobi e 0,5/2.000 simulídeos (95% LSIC \2013 1,4) no polo base Balawaú. Simulim incrustatum e S. oyapockense não apresentaram nenhum pool considerado positivo. Neste estudo, o uso de diagnóstico molecular combinado com a identificação e aspectos bionômicos das espécies vetores e análises epidemiológicas possibilitou avaliar as informações sobre o estado da transmissão da oncocercose após a intervenção de 15 anos de tratamento com ivermectina. Com base nos parâmetros entomológicos de prevalência da infectividade (TI) e o potencial de transmissão anual (PTA) detectado abaixo do preconizado para as áreas sentinela foi possível verificar para período estudado a supressão da transmissão da oncocercose nas três áreas sentinela \2013 Xitei, Balawaú e Toototobi / Abstract: In Brazil, the onchocerciasis endemic area is restricted to the Amazon region, in the Yanomami area. The Brazilian Program to Eliminate Onchocerciasis (PBEO) adopts as the most important strategy to control the mass treatment of this population with Ivermectina, and so does the Onchocerciasis Elimination Program for the Americas (OEPA), as well as the other five endemic countries in Latin America - Mexico, Guatemala, Equador, Colombia and Venezuela. According to the strategies defined by the World Health Organization (WHO) this study was done aiming the availability of tools for the evaluation of the epidemiological status onchocerciasis based on entomologic parameters, offering subsidies for the study and effective controlling actions. Were conducted, from 2009 to 2011, entomological, molecular and epidemiological evaluations in three sentinels areas under surveillance of PBEO \2013 Xitei (Ketaa and Watatase), Balawau (Maxapapi and Wanapiu) and Toototobi (Xiroxiropiu) to monitor the PBEO/OEPA controlling strategies impact in the onchocerciasis transmission in Brazil. To analyze the entomological parameters, monthly, simulids captures from 4-8 consecutive days, between the hours of 7am\20136pm were done. For the molecular analysis, groups of up to 50 females were organized according to species and other parameters, separated in heads and bodies and each group was tested through PCR-ELISA, for the presence of the DNA parasite From a total of 74,397 captured simulids (54% S. guianense, 40% S. incrustatum; 3% S. oyapockense); 16,971 were captured from Xitei/ Watatase, 22,910 from Xitei/Ketaa, 2,301 from Balawau/Maxapapi, 10,986 from Balawau/Wanapiu and 21,229 from Toototobi. In a total of 1,559 pools examined (821 from Xitei, 302 from Balawau and 436 from Toototobi), the estimated prevalence of infected S. guianense was 1.0/2,000 simulids (95% L SIC\20132.2) in the Toototobi area, and 0.5/2,000, simulids (95% L SIC \2013 1.4) in the Balawau area. Simulium incrustatum and S. oyapockense did not present any pool considered positive. In this study, the use of molecular diagnostic combined with the identification and bionomic aspects of the vector species, and epidemiologic analysis, assisted in the evaluation of the Onchocerciasis transmission after 15 years of Ivermectina treatment, there are presenting an updated epidemiological profile of the disease in the Brazilian Amazon. Based on entomological parameters of prevalence of infectivity (IT) and the potential for transmission (PTA) detected below levels considered the sentinel areas was verified for the period studied the suppression of onchocerciasis transmission in the three sentinel areas\2013 Xitei, Balawaú e Toototobi

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No description available.

The occurrence of human Onchocerciasis has not previously been reported from the Anglo-Egyptian Sudan. It is my intention to show that the nematode Onchocerca Volvulus (Leuckart 1893) has a wide distribution in the southern Sudan, and that it, with its microfilariae, give rise to a variety of pathological conditions including grave ocular troubles, and that it is a parasite of serious economic importance, and not merely of academic interest. For the past six years I have been engaged on sleeping Sickness and Leprosy campaigns in the southern Sudan, and have come into close touch with the native population and learned much of their habits and customs, and got a working knowledge of some of their dialects. This experience has been invaluable in the necessarily rather interrupted researches into onchocerciasis, and has allowed one to pass fairer and more critical judgment on the often highly imaginative statements of natives. Microfilariae were often found in gland juice and blood smears during routine Trypanosomiasis inspections, and elephantiasis and hydroceles formed the basis of ones surgical practice, but the former were always imagined to be Microfilariae Wuchereria Bancrofii and the latter their sequelae. Some three years ago it was noticed that in certain arears an appreciable proportion of the young adult population were blind or had serious eye symptoms or complained of »night blindness». In 1932 Dr. Bryant, a colleague of mine working in Rumbek district described the disease as a retino-choroiditis and sometimes associated with signs of keratitis, iritis, iridocyclitis and synechiae. He pointed out that although "Simulium Damnosum", the vector, had not been found, the possibility of Onchocerca Volvulus as the cause must be borne in mind. Later, in 1933, he identified microfilariae of Onchocerca Volvulus in a case of hydrocele in the Heridi district. Since then a careful study of cases of elephantiasis, hydroceles, skin diseases, tumour formations, and of eye conditions has been made, the incidence and distribution of Onchocerca Volvulus roughly mapped out, and also the incidence of the insect vector. Other possible causal factors have been investigated, and a certain amount of experimental work done. Pathological specimens have been collected and examined by experts in London. The results of these investigations will be given and my arguments for considering Onchocerca Volvulus capable of causing those serious pathological changes set forth. A digest and historical review of the more important literature and a description of the parasite precedes the main Thesis.

Thèse--Sc. nat.--Rennes, 1979. / Bibliogr. p. 247-268.

Ivermectin (IVM) is the only safe drug for mass-treatment of onchocerciasis. IVM-resistance has been reported in gastrointestinal nematode parasites of animals. A reduction in response to IVM in Onchocerca volvulus could have significant consequences for the onchocerciasis control programs. Over the last few years, studies have reported genetic selection or reduced responses to IVM in some O. volvulus populations. The risk of a recrudescence of the disease was recently reported with the emergence of resistant adult parasite population in Ghana. It is important to understand the effects of IVM on O. volvulus populations to be able to identify genetic markers to follow IVM selection in the field. In this study, O. volvulus samples were derived from a clinical trial in Cameroon, in which patients were sampled before, and following three years (1994-1997) of IVM treatments. There were four treatment groups: 150mug/kg (1xp.a. or 4xp.a.) and 800mug/kg (1xp.a. or 4xp.a.). DNA from macrofilariae was genotyped for beta-tubulin and P-glycoprotein-like protein (PLP) gene, as well as two control genes and other loci. Reproductive organs of female worms were analyzed by microscopy. A correlation was established between the reproductive status of the female worms and beta-tubulin genotype with the beta-tubulin heterozygous female worms being less fertile than the homozygous female worms. This disadvantage in fertility seemed to disappear after repeated exposure with IVM. We have found evidence that repeated IVM treatment selects for specific alleles of beta-tubulin and PLP. We observed that IVM selection pressure was higher in the female worms than in the male worms. Additionally, loss of polymorphism and selection pressure were higher following thirteen three-monthly doses of IVM compared to annual doses of IVM. Moreover, we found evidence of excess of homozygosity in O. volvulus population, that may be caused by non-random mating and/or subdivision population, which may have implication for the control program. PLP and beta-tubulin genes appear to be promising DNA markers for field use to follow IVM selection. In this perspective, alternative control measures could be considered locally in regions where gene selection is apparent, reducing the likelihood that IVM resistance would develop further and spread.

Onchocerciasis, commonly known as "River blindness" is a disease affecting over 37 million people, worldwide. It is caused by the parasitic nematode Onchocerca volvulus and transmitted by the blackfly vector of the genus Simulium. The drug ivermectin (IVM) is the principal means of controlling the disease. As a result of recent reports on sub-optimal response to ivermectin and genetic selection in O. volvulus, we carried out a 21 month epidemiological study to investigate the response of O. volvulus to repeated rounds of IVM treatments in 2501 subjects from 19 Ghanaian communities that have received between 6 and 18 annual treatments and one IVM naive community. Skin microfilaria (mt) assessments were done before IVM treatment and at days 30, 90, 180 and 364 post-IVM treatment. At day 90 after the second IVM treatment, nodulectomies were carried out on 140 patients and embryogrammes constructed on female worms. We found IVM is still an effective microfilaricide, with efficacy of 98-100%. However, its effect on adult worm fertility has been reduced. Day 90 and 180 post-treatment showed significantly higher (p<0.05) skin mf repopulation of 7.1% to 53.9%, and >100% of pretreatment counts at day 364 post-treatment in four communities compared with the other six communities, which had <80% of pretreatment mf counts on day 364. From these results we classified the 10 communities into good IVM response (four communities), intermediate IVM response (two communities), poor IVM response (three communities), and the previously IVM naive community. Nodule and worm viability and worm densities were significantly higher (p<0.05) in the poor response communities compared with the good response communities, with the intermediate falling between the two. Embryogramme analysis showed significantly higher reproductive activity and output in worms from poor response communities with up to 41% of females having live stretched mf in utero compared with good response communities which had no intra-uterine stretched mf. These results show evidence of lack of sustained response of adult O. volvulus to IVM in the poor response communities, manifested as a rapid return to fertility after IVM treatment. We conclude that IVM resistance is emerging in onchocerciasis and is manifested as a loss of effect of IVM on suppression of parasite reproduction. / Beta tubulin isotype 1 gene has been shown to be linked to IVM treatment and selection in O. volvulus and veterinary nematodes. Genetic analysis of the full length genomic DNA sequence of beta-tubulin from worms obtained in the three IVM response categories and IVM naive community showed single nucleotide polymorphisms (SNPs) at 24 sites on the entire 3696 bp. The frequencies of eight SNPs were significantly different (p< 0.05) between the poor response communities and the good response/naive communities. Four SNPs, 183 T/G, 1188 T/C, 1309 CIT and 1545 A/G resulting in a genotype configuration GG/CC/TT/GG (183/1188/1309/1545) was significantly higher in the poor IVM response communities than the other communities. The phenotypic and genotypic analyses are consistent with a conclusion that IVM resistance has been selected. These four SNPs could be used to develop a genetic marker for early detection of IVM resistance. This study has shown for the first time that IVM resistance is emerging in Onchocerca volvulus and that there are genetic changes associated with IVM resistance which could be used for epidemiological monitoring for emerging resistance.

Onchocerca volvulus is a parasitic filarial nematode responsible for human onchocerciasis, a disease commonly known as "River Blindness". Although there are no well documented cases of ivermectin resistance in O. volvulus, reports of suboptimal responses to ivermectin have appeared. The purpose of this thesis was to examine genetic polymorphisms in O. volvulus and to determine whether there was genetic evidence of ivermectin selection on O. volvulus genes. Analysis of 17 genes from O. volvulus was undertaken in two populations of worms, either from ivermectin-naive patients or from patients who had been repeatedly treated with ivermectin annually. In 14 of the genes no differences in genetic polymorphism were found (although polymorphisms were identified). However, chi square analysis (chi2=0.05) indicated significant differences in allele frequencies for a P-glycoprotein, a beta-tubulin and a putative dyf=8 gene. Analysis of the O. volvulusbeta-tubulin alleles identified three amino acid substitutions in the H3 region with ivermectin selection. Microtubules play a key structural role in the formation of neurons, and in ivermectin-resistant Haemonchus contortus, amphidial neurons show distorted microtubule bundles. Polymerization and depolymerization assays of the recombinant O. volvulus beta-tubulin alleles showed interesting differences between the polymerized tubulin using the two different alleles. It is speculated that similar differences could cause the disorganization of the microtubules identified in the amphidial neurons in ivermectin resistant H. contortus. In addition to the coding mutations, a 24 bp deletion in the adjacent intron to the H3 was detected. A PCR diagnostic assay was developed to genotype individual macro- and microfilariae. Further analyses were conducted to investigate the possibility of a direct relationship between ivermectin and beta-tubulin. Data obtained from equilibrium dialysis experiments indicated that BODIPY FL ivermectin bound to purified O. volvulus alpha- and beta-tubulins. More interesting, non-fluorescent ivermectin and taxol competed with the BODIPY FL ivermectin. The work presented in this thesis provides evidence of genetic selection by ivermectin on O. volvulus and suggests a putative binding site for ivermectin on tubulin. These data provide novel information on ivermectin selection in O. volvulus and on the possible involvement of tubulin in ivermectin resistance.