p. 1-103 / Submitted by Santiago Fabio (fabio.ssantiago@hotmail.com) on 2013-05-08T18:50:57Z No. of bitstreams: 1 Diss_Sheila_Alvim5656565.pdf: 645345 bytes, checksum: d86f8f5b18f601785339bb6089fdefc4 (MD5) / Approved for entry into archive by Rodrigo Meirelles(rodrigomei@ufba.br) on 2013-05-11T15:36:48Z (GMT) No. of bitstreams: 1 Diss_Sheila_Alvim5656565.pdf: 645345 bytes, checksum: d86f8f5b18f601785339bb6089fdefc4 (MD5) / Made available in DSpace on 2013-05-11T15:36:48Z (GMT). No. of bitstreams: 1 Diss_Sheila_Alvim5656565.pdf: 645345 bytes, checksum: d86f8f5b18f601785339bb6089fdefc4 (MD5) Previous issue date: 2006 / Com o objetivo de avaliar a influência que a infecção por parasitas intestinais exerce sobre o estado antropométrico na infância, na cidade de Salvador-BA, foi desenvolvido um estudo seccional em uma amostra probabilística de 629 crianças entre 12 e 48 meses. Aplicou-se a técnica de regressão logística multivariada, tendo como variáveis dependentes os indicadores antropométricos peso por idade (P/I) e altura/comprimento por idade (A/I). A presença ou ausência de diferentes parasitas nas fezes (A. lumbricoides, T. trichiura, G. duodenalis), constituiu-se na variável independente. O parasita mais freqüente foi A. lumbricoides (23,1%), seguido de T. trichiura (16,5%) e G. duodenalis (13,5%). Entre as crianças infectadas a média de z-scores dos indicadores antropométricos foi menor do que a observada entre as crianças não infectadas. Dentre as infecções estudadas, apenas a giardíase esteve significativamente associada ao déficit de crescimento linear, incluindo quando a freqüência da coleta pública de lixo era inadequada (modificador de efeito). Observam-se associações com o déficit antropométrico, segundo o indicador P/I, na presença da giardíase quando a criança não era amamentada por mais de seis meses (OR=2,92; IC95%=1,62-5,24) e quando residia em domicílio cuja pavimentação da rua/calçada era inadequada (OR=3,04; IC95%=1,53-6,07), após ajuste por confundidores. Os resultados deste estudo evidenciaram que, em crianças com giardíase, a amamentação durante o período recomendado pela OMS constituiu-se em um fator de proteção para o estado antropométrico e que o efeito negativo das condições ambientais indesejadas referentes ao lixo urbano e à pavimentação sobre o estado antropométrico pode ser devido à provável mais elevada carga parasitária existente nas crianças infectadas por este protozoário, vivendo neste contexto insalubre. / Salvador

Submitted by Miriam Lucas (miriam.lucas@unioeste.br) on 2018-04-18T16:58:28Z No. of bitstreams: 2 Fernanda_Ferreira_de_Carvalho_2017.pdf: 1245761 bytes, checksum: f7d305fe3b4c55009b31a1ca0fd7ec70 (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) / Made available in DSpace on 2018-04-18T16:58:29Z (GMT). No. of bitstreams: 2 Fernanda_Ferreira_de_Carvalho_2017.pdf: 1245761 bytes, checksum: f7d305fe3b4c55009b31a1ca0fd7ec70 (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) Previous issue date: 2017-08-10 / Foz do Iguaçu has today more than 81 ethnicities, associated to a wide cultural diversity they contribute to the divergence of hygienic habits, resulting in precarious sanitary conditions. Fir this reason, the city government has been presenting several changes in public health area. The city is also located on a tri-border border area, about 7 km from Ciudad Del Este (Paraguay) and 10 km from Puerto Iguazu (Argentina). Its main economic product is Tourism, due to the relevance of the Iguazu Falls, considered one of the seven wonders of nature and Itaipu Hydroelectric Power Plant, which is on the border of Brazil and Paraguay. Because of the construction of the Hydroelectric Power Plant, in just 20 years, Foz do Iguaçu has grown by 383% in the total population of the city, generating a great job offer. Due to the increasing number of women entering the labor market, the Municipal Early Childhood Centers (CMEIs) have been a reality in the life of families, as this is the place where many children spend most of their time while mothers are at work. These institutions play an important role in the development of children as a whole, mainly referring to the intellectual, physical, social and psychological aspects. Children attending CMEIs are more susceptible to infections than those who are kept in their homes, which can be explained by the large interpersonal contact provided by the collective environments. To verify the prevalence of parasitic diseases among children attending CMEIs in Foz do Iguaçu and to analyze the possible relationship with the socioeconomic, environmental and hygienic scenario of children, parents and caregivers. This is a field research, with descriptive and quantitative approaches. The studied population consisted of all pre-school children aged from 3 to 5 years old, enrolled in CMEIs A and B, and parents and/or guardians working with sampling by convenience. The feces of the children were collected and then sent for analysis in the Environmental Laboratory of the Itaipu Hydroelectric Power Plant. In addition, a questionnaire was applied to the parents or guardians of the children, containing 19 closed questions, regarding the factors that contribute to the merge of parasitic diseases. Data were analyzed by the BioEstat 5.3® program and, the level of significance was considered at 5%. Results: A total of 82 fecal samples were analyzed from February to June 2016. Positive results for parasites were found in 38.8% of them. The following etiological agents were found: Giardia duodenalis (28,0%), Hymenolepis nana (3,6%), Ascaris lumbricoides (2,4%), Entamoeba coli (2,4%) and Enterobius vermicularis (2,4%). Regarding the degree of parasitism, 35.0% of the children presented monoparasitism and only 4% presented biparasitism, and there were no cases of polyparasitism. The parents and guardians of the parasitized children (32/82) received the antiparasitic medication prescription (metronidazole) and the medicament for the treatment of the children. In addition, they received guidance in order to prevent the emergence of new cases. The most affected gender by parasites was the male (20/41), followed by the female (12/41), (p<0.05) in the age range of 3 to 4 years (84.3%). The socioeconomic profile revealed that the level of schooling of mothers, complete secondary education (15/29 - p<0.05), and incomplete high school parents (13/24 - p>0.05), family income up to one Minimum wage (19/47 - p<0.05), residential structure with piped water and treated with chlorine (32/80 - p<0.05), waste disposal by means of cesspool (12/36 - p<0.05), may have influenced the average incidence of parasitism, as well as the aspects related to the habit of washing hands after diaper change (21/47 - p<0.05) and the health of the child, the taking of feces exams (19/45), abdominal pain (25/65), nausea (31/79), vomiting (31/78), weight loss (28/71) and presence of formed faeces (17/43) ) (P<0.05). However, the prevalence of parasitoses was related to the form of cleaning foods (fruits, and vegetables), only with water (23/60 - p<0.05), with children who do not have the habit of washing their hands with soap before meals and after using the toilet (6/15 - p>0.05). Conclusion: The results indicate an important prevalence of intestinal parasitoses in children who live on the border area, in the age range of 3 to 5 years old, mainly giardiasis. The presence of parasitic diseases in this population provides an epidemiological profile that can help in the construction of public health policies which are more appropriate to this social reality, in order to contribute to the improvement of health education of the population, especially the family members responsible for the children who are the most affected by the to intestinal parasites. Keywords: parasitic / Foz do Iguaçu presenta 81 etnias, con una amplia diversidad cultural, contribuyendo a la divergencia de hábitos higiénicos, resultando en precarias condiciones sanitarias. Como consecuencia, el municipio viene presentando diversas transformaciones en el área de la salud. Además, se encuentra en la triple frontera, aproximadamente siete kilómetros de Ciudad Del Este (Paraguay) y a 10 km de Puerto Iguazu (Argentina). Su economía se basa principalmente en el turismo, debido a las Cataratas del Iguazú, la cual es considerada como una de las siete maravillas de la naturaleza y la Usina Hidroeléctrica de Itaipú. En los últimos 20 años, debido a la construcción de la Usina Hidroeléctrica Itaipú, el municipio de Foz do Iguaçu tuvo un crecimiento poblacional del 383%, convirtiéndose además en unas de las principales fuente de empleo. En razón del número cada vez mayor de mujeres insertas en el mercado de trabajo, los Centros Municipales de Educación Infantil (CMEIs) han sido una realidad en la vida de las familias, por ser este el lugar donde muchos niños pasan la mayor parte de su tiempo. Estas instituciones desempeñan un papel importantísimo en el desarrollo integral de los niños, principalmente relacionados con los aspectos intelectual, físico, social y psicológico. Los niños que frecuentan los CMEIs son más susceptibles a las infecciones que las que se mantienen en sus hogares, lo que puede ser explicado por el gran contacto interpersonal propiciado por los ambientes colectivos. Verificar la prevalencia de parasitosis en niños matriculadas en los CMEIs de Foz do Iguaçu y analizar la posible relación con el escenario socioeconómico, ambiental e higiénico de los niños, de los padres y/o responsables. Materiales y Métodos: Se trata de una investigación de campo, descriptiva y de abordaje cuantitativo. La población del estudio se constituyó de todos los niños con edad preescolar comprendida entre los 3 y 5 años, matriculados en los CMEIs A y B, y de los padres y / o responsables donde se trabajó con el muestreo de conveniencia. Se realizó la recolección de examen parasitológico de heces en los niños matriculados en los CMEIs y posteriormente procesados en el Laboratorio Ambiental de la Usina Hidroeléctrica de Itaipú. Además, se aplicó un cuestionario a los padres o responsables de los niños, conteniendo 19 preguntas cerradas, referentes a los factores contribuyentes para el surgimiento de enfermedades parasitarias. Los datos fueron analizados por el programa BioEstat 5.3® y, considerado el nivel de significancia en el 5%. Se analizaron 82 muestras fecales en el período de febrero a junio de 2016, obteniéndose positividad para parásitos en (38,8%), siendo encontrados los siguientes agentes etiológicos: Giardia duodenalis (28,0%), Hymenolepis nana (3,6%), Ascaris lumbricoides (2,4%), Entamoeba coli (2,4%) y Enterobius vermicular (2,4%). En cuanto al grado de parasitismo, el 35,0% de los niños presentaron monoparasitismo y sólo el 4% presentó biparasitismo, no habiendo casos de poliparasitismo. Los padres y/o los responsables de los niños parásitos (32/82), recibieron la prescripción medicamentosa de antiparasitario (metronidazol) y el medicamento para el tratamiento de los niños. Además, recibieron orientaciones para prevenir el surgimiento de nuevos casos. El género, más propicio por parasitosis fue el masculino (20/41), seguido del femenino (12/41), (P<0,05) con rango de edad de 3 a 4 años (84,3%). El perfil socioeconómico reveló que el grado de escolaridad de las madres, enseñanza media completa (15/29 - p<0,05), y de los padres de secundaria incompleta (13/24 - p>0,05), ingreso familiar de hasta un año Salario mínimo (19/47 - p<0,05), estructura residencial con agua potable y tratada con cloro (32/80 - p<0,05), eliminación de los desechos por medio de fosa (12/36 - p<0,5), puede haber influido en la incidencia media de parasitismo, así como los aspectos relacionados con la higienización de las manos de los responsables de los niños después del cambio de pañales (21/47 - p<0,05) y de la salud del niño, realización De examen parasitológico de heces (19/45), dolor abdominal (25/65), náuseas (31/79), vómitos (31/78), pérdida de peso (28/71) y presencia de heces formadas (17/43) (P<0,05). Sin embargo, la prevalencia de parasitosis fue relacionada con la forma de higienización de alimentos (frutas, verduras y legumbres), solamente con agua (23/60 - p<0,05), con los niños que no tienen el hábito de lavarse las manos Con uso de jabón antes de las comidas y después del uso del sanitario (6/15 - p>0,05). Conclusión: Los resultados apuntan a una prevalencia importante de parasitosis intestinal en niños de la región de frontera con edad comprendida entre los 3 a 5 años, principalmente de giardiasis. La presencia de enfermedades parasitarias en esta población proporciona un perfil epidemiológico que puede ayudar en la construcción de políticas públicas de salud más adecuadas a esa realidad social, para contribuir a mejorar la educación en salud de la población, en especial a los familiares responsables por los niños afectados por parasitosis intestinales. / Foz do Iguaçu apresenta mais de 81 etnias, associadas a uma ampla diversidade cultural, contribuindo para a divergência de hábitos higiênicos, resultando em precárias condições sanitárias. Como decorrência, o município vem apresentando diversas transformações na área da saúde. Além disso, encontra-se na tríplice fronteira, a cerca de sete km de Ciudad Del Este (Paraguai) e 10 km de Puerto Iguazu (Argentina). Tem como principal base de economia o turismo, devido a Cataratas do Iguaçu, considerada umas das sete maravilhas da natureza e a Usina Hidrelétrica de Itaipu, que fica na divisa do Brasil com o Paraguai. Devido a construção da Usina Hidrelétrica, em apenas 20 anos, Foz do Iguaçu apresentou um crescimento de 383% de habitantes no total da população do município, gerando grande oferta de emprego. Em razão do número cada vez maior de mulheres inseridas no mercado de trabalho, os Centros Municipais de Educação Infantil (CMEIs) tem sido uma realidade na vida das famílias, por ser este o local onde muitas crianças passam a maior parte do seu tempo. Estas instituições têm um papel importantíssimo no desenvolvimento integral das crianças, principalmente relacionados aos aspectos intelectual, físico, social e psicológico. As crianças que frequentam os CMEIs são mais suscetíveis às infecções do que aquelas que são mantidas em suas residências, o que pode ser explicado pelo grande contato interpessoal propiciado pelos ambientes coletivos. Objetivou-se verificar a prevalência de parasitoses em crianças atendidas em CMEIs de Foz do Iguaçu e analisar a possível relação com o cenário socioeconômico, sanitário e higiênico das crianças, dos pais e ou responsáveis. Trata-se de uma pesquisa de campo, descritiva e de abordagem quantitativa. A população do estudo foi constituída de todas as crianças com idade pré-escolar de 3 a 5 anos, matriculadas nos CMEIs A e B, e dos pais e ou responsáveis onde se trabalhou com amostragem de conveniência. Foi realizada a coleta de exame parasitológico de fezes nas crianças matriculadas nos CMEIs e, posteriormente processadas no Laboratório Ambiental da Usina Hidrelétrica de Itaipu. Além disso, foi aplicado um questionário aos pais ou responsáveis das crianças, contendo 19 questões perguntas fechadas, referentes aos fatores contribuintes para o surgimento de doenças parasitárias. Os dados foram analisados pelo programa BioEstat 5.3® e, considerado o nível de significância em 5%. Foram analisadas 82 amostras fecais no período de fevereiro a junho de 2016, obtendo-se positividade para parasitos em 38,8%, sendo encontrados os seguintes agentes etiológicos: Giardia duodenalis (28,0%), Hymenolepis nana (3,6%), Ascaris lumbricoides (2,4%), Entamoeba coli (2,4%) e Enterobius vermicularis (2,4%). Em relação ao grau de parasitismos, 35,0% das crianças apresentaram monoparasitismo e apenas 4% apresentaram biparasitismo, não havendo casos de poliparasitismo. Os pais e ou responsáveis das crianças parasitadas (32/82), receberam a prescrição medicamentosa de antiparasitário (metronidazol) e o medicamento para o tratamento das crianças. Além disso, receberam orientações no intuito de se prevenir o surgimento de novos casos. O gênero, mais acometido por parasitoses foi o masculino (20/41), seguido do feminino (12/41), (p<0,05) com faixa etária de 3 a 4 anos (84,3%). O perfil socioeconômico revelou que o grau de escolaridade das mães, ensino médio completo (15/29 - p<0,05), e dos pais ensino médio incompleto (13/24 - p>0,05), renda familiar de até um salário mínimo (19/47 – p<0,05), estrutura residencial com água encanada e tratada com cloro (32/80 – p<0,05), eliminação dos dejetos por meio de fossa (12/36 - p<0,05), pode ter influenciado para a incidência média de parasitismo, assim como os aspectos relacionados a higienização das mãos dos responsáveis das crianças após a troca de fraldas (21/47 - p<0,05) e da saúde da criança, realização de exame parasitológico de fezes (19/45), dor abdominal (25/65), náuseas (31/79), vômitos (31/78), perda de peso (28/71) e presença de fezes formadas (17/43) (p<0,05). No entanto, a prevalência de parasitoses foi relacionada à forma de higienização de alimentos (frutas, verduras e legumes), somente com água (23/60 - p<0,05), com as crianças que não tem o hábito de lavar as mãos com uso de sabão antes das refeições e após o uso do sanitário (6/15 – p>0,05). Os resultados apontam uma prevalência importante de parasitoses intestinais em crianças fronteiriças na faixa etária de 3 a 5 anos, principalmente de giardíase. A presença de doenças parasitárias nesta população fornece um perfil epidemiológico que pode auxiliar na construção de políticas públicas de saúde mais adequadas a essa realidade social, de forma a contribuir na melhoria da educação em saúde da população, em especial dos familiares responsáveis das crianças atingidas pelas parasitoses intestinais.

A giardíase é uma doença entérica de alta prevalência particularmente nos países em desenvolvimento. Diferentes espécies foram descritas em função de seus hospedeiros. A G. duodenalis é a espécie que parasito não só o homem, mas também animais domésticos e silvestres. Recentemente, com a aplicação de técnicas moleculares, foi possível identificar sete diferentes agrupamentos sendo que cada um deles tem predileção por determinadas espécies de hospedeiro. Essa discriminação não é possível por meio da microscopia. A identificação do(s) agrupamento(s) presente(s) na população de determinada região pode ser de grande importância na elaboração de políticas de saúde pública já que revela o perfil de transmissão desse protozoário. Este estudo teve por objetivos verificar o grau de positividade de enteroparasitos, a contribuição de cada um deles nesse índice e o genótipo dos cistos de G. duodenalis presentes em amostras de fezes de indivíduos atendidos no Hospital Universitário da Universidade de São Paulo. Foram analisadas 6717 amostras com positividade para algum enteroparasito de 12,5%, sendo S.stercoralis e G. duodenais respectivamente o helminto e protozoário patogênico mais frequentes. A maioria dos casos de G. duodenalis genotipada era dos agrupamentos AII e B. / Giardiasis is a very prevalent enteric disease occurring mainly in developing countries. Different species have been described concerning their hosts. G. duodenalis parasites not only men but also domestic and wild animals. Recently molecular techniques have been used to identify seven different assemblages which parasites specific hosts. The microscopic analysis do not allows that discrimination. It can be useful to know the assemblage profile of specific population in order to provide some important information to built public health politicies once it reveals the transmition of the protozoan The aim of this study was to verify the enteroparasites infection degree, the individual contribution of each one in this picture and to genotyping G. duodenalis cysts from human faecal samples of individuals from University Hospital from Universty of São Paulo. So, 6717 faecal samples were analysed and 12,5% were positive for some enteroparasite and S.stercoralis and G. duodenais were the helminth e pathogenic protozoan more frequently found. Most of the cases of G. duodenalis were from the assemblages AII and B.

A giardíase é uma doença entérica de alta prevalência particularmente nos países em desenvolvimento. Diferentes espécies foram descritas em função de seus hospedeiros. A G. duodenalis é a espécie que parasito não só o homem, mas também animais domésticos e silvestres. Recentemente, com a aplicação de técnicas moleculares, foi possível identificar sete diferentes agrupamentos sendo que cada um deles tem predileção por determinadas espécies de hospedeiro. Essa discriminação não é possível por meio da microscopia. A identificação do(s) agrupamento(s) presente(s) na população de determinada região pode ser de grande importância na elaboração de políticas de saúde pública já que revela o perfil de transmissão desse protozoário. Este estudo teve por objetivos verificar o grau de positividade de enteroparasitos, a contribuição de cada um deles nesse índice e o genótipo dos cistos de G. duodenalis presentes em amostras de fezes de indivíduos atendidos no Hospital Universitário da Universidade de São Paulo. Foram analisadas 6717 amostras com positividade para algum enteroparasito de 12,5%, sendo S.stercoralis e G. duodenais respectivamente o helminto e protozoário patogênico mais frequentes. A maioria dos casos de G. duodenalis genotipada era dos agrupamentos AII e B. / Giardiasis is a very prevalent enteric disease occurring mainly in developing countries. Different species have been described concerning their hosts. G. duodenalis parasites not only men but also domestic and wild animals. Recently molecular techniques have been used to identify seven different assemblages which parasites specific hosts. The microscopic analysis do not allows that discrimination. It can be useful to know the assemblage profile of specific population in order to provide some important information to built public health politicies once it reveals the transmition of the protozoan The aim of this study was to verify the enteroparasites infection degree, the individual contribution of each one in this picture and to genotyping G. duodenalis cysts from human faecal samples of individuals from University Hospital from Universty of São Paulo. So, 6717 faecal samples were analysed and 12,5% were positive for some enteroparasite and S.stercoralis and G. duodenais were the helminth e pathogenic protozoan more frequently found. Most of the cases of G. duodenalis were from the assemblages AII and B.

An enhanced notified enteric disease surveillance trial began on 1 July 2007 and continued until 30 June 2008. The aim of the trial was to measure the quality, timeliness and completeness of data collected and submitted by a regional Public Health Service (PHS) to the Institute of Environmental Science and Research Limited (ESR), via the national disease database (EpiSurv) for notified cases of enteric diseases. The trial evaluated two different methods of data collection: postal questionnaires and telephone interviews. Telephone interview techniques were used to improve the contact rate, timeliness and completeness of data gathered from all notified cases of campylobacteriosis in the Manawatu, Horowhenua and Tararua regions. The target set for the project was to achieve a 95% contact rate with 90% full completion of all EpiSurv data fields. For all notified cases of campylobacteriosis a 97% contact rate was achieved in a time frame of between zero to 20 days (three day median) and completeness of all the EpiSurv case report fields ranged between 96 – 100% in the final data. Prior to the commencement of the study, between 1 July 2004 to 30 June 2005, MidCentral PHS (MCPHS) made contact with around 58% of all notified cases of campylobacteriosis and 77% of all other notified enteric disease cases1 . A short pre-screen mail questionnaire, with reply-paid envelope, was sent to all notified cases of cryptosporidiosis, giardiasis, salmonellosis and yersiniosis in the MCPHS regions. EpiSurv case report fields were completed using information supplied in the returned questionnaires. Return rate, timeliness, and completeness were compared with the telephone interview group. Fifty three percent of cases we attempted to contact via mail questionnaire responded within two to 63 days (six day median) and completeness of all the EpiSurv case report fields ranged between 81 – 100%. In addition, we monitored the newly introduced ESR Early Aberration Reporting System (EARS) flags for increased levels of disease compared to historical disease rates, and assessed its usefulness as a tool to identify potential outbreaks in the region. While no outbreaks that had not already been identified by PHS staff were found by monitoring the EARS system, EARS has become an important tool in the MCPHS for comparing our rates of disease with bordering PHSs. EARS also provided a good quick reference tool for media enquiries and the graphs produced in EARS have been well utilised as visual aids for training and seminars presented during the trial period. The results of the surveillance trial initiatives were compared to the rest of New Zealand (NZ) over the same time frame and with a comparable, medium-sized, PHS. While the results of the telephone interviews from the MCPHS trial were close to the comparable PHS, they were significantly higher than for the rest of NZ. The postal questionnaires achieved a lower contact rate than the comparable PHS but similar to the rest of NZ. However, the quality of data gathered in the returned MCPHS postal questionnaire was significantly higher in most fields. Additional analysis was undertaken which indicated that those cases living in higher deprivation and rural areas were less likely to respond to a postal questionnaire. An over-representation of common enteric disease notifications from rural areas in the MCPHS was also highlighted by our research. This trial has shown the effectiveness of utilising telephone interviews and telemarketing techniques for gathering timely and complete data for human enteric disease surveillance within the MCPHS. It has also demonstrated that a short pre-screen questionnaire can be effective in collecting good quality data needed to complete the standard EpiSurv case report form.

An enhanced notified enteric disease surveillance trial began on 1 July 2007 and continued until 30 June 2008. The aim of the trial was to measure the quality, timeliness and completeness of data collected and submitted by a regional Public Health Service (PHS) to the Institute of Environmental Science and Research Limited (ESR), via the national disease database (EpiSurv) for notified cases of enteric diseases. The trial evaluated two different methods of data collection: postal questionnaires and telephone interviews. Telephone interview techniques were used to improve the contact rate, timeliness and completeness of data gathered from all notified cases of campylobacteriosis in the Manawatu, Horowhenua and Tararua regions. The target set for the project was to achieve a 95% contact rate with 90% full completion of all EpiSurv data fields. For all notified cases of campylobacteriosis a 97% contact rate was achieved in a time frame of between zero to 20 days (three day median) and completeness of all the EpiSurv case report fields ranged between 96 – 100% in the final data. Prior to the commencement of the study, between 1 July 2004 to 30 June 2005, MidCentral PHS (MCPHS) made contact with around 58% of all notified cases of campylobacteriosis and 77% of all other notified enteric disease cases1 . A short pre-screen mail questionnaire, with reply-paid envelope, was sent to all notified cases of cryptosporidiosis, giardiasis, salmonellosis and yersiniosis in the MCPHS regions. EpiSurv case report fields were completed using information supplied in the returned questionnaires. Return rate, timeliness, and completeness were compared with the telephone interview group. Fifty three percent of cases we attempted to contact via mail questionnaire responded within two to 63 days (six day median) and completeness of all the EpiSurv case report fields ranged between 81 – 100%. In addition, we monitored the newly introduced ESR Early Aberration Reporting System (EARS) flags for increased levels of disease compared to historical disease rates, and assessed its usefulness as a tool to identify potential outbreaks in the region. While no outbreaks that had not already been identified by PHS staff were found by monitoring the EARS system, EARS has become an important tool in the MCPHS for comparing our rates of disease with bordering PHSs. EARS also provided a good quick reference tool for media enquiries and the graphs produced in EARS have been well utilised as visual aids for training and seminars presented during the trial period. The results of the surveillance trial initiatives were compared to the rest of New Zealand (NZ) over the same time frame and with a comparable, medium-sized, PHS. While the results of the telephone interviews from the MCPHS trial were close to the comparable PHS, they were significantly higher than for the rest of NZ. The postal questionnaires achieved a lower contact rate than the comparable PHS but similar to the rest of NZ. However, the quality of data gathered in the returned MCPHS postal questionnaire was significantly higher in most fields. Additional analysis was undertaken which indicated that those cases living in higher deprivation and rural areas were less likely to respond to a postal questionnaire. An over-representation of common enteric disease notifications from rural areas in the MCPHS was also highlighted by our research. This trial has shown the effectiveness of utilising telephone interviews and telemarketing techniques for gathering timely and complete data for human enteric disease surveillance within the MCPHS. It has also demonstrated that a short pre-screen questionnaire can be effective in collecting good quality data needed to complete the standard EpiSurv case report form.

An enhanced notified enteric disease surveillance trial began on 1 July 2007 and continued until 30 June 2008. The aim of the trial was to measure the quality, timeliness and completeness of data collected and submitted by a regional Public Health Service (PHS) to the Institute of Environmental Science and Research Limited (ESR), via the national disease database (EpiSurv) for notified cases of enteric diseases. The trial evaluated two different methods of data collection: postal questionnaires and telephone interviews. Telephone interview techniques were used to improve the contact rate, timeliness and completeness of data gathered from all notified cases of campylobacteriosis in the Manawatu, Horowhenua and Tararua regions. The target set for the project was to achieve a 95% contact rate with 90% full completion of all EpiSurv data fields. For all notified cases of campylobacteriosis a 97% contact rate was achieved in a time frame of between zero to 20 days (three day median) and completeness of all the EpiSurv case report fields ranged between 96 – 100% in the final data. Prior to the commencement of the study, between 1 July 2004 to 30 June 2005, MidCentral PHS (MCPHS) made contact with around 58% of all notified cases of campylobacteriosis and 77% of all other notified enteric disease cases1 . A short pre-screen mail questionnaire, with reply-paid envelope, was sent to all notified cases of cryptosporidiosis, giardiasis, salmonellosis and yersiniosis in the MCPHS regions. EpiSurv case report fields were completed using information supplied in the returned questionnaires. Return rate, timeliness, and completeness were compared with the telephone interview group. Fifty three percent of cases we attempted to contact via mail questionnaire responded within two to 63 days (six day median) and completeness of all the EpiSurv case report fields ranged between 81 – 100%. In addition, we monitored the newly introduced ESR Early Aberration Reporting System (EARS) flags for increased levels of disease compared to historical disease rates, and assessed its usefulness as a tool to identify potential outbreaks in the region. While no outbreaks that had not already been identified by PHS staff were found by monitoring the EARS system, EARS has become an important tool in the MCPHS for comparing our rates of disease with bordering PHSs. EARS also provided a good quick reference tool for media enquiries and the graphs produced in EARS have been well utilised as visual aids for training and seminars presented during the trial period. The results of the surveillance trial initiatives were compared to the rest of New Zealand (NZ) over the same time frame and with a comparable, medium-sized, PHS. While the results of the telephone interviews from the MCPHS trial were close to the comparable PHS, they were significantly higher than for the rest of NZ. The postal questionnaires achieved a lower contact rate than the comparable PHS but similar to the rest of NZ. However, the quality of data gathered in the returned MCPHS postal questionnaire was significantly higher in most fields. Additional analysis was undertaken which indicated that those cases living in higher deprivation and rural areas were less likely to respond to a postal questionnaire. An over-representation of common enteric disease notifications from rural areas in the MCPHS was also highlighted by our research. This trial has shown the effectiveness of utilising telephone interviews and telemarketing techniques for gathering timely and complete data for human enteric disease surveillance within the MCPHS. It has also demonstrated that a short pre-screen questionnaire can be effective in collecting good quality data needed to complete the standard EpiSurv case report form.

Heat shock proteins (Hsps) are a class of molecular chaperones which were first discovered as proteins up-regulated in response to heat stress in Drosophila. Later, it was found that these set of proteins get up-regulated as a general stress response associated with destabilization of native protein structures. Over a period of time, intricate involvement of Hsps in various biological processes has been well established. Heat shock protein 90 (Hsp90) is one of the important representative of this class of proteins. Hsp90 is an essential molecular chaperone which is evolutionarily conserved. It has a selective set of proteins to chaperone called as clients, which majorly include transcription factors and protein kinases. Through its interaction with its clients it modulates cell cycle, signal transduction, differentiation, development and evolution. Previous studies from Candida, Leishmania and Plasmodium have implicated Hsp90 to be involved in stage transition and growth. It is also critically involved in regulating growth of other protozoans such as Dictyostelium, Entamoeba and Trypanosoma. Thus, selective inhibition of Hsp90 has been explored as an intervention strategy against important human diseases such as cancer, malaria and other protozoan diseases. In Plasmodium falciparum, Hsp90 plays a critical role in stage transition. The parasite inside the human RBC develops from ring to trophozoite to schizont stage and inhibition of Hsp90 using specific pharmacological inhibitor arrests the growth of parasite at ring stage. In Dictyostelium, it has been observed that Hsp90 function is required for development. Inhibition of Hsp90 causes mound arrest and stops the cells from entering to its next developmental stage, fruiting bodies. In parallel, Hsp90 in Candida has been shown to be involved in morphogenesis. In nature Candida exists as a single cell yeast form and upon entry into the human host these yeast forms undergo morphogenesis to form virulent filamentous fungi. Inhibition of Hsp90 mimics temperature mediated morphogenesis. All together, these studies suggest that Hsp90 functions in a context dependent manner and each biological system explored has given new insights into the Hsp90 biology. Giardia lamblia, a protozoan parasite of humans and animals, is an important cause of diarrheal disease causing significant morbidity and also mortality in tropical countries. In the present study we focus on the biology of Hsp90 from Giardia lamblia. Giardia has a biphasic life cycle with infective cyst stage and pathogenic trophozoite stage. These cysts are present in the environment and enter mammalian host through oral route. They undergo a process called as excystation in the intestine giving rise to trophozoites. The trophozoites so formed colonize the upper part of the small intestine which causes the symptoms of giardiasis. Some of the trophozoites escape from the nutrition rich milieu of the upper part of small intestine to the lower part. In this region, trophozoites undergo a process called as encystation, wherein each trophozoite forms a cyst which escapes through faeces back into the environment. As seen in the life cycle of Giardia there are two major biological transitions, excystation and encystation; and till date no definitive player or pathway is known to regulate these processes. With the knowledge of Hsp90 playing an important role in similar biological transitions in other organisms we were encouraged to study role of Hsp90 in Giardia lamblia. Trans-splicing based generation of a full length Hsp90 in Giardia lamblia To understand the role of Hsp90, we first carried out sequence alignment of Hsp90 predicted ORFs in Giardia genome with yeast Hsp90. On alignment we observed that Hsp90 in Giardia is discontinuous and is annotated to be encoded by two different ORFs. Hsp90 in most organisms is coded by a single ORF with none to many cis-spliced introns. In a relatively intron poor organism G. lamblia, cytosolic Hsp90 is coded by two different ORFs separated by 777 kb in the genome. On multiple sequence alignment, we noticed that these two ORFs correspond to two independent regions of the Hsp90 protein. The ORFs are designated as hspN and hspC, containing the N-terminal and the C-terminal region of the protein respectively. We began our study by sequencing whole genome of Giardia lamblia clinical strain. Our genome sequencing confirmed the split nature of hsp90 and showed high ‘synteny’ between the other sequenced isolates. Using PCR based approach we have ruled out the possibility of having a full length gene in the genome. In contradiction to the genome result, we have observed a higher molecular weight protein in the lysate on proteomic analysis which was further confirmed by western blotting. The protein was observed to have a molecular weight of 80 kDa which could be a resultant of combination of two ORFs, suggesting the presence of a full length mRNA for Hsp90. PCR amplification using primers against both the fragments resulted in amplification of 2.1 kb product from the RNA pool of Giardia. Sequencing of this product showed that hspN and hspC were stitched together to form a mature messenger for full length Hsp90. In total our results suggest a post transcriptional process, trans-splicing, to be involved in the construction of Hsp90. The transition marked by this fusion coincides with the canonical GU¬AG splice site transitions as observed in other eukaryotes. Interestingly, a 26 nt near-complementary region was observed inside and upstream of hspN and hspC ORFs respectively. Put together these results suggest that the 26 nt complementary region acts as the positioning element to bring these two precursors in spatial proximity. With efficient spliceosomal activity these two precursor forms are trans-spliced to generate a full length cytosolic Hsp90 in Giardia. There are only four genes which have cis-spliced introns in the Giardia genome and the core components of the spliceosomal machinery are also present. The presence of canonical splice site in both the transcripts suggests that these transcripts are fused together by the spliceosomal machinery by the phenomenon of trans-splicing. The formation of full length Hsp90 RNA by its fragmented gene is the first example of trans-splicing in Giardia. To understand, are there any other genes which are also similarly trans-spliced we have carried out shotgun proteomic analysis of the total cell lysate obtained from Giardia trophozoites. Using Hsp90 as template, in our proteomic datasets, we have designed an algorithm for identification of additional trans-spliced gene products at the protein level. We have identified a total of 476 proteins of which hypothetical proteins constitute the major class followed by metabolic enzymes. We have compared the theoretical molecular weights for the identified proteins with the experimentally determined mass. Any discrepancy in the molecular mass was further analyzed and we assigned a gene to be potentially trans-spliced based on three criteria: if they were encoded by two or more different ORFs (loci), absence of a single full length counterpart and presence of splice sites with branch point and positional elements. Using this algorithm we were able to identify dynein as a potential candidate of trans-splicing reaction which was confirmed by the nucleotide sequence analysis of the predicted ORFs. Interestingly, dynein gene fragments were observed to be scattered on different chromosomes with minor splice sites unlike hsp90 genes. In vivo Expression of Hsp90 sub-fragments, HspN and HspC In the mature Hsp90 mRNA formed upon trans-splicing, 33 additional codons are present right between hspN and hspC sequences and they were acquired from the upstream region of hspC ORF. The 33 codons encode for an important region of Hsp90 which harbours the conserved catalytic “Arg” residue; suggesting that the full length Giardia Hsp90 (GlHsp90) formed could be an active ATPase. To confirm the same we have carried out in vitro characterization of trans-spliced Hsp90. Towards this, we have cloned, expressed and purified His tag-GlHsp90. As a first step, highly purified protein was used to assess its efficiency in binding to it cognate ligand, ATP, and the known inhibitors. Our binding studies show that GlHsp90 binds to ATP with a dissociation constant of 628 M and to its inhibitors, GA and 17AAG with 1.5 μM and 17.5 μM respectively. The bound ATP will be subsequently cleaved by Hsp90 which is an essential step in the chaperone cycle. As determined in our ATPase assay we observed that GlHsp90 hydrolyzes bound ATP with the catalytic efficiency of 4.4 × 10-5μM-1.min-1which confirms that Hsp90 generated upon trans-splicing is an active ATPase. The uniqueness of the hsp90 gene arrangement in Giardia posed a new question. Do these gene fragments also get translated? Our results suggest that HspN and HspC are poly¬adenylated. In order to determine the levels of these transcripts we performed qRT-PCR using primers specific to HspN, HspC and GlHsp90. We have observed that, in comparison with HspN transcript level, HspC and GlHsp90 transcripts are 15 and 75 folds higher respectively. To check for the presence of translation products of these transcripts, we have re-analyzed our proteomic datasets wherein we could identify peptides corresponding to HspN and HspC in their respective molecular weight region, 45 to 35 kDa. To confirm the proteomic data, western blot analysis was performed for trophozoite lysate on both 1D and 2D gels using anti-HspN antibody. Two specific bands (1D) / spots (2D) corresponding to the full length Hsp90 and HspN were identified. Gel filtration analysis revealed that HspN co¬eluted with full length Hsp90 thereby suggesting that both the proteins are in a same complex. With the background that HspN and HspC are present at the protein level, we asked if these fragments in combination can hydrolyse ATP. We reconstituted recombinant HspN and HspC in equimolar amounts and scored for the hydrolysis of ATP. However, no Pi release was observed. To determine whether HspN and HspC could modulate Hsp90 function, ATPase activity was monitored in the presence of HspN or HspC, in vitro. It was observed that ATPase activity was inhibited by both the fragments thus suggesting that HspN and HspC negatively regulate Hsp90 ATPase activity. Role of Hsp90 in Giardia encystation Giardia has a biphasic life cycle with proliferative trophozoites and latent cyst stage. In Giardia, in vitro encystation was established nearly two decades back by modulating the medium conditions. However, the mechanism and triggers underlying this transition are not well characterized. To understand whether Hsp90 has any role in this transition, in vitro conversion of trophozoites to cysts was achieved. The cysts obtained showed all the characteristic features of mature Giardia cyst with cyst wall protein 1 (CWP1) on the cyst wall and four nuclei as determined by immunofluorescence analysis. Further, the levels of Hsp90 in trophozoites were compared with mature cysts at both transcript and protein levels and it was found that cysts show more than 50% reduction in the level of Hsp90 in comparison with normal trophozoites. In accordance, exogenous inhibition of Hsp90 using 17AAG promoted the formation of cysts in vitro by 60 folds in a dose dependent manner; however, the window period of Hsp90 function compromise plays an important role in this process. Higher numbers of cysts were obtained from the cells treated with inhibitors during pre-encystation condition but inhibition of Hsp90 during encystation did not affect the formation of cysts, suggesting that Hsp90 down-regulation plays an important role during commitment towards encystation. To further show that cyst formation is a specific response to Hsp90 inhibition we have carried out encystation in the presence of metranidazole and from heat shocked cells; however, in both the conditions we did not observe any significant change in cyst formation, thus confirming that Hsp90 plays an important role during encystation in Giardia lamblia. Summary In Conclusion, Our study throws light on a unique aspect of Hsp90 biology in Giardia Lamblia, wherein the formation of the full length protein is dependent on a unique trans splicing reaction of its gene components representing different domains. We have also shown that HsP90 fragments, HspN and HspC, are also expressed in Trophozoites. Our in vitro data suggests that these fragments possibly regulate the function of Hsp90. Furthermore, the full length of Hsp90 plays an important role in stage transition in Giardia wherein inhibition of Hsp90 induces encystations. The study has opened many new avenues for research. Understanding the exact role of HspN and HspC in vivo will provide better appreciation for the evolution of such a complex biogenesis of an essential protein.

Made available in DSpace on 2016-01-26T12:51:36Z (GMT). No. of bitstreams: 1 juarezeliassantosjunior_dissert.pdf: 1690383 bytes, checksum: 9dd87174c2d25f09841337ec79d6968f (MD5) Previous issue date: 2010-11-25 / Conselho Nacional de Desenvolvimento Científico e Tecnológico / The parasite Giardia duodenalis, responsible for giardiasis, is commonly found in intestines of mammals, including man, currently considered to be important public health. Molecular research showed that G. duodenalis presents seven genotypes: A, B, C, D, E, F and G. Only genotypes A, which owns the subgenotypes A1 and A2 and B were detected in humans but also in other mammalian hosts. Objective: Evaluate the genotypic frequency of this parasite in humans in the Northwest region and to correlate the presence of diarrhea and their genotypes. Material and Method: Fecal samples were collected in the city of São José do Rio Preto (n = 150) and Araçatuba (n = 154), Northwest of São Paulo, Brazil. The parasitologic diagnosis was done by light microscopy and genotyping of &#946;-Giardin gene by PCR-RFLP. Results: The subgenotype A1 was the most prevalent, however, the subgenotype A2 more frequent in Araçatuba. Genotype B was not found. No significant result was observed between significant stool consistency and subgenotype detected. Conclusion: The distribution of these genotypes may be related to host-parasite interactions in several areas and may influence the clinical and epidemiologic each region. Keywords: 1. Molecular epidemiology; 2. Gene &#946;-giardin; 3. Giardiasis; 4. Northwest of São Paulo. / O parasito Giardia duodenalis, responsável pela giardíase, é comumente encontrado no intestino de mamíferos, inclusive o homem, sendo de importância em saúde pública. Pesquisas moleculares evidenciaram que Giardia duodenalis apresenta sete genótipos: A, B, C, D, E, F e G. Apenas o genótipo A, que possui os subgenótipos A1 e A2, e B foram detectados em humanos, mas também em outros hospedeiros mamíferos. Objetivo: Avaliar a freqüência genotípica deste parasito em humanos no Noroeste Paulista e correlacionar a presença deste enteropatógeno e seus genótipos à diarréia. Material e Método: Amostras fecais foram coletadas nos municípios de São José do Rio Preto (n=150 ) e Araçatuba (n=154), Noroeste do Estado de São Paulo, Brasil. O diagnóstico parasitológico foi feito por meio de microscopia ótica e a genotipagem do gene &#946;-giardina pela técnica de PCR-RFLP. Resultados: O subgenótipo A1 foi o mais prevalente, no entanto, o subgenótipo A2 mais freqüente em Araçatuba. O genótipo B não foi encontrado e nenhum resultado significativo foi observado entre a consistência das fezes e o subgenótipo detectado. Conclusão: A distribuição destes genótipos pode ser relacionada as interações parasito-hospedeiro em diversas áreas, podendo influenciar as características clínico-epidemiológicas de cada região.