Twelve isolates of Ichthyophthirius multifiliis were successfully established and maintained by serial passage through naïve carp, for a maximum of 39 laboratory cycles. The management system employed was such that large numbers of the parasite were available for all investigations. The ability to induce exit of immature trophonts through media incubation was used to confirm events in the initial stages of host colonisation. The normal course of primary infection was also established providing useful criteria for assessing success of the in vitro systems tested. Survival of both theronts and tomonts within selected monophasic media was investigated. Theronts in Eagles Minimum Essential medium (EMEM), survived and were viable for 120 hours, 72 hours longer than water controls. No further development of the theronts was observed. Tomonts also demonstrated an increased survival time in comparison to the controls with tomites surviving within the cyst for 22 days within EMEM-S media diluted 50:50 with sterile distilled water. Division of tomonts was identified as being precystic, post divisional cystic or cystic, and the frequency of such divisions was dependent upon dilution of media. Sterile viable theronts were recovered at 168h from tomonts that had been incubated within EMEM diluted 30:70 with distilled water. Delayed encystment was achieved by incubation in concentrated media, theront production being delayed for 96h, 72h later than seen in the aquatic environment. Cultured cell monolayers were used as associates within culture systems. Behaviour of theronts on introduction into the culture systems indicated recognition of the cultured tissue as potential host material, sustained contact of up to l20hours was observed between the introduced parasite and cells. However, no developmental markers were identified within the cultured parasite and no significant growth was achieved. Attempts to simulate the situation in vivo by use of multilayered systems and crude cell explants were also unsuccessful. Transmission electron microscopy of the parasite within a cell aggregate system was undertaken at daily intervals up to 120h providing evidence that the parasite was attempting to gain nutrients by phagocytosis. However, increased vacuolation of the parasite during the period of culture was clearly evident leading eventually to parasite death. The significance of the results is discussed in relation to the normal course of infection and the future promise of a long term culture method for this important pathogen.

Orientador: Cassiano Victória / Resumo: A raiva é uma zoonose transmitida pelo vírus do gênero Lyssavirus, e é considerada uma das enfermidades mais temidas no mundo devido à sua alta letalidade e inexistência de tratamento eficaz. Anualmente morrem mais de 50 mil pessoas por raiva em países subdesenvolvidos. O cão é o principal transmissor da raiva humana nos centros urbanos, responsável por 99% dos casos e 92% dos tratamentos pós-exposição. No Brasil, a raiva animal se apresenta de forma endêmica. A vacinação é o método mais eficiente de prevenção da raiva em cães e, por consequência, a proteção à população humana. No município de Botucatu, SP, há mais de 30 anos não são diagnosticados casos de raiva canina, devido às campanhas anuais de vacinação contra a raiva realizadas há 50 anos. Em 2010, o Ministério da Saúde preconizou o uso da vacina contra a raiva animal de cultivo celular nas campanhas de vacinação em massa. Este estudo teve como objetivo avaliar a resposta imunológica de cães que receberam apenas doses da vacina de cultivo celular contra o vírus da raiva. Para consolidação do tamanho amostral considerou-se uma porcentagem de 80% de adesão com consentimento livre e esclarecido, erro de estimação da ordem de 4% e nível de confiança de 95%. O título de anticorpos para a raiva foi detectado pelo Microteste Simplificado de Inibição de Fluorescência - SFIMT. Todas as discussões analíticas no plano estatístico foram realizadas no nível de significância de 5%. Observou-se que 59,12% (428/724) dos cães possuíam... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: Rabies is a zoonosis transmitted by the virus of the genus Lyssavirus and is considered one of the most feared diseases in the world due to its high lethality and lack of effective treatment. Annually more than 50,000 people die from rabies in underdeveloped countries. The dog is the main transmitter of human rabies, accounting for 99% of cases and 92% of post-exposure treatments. In Brazil, animal rabies presents itself endemic. Vaccination is the most effective method of preventing rabies in dogs and, consequently, protecting the human population. In the municipality of Botucatu, SP, for more than 30 years no cases of canine rabies have been diagnosed due to the annual campaigns of rabies vaccination applied 50 years. In 2010, the Ministry of Health advocated the use of the antirabies animal cell culture vaccine in mass vaccination campaigns. This study aimed to evaluate the immunological response of dogs that received only doses of the cell culture vaccine against the rabies virus. To consolidate the sample size, a percentage of 80% adherence with free and informed consent, an error of the order of 4% and a 95% confidence level were considered. The antibody titers for rabies were detected by the Simplified Fluorescence Inhibition Microtest - SFIMT. All statistical analyses were performed at a significance level of 5%. It was observed that 59.12% (428/724) of the dogs had a protective titer (≥0.5 IU / mL) and 40.88% (296/724) had not developed a protective titer of anti-rab... (Complete abstract click electronic access below) / Mestre