Rickettsia typhi IN RODENTS AND R. felis IN FLEAS IN YUCATÁN AS A POSSIBLE CAUSAL AGENT OF UNDEFINED FEBRILE CASES

Rickettsia typhi is the causal agent of murine typhus; a worldwide zoonotic and vector-borne infectious disease, commonly associated with the presence of domestic and wild rodents. Human cases of murine typhus in the state of Yucatán are frequent. However, there is no evidence of the presence of Rickettsia typhi in mammals or vectors in Yucatán. The presence of Rickettsia in rodents and their ectoparasites was evaluated in a small municipality of Yucatán using the conventional polymerase chain reaction technique and sequencing. The study only identified the presence of Rickettsia typhi in blood samples obtained from Rattus rattus and it reported, for the first time, the presence of R. felis in the flea Polygenis odiosus collected from Ototylomys phyllotis rodent. Additionally, Rickettsia felis was detected in the ectoparasite Ctenocephalides felis fleas parasitizing the wild rodent Peromyscus yucatanicus. This study’s results contributed to a better knowledge of Rickettsia epidemiology in Yucatán.

Expressão em Escherichia coli de antigénios do Cell fusing agent virus (Flaviviridae: Flavivirus) como proteína de fusão

RESUMO: O Cell Fusing Agent Vírus (CFAV), considerado como o primeiro “flavivírus específicos de insectos” (ISF), parece estar exclusivamente adaptado aos seus hospedeiros, não replicando em células de vertebrados. Apesar de ter sido identificado há mais de três décadas (1975), a verdade é que muito pouco se conhece sobre a sua biologia. Dado o seu parentesco filogenético com alguns outros flavivírus encontrados naturalmente em mosquitos de diferentes géneros colhidos em diferentes regiões do globo, este vírus poderá ser usado como modelo para o estudo de ISF. No entanto, necessitam do desenvolvimento de ferramentas básicas, tais como clones moleculares ou baterias de soros contendo anticorpos que reconheçam uma ou mais proteínas codificadas pelo genoma viral, produzidas, por exemplo, a partir de antigénios virais produzidos de forma recombinante. Com este trabalho pretendeu-se a optimização de protocolos que permitiram a expressão e purificação parcial de quatro proteínas [duas proteínas estruturais (C e E) e duas não estruturais (NS3hel e NS5B)] do CFAV em E. coli, todas elas produzidas como proteínas de fusão com “caudas” (tags) de hexahistidina nos seus extremos carboxilo. Para a expansão do CFAV foram utilizadas células Aedes albopictus (C6/36). Após a realização da extracção do RNA viral e a obtenção de cDNA, procedeu-se amplificação, por RT-PCR, das regiões codificantes das proteínas C, E, NS3hel e NS5B, utilizando primers específicos. Os quatro fragmentos de DNA foram independentemente inseridos no vector pJTE1.2/blunt usando E. coli NovaBlue como hospedeira de clonagem e, posteriormente, inseridos em vectores de expressão pET-28b e pET-29b usando E. coli BL21(DE3)pLysS e Rosetta(DE3)pLysS como hospedeiras de expressão. Após da indução, expressão e purificação das proteínas recombinantes C, E, NS3hel e NS5B, foi confirmada a autenticidade destas proteínas produzidas através do método Western Blot com um anticorpo anti-histidina. --------- ABSTRACT: The Cell Fusing Agent virus (CFAV) considered as the first "insect- specific flavivirus" (ISF) and seems to be uniquely adapted to their hosts, not replicating in vertebrate cells. Although it has been known for more than three decades (1975), the truth is very little is known about its biology. Given its close phylogenetic relationship with other flavivirus naturally circulating in various genera of mosquitoes collected from different regions of the globe, this virus could be used as a model for the study of ISF. However, such studies require the development of experimental basic tools, such as molecular clones or serum batteries containing antibodies that recognize one or more proteins encoded by the viral genome, produced, for example, from viral antigens recombinant produced. In this work, we carried out the optimization of protocols that allowed the expression and partial purification of four proteins [two structural proteins (C and E) and two nonstructural proteins (NS3hel and NS5B)] CFAV in E. coli as fusion protein for c-terminal hexahistidine tags. For the expansion of the CFAV we used Aedes albopictus (C6/36) cells. After completion of the viral RNA extraction and cDNA obtained, amplification of the coding regions of the C, E, NS5B and NS3hel proteins was carried out by RT-PCR using specific primers. The four DNA fragments were independently inserted into the vector pJTE1.2/blunt using E. coli NovaBlue as cloning host and then inserted into expression vectors pET-28b and pET-29b using E. coli BL21(DE3)pLysS and Rosetta(DE3)pLysS as expression host. After induction, expression and purification of recombinant C, E, NS3hel and NS5B proteins Western Blot analyses with an anti-histidine antibody confirmed the authenticity of these proteins produced.

HUMAN TOXOPLASMOSIS OUTBREAKS AND THE AGENT INFECTING FORM. FINDINGS FROM A SYSTEMATIC REVIEW

SUMMARY Toxoplasmosis, a worldwide highly prevalent zoonotic infection, is transmitted either by the oocysts, from water and soil, or the tissue cysts, in raw or undercooked infected meat, of Toxoplasma gondii. An ongoing debate is whether there are differences between the clinical and epidemiological characteristics of the outbreaks due to one or the other infective form of the agent. We performed a systematic review, recovering 437 reported outbreaks of which 38 were selected. They were complete reports containing ascribed Toxoplasma infecting form, and clinical and demographic data. There was no gender or age group selection in the outbreaks, which were described more often in the Americas. A large number of individuals were affected when oocysts, associated with soil and water contaminated with cat feces, were considered the transmission source. Onset of symptoms occurred early when the infection was ascribed to meat tissue cysts (11.4 ± 6.7 days) with sharpened temporal distribution of cases, while a broader and prolonged appearance of new cases was observed when oocysts in water were the source of the infection (20 ± 7 days, p < 0.001). Such information may be useful in the design and implementation of control strategies.

Leishmania infantum AS A CAUSATIVE AGENT OF CUTANEOUS LEISHMANIASIS IN THE STATE OF MATO GROSSO DO SUL, BRAZIL

Cutaneous leishmaniasis is caused by different species of theLeishmania genus. Leishmania(Leishmania) infantum, causing cutaneous leishmaniasis, has been described in patients living in areas where visceral leishmaniasis is endemic. In this study, it was possible to characterize this species in seven slides from cutaneous tissue imprints from patients with cutaneous leishmaniasis in the State of Mato Grosso do Sul, Brazil.

An attempt to control schistosomiasis mansoni in an endemic area by the use of Hycanthone as chemotherapeutic agent

Hycanthone in a single dose of 3.3 mg/kg of body weight was used to treal mansoni schistosomiasis in 597 persons (83%) of the population of the endemic vilage of Canabrava. Ninety two patients received a 2nd course 14 months later. There was one death em the 2nd treatment. The cure rate after one stool examination was smaller after the 2ni treatment in comparison with the first one. Re-infections did not occur imediately after the treatment. Three years later the prevalence of persons passing eggs through one stool examination was 19% compared to the 46.3% before the treatment. Adults are more resistant to the re-infections than younger.

Customer Lifetime Value: a framework for application in the insurance industry - Building a Business Process to Generate and Maintain an Automatic Estimation Agent

Research Project submited as partial fulfilment for the Master Degree in Statistics and Information Management

An agent based architecture for material handling systems

Dissertação para obtenção do Grau de Mestre em Engenharia Electrotécnica e de Computadores

Lightweight robust behavior industrial agent methodology

Dissertação para obtenção do Grau de Mestre em Engenharia Electrotécnica e de Computadores

Novel ready-to-eat mango product using gellan gum as gelling agent: physico-chemical, microbial and sensory characteristics

Este trabalho foi efectuado com o apoio da Universidade de Lisboa, Instituto Superior de Agronomia com o Centro de Engenharia dos Biossistemas (CEER

Agent-based parking occupancy simulation

The existing parking simulations, as most simulations, are intended to gain insights of a system or to make predictions. The knowledge they have provided has built up over the years, and several research works have devised detailed parking system models. This thesis work describes the use of an agent-based parking simulation in the context of a bigger parking system development. It focuses more on flexibility than on fidelity, showing the case where it is relevant for a parking simulation to consume dynamically changing GIS data from external, online sources and how to address this case. The simulation generates the parking occupancy information that sensing technologies should eventually produce and supplies it to the bigger parking system. It is built as a Java application based on the MASON toolkit and consumes GIS data from an ArcGis Server. The application context of the implemented parking simulation is a university campus with free, on-street parking places.

Cryptococcus gattii molecular type VGII as agent of meningitis in a healthy child in Rio de Janeiro, Brazil: report of an autochthonous case

Cryptococcus gattii causes meningoencephalitis in immunocompetent hosts, occurring endemically in some tropical and subtropical regions. Recently, this fungus was involved in an outbreak in Vancouver Island and British Columbia (Canada). In this temperate region, the VGII type is predominant. The paper describes an autochthonous case of meningoencephalitis by C. gattii VGII in a previously health child in Rio de Janeiro, considered nonendemic region of Brazil. The fungus was identified by biochemical tests and the molecular type was determined by URA5-RFLP. The present report highlights the need for clinical vigilance for primary cryptococcal meningitis in nonendemic areas.

Integration of a mobile autonomous robot in a surveillance multi-agent system

This dissertation aims to guarantee the integration of a mobile autonomous robot equipped with many sensors in a multi-agent distributed and georeferenced surveillance system. The integration of a mobile autonomous robot in this system leads to new features that will be available to clients of surveillance system may use. These features may be of two types: using the robot as an agent that will act in the environment or by using the robot as a mobile set of sensors. As an agent in the system, the robot can move to certain locations when alerts are received, in order to acknowledge the underlying events or take to action in order to assist in resolving this event. As a sensor platform in the system, it is possible to access information that is read from the sensors of the robot and access complementary measurements to the ones taken by other sensors in the multi-agent system. To integrate this mobile robot in an effective way it is necessary to extend the current multi-agent system architecture to make the connection between the two systems and to integrate the functionalities provided by the robot into the multi-agent system.