Análise da variabilidade gênica de antígenos de T. cruzi com aplicação para o diagnóstico sorológico da doença de Chagas

A doença de Chagas é uma zoonose causada pelo protozoário flagelado Trypanosoma cruzi. Estima-se que 8 milhões de pessoas estão infectadas com o T. cruzi em todo o mundo, principalmente na América Latina. Testes tradicionais de diagnóstico estão sendo gradualmente substituídos por métodos inovadores. A utilização de antígenos recombinantes foi proposta nos anos 90, e várias combinações foram testadas com soros de pacientes com diferentes formas clínicas de diferentes regiões da América Latina. Apesar do ganho em especificidade, estes testes apresentaram menor sensibilidade, frustrando expectativas. Este estudo objetivou analisar a variabilidade genética dos genes KMP11 e 1F8 que codificam antígenos comumente utilizados em diagnóstico experimental. Cepas de T. cruzi pertencentes a diferentes sub-grupos taxonômicos e de diferentes regiões foram analisadas para avaliar o impacto da variação antigênica em testes de diagnóstico. Maximizando a sensibilidade, evitar reatividade cruzada com epítopos de outros agentes patogênicos deve permitir a concepção de melhores testes rápidos. Num primeiro passo, DNA genômico foi extraído das seguintes cepas: Dm28c, Colombiana, Y, 3663, 4167, LL014 e CL Brener e foi realizada a amplificação dos genes 1F8 e KMP11 que codificam antígenos a partir destas cepas. Em seguida foram realizadas a clonagem, sequenciamento, expressão e detecção dos antígenos recombinantes. Na etapa final, análises de estruturas secundárias e terciárias, a última apenas para o antígeno 1F8, visualizaram as diferenças nas sequências de aminoácidos obtidas a partir de sequenciamento de DNA. Os resultados apresentados neste estudo mostram que o antígeno KMP11 de T. cruzi possui uma similaridade na sequência de aminoácidos muito elevada com o T. rangeli, mostrando a necessidade de um mapeamento antigênico desta proteína em todos os tripanosomatídeos que apresentaram alta similaridade com o antígeno de T. cruzi, como o T. rangeli, para verificar a presença de epítopos específicos de T. cruzi. O antígeno 1F8 pode ser uma ferramenta útil no diagnóstico da doença de Chagas e que será necessário aprofundar os conhecimentos sobre os determinantes antigênicos para futuramente elaborar poli-epítopos sintéticos adaptados de um maior número possível de antígenos, obtendo a maior especificidade e sensibilidade nos testes de diagnóstico sorológico e de teste rápido da doença de Chagas. Este estudo representa um passo crucial para a otimização de antígenos recombinantes para o diagnóstico da doença de Chagas.

Sources of variability in the free-ball micro-scale abrasion test

In the 'free-ball' version of the micro-scale abrasion or ball-cratering test the rotating ball rests against a tilted sample and a grooved drive shaft. Tests under nominally identical conditions with different apparatus commonly show small but significant differences in measured wear rate. An indirect method has been developed and demonstrated for continuous on-line measurement of the coefficient of friction in the free-ball test. Experimental investigation of the effects of sample tilt angle and drive shaft groove width shows that both these factors influence the stability of the rotation of the ball, and the shape of the abrasive slurry pool, which in turn affect the coefficient of friction in the wear scar area and the measured wear rate. It is suggested that in order to improve the reproducibility of this method the geometry of the apparatus should be specified. For the apparatus used in this work with a steel ball of 25 mm diameter, a sample tilt angle of 60-75° and a shaft groove width of about 10mm provided the most stable ball motion and a wear rate which showed least variability. © 2004 Elsevier B.V. All rights reserved.

Mitochondrial gene sequences useful for species identification of western North Atlantic Ocean sharks

Molecular-based approaches for shark species identification have been driven largely by issues specific to the fishery. In an effort to establish a more comprehensive identification data set, we investigated DNA sequence variation of a 1.4-kb region from the mitochondrial genome covering partial sequences from the 12S rDNA, 16S rDNA, and the complete valine tRNA from 35 shark species from the Atlantic fishery. Generally, within-species variability was low in relation to interspecific divergence because species haloptypes formed monophyletic groups. Phylogenetic analyses resolved ordinal relationships among Carcharhiniformes and Lamniformes, and revealed support for the families Sphyrnidae and Triakidae (within Carcharhiniformes) and Lamnidae and Alopidae (within Lamniformes). The combination of limited intraspecific variability and sufficient between-species divergence indicates that this locus is suitable for species identification.

Variability in spawning frequency and reproductive development of the narrow-barred Spanish mackerel (Scomberomorus commerson) along the west coast of Australia

The narrow-barred Spanish mackerel (Scomberomorus commerson) is widespread throughout the Indo-West Pacific region. This study describes the reproductive biology of S. commerson along the west coast of Australia, where it is targeted for food consumption and sports fishing. Development of testes occurred at a smaller body size than for ovaries, and more than 90% of males were sexually mature by the minimum legal length of 900 mm TL compared to 50% of females. Females dominated overall catches although sex ratios within daily catches vary considerably and females were rarely caught when spaw n ing. Scomberomorus commerson are seasonally abundant in coastal waters and most of the commercial catch is taken prior to the reproductive season. Spawning occurs between about August and November in the Kimberley region and between October and January in the Pilbara region. No spawning activity was recorded in the more southerly West Coast region, and only in the north Kimberley region were large numbers of fish with spawning gonads collected. Catches dropped to a minimum when spawning began in the Pilbara region, when fish became less abundant in inshore waters and inclement weather conditions limited fishing on still productive offshore reefs. Final maturation and ovulation of oocytes took place within a 24-hour period, and females spawned in the afternoon-evening every three days. A third of these spawning females released batches of eggs on consecutive days. Relationships between length, weight, and batch fecundity are presented.

Maximum likelihood estimation of mortality and growth with individual variability from multiple length-frequency data

We consider estimation of mortality rates and growth parameters from length-frequency data of a fish stock and derive the underlying length distribution of the population and the catch when there is individual variability in the von Bertalanffy growth parameter L∞. The model is flexible enough to accommodate 1) any recruitment pattern as a function of both time and length, 2) length-specific selectivity, and 3) varying fishing effort over time. The maximum likelihood method gives consistent estimates, provided the underlying distribution for individual variation in growth is correctly specified. Simulation results indicate that our method is reasonably robust to violations in the assumptions. The method is applied to tiger prawn data (Penaeus semisulcatus) to obtain estimates of natural and fishing mortality.