Molecular Characterization of Viruses Causing the Cassava Brown Streak Disease Epidemic in Eastern Africa

Cassava brown streak disease (CBSD) was described for the first time in Tanganyika (now Tanzania) about seven decades ago. Tanganyika (now Tanzania) about seven decades ago. It was endemic in the lowland areas of East Africa and inland parts of Malawi and caused by Cassava brown streak virus (CBSV; genus Ipomovirus; Potyviridae). However, in 1990s CBSD was observed at high altitude areas in Uganda. The causes for spread to new locations were not known.The present work was thus initiated to generate information on genetic variability, clarify the taxonomy of the virus or viruses associated with CBSD in Eastern Africa as well as to understand the evolutionary forces acting on their genes. It also sought to develop a molecular based diagnostic tool for detection of CBSD-associated virus isolates. Comparison of the CP-encoding sequences of CBSD-associated virus isolates collected from Uganda and north-western Tanzania in 2007 and the partial sequences available in Genbank revealed occurrence of two genetically distinct groups of isolates. Two isolates were selected to represent the two groups. The complete genomes of isolates MLB3 (TZ:Mlb3:07) and Kor6 (TZ:Kor6:08) obtained from North-Western (Kagera) and North-Eastern (Tanga) Tanzania, respectively, were sequenced. The genomes were 9069 and 8995 nucleotides (nt), respectively. They translated into polyproteins that were predicted to yield ten mature proteins after cleavage. Nine proteins were typical in the family Potyviridae, namely P1, P3, 6K1, CI, 6K2, VPg, NIa-Pro, NIb and CP, but the viruses did not contain HC-Pro. Interestingly, genomes of both isolates contained a Maf/HAM1-like sequence (HAM1h; 678 nucleotides, 25 kDa) recombined between the NIb and CP domains in the 3’-proximal part of the genomes. HAM1h was also identified in Euphorbia ringspot virus (EuRSV) whose sequence was in GenBank. The HAM1 gene is widely spread in both prokaryotes and eukaryotes. In yeast (Saccharomyces cerevisiae) it is known to be a nucleoside triphosphate (NTP) pyrophosphatase. Novel information was obtained on the structural variation at the N-termini of polyproteins of viruses in the genus Ipomovirus. Cucumber vein yellowing virus (CVYV) and Squash vein yellowing virus (SqVYV) contain a duplicated P1 (P1a and P1b) but lack the HC-Pro. On the other hand, Sweet potato mild mottle virus (SPMMV), has a single but large P1 and has HC-Pro. Both virus isolates (TZ:Mlb3:07 & TZ:Kor6:08) characterized in this study contained a single P1 and lacked the HC-Pro which indicates unique evolution in the family Potyviridae. Comparison of 12 complete genomes of CBSD-associated viruses which included two genomes characterized in this study, revealed genetic identity of 69.0–70.3% (nt) and amino acid (aa) identities of 73.6–74.4% at polyprotein level. Comparison was also made among 68 complete CP sequences, which indicated 69.0-70.3 and 73.6-74.4 % identity at nt and aa levels, respectively. The genetic variation was large enough for dermacation of CBSD-associated virus isolates into two distinct species. The name CBSV was retained for isolates that were related to CBSV isolates available in database whereas the new virus described for the first time in this study was named Ugandan cassava brown streak virus (UCBSV) by the International Committee on Virus Taxonomy (ICTV). The isolates TZ:Mlb3:07 and TZ:Kor6:08 belong to UCBSV and CBSV, respectively. The isolates of CBSV and UCBSV were 79.3-95.5% and 86.3-99.3 % identitical at nt level, respectively, suggesting more variation amongst CBSV isolates. The main sources of variation in plant viruses are mutations and recombination. Signals for recombination events were detected in 50% of isolates of each virus. Recombination events were detected in coding and non-coding (3’-UTR) sequences except in the 5’UTR and P3. There was no evidence for recombination between isolates of CBSV and UCBSV. The non-synonomous (dN) to synonomous (dS) nucleotide substitution ratio (ω) for the HAM1h and CP domains of both viruses were ≤ 0.184 suggesting that most sites of these proteins were evolving under strong purifying selection. However, there were individual amino acid sites that were submitted to adaptive evolution. For instance, adaptive evolution was detected in the HAM1h of UCBSV (n=15) where 12 aa sites were under positive selection (P< 0.05) but not in CBSV (n=12). The CP of CBSV (n=23) contained 12 aa sites (p<0.01) while only 5 aa sites in the CP gene of UCBSV were predicted to be submitted to positive selection pressure (p<0.01). The advantages offered by the aa sites under positive selection could not be established but occurrence of such sites in the terminal ends of UCBSV-HAMIh, for example, was interpreted as a requirement for proteolysis during polyprotein processing. Two different primer pairs that simultaneously detect UCBSV and CBSV isolates were developed in this study. They were used successfully to study distribution of CBSV, UCBSV and their mixed infections in Tanzania and Uganda. It was established that the two viruses co-infect cassava and that incidences of co-infection could be as high as 50% around Lake Victoria on the Tanzanian side. Furthermore, it was revealed for the first time that both UCBSV and CBSV were widely distributed in Eastern Africa. The primer pair was also used to confirm infection in a close relative of cassava, Manihot glaziovii (Müller Arg.) with CBSV. DNA barcoding of M. glaziovii was done by sequencing the matK gene. Two out of seven M. glaziovii from the coastal areas of Korogwe and Kibaha in north eastern Tanzania were shown to be infected by CBSV but not UCBSV isolates. Detection in M. glaziovii has an implication in control and management of CBSD as it is likely to serve as virus reservoir. This study has contributed to the understanding of evolution of CBSV and UCBSV, which cause CBSD epidemic in Eastern Africa. The detection tools developed in this work will be useful in plant breeding, verification of the phytosanitary status of materials in regional and international movement of germplasm, and in all diagnostic activities related to management of CBSD. Whereas there are still many issues to be resolved such as the function and biological significance of HAM1h and its origin, this work has laid a foundation upon which the studies on these aspects can be based.

Regulation of respiratory activity of brown adipose tissue mitochondria through changes in cytochromes.

Exposure of cold-acclimatized rats to heat (37 degrees C) for a short period decreased brown adipose tissue (BAT) mitochondrial substrate-dependent oxygen uptake and H2O2 generation. Both the concentration and substrate-dependent rate of cytochrome b reduction decreased as early as 3 h of heat exposure. These results identify cytochrome b as the locus of regulation of electron transport in BAT mitochondria under conditions of heat stress.

Microsomal redox systems in brown adipose tissue: high lipid peroxidation, low cholesterol biosynthesis and no detectable cytochrome P-450

The presence of redox systems in microsomes of brown adipose tissue (BAT) in cold exposed rats was investigated and compared with liver. BAT microsomes showed high activity of lipid peroxidation measured both by the formation of malondialdehyde (MDA) and by oxygen uptake. NADH and NADPH dependent cytochrome c reductase activity were present in both BAT and liver microsomes. Aminopyrine demethylase and aniline hydroxylase activities, the characteristic detoxification enzymes in liver microsomes could not be detected in BAT microsomes. BAT minces showed very poor incorporation of [1-14C]acetate and [2-14C]-mevalonate in unsaponifiable lipid fraction compared to liver. Biosynthesis of cholesterol and ubiquinone, but not fatty acids, and the activity of 3-hydroxy-3-methyl glutaryl CoA reductase appear to be very low in BAT. Examination of difference spectra showed the presence of only cytochrome b 5 in BAT microsomes. In addition to the inability to detect the enzyme activities dependent on cytochrome P-450, a protein with the characteristic spectrum, molecular size in SDS-PAGE and interaction with antibodies in double diffusion test, also could not be detected in BAT microsomes. The high activity of lipid peroxidation in microsomes, being associated with large oxygen uptake and oxidation of NADPH, will also contribute to the energy dissipation as heat in BAT, considered important in thermogenesis.

Evaluación de pollitas Hy-Line variedad Brown, en etapa de inicio a rompimiento de postura, bajo manejo tradicional

Se evaluó el comportamiento productivo de pollitas Hy- Line var. Brown, durante las primeras diecisiete semanas de desarrollo con la guía de manejo comercial para estas aves. Se utilizó un lote de 4,591 aves distribuidas en dos galeras. Se muestreó el 4% de la población de forma aleatoria y sin reemplazo, registrando peso, alimento consumido y mortalidad. Se realizaron los cálculos de consumo, uniformidad de peso, mortalidad de lote y se registraron las prácticas sanitarias de la granja. El consumo total obtenido fue un 15.6% inferior al reportado por la guía de manejo (803 g), viéndose afectado por cambios en el manejo del alimento, el que fue suministrado de acuerdo al peso vivo y no a la edad de las aves. El peso de las aves resultó 9.3% inferior al estipulado por la guía de manejo. La uniformidad del lote fue del 80% con una variación de ±10% igual al esperado para este tipo de aves. La mortalidad de la granja fue inferior al 0.04% expresado por la guía de manejo comercial. La aplicación del plan sanitario sufrió alteración en el tiempo de ejecución, sin embargo no tuvo efecto sobre la mortalidad de la granja. El manejo zootécnico de la granja está dentro de lo esperado en granjas de pequeños productores avícolas en condiciones tropicales.

Classificação de projetos de lei relacionados à saúde apresentados na Câmara dos Deputados entre 2001 e 2004

Consultoria Legislativa - Área XVI - Saúde Pública, Sanitarismo.

Classificação de projetos de lei relacionados à saúde apresentados na Câmara dos Deputados em 2001

Propõe o 'desenvolvimento de uma metodologia para classificação de projetos de lei relacionados à saúde e a demonstração de sua utilidade por meio de sua aplicação no conjunto de projetos de lei ordinária apresentados em 2001'.

Classificação de documentos sigilosos e regulamentação da liberdade de informação

Faz uma análise sobre a classificação e utilização da informação sigilosa na Suécia, Finlândia, Norugega, Dinamarca, Grã-Bretanha e Estados Unidos.

Sobre a classificação dos protetores solares na categoria de medicamentos

Consultoria Legislativa - Área XVI - Saúde Pública, Sanitarismo.

Proposição, aplicação e avaliação de um método de classificação temática em bases de dados textuais indexadas com auxílio de vocabulários controlados

Descreve a proposição, a aplicação e a avaliação de um método de classificação temática em uma base de dados com discursos proferidos por deputados federais no plenário da Câmara dos Deputados da República Federativa do Brasil entre outubro de 2000 e outubro de 2002, indexada com auxílio de um vocabulário controlado.

Manual para classificação de proposições legislativas relacionadas à saúde

Objetiva oferecer uma metodologia que permita a classificação de proposições legislativas relacionadas à saúde, que tramitam no Congresso Nacional, facilitando estudos em áreas como a política de saúde e a ciência política.

O Plano de Classificação Funcional da Câmara dos Deputados como artefato do modelo de arquitetura da informação

Analisa o Plano de Classificação Funcional da Câmara dos Deputados (PFC-CD) com a finalidade de verificar sua usabilidade como artefato do modelo de Arquitetura da Informação, a ser implementado por essa instituição.

Classificação Ranganathiana da tabela de tipos de proposição do Sistema de Informação Legislativa da Câmara dos Deputados : uma proposta metodológica

Aborda os problemas estruturais e informacionais da Tabela de Tipos de Proposição do Sistema de Informação Legislativa, o Sileg, da Câmara dos Deputados, relacionada à classificação dos tipos documentais daquela instituição, e utilizada, como referência, no registro (entrada de dados) e na recuperação de informações, ditas legislativas. Discute os fundamentos da Arquitetura da Informação, como disciplina da Ciência da Informação, para a organização da Informação Legislativa, especialmente, em relação à significação, à representação informacional e à teoria de classificação. Oferece uma definição para Informação Legislativa, que possui limitação material e formal, e aborda os antecedentes históricos de sua disponibilização à sociedade brasileira, em cumprimento aos princípios constitucionais da publicidade e transparência dos atos dos órgãos públicos e do direito de o cidadão brasileiro ter acesso e receber, desses órgãos públicos, informações de seu interesse particular. Aplica e comprova a viabilidade de utilização da teoria de classificação facetada, de Ranganathan, no estudo e na reorganização da Tabela de Tipos de Proposição do Sileg, que contemple as características multifacetada das proposições legislativas.

Development of kelp rockfish, Sebastes atrovirens (Jordan and Gilbert 1880), and brown rockfish, S. auriculatus (Girard 1854), from birth to pelagic juvenile stage, with notes on early larval development of black-and-yellow rockfish, S. chrysomelas (Jordan and Gilbert 1880), reared in the laboratory (Pisces: Sebastidae).

Larval kelp (Sebastes atrovirens), brown (S. auriculatus), and blackand-yellow (S. chrysomelas) rockfish were reared from known adults, to preflexion stage, nine days after birth for S. chrysomelas, to late postflexion stage for S. atrovirens, and to pelagic juvenile stage for S. auriculatus. Larval S. atrovirens and S. chrysomelas were about 4.6 mm body length (BL) and S. auriculatus about 5.2 mm BL at birth. Both S. atrovirens and S. auriculatus underwent notochord flexion at about 6–9 mm BL. Sebastes atrovirens transform to the pelagic juvenile stage at about 14–16 mm BL and S. auriculatus transformed at ca. 25 mm BL. Early larvae of all three species were characterized by melanistic pigment dorsally on the head, on the gut, on most of the ventral margin of the tail, and in a long series on the dorsal margin of the tail. Larval S. atrovirens and S. auriculatus developed a posterior bar on the tail during the flexion or postflexion stage. In S. atrovirens xanthic pigment resembled the melanistic pattern throughout larval development. Larval S. auriculatus lacked xanthophores except on the head until late preflexion stage, when a pattern much like the melanophore pattern gradually developed. Larval S. chrysomelas had extensive xanthic pigmentation dorsally, but none ventrally, in preflexion stage. All members of the Sebastes subgenus Pteropodus (S. atrovirens, S. auriculatus, S. carnatus, S. caurinus, S. chrysomelas, S. dalli, S. maliger, S. nebulosus, S. rastrelliger) are morphologically similar and all share the basic melanistic pigment pattern described here. Although the three species reared in this study can be distinguished on the basis of xanthic pigmentation, it seems unlikely that it will be possible to reliably identify field-collected larvae to species using traditional morphological and melanistic pigmentation characters. (PDF file contains 36 pages.)