The dynamics of RNA participation in the vitellogenesis of Rhipicephalus sanguineus ticks Latreille 1806 (Acari:Ixodidae). I. Nucleoli or Cajal bodies?

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

The dynamics of RNA participation in the vitellogenesis of Rhipicephalus sanguineus ticks Latreille 1806 (Acari: Ixodidae). I. Nucleoli or Cajal bodies?

To understand the morphological and histological aspects of internal systems of ticks has become important matter since these arthropods have an impact in the areas of the economy and public health. In this context, this study has provided morphological data on female germinative cells of Rhipicephalus sanguineus ticks, ectoparasites of dogs that maintain a close relationship with human on a daily basis. Oocytes of engorged females were analyzed, through the PAS reaction (detection of polysaccharides) counterstained by methyl green (detection of RNA) revealing information that allowed to infer for the first time the presence of Cajal bodies, in the germinal vesicles (nuclei) of developing oocytes, as well as showing how the RNA and the polysaccharides are involved in the dynamics of the vitellogenesis in this species. [copyright] 2010 Elsevier Ltd. All rights reserved.

Study of the nucleolar cycle and ribosomal RNA distribution during meiosis in triatomines (Triatominae, Heteroptera)

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Evidence supporting the recognition of Cole latent virus as a distinct carlavirus

Some molecular properties are described of Cole latent virus (CoLV), hitherto designated a tentative species of the Carlavirus genus. CoLV genomic RNA (Ribonucleic acid) of 8.3 Kb is polyadenylated. Two unencapsidated polyadenylated subgenomic RNAs (2.6 and 1.3 Kb) and three double-stranded RNAs (dsRNAs) (8.3, 2.6 and 1.3 Kbp), which are twice the size of the genomic and subgenomics ssRNAs, are produced in CoLV-infected plants, two additional dsRNAs (7.2 and 6.3 Kbp) were also detected plant extracts. By using a Carlavirus specific primer and a CoLV cDNA, a-3'-terminus fragment of 116 bp was amplified; it had homology with the carlaviruses Potato virus M (62%)., Hop latent virus (37%) and Blueberry scorch virus (36%) but no significant homology with 11 other carlaviruses. These results support the classification of CoLV as a distinct species of the Carlavirus genus.

Propriedades moleculares de um isolado brasileiro do Southern bean mosaic virus

Um isolado do Southern bean mosaic virus (SBMV), gênero Sobemovirus, encontrado em feijoeiro (Phaseolus vulgaris) no Estado de São Paulo, foi purificado e algumas de suas propriedades moleculares determinadas. As partículas virais apresentam diâmetro de 28-30 nm e proteína capsidial com massa molecular estimada em 30 kDa. Das partículas virais foi extraído RNA de vários tamanhos (4,2 Kb, 3,1 Kb, 2,65 Kb, 2,15 Kb, 1,64 Kb, 1,36 Kb e 1,0 Kb) sendo aquele de 4,2 Kb o RNA genômico e o de 1,0 Kb supostamente um subgenômico que codifica para a proteína capsidial. Ácidos ribonucleicos de mesmo tamanho foram também detectados in vivo, indicando estar associados à replicação viral. Na análise do RNA de fita dupla (dsRNA), somente duas espécies foram detectadas (4,2 Kpb e 1,0 Kpb) correspondendo às formas replicativas do RNA genômico e do subgenômico para proteína capsidial. Os resultados indicam que somente estes dois RNA são replicados por meio de formas replicativas (RFs), enquanto os demais devem ser formados talvez por iniciação interna da fita negativa do RNA genômico. O SBMV-B SP apresentou propriedades moleculares análogas àquelas do SBMV descrito na América do Norte.

Label-free DNA detection of hepatitis C virus based on modified conducting polypyrrole films at microelectrodes and atomic force microscopy tip-integrated electrodes

We present a new strategy for the label-free electrochemical detection of DNA hybridization for detecting hepatitis C virus based on electrostatic modulation of the ion-exchange kinetics of a polypyrrole film deposited at microelectrodes. Synthetic single-stranded 18-mer HCV genotype-1-specific probe DNA has been immobilized at a 2,5-bis(2-thienyl)-N-(3-phosphoryl-n-alkyl)pyrrole film established by electropolymerization at the previously formed polypyrrole layer. HCV DNA sequences (244-mer) resulting from the reverse transcriptase-linked polymerase chain reaction amplification of the original viral RNA were monitored by affecting the ion-exchange properties of the polypyrrole film. The performance of this miniaturized DNA sensor system was studied in respect to selectivity, sensitivity, and reproducibility. The limit of detection was determined at 1.82 x 10(-21) mol L-1. Control experiments were performed with cDNA from HCV genotypes 2a/c, 2b, and 3 and did not show any unspecific binding. Additionally, the influence of the spacer length of 2,5-bis(2-thienyl)-N-(3-phosphoryl-n-alkyl)pyrrole on the behavior of the DNA sensor was investigated. This biosensing scheme was finally extended to the electrochemical detection of DNA at submicrometer-sized DNA biosensors integrated into bifunctional atomic force scanning electrochemical microscopy probes. The 18-mer DNA target was again monitored by following the ion-exchange properties of the polypyrrole film. Control experiments were performed with 12-base pair mismatched sequences.

Improvement in RNA extraction from S. cerevisie by optimization in the autolysis and NH3 hydrolysis

The optimization of autolysis of Saccharomyces cerevisiae from brewery was studied aiming at the maximum ribonucleic acid extraction and yeast extract production. The best conditions for yeast autolysis was 55.2ºC, pH= 5.1 and 9.8% NaCl for 24h of processing, without the NH3 use. In these conditions, the RNA yield was 89.7%, resulting in 51.3% of dehydrated yeast extract with 57.9% protein. The use of 12.2% NH3 at 60ºC after autolysis (8h) and plasmolysis (8h) was not viable due to the reduction in the RNA yield from 89.7to78.4%. on the other hand, the thermal shock at 60ºC for 15 minutes prior to autolysis provided an increase in the yield from 89.7 to 91.4%. The autolysis, including NaCl plasmolysis in the optimized conditions was efficient, economic and with short time, thus usable for industrial purpose to obtain more valuable products such as yeast extract enriched in RNA and/or protein, for different applications.

COMPENSATORY LUNG GROWTH: LUNG PROTEIN,DNA and RNA CONTENTS IN TRILOBECTOMIZED RATS

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Compensatory lung growth: protein, DNA and RNA lung contents in undernourished trilobectomized rats

OBJETIVO: demonstrar se ocorre crescimento pulmonar compensatório (CPC) representado pelos conteúdos de proteínas, DNA e RNA no rato adulto jovem, subnutrido, submetido à trilobectomia pulmonar. MÉTODOS: Utilizamos 137 ratos Wistar, machos, distribuídos por sorteio, em 9 grupos, submetidos a três tratamentos (controle, toracotomia, trilobectomia), sacrificados em três momentos (7, 30 e 90 dias). Na trilobectomia foram extirpados os lobos médio, acessório e caudal direitos. Variáveis estudadas: conteúdos pulmonares de proteínas, DNA e RNA. RESULTADOS: No lobo cranial e pulmão esquerdo o conteúdo protéico foi maior nos trilobectomizados. Ocorreu CPC insuficiente para suprir a perda desta variável, sendo menor nos pulmões dos trilobectomizados. O incremento nos conteúdos de DNA do lobo cranial e pulmão esquerdo dos trilobectomizados foram suficientes para compensar a perda desta variável, resultando num conteúdo de DNA dos pulmões semelhante aos controle. O conteúdo de RNA, nos trilobectomizados, foi maior no lobo cranial e pulmão esquerdo, com maior eficiência no primeiro, insuficiente para que se aproximassem aos obtidos nos demais grupos, ficando menores. CONCLUSÃO: Nos trilobectomizados ocorreu CPC, provavelmente com hiperplasia celular e pouca hipertrofia, devido a grande compensação do DNA e pequena do RNA. Esta foi a grande diferença quando comparamos este resultado ao obtido com animais nutridos, que apresentavam hipertrofia pronunciada.

Effect of acute heat stress on heat shock protein 70 messenger RNA and on heat shock protein expression in the liver of broilers

1. The synthesis of heat shock protein 70 (Hsp70) mRNA and the expression of Hsp70 in the liver of broiler chickens submitted to acute heat stress (35 degrees C for 5 h) was investigated.2. Hsp70 expression was detected by SDS-PAGE and Western blot analysis using a polyclonal antiserum against Hsp70 of Blastocladiella emersonii. The specific signal of Hsp70 mRNA was analysed by Northern blot using as probe a Hsp70 cDNA of B. emersonii.3. An increase in the amount of Hsp70 was detected from the first up to the fifth hour of acute heat exposure. This increase in the amount of Hsp70 was accompanied by an increase in Hsp70 mRNA which peaked at 3 h.4. This study shows that the heat induced increase in Hsp70 mRNA and protein in broiler liver, in vivo, are time dependent, similar to that in mammals.

GENETIC INTERRELATIONSHIPS OF PROTEOLYTIC CLOSTRIDIUM-BOTULINUM TYPE-A, TYPE-B, AND TYPE-F AND OTHER MEMBERS OF THE CLOSTRIDIUM-BOTULINUM COMPLEX AS REVEALED BY SMALL-SUBUNIT RIBOSOMAL-RNA GENE-SEQUENCES

The phylogenetic interrelationships of members of the Clostridium botulinum complex of species was investigated by direct sequencing of their 16S rRNA genes. Comparative analysis of the 16S rRNA sequences demonstrated the presence of four phylogenetically distinct lineages corresponding to: i) proteolytic C. botulinum types Al B, and F, and C. sporogenes, ii) saccharolytic types B, E and F, iii) types C and D and C. novyi type A, and iv) type G and C. subterminale. The phylogenetic groupings obtained from the 16S rRNA were in complete agreement with the four divisions recognised within the 'species complex' on the basis of phenotypic criteria.