Biomedical subjects

Osvaldo Marinotti

Publications and source records attributed to Osvaldo Marinotti.

9 recordsLinked to original sources

PubMed2006

Functional characterization of the promoter of the vitellogenin gene, AsVg1, of the malaria vector, Anopheles stephensi.

Some genetic strategies for controlling transmission of mosquito-borne diseases call for the introgression of antipathogen effector genes into vector populations. Endogenous mosquito promoter and other cis-acting DNA sequences are needed to direct the expression of the effector molecules to maximize their efficacy. Vitellogenin (Vg)-encoding gene control sequences are candidates for driving tissue-, stage- and sex-specific expression of exogenous genes. One of the Anopheles stephensi Vg genes, AsVg1, was cloned and a full-length cDNA, as well as 850 base pairs adjacent to the 5'-end, were sequenced and characterized. Expression of AsVg1 is restricted to the fat body tissues of blood-fed females, and the amino acid sequence of the conceptual translation product is >85% identical to those of other anopheline Vgs. These characteristics support the conclusion that AsVg1 is a Vg-encoding gene. Functional analyses of the AsVg1 putative cis-regulatory sequences were performed using transgenic mosquitoes. The results showed that DNA fragments encompassing the 850 base pairs immediately adjacent to the 5'-end of the gene and the 3'-end untranslated region are sufficient to direct sex-, stage- and tissue-specific expression of a reporter gene. These data indicate that the AsVg1 promoter is a good candidate for controlling the expression of anti-pathogen effector molecules in this malaria vector mosquito.

Animals↗

PubMed2006

angaGEDUCI: Anopheles gambiae gene expression database with integrated comparative algorithms for identifying conserved DNA motifs in promoter sequences.

BACKGROUND: The completed sequence of the Anopheles gambiae genome has enabled genome-wide analyses of gene expression and regulation in this principal vector of human malaria. These investigations have created a demand for efficient methods of cataloguing and analyzing the large quantities of data that have been produced. The organization of genome-wide data into one unified database makes possible the efficient identification of spatial and temporal patterns of gene expression, and by pairing these findings with comparative algorithms, may offer a tool to gain insight into the molecular mechanisms that regulate these expression patterns. DESCRIPTION: We provide a publicly-accessible database and integrated data-mining tool, angaGEDUCI, that unifies 1) stage- and tissue-specific microarray analyses of gene expression in An. gambiae at different developmental stages and temporal separations following a bloodmeal, 2) functional gene annotation, 3) genomic sequence data, and 4) promoter sequence comparison algorithms. The database can be used to study genes expressed in particular stages, tissues, and patterns of interest, and to identify conserved promoter sequence motifs that may play a role in the regulation of such expression. The database is accessible from the address http://www.angaged.bio.uci.edu. CONCLUSION: By combining gene expression, function, and sequence data with integrated sequence comparison algorithms, angaGEDUCI streamlines spatial and temporal pattern-finding and produces a straightforward means of developing predictions and designing experiments to assess how gene expression may be controlled at the molecular level.

Algorithms↗

PubMed2006

Structure and expression of the lipophorin-encoding gene of the malaria vector, Anopheles gambiae.

Lipophorin is the major hemolymph protein responsible for lipid transport among tissues of insects. This protein may be a lipid source for the development and reproduction of human malaria parasites in mosquitoes, and therefore could be a target to disrupt malaria parasite development in the vector. The lipophorin of Anopheles gambiae was purified by KBr gradient ultracentrifugation and showed variation in density from 1.111 to 1.143 g/ml during development. The amount and density of lipophorin increase in blood-fed females, indicating an adaptation of vitellogenic mosquitoes to an elevated rate of lipid transport to the developing eggs. The A. gambiae lipophorin gene is composed of eight exons and transcribes an mRNA that is 10,516 nucleotides in length. The predicted initial translation product is a preproapoliphorin consisting of 3332 amino acids, which is processed by proteolysis to generate two mature apolipophorins: apolipophorin-I (Mr = 280,000) and apolipophorin-II (Mr = 81,000). The gene is expressed in the fat body tissues throughout development. An elevated transcriptional activity of the lipophorin gene during vitellogenesis is consistent with the presence of putative cis-regulatory elements (GATA and ecdysone responsive elements) in its 3'-end flanking DNA sequence.

Amino Acid Sequence↗

PubMed2005

Characterization of the c-type lysozyme gene family in Anopheles gambiae.

Seven new c-type lysozyme genes were found using the Anopheles gambiae genome sequence, increasing to eight the total number of genes in this family identified in this species. The eight lysozymes in An. gambiae have considerable variation in gene structure and expression patterns. Lys c-6 has the most unusual primary amino acid structure as the predicted protein consists of five lysozyme-like domains. Transcript abundance of each c-type lysozyme was determined by semiquantitative RT-PCR. Lys c-1, c-6 and c-7 are expressed constitutively in all developmental stages from egg to adult. Lys c-2 and c-4 also are found in all stages, but with relatively much higher levels in adults. Conversely, Lys c-3 and c-8 transcripts are highest in larvae. Lys c-1, c-6 and c-7 transcripts are found in nearly all the adult tissue samples examined while Lys c-2 and Lys c-4 are more restricted in their expression. Lys c-1 and c-2 transcripts are clearly immune responsive and are increased significantly 6-12 h post challenge with bacteria. The functional adaptive changes that may have evolved during the expansion of this gene family are briefly discussed in terms of the expression patterns, gene and protein structures.

Amino Acid Sequence↗

PubMed2005

Nanos (nos) genes of the vector mosquitoes, Anopheles gambiae, Anopheles stephensi and Aedes aegypti.

A number of genetics-based strategies for the control of vector-borne diseases require the development of genetic drive systems for introgressing antipathogen effector genes into wild populations of insects. Modified transposons whose mobilization is controlled by the DNA elements of developmentally regulated genes offer a potential solution for introducing effector genes into mosquitoes. Such elements could exhibit sex-, stage- and species-specific transposition, thus mitigating some of the concerns associated with autonomous transposition. Hybridizations in situ show that the transcription products of the nanos orthologous genes of Anopheles gambiae (Anga nos), An. stephensi (Anst nos) and Aedes aegypti (Aeae nos) accumulate in developing oocytes in adult females and localize to the posterior pole in early embryos. These features make nos genes promising candidates for donating control sequences to modified transposons.

Aedes↗

PubMed2005

Morphological and enzymatic analysis of the midgut of Anopheles darlingi during blood digestion.

The midgut of adult female Anopheles darlingi is comprised of narrow anterior and dilated posterior regions, with a single layered epithelium composed by cuboidal digestive cells. Densely packed apical microvilli and an intricate basal labyrinth characterize each cell pole. Before blood feeding, apical cytoplasm contains numerous round granules and whorled profiles of rough endoplasmic reticulum. Engorgement causes a great distension of midgut. This provokes the flattening of digestive cells and their nuclei. Simultaneously, apical granules disappear, the whorls of endoplasmic reticulum disassemble and 3h post bloodmeal (PBM), nucleoli enlarge manyfold. An intense absorptive process takes place during the first 24 h PBM, with the formation of large glycogen inclusions, which persist after the end of the digestive process. Endoproteases activities are induced after bloodmeal and attain their maximum values between 10 and 36 h PBM. At least two different aminopeptidases seem to participate in the digestive process, with their maximum activity values at 36 and 48 h PBM, respectively. Coarse electrondense aggregates, possibly debris from digested erythrocytes, begin to appear on the luminal face of the peritrophic membrane from 18 h PBM and persist during all the digestive process, and are excreted at its end. We suggest that these aggregates could contain some kind of insoluble form of haem, in order of neutralize its toxicity.

Animals↗

PubMed2005

An updated catalogue of salivary gland transcripts in the adult female mosquito, Anopheles gambiae.

Salivary glands of blood-sucking arthropods contain a variety of compounds that prevent platelet and clotting functions and modify inflammatory and immunological reactions in the vertebrate host. In mosquitoes, only the adult female takes blood meals, while both sexes take sugar meals. With the recent description of the Anopheles gambiae genome, and with a set of approximately 3000 expressed sequence tags from a salivary gland cDNA library from adult female mosquitoes, we attempted a comprehensive description of the salivary transcriptome of this most important vector of malaria transmission. In addition to many transcripts associated with housekeeping functions, we found an active transposable element, a set of Wolbachia-like proteins, several transcription factors, including Forkhead, Hairy and doublesex, extracellular matrix components and 71 genes coding for putative secreted proteins. Fourteen of these 71 proteins had matching Edman degradation sequences obtained from SDS-PAGE experiments. Overall, 33 transcripts are reported for the first time as coding for salivary proteins. The tissue and sex specificity of these protein-coding transcripts were analyzed by RT-PCR and microarray experiments for insight into their possible function. Notably, two gene products appeared to be differentially spliced in the adult female salivary glands, whereas 13 contigs matched predicted intronic regions and may include additional alternatively spliced transcripts. Most An. gambiae salivary proteins represent novel protein families of unknown function, potentially coding for pharmacologically or microbiologically active substances. Supplemental data to this work can be found at http://www.ncbi.nlm.nih.gov/projects/omes/index.html#Ag2.

Amino Acid Sequence↗

PubMed2003

The major salivary gland antigens of Culex quinquefasciatus are D7-related proteins.

The sera of persons with strong allergic responses to the bites of the mosquito, Culex quinquefasciatus, contained IgE antibodies reactive with two major salivary gland proteins with molecular weights of 35 and 28 kDa. These antigens were purified, their amino termini sequenced, and the sequences were used to search for similar sequences in public databases. Two cDNAs, CuQu-D7Clu1 and CuQu-D7Clu12, which encode D7-related proteins, were identified as containing predicted amino acid sequences identical to the 35 and 28 kDa antigens, respectively. These proteins are expressed specifically in adult female salivary glands and, their predicted tertiary structures are consistent with a role as carriers of hydrophobic molecules in mosquito saliva.

Adolescent↗

PubMed2002

Taxonomic status of Ixodes didelphidis (Acari: Ixodidae).

Ixodes didelphilis Fonseca & Aragão was described in Brazil in 1952 as a new tick species that differed from Ixodes loricatus Neumann by the spiracular plate pattern. We have reared four tick colonies from different geographic areas in the laboratory that were started from single engorged females originally identified as I. didelphidis (BMG, colony ) and I. Iocicatus (CSP, PSP, and TRJ colonies). We analyzed the spiracular plate morphology of F1 adult ticks from each tick colony, compared their biological 11th, and performed a molecular analysis of the second internal transcribed rDNA spacer (ITS2) to test the validity of the species I. didelphidis. The spiracular plate analysis of laboratory F1 adult ticks yielded single females from the four colonies showed variations that invalidate morphological parameters for differentiation of loricatus and I. didelphidis. Biological data of the BMC, CSP, and TRJ colonies were similar. The biology ofthe PSP colony was not evaluated. The ITS2 sequence variations observed between the tick colonies ranged from 1.3 to 4.9%, and the similarity tree constructed by the neighbor-joining method with nucleotide distances showerl that the distances between the samples were similar to what is expected for intraspecific variations found in other ticks species. The morphological and biological results, in conjunction with the ITS2 analysis, supported the conspecificity I. loricatus and I. didelphidis.

Animals↗