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Construction and characterization of an expressed sequenced tag library for the mosquito vector Armigeres subalbatus.

Mayhew GF, Bartholomay LC, Kou HY, Rocheleau TA, Fuchs JF, Aliota MT, Tsao IY, Huang CY, Liu TT, Hsiao KJ, Tsai SF, Yang UC, Perna NT, Cho WL, Christensen BM, Chen CC - BMC Genomics (2007)

Bottom Line: All of these sequences were collapsed into clusters resulting in 8,020 unique sequence clusters or singletons.EST clusters were annotated and curated manually within ASAP (A Systematic Annotation Package for Community Analysis of Genomes) web portal according to BLAST results from comparisons to Genbank, and the Anopheles gambiae and Drosophila melanogaster genome projects.The resulting dataset is the first of its kind for this mosquito vector and provides a basis for future studies of mosquito vectors regarding the cascade of events that occurs in response to infection, and thereby providing insight into vector competence and innate immunity.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Pathobiological Sciences, University of Wisconsin-Madison, 1656 Linden Dr,, Madison Wisconsin, 53706, USA. mayhew@svm.vetmed.wisc.edu

ABSTRACT

Background: The mosquito, Armigeres subalbatus, mounts a distinctively robust innate immune response when infected with the nematode Brugia malayi, a causative agent of lymphatic filariasis. In order to mine the transcriptome for new insight into the cascade of events that takes place in response to infection in this mosquito, 6 cDNA libraries were generated from tissues of adult female mosquitoes subjected to immune-response activation treatments that lead to well-characterized responses, and from aging, naïve mosquitoes. Expressed sequence tags (ESTs) from each library were produced, annotated, and subjected to comparative analyses.

Results: Six libraries were constructed and used to generate 44,940 expressed sequence tags, of which 38,079 passed quality filters to be included in the annotation project and subsequent analyses. All of these sequences were collapsed into clusters resulting in 8,020 unique sequence clusters or singletons. EST clusters were annotated and curated manually within ASAP (A Systematic Annotation Package for Community Analysis of Genomes) web portal according to BLAST results from comparisons to Genbank, and the Anopheles gambiae and Drosophila melanogaster genome projects.

Conclusion: The resulting dataset is the first of its kind for this mosquito vector and provides a basis for future studies of mosquito vectors regarding the cascade of events that occurs in response to infection, and thereby providing insight into vector competence and innate immunity.

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Related in: MedlinePlus

Homologous sequences for Ar. subalbatus found in fly databases. A) Comparative analysis of Ar. subalbatus EST clusters with predicted peptides from 3 other mosquito species with completed genomes: Ae. aegypti, An. gambiae, and C.p. quinquefasciatus. Overlapping regions indicate homologous sequences from blastx searches against the peptide databases. A homolog is defined as having an e-value cutoff of 1e-20, a percent match of 40% (true matches, not conserved), and a minimum match length of 30 for the high-scoring segment pair (HSP). This comparison includes 8,020 possible cluster sequences from Ar. subalbatus (brackets), of which 3,013 had no homolog. Boxes directly adjacent to circles indicate 1) the species being compared to Ar. subalbatus, and 2) the total # of homologous sequences between that species and Ar. subalbatus. B) A gene list of the total of overlapping and non-overlapping Ar. subalbatus homologs to Ae. aegypti, An. gambiae, and C. p. quinquefasciatus was compared to a gene list of homologs found to D. melanogaster. A significant number of genes (2,074) from Ar. subalbatus have no homolog in the fruit fly, but qualify as homologs to genes in other mosquito species.
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Figure 3: Homologous sequences for Ar. subalbatus found in fly databases. A) Comparative analysis of Ar. subalbatus EST clusters with predicted peptides from 3 other mosquito species with completed genomes: Ae. aegypti, An. gambiae, and C.p. quinquefasciatus. Overlapping regions indicate homologous sequences from blastx searches against the peptide databases. A homolog is defined as having an e-value cutoff of 1e-20, a percent match of 40% (true matches, not conserved), and a minimum match length of 30 for the high-scoring segment pair (HSP). This comparison includes 8,020 possible cluster sequences from Ar. subalbatus (brackets), of which 3,013 had no homolog. Boxes directly adjacent to circles indicate 1) the species being compared to Ar. subalbatus, and 2) the total # of homologous sequences between that species and Ar. subalbatus. B) A gene list of the total of overlapping and non-overlapping Ar. subalbatus homologs to Ae. aegypti, An. gambiae, and C. p. quinquefasciatus was compared to a gene list of homologs found to D. melanogaster. A significant number of genes (2,074) from Ar. subalbatus have no homolog in the fruit fly, but qualify as homologs to genes in other mosquito species.

Mentions: Those clusters that were homologous were subjected to Venn analysis (Figure 3A) to discover overlapping predicted peptides in 3 other mosquito species: Ae. aegypti, (Ae Vectorbase AaegL1.1), An. gambiae (Anoph Vectorbase AgamP3), and C.p. quinquefasciatus (Cpip Vectorbase CpipJ1.0_5), and the fruit fly, D. melanogaster. The mosquito with the largest number of gene products that are uniquely homologous to Ar. subalbatus is Ae. aegypti, as would be predicted by the degree of relatedness of these two mosquitoes. In comparing Ar. subalbatus to all available mosquito and Drosophila homologous predicted peptides, 2908 sequences are represented in all fly species. A significant number (2,074) of clusters from Ar. subalbatus qualify as homologs to genes in other mosquito species, but have no homolog in the fruit fly (Figure 3B).


Construction and characterization of an expressed sequenced tag library for the mosquito vector Armigeres subalbatus.

Mayhew GF, Bartholomay LC, Kou HY, Rocheleau TA, Fuchs JF, Aliota MT, Tsao IY, Huang CY, Liu TT, Hsiao KJ, Tsai SF, Yang UC, Perna NT, Cho WL, Christensen BM, Chen CC - BMC Genomics (2007)

Homologous sequences for Ar. subalbatus found in fly databases. A) Comparative analysis of Ar. subalbatus EST clusters with predicted peptides from 3 other mosquito species with completed genomes: Ae. aegypti, An. gambiae, and C.p. quinquefasciatus. Overlapping regions indicate homologous sequences from blastx searches against the peptide databases. A homolog is defined as having an e-value cutoff of 1e-20, a percent match of 40% (true matches, not conserved), and a minimum match length of 30 for the high-scoring segment pair (HSP). This comparison includes 8,020 possible cluster sequences from Ar. subalbatus (brackets), of which 3,013 had no homolog. Boxes directly adjacent to circles indicate 1) the species being compared to Ar. subalbatus, and 2) the total # of homologous sequences between that species and Ar. subalbatus. B) A gene list of the total of overlapping and non-overlapping Ar. subalbatus homologs to Ae. aegypti, An. gambiae, and C. p. quinquefasciatus was compared to a gene list of homologs found to D. melanogaster. A significant number of genes (2,074) from Ar. subalbatus have no homolog in the fruit fly, but qualify as homologs to genes in other mosquito species.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2262096&req=5

Figure 3: Homologous sequences for Ar. subalbatus found in fly databases. A) Comparative analysis of Ar. subalbatus EST clusters with predicted peptides from 3 other mosquito species with completed genomes: Ae. aegypti, An. gambiae, and C.p. quinquefasciatus. Overlapping regions indicate homologous sequences from blastx searches against the peptide databases. A homolog is defined as having an e-value cutoff of 1e-20, a percent match of 40% (true matches, not conserved), and a minimum match length of 30 for the high-scoring segment pair (HSP). This comparison includes 8,020 possible cluster sequences from Ar. subalbatus (brackets), of which 3,013 had no homolog. Boxes directly adjacent to circles indicate 1) the species being compared to Ar. subalbatus, and 2) the total # of homologous sequences between that species and Ar. subalbatus. B) A gene list of the total of overlapping and non-overlapping Ar. subalbatus homologs to Ae. aegypti, An. gambiae, and C. p. quinquefasciatus was compared to a gene list of homologs found to D. melanogaster. A significant number of genes (2,074) from Ar. subalbatus have no homolog in the fruit fly, but qualify as homologs to genes in other mosquito species.
Mentions: Those clusters that were homologous were subjected to Venn analysis (Figure 3A) to discover overlapping predicted peptides in 3 other mosquito species: Ae. aegypti, (Ae Vectorbase AaegL1.1), An. gambiae (Anoph Vectorbase AgamP3), and C.p. quinquefasciatus (Cpip Vectorbase CpipJ1.0_5), and the fruit fly, D. melanogaster. The mosquito with the largest number of gene products that are uniquely homologous to Ar. subalbatus is Ae. aegypti, as would be predicted by the degree of relatedness of these two mosquitoes. In comparing Ar. subalbatus to all available mosquito and Drosophila homologous predicted peptides, 2908 sequences are represented in all fly species. A significant number (2,074) of clusters from Ar. subalbatus qualify as homologs to genes in other mosquito species, but have no homolog in the fruit fly (Figure 3B).

Bottom Line: All of these sequences were collapsed into clusters resulting in 8,020 unique sequence clusters or singletons.EST clusters were annotated and curated manually within ASAP (A Systematic Annotation Package for Community Analysis of Genomes) web portal according to BLAST results from comparisons to Genbank, and the Anopheles gambiae and Drosophila melanogaster genome projects.The resulting dataset is the first of its kind for this mosquito vector and provides a basis for future studies of mosquito vectors regarding the cascade of events that occurs in response to infection, and thereby providing insight into vector competence and innate immunity.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Pathobiological Sciences, University of Wisconsin-Madison, 1656 Linden Dr,, Madison Wisconsin, 53706, USA. mayhew@svm.vetmed.wisc.edu

ABSTRACT

Background: The mosquito, Armigeres subalbatus, mounts a distinctively robust innate immune response when infected with the nematode Brugia malayi, a causative agent of lymphatic filariasis. In order to mine the transcriptome for new insight into the cascade of events that takes place in response to infection in this mosquito, 6 cDNA libraries were generated from tissues of adult female mosquitoes subjected to immune-response activation treatments that lead to well-characterized responses, and from aging, naïve mosquitoes. Expressed sequence tags (ESTs) from each library were produced, annotated, and subjected to comparative analyses.

Results: Six libraries were constructed and used to generate 44,940 expressed sequence tags, of which 38,079 passed quality filters to be included in the annotation project and subsequent analyses. All of these sequences were collapsed into clusters resulting in 8,020 unique sequence clusters or singletons. EST clusters were annotated and curated manually within ASAP (A Systematic Annotation Package for Community Analysis of Genomes) web portal according to BLAST results from comparisons to Genbank, and the Anopheles gambiae and Drosophila melanogaster genome projects.

Conclusion: The resulting dataset is the first of its kind for this mosquito vector and provides a basis for future studies of mosquito vectors regarding the cascade of events that occurs in response to infection, and thereby providing insight into vector competence and innate immunity.

Show MeSH
Related in: MedlinePlus