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Transcriptome Analysis of the Emerald Ash Borer (EAB), Agrilus planipennis: De Novo Assembly, Functional Annotation and Comparative Analysis.

Duan J, Ladd T, Doucet D, Cusson M, vanFrankenhuyzen K, Mittapalli O, Krell PJ, Quan G - PLoS ONE (2015)

Bottom Line: The EAB transcriptome assembly was compared with 13 other sequenced insect species, resulting in the prediction of 536 unigenes that are Coleoptera-specific.This study provides one of the most fundamental and comprehensive transcriptome resources available for EAB to date.Identification of the tissue- stage- or species- specific unigenes will benefit the further study of gene functions during growth and metamorphosis processes in EAB and other pest insects.

View Article: PubMed Central - PubMed

Affiliation: Great Lakes Forestry Centre, Canadian Forest Service, Natural Resources Canada, Sault Ste. Marie, Ontario, Canada; Department of Molecular and Cellular Biology, University of Guelph, Guelph, Ontario, Canada.

ABSTRACT

Background: The Emerald ash borer (EAB), Agrilus planipennis, is an invasive phloem-feeding insect pest of ash trees. Since its initial discovery near the Detroit, US- Windsor, Canada area in 2002, the spread of EAB has had strong negative economic, social and environmental impacts in both countries. Several transcriptomes from specific tissues including midgut, fat body and antenna have recently been generated. However, the relatively low sequence depth, gene coverage and completeness limited the usefulness of these EAB databases.

Methodology and principal findings: High-throughput deep RNA-Sequencing (RNA-Seq) was used to obtain 473.9 million pairs of 100 bp length paired-end reads from various life stages and tissues. These reads were assembled into 88,907 contigs using the Trinity strategy and integrated into 38,160 unigenes after redundant sequences were removed. We annotated 11,229 unigenes by searching against the public nr, Swiss-Prot and COG. The EAB transcriptome assembly was compared with 13 other sequenced insect species, resulting in the prediction of 536 unigenes that are Coleoptera-specific. Differential gene expression revealed that 290 unigenes are expressed during larval molting and 3,911 unigenes during metamorphosis from larvae to pupae, respectively (FDR< 0.01 and log2 FC>2). In addition, 1,167 differentially expressed unigenes were identified from larval and adult midguts, 435 unigenes were up-regulated in larval midgut and 732 unigenes were up-regulated in adult midgut. Most of the genes involved in RNA interference (RNAi) pathways were identified, which implies the existence of a system RNAi in EAB.

Conclusions and significance: This study provides one of the most fundamental and comprehensive transcriptome resources available for EAB to date. Identification of the tissue- stage- or species- specific unigenes will benefit the further study of gene functions during growth and metamorphosis processes in EAB and other pest insects.

No MeSH data available.


Related in: MedlinePlus

Phylogenetic relationships of orthologs between EAB and other insect species.(A) Phylogenetic relations of EAB with other insect species. A maximum likelihood evolutionary tree was inferred based on concatenated single copy orthologs among EAB and 13 insect species. The vertebrates H. sapiens and G. gallus were used as outgroups. Bar shows a genetic distance of 0.1. (B) The comparison of orthologous gene among EAB and other species. 1:1:1 orthologs include the common orthologs with only one copy in all surveyed species, N:N:N orthologs include the common orthologs with different copy numbers in the different species, patchy orthologs include the orthologs existing in at least one species of vertebrates and insects, Coleoptera-specific orthologs include the orthologs existing only in EAB and T. castaneum, insect-specific orthologs includes the orthologs existing only in insects, species specific orthologs represent the genes from only one species, other orthologs include the unclassified orthologs, and unclustered orthologs include these that cannot be clustered into known gene families.
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pone.0134824.g005: Phylogenetic relationships of orthologs between EAB and other insect species.(A) Phylogenetic relations of EAB with other insect species. A maximum likelihood evolutionary tree was inferred based on concatenated single copy orthologs among EAB and 13 insect species. The vertebrates H. sapiens and G. gallus were used as outgroups. Bar shows a genetic distance of 0.1. (B) The comparison of orthologous gene among EAB and other species. 1:1:1 orthologs include the common orthologs with only one copy in all surveyed species, N:N:N orthologs include the common orthologs with different copy numbers in the different species, patchy orthologs include the orthologs existing in at least one species of vertebrates and insects, Coleoptera-specific orthologs include the orthologs existing only in EAB and T. castaneum, insect-specific orthologs includes the orthologs existing only in insects, species specific orthologs represent the genes from only one species, other orthologs include the unclassified orthologs, and unclustered orthologs include these that cannot be clustered into known gene families.

Mentions: To identify the species-specific genes of EAB, the 15,079 transcripts containing an ORF longer than 300 bp were compared against 13 other sequenced insect species (Materials and Methods). Two vertebrate genomes, G. gallus and H. sapiens, were used as outgroups. Orthologous relationships among these species were classified based on their sequence similarity (Fig 5). A total of 2,541 EAB unigenes belonging to the ancient group have orthologous relations among all the surveyed insects and vertebrates, of which 507 unigenes contain only one copy in all examined species. These unigenes may belong to essential gene categories and may be subject to strict evolutionary constraints. A maximum likelihood evolutionary tree was inferred based these single copy orthologs. The results showed that EAB has the closest relationship with T. castaneum (Fig 5). In the comparison against G. gallus and H. sapiens, 2,896 EAB unigenes were predicted as insect-specific genes because they had orthologs only in other examined insects but not in G. gallus and H. sapiens. We found 536 EAB unigenes that only had orthologs in T. castaneum but not found in other insect species, suggesting they might be Coleoptera-specfic orthologs (Fig 5). Most of the identified Coleoptera-specfic genes were new and predicted as hypothetical proteins. Only a few of the hypothetical proteins had a predicted function, such as the ecdysis triggering hormone preprotein gene (NP_001165744), odorant binding protein C20, serine protease P72 (EFA09224) and EMSY (XP_001811996), but little is known about their functions in beetles.


Transcriptome Analysis of the Emerald Ash Borer (EAB), Agrilus planipennis: De Novo Assembly, Functional Annotation and Comparative Analysis.

Duan J, Ladd T, Doucet D, Cusson M, vanFrankenhuyzen K, Mittapalli O, Krell PJ, Quan G - PLoS ONE (2015)

Phylogenetic relationships of orthologs between EAB and other insect species.(A) Phylogenetic relations of EAB with other insect species. A maximum likelihood evolutionary tree was inferred based on concatenated single copy orthologs among EAB and 13 insect species. The vertebrates H. sapiens and G. gallus were used as outgroups. Bar shows a genetic distance of 0.1. (B) The comparison of orthologous gene among EAB and other species. 1:1:1 orthologs include the common orthologs with only one copy in all surveyed species, N:N:N orthologs include the common orthologs with different copy numbers in the different species, patchy orthologs include the orthologs existing in at least one species of vertebrates and insects, Coleoptera-specific orthologs include the orthologs existing only in EAB and T. castaneum, insect-specific orthologs includes the orthologs existing only in insects, species specific orthologs represent the genes from only one species, other orthologs include the unclassified orthologs, and unclustered orthologs include these that cannot be clustered into known gene families.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4526369&req=5

pone.0134824.g005: Phylogenetic relationships of orthologs between EAB and other insect species.(A) Phylogenetic relations of EAB with other insect species. A maximum likelihood evolutionary tree was inferred based on concatenated single copy orthologs among EAB and 13 insect species. The vertebrates H. sapiens and G. gallus were used as outgroups. Bar shows a genetic distance of 0.1. (B) The comparison of orthologous gene among EAB and other species. 1:1:1 orthologs include the common orthologs with only one copy in all surveyed species, N:N:N orthologs include the common orthologs with different copy numbers in the different species, patchy orthologs include the orthologs existing in at least one species of vertebrates and insects, Coleoptera-specific orthologs include the orthologs existing only in EAB and T. castaneum, insect-specific orthologs includes the orthologs existing only in insects, species specific orthologs represent the genes from only one species, other orthologs include the unclassified orthologs, and unclustered orthologs include these that cannot be clustered into known gene families.
Mentions: To identify the species-specific genes of EAB, the 15,079 transcripts containing an ORF longer than 300 bp were compared against 13 other sequenced insect species (Materials and Methods). Two vertebrate genomes, G. gallus and H. sapiens, were used as outgroups. Orthologous relationships among these species were classified based on their sequence similarity (Fig 5). A total of 2,541 EAB unigenes belonging to the ancient group have orthologous relations among all the surveyed insects and vertebrates, of which 507 unigenes contain only one copy in all examined species. These unigenes may belong to essential gene categories and may be subject to strict evolutionary constraints. A maximum likelihood evolutionary tree was inferred based these single copy orthologs. The results showed that EAB has the closest relationship with T. castaneum (Fig 5). In the comparison against G. gallus and H. sapiens, 2,896 EAB unigenes were predicted as insect-specific genes because they had orthologs only in other examined insects but not in G. gallus and H. sapiens. We found 536 EAB unigenes that only had orthologs in T. castaneum but not found in other insect species, suggesting they might be Coleoptera-specfic orthologs (Fig 5). Most of the identified Coleoptera-specfic genes were new and predicted as hypothetical proteins. Only a few of the hypothetical proteins had a predicted function, such as the ecdysis triggering hormone preprotein gene (NP_001165744), odorant binding protein C20, serine protease P72 (EFA09224) and EMSY (XP_001811996), but little is known about their functions in beetles.

Bottom Line: The EAB transcriptome assembly was compared with 13 other sequenced insect species, resulting in the prediction of 536 unigenes that are Coleoptera-specific.This study provides one of the most fundamental and comprehensive transcriptome resources available for EAB to date.Identification of the tissue- stage- or species- specific unigenes will benefit the further study of gene functions during growth and metamorphosis processes in EAB and other pest insects.

View Article: PubMed Central - PubMed

Affiliation: Great Lakes Forestry Centre, Canadian Forest Service, Natural Resources Canada, Sault Ste. Marie, Ontario, Canada; Department of Molecular and Cellular Biology, University of Guelph, Guelph, Ontario, Canada.

ABSTRACT

Background: The Emerald ash borer (EAB), Agrilus planipennis, is an invasive phloem-feeding insect pest of ash trees. Since its initial discovery near the Detroit, US- Windsor, Canada area in 2002, the spread of EAB has had strong negative economic, social and environmental impacts in both countries. Several transcriptomes from specific tissues including midgut, fat body and antenna have recently been generated. However, the relatively low sequence depth, gene coverage and completeness limited the usefulness of these EAB databases.

Methodology and principal findings: High-throughput deep RNA-Sequencing (RNA-Seq) was used to obtain 473.9 million pairs of 100 bp length paired-end reads from various life stages and tissues. These reads were assembled into 88,907 contigs using the Trinity strategy and integrated into 38,160 unigenes after redundant sequences were removed. We annotated 11,229 unigenes by searching against the public nr, Swiss-Prot and COG. The EAB transcriptome assembly was compared with 13 other sequenced insect species, resulting in the prediction of 536 unigenes that are Coleoptera-specific. Differential gene expression revealed that 290 unigenes are expressed during larval molting and 3,911 unigenes during metamorphosis from larvae to pupae, respectively (FDR< 0.01 and log2 FC>2). In addition, 1,167 differentially expressed unigenes were identified from larval and adult midguts, 435 unigenes were up-regulated in larval midgut and 732 unigenes were up-regulated in adult midgut. Most of the genes involved in RNA interference (RNAi) pathways were identified, which implies the existence of a system RNAi in EAB.

Conclusions and significance: This study provides one of the most fundamental and comprehensive transcriptome resources available for EAB to date. Identification of the tissue- stage- or species- specific unigenes will benefit the further study of gene functions during growth and metamorphosis processes in EAB and other pest insects.

No MeSH data available.


Related in: MedlinePlus