scholarly journals Transcriptome landscape of the developing olive fruit fly embryo delineated by Oxford Nanopore long-read RNA-Seq

2018 ◽  
Author(s):  
Anthony Bayega ◽  
Spyros Oikonomopoulos ◽  
Eleftherios Zorbas ◽  
Yu Chang Wang ◽  
Maria-Eleni Gregoriou ◽  
...  
Keyword(s):  
Embryo Development ◽  
De Novo ◽  
Transcriptome Assembly ◽  
Fruit Fly ◽  
Full Length ◽  
Olive Fruit Fly ◽  
Rna Seq ◽  
Olive Fruit ◽  
Olive Fly ◽  
Oxford Nanopore

AbstractThe olive fruit fly or olive fly (Bactrocera oleae) is the most important pest of cultivated olive trees. Like all insects the olive fly undergoes complete metamorphosis. However, the transcription dynamics that occur during early embryonic development have not been explored, while detailed transcriptomic analysis in the absence of a fully annotated genome is challenging. We collected olive fly embryos at hourly intervals for the first 6 hours of development and performed full-length cDNA-Seq using a purpose designed SMARTer cDNA synthesis protocol followed by sequencing on the MinION (Oxford Nanopore Technologies). We generated 31 million total reads across the timepoints (median yield 4.2 million per timepoint). The reads showed 98 % alignment rate to the olive fly genome and 91 % alignment rate to the NBCI predicted B. oleae gene models. Over 50 % of the expressed genes had at least one read covering its entire length validating our full-length RNA-Seq procedure. Expression of 68 % of the predicted B. oleae genes was detected in the first six hours of development. We generated a de novo transcriptome assembly of the olive fly and identified 3553 novel genes and a total of 79,810 transcripts; a fourfold increase in transcriptome diversity compared to the NCBI predicted transcriptome. On a global scale, the first six hours of embryo development were characterized by dramatic transcriptome changes with the total number of transcripts per embryo dropping to half from the first hour to the second hour of embryo development. Clustering of genes based on temporal co-expression followed by gene-set enrichment analysiss of genes expressed in the first six hours of embryo development showed that genes involved in transcription and translation, macro-molecule biosynthesis, and neurodevelopment were highly enriched. These data provide the first insight into the transcriptome landscape of the developing olive fly embryo. The data also reveal transcript signatures of sex development. Overall, full-length sequencing of the cDNA molecules permitted a detailed characterization of the isoform complexity and the transcriptional dynamics of the first embryonic stages of the B. oleae.

scholarly journals De novo genome assembly of the olive fruit fly (Bactrocera oleae) developed through a combination of linked-reads and long-read technologies

2018 ◽  
Author(s):  
Haig Djambazian ◽  
Anthony Bayega ◽  
Konstantina T. Tsoumani ◽  
Efthimia Sagri ◽  
Maria-Eleni Gregoriou ◽  
...  
Keyword(s):  
Y Chromosome ◽  
De Novo ◽  
Fruit Fly ◽  
Bactrocera Oleae ◽  
Olive Fruit Fly ◽  
Olive Fruit ◽  
Long Reads ◽  
Oxford Nanopore ◽  
Long Read ◽  
Oxford Nanopore Technologies

AbstractLong-read sequencing has greatly contributed to the generation of high quality assemblies, albeit at a high cost. It is also not always clear how to combine sequencing platforms. We sequenced the genome of the olive fruit fly (Bactrocera oleae), the most important pest in the olive fruits agribusiness industry, using Illumina short-reads, mate-pairs, 10x Genomics linked-reads, Pacific Biosciences (PacBio), and Oxford Nanopore Technologies (ONT). The 10x linked-reads assembly gave the most contiguous assembly with an N50 of 2.16 Mb. Scaffolding the linked-reads assembly using long-reads from ONT gave a more contiguous assembly with scaffold N50 of 4.59 Mb. We also present the most extensive transcriptome datasets of the olive fly derived from different tissues and stages of development. Finally, we used the Chromosome Quotient method to identify Y-chromosome scaffolds and show that the long-reads based assembly generates very highly contiguous Y-chromosome assembly.JR is a member of the MinION Access Program (MAP) and has received free-of-charge flow cells and sequencing kits from Oxford Nanopore Technologies for other projects. JR has had no other financial support from ONT.AB has received re-imbursement for travel costs associated with attending Nanopore Community meeting 2018, a meeting organized my Oxford Nanopore Technologies.

scholarly journals Nanopore long-read RNA-seq and absolute quantification delineate transcription dynamics in early embryo development of an insect pest

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Anthony Bayega ◽  
Spyros Oikonomopoulos ◽  
Maria-Eleni Gregoriou ◽  
Konstantina T. Tsoumani ◽  
Antonis Giakountis ◽  
...  
Keyword(s):  
Embryo Development ◽  
Fruit Fly ◽  
Early Embryo ◽  
Olive Fruit Fly ◽  
Rna Seq ◽  
Early Embryo Development ◽  
Olive Fruit ◽  
Transcript Quantification ◽  
Long Read ◽  
Insight Into

AbstractThe olive fruit fly, Bactrocera oleae, is the most important pest for the olive fruit but lacks adequate transcriptomic characterization that could aid in molecular control approaches. We apply nanopore long-read RNA-seq with internal RNA standards allowing absolute transcript quantification to analyze transcription dynamics during early embryo development for the first time in this organism. Sequencing on the MinION platform generated over 31 million reads. Over 50% of the expressed genes had at least one read covering its entire length validating our full-length approach. We generated a de novo transcriptome assembly and identified 1768 new genes and a total of 79,810 isoforms; a fourfold increase in transcriptome diversity compared to the current NCBI predicted transcriptome. Absolute transcript quantification per embryo allowed an insight into the dramatic re-organization of maternal transcripts. We further identified Zelda as a possible regulator of early zygotic genome activation in B. oleae and provide further insights into the maternal-to-zygotic transition. These data show the utility of long-read RNA in improving characterization of non-model organisms that lack a fully annotated genome, provide potential targets for sterile insect technic approaches, and provide the first insight into the transcriptome landscape of the developing olive fruit fly embryo.

scholarly journals Functional Genomics of a Symbiotic Community: Shared Traits in the Olive Fruit Fly Gut Microbiota

2019 ◽  
Vol 12 (2) ◽  
pp. 3778-3791 ◽  
Author(s):  
Frances Blow ◽  
Anastasia Gioti ◽  
Ian B Goodhead ◽  
Maria Kalyva ◽  
Anastasia Kampouraki ◽  
...  
Keyword(s):  
Gut Microbiota ◽  
Transcriptome Assembly ◽  
Indirect Evidence ◽  
Fruit Fly ◽  
Underlying Mechanism ◽  
Olive Fruit Fly ◽  
Olive Fruit ◽  
Dietary Nitrogen ◽  
Obligate Symbiont ◽  
Insect Gut

Abstract The olive fruit fly Bactrocera oleae is a major pest of olives worldwide and houses a specialized gut microbiota dominated by the obligate symbiont “Candidatus Erwinia dacicola.” Candidatus Erwinia dacicola is thought to supplement dietary nitrogen to the host, with only indirect evidence for this hypothesis so far. Here, we sought to investigate the contribution of the symbiosis to insect fitness and explore the ecology of the insect gut. For this purpose, we examined the composition of bacterial communities associated with Cretan olive fruit fly populations, and inspected several genomes and one transcriptome assembly. We identified, and reconstructed the genome of, a novel component of the gut microbiota, Tatumella sp. TA1, which is stably associated with Mediterranean olive fruit fly populations. We also reconstructed a number of pathways related to nitrogen assimilation and interactions with the host. The results show that, despite variation in taxa composition of the gut microbial community, core functions related to the symbiosis are maintained. Functional redundancy between different microbial taxa was observed for genes involved in urea hydrolysis. The latter is encoded in the obligate symbiont genome by a conserved urease operon, likely acquired by horizontal gene transfer, based on phylogenetic evidence. A potential underlying mechanism is the action of mobile elements, especially abundant in the Ca. E. dacicola genome. This finding, along with the identification, in the studied genomes, of extracellular surface structure components that may mediate interactions within the gut community, suggest that ongoing and past genetic exchanges between microbes may have shaped the symbiosis.

scholarly journals De novo assembly of the olive fruit fly (Bactrocera oleae) genome with linked-reads and long-read technologies minimizes gaps and provides exceptional Y chromosome assembly

BMC Genomics ◽  
2020 ◽  
Vol 21 (1) ◽  
Author(s):  
Anthony Bayega ◽  
Haig Djambazian ◽  
Konstantina T. Tsoumani ◽  
Maria-Eleni Gregoriou ◽  
Efthimia Sagri ◽  
...  
Keyword(s):  
Y Chromosome ◽  
De Novo Assembly ◽  
De Novo ◽  
Fruit Fly ◽  
Bactrocera Oleae ◽  
Olive Fruit Fly ◽  
Olive Fruit ◽  
Long Read

scholarly journals Yeasts Associated with the Olive Fruit Fly Bactrocera oleae (Rossi) (Diptera: Tephritidae) Lead to New Attractants

Agronomy ◽  
2020 ◽  
Vol 10 (10) ◽  
pp. 1501
Author(s):  
Elda Vitanović ◽  
Julian M. Lopez ◽  
Jeffrey R. Aldrich ◽  
Maja Jukić Špika ◽  
Kyria Boundy-Mills ◽  
...  
Keyword(s):  
Candida Utilis ◽  
Insect Pest ◽  
Fruit Fly ◽  
Bactrocera Oleae ◽  
Olive Fruit Fly ◽  
Olive Fruit ◽  
Torula Yeast ◽  
Yeast Cultures ◽  
Industry Standard ◽  
Olive Fly

The olive fruit fly (Bactrocera oleae Rossi) is the primary insect pest in all olive-growing regions worldwide. New integrated pest management (IPM) techniques are needed for B. oleae to mitigate reliance on pesticides used for its control which can result in negative environmental impacts. More effective lures for monitoring olive flies would help to know when and where direct chemical applications are required. The aim of this research was to find new, more effective methods for B. oleae detection and monitoring. Twelve insect-associated yeasts were selected and tested as living cultures in McPhail traps for the attraction of olive flies. Certain yeasts were more attractive than others to B. oleae; specifically, Kuraishia capsulata, Lachancea thermotolerans, Peterozyma xylosa, Scheffersomyces ergatensis, and Nakazawae ernobii, than the industry-standard dried torula yeast (Cyberlindnera jadinii; syn. Candida utilis). The attractiveness of dry, inactive (i.e., non-living) formulations of these five yeasts was also tested in the field. Inactive formulations of K. capsulata, P. xylosa, N. ernobii, and L. thermotolerans were significantly more attractive to B. oleae than commercially available torula yeast. Green lacewing, Chrysoperla comanche (Stephens) (Neuroptera: Chrysopidae), adults were incidentally caught in traps baited with the live yeast cultures. This is the first field study that compares olive fly attraction to yeast species other than torula yeast. Commercialization of yeasts that are more attractive than the torula standard would improve monitoring and associated control of the olive fruit fly.

scholarly journals Extending rnaSPAdes functionality for hybrid transcriptome assembly

2020 ◽  
Author(s):  
Andrey D. Prjibelski ◽  
Giuseppe D. Puglia ◽  
Dmitry Antipov ◽  
Elena Bushmanova ◽  
Daniela Giordano ◽  
...  
Keyword(s):  
De Novo ◽  
Transcriptome Assembly ◽  
Rna Seq ◽  
Structure Information ◽  
Oxford Nanopore ◽  
Complete Sequences ◽  
Novel Method ◽  
Hybrid Assemblies ◽  
Oxford Nanopore Technologies ◽  
Alternative Isoforms

AbstractBackgroundDe novo RNA-Seq assembly is a powerful method for analysing transcriptomes when the reference genome is not available or poorly annotated. However, due to the short length of Illumina reads it is usually impossible to reconstruct complete sequences of complex genes and alternative isoforms. Recently emerged possibility to generate long RNA reads, such as PacBio and Oxford Nanopores, may dramatically improve the assembly quality, and thus the consecutive analysis. While reference-based tools for analysing long RNA reads were recently developed, there is no established pipeline for de novo assembly of such data.ResultsIn this work we present a novel method that allows to perform high-quality de novo transcriptome assemblies by combining accuracy and reliability of short reads with exon structure information carried out from long error-prone reads. The algorithm is designed by incorporating existing hybridSPAdes approach into rnaSPAdes pipeline and adapting it for transcriptomic data.ConclusionTo evaluate the benefit of using long RNA reads we selected several datasets containing both Illumina and Iso-seq or Oxford Nanopore Technologies (ONT) reads. Using an existing quality assessment software, we show that hybrid assemblies performed with rnaSPAdes contain more full-length genes and alternative isoforms comparing to the case when only short-read data is used.Availability and implementationrnaSPAdes is implemented in C++ and Python and is freely available for Linux and MacOS under GPLv2 license at cab.spbu.ru/software/rnaspades/ and github.com/ablab/spades.

scholarly journals Baltica: integrated splice junction usage analysis

2021 ◽  
Author(s):  
Thiago Britto-Borges ◽  
Volker Boehm ◽  
Niels H Gehring ◽  
Christoph Dieterich
Keyword(s):  
Data Integration ◽  
De Novo ◽  
Transcriptome Assembly ◽  
Splice Junction ◽  
Rna Seq ◽  
Oxford Nanopore ◽  
Single Method ◽  
Usage Analysis ◽  
First Time ◽  
Transcriptional Process

Alternative splicing is a tightly regulated co- and post-transcriptional process contributing to the transcriptome diversity observed in eukaryotes. Several methods for detecting differential junction usage (DJU) from RNA sequencing (RNA-seq) datasets exist. Yet, efforts to integrate the results from DJU methods are lacking. Here, we present Baltica, a framework that provides workflows for quality control, de novo transcriptome assembly with StringTie2, and currently 4 DJU methods: rMATS, JunctionSeq, Majiq, and LeafCutter. Baltica puts the results from different DJU methods into context by integrating the results at the junction level. We present Baltica using 2 datasets, one containing known artificial transcripts (SIRVs) and the second dataset of paired Illumina and Oxford Nanopore Technologies RNA-seq. The data integration allows the user to compare the performance of the tools and reveals that JunctionSeq outperforms the other methods, in terms of F1 score, for both datasets. Finally, we demonstrate for the first time that meta-classifiers trained on scores of multiple methods outperform classifiers trained on scores of a single method, emphasizing the application of our data integration approach for differential splicing identification. Baltica is available at https://github.com/dieterich-lab/Baltica under MIT license.

scholarly journals Comparative Field Studies of Various Traps and Attractants for the Olive Fruit fly, Bαctrocerα oleαe

2017 ◽  
Vol 12 ◽  
pp. 71 ◽  
Author(s):  
G.E. Haniotakis ◽  
ΤΗ. Broumas ◽  
C. Liaropoulos
Keyword(s):  
Uv Light ◽  
Fruit Fly ◽  
Field Studies ◽  
Ammonium Bicarbonate ◽  
Protein Hydrolysates ◽  
Olive Fruit Fly ◽  
Mass Trapping ◽  
Olive Fruit ◽  
Olive Fly ◽  
High Concentrations

In the framework of an ongoing effort for development of an effective trap for the control of the olive fruit fly, Bactrocera oleae (Gmelin) (Diptera, Tephrifidae), by mass trapping, four trap types (wood, cloth, laminate, McPhail), seven different colors of laminate traps, different food attractants, attractant combinations or formulations (total of ten cases), and two insecticides (deltamethrin, b-cyfluthrin) used as killing agents on the traps, were compared under field conditions. The following conclusions may be drawn: Laminate traps (paper envelopes with a polyethylene lining inside, 15×20 cm in size, containing 70 g of ammonium bicarbonate salt and impregnated with 15 mg (a.i.) of an insecticide), were more effective than wood traps. Effectiveness of laminate traps can be enhanced by using an attracting color or in the case of the food attractant ammonium bicarbonate, by incorporating copper oxychloride. Combinations of two protein hydrolysates were more effective than one hydrolysate alone. Effectiveness of protein hydrolysates is not constantly higher than that of ammonium bicarbonate, the use of which is more economic and convenient. The insecticide b-cyfluthrin may be used on traps instead of deltamethrin, which is known to have a repellent effect on the olive fly, especially Linder high concentrations, and is unstable under natural UV light.

scholarly journals Full-Length Transcriptome Assembly of Italian Ryegrass Root Integrated with RNA-Seq to Identify Genes in Response to Plant Cadmium Stress

2020 ◽  
Vol 21 (3) ◽  
pp. 1067 ◽  
Author(s):  
Zhaoyang Hu ◽  
Yufei Zhang ◽  
Yue He ◽  
Qingqing Cao ◽  
Ting Zhang ◽  
...  
Keyword(s):  
De Novo ◽  
Transcriptome Assembly ◽  
Italian Ryegrass ◽  
Full Length ◽  
Rna Seq ◽  
Cd Stress ◽  
Toxic Heavy Metal ◽  
Regulatory Pathways ◽  
A Genome ◽  
Cd Translocation

Cadmium (Cd) is a toxic heavy metal element. It is relatively easily absorbed by plants and enters the food chain, resulting in human exposure to Cd. Italian ryegrass (Lolium multiflorum Lam.), an important forage cultivated widely in temperate regions worldwide, has the potential to be used in phytoremediation. However, genes regulating Cd translocation and accumulation in this species are not fully understood. Here, we optimized PacBio ISO-seq and integrated it with RNA-seq to construct a de novo full-length transcriptomic database for an un-sequenced autotetraploid species. With the database, we identified 2367 differentially expressed genes (DEGs) and profiled the molecular regulatory pathways of Italian ryegrass with Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis in response to Cd stress. Overexpression of a DEG LmAUX1 in Arabidopsis thaliana significantly enhanced plant Cd concentration. We also unveiled the complexity of alternative splicing (AS) with a genome-free strategy. We reconstructed full-length UniTransModels using the reference transcriptome, and 29.76% of full-length models had more than one isoform. Taken together, the results enhanced our understanding of the genetic diversity and complexity of Italian ryegrass under Cd stress and provided valuable genetic resources for its gene identification and molecular breeding.

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