Search results for: genome analysis

Authors: S. Najafi, E. Bertini, M. Pezzotti, G.B. Tornielli, S. Zenoni

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Grapevine is an important agricultural fruit crop plant consumed worldwide and with a key role in the global economy. Grapevine is strongly affected by both biotic and abiotic stresses, which impact grape growth at different stages, such as during plant and berry development and pre- and post-harvest, consequently causing significant economic losses. Recently global warming has propelled the anticipation of the onset of berry ripening, determining the reduction of a grape color and increased volatilization of aroma compounds. Climate change could negatively alter the physiological characteristics of the grape and affect the berry and wine quality. Modern plant breeding can provide tools such as genome editing for improving grape resilience traits while maintaining intact the viticultural and oenological quality characteristics of the genotype. This study aims at developing a platform for genome editing application in grapevine plants with the final goal to improve berry quality, biotic, and abiotic resilience traits. We chose to directly deliver ribonucleoproteins (RNP, preassembled Cas protein and guide RNA) into plant protoplasts, and, from these cell structures, regenerate grapevine plants edited in specific selected genes controlling traits of interest. Edited plants regenerated by somatic embryogenesis from protoplasts will then be sequenced and molecularly characterized. Embryogenic calli of Sultana and Shiraz cultivars were initiated from unopened leaves of in-vitro shoot tip cultures and from stamens, respectively. Leaves were placed on NB2 medium while stamens on callus initiation medium (PIV) medium and incubated in the dark at 28 °C for three months. Viable protoplasts, tested by FDA staining, isolated from embryogenic calli were cultured by disc method at 1*105 protoplasts/ml. Mature well-shaped somatic embryos developed directly in the protoplast culture medium two months later and were transferred in the light into to shooting medium for further growth. Regenerated plants were then transferred to the greenhouse; no phenotypic alterations were observed when compared to non in-vitro cultured plants. The performed experiments allowed to established an efficient protocol of embryogenic calli production, protoplast isolation, and regeneration of the whole plant through somatic embryogenesis in both Sultana and Shiraz. Regenerated plants, through direct somatic embryogenesis deriving from a single cell, avoid the risk of chimerism during the regeneration process, therefore improving the genome editing process. As pre-requisite of genome editing, an efficient method for transfection of protoplast by yellow fluorescent protein (YFP) marker genes was also established and experiments of direct delivery of CRISPR–Cas9 ribonucleoproteins (RNPs) in protoplasts to achieve efficient DNA-free targeted mutations are in progress.

Keywords: CRISPR-cas9, plant regeneration, protoplast isolation, Vitis vinifera

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Authors: Niousha Bagheri Khulenjani, Mohammad Saniee Abadeh

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Learning from very big datasets is a significant problem for most present data mining and machine learning algorithms. MicroRNA (miRNA) is one of the important big genomic and non-coding datasets presenting the genome sequences. In this paper, a hybrid method for the classification of the miRNA data is proposed. Due to the variety of cancers and high number of genes, analyzing the miRNA dataset has been a challenging problem for researchers. The number of features corresponding to the number of samples is high and the data suffer from being imbalanced. The feature selection method has been used to select features having more ability to distinguish classes and eliminating obscures features. Afterward, a Convolutional Neural Network (CNN) classifier for classification of cancer types is utilized, which employs a Genetic Algorithm to highlight optimized hyper-parameters of CNN. In order to make the process of classification by CNN faster, Graphics Processing Unit (GPU) is recommended for calculating the mathematic equation in a parallel way. The proposed method is tested on a real-world dataset with 8,129 patients, 29 different types of tumors, and 1,046 miRNA biomarkers, taken from The Cancer Genome Atlas (TCGA) database.

Keywords: cancer classification, feature selection, deep learning, genetic algorithm

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Authors: Yugander Arra, Florence Auguy, Melissa Stiebner, Sophie Chéron, Michael M. Wudick, Van Schepler-Luu, Sébastien Cunnac, Wolf B. Frommer, Laurence Albar

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Rice yellow mottle virus (RYMV) is one of the most important diseases affecting rice in Africa. The most promising strategy to reduce yield losses is the use of highly resistant varieties. The resistance gene RYMV2 is homolog of the Arabidopsis constitutive expression of pathogenesis related protein-5 (AtCPR5) nucleoporin gene. Resistance alleles are originating from African cultivated rice Oryza glaberrima, rarely cultivated, and are characterized by frameshifts or early stop codons, leading to a non-functional or truncated protein. Rice possesses two paralogs of CPR5 and function of these genes are unclear. Here, we evaluated the role of the two rice candidate nucleoporin paralogs OsCPR5.1 (pathogenesis-related gene 5; RYMV2) and OsCPR5.2 by CRISPR/Cas9 genome editing. Despite striking sequence and structural similarity, only loss-of-function of OsCPR5.1 led to full resistance, while loss-of-function oscpr5.2 mutants remained susceptible. Short N-terminal deletions in OsCPR5.1 also did not lead to resistance. In contrast to Atcpr5 mutants, neither OsCPR5.1 nor OsCPR5.2 knock out mutants showed substantial growth defects. Taken together, the candidate nucleoporin OsCPR5.1, but not its close homolog OsCPR5.2, plays a specific role for the susceptibility to RYMV, possibly by impairing the import of viral RNA or protein into the nucleus. Whereas gene introgression from O. glaberrima to high yielding O. sativa varieties is impaired by strong sterility barriers and the negative impact of linkage drag, genome editing of OsCPR5.1, while maintaining OsCPR5.2 activity, thus provides a promising strategy to generate O. sativa elite lines that are resistant to RYMV.

Keywords: CRISPR Cas9, genome editing, knock out mutant, recessive resistance, rice yellow mottle virus

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Authors: Lien Gysens, Bert Vanmechelen, Maarten Haspeslagh, Piet Maes, Ann Martens

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Bovine papillomavirus (BPV) types 1 and 2 play a central role in the etiology of the most common neoplasm in horses, the equine sarcoid. The unknown mechanism behind the unique variety in a clinical presentation on the one hand and the host-dependent clinical outcome of BPV-1 infection, on the other hand, indicate the involvement of additional factors. Earlier studies have reported the potential functional significance of intratypic sequence variants, along with the existence of sarcoid-sourced BPV variants. Therefore, intratypic sequence variation seems to be an important emerging viral factor. This study aimed to give a broad insight in sarcoid-sourced BPV variation and explore its potential association with disease presentation. In order to do this, a nanopore sequencing approach was successfully optimized for screening a wide spectrum of clinical samples. Specimens of each tumour were initially screened for BPV-1/-2 by quantitative real-time PCR. A custom-designed primer set was used on BPV-positive samples to amplify the complete viral genome in two multiplex PCR reactions, resulting in a set of overlapping amplicons. For phylogenetic analysis, separate alignments were made of all available complete genome sequences for BPV-1/-2. The resulting alignments were used to infer Bayesian phylogenetic trees. We found substantial genetic variation among sarcoid-derived BPV-1, although this variation could not be linked to disease severity. Several of the BPV-1 genomes had multiple major deletions. Remarkably, the majority of the cluster within the region coding for late viral genes. Together with the extensiveness (up to 603 nucleotides) of the described deletions, this suggests an altered function of L1/L2 in disease pathogenesis. By generating a significant amount of complete-length BPV genomes, we succeeded in introducing next-generation sequencing into veterinary research focusing on the equine sarcoid, thus facilitating the first report of both nanopore-based sequencing of complete sarcoid-sourced BPV-1/-2 and the simultaneous nanopore sequencing of multiple complete genomes originating from a single clinical sample.

Keywords: Bovine papillomavirus, equine sarcoid, horse, nanopore sequencing, phylogenetic analysis

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Authors: Fahimeh Palizban, Farshad Noravesh, Amir Hossein Saeidian, Mahya Mehrmohamadi

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In the recent decade, the emergence of liquid biopsy has significantly improved cancer monitoring and detection. Dying cells, including those originating from tumors, shed their DNA into the blood and contribute to a pool of circulating fragments called cell-free DNA. Accordingly, identifying the tissue origin of these DNA fragments from the plasma can result in more accurate and fast disease diagnosis and precise treatment protocols. Open chromatin regions are important epigenetic features of DNA that reflect cell types of origin. Profiling these features by DNase-seq, ATAC-seq, and histone ChIP-seq provides insights into tissue-specific and disease-specific regulatory mechanisms. There have been several studies in the area of cancer liquid biopsy that integrate distinct genomic and epigenomic features for early cancer detection along with tissue of origin detection. However, multimodal analysis requires several types of experiments to cover the genomic and epigenomic aspects of a single sample, which will lead to a huge amount of cost and time. To overcome these limitations, the idea of predicting OCRs from WGS is of particular importance. In this regard, we proposed a computational approach to target the prediction of open chromatin regions as an important epigenetic feature from cell-free DNA whole genome sequence data. To fulfill this objective, local sequencing depth will be fed to our proposed algorithm and the prediction of the most probable open chromatin regions from whole genome sequencing data can be carried out. Our method integrates the signal processing method with sequencing depth data and includes count normalization, Discrete Fourie Transform conversion, graph construction, graph cut optimization by linear programming, and clustering. To validate the proposed method, we compared the output of the clustering (open chromatin region+, open chromatin region-) with previously validated open chromatin regions related to human blood samples of the ATAC-DB database. The percentage of overlap between predicted open chromatin regions and the experimentally validated regions obtained by ATAC-seq in ATAC-DB is greater than 67%, which indicates meaningful prediction. As it is evident, OCRs are mostly located in the transcription start sites (TSS) of the genes. In this regard, we compared the concordance between the predicted OCRs and the human genes TSS regions obtained from refTSS and it showed proper accordance around 52.04% and ~78% with all and the housekeeping genes, respectively. Accurately detecting open chromatin regions from plasma cell-free DNA-seq data is a very challenging computational problem due to the existence of several confounding factors, such as technical and biological variations. Although this approach is in its infancy, there has already been an attempt to apply it, which leads to a tool named OCRDetector with some restrictions like the need for highly depth cfDNA WGS data, prior information about OCRs distribution, and considering multiple features. However, we implemented a graph signal clustering based on a single depth feature in an unsupervised learning manner that resulted in faster performance and decent accuracy. Overall, we tried to investigate the epigenomic pattern of a cell-free DNA sample from a new computational perspective that can be used along with other tools to investigate genetic and epigenetic aspects of a single whole genome sequencing data for efficient liquid biopsy-related analysis.

Keywords: open chromatin regions, cancer, cell-free DNA, epigenomics, graph signal processing, correlation clustering

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Authors: Yunfeng Shan, Xiaoliang Wang, Youjun Zhou

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Whole-genome data from two lungfish species, along with other species, present a valuable opportunity to reassess the longstanding debate regarding the evolutionary relationships among tetrapods, lungfishes, and coelacanths. However, the use of bootstrap support has become outdated for large-scale phylogenomic data. Without robust phylogenetic support, the phylogenetic trees become meaningless. Therefore, it is necessary to re-evaluate the phylogenies of tetrapods, lungfishes, and coelacanths using novel measures of phylogenetic support specifically designed for phylogenomic data, as the previous phylogenies were based on 100% bootstrap support. Our findings consistently provide strong evidence favoring lungfish as the closest living relative of tetrapods. This conclusion is based on high gene support confidence with confidence intervals exceeding 95%, high internode certainty, and high gene concordance factor. The evidence stems from two datasets containing recently deciphered whole genomes of two lungfish species, as well as five previous datasets derived from lungfish transcriptomes. These results yield fresh insights into the three hypotheses regarding the phylogenies of tetrapods, lungfishes, and coelacanths. Importantly, these hypotheses are not mere conjectures but are substantiated by a significant number of genes. Analyzing real biological data further demonstrates that the inclusion of additional taxa diminishes the number of orthologues and leads to more diverse tree topologies. Consequently, gene trees and species trees may not be identical even when whole-genome sequencing data is utilized. However, it is worth noting that many gene trees can accurately reflect the species tree if an appropriate number of taxa, typically ranging from six to ten, are sampled. Therefore, it is crucial to carefully select the number of taxa and an appropriate outgroup while excluding fast-evolving taxa as outgroups to mitigate the adverse effects of long-branch attraction (LBA) and achieve an accurate reconstruction of the species tree. This is particularly important as more whole-genome sequencing data becomes available.

Keywords: gene support confidence (GSC), origin of land vertebrates, coelacanth, two whole genomes of lungfishes, confidence intervals

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Authors: Narin Salehiyan

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The later sequencing of the complete genomes of Mycoplasma genitalium and M. pneumoniae has pulled in significant consideration to the atomic science of mycoplasmas, the littlest self-replicating living beings. It shows up that we are presently much closer to the objective of defining, in atomic terms, the complete apparatus of a self-replicating cell. Comparative genomics based on comparison of the genomic cosmetics of mycoplasmal genomes with those of other microbes, has opened better approaches of looking at the developmental history of the mycoplasmas. There's presently strong hereditary bolster for the speculation that mycoplasmas have advanced as a department of gram-positive microbes by a handle of reductive advancement. Amid this prepare, the mycoplasmas misplaced significant parcels of their ancestors’ chromosomes but held the qualities basic for life. In this way, the mycoplasmal genomes carry a tall rate of preserved qualities, incredibly encouraging quality comment. The critical genome compaction that happened in mycoplasmas was made conceivable by receiving a parasitic mode of life. The supply of supplements from their has clearly empowered mycoplasmas to lose, amid advancement, the qualities for numerous assimilative forms. Amid their advancement and adjustment to a parasitic mode of life, the mycoplasmas have created different hereditary frameworks giving a profoundly plastic set of variable surface proteins to avoid the have safe framework.

Keywords: mycoplasma, plasma, pathogen, genome

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Authors: Fatimazahrae Elbakri, Azeddine Ibrahimi, Meryem Lemrani, Dris Belghyti

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Leishmaniasis represents a major public health problem because of the number of cases recorded each year and the wide distribution of the disease. It is a parasitic disease of flagellated protozoa transmitted by the bite of certain species of sandfly, causing a spectrum of clinical pathology in humans ranging from disfiguring skin lesions to fatal visceral leishmaniasis. Cutaneous leishmaniasis due to Leishmania major is a polymorphic disease; in fact, the infection can be asymptomatic, localized, or disseminated. The objective of this work is to determine the genomic diversity that contributes to clinical variability by trying to identify the variation in chromosome number and to extract SNPs and SNPs and InDels; it is based on four sequences (WGS) of Leishmania major available on NCBI in Fastq form, from three countries: Tunisia, Algeria, and Israel, the analysis is set up from a pipeline to facilitate the discovery of genetic diversity, in particular SNP and chromosomal somy.

Keywords: Leshmania major, cutaneous Leishmania, NGS, genomic, somy, variant calling

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Authors: Benjamin A. Ha, Marco Morselli, Xinhui Paige Zhang, Elizabeth A. C. Heath-Heckman, Jonathan B. Puritz, David K. Jacobs

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Next-generation sequencing has enhanced the ability to acquire massive amounts of sequence data to address classic population genetic questions for non-model organisms. Targeted approaches allow for cost effective or more precise analyses of relevant sequences; although, many such techniques require a known genome and it can be costly to purchase probes from a company. This is challenging for non-model organisms with no published genome and can be expensive for large population genetic studies. Expressed exome capture sequencing (EecSeq) synthesizes probes in the lab from expressed mRNA, which is used to capture and sequence the coding regions of genomic DNA from a pooled suite of samples. A normalization step produces probes to recover transcripts from a wide range of expression levels. This approach offers low cost recovery of a broad range of genes in the genome. This research project expands on EecSeq to investigate if mRNA from one taxon may be used to capture relevant sequences from a series of increasingly less closely related taxa. For this purpose, we propose to use the endangered Northern Tidewater goby, Eucyclogobius newberryi, a non-model organism that inhabits California coastal lagoons. mRNA will be extracted from E. newberryi to create probes and capture exomes from eight other taxa, including the more at-risk Southern Tidewater goby, E. kristinae, and more divergent species. Captured exomes will be sequenced, analyzed bioinformatically and phylogenetically, then compared to previously generated phylogenies across this group of gobies. This will provide an assessment of the utility of the technique in cross-species studies and for analyzing low genetic variation within species as is the case for E. kristinae. This method has potential applications to provide economical ways to expand population genetic and evolutionary biology studies for non-model organisms.

Keywords: coastal lagoons, endangered species, non-model organism, target capture method

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Authors: Kesavan Markkandan, Ha Young Park, Seung-Il Yoo, Sin-Gi Park, Junhyung Park

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For genetic identification of crop cultivars, insertions/deletions (InDel) markers have been preferred currently because they are easy to use, PCR based, co-dominant and relatively abundant. However, new InDels need to be developed for genetic studies of new varieties due to the difference of allele frequencies in InDels among the population groups. These new varieties are evolved with low levels of genetic diversity in specific genome loci with high recombination rate. In this study, we described soybean barcode system approach based on InDel makers, each of which is specific to a variation block (VB), where the genomes split by all assumed recombination sites. Firstly, VBs in crop cultivars were mined for transferability to VB-specific InDel markers. Secondly, putative InDels in the VB regions were identified for the development of barcode system by analyzing particular cultivar’s whole genome data. Thirdly, common VB-specific InDels from all cultivars were selected by gel electrophoresis, which were converted as 2D barcode types according to comparing amplicon polymorphisms in the five cultivars to the reference cultivar. Finally, the polymorphism of the selected markers was assessed with other cultivars, and the barcode system that allows a clear distinction among those cultivars is described. The same approach can be applicable for other commercial crops. Hence, VB-based genetic identification not only minimize the molecular markers but also useful for assessing cultivars and for marker-assisted breeding in other crop species.

Keywords: variation block, polymorphism, InDel marker, genetic identification

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Authors: Zewdneh Zana Zate

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The study focuses on the evolutionary and ecological genomics of both wild and cultivated Coffea arabica L. at its center of origin, Ethiopia, aiming to uncover how this vital species may withstand future climate changes. Utilizing bioclimatic models, we project the future distribution of Arabica under varied climate scenarios for 2050 and 2080, identifying potential conservation zones and immediate risk areas. Through whole-genome resequencing of accessions from Ethiopian gene banks, this research assesses genetic diversity and divergence between wild and cultivated populations. It explores relationships, demographic histories, and potential hybridization events among Coffea arabica accessions to better understand the species' origins and its connection to parental species. This genomic analysis also seeks to detect signs of natural or artificial selection across populations. Integrating these genomic discoveries with ecological data, the study evaluates the current and future ecological and genomic vulnerabilities of wild Coffea arabica, emphasizing necessary adaptations for survival. We have identified key genomic regions linked to environmental stress tolerance, which could be crucial for breeding more resilient Arabica varieties. Additionally, our ecological modeling predicted a contraction of suitable habitats, urging immediate conservation actions in identified key areas. This research not only elucidates the evolutionary history and adaptive strategies of Arabica but also informs conservation priorities and breeding strategies to enhance resilience to climate change. By synthesizing genomic and ecological insights, we provide a robust framework for developing effective management strategies aimed at sustaining Coffea arabica, a species of profound global importance, in its native habitat under evolving climatic conditions.

Keywords: coffea arabica, climate change adaptation, conservation strategies, genomic resilience

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Authors: Moneesh Thakur, Hridayesh Prasad, Nikitasha Bora, Parimal Roy Choudhary, A. K. Samanta, Sanjeev Kumar

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Canine demodicosis is a common parasitic condition which involves dog skin. Demodicosis in dogs is due the prominent growth of Demodex. Out of various canine Demodex spp., Demodex canis is the most often involved species. Canine demodicosis can occur as either a localized or generalized form of demodicosis severely affect the dogs and in non-treated dogs may cause death. This study was planned with the aim to screen and characterize the 18S rRNA gene of isolated Demodex canis. A total of 1200 dogs were screened during this study period. The skin scrapings of all the suspected dogs were examined under a microscope at 100X magnification for the presence of Demodex canis. The skin scrapings positive for Demodex canis were examined using PCR for confirmation. A total of 35 dogs were confirmed a positive result for D. canis based on 18S rRNA gene amplification by PCR. Further, the 18S rRNA gene of isolated Demodex canis was cloned and sequenced for genome analysis. On the sequence analysis, it was found that isolated sequence (GenBank Accession No. MK177513) had close similarity (99.7%) to that of D. canis genotype of China (Accession No. MG372254).

Keywords: PCR, phylogenetic analysis, cloning and sequening, Demodex canis

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Authors: Maxence Queyrel, Edi Prifti, Alexandre Templier, Jean-Daniel Zucker

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Analysis of the human microbiome using metagenomic sequencing data has demonstrated high ability in discriminating various human diseases. Raw metagenomic sequencing data require multiple complex and computationally heavy bioinformatics steps prior to data analysis. Such data contain millions of short sequences read from the fragmented DNA sequences and stored as fastq files. Conventional processing pipelines consist in multiple steps including quality control, filtering, alignment of sequences against genomic catalogs (genes, species, taxonomic levels, functional pathways, etc.). These pipelines are complex to use, time consuming and rely on a large number of parameters that often provide variability and impact the estimation of the microbiome elements. Training Deep Neural Networks directly from raw sequencing data is a promising approach to bypass some of the challenges associated with mainstream bioinformatics pipelines. Most of these methods use the concept of word and sentence embeddings that create a meaningful and numerical representation of DNA sequences, while extracting features and reducing the dimensionality of the data. In this paper we present an end-to-end approach that classifies patients into disease groups directly from raw metagenomic reads: metagenome2vec. This approach is composed of four steps (i) generating a vocabulary of k-mers and learning their numerical embeddings; (ii) learning DNA sequence (read) embeddings; (iii) identifying the genome from which the sequence is most likely to come and (iv) training a multiple instance learning classifier which predicts the phenotype based on the vector representation of the raw data. An attention mechanism is applied in the network so that the model can be interpreted, assigning a weight to the influence of the prediction for each genome. Using two public real-life data-sets as well a simulated one, we demonstrated that this original approach reaches high performance, comparable with the state-of-the-art methods applied directly on processed data though mainstream bioinformatics workflows. These results are encouraging for this proof of concept work. We believe that with further dedication, the DNN models have the potential to surpass mainstream bioinformatics workflows in disease classification tasks.

Keywords: deep learning, disease prediction, end-to-end machine learning, metagenomics, multiple instance learning, precision medicine

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Authors: Ghada Badr, Manar Hosny, Nuha Bintayyash, Eman Albilali, Souad Larabi Marie-Sainte

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The median problem is significantly applied to derive the most reasonable rearrangement phylogenetic tree for many species. More specifically, the problem is concerned with finding a permutation that minimizes the sum of distances between itself and a set of three signed permutations. Genomes with equal number of genes but different order can be represented as permutations. In this paper, an algorithm, namely BeamGA median, is proposed that combines a heuristic search approach (local beam) as an initialization step to generate a number of solutions, and then a Genetic Algorithm (GA) is applied in order to refine the solutions, aiming to achieve a better median with the smallest possible reversal distance from the three original permutations. In this approach, any genome rearrangement distance can be applied. In this paper, we use the reversal distance. To the best of our knowledge, the proposed approach was not applied before for solving the median problem. Our approach considers true biological evolution scenario by applying the concept of common intervals during the GA optimization process. This allows us to imitate a true biological behavior and enhance genetic approach time convergence. We were able to handle permutations with a large number of genes, within an acceptable time performance and with same or better accuracy as compared to existing algorithms.

Keywords: median problem, phylogenetic tree, permutation, genetic algorithm, beam search, genome rearrangement distance

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Authors: Mona Heydari, Ehsan Motamedian, Seyed Abbas Shojaosadati

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Prediction of perturbations after genetic manipulation (especially gene knockout) is one of the important challenges in systems biology. In this paper, a new algorithm is introduced that integrates microarray data into the metabolic model. The algorithm was used to study the change in the cell phenotype after knockout of Gss gene in Escherichia coli BW25113. Algorithm implementation indicated that gene deletion resulted in more activation of the metabolic network. Growth yield was more and less regulating gene were identified for mutant in comparison with the wild-type strain.

Keywords: metabolic network, gene knockout, flux balance analysis, microarray data, integration

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Authors: Daniel Osorio, Janneth Gonzalez, Andres Pinzon

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In this work, proteins and metabolic pathways associated with the neuroprotective response mediated by the synthetic neurosteroid tibolone under a palmitate-induced inflammatory model were identified by flux balance analysis (FBA). Three different metabolic scenarios (‘healthy’, ‘inflamed’ and ‘medicated’) were modeled over a gene expression data-driven constructed tissue-specific metabolic reconstruction of mature astrocytes. Astrocyte reconstruction was built, validated and constrained using three open source software packages (‘minval’, ‘g2f’ and ‘exp2flux’) released through the Comprehensive R Archive Network repositories during the development of this work. From our analysis, we predict that tibolone executes their neuroprotective effects through a reduction of neurotoxicity mediated by L-glutamate in astrocytes, inducing the activation several metabolic pathways with neuroprotective actions associated such as taurine metabolism, gluconeogenesis, calcium and the Peroxisome Proliferator Activated Receptor signaling pathways. Also, we found a tibolone associated increase in growth rate probably in concordance with previously reported side effects of steroid compounds in other human cell types.

Keywords: astrocytes, flux balance analysis, genome scale metabolic reconstruction, inflammation, neuroprotection, tibolone

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Authors: Adel Alblihy, Reem Ali, Mashael Algethami, Ahmed Shoqafi, Michael S. Toss, Juliette Brownlie, Natalie J. Tatum, Ian Hickson, Paloma Ordonez Moran, Anna Grabowska, Jennie N. Jeyapalan, Nigel P. Mongan, Emad A. Rakha, Srinivasan Madhusudan

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Platinum resistance is a clinical challenge in ovarian cancer. Platinating agents induce DNA damage which activate Mre11 nuclease directed DNA damage signalling and response (DDR). Upregulation of DDR may promote chemotherapy resistance. Here we have comprehensively evaluated Mre11 in epithelial ovarian cancers. In clinical cohort that received platinum- based chemotherapy (n=331), Mre11 protein overexpression was associated with aggressive phenotype and poor progression free survival (PFS) (p=0.002). In the ovarian cancer genome atlas (TCGA) cohort (n=498), Mre11 gene amplification was observed in a subset of serous tumours (5%) which correlated highly with Mre11 mRNA levels (p<0.0001). Altered Mre11 levels was linked with genome wide alterations that can influence platinum sensitivity. At the transcriptomic level (n=1259), Mre11 overexpression was associated with poor PFS (p=0.003). ROC analysis showed an area under the curve (AUC) of 0.642 for response to platinum-based chemotherapy. Pre-clinically, Mre11 depletion by gene knock down or blockade by small molecule inhibitor (Mirin) reversed platinum resistance in ovarian cancer cells and in 3D spheroid models. Importantly, Mre11 inhibition was synthetically lethal in platinum sensitive XRCC1 deficient ovarian cancer cells and 3D-spheroids. Selective cytotoxicity was associated with DNA double strand break (DSB) accumulation, S-phase cell cycle arrest and increased apoptosis. We conclude that pharmaceutical development of Mre11 inhibitors is a viable clinical strategy for platinum sensitization and synthetic lethality in ovarian cancer.

Keywords: MRE11; XRCC1, ovarian cancer, platinum sensitization, synthetic lethality

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Authors: Debbarma Asis, Guha Chandan

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Tumor Protein-53 (P53) is one of the tumor suppressor proteins. P53 regulates the cell cycle that conserves stability by preventing genome mutation. It is named so as it runs as 53-kilodalton (kDa) protein on Polyacrylamide gel electrophoresis although the actual mass is 43.7 kDa. Experimental evidence has indicated that P53 cancer mutants loses tumor suppression activity and subsequently gain oncogenic activities to promote tumourigenesis. Tumor-specific DNA has recently been detected in the plasma of breast cancer patients. Detection of tumor-specific genetic materials in cancer patients may provide a unique and valuable tumor marker for diagnosis and prognosis. Commercially available MDA-468 breast cancer cell line was used for the proposed study.

Keywords: tumor protein (P53), cancer mutants, MDA-468, tumor suppressor gene

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Authors: Mohammad Vahed, Ana Sadeghitohidi, Majid Vahed, Hiroki Takahashi

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In the genomics field, the huge amounts of data have produced by the next-generation sequencers (NGS). Data volumes are very rapidly growing, as it is postulated that more than one billion bases will be produced per year in 2020. The growth rate of produced data is much faster than Moore's law in computer technology. This makes it more difficult to deal with genomics data, such as storing data, searching information, and finding the hidden information. It is required to develop the analysis platform for genomics big data. Cloud computing newly developed enables us to deal with big data more efficiently. Hadoop is one of the frameworks distributed computing and relies upon the core of a Big Data as a Service (BDaaS). Although many services have adopted this technology, e.g. amazon, there are a few applications in the biology field. Here, we propose a new algorithm to more efficiently deal with the genomics big data, e.g. sequencing data. Our algorithm consists of two parts: First is that BDaaS is applied for handling the data more efficiently. Second is that the hybrid method of MapReduce and Fuzzy logic is applied for data processing. This step can be parallelized in implementation. Our algorithm has great potential in computational analysis of genomics big data, e.g. de novo genome assembly and sequence similarity search. We will discuss our algorithm and its feasibility.

Keywords: big data, fuzzy logic, MapReduce, Hadoop, cloud computing

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Authors: L. Salomon, P. Sklenar

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The Andes hold the highest plant species diversity in the world. How this occurred is one of the most intriguing questions in studies addressing the origin and patterning of plant diversity worldwide. Recently, the explosive adaptive radiations found in high Andean groups have been pointed as triggers to this spectacular diversity. The Andes is the species-richest area for the biggest genus from the Asteraceae family: Senecio. There, the genus presents an incredible diversity of species, striking growth form variation, and large niche span. Even when some studies tried to disentangle the evolutionary story for some Andean species in Senecio, they obtained partially resolved and low supported phylogenies, as expected for recently radiated groups. The high-throughput sequencing (HTS) approaches have proved to be a powerful tool answering phylogenetic questions in those groups whose evolutionary stories are recent and traditional techniques like Sanger sequencing are not informative enough. Although these tools have been used to understand the evolution of an increasing number of Andean groups, nowadays, their scope has not been applied for Senecio. This project aims to contribute to a better knowledge of the mechanisms shaping the hyper diversity of Senecio in the Andean region, using HTS focusing on Senecio ser. Culcitium (Asteraceae), recently recircumscribed. Firstly, reconstructing a highly resolved and supported phylogeny, and after assessing the role of allopatric differentiation, hybridization, and genome duplication in the diversification of the group. Using the Hyb-Seq approach, combining target enrichment using Asteraceae COS loci baits and genome skimming, more than 100 new accessions were generated. HybPhyloMaker and HybPiper pipelines were used for the phylogenetic analyses, and another pipeline in development (Paralogue Wizard) was used to deal with paralogues. RAxML was used to generate gene trees and Astral for species tree reconstruction. Phyparts were used to explore as first step of gene tree discordance along the clades. Fully resolved with moderated supported trees were obtained, showing Senecio ser. Culcitium as monophyletic. Within the group, some species formed well-supported clades with morphologically related species, while some species would not have exclusive ancestry, in concordance with previous studies using amplified fragment length polymorphism (AFLP) showing geographical differentiation. Discordance between gene trees was detected. Paralogues were detected for many loci, indicating possible genome duplications; ploidy level estimation using flow cytometry will be carried out during the next months in order to identify the role of this process in the diversification of the group. Likewise, TreeSetViz package for Mesquite, hierarchical likelihood ratio congruence test using Concaterpillar, and Procrustean Approach to Cophylogeny (PACo), will be used to evaluate the congruence among different inheritance patterns. In order to evaluate the influence of hybridization and Incomplete Lineage Sorting (ILS) in each resultant clade from the phylogeny, Joly et al.'s 2009 method in a coalescent scenario and Paterson’s D-statistic will be performed. Even when the main discordance sources between gene trees were not explored in detail yet, the data show that at least to some degree, processes such as genome duplication, hybridization, and/or ILS could be involved in the evolution of the group.

Keywords: adaptive radiations, Andes, genome duplication, hybridization, Senecio

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Authors: Felice Elefant, Akanksha Bhatnaghar, Keegan Krick, Elizabeth Heller

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Context: The severity of Alzheimer’s Disease (AD) progression involves an interplay of genetics, age, and environmental factors orchestrated by histone acetyltransferase (HAT) mediated neuroepigenetic mechanisms. While disruption of Tip60 HAT action in neural gene control is implicated in AD, alternative mechanisms underlying Tip60 function remain unexplored. Altered RNA splicing has recently been highlighted as a widespread hallmark in the AD transcriptome that is implicated in the disease. Research Aim: The aim of this study was to identify a novel RNA binding/splicing function for Tip60 in human hippocampus and impaired in brains from AD fly models and AD patients. Methodology/Analysis: The authors used RNA immunoprecipitation using RNA isolated from 200 pooled wild type Drosophila brains for each of the 3 biological replicates. To identify Tip60’s RNA targets, they performed genome sequencing (DNB-SequencingTM technology, BGI genomics) on 3 replicates for Input RNA and RNA IPs by Tip60. Findings: The authors' transcriptomic analysis of RNA bound to Tip60 by Tip60-RNA immunoprecipitation (RIP) revealed Tip60 RNA targets enriched for critical neuronal processes implicated in AD. Remarkably, 79% of Tip60’s RNA targets overlap with its chromatin gene targets, supporting a model by which Tip60 orchestrates bi-level transcriptional regulation at both the chromatin and RNA level, a function unprecedented for any HAT to date. Since RNA splicing occurs co-transcriptionally and splicing defects are implicated in AD, the authors investigated whether Tip60-RNA targeting modulates splicing decisions and if this function is altered in AD. Replicate multivariate analysis of transcript splicing (rMATS) analysis of RNA-Seq data sets from wild-type and AD fly brains revealed a multitude of mammalian-like AS defects. Strikingly, over half of these altered RNAs were bonafide Tip60-RNA targets enriched for in the AD-gene curated database, with some AS alterations prevented against by increasing Tip60 in fly brain. Importantly, human orthologs of several Tip60-modulated spliced genes in Drosophila are well characterized aberrantly spliced genes in human AD brains, implicating disruption of Tip60’s splicing function in AD pathogenesis. Theoretical Importance: The authors' findings support a novel RNA interaction and splicing regulatory function for Tip60 that may underlie AS impairments that hallmark AD etiology. Data Collection: The authors collected data from RNA immunoprecipitation experiments using RNA isolated from 200 pooled wild type Drosophila brains for each of the 3 biological replicates. They also performed genome sequencing (DNBSequencingTM technology, BGI genomics) on 3 replicates for Input RNA and RNA IPs by Tip60. Questions: The question addressed by this study was whether Tip60 has a novel RNA binding/splicing function in human hippocampus and whether this function is impaired in brains from AD fly models and AD patients. Conclusions: The authors' findings support a novel RNA interaction and splicing regulatory function for Tip60 that may underlie AS impairments that hallmark AD etiology.

Keywords: Alzheimer's disease, cognition, aging, neuroepigenetics

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Authors: Mina Hojat Ansari, Kamran Bagheri Lankarani, Mohammad Reza Fattahi, Ali Reza Safarpour

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Irritable bowel syndrome (IBS) is a common bowel disorder which is usually diagnosed through the abdominal pain, fecal irregularities and bloating. Alteration in the intestinal microbial composition is implicating to inflammatory and functional bowel disorders which is recently also noted as an IBS feature. Owing to the potential importance of microbiota implication in both efficiencies of the treatment and prevention of the diseases, we examined the association between the intestinal microbiota and different bowel patterns in a cohort of subjects with IBS and healthy controls. Fresh fecal samples were collected from a total of 50 subjects, 30 of whom met the Rome IV criteria for IBS and 20 Healthy control. Total DNA was extracted and library preparation was conducted following the standard protocol for small whole genome sequencing. The pooled libraries sequenced on an Illumina Nextseq platform with a 2 × 150 paired-end read length and obtained sequences were analyzed using several bioinformatics programs. The majority of sequences obtained in the current study assigned to bacteria. However, our finding highlighted the significant microbial taxa variation among the studied groups. The result, therefore, suggests a significant association of the microbiota with symptoms and bowel characteristics in patients with IBS. These alterations in fecal microbiota could be exploited as a biomarker for IBS or its subtypes and suggest the modification of the microbiota might be integrated into prevention and treatment strategies for IBS.

Keywords: irritable bowel syndrome, intestinal microbiota, small whole genome sequencing, fecal samples, Illumina

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Authors: Olga Blagova, Yuliya Osipova, Evgeniya Kogan, Alexander Nedostup

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Purpose: to quantify the value of various clinical, laboratory and instrumental signs in the diagnosis of myocarditis in comparison with morphological studies of the myocardium. Methods: in 100 patients (65 men, 44.7±12.5 years) with «idiopathic» arrhythmias (n = 20) and dilated cardiomyopathy (DCM, n = 80) were performed 71 endomyocardial biopsy (EMB), 13 intraoperative biopsy, 5 study of explanted hearts, 11 autopsy with virus investigation (real-time PCR) of the blood and myocardium. Anti-heart antibodies (AHA) were also measured as well as cardiac CT (n = 45), MRI (n = 25), coronary angiography (n = 47). The comparison group included of 50 patients (25 men, 53.7±11.7 years) with non-inflammatory heart diseases who underwent open heart surgery. Results. Active/borderline myocarditis was diagnosed in 76.0% of the study group and in 21.6% of patients of the comparison group (p < 0.001). The myocardial viral genome was observed more frequently in patients of comparison group than in study group (group (65.0% and 40.2%; p < 0.01. Evaluated the diagnostic value of noninvasive markers of myocarditis. The panel of anti-heart antibodies had the greatest importance to identify myocarditis: sensitivity was 81.5%, positive and negative predictive value was 75.0 and 60.5%. It is defined diagnostic value of non-invasive markers of myocarditis and diagnostic algorithm providing an individual assessment of the likelihood of myocarditis is developed. Conclusion. The greatest significance in the diagnosis of latent myocarditis in patients with 'idiopathic' arrhythmias and DCM have AHA. The use of complex of noninvasive criteria allows estimate the probability of myocarditis and determine the indications for EMB.

Keywords: myocarditis, "idiopathic" arrhythmias, dilated cardiomyopathy, endomyocardial biopsy, viral genome, anti-heart antibodies

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Authors: Shagufta Jabeen, Faez Jesse Firdaus Abdullah, Zunita Zakaria, Nurulfiza Mat Isa, Yung Chie Tan, Wai Yan Yee, Abdul Rahman Omar

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Pasteurella multocida is associated with acute, as well as, chronic infections in avian and bovine such as pasteurellosis and hemorrhagic septicemia (HS) in cattle and buffaloes. Iron is one of the most important nutrients for pathogenic bacteria including Pasteurella and acts as a cofactor or prosthetic group in several essential enzymes and is needed for amino acid, pyrimidine, and DNA biosynthesis. In our recent study, we showed that 2% of Pasteurella multocida serotype A strain PMTB2.1 encode for iron regulating genes (Accession number CP007205.1). Genome sequencing of other Pasteurella multocida serotypes namely PM70 and HB01 also indicated up to 2.5% of the respective genome encode for iron regulating genes, suggesting that Pasteurella multocida genome comprises of multiple systems for iron uptake. Since P. multocida PMTB2.1 has more than 40 CDs out of 2097 CDs (approximately 2%), encode for iron-regulated. The gene expression profiling of four iron-regulating genes namely fbpb, yfea, fece and fur were characterized under iron-restricted environment. The P. multocida strain PMTB2.1 was grown in broth with and without iron chelating agent and samples were collected at different time points. Relative mRNA expression profile of these genes was determined using Taqman probe based real-time PCR assay. The data analysis, normalization with two house-keeping genes and the quantification of fold changes were carried out using Bio-Rad CFX manager software version 3.1. Results of this study reflect that iron reduced environment has significant effect on expression profile of iron regulating genes (p < 0.05) when compared to control (normal broth) and all evaluated genes act differently with response to iron reduction in media. The highest relative fold change of fece gene was observed at early stage of treatment indicating that PMTB2.1 may utilize its periplasmic protein at early stage to acquire iron. Furthermore, down-regulation expression of fece with the elevated expression of other genes at later time points suggests that PMTB2.1 control their iron requirements in response to iron availability by down-regulating the expression of iron proteins. Moreover, significantly high relative fold change (p ≤ 0.05) of fbpb gene is probably associated with the ability of P. multocida to directly use host iron complex such as hem, hemoglobin. In addition, the significant increase (p ≤ 0.05) in fbpb and yfea expressions also reflects the utilization of multiple iron systems in P. multocida strain PMTB2.1. The findings of this study are very much important as relative scarcity of free iron within hosts creates a major barrier to microbial growth inside host and utilization of outer-membrane proteins system in iron acquisition probably occurred at early stage of infection with P. multocida. In conclusion, the presence and utilization of multiple iron system in P. multocida strain PMTB2.1 revealed the importance of iron in the survival of P. multocida.

Keywords: iron-related genes, real-time PCR, gene expression profiling, fold changes

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Authors: Ghazi Chabchoub, Mouna Feki, Mohamed Abid, Hammadi Ayadi

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Autoimmune thyroid diseases (AITDs), including Graves’ disease (GD) and Hashimoto’s thyroiditis (HT), are inherited as complex traits. Genetic factors associated with AITDs have been tentatively identified by candidate gene and genome scanning approaches. We analysed three intragenic microsatellite markers in the thyroid hormone receptor associated protein 2 gene (THRAP2), mapped near D12S79 marker, which have a potential role in immune function and inflammation [THRAP2-1(TG)n, THRAP2-2 (AC)n and THRAP2-3 (AC)n]. Our study population concerned 12 patients affected with AITDs belonging to a multiplex Tunisian family with high prevalence of AITDs. Fluorescent genotyping was carried out on ABI 3100 sequencers (Applied Biosystems USA) with the use of GENESCAN for semi-automated fragment sizing and GENOTYPER peak-calling software. Statistical analysis was performed using the non parametric Lod score (NPL) by Merlin software. Merlin outputs non-parametric NPLall (Z) and LOD scores and their corresponding asymptotic P values. The analysis for three intragenic markers in the THRAP2 gene revealed strong evidence for linkage (NPL=3.68, P=0.00012). Our results suggested the possible role of THRAP2 gene in AITDs susceptibility in this family.

Keywords: autoimmunity, autoimmune disease, genetic, linkage analysis

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Authors: Xiaoping Su, Gabriel G. Malouf

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Introduction: Breast cancer is a heterogeneous disease that can be classified in 4 subgroups using transcriptional profiling. The role of lncRNA expression in human breast cancer biology, prognosis, and molecular classification remains unknown. Methods and results: Using an integrative comprehensive analysis of lncRNA, mRNA and DNA methylation in 900 breast cancer patients from The Cancer Genome Atlas (TCGA) project, we unraveled the molecular portraits of 1,700 expressed lncRNA. Some of those lncRNAs (i.e, HOTAIR) are previously reported and others are novel (i.e, HOTAIRM1, MAPT-AS1). The lncRNA classification correlated well with the PAM50 classification for basal-like, Her-2 enriched and luminal B subgroups, in contrast to the luminal A subgroup which behaved differently. Importantly, estrogen receptor (ESR1) expression was associated with distinct lncRNA networks in lncRNA clusters III and IV. Gene set enrichment analysis for cis- and trans-acting lncRNA showed enrichment for breast cancer signatures driven by breast cancer master regulators. Almost two third of those lncRNA were marked by enhancer chromatin modifications (i.e., H3K27ac), suggesting that lncRNA expression may result in increased activity of neighboring genes. Differential analysis of gene expression profiling data showed that lncRNA HOTAIRM1 was significantly down-regulated in basal-like subtype, and DNA methylation profiling data showed that lncRNA HOTAIRM1 was highly methylated in basal-like subtype. Thus, our integrative analysis of gene expression and DNA methylation strongly suggested that lncRNA HOTAIRM1 should be a tumor suppressor in basal-like subtype. Conclusion and significance: Our study depicts the first lncRNA molecular portrait of breast cancer and shows that lncRNA HOTAIRM1 might be a novel tumor suppressor.

Keywords: lncRNA profiling, breast cancer, HOTAIRM1, tumor suppressor

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Authors: Siddhant Sethi, Yasuharu Takashima, Shigetaka Nakamura, Kenzo Fujimoto

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Site-directed mutagenesis is a renowned technique to introduce specific mutations in the genome. To achieve site-directed mutagenesis, many chemical and enzymatic approaches have been reported in the past like disulphite induced genome editing, CRISPR-Cas9, TALEN etc. The chemical methods are invasive whereas the enzymatic approaches are time-consuming and expensive. Most of these techniques are unusable in the cellular application due to their toxicity and other limitations. Photo-chemical cytosine deamination, introduced in 2010, is one of the major technique for enzyme-free single-point mutation of cytosine to uracil in DNA and RNA, wherein, 3-cyanovinylcarbazole nucleoside (CNVK) containing oligodeoxyribonucleotide (ODN) having CNVK at -1 position to that of target cytosine is reversibly crosslinked to target DNA strand using 366 nm and then incubated at 90ºC to accommodate deamination. This technique is superior to enzymatic methods of site-directed mutagenesis but has a disadvantage that it requires the use of high temperature for the deamination step which restricts its applicability in the in vivo applications. This study has been focused on improving the technique by reducing the temperature required for deamination. Firstly, the photo-cross-linker, CNVK has been modified by replacing cyano group attached to vinyl group with methyl ester (OMeVK), amide (NH2VK), and carboxylic acid (OHVK) to observe the acceleration in the deamination of target cytosine cross-linked to vinylcarbazole derivative. Among the derivatives, OHVK has shown 2 times acceleration in deamination reaction as compared to CNVK, while the other two derivatives have shown deceleration towards deamination reaction. The trend of rate of deamination reaction follows the same order as that of hydrophilicity of the vinylcarbazole derivatives. OHVK being most hydrophilic has shown highest acceleration while OMeVK is least hydrophilic has proven to be least active for deamination. Secondly, in the related study, the counter-base of the target cytosine, guanine has been replaced by inosine, 2-aminopurine, nebularine, and 5-nitroindole having distinct hydrogen bonding patterns with target cytosine. Among the ODNs with these counter bases, ODN with inosine has shown 12 fold acceleration towards deamination of cytosine cross-linked to CNVK at physiological conditions as compared to guanosine. Whereas, when 2-aminopurine, nebularine, and 5-nitroindole were used, no deamination reaction took place. It can be concluded that inosine has potential to be used as the counter base of target cytosine for the CNVK mediated photo-cross-linking induced deamination of cytosine. The increase in rate of deamination reaction has been attributed to pattern and number of hydrogen bonding between the cytosine and counter base. One of the important factor is presence of hydrogen bond between exo-cyclic amino group of cytosine and the counter base. These results will be useful for development of more efficient technique for site-directed mutagenesis for C → U transformations in the DNA/RNA which might be used in the living system for treatment of various genetic disorders and genome engineering for making designer and non-native proteins.

Keywords: C to U transformation, DNA editing, genome engineering, ultra-fast photo-cross-linking

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Authors: Jianhua Zhang, Weiping Chen, Guanjie Chen, Jason Flannick, Emma Fikse, Glenda Smerin, Yanqin Yang, Yulong Li, John A. Hanover, William F. Simonds

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Ethnic disparities in many diseases are well recognized and reflect the consequences of genetic, behavior, and environmental factors. However, direct scientific evidence connecting the ethnic genetic variations and the disease disparities has been elusive, which may have led to the ethnic inequalities in large scale genetic studies. Through the genome-wide analysis of data representing 185,934 subjects, including 14,955 from our own studies of the African America Diabetes Mellitus, we discovered sets of genetic variants either unique to or conserved in all ethnicities. We further developed a quantitative gene function-based high-risk variant index (hrVI) of 20,428 genes to establish profiles that strongly correlate with the subjects' self-identified ethnicities. With respect to the ability to detect human essential and pathogenic genes, the hrVI analysis method is both comparable with and complementary to the well-known genetic analysis methods, pLI and VIRlof. Application of the ethnicity-specific hrVI analysis to the type 2 diabetes mellitus (T2DM) national repository, containing 20,791 cases and 24,440 controls, identified 114 candidate T2DM-associated genes, 8.8-fold greater than that of ethnicity-blind analysis. All the genes identified are defined as either pathogenic or likely-pathogenic in ClinVar database, with 33.3% diabetes-associated and 54.4% obesity-associated genes. These results demonstrate the utility of hrVI analysis and provide the first genetic evidence by clustering patterns of how genetic variations among ethnicities may impede the discovery of diabetes and foreseeably other disease-associated genes.

Keywords: diabetes-associated genes, ethnic health disparities, high-risk variant index, hrVI, T2DM

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Authors: Chul Lee, Seoae Cho, Erich D. Jarvis, Heebal Kim

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Vocal learning, the ability to imitate vocalizations based on auditory experience, is a homoplastic character state observed in different independent lineages of animals such as songbirds, parrots, hummingbirds and human. It has now become possible to perform genome-wide molecular analyses across vocal learners and vocal non-learners with the recent expansion of avian genome data. It was analyzed the whole genomes of human and 48 avian species including those belonging to the three avian vocal learning lineages, to determine if behavior and neural convergence are associated with molecular convergence in divergent species of vocal learners. Analyses of 8295 orthologous genes across bird species revealed 141 genes with amino acid substitutions specific to vocal learners. Out of these, 25 genes have vocal learner specific genetic homoplasies, and their functions were enriched for learning. Several sites in these genes are estimated under convergent evolution and positive selection. A potential role for a subset of these genes in vocal learning was supported by associations with gene expression profiles in vocal learning brain regions of songbirds and human disease that cause language dysfunctions. The key candidate gene with multiple independent lines of the evidences specific to vocal learners was DRD5. Our findings suggest cAMP-based learning pathway in avian vocal learners, indicating molecular homoplastic changes associated with a complex behavioral trait, vocal learning.

Keywords: amino acid substitutions, convergent evolution, positive selection, vocal learning

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Authors: Kaomud Tyagi, Rajasree Chakraborty, Shantanu Kundu, Devkant Singha, Kailash Chandra, Vikas Kumar

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The melon thrips, Thrips palmi is a serious pest of a wide range of agriculture crops and also act as vectors for plant viruses (genus Tospovirus, family Bunyaviridae). More molecular data on this species is required to understand the cryptic speciation and evolutionary affiliations. Mitochondrial genomes have been widely used in phylogenetic and evolutionary studies in insect. So far, mitogenomes of five thrips species (Anaphothrips obscurus, Frankliniella intonsa, Frankliniella occidentalis, Scirtothrips dorsalis and Thrips imaginis) is available in the GenBank database. In this study, we sequenced the first complete mitogenome T. palmi and compared it with available thrips mitogenomes. We assembled the mitogenome from the whole genome sequencing data generated using Illumina Hiseq2500. Annotation was performed using MITOS web-server to estimate the location of protein coding genes (PCGs), transfer RNA (tRNAs), ribosomal RNAs (rRNAs) and their secondary structures. The boundaries of PCGs and rRNAs was confirmed manually in NCBI. Phylogenetic analyses were performed using the 13 PCGs data using maximum likelihood (ML) in PAUP, and Bayesian inference (BI) in MrBayes 3.2. The complete mitogenome of T. palmi was 15,333 base pairs (bp), which was greater than the genomes of A. obscurus (14,890bp), F. intonsa (15,215 bp), F. occidentalis (14,889 bp) and S. dorsalis South Asia strain (SA1) (14,283 bp), but smaller than the genomes of T. imaginis (15,407 bp) and S. dorsalis East Asia strain (EA1) (15,343bp). Like in other thrips species, the mitochondrial genome of T. palmi was represented by 37 genes, including 13 PCGs, large and small ribosomal RNA (rrnL and rrnS) genes, 22 transfer RNA (tRNAs) genes (with one extra gene for trn-Serine) and two A+T-rich control regions (CR1 and CR2). Thirty one genes were observed on heavy (H) strand and six genes on the light (L) strand. The six tRNA genes (trnG,trnK, trnY, trnW, trnF, and trnH) were found to be conserved in all thrips species mitogenomes in their locations relative to a protein-coding or rRNA gene upstream or downstream. The gene arrangements of T. palmi is very close to T. imaginis except the rearrangements in tRNAs genes: trnR (arginine), and trnE (glutamic acid) were found to be located between cox3 and CR2 in T. imaginis which were translocated between atp6 and CR1 in T. palmi; trnL1 (Leucine) and trnS1(Serine) were located between atp6 and CR1 in T. imaginis which were translocated between cox3 and CR2 in T. palmi. The location of CR1 upstream of nad5 gene was suggested to be ancestral condition of the thrips species in subfamily Thripinae, was also observed in T. palmi. Both the Maximum likelihood (ML) and Bayesian Inference (BI) phylogenetic trees generated resulted in similar topologies. The T. palmi was clustered with T. imaginis. We concluded that more molecular data on the diverse thrips species from different hierarchical level is needed, to understand the phylogenetic and evolutionary relationships among them.

Keywords: thrips, comparative mitogenomics, gene rearrangements, phylogenetic analysis

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