MRF: a tool to overcome the barrier of inconsistent genome annotations and perform comparative genomics studies for the largest animal DNA virus.

BACKGROUND: The genome of the largest known animal virus, the white spot syndrome virus (WSSV) responsible for huge economic losses and loss of employment in aquaculture, suffers from inconsistent annotation nomenclature. Novel genome sequence, circular genome and variable genome length led to nomenclature inconsistencies. Since vast knowledge has already accumulated in the past two decades with inconsistent nomenclature, the insights gained on a genome could not be easily extendable to other genomes. Therefore, the present study aims to perform comparative genomics studies in WSSV on uniform nomenclature. METHODS: We have combined the standard mummer tool with custom scripts to develop missing regions finder (MRF) that documents the missing genome regions and coding sequences in virus genomes in comparison to a reference genome and in its annotation nomenclature. The procedure was implemented as web tool and in command-line interface. Using MRF, we have documented the missing coding sequences in WSSV and explored their role in virulence through application of phylogenomics, machine learning models and homologous genes. RESULTS: We have tabulated and depicted the missing genome regions, missing coding sequences and deletion hotspots in WSSV on a common annotation nomenclature and attempted to link them to virus virulence. It was observed that the ubiquitination, transcription regulation and nucleotide metabolism might be essentially required for WSSV pathogenesis; and the structural proteins, VP19, VP26 and VP28 are essential for virus assembly. Few minor structural proteins in WSSV would act as envelope glycoproteins. We have also demonstrated the advantage of MRF in providing detailed graphic/tabular output in less time and also in handling of low-complexity, repeat-rich and highly similar regions of the genomes using other virus cases. CONCLUSIONS: Pathogenic virus research benefits from tools that could directly indicate the missing genomic regions and coding sequences between isolates/strains. In virus research, the analyses performed in this study provides an advancement to find the differences between genomes and to quickly identify the important coding sequences/genomes that require early attention from researchers. To conclude, the approach implemented in MRF complements similarity-based tools in comparative genomics involving large, highly-similar, length-varying and/or inconsistently annotated viral genomes.

Transcriptomic variations associated with salinity stress in Penaeus indicus.

BACKGROUND: The Indian white shrimp, Penaeus indicus a native species of India, is important brackishwater aquaculture species. Shrimps are euryhaline in nature and they regulate osmotic and ionic concentrations by osmoregulatory process. However, variations in abiotic factors such as salinity result in stress to the shrimps during culture period affecting their growth and immunity. METHODS AND RESULTS: To understand the adaptive mechanism to stress in low salinity conditions, RNA-seq was used to compare the transcriptomic response of P. indicus upto 3 weeks. De novo assembly using Trinity assembler generated a total of 173,582 transcripts. The assembly had a mean length of 854 bp, N50 value of 1243 bp and GC content of 42.33%. Differential gene expression analysis, resulted in identification of 2130, 3090, and 5351 DEGs in 7 days, 14 days and 21 days respectively of salinity stress period. The pathway prediction of the assembled trinity transcripts using KEGG database showed total number of 329 pathways linking 12,430 transcripts. KEGG pathway enrichment analyses led to the identification of several enriched pathways related to lipid metabolism, amino acid metabolism, glycolysis, signalling pathways etc. Selected genes involved in osmoregulatory process and immune response in shrimps were validated and analysed for the gene expression levels by quantitative real-time PCR (qPCR). CONCLUSION: This study on the adaptive transcriptomic response of P. indicus to low salinity, will further help in our understanding of the molecular mechanisms underlying osmoregulation mechanism in shrimps.

Genome assembly, Full-length transcriptome, and isoform diversity of Red Snapper, Lutjanus argentimaculatus.

The mangrove red snapper, Lutjanus argentimaculatus, is a marine food fish of economic and aquaculture importance. The application of genomic selection-based breeding programs for this species is limited by the absence of a reference genome and transcriptome profiles. The current study attempted to fill this void by generating genomic and transcriptomic resources for red snapper. Using PacBio long reads, and Arima Hi-C linked reads, a scaffold-level genome assembly was generated for L. argentimaculatus. The assembly is of 1.03 Gb comprising of 400 scaffolds with N50 of 33.8 Mb and was assessed to be 97.2% complete upon benchmarking with BUSCO. Full-length transcriptome generated with PacBio Iso-Sequencing strategy using six tissues (muscle, gills, liver, kidney, stomach, and gonad) contained 56,515 isoforms belonging to 18,108 unique genes with N50 length of 3,973 bp. The resources generated will have potential applications in the functional studies, conservation, broodstock management and selective breeding programmes of L. argentimaculatus.

Phylogenetic relations and mitogenome-wide similarity metrics reveal monophyly of Penaeus sensu lato.

Splitting of the genus Penaeus sensu lato into six new genera based on morphological features alone has been controversial in penaeid shrimp taxonomy. Several studies focused on building phylogenetic relations among the genera of Penaeus sensu lato. However, they lack in utilizing full mitochondrial DNA genome of shrimp representing all the six controversial genera. For the first time, the present study targeted the testing of all the six genera of Penaeus sensu lato for phylogenetic relations utilizing complete mitochondrial genome sequence. In addition, the study reports for the first time about the complete mitochondrial DNA genome sequence of Fenneropenaeus indicus, an important candidate species in aquaculture and fisheries, and utilized it for phylogenomics. The maximum likelihood and Bayesian approaches were deployed to generate and comprehend the phylogenetic relationship among the shrimp in the suborder, Dendrobranchiata. The phylogenetic relations established with limited taxon sampling considered in the study pointed to the monophyly of Penaeus sensu lato and suggested collapsing of the new genera to a single genus. Further, trends in mitogenome-wide estimates of average amino acid identity in the order Decapoda and the genus Penaeus sensu lato supported restoration of the old genus, Penaeus, rather promoting the creation of new genera.

The complete chloroplast genome data of Areca catechu (Arecaceae).

Areca is a genus comprising about 50 species endemic to the humid tropics. Arecanut (Areca catechu L.) is a commercially and economically important crop in South and Southeast Asia. In addition to its contribution to the agricultural economies of countries where the crop is grown, arecanut holds an important place in the religious, cultural, and social milieu of the rural folks. The nuts have been used since time immemorial in traditional Indian (Unani and Ayurveda) and Chinese herbal systems of medicine for the treatment of various disorders like rheumatism, parasitic infection, diseases of gastrointestinal tracts, and depression. Here, we report the complete chloroplast (cp) genome sequence of arecanut. The cp genome of A. catechu was a typical circular DNA molecule with a size of 158,689 bp in length. The genome possessed a typical quadripartite structure composed of a pair of inverted repeats (IRa and IRb) of 27,137 bp separated by a large single-copy (LSC) region of 86,814 bp and a small single-copy (SSC) region of 17,601 bp and a GC content of 37.3%. The cp genome of arecanut encodes a set of 133 genes, comprising 88 protein-coding genes, 37 tRNA genes, and eight rRNA genes; among these, 21 contained introns. A total of 70 SSR loci were detected, the majority being in inter-genic regions. Phylogenetic analysis revealed that A. catechu was closely related to A. vestiaria.

RNA sequencing and de novo assembly of Solanum trilobatum leaf transcriptome to identify putative transcripts for major metabolic pathways.

Solanum trilobatum L. is an important medicinal plant in traditional Indian system of medicine belonging to Solanaceae family. However, non-availability of genomic resources hinders its research at the molecular level. We have analyzed the S. trilobatum leaf transcriptome using high throughput RNA sequencing. The de novo assembly of 136,220,612 reads produced 128,934 non-redundant unigenes with N50 value of 1347 bp. Annotation of unigenes was performed against databases such as NCBI nr database, Gene Ontology, KEGG, Uniprot, Pfam, and plnTFDB. A total of 60,097 unigenes were annotated including 48 Transcription Factor families and 14,490 unigenes were assigned to 138 pathways using KEGG database. The pathway analysis revealed the transcripts involved in the biosynthesis of important secondary metabolites contributing for its medicinal value such as Flavonoids. Further, the transcripts were quantified using RSEM to identify the highly regulated genes for secondary metabolism. Reverse-Transcription PCR was performed to validate the de novo assembled unigenes. The expression profile of selected unigenes from flavonoid biosynthesis pathway was analyzed using qRT-PCR. We have also identified 13,262 Simple Sequence Repeats, which could help in molecular breeding. This is the first report of comprehensive transcriptome analysis in S. trilobatum and this will be an invaluable resource to understand the molecular basis related to the medicinal attributes of S. trilobatum in further studies.

The complete chloroplast genome sequence of Indian mustard (Brassica juncea L.).

Brassica juncea L. is the second most important edible oil seed crop of India. In the current study, we report the complete chloroplast genome sequence of Indian mustard, Brassica juncea L. The size of the complete chloroplast genome was found to be 153 483 bp long with an overall GC content of 36.36%. A large single copy (LSC) region of 83 286 bp and a short single copy region (SSC) region of 17 775 bp were separated by a pair of inverted repeat (IR) regions of 26 211 bp. A total of 113 unique genes were annotated, which includes 79 protein-coding genes, 30 tRNA genes, and four rRNA genes. Either one or two introns were present in nine protein-coding genes and six tRNA genes. Phylogenetic analysis with the complete chloroplast genomes of other related species revealed that B. juncea is much closer to Brassica rapa.