Mitogenome-wise codon usage pattern from comparative analysis of the first mitogenome of Blepharipa sp. (Muga uzifly) with other Oestroid flies.

Uziflies (Family: Tachinidae) are dipteran endoparasites of sericigenous insects which cause major economic loss in the silk industry globally. Here, we are presenting the first full mitogenome of Blepharipa sp. (Acc: KY644698, 15,080 bp, A + T = 78.41%), a dipteran parasitoid of Muga silkworm (Antheraea assamensis) found in the Indian states of Assam and Meghalaya. This study has confirmed that Blepharipa sp. mitogenome gene content and arrangement is similar to other Tachinidae and Sarcophagidae flies of Oestroidea superfamily, typical of ancestral Diptera. Although, Calliphoridae and Oestridae flies have undergone tRNA translocation and insertion, forming unique intergenic spacers (IGS) and overlapping regions (OL) and a few of them (IGS, OL) have been conserved across Oestroidea flies. The Tachinidae mitogenomes exhibit more AT content and AT biased codons in their protein-coding genes (PCGs) than the Oestroidea counterpart. About 92.07% of all (3722) codons in PCGs of this new species have A/T in their 3rd codon position. The high proportion of AT and repeats in the control region (CR) affects sequence coverage, resulting in a short CR (Blepharipa sp.: 168 bp) and a smaller tachinid mitogenome. Our research unveils those genes with a high AT content had a reduced effective number of codons, leading to high codon usage bias. The neutrality test shows that natural selection has a stronger influence on codon usage bias than directed mutational pressure. This study also reveals that longer PCGs (e.g., nad5, cox1) have a higher codon usage bias than shorter PCGs (e.g., atp8, nad4l). The divergence rates increase nonlinearly as AT content at the 3rd codon position increases and higher rate of synonymous divergence than nonsynonymous divergence causes strong purifying selection. The phylogenetic analysis explains that Blepharipa sp. is well suited in the family of insectivorous tachinid maggots. It's possible that biased codon usage in the Tachinidae family reduces the effective number of codons, and purifying selection retains the core functions in their mitogenome, which could help with efficient metabolism in their endo-parasitic life style and survival strategy.

The mitochondrial genome of Muga silkworm (Antheraea assamensis) and its comparative analysis with other lepidopteran insects.

Muga (Antheraea assamensis) is an economically important silkmoth endemic to the states of Assam and Meghalaya in India and is the producer of the strongest known commercial silk. However, there is a scarcity of genomic and proteomic data for understanding the organism at a molecular level. Our present study is on decoding the complete mitochondrial genome (mitogenome) of A. assamensis using next generation sequencing technology and comparing it with other available lepidopteran mitogenomes. Mitogenome of A. assamensis is an AT rich circular molecule of 15,272 bp (A+T content ~80.2%). It contains 37 genes comprising of 13 protein coding genes (PCGs), 22 tRNA and 2 rRNA genes along with a 328 bp long control region. Its typical tRNAMet-tRNAIle-tRNAGln arrangement differed from ancestral insects (tRNAIle-tRNAGln-tRNAMet). Two PCGs cox1 and cox2 were found to have CGA and GTG as start codons, respectively as reported in some lepidopterans. Interestingly, nad4l gene showed higher transversion mutations at intra-species than inter-species level. All PCGs evolved under strong purifying selection with highest evolutionary rates observed for atp8 gene while lowest for cox1 gene. We observed the typical clover-leaf shaped secondary structures of tRNAs with a few exceptions in case of tRNASer1 and tRNATyr where stable DHU and TΨC loop were absent. A significant number of mismatches (35) were found to spread over 19 tRNA structures. The control region of mitogenome contained a six bp (CTTAGA/G) deletion atypical of other Antheraea species and lacked tandem repeats. Phylogenetic position of A. assamensis was consistent with the traditional taxonomic classification of Saturniidae. The complete annotated mitogenome is available in GenBank (Accession No. KU379695). To the best of our knowledge, this is the first report on complete mitogenome of A. assamensis.

De novo transcriptome of the muga silkworm, Antheraea assamensis (Helfer).

Antheraea assamensis (Lepidoptera: Saturniidae), is a semi-domesticated silkworm known to be endemic to Assam and the adjoining hilly areas of Northeast India. It is the only producer of a unique, commercially important variety of golden silk called "muga silk". Herein, we report the de novo transcriptome of A. assamensis reared on Machilus bombycina leaves for the first time. Short reads generated by high throughput sequencing of cDNA libraries from multiple tissues, viz. alimentary canal, silk gland and residual body of the 5th instar of muga silkworm were assembled into transcripts via a de novo assembly pipeline followed by functional annotation and classification. A total of 1,21,433 transcripts were generated from ~231 million raw reads of which ~74% (89,583) were either allocated a functional annotation or categorized under Pfam/COG/KEGG categories. Identification of differentially expressed transcripts and their comparative sequence analysis revealed candidate genes related to silk synthesis, viz. silk gland factor-1 and 3, sericin-like transcript, etc. with conserved forkhead, homeo- and POU domains. Several candidate anti-microbial peptides which may have potential anti-bacterial, anti-fungal or anti-parasitic activity in A. assamensis were also identified. T/A and AT/TA were predicted to be the most abundant mono- and di-nucleotide simple sequence repeat markers in the transcriptome. Transcriptome validation was carried out by quantitative real-time PCR (qPCR) amplification of eight transcripts. The resources generated by this study will expand the periphery of existing genomic data on A. assamensis facilitating future in-depth studies on its unknown aspects.