Purifying selection drove the adaptation of mitochondrial genes along with correlation of gene rearrangements and evolutionary rates in two subfamilies of Whitefly (Insecta: Hemiptera).

Insect mitochondrial genomes (mitogenomes) are usually represented by a conserved gene order. Whiteflies exhibit gene rearrangement in their mitogenomes; however, understanding how nucleotide substitution rates shape gene rearrangement in whiteflies is unclear due to the limited number of mitogenomes. Additionally, the mechanisms by which selection pressure drives adaptations in mitochondrial genes in the two subfamilies of whiteflies are not yet known. Here, we analyzed 18 whitefly mitogenomes, including one newly generated mitogenome, to compare nucleotide substitution rates, selection pressure, and gene arrangements. The newly generated mitogenome is reported along with reannotation of Pealius mori and comparisons to other whitefly mitogenomes. Comparative studies on nucleotide composition of 18 whiteflies revealed the positive GC skewness, confirming the reversal of strand asymmetry. We found 11 rearranged gene orders within two subfamilies of whiteflies with 8-18 breakpoints of gene rearrangements. Members of the subfamily Aleyrodinae exhibit more complex pathways in the evolution of gene order as compared to the subfamily Aleurodicinae. Our findings also revealed that the increase or reduction of nucleotide substitution rates does not have an impact on any of the gene rearrangement scenarios depicting neutral correlation. Selection pressure analysis revealed that the mitogenomes from members of both the subfamilies Aleurodicinae and Aleyrodinae are characterized by intense purifying selection pressure.

Comparative analysis of the two suborders of Thysanoptera and characterization of the complete mitochondrial genome of Thrips parvispinus.

Thrips parvispinus is a serious sucking pest on a number of economically important crops in the oriental region. It has gained importance recently for its drastic range extension distribution as an invasive pest. Here, the complete mitochondrial genome (15,067 bp) of Thrips parvispinus was sequenced and characterized. It possesses 37 genes and the putative noncoding region is duplicated. Comparative analyses of nucleotide diversity, skewness, codon usage bias, and selection pressure in mitochondrial protein-coding genes of the available 31 thrips mitogenomes (24 Terebrantia + 7 Tubulifera) were performed. Phylogenetic analysis showed a sister relationship of T. parvispinus to the clade (T. florum + T. hawaiiensis). Phylogenetic analyses formed the monophyly of subfamilies Phlaeothripinae and Idolothripinae within the family Phlaeothripidae (Suborder Tubulifera). Low nucleotide diversity was indicative of reversal of strand asymmetry in the Tubulifera. Neutrality analysis showed that directional mutation plays a major role in shaping codon usage bias in both suborders. Principal component analysis indicated distinct codon usage patterns in each suborder. Our data suggested weaker selection constrains on Terebrantia than in the Tubulifera. More tubuliferan mitogenomes are required to resolve previous classification hypotheses and elucidate genome evolution in these two suborders.

Morphological and DNA Barcoding Evidence for Invasive Pest Thrips, Thrips parvispinus (Thripidae: Thysanoptera), Newly Recorded From India.

South East Asia pest thrips species, Thrips parvispinus (Karny), is a serious pest on a number of agricultural and horticultural crops in a number of plant families. Based on an integrated approach of morphology and DNA barcoding, invasion of this serious pest is reported first time from India on papaya plantations. Molecular data have corroborated with the morphological identification. Haplotyping data suggested that the Indonesia may be a probable source of invasion of this pest to India.

A new species of Psephenothrips (Thysanoptera:Phlaeothripidae) with one new record from India.

Psephenothrips uttarakhandensis sp. n. is described and illustrated from India. Also, Neohydatothrips xestosternitus (Han Cui) is newly recorded from India. The DNA barcode data using partial mitochondrial cytochrome c oxidase subunit I (mtCOI) from the holotype of the new species and one sequence of Neohydatothrips xestosternitus were generated and submitted to The Barcode of Life Data Systems (BOLD).

Gut microbial composition in developmental stages of gall inducing thrips Gynaikothrips uzeli and associated plant pathogenesis.

Gut bacteria play a crucial role in the several metabolic activity of the insects. In the present work, effort has been made to decipher the gut microbiota associated with the developmental stages of Gynaikothrips uzeli a gall inducing thrips along with their predicted functional role. Further, an effort has been made to correlate the bacterial communities with plant pathogenesis and thelytoky behaviour of G. uzeli. Findings obtained revealed that genus Arsenophonus dominated the total bacterial diversity and was transmitted vertically through the developmental stages. Further, it was observed that the high abundance of genus Arsenophonus promotes the thelytoky behaviour in G. uzeli and results in the killing of males. Furthermore, strong connecting link between Arsenophonus abundance and gall induction in F. benjamina was observed in the current dataset. G. uzeli being in the category of phloem sucking insect was known for the induction of galls and the current findings for the first time unveiled the facts that high abundance of genus Arsenophonus a well-known plant pathogen may be one of the major reason for inducing galls in F. benjamina. Moreover, PICRUSt2 analysis revealed that predicted functional pathways like biosynthesis of amino acids, and metabolism of carbon, nitrogen, carbohydrates and amino acids (e.g. Arginine, Alanine, Aspartate, Glutamate, Proline, Cysteine, Methionine, Glycine, Threonine, and Serine) were frequently noticed in profiles associated with all the developmental stages of G. uzeli. More to this, the high abundance of Arsenophonus in G. uzeli suggest that representatives of this genus may be resistant and/or tolerant to different antibacterial agents, alkaloids, flavonoids, and glycosides (e.g. quercetin). The correlation of bacterial diversity in pathogenicity can be extrapolated in different pest and vector species of other arthropods.

Complete mitogenome of the endangered and endemic Nicobar treeshrew (Tupaia nicobarica) and comparison with other Scandentians.

The Nicobar treeshrew (Tupaia nicobarica) is an endangered small mammal endemic to the Nicobar Island of the Andaman Sea, India regarded as an alternative experimental animal model in biomedical research. The present study aimed to assemble the first mitochondrial genome of T. nicobarica to elucidate its phylogenetic position with respect to other Scandentians. The structure and variation of the novel mitochondrial genome were analyzed and compared with other Scandentians. The complete mitogenome (17,164 bp) encodes 37 genes, including 13 protein-coding genes (PCGs), 22 transfer RNA (tRNAs), two ribosomal RNA (rRNAs), and one control region (CR). Most of the genes were encoded on majority strand, except nad6 and eight tRNAs. The nonsynonymous/synonymous ratio in all PCGs indicates strong negative selection among all Tupaiidae species. The comparative study of CRs revealed the occurrence of tandem repeats (CGTACA) found in T. nicobarica. The phylogenetic analyses (Maximum Likelihood and Bayesian Inference) showed distinct clustering of T. nicobarica with high branch supports and depict a substantial divergence time (12-19 MYA) from the ancestor lineage of Tupaiidae. The 16S rRNA dataset corroborates the taxonomic rank of two subspecies of T. nicobarica from the Great and Little Nicobar Islands. In the future, whole nuclear genome sequencing is necessary to further improve our understanding of evolutionary relationships among treeshrews, and will have implications for biomedical research.

A new species of Oneilliella from India with description of the female of Trachynotothrips brevispinis Masumoto amp; Okajima (Thysanoptera: Thripidae).

Oneilliella shivii sp.n. (Panchaetothripinae) is described from India as the second species in this genus. Trachynotothrips brevispinis and T. striatus (Thripinae) are newly recorded from India, and the first description provided of the female of T. brevispinis. Partial sequence data of mitochondrial cytochrome c oxidase (mtCOI) of these species were generated.

A new species of Mycterothrips Trybom (Thysanoptera: Thripidae) from India with new record of the genus Paithrips Nonaka amp; Jangvitaya.

Mycterothrips nainiae sp. n. (Thripinae) is described and illustrated from India, and one genus and species, Paithrips circularis Nonaka and Jangvitaya, is newly recorded from India. A key to species of Mycterothrips from India is also provided. The DNA barcode data using partial mitochondrial cytochrome c oxidase subunit I (mtCOI) from the holotype also five sequences of Paithrips circularis were generated.

Tryphactothrips rutherfordi (Bagnall) (Thysanoptera, Panchaetothripinae): first description of the male.

The subfamily Panchaetothripinae (family Thripidae) is represented by 140 species under 40 genera (Thrips Wiki 2019). In India, 36 species under 16 genera are recorded (Tyagi Kumar 2016, Tyagi et al. 2017, Rachana and Varatharajan, 2018, Johnson et al. 2019). The members of this subfamily are leaf-feeders and usually dark brown in colour with strongly reticulate sculpture on body, terminal antennal segments needle-shaped, tarsi 1- or 2-segmented, fore wing upper vein fused with costa. The genus Tryphactothrips was established by Bagnall (1919), and this genus remains monobasic with only Dinurothrips rutherfordi Bagnall from Sri Lanka as the type species. The genus Tryphactothrips can be distinguished from related genera by the presence of sculptured round areolae on abdominal segments. It is closely related to Anisopilothrips Stannard Mitri but can be identified by paired sigmoidal setae on abdominal tergites (absent in Anisopilothrips), mesonotum without complete median longitudinal split (complete median longitudinal in Anisopilothrips). Recently, a series of both sexes of Tryphactothrips rutherfordi with banded fore wings was collected on fern from Kerala state of India. Females were identified using published keys (Wilson 1975), and the male is here described for the first time. DNA was isolated from the studied specimens and partial fragment of mtCOI gene was amplified and sequenced (Tyagi et al. 2017). Four sequences were submitted in the GenBank (Accession No. MN627201 to MN627204). Photographs and illustrations were taken through a Leica Trinocular Microscope (Leica DM-1000) using Leica software application suite (LAS EZ 2.1.0). The studied specimens were deposited in the National Zoological Collections (NZC), Zoological Survey of India, Kolkata, India.

A new species of Apelaunothrips (Thysanoptera: Phlaeothripidae) from India with new synonym of Podothrips sasacola Kurosawa.

The first objective of this paper is to describe a new fungus-feeding species in the Phlaeothripinae genus Apelaunothrips, and to provide a key to the six species of this genus recorded from India. The second objective is to establish a new synonym in the Poaceae-associated genus, Podothrips, based on structural variation observed among recently collected samples from bamboo. The slide-mounted specimens were studied with a Leica Trinocular Microscope (Leica DM-1000) and a Leica software application suite (LAS EZ 2.1.0) was used to make line drawing and photographs.

The first complete mitochondrial genome of marigold pest thrips, Neohydatothrips samayunkur (Sericothripinae) and comparative analysis.

Complete mitogenomes from the order Thysanoptera are limited to representatives of the subfamily Thripinae. Therefore, in the present study, we sequenced the mitochondrial genome of Neohydatothrips samayunkur (15,295 bp), a member of subfamily Sericothripinae. The genome possesses the canonical 13 protein-coding genes (PCGs), 22 transfer RNA genes (tRNAs), and two ribosomal RNA genes (rRNAs) as well as two putative control regions (CRs). The majority strand was 77.42% A + T content, and 22.58% G + C with weakly positive AT skew (0.04) and negative GC skew (-0.03). The majority of PCGs start with ATN codons as observed in other insect mitochondrial genomes. The GCG codon (Alanine) was not used in N. samayunkur. Most tRNAs have the typical cloverleaf secondary structure, however the DHU stem and loop were absent in trnV and trnS1, while the TΨC loop was absent in trnR and trnT. The two putative control regions (CR1 and CR2) show 99% sequence similarity indicated a possible duplication, and shared 57 bp repeats were identified. N. samayunkur showed extensive gene rearrangements, with 11 PCGs, 22 tRNAs, and two rRNAs translocated when compared to the ancestral insect. The gene trnL2 was separated from the 'trnL2-cox2' gene block, which is a conserved, ancestral gene order found in all previously sequenced thrips mitogenomes. Both maximum likelihood (ML) and Bayesian inference (BI) phylogenetic trees resulted in similar topologies. The phylogenetic position of N. samayunkur indicates that subfamily Sericothripinae is sister to subfamily Thripinae. More molecular data from different taxonomic groups is needed to understand thrips phylogeny and evolution.

Molecular footprint of Frankliniella occidentalis from India: a vector of Tospoviruses.

The western flower thrips, F. occidentalis is a vector of Tospoviruses and native to Western North America and Mexico. The present study is based on collected F. occidentalis specimens from Karnataka state in southern India and morphologically identified through available keys. The generated DNA barcode data show 99-100% similarity with the database sequences of F. occidentalis. The phylogenetic analysis (NJ, ML, and BA) shows three distinct clades of F. occidentalis in the present dataset with high bootstrap supports and posterior probabilities. The K2P genetic distances further depicted high similarity of the generated sequences from India and Netherlands. The Clade-1 (India + Netherlands) also shows a close relationship with Clade-2 (Kenya) rather than Clade-3 (Canada + USA). This study recorded the first genetic footprint of F. occidentalis in India and indicated the gene flow from the Netherlands to India. The similar molecular techniques may help to detect the invasion of many alien species in the near future and assists the quarantine regulations to protect the native ecosystem.

DNA barcoding of selected Scirtothrips species (Thysanoptera) from India.

The members of the genus Scirtothrips are highly polyphagous, including major pest and vector species. We applied both morphology and molecular approaches to delimit the selected Scirtothrips species from India. Out of 43 generated barcode sequences, six sequences of three species (S. hitam, S. mangiferae, and S. malayensis) are the novel contribution in global database. The Bayesian (BA) phylogeny clearly distinguishes all the studied species with reciprocal monophyletic criteria and represents multiple clades in S. dorsalis and S. oligochaetus. The high Kimura-2-Parameter (K2P) genetic divergences were observed between the multiple clades of S. dorsalis (4.5-8.8%) and S. oligochaetus (6.4%), which indicating possible existence of cryptic diversity. The current study also provided the morphological keys for six Scirtothrips species including S. hitam as a new record to India.

One generic synonym and one new species of Phlaeothripidae from India (Thysanoptera).

Haplothripsshivendraii Tyagi & Kumar, sp. n. is described from Rajasthan state of India. The monobasic Austro-oriental genus Dyothrips Kudô is formally synonymised with Haplothrips.

Complete mitochondrial genome of Black Soft-shell Turtle (Nilssonia nigricans) and comparative analysis with other Trionychidae.

The characterization of mitochondrial genome has been evidenced as an efficient field of study for phylogenetic and evolutionary analysis in vertebrates including turtles. The aim of this study was to distinguish the structure and variability of the Trionychidae species mitogenomes through comparative analysis. The complete mitogenome (16796 bp) of an endangered freshwater turtle, Nilssonia nigricans was sequenced and annotated. The mitogenome encoded for 37 genes and a major non-coding control region (CR). The mitogenome was A + T biased (62.16%) and included six overlapping and 19 intergenic spacer regions. The Relative synonymous codon usage (RSCU) value was consistent among all the Trionychidae species; with the exception of significant reduction of Serine (TCG) frequency in N. nigricans, N. formosa, and R. swinhoei. In N. nigricans, most of the transfer RNAs (tRNAs) were folded into classic clover-leaf secondary structures with Watson-Crick base pairing except for trnS1 (GCT). The comparative analysis revealed that most of the tRNAs were structurally different, except for trnE (TTC), trnQ (TTG), and trnM (CAT). The structural features of tRNAs resulted ≥ 10 mismatched or wobble base pairings in 12 tRNAs, which reflects the nucleotide composition in both H- and L-strands. The mitogenome of N. nigricans also revealed two unique tandem repeats (ATTAT)8, and (TATTA)20 in the CR. Further, the conserved motif 5'-GACATA-3' and stable stem-loop structure was detected in the CRs of all Trionychidae species, which play an significant role in regulating transcription and replication in the mitochondrial genome. Further, the comparative analysis of Ka/Ks indicated negative selection in most of the protein coding genes (PCGs). The constructed Maximum Likelihood (ML) phylogeny using all PCGs showed clustering of N. nigricans with N. formosa. The resulting phylogeny illustrated the similar topology as described previously and consistent with the taxonomic classification. However, more sampling from different taxonomic groups of Testudines and studies on their mitogenomics are desirable for better understanding of the phylogenetic and evolutionary relationships.

The complete mitochondrial genome of Melon thrips, Thrips palmi (Thripinae): Comparative analysis.

The melon thrips, Thrips palmi is a serious pest and vector for plant viruses on a wide range of economically important crops. DNA barcoding evidenced the presence of cryptic diversity in T. palmi and that warrants exhaustive molecular studies. Our present study is on decoding the first complete mitochondrial genome of T. palmi (15,333 bp) through next-generation sequencing (NGS). The T. palmi mt genome contains 37 genes, including 13 Protein coding genes (PCGs), two ribosomal RNA (rRNAs), 22 transfer RNA (tRNAs), and two control regions (CRs). The majority strand of T. palmi revealed 78.29% A+T content, and 21.72% G+C content with positive AT skew (0.09) and negative GC skew (-0.06). The ATN initiation codons were observed in 12 PCGs except for cox1 which have unique start codon (TTG). The relative synonymous codon usage (RSCU) analysis revealed Phe, Leu, Ile, Tyr, Asn, Lys and Met were the most frequently used amino acids in all PCGs. The codon (CGG) which is assigned to Arginine in most insects but absent in T. palmi. The Ka/Ks ratio ranges from 0.078 in cox1 to 0.913 in atp8. We observed the typical cloverleaf secondary structure in most of the tRNA genes with a few exceptions; absence of DHU stem and loop in trnV and trnS, absence of DHU loop in trnE, lack of T-arm and loop in trnN. The T. palmi gene order (GO) was compared with ancestral GO and observed an extensive gene arrangement in PCGs, tRNAs and rRNAs. The cox2 gene was separated from the gene block 'cox2-trnL2' in T. palmi as compared with the other thrips mt genomes, including ancestor GO. Further, the nad1, trnQ, trnC, trnL1, trnV, trnF, rrnS, and rrnL were inversely transpositioned in T. palmi GO. The gene blocks 'trnQ-trnS2-trnD' and 'trnN-trnE-trnS1-trnL1' seems to be genus specific. The T. palmi mt genome contained 24 intergenic spacer regions and 12 overlapping regions. The 62 bp of CR2 shows the similarity with CR1 indicating a possible duplication. The occurrence of multiple CRs in thrips mt genomes seems to be a derived trait which needs further investigation. Although, the study depicted extensive gene rearrangements in T. palmi mt genome, but the negative GC skew reflects only strand asymmetry. Both the ML and BI phylogenetic trees revealed the close relationships of Thrips with Scirtothrips as compared to Frankliniella. Thus, more mt genomes of the diverse thrips species are required to understand the in-depth phylogenetic and evolutionary relationships.

DNA testing of edible crabs from seafood shops on the Odisha coast, India.

Seafood consumption is highly demanding due to the important source of protein it contains, as well as being rich in omega-3 fatty acids. However, the adulteration of seafood is an alarming issue worldwide, including India. This study deals with edible crabs from seafood shops on the Odisha state coast in eastern India. The generated DNA barcode sequences successfully identified most of the studied brachyuran crab species by similarity search results in global databases. The species were also delimited by significant genetic divergence and Neighbour-Joining phylogeny. Additionally, the study detected the contamination of unknown organisms in the commercialized crab recipes from seafood shops. The DNA based species detection of brachyuran crab may be useful to resolve many ambiguities in species identification and monitoring of commercialized seafood concerning food safety.

Danothrips litseae sp.n. (Thysanoptera, Thripidae) from India on leaves of Litsea.

Danothrips litseae sp. n. is described from Litsea sp. in North East India. This is the third species of the genus with distinct tergal craspeda, also weak, indistinct, irregular scallop-like projections on the sternites. Molecular data of mitochondrial cytochrome c oxidase (mtCOI) from specimens of this species are also generated as additional data for future studies.

A new species of Trichromothrips (Thysanoptera) from India with four new records.

Globally 6100 species of the Order Thysanoptera are reported, of which 739 are known from India (Tyagi & Kumar 2016). The purpose here is to describe from India one new species and record for the first time from this country four other species, representing three different families, and full nomenclatural details are available at ThripsWiki (2017). From three of the species, including the holotype of the new species, DNA was isolated and amplification of partial fragment of mtCOI gene was performed (Tyagi et al. 2017) with the sequences submitted to the Barcode of Life Database. Photographs and illustrations were taken through a Leica Trinocular Microscope (Leica DM-1000) using Leica software application suite (LAS EZ 2.1.0). Voucher specimens, also the new holotype, are deposited in the National Zoological Collections (NZC), Zoological Survey of India, Kolkata, India.

DNA Barcoding studies on Thrips in India: Cryptic species and Species complexes.

Thrips are one of the major sucking pest and vector of plant viruses causing huge economic loss in agriculture. The accurate identification of thrips is crucial for effective pest management strategies. However, morphology based identification has limitations and warrants integration of molecular data. We attempted the largest DNA barcoding initiative on 370 sequences of 89 thrips morphospecies including 104 novel sequences from 39 morphospecies, including the type specimens of four species. The results of multiple species delimitation methods (BIN, ABGD, GMYC and bPTP) were consistent for 73 species (82%) with their morphological identifications. A total of 107 molecular operational taxonomic units (MOTUs) was recovered for 89 morphospecies by superimposing multiple methods and applying a three level nomenclature system. We detected more than one MOTU in 14 morphospecies indicating to have cryptic diversity including, two major vector species (Frankliniella schultzei and Thrips palmi). However, four morphospecies (Thrips moundi, Thrips carthami, Haplothrips andersi and Haplothrips gowdeyi) showed low genetic distances between them with overlapping in barcode gap that requires further analysis with multiple molecular markers and more specimens from wide geographical areas for better taxonomic judgment. We also presented the advantage of simultaneous use of multiple delimitation methods for detection and identification of cryptic species.