Publications by authors named "Mohammad Vatanparast"

22 Publications

  • Page 1 of 1

Comparative RNA-Seq Analyses of (Hymenoptera: Formicidae) Reveal Gene in Response to Cold Stress.

Genes (Basel) 2021 Oct 13;12(10). Epub 2021 Oct 13.

Plant Quarantine Technology Center, Animal and Plant Quarantine Agency, Gimcheon 39660, Korea.

, as a fire ant species, shows some predatory behavior towards earthworms and woodlice, and preys on the larvae of other ant species by tunneling into a neighboring colony's brood chamber. This study focused on the molecular response process and gene expression profiles of to low (9 °C)-temperature stress in comparison with normal temperature (25 °C) conditions. A total of 89,657 unigenes (the clustered non-redundant transcripts that are filtered from the longest assembled contigs) were obtained, of which 32,782 were annotated in the NR (nonredundant protein) database with gene ontology (GO) terms, gene descriptions, and metabolic pathways. The results were 81 GO subgroups and 18 EggNOG (evolutionary genealogy of genes: Non-supervised Orthologous Groups) keywords. Differentially expressed genes (DEGs) with logfold change (FC) > 1 and logFC < -1 with -value ≤ 0.05 were screened for cold stress temperature. We found 215 unigenes up-regulated and 115 unigenes down-regulated. Comparing transcriptome profiles for differential gene expression resulted in various DE proteins and genes, including fatty acid synthases and lipid metabolism, which have previously been reported to be involved in cold resistance. We verified the RNA-seq data by qPCR on 20 up- and down-regulated DEGs. These findings facilitate the basis for the future understanding of the adaptation mechanisms of and the molecular mechanisms underlying the response to low temperatures.
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http://dx.doi.org/10.3390/genes12101610DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8535336PMC
October 2021

Knockdown of Helicoverpa armigera protease genes affects its growth and mortality via RNA interference.

Arch Insect Biochem Physiol 2021 Nov 26;108(3):e21840. Epub 2021 Sep 26.

Department of Plant Quarantine Technology Center, Animal and Plant Quarantine Agency, Gimcheon, South Korea.

Helicoverpa armigera (Hübner) (Lepidoptera: Noctuidae), the cotton bollworm, is a destructive pest which is famous for its resistance to a variety of insecticides. RNA interference is a posttranscriptional gene silencing mechanism that has become a popular tool to control insect pests, triggered by double-stranded RNAs (dsRNAs). The effect of ingestion and injection delivery methods of dsRNA related to some protease genes including Trypsin (Ha-TRY39 and Ha-TRY96), Chymotrypsin (Ha-CHY), and Cathepsin L (Ha-CAT) on growth and development of H. armigera was investigated in this study. All protease genes encoded full ORFs and were expressed in all H. armigera larvae stages and tissues. In both injection and feeding bioassays, Ha-RNAi CHY's performance outperformed that of other protease genes. CHY enzyme activity in the midgut of larvae was significantly reduced after treatment with ds-HaCHY. Oral administration of ds-CHY also resulted in significant mortality of H. armigera larvae. However, because of the high RNase activity in the midgut lumen of lepidoptera, a large amount of dsRNA was needed to effectively kill instars of H. armigera. To reduce dsRNA degradation, bacterial expression and dsRNA formulation were used. After oral administration, it was toxic to H. armigera larvae. Before oral administration, bacterial cells were sonicated to increase dsRNA release. The RNA interference efficiency of sonicated bacteria was significantly increased, resulting in higher larval mortality when administered orally. All of these findings point to Ha-CHY as a new candidate for developing an effective dsRNA-based pesticide for H. armigera control.
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http://dx.doi.org/10.1002/arch.21840DOI Listing
November 2021

Comparison of gene expression in the red imported fire ant (Solenopsis invicta) under different temperature conditions.

Sci Rep 2021 08 13;11(1):16476. Epub 2021 Aug 13.

Plant Quarantine Technology Center, Animal and Plant Quarantine Agency, Gimcheon, 39660, Republic of Korea.

The red imported fire ant (RIFA), Solenopsis invicta Buren is native to South America and is known as a global problematic invasive species. This study focused on the molecular response of RIFA by comparing gene expression profiles after exposing ants to low (10 °C) and high (40 °C) temperature stress and comparing them to untreated controls (30 °C). A total of 99,085 unigenes (the clustered non-redundant transcripts that are filtered from the longest assembled contigs) were obtained, of which 19,154 were annotated with gene descriptions, gene ontology terms, and metabolic pathways. 86 gene ontology (GO) functional sub-groups and 23 EggNOG terms resulted. Differentially expressed genes (DEGs) with logFC ≥ 10 were screened and were compared at different temperatures. We found 203, 48, and 66 specific DEGs co-regulated at 10, 20, and 40 °C. Comparing transcriptome profiles for differential gene expression resulted in various DE genes, including cytochrome P450, NADH dehydrogenase subunit 1, cuticle protein and heat shock protein (HSP), which have previously been reported to be involved in cold and high temperature resistance. GO analysis revealed that antioxidant activity is up-regulated under high temperature stress. We verified the RNA-seq data by qPCR on 20 up- and down-regulated DEGs. These findings provide a basis for future understanding of the adaptation mechanisms of RIFA and the molecular mechanisms underlying the response to low and high temperatures.
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http://dx.doi.org/10.1038/s41598-021-95779-wDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8363622PMC
August 2021

Horizontally transmitted parasitoid killing factor shapes insect defense to parasitoids.

Science 2021 07;373(6554):535-541

Tokyo University of Agriculture and Technology, Fuchu, Tokyo 183-8509, Japan.

Interkingdom competition occurs between hymenopteran parasitoids and insect viruses sharing the same insect hosts. It has been assumed that parasitoid larvae die with the death of the infected host or as result of competition for host resources. Here we describe a gene family, (), that encodes proteins toxic to parasitoids of the Microgastrinae group and determines parasitism success. are found in several entomopathogenic DNA virus families and in some lepidopteran genomes. We provide evidence of equivalent and specific toxicity against endoparasites for PKFs found in entomopoxvirus, ascovirus, baculovirus, and Lepidoptera through a mechanism that elicits apoptosis in the cells of susceptible parasitoids. This highlights the evolutionary arms race between parasitoids, viruses, and their insect hosts.
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http://dx.doi.org/10.1126/science.abb6396DOI Listing
July 2021

Repat33 Acts as a Downstream Component of Eicosanoid Signaling Pathway Mediating Immune Responses of , a Lepidopteran Insect.

Insects 2021 May 14;12(5). Epub 2021 May 14.

Department of Plant Medicals, College of Life Sciences, Andong National University, Andong 36729, Korea.

Repat (=response to pathogen) is proposed for an immune-associated gene family from , a lepidopteran insect. In this gene family, 46 members (-) have been identified. They show marked variations in their inducible expression patterns in response to infections by different microbial pathogens. However, their physiological functions in specific immune responses and their interactions with other immune signaling pathways remain unclear. is a gene highly inducible by bacterial infections. The objective of this study was to analyze the physiological functions of in mediating cellular and humoral immune responses. Results showed that was expressed in all developmental stages and induced in immune-associated tissues such as hemocytes and the fat body. RNA interference (RNAi) of expression inhibited the hemocyte-spreading behavior which impaired nodule formation of hemocytes against bacterial infections. Such RNAi treatment also down-regulated expression levels of some antimicrobial genes. Interestingly, expression was controlled by eicosanoids. Inhibition of eicosanoid biosynthesis by RNAi against a phospholipase A (PLA) gene suppressed expression while an addition of arachidonic acid (a catalytic product of PLA) to RNAi treatment recovered such suppression of expression. These results suggest that Repat33 is a downstream component of eicosanoids in mediating immune responses of .
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http://dx.doi.org/10.3390/insects12050449DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8156158PMC
May 2021

EpOMEs act as immune suppressors in a lepidopteran insect, Spodoptera exigua.

Sci Rep 2020 11 19;10(1):20183. Epub 2020 Nov 19.

Department of Plant Medicals, Andong National University, Andong, 37629, South Korea.

Epoxyoctadecamonoenoic acids (EpOMEs) are epoxide derivatives of linoleic acid (9,12-octadecadienoic acid) and include 9,10-EpOME and 12,13-EpOME. They are synthesized by cytochrome P450 monooxygenases (CYPs) and degraded by soluble epoxide hydrolase (sEH). Although EpOMEs are well known to play crucial roles in mediating various physiological processes in mammals, their role is not well understood in insects. This study chemically identified their presence in insect tissues: 941.8 pg/g of 9,10-EpOME and 2,198.3 pg/g of 12,13-EpOME in fat body of a lepidopteran insect, Spodoptera exigua. Injection of 9,10-EpOME or 12,13-EpOME into larvae suppressed the cellular immune responses induced by bacterial challenge. EpOME treatment also suppressed the expression of antimicrobial peptide (AMP) genes. Among 139 S. exigua CYPs, an ortholog (SE51385) to human EpOME synthase was predicted and its expression was highly inducible upon bacterial challenge. RNA interference (RNAi) of SE51385 prevented down-regulation of immune responses at a late stage (> 24 h) following bacterial challenge. A soluble epoxide hydrolase (Se-sEH) of S. exigua was predicted and showed specific expression in all development stages and in different larval tissues. Furthermore, its expression levels were highly enhanced by bacterial challenge in different tissues. RNAi reduction of Se-sEH interfered with hemocyte-spreading behavior, nodule formation, and AMP expression. To support the immune association of EpOMEs, urea-based sEH inhibitors were screened to assess their inhibitory activities against cellular and humoral immune responses of S. exigua. 12-(3-adamantan-1-yl-ureido) dodecanoic acid (AUDA) was highly potent in suppressing the immune responses. The addition of AUDA to a pathogenic bacterium significantly increased bacterial pathogenicity by suppressing host immune defense. In sum, this study demonstrated that EpOMEs play a crucial role in facilitating anti-inflammatory responses in S. exigua.
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http://dx.doi.org/10.1038/s41598-020-77325-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7677322PMC
November 2020

Male-biased Adult Production of the Striped Fruit Fly, , by Feeding dsRNA Specific to .

Insects 2020 Mar 28;11(4). Epub 2020 Mar 28.

Department of Plant Medicals, Andong National University, Andong 36729, Korea.

Sterile insect release technique (SIT) is effective for eradicating quarantine insects including various tephritid fruit flies. When SIT is used for fruit flies, it is challenging to remove females from sterile males due to oviposition-associated piercing damage. This study developed a sex transition technique by feeding double-stranded RNA (dsRNA) specific to a sex-determining gene, () of the striped fruit fly, . is homologous to other fruit fly orthologs. It is highly expressed in female adults. RNA interference (RNAi) of by injecting or feeding its specific dsRNA to larvae significantly increased male ratio. Recombinant cells expressing dsRNA specific to were prepared and used to feed larvae to suppress gene expression levels. When these recombinant bacteria were fed to larvae during the entire feeding stage, the test population was significantly male-biased. Some females treated with such recombinant exhibited mosaic morphological characters such as the presence of male-specific abdominal setae in females. This study proposes a novel technique by feeding dsRNA specific to to reduce female production during mass-rearing of tephritid males for SIT.
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http://dx.doi.org/10.3390/insects11040211DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7240746PMC
March 2020

Biosynthesis and immunity of epoxyeicosatrienoic acids in a lepidopteran insect, Spodoptera exigua.

Dev Comp Immunol 2020 06 14;107:103643. Epub 2020 Feb 14.

Department of Plant Medicals, Andong National University, Andong, 36729, South Korea. Electronic address:

Eicosanoids mediate both cellular and humoral immune responses in insects. Epoxyeicosatrienoic acids (EETs) are a group of eicosanoids containing epoxide formed by epoxygenase (EPX) activity of cytochrome P450 (CYP). Although EETs have been considered to mediate immune responses in some insects, their synthetic machinery was little understood in insects. This study monitored EETs in a lepidopteran insect, Spodoptera exigua, immunized with bacteria and found all four EETs (5,6-EET, 8,9-EET, 11,12-EET, and 14,15-EET) from larval fat body at 247-1,736 pg/g levels. Then to predict EPXs, 140 CYPs were collected from S. exigua transcriptomes and compared with human EPXs. Four CYPs (SeEPX1-SeEPX4) sharing homologies with human EPXs were chosen and assessed in subsequent expression and functional analyses. All four EPXs were expressed in all development stages. In larval stage, all four EPXs were expressed in immune-associated tissues such as fat body and hemocytes. Furthermore, their expression levels were highly enhanced by bacterial challenge in different tissues. RNA interference (RNAi) using gene-specific double stranded RNA injection suppressed their expression levels by more than 55%. RNAi treatments interfered with hemocyte-spreading behavior and nodule formation upon bacterial challenge except RNAi treatment against SeEPX2. All four EETs stimulated cellular immune response measured by nodule formation in S. exigua. The suppressed immune responses by the RNAi treatments against three SeEPXs were rescued by the addition of 8,9-EET. However, other three EETs gave their specific rescue effect depending on SeEPX types under RNAi. In humoral immune response, all four RNAi treatments suppressed expression of antimicrobial peptide genes. This study reports the presence of all four EETs in larval fat body of S. exigua and suggests that four SeEPXs are associated with immune responses mediated by EETs.
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http://dx.doi.org/10.1016/j.dci.2020.103643DOI Listing
June 2020

Functional interaction of bacterial virulence factors of Xenorhabdus nematophila with a calcium-independent cytosolic PLA of Spodoptera exigua.

J Invertebr Pathol 2020 01 16;169:107309. Epub 2019 Dec 16.

Department of Plant Medicals, College of Life Sciences, Andong National University, Andong 36729, Republic of Korea. Electronic address:

Phospholipase A (PLA) hydrolyzes the ester bond of phospholipids (PLs) at sn-2 and releases free fatty acids and lysophospholipids that are subsequently changed into various signal molecules to mediate various physiological processes. Numerous PLAs are known in various biological systems and can be divided into at least 16 groups. Although different PLAs recently have been annotated from several insect species, physiological roles are known for only a few genes. Two calcium-independent PLAs (Se-iPLAA and Se-iPLAB) are known in the beet armyworm, Spodoptera exigua (Lepidoptera: Noctuidae). We generated and purified a recombinant Se-iPLAB (rSe-iPLAB) using a bacterial expression system and analyzed the enzyme kinetics. rSe-iPLAB exhibited catalytic activities against both arachidonyl (AA)-PL and non-AA-PL substrates. It was highly susceptible to iPLA-specific inhibitor, but insensitive to inhibitors specific to secretory PLAs or calcium-dependent cytosolic PLAs. Increasing calcium concentrations prevented enzyme activity, and culture medium of an entomopathogenic bacterium, Xenorhabdus nematophila, or its organic extracts significantly inhibited enzyme activity. Binding assays of rSe-iPLAB with known secondary metabolites identified from X. nematophila indicated that benzylideneacetone was the most potent inhibitor with a high binding affinity at 0.2 μM against rSe-iPLAB. Furthermore, rSe-iPLAB catalyzed the release of fatty acids from PLs extracted from S. exigua fat body, suggesting its physiological role in maintaining PL integrity. All these catalytic activities indicate that Se-iPLAB has the typical biochemical properties of other iPLAs. Its high binding affinity to secondary metabolites of X. nematophila suggests that it is a molecular target of X. nematophila, an entomopathogen.
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http://dx.doi.org/10.1016/j.jip.2019.107309DOI Listing
January 2020

Toll/IMD signal pathways mediate cellular immune responses via induction of intracellular PLA expression.

Arch Insect Biochem Physiol 2019 Jul 6;101(3):e21559. Epub 2019 May 6.

Department of Plant Medicals, College of Life Sciences, Andong National University, Andong, Korea.

Phospholipase A (PLA ) hydrolyzes fatty acids from phospholipids at the sn-2 position. Two intracellular PLA s, iPLA A and iPLA B, have been found in Spodoptera exigua. Both are calcium-independent cellular PLA . Their orthologs have been found in other insects. These two iPLA s are different in ankyrin motif of N terminal region. The objective of this study was to determine whether Toll/immune deficiency (IMD) signal pathways could mediate cellular immune responses via induction of iPLA expression. Both iPLA s were expressed in all developmental stages of S. exigua, showing the highest expression in the adult stage. During larval stage, hemocyte is the main tissue showing expression of these iPLA s. Both iPLA s exhibited similar expression patterns after immune challenge with different microbial pathogens such as virus, bacteria, and fungi. Promoter component analysis of orthologs encoded in S. frugiperda indicated nuclear factor-κB- and Relish-responsible elements on their promoters, suggesting their expression in S. exigua under Toll/IMD immune signaling pathways. RNA interference (RNAi) of MyD88 or Pelle under Toll pathway suppressed inducible expression levels of both iPLA s in response to Gram-positive bacteria containing Lys-type peptidoglycan or fungal infection. In contrast, RNAi against Relish under IMD pathway suppressed both iPLA s in response to infection with Gram-negative bacteria. Under RNAi conditions, hemocytes significantly lost cellular immune response measured by nodule formation. However, addition of arachidonic acid (a catalytic product of PLA ) rescued such immunosuppression. These results suggest that Toll/IMD signal pathways can mediate cellular immune responses via eicosanoid signaling by inducing iPLA expression.
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http://dx.doi.org/10.1002/arch.21559DOI Listing
July 2019

A prophylactic role of a secretory PLA of Spodoptera exigua against entomopathogens.

Dev Comp Immunol 2019 06 16;95:108-117. Epub 2019 Feb 16.

Department of Plant Medicals, Andong National University, Andong, 36729, South Korea. Electronic address:

Phospholipase A (PLA) hydrolyses phospholipids at sn-2 position to release free fatty acids and lysophospholipids. Secretory type of PLA (sPLA) has been found in many different animals including insects. Insect sPLAs have been divided into venomous and nonvenomous PLAs. A non-venomous sPLA (Se-sPLA) has been identified in beet armyworm, Spodoptera exigua. Its high enzyme activity is detected in hemolymph of naïve larvae. However, the physiological role of high sPLA activity in hemolymph remains unclear. To determine the physiological role of sPLA in hemolymph, a recombinant Se-sPLA (rSe-sPLA) was expressed in a bacterial expression system and purified to test antimicrobial activity against various microbes. Purified rSe-sPLA exhibited typical enzyme kinetic properties, including becoming saturated at high substrate concentrations, exhibiting optimal activity at pH 7-9, and being inactivated at high temperatures. However, a reducing agent (dithiothreitol) or calcium chelator treatment inhibited the catalytic activity. A specific inhibitor to sPLA also inhibited the enzyme activity of rSe-sPLA while other type PLA inhibitors did not. Furthermore, eight bacterial metabolites of Xenorhabdus and Photorhabdus known to be inhibitory against insect PLA significantly inhibited the enzyme activity of rSe-sPLA. High concentrations of rSe-sPLA (above 0.5 mM) showed significant cytotoxicity to hemocytes of S. exigua. At concentrations without showing cytotoxicity, rSe-sPLA possessed significant antimicrobial activities against entomopathogenic bacteria (Serratia marscens and Entercoccus mondtii) and fungi (Beauveria bassiana and Metarhyzium rileyi). Hemolymph obtained from larvae treated with RNA interference specific to Se-sPLA significantly lost such antimicrobial activities. However, the addition of rSe-sPLA to the hemolymph significantly rescued such antimicrobial activities. These results indicate that Se-sPLA possesses antimicrobial activity, suggesting that it might act as a prophylactic agent against microbial pathogens in the hemolymph of S. exigua.
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http://dx.doi.org/10.1016/j.dci.2019.02.008DOI Listing
June 2019

Correction to: Ultrastructure and secretion of glandular trichomes in species of subtribe Cajaninae Benth (Leguminosae, Phaseoleae).

Protoplasma 2019 03;256(2):447

Departamento de Botânica, Instituto de Biociências, Programa de Pós-Graduação em Ciências Biológicas (Botânica), Instituto de Biociências, Universidade Estadual Paulista, Botucatu, São Paulo, 18618-000, Brazil.

In the original version of this article unfortunately some symbols did not appear in the plates caused by technical problems of the journal. We apologize for any inconvenience caused.
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http://dx.doi.org/10.1007/s00709-018-1313-2DOI Listing
March 2019

Ultrastructure and secretion of glandular trichomes in species of subtribe Cajaninae Benth (Leguminosae, Phaseoleae).

Protoplasma 2019 Mar 10;256(2):431-445. Epub 2018 Sep 10.

Departamento de Botânica, Instituto de Biociências, Programa de Pós-Graduação em Ciências Biológicas (Botânica), Instituto de Biociências, Universidade Estadual Paulista, Botucatu, São Paulo, 18618-000, Brazil.

The subtribe Cajaninae of papilionoid legumes has a pantropical distribution and comprises approximately 490 species. These species have diversified throughout dry environments where there are high temperatures and strong light. The subtribe stands out because all its representatives have vesicular glands. In addition, bulbous-based and capitate trichomes are important secretory structures present in all genera of the Cajaninae. We analyzed the ultrastructure and histochemistry of these glandular trichome types in leaflets of the three species of the subtribe. Using transmission electron microscopy and histochemical analyses, we link the glandular secretions to subcellular structures. We here report for the first time the type of exudate and ultrastructure of the glands of subtribe Cajaninae. Terpenoids and phenolics were confirmed by histochemistry tests, and we observed that the organelles responsible for biosynthesis of oils are the most representative in these glands. Each glandular trichome showed particular ultrastructural features compatible with the compounds produced. We suggest that these glandular trichomes, with their respective exudates, act in defense against herbivory and against possible damage by ultraviolet radiation.
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http://dx.doi.org/10.1007/s00709-018-1307-0DOI Listing
March 2019

A non-venomous sPLA of a lepidopteran insect: Its physiological functions in development and immunity.

Dev Comp Immunol 2018 12 11;89:83-92. Epub 2018 Aug 11.

Department of Plant Medicals, Andong National University, Andong, 36729, South Korea. Electronic address:

Eicosanoids are oxygenated C20 polyunsaturated fatty acids that mediate various physiological processes in insects. Eicosanoid biosynthesis begins with a C20 precursor, arachidonic acid (5,8,11,14-eicosatetraenoic acid: AA). AA is usually released from phospholipids at sn-2 position by catalytic activity of phospholipase A (PLA). Although various PLAs classified into 16 gene families (= Groups) are known in various biological systems, few PLAs are known in insects. Only two PLAs involved in intracellular calcium independent PLA (iPLA) group have been identified in lepidopteran insects with well known eicosanoid physiology. This study reports the first secretory PLA (sPLA) in lepidopteran insects. A partial open reading frame (ORF) of PLA was obtained by interrogating Spodoptera exigua transcriptome. Subsequent 3'-RACE resulted in a full ORF (Se-sPLA2A) encoding 194 amino acid sequence containing signal peptide, calcium-binding domain, and catalytic site. Phylogenetic analysis indicated that Se-sPLAA was clustered with other Group III sPLAs. Se-sPLAA was expressed in most larval instars except late last instar. Its expression was inducible by immune challenge and juvenile hormone analog injection. RNA interference of Se-sPLAA significantly suppressed cellular immunity and impaired larval development. These results suggest that non-venomous sPLA plays a crucial role in immune and developmental processes in S. exigua, a lepidopteran insect.
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http://dx.doi.org/10.1016/j.dci.2018.08.008DOI Listing
December 2018

De novo transcriptome assembly of Pueraria montana var. lobata and Neustanthus phaseoloides for the development of eSSR and SNP markers: narrowing the US origin(s) of the invasive kudzu.

BMC Genomics 2018 Jun 5;19(1):439. Epub 2018 Jun 5.

Computational Biology Institute, Milken Institute School of Public Health, George Washington University, Washington, DC, USA.

Background: Kudzu, Pueraria montana var. lobata, is a woody vine native to Southeast Asia that has been introduced globally for cattle forage and erosion control. The vine is highly invasive in its introduced areas, including the southeastern US. Modern molecular marker resources are limited for the species, despite its importance. Transcriptomes for P. montana var. lobata and a second phaseoloid legume taxon previously ascribed to genus Pueraria, Neustanthus phaseoloides, were generated and mined for microsatellites and single nucleotide polymorphisms.

Results: Roche 454 sequencing of P. montana var. lobata and N. phaseoloides transcriptomes produced read numbers ranging from ~ 280,000 to ~ 420,000. Trinity assemblies produced an average of 17,491 contigs with mean lengths ranging from 639 bp to 994 bp. Transcriptome completeness, according to BUSCO, ranged between 64 and 77%. After vetting for primer design, there were 1646 expressed simple sequence repeats (eSSRs) identified in P. montana var. lobata and 1459 in N. phaseoloides. From these eSSRs, 17 identical primer pairs, representing inter-generic phaseoloid eSSRs, were created. Additionally, 13 primer pairs specific to P. montana var. lobata were also created. From these 30 primer pairs, a final set of seven primer pairs were used on 68 individuals of P. montana var. lobata for characterization across the US, China, and Japan. The populations exhibited from 20 to 43 alleles across the seven loci. We also conducted pairwise tests for high-confidence SNP discovery from the kudzu transcriptomes we sequenced and two previously sequenced P. montana var. lobata transcriptomes. Pairwise comparisons between P. montana var. lobata ranged from 358 to 24,475 SNPs, while comparisons between P. montana var. lobata and N. phaseoloides ranged from 5185 to 30,143 SNPs.

Conclusions: The discovered molecular markers for kudzu provide a starting point for comparative genetic studies within phaseoloid legumes. This study both adds to the current genetic resources and presents the first available genomic resources for the invasive kudzu vine. Additionally, this study is the first to provide molecular evidence to support the hypothesis of Japan as a source of US kudzu and begins to narrow the origin of US kudzu to the central Japanese island of Honshu.
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http://dx.doi.org/10.1186/s12864-018-4798-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5989403PMC
June 2018

Targeting legume loci: A comparison of three methods for target enrichment bait design in Leguminosae phylogenomics.

Appl Plant Sci 2018 Mar 2;6(3):e1036. Epub 2018 Apr 2.

Department of Botany National Museum of Natural History Smithsonian Institution P.O. Box 37012, MRC 166 Washington DC 20560 USA.

Premise Of The Study: The development of pipelines for locus discovery has spurred the use of target enrichment for plant phylogenomics. However, few studies have compared pipelines from locus discovery and bait design, through validation, to tree inference. We compared three methods within Leguminosae (Fabaceae) and present a workflow for future efforts.

Methods: Using 30 transcriptomes, we compared Hyb-Seq, MarkerMiner, and the Yang and Smith (Y&S) pipelines for locus discovery, validated 7501 baits targeting 507 loci across 25 genera via Illumina sequencing, and inferred gene and species trees via concatenation- and coalescent-based methods.

Results: Hyb-Seq discovered loci with the longest mean length. MarkerMiner discovered the most conserved loci with the least flagged as paralogous. Y&S offered the most parsimony-informative sites and putative orthologs. Target recovery averaged 93% across taxa. We optimized our targeted locus set based on a workflow designed to minimize paralog/ortholog conflation and thus present 423 loci for legume phylogenomics.

Conclusions: Methods differed across criteria important for phylogenetic marker development. We recommend Hyb-Seq as a method that may be useful for most phylogenomic projects. Our targeted locus set is a resource for future, community-driven efforts to reconstruct the legume tree of life.
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http://dx.doi.org/10.1002/aps3.1036DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5895186PMC
March 2018

Differential Inhibition of Helicoverpa armigera (Lep.: Noctuidae) Gut Digestive Trypsin by Extracted and Purified Inhibitor of Datura metel (Solanales: Solanaceae).

J Econ Entomol 2018 02;111(1):178-186

Department of Plant Protection, Faculty of Agriculture, Bu-Ali Sina University, Hamedan, I. R. Iran.

The cotton bollworm, Helicoverpa armigera Hubner (Lep: Noctuidae), is an economically important pest of numerous major food crops worldwide. Protease inhibitors from plants, expressed constitutively in transgenic crops, have potential for pest management as an alternative to chemical pesticides. In this study, a protease inhibitor was isolated, purified, and characterized from Datura metel L. seeds. The purity of the isolated inhibitor was confirmed by reverse-phase high-performance liquid chromatography, and activity staining showed one major peak and one clear activity band for the protein. Electrophoretic studies following gel filtration and ion-exchange chromatography revealed two and one bands for purified proteins, respectively. Partial biochemical characterizations of the purified inhibitor were determined. Maximum inhibitory activity was observed at 40-45°C (optimal temperature) when tested against gut extracts of fourth to sixth instar H. armigera larvae. Thermo-stability of the trypsin inhibitor against sixth instar larval midgut trypsin was observed up to 50°C when incubated for 30 min and 2 h. Among metal ions tested, Fe2+, Cu2+, and Mn2+ were found to decrease the trypsin inhibitory activity, whereas Hg2+, Mg2+, K+, Zn2+, Na+, Ca2+, and Cd2+ were found to significantly increase the inhibitory effect. This trypsin inhibitor showed competitive inhibition where the apparent value of Michaelis-Menten Km increased, but the value of Vmax remained unchanged.
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http://dx.doi.org/10.1093/jee/tox209DOI Listing
February 2018

Optimization of recombinant bacteria expressing dsRNA to enhance insecticidal activity against a lepidopteran insect, Spodoptera exigua.

PLoS One 2017 11;12(8):e0183054. Epub 2017 Aug 11.

Department of Plant Medicals, Andong National University, Andong, Korea.

Double-stranded RNA (dsRNA) has been applied to control insect pests due to its induction of RNA interference (RNAi) of a specific target gene expression. However, developing dsRNA-based insecticidal agent has been a great challenge especially against lepidopteran insect pests due to variations in RNAi efficiency. The objective of this study was to screen genes of chymotrypsins (SeCHYs) essential for the survival of the beet armyworm, Spodoptera exigua, to construct insecticidal dsRNA. In addition, an optimal oral delivery method was developed using recombinant bacteria. At least 7 SeCHY genes were predicted from S. exigua transcriptomes. Subsequent analyses indicated that SeCHY2 was widely expressed in different developmental stages and larval tissues by RT-PCR and its expression knockdown by RNAi caused high mortality along with immunosuppression. However, a large amount of dsRNA was required to efficiently kill late instars of S. exigua because of high RNase activity in their midgut lumen. To minimize dsRNA degradation, bacterial expression and formulation of dsRNA were performed in HT115 Escherichia coli using L4440 expression vector. dsRNA (300 bp) specific to SeCHY2 overexpressed in E. coli was toxic to S. exigua larvae after oral administration. To enhance dsRNA release from E. coli, bacterial cells were sonicated before oral administration. RNAi efficiency of sonicated bacteria was significantly increased, causing higher larval mortality at oral administration. Moreover, targeting young larvae possessing weak RNase activity in the midgut lumen significantly enhanced RNAi efficiency and subsequent insecticidal activity against S. exigua.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0183054PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5553977PMC
October 2017

Parsing polyphyletic Pueraria: Delimiting distinct evolutionary lineages through phylogeny.

Mol Phylogenet Evol 2016 11 2;104:44-59. Epub 2016 Aug 2.

East Carolina University, Department of Biology, Howell Science Complex, Greenville, NC 27858, USA(1).

Several taxonomic and phylogenetic studies have hypothesized polyphyly within Pueraria DC., a genus comprising 19 species (24 with varieties) including the highly invasive Pueraria montana var. lobata (Kudzu) introduced to the U.S.A. about 150years ago. Previous efforts to investigate monophyly of the genus have been hampered by limited taxon sampling or a lack of comprehensive evolutionary context that would enable definitive taxonomic associations. This work presents a comprehensive phylogenetic investigation of Pueraria within the context of tribe Phaseoleae (Leguminosae). Polyphyly was found to be more extensive than previously thought, with five distinct lineages spread across the tribe and spanning over 25mya of divergence strongly supported by two chloroplast and one nuclear marker, AS2, presented here as a phylogenetic marker for the first time. Our phylogenies support taxonomic revisions to rectify polyphyly within Pueraria, including the resurrection of Neustanthus, moving one species to Teyleria, and the creation of two new genera, Haymondia and Toxicopueraria (taxonomic revisions published elsewhere).
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http://dx.doi.org/10.1016/j.ympev.2016.08.001DOI Listing
November 2016

Transcriptome sequencing and marker development in winged bean (Psophocarpus tetragonolobus; Leguminosae).

Sci Rep 2016 06 30;6:29070. Epub 2016 Jun 30.

US National Herbarium (US), Department of Botany, Smithsonian Institution-NMNH, 10th and Constitution Ave, Washington DC, 20013, USA.

Winged bean, Psophocarpus tetragonolobus (L.) DC., is similar to soybean in yield and nutritional value but more viable in tropical conditions. Here, we strengthen genetic resources for this orphan crop by producing a de novo transcriptome assembly and annotation of two Sri Lankan accessions (denoted herein as CPP34 [PI 491423] and CPP37 [PI 639033]), developing simple sequence repeat (SSR) markers, and identifying single nucleotide polymorphisms (SNPs) between geographically separated genotypes. A combined assembly based on 804,757 reads from two accessions produced 16,115 contigs with an N50 of 889 bp, over 90% of which has significant sequence similarity to other legumes. Combining contigs with singletons produced 97,241 transcripts. We identified 12,956 SSRs, including 2,594 repeats for which primers were designed and 5,190 high-confidence SNPs between Sri Lankan and Nigerian genotypes. The transcriptomic data sets generated here provide new resources for gene discovery and marker development in this orphan crop, and will be vital for future plant breeding efforts. We also analyzed the soybean trypsin inhibitor (STI) gene family, important plant defense genes, in the context of related legumes and found evidence for radiation of the Kunitz trypsin inhibitor (KTI) gene family within winged bean.
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http://dx.doi.org/10.1038/srep29070DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4928180PMC
June 2016

A dated phylogeny of the papilionoid legume genus Canavalia reveals recent diversification by a pantropical liana lineage.

Mol Phylogenet Evol 2016 May 6;98:133-46. Epub 2016 Feb 6.

Universidade Estadual de Feira de Santana, Departamento de Ciências Biológicas, Av. Transnordestina, s/n, Novo Horizonte, Feira de Santana 44036-900, Bahia, Brazil. Electronic address:

Canavalia is a pantropical legume genus of lianas comprising approximately 60 species distributed in a wide range of habitats. In the last taxonomic revision, the genus was divided into four subgenera: Canavalia (Pantropical), Catodonia (Neotropical, excepting one species also found in the Old World), Maunaloa (Hawaiian), and Wenderothia (Neotropical). In this study, we reconstructed the phylogeny of Canavalia using a broad taxon sampling and analyses of nuclear (ETS and ITS) and plastid markers (trnK/matK). We evaluated the infrageneric classification of the genus and investigated its biogeographical history using molecular dating analyses and ancestral area reconstructions. The phylogenetic analyses resolved subgenus Wenderothia as monophyletic. Subgenus Catodonia needs to be recircumscribed and the relationships between subgenera Canavalia and Maunaloa remain unclear. Canavalia arose during the Miocene with a mean stem age estimate of 13.8Ma and mean crown age estimate of 8.7Ma, and most extant species evolved during the Pleistocene. Several climatic and geological events are chronologically coincident with the divergence of the major clades of Canavalia (glacial/interglacial periods, Andes uplift and the formation of Pebas and post-Pebas systems, closure of the Isthmus of Panama, and change in the direction of ocean currents). Ancestral area reconstructions for the early divergence of the genus are equivocal, although, some evidence suggests Canavalia originated in the wet forests of South America and achieved its current pantropical distribution through recent transoceanic dispersal. The evolution of Canavalia is better explained by a series of several processes than by discrete historical events.
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http://dx.doi.org/10.1016/j.ympev.2016.02.001DOI Listing
May 2016

A multilocus phylogenetic analysis reveals the monophyly of a recircumscribed papilionoid legume tribe Diocleae with well-supported generic relationships.

Mol Phylogenet Evol 2015 Sep 28;90:1-19. Epub 2015 Apr 28.

Smithsonian Institution, 10th and Constitution Ave, Washington, DC 20013-7012, USA.

Deciphering the phylogenetic relationships within the species-rich Millettioid clade has persisted as one of the major challenges in the systematics and evolutionary history of papilionoid legumes (Leguminosae, Papilionoideae). Historically, the predominantly neotropical lianas of subtribe Diocleinae in the Millettioid legumes have been taxonomically tangled together with the largely heterogeneous tribe Phaseoleae. This work presents a comprehensive molecular phylogenetic analysis based on nuclear and chloroplast markers and includes all genera ever referred to Diocleae except for the monospecific Philippine Luzonia, resolving several key generic relationships within the Millettioid legumes. The first of two separate analyses includes 310 matK accessions and strongly supports the reestablishment of tribe Diocleae as a branch of the Millettioid clade. This work sheds greater light on the higher-level phylogeny of Diocleae and allows the recognition of three major lineages: the Canavalia, Dioclea, and Galactia clades. The second set of phylogenetic analyses utilized nuclear (ITS/5.8S and ETS) and plastid (matK and trnT-Y) DNA sequences to reveal (i) the monophyly of Canavalia and Cleobulia; (ii) the monophyly of Bionia with the exclusion of Bionia bella; (iii) the paraphyly of Dioclea with respect to Cleobulia, Cymbosema, and Macropsychanthus; (iv) the paraphyly of Cratylia with respect to the broadly polyphyletic Camptosema; and (v) the polyphyly of Galactia with species scattered widely across the tree.
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http://dx.doi.org/10.1016/j.ympev.2015.04.016DOI Listing
September 2015
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