Novel and environmental friendly approach; Impact of Neem (<i>Azadirachta indica</i>) gum nano formulation (NGNF) on <i>Helicoverpa armigera</i> (Hub.) and <i>Spodoptera litura</i> (Fab.)

Kamaraj, Chinnaperumal,Gandhi, Pachiyappan Rajiv,Elango, Gandhi,Karthi, Sengodan,Chung, Ill-Min,Rajakumar, Govindasamy Elsevier 2018 INTERNATIONAL JOURNAL OF BIOLOGICAL MACROMOLECULES Vol.107 No.1

<P><B>Abstract</B></P> <P>The future of this study was to prepare a natural pesticide which will not harm the environment and yet control pests. Neem gum nano formulation (NGNF), a novel biopesticide prepared from the Neem gum extract (<I>Azadirachta indica</I>) (NGE) was evaluated for its antifeedant, larvicidal and pupicidal activities against <I>Helicoverpa armigera</I> (Hub.) and <I>Spodoptera litura</I> (Fab.) at 100ppm. The NGNF showed significant (100%) antifeedant, larvicidal and pupicidal activities against <I>H. armigera</I> and <I>S. litura</I>. The LC<SUB>50</SUB> values of 10.20, 12.49 and LC<SUB>90</SUB> values of 32.68, 36.68ppm on <I>H. armigera</I> and <I>S. litura</I>, respectively at 100ppm. The NGNF treatments showed differences in the activities of detoxifying enzymes, carboxylesterases, glucosidases and glutathione S-transferases in the larval gut. Earthworm toxicity illustrated that 6.25ppm of chemical insecticides (cypermethrin) varied widely in their contact toxicities compared to 100ppm of NGNF and control in both contact filter paper and artificial soil test. The NGNF were characterized and confirmed by FTIR, XRD, SEM and EDX analysis. Ten compounds were identified from the Neem gum extract (NGE) by Gas Chromatography-Mass Spectrometry (GC–MS) analysis. The major compounds were fatty acids like Hexadecanoic acid, oleic acid, and ricinoleic acid. NGNF could be used as an agent to prepare novel bio-pesticides formulations.</P> <P><B>Highlights</B></P> <P> <UL> <LI> Neem gum mediated nano formulation (NGNF) as a bio-pesticide agent against <I>Helicoverpa armigera</I> and <I>Spodoptera litura</I>. </LI> <LI> NGNF exhibited potential antifeedant, larvicidal and pupicidal activities against <I>Helicoverpa armigera</I> and <I>Spodoptera litura</I>. </LI> <LI> Environmental toxicity was assessed by earthworm (<I>Eudrilus eugeniae</I>) there is no significant toxicity observed in NGNF treatments. </LI> </UL> </P>

Isolation and Characterization of the C-type Lysozyme Gene from the Common Cutworm Spodoptera litura

김종완,Sung Moon Yoe 한국통합생물학회 2009 Animal cells and systems Vol.13 No.3

We have isolated and characterized a new insect chicken type (c-type) lysozyme gene from the common cutworm, Spodoptera litura. The full-length cDNA of Spodoptera lysozyme is cloned by rapid amplification of cDNA ends PCR (RACE-PCR). The isolated cDNA consists of 1039 bp including the coding region for a 142-amino acid residue polypeptide, which included a signal peptide of 21-amino acid residue and a mature protein of 121-amino acid residue. The predicted molecular weight of mature lysozyme and its theoretical isoelectric point from amino acid composition is 13964.8 Da and 9.05, respectively. The deduced amino acid sequence of Spodoptera lysozyme gene shows the highest similarity (96.7%) to Spodoptera exigua lysozyme among other lepidopteran species. Amino acid sequence comparison with other the c-type lysozymes, Spodoptera lysozyme has the completely conserved Glu32 and Asp50 of the active site and eight Cys residues are completely conserved in the same position as that of other lepidopteran lysozymes. We have isolated and characterized a new insect chicken type (c-type) lysozyme gene from the common cutworm, Spodoptera litura. The full-length cDNA of Spodoptera lysozyme is cloned by rapid amplification of cDNA ends PCR (RACE-PCR). The isolated cDNA consists of 1039 bp including the coding region for a 142-amino acid residue polypeptide, which included a signal peptide of 21-amino acid residue and a mature protein of 121-amino acid residue. The predicted molecular weight of mature lysozyme and its theoretical isoelectric point from amino acid composition is 13964.8 Da and 9.05, respectively. The deduced amino acid sequence of Spodoptera lysozyme gene shows the highest similarity (96.7%) to Spodoptera exigua lysozyme among other lepidopteran species. Amino acid sequence comparison with other the c-type lysozymes, Spodoptera lysozyme has the completely conserved Glu32 and Asp50 of the active site and eight Cys residues are completely conserved in the same position as that of other lepidopteran lysozymes.

Comparative Analysis of the Difference in the Midgut Microbiota between the Laboratory Reared and the Field-caught Populations of Spodoptera litura

( Neeti Pandey ),( Raman Rajagopal ) 한국미생물생명공학회(구 한국산업미생물학회) 2019 한국미생물·생명공학회지 Vol.47 No.3

Midgut microbiota is known to play a fundamental role in the biology and physiology of the agricultural pest, Spodoptera litura. This study reports the difference in the larval midgut microbiota of field-caught and laboratory-reared populations of S. litura by performing 16S rDNA amplicon pyrosequencing. Field populations for the study were collected from castor crops, whereas laboratory-reared larvae were fed on a regular chickpea based diet. In total, 23 bacterial phylotypes were observed from both laboratory-reared and field-caught caterpillars. Fisher’s exact test with Storey’s FDR multiple test correction demonstrated that bacterial genus, Clostridium was significantly abundant (p < 0.05) in field-caught larvae of S. litura as compared to that in the laboratory-reared larvae. Similarly, bacterial genera, such as Bradyrhizobium, Burkholderia, and Fibrisoma were identified (p < 0.05) predominantly in the laboratory-reared population. The Bray-Curtis dissimilarity matrix depicted a value of 0.986, which exhibited the maximum deviation between the midgut microbiota of the laboratory-reared and field-caught populations. No significant yeast diversity was seen in the laboratory-reared caterpillars. However, two yeast strains, namely Candida rugosa and Cyberlindnera fabianii were identified by PCR amplification and molecular cloning of the internal transcribed space region in the field-caught caterpillars. These results emphasize the differential colonization of gut residents based on environmental factors and diet.

담배거세미나방(Spodoptera litura) 유충 혈림프내 Phenoloxidase의 특성

이상용,이상대,강창수 한국곤충학회 1997 Entomological Research Vol.27 No.2

담배거세미나방(Spodoptera litura)유충의 혈림프내에 존재하는 phenoloxidase는 전기영 동상에서 단일의 단백질형태로 존재하는 것으로 나타났다. 또한, 이 효소는 기질반응을 이용한 파장 430nm에서의 dopachrome 축적을 통해 L-dopa를 O-dopaquinone으로 전환시키는 diphenoloxidase activity를 가지고 있음이 확인되었다. 이 효소는 L-dopa를 기질로 할 때 pH8.0에서 최적의 활성을 나타내었고, 7$0^{\circ}C$까지의 열안정성을 보였다. L-dopa에 대한 Km값은 pH 7.4 조건하에서 약 1.5mM로 나타났고, 등전점은 약 8.2로 판명되었다. PTU, EGTA, EDTA등을 사용한 효소활성의 억제실험에서 PTU의 경우 약 12 $\mu$M에서 50%정도의 저해효과를 나타내었는데 이것으로 보아 phenoloxidase의 활성에는 구리이온이 중요 한 역할을 하는 것으로 보여진다. 또한, phenoloxidase는 일부 알코올류와 유기물질들의 첨가 그리고 여러환경 조건에 따라 활성의 변화를 보이는데, 이것으로 보아 phenoloxidase 효소계에는 체내에서 비활성형의 phenoloxidase 형태로 존재하다가 외부자극에 의하여 활성형으로 전환되는 효소활성계를 최소한 하나 이상 포함하는 것으로 생각된다. For studies of non-self recognition in insects, we investigated the phenoloxidase (PO) system from the larval haemolymph of Spodoptera litura. PO is present in haemolymph and detected as an intense band with high molecular weight by electrophoresis. The enzyme catalysed the oxidation of other di-phenols, besides dopa. The enzyme activity was stable up to 70$^{\circ}C$ and showed a pH optimum 8.0 in the reaction mixture containing L-dopa. The Km value fer PO was 1.5 mM at pH 7.4 and pI value was 8.2. PTU, the specific inhibitor for dopamin ${\beta}$-hydroxylase, strongly inhibited the PO activity: half maximal inhibition by PTU occurred at 12 ${\mu}$M, suggesting that Cu$\^$2+/ is important for the activity of PO. While PO can be activated by organic solvents, heat treatment or other environmental effect, suggesting in vivo conversion system that inactive PO might be changed to active PO by external stimulus.

Effects of Electron Beam on Life Stages, DNA Damage and DNA Recovery of Spodoptera litura

Seung-Hwan Yun,Hyun-Na Koo,Gil-Hah Kim 한국응용곤충학회 2012 한국응용곤충학회 학술대회논문집 Vol.2012 No.10

The effects of electron beam irradiation on life stage and reproduction of Spodoptera litura were examined. Eggs, larvae, pupae, and adults were irradiated at target doses of 30, 50, 100, 150, 200, and 250 Gy. When eggs were irradiated with 100 Gy, egg hatching was completely inhibited. When irradiated to the larvae, pupation was inhibited at 100 Gy and larval period was delayed. When irradiated to the pupae, emergence was inhibited at over 100 Gy. When irradiated to the adults, longevity and fecundity did not show any differences. However, egg hatching was significantly decreased at 100 Gy and above. Also, electron beam irradiation was not induced the rapid death of S. litura. Reciprocal crosses between irradiated and unirradiated moths demonstrated that males were more radiotolerant than females. Adult longevity was not affected in all stages. The levels of DNA damage in S. litura adults were evaluated using the alkaline comet assay. Our results indicate that electron beam irradiation increased levels of DNA damage. The recovery of DNA damage in S. litura adults increased as time passed. but DNA damage hasn’t recovered fully. These results indicate that electron beam irradiation induced abnormal development and reproduction by DNA damage in S. litura. and as time goes by, the DNA damage was decresed.

X-ray Irradiation Induced Abnormal Development and Reproduction by DNA Damage in Spodoptera litura

Min-Jun Kim,Seung-Hwan Yun,Seon-Woo Lee,Soon-Kyu Lee,Hyun Kyung Kim,Hyun-Na Koo,Gil-Hah Kim 한국응용곤충학회 2014 한국응용곤충학회 학술대회논문집 Vol.2014 No.04

The effects of X-ray irradiation on development and reproduction of Spodoptera litura were examined. Eggs, larvae, pupae, and adults were irradiated at target doses of 10 - 250 Gy. When eggs were irradiated with 30 Gy, egg hatching was completely inhibited. When irradiated to the larvae, pupation was inhibited at 100 Gy and larval period was delayed. When irradiated to the pupae, emergence was inhibited at over 100 Gy. When irradiated to the adults, longevity and fecundity did not show any differences. However, egg hatching was significantly decreased at 70 Gy and above. Also, X-ray irradiation was not induced the rapid death of S. litura. Reciprocal crosses between irradiated and unirradiated moths demonstrated that males were more radiotolerant than females. The levels of DNA damage in S. litura adults were evaluated using the alkaline comet assay. Our results indicate that X-ray irradiation increased levels of DNA damage. The recovery of DNA damage in S. litura adults increased as time passed. But DNA damage hasn't recovered fully. These results indicate that X-ray irradiation induced abnormal development and reproduction by DNA damage in S. litura.

Isolation and Characterization of the C-type Lysozyme Gene from the Common Cutworm Spodoptera litura

Kim, Jong-Wan,Yoe, Sung-Moon The Korean Society for Integrative Biology 2009 Animal cells and systems Vol.13 No.3

We have isolated and characterized a new insect chicken type (c-type) lysozyme gene from the common cutworm, Spodoptera litura. The full-length cDNA of Spodoptera lysozyme is cloned by rapid amplification of cDNA ends PCR (RACE-PCR). The isolated cDNA consists of 1039 bp including the coding region for a 142-amino acid residue polypeptide, which included a signal peptide of 21-amino acid residue and a mature protein of 121-amino acid residue. The predicted molecular weight of mature lysozyme and its theoretical isoelectric point from amino acid composition is 13964.8 Da and 9.05, respectively. The deduced amino acid sequence of Spodoptera lysozyme gene shows the highest similarity (96.7%) to Spodoptera exigua lysozyme among other lepidopteran species. Amino acid sequence comparison with other the c-type lysozymes, Spodoptera lysozyme has the completely conserved $Glu^{32}$ and $Asp^{50}$ of the active site and eight Cys residues are completely conserved in the same position as that of other lepidopteran lysozymes.

Identification and Characterization of Baculoviruses isolated from Spodoptera litura

Yong Wang,Jae Young Choi,Jong Yul Roh,Qin Liu,Xueying Tao,Jong Bin Park,Soo Dong Woo,Byung Rae Jin,Yeon Ho Je 한국응용곤충학회 2010 한국응용곤충학회 학술대회논문집 Vol.2010 No.10

We isolated two baculoviruses, Spodoptera litura granulovirus (SlGV) and S. litura nucleopolyhedrovirus (SlNPV) in the dead larvae of S. litura. The granule of SlGV were ovoidal shape with an approximate measure of 240-340 nm×140-180 nm, and each granule contained one single rod-shape virion with a mean size of 180-200 nm×20-40 nm. Whereas, the polyhedra of SlNPV were irregular in shape with a approximate diameter of 1.0-1.5 ㎛, and numerous virions comprised of the multinucleocapsid were contained in each polyhedra. The major component of occlusion bodies produced by SlGV and SlNPV were about 29 and 30 kDa, respectively. When the phylogenic relationship between these viruses were analyzed using the nucleotide sequences of granulin gene from SlGV and polyhedrin gene from SlNPV, they were not closely related to each other. We also found that the two viruses showed similar insecticidal activity against 2nd instar larvae of Spodotera litura in terms of dose-response, but SlGV showed much longer LT50 than that of SlNPV. The two baculoviruses might be cooperatively be applied as biological control agent for the control of S. litura

Genetics of resistance to Cry1C toxin from Bacillus thuringiensis in Spodoptera litura (Fab.)

Atul S.Thakare,Umesh P. BARKHADE,Vrushali N. PAWADE,Shivcharan M. THAKARE 한국곤충학회 2010 Entomological Research Vol.40 No.3

The present study was undertaken to determine the genetics of Cry1C resistance in Spodoptera litura. Selection of S. litura (Fab.) with Cry1C was done for eight generations to develop resistance. Reciprocal crosses between resistant and susceptible populations were made to understand the population genetics of Cry1C resistance in S. litura. Generation wise selection with Cry1C was evaluated for resistance development in S. litura. The LC50 of Cry1C was 0.14 µg/cm2 for the first selected generation and it increased to 23.98 µg/cm2 after eight selected generations, which is a 285.47-fold increase in resistance compared with the susceptible strain. The estimated realized heritability (h2) after eight generations of selection with Cry1C insecticidal protein was 0.44. The number of generations required for the tenfold increase in LC50 (1/R) was estimated to be 3.33. Response to Cry1C selection in S. litura was 0.30, the estimated selection differential was 0.69 and the pheonotypic standard deviation (dP) was 0.24. Reciprocal crosses between Cry1C resistant and susceptible strain of S. litura showed autosomal resistance.

Knockdown of dorsal/relish increases larvae susceptibility to microbes in Spodoptera litura

Yangyang Cai,Zimo Guo,Chunping Ma,Jianya Su 한국응용곤충학회 2019 Journal of Asia-Pacific Entomology Vol.22 No.1

The tobacco cutworm, Spodoptera litura has developed high levels of resistance to frequently-used insecticides, and failures of chemical control has frequently been reported in China. Moreover, the control efficiency of biological control agents was not satisfactory by farmers, and thus the development of novel or combined techniques for such pest control becomes an imperative requirement. In this study the potential application of innate immune-related genes in insect control was investigated. Sldorsal and Slrelish were identified from S. litura. Sldorsals were mainly expressed in epidermal and fatbody of larvae, and Slrelish highly expressed in fatbody and haemolymph of larvae. Their expression characteristic suggests potential roles in defense against fungi or bacteria. The expression of Sldorsal was up-regulated by the challenge of entomogenous fungi Isaria javanica and Beauveria bassiana, however, the expression of Slrelish could only be induced by Escherichia coli challenge. Injection of Sldorsal or Slrelish dsRNA down-regulated the expression level of the corresponding gene and broke specific signaling transduction pathway of innate immunity in S. litura. Knockdown of Sldorsal by RNAi significantly increased the mortality of S. litura larvae caused by I. javanica and B. bassiana, and Slrelish RNAi highly increased the mortality caused by E. coli. Sldorsal and Slrelish are essential genes for innate immunity, their knockdown by RNAi increases S. litura susceptibility to insect pathogens, therefore enhances the mortalities caused by I. javanica, B. bassiana and E. coli respectively.