Systematic induced resistance in Solanum lycopersicum (L.) against vascular wilt pathogen (Fusarium oxysporum f. sp. lycopersici) by Citrullus colocynthis and Trichoderma viride.

The antifungal effects of Citrullus colocynthis extract (Hexane, chloroform, methanol, and water) were tested in vitro on Fusarium oxysporum f. sp. lycopersici (Sacc.) W. C. Snyder & H. N. Hans (FOL), the causal agent of Fusarium wilt. Of these, methanol and water extract at 10% showed the highest inhibition of mycelial growth of FOL by 12.32 and 23.61 mm respectively. The antifungal compounds were identified through Fourier transform infrared (FT-IR) spectroscopy and gas chromatography-mass spectroscopy (GC-MS). The methanol extract was compatible with the biocontrol agent Trichoderma viride. The antagonistic fungi were mass-cultured under laboratory conditions using sorghum seeds. Both T. viride and C. colocynthis methanol extract was also tested alone and together against FOL under both in vitro and in vivo conditions. The combination of T. viride and C. colocynthis showed the highest percentage of antifungal activity (82.92%) against FOL under in vitro conditions. This study revealed that induced systemic resistance (ISR) in enhancing the disease resistance in tomato plants against Fusarium wilt disease. The combined treatment of T. viride and C. colocynthis significantly reduced the disease incidence and index by 21.92 and 27.02% in greenhouse conditions, respectively. Further, the induction of defense enzymes, such as peroxidase (PO), polyphenol oxidase (PPO), ß-1,3-glucanase, and chitinase were studied. The accumulation of defense enzyme was greater in plants treated with a combination of T. viride and C. colocynthis compared to the control. Reduction of wilt disease in tomato plants due to the involvement of defense-related enzymes is presumed through this experiment.

Toxic effect of essential oil and its compounds isolated from Sphaeranthus amaranthoides Burm. f. against dengue mosquito vector Aedes aegypti Linn.

Aedes aegypti is a major mosquito vector that can transfer many deadly diseases such as dengue, chikungunya, Zika, and yellow fever viruses. Due to the developing resistance among the vector populations by the application of chemical insecticides, alternative eco-friendly vector management strategies are being focused. In this aspect, the present study was carried out to evaluate the mosquitocidal potentials of essential oil of Sphaeranthus amaranthoides (EO-Sa). EO-Sa was found to be effective against Ae. aegypti mosquito vector by exhibiting significant larvicidal, adulticidal and repellent activities. GCMS analysis of EO-Sa revealed the presence of Carvone as the major component (peak area of 89.7%). The larvicidal bioassays performed revealed that the second instar larvae were relatively more susceptible (94.32% mortality) to EO-Sa treatments (75 ppm), LC50, 20.38 ppm.The sub lethal treatment concentration (20 ppm) significantly affected the oviposition, fecundity and morphology of Ae. aegypti. At sub lethal treatment concentration, EO-Sa down regulated α- and ß carboxylesterase and up regulated the GST and CYP450 level of third and fourth instar larvae. Thus the present results illustrates that EO-Sa can deliver a durable larvicidal, repellent and adulticidal activity against Ae. aegypti in an effective and eco-friendly manner.

Target and non-target botanical pesticides effect of Trichodesma indicum (Linn) R. Br. and their chemical derivatives against the dengue vector, Aedes aegypti L.

The effects of crude ethanol derived leaf extract Trichodesma indicum (Linn) (Ex-Ti) and their chief derivatives were accessed on the survival and development of the dengue mosquito Ae. aegypti also their non-toxic activity against mosquito predator. T. indicum is recognized to be the vital weed plant and a promising herb in the traditional ayurvedic medicine. In this study, the GC-MS chromatogram of Ex-Ti showed higher peak area percentage for cis-10-Heptadecenoic acid (21.83%) followed by cycloheptadecanone (14.32%). The Ex-Ti displayed predominant mortality in larvae with 96.45 and 93.31% at the prominent dosage (200 ppm) against III and IV instar. Correspondingly, sub-lethal dosage against the enzymatic profile of III and IV instar showed downregulation of α,ß-carboxylesterase and SOD protein profiles at the maximum concentration of 100 ppm. However, enzyme level of GST as well as CYP450 increased significantly dependent on sub-lethal concentration. Likewise, fecundity and hatchability of egg rate of dengue mosquito decreased to the sub-lethal concentration of Ex-Ti. Repellent assay illustrates that Ex-Ti concentration had greater protection time up to 210 min at 100 ppm. Also, activity of Ex-Ti on adult mosquito displayed 100% mortality at the maximum dosage of 600, 500 and 400 ppm within the period of 50, 60 and 70 min, respectively. Photomicrography screening showed that lethal dosage of Ex-Ti (100 ppm) produced severe morphological changes with dysregulation in their body parts as matched to the control. Effects of Ex-Ti on the Toxorhynchites splendens IV instar larvae showed less mortality (43.47%) even at the maximum dosage of 1500 ppm as matched to the chemical pesticide Temephos. Overall, the present research adds a toxicological valuation on the Ex-Ti and their active constituents as a larvicidal, repellent and adulticidal agents against the global burdening dengue mosquito.

Target and non-target response of Swietenia Mahagoni Jacq. chemical constituents against tobacco cutworm Spodoptera litura Fab. and earthworm, Eudrilus eugeniae Kinb.

Toxicological screening of Swietenia mahagoni Jacq. (Meliaceae, West Indies Mahogany) against the lepidopteran pest Spodoptera litura was examined. Phytochemical screening through GC-MS analysis revealed nine peaks with prominent peak area % in Bis (2-ethylhexyl) phthalate (31.5%) was observed. The larvae exposed to discriminating dosage of 100 ppm deliver significant mortality rate compare to other treatment concentrations. The lethal concentrations (LC50 and LC90) was observed at the dosage of 31.04 and 86.82 ppm respectively. Sub-lethal concentrations (30 ppm) showed higher larval and pupal durations. However, pupal weight and mean fecundity rate reduced significantly. Similarly, the adult longevity reduced significantly in dose dependent manner. Midgut histology studies showed that the methanolic extracts significantly disturbs the gut epithelial layer, lumen and brush border membrane compare to the control. The soil assay on a non-target beneficial organism, the soil indicator earthworm Eudrilus eugeniae, with extracts from S. mahagoni (200 mg/kg) showed no toxicity compared to Monocrotophos at the dosage of 10 ppm/kg. Current results suggest that this bio-rational plant product from S. mahagoni displays a significant effect to reduce lepidopteran pests with low toxicity to other beneficial species.

Acute toxicity of chemical pesticides and plant-derived essential oil on the behavior and development of earthworms, Eudrilus eugeniae (Kinberg) and Eisenia fetida (Savigny).

Comparative toxicity of two chemical pesticides (temephos and monocrotophos) versus a plant-derived betel leaf oil Piper betle (L.) to earthworm Eudrilus eugeniae (Kinberg) and redworm Eisenia fetida Savigny, historically: Eisenia foetida (Savigny 1826), was evaluated. Mortality rate was more prominent in temephos at 100 µg concentration to both the earthworms in filter paper test (FPT) as well as 10 mg concentration in artificial soil test (AST). In contrast, P. betle does not display much mortality rate to both the earthworms even at 1000 mg of treatment concentrations. The lethal concentration (LC50) value was observed at 3.89 and 5.26 mg/kg for temephos and monocrotophos against E. eugeniae and 3.81 and 5.25 mg/kg to E. fetida, respectively. Whereas, LC50 value of betel leaf oil was only observed at 3149 and 4081 mg/kg to E. eugeniae and E. fetida, respectively. Correspondingly, the avoidance or attraction assay also displayed that earthworms were more sensitive to the soil containing chemical pesticides. Whereas, the avoidance percentage was decreased in the P. betle oil. Similarly, sublethal concentration of chemical pesticides (5 and 6.5 mg) significantly reduced the earthworm weight and growth rate. However, P. betle oil did not change the developmental rate in the duration of the assay (2, 7 and 14 days) even at 4000 mg treatment concentration. The enzyme ratio of CAT and SOD was also affected significantly after exposure to the chemical pesticides (6.5 mg/kg). Hence, our study implied the risk assessment associated with the chemical pesticides and also recommends plant-derived harmless P. betle oil against beneficial species as an alternative pest control agent.

Toxicological effects of Sphaeranthus indicus Linn. (Asteraceae) leaf essential oil against human disease vectors, Culex quinquefasciatus Say and Aedes aegypti Linn., and impacts on a beneficial mosquito predator.

Use of environmentally friendly, decomposable natural products for effective vector control has gained considerable momentum in modern society. In this study, essential oil of Sphaeranthus indicus (Si-EO) was extracted and further phytochemical screening revealed fourteen compounds with prominent peak area percentage of 24.9 and 22.54% in 3,5-di-tert-butyl-4-hydroxybenzaldehyde and benzene,2-(1,1-dimethylethyl)-1,4-dimethoxy, respectively. The Si-EO was further evaluated for their larvicidal response against Culex quinquefasciatus and Aedes aegypti at different dosages (62.5, 125, 250 and 500 ppm). The Si-EO displayed prominent larvicidal activity at higher concentration (500 ppm) against both species of mosquitoes. The LC50 and LC90 values of oils were observed at 130 and 350 ppm against C. quinquefasciatus larvae and at 140 and 350 ppm against A. aegypti larvae, respectively. Repellent bioassay established higher protection rate at 200 ppm up to 120 min against both the mosquitoes. However, adulticidal response displayed higher mortality rate only at 700 and 800 ppm against C. quinquefasciatus and A. aegypti, respectively. Toxicological screening against mosquito predator Toxorhynchites splendens revealed that the Si-EO was harmless even at the concentration of 1500 ppm. Overall, these results suggest that the Si-EO plays a significant role as a new bio-rational product against ecological burden mosquito vectors which provides an eco-friendly alternative to synthetic pesticides.

Comparative analysis of mosquito (Diptera: Culicidae: Aedes aegypti Liston) responses to the insecticide Temephos and plant derived essential oil derived from Piper betle L.

Resistance to treatments with Temephos or plant derived oil, Pb-CVO, between a field collected Wild Strain (WS) and a susceptible Laboratory Strain (LS) of Ae. aegypti were measured. The Temephos (0.1mg/L) showed the greatest percentage of mosquito mortality compared to Pb-CVO (1.5mg/L) in LS Ae. aegypti. However, WS Ae. aegypti was not significantly affected by Temephos (0.1mg/L) treatment compare to the Pb-CVO (1.5mg/L). However, both strains (LS and WS) when treated with Pb-CVO (1.5mg/L) displayed steady larval mortality rate across all instars. The LC50 of Temephos was 0.027mg in LS, but increased in WS to 0.081mg/L. The LC50 of Pb-CVO treatment was observed at concentrations of 0.72 and 0.64mg/L for LS and WS strains respectively. The enzyme level of α- and ß-carboxylesterase was reduced significantly in both mosquito strains treated with Pb-CVO. Whereas, there was a prominent deviation in the enzyme ratio observed between LS and WS treated with Temephos. The GST and CYP450 levels were upregulated in the LS, but decreased in WS, after treatment with Temephos. However, treatment with Pb-CVO caused both enzyme levels to increase significantly in both the strains. Visual observations of the midgut revealed cytotoxicity from sub-lethal concentrations of Temephos (0.04mg/L) and Pb-CVO (1.0mg/L) in both strains of Ae. aegypti compared to the control. The damage caused by Temephos was slightly less in WS compared to LS mosquito strains.

Developmental response of Spodoptera litura Fab. to treatments of crude volatile oil from Piper betle L. and evaluation of toxicity to earthworm, Eudrilus eugeniae Kinb.

Evaluations of biological effects of (Pb-CVO) the crude volatile oil of Piper betle leaves on the tobacco cutworm Spodoptera litura were conducted. Pb-CVO was subjected to GC-MS analysis and twenty vital compounds were isolated from the betel leaf oil. Pb-CVO was tested at four different concentrations (0.25, 0.5, 1.0 and 1.5%) against S. litura. The treated insects exhibited dose depended mortality. The mortality rate was significantly higher at the 1.0 and 1.5% Pb-CVO. The LC50 (Lethal concentration) were observed at 0.48% Pb-CVO. Larval and pupal durations increased in all treatment concentrations (0.25, 0.3, 0.4 and 0.5%) whereas, pupal weight decreased compared to control. Adult longevity of S. litura was reduced in all treatments but predominantly in the 0.4 and 0.5% Pb-CVO. Correspondingly, mean fecundity rate was reduced at all concentrations compared to control. Histological studies of larvae mid-gut profiles of S. litura were severely damaged in 1.0 and 1.5% and showed abnormalities in mid-gut cells with 0.25 and 0.5% Pb-CVO treatments. Earthworm toxicity illustrated that 0.1% of chemical insecticides (monocrotophos and cypermethrin) varied widely in their contact toxicities compared to 0.5 and 1.0% Pb-CVO and control in both contact filter paper and artificial soil test. These findings suggest that twenty essential compounds of betel leaf oil were significant inhibitors of the development and caused behavioral changes of S. litura. Treatment with betel leaf oil at these concentrations had no adverse effect on earthworm populations.

Toxicity of Alangium salvifolium Wang chemical constituents against the tobacco cutworm Spodoptera litura Fab.

Widespread use of synthetic pesticides has resulted in the development of insecticide-resistant populations of pests and harmful effects on human health and the environment. There is a need to identify alternative pest management strategies to reduce our reliance on conventional chemical pesticides. In recent years the use of botanical pesticides for protecting crops from insect pests has assumed greater importance. Methanol extract of Alangium salvifolium (L.f.) Wang has potential insecticidal activity against Spodoptera litura Fab. The active fractions were identified through chromatographic techniques as F-IV (Rf value=0.45) and F-VI (Rf value=0.63) and were subjected to GC-MS (GCMATE II). Fifty, 100 and 200ppm of active fractions were applied to fourth instar larvae and the mortality increased with higher concentrations. Relative consumption rate, relative growth rate, efficiency of conversion of ingested food and efficiency of conversion of digested food values all decreased in treated larvae, but approximate digestibility rate increased after treatment. The hydrolytic enzymes, such as acid phosphatase, alkaline phosphatase and the glycolytic enzyme lactate dehydrogenase were inhibited in treated larvae compared with controls. The histopathology study revealed that the epithelial columnar cells were enlarged, completely atrophied; intercellular spaces were swollen, and also noted a cytoplasmic ooze of cell material that mixed with food column. The present study clearly showed the active fractions from A. salvifolium as potential botanicals to control the larvae of S. litura. This is the first report for nutritional indices, enzymatic activities and histological effects of A. salvifolium chemical constituents against S. litura. Thus probably, this will be used as an alternative for synthetic pesticides against the polyphagous pest like S. litura.

Bacillus subtilis chitinase identified by matrix-assisted laser desorption/ionization time-of flight/time of flight mass spectrometry has insecticidal activity against Spodoptera litura Fab.

An extracellular chitinase was identified and purified (CS1 and CS2) from Bacillus subtilis. The 16S rRNA sequencing was submitted in GenBank (accession numbers KC336487 and KC412256). The purified crude enzymes were identified through matrix-assisted laser desorption/ionization time-of-flight/time-of-flight mass spectrometry (MALDI-TOF MS) analysis. The peptide sequences were matched with chitinase sequences. The peak m/z with 1297. 592 and 3094.570 mascot search resulted sequence was blasted with NCBI protein sequences and confirmed that it is a chitinase enzyme. The effects of chitinase on gut enzymes lactate dehydrogenase, acid phosphatase, alkaline phosphatase and adenosine triphosphatase of the tobacco cutworm Spodoptera litura larvae were investigated. At all concentrations tested, chitinase decreased the activities of these gut enzymes relative to the control. When chitinase treated leaves were fed to larvae in bioassays, gut tissue and gut enzymes were affected. The histological study clearly shows the chitinase treated larval gut, peritrophic membrane and epithelial cells were affected significantly. Chitinase isolated from B. subtilis has effectively reduced the gut enzyme activity and growth of S. litura. The chitin based bioformulation may serve as an effective biocide against the polyphagous pest like S. litura.