Chemicals isolated from Justicia adhatoda Linn reduce fitness of the mosquito, Aedes aegypti L.

Extracts from Justicia adhatoda L. (Acanthaceae) strongly reduced the fitness of the mosquito, Aedes aegypti Linn. The methanolic extracts inhibited several enzymes responsible for protecting insects from oxidative and other damage, including glutathione-S-transferase, superoxide dismutase, cytochrome P450, and α- and ß-esterases. They increased repellency (maximum repellency at 100 ppm) in host-seeking adult females using the "arm-in cage assay." Histopathological examination showed the extracts led to serious midgut cell damage. Justicia adhatoda extracts led to reduced fecundity and oviposition of gravid females compared to controls. The extracts led to substantially reduced A. aegypti survival. We infer that the extracts have potential to reduce pathogen transmission by suppressing population growth of A. aegypti, and possibly other mosquito species.

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.

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.

Potential mode of action of a novel plumbagin as a mosquito repellent against the malarial vector Anopheles stephensi, (Culicidae: Diptera).

Plumbagin was isolated and characterized from the roots of Plumbago zeylanica using chromatography: TLC, Column chromatogram, HPLC, FTIR and 1H NMR. The isolated pure compounds were assayed for potency as inhibitors of: acetylcholine esterase (AchE), glutathione S-transferases (GST), superoxide dismutase (SOD), cytochrome P450 and α, ß-esterase, and for repellency with Anopheles stephensi at four different concentrations (25, 50, 75 and 100ppm). The enzyme assay against the pure compound reveals that the level of esterase and SOD was decreased significantly in contrast the level of GST and cytochrome P450 was increased significantly. Our results suggests that novel Plumbagin has significantly alters the level of enzyme comparable to the control. Evaluations resulted in Plumbagin producing maximum repellency scores against An. stephensi mosquitoes in dose dependent manner with highest repellence was observed in the 100ppm. Histological examination showed that the midgut, hindgut and muscles are the most affected tissues. These tissues affected with major changes including separation and collapse of epithelial layer and cellular vacuolization. The results support the utility of plant compound Plumbagin for vector control as an alternative to synthetic insecticides, however, more vigorous field trials are needed to determine viability under natural conditions.

Toxicity of aristolochic acids isolated from Aristolochia indica Linn (Aristolochiaceae) against the malarial vector Anopheles stephensi Liston (Diptera: Culicidae).

With the growth of resistance to overused insecticides, vector management has become highly problematic. Hence more concentration has been focused on botanicals. Therefore our present study was aimed to evaluate the toxicity of compounds, aristolochic acid I and aristolochic acid II from the methanol extract of Aristolochia indica L. (Aristolochiaceae) leaves on larvae of Anopheles stephensi L. (Diptera: Culicidae) employing World Health Organization standard larvicide testing procedures. The soxhlet extraction was carried out using polar solvent, methanol. The isolated toxic compounds were purified through RP-HPLC. The FTIR spectroscopic studies revealed different peak values with functional groups in the mixed compounds (AA-I and AA- II). These two aristolochic acids were further studied through (13)C and (1)HNMR analysis with confirmed by structures. Bioassay-guided fractionation through flash chromatography lead to the isolation of two larvicidal compounds namely aristolochic acid I and II. In these bioassays, the larvae were exposed to concentrations of 100, 250, 500,750 and 1000 ppm for each compound. Between the two, AA-I exerted no significant toxicity difference (P < 0.05) on mosquito larvae with LC50- 171.3, 209.8, 269.1, 502.3 ppm and LC90-751.6, 963.8, 972.7, 990.8 ppm compared to AA-II with LC50-134.8, 166.7, 240.4,543.2 ppm and LC90- 636.7, 792.5, 990.8, 986.2 ppm against first, second, third and fourth instars, respectively. Further, the isolated compounds were severely affecting the mosquito gut. From the results, A. indica toxic compounds could be considered as one of the influential applicant to bring about useful botanicals so as to prevent the resurrection of mosquito vectors.

Physiological and biochemical effects of botanical extract from Piper nigrum Linn (Piperaceae) against the dengue vector Aedes aegypti Liston (Diptera: Culicidae).

The leaves of Piper nigrum L. (Piperaceae) were evaluated for chemical constituents and mosquito larvicidal activity against the larvae of Aedes aegypti. GC and GC-MS analyses revealed that the crude extracts contain 16 compounds. Thymol (20.77%) and ç-elemene (10.42%) were identified as the major constituents followed by cyclohexene, 4-ethenyl-4-methyl-3-(1-methylethenyl)-1-(1 methylethyl)-, (3R-trans) (7.58%), 4,6-octadienoic acid, 2-acetyl-2-methyl-, ethyl ester (6.98), 2(3H)-furanone, 3,4-bis(1,3-benzodioxol-5-ylmethyl) dihydro-, (3R-trans) (6.95%), 1-naphthalenol, 1,2,3,4,4a,7,8,8a-octahydro-1,6-dimethyl-4-(1-methylethyl)-, [1R-(1à,4á,4aá,8aá)]-(Cedreanol) (5.30%), trans-2-undecen-1-ol (4.48%), phytol (4.22%), 1,6-cyclodecadiene, 1-methyl-5-methylene-8-(1-methylethyl)-,[s-(E,E)] (3.78%) and 2,6-dimethyl-3,5,7-octatriene-2-ol, Z,Z (2.39%). Larval mortality was observed after 3 h of exposure period. The crude extract showed remarkable larvicidal activity against Ae. aegypti (LC50 = 34.97). The larvae of Ae. aegypti exposed to the P. nigrum, significantly reduced the activities of α- and ß-carboxylesterases and superdioxide. Further, P. nigrum extract was severely affecting the mosquito gut cellular organelles. Based on the results, the chemical constituents of crude extracts of P. nigrum can be considered as a new source of larvicide for the control of Ae. aegypti.

Antimalarial efficacy of dynamic compound of plumbagin chemical constituent from Plumbago zeylanica Linn (Plumbaginaceae) against the malarial vector Anopheles stephensi Liston (Diptera: Culicidae).

In the present investigation, the effective root compound of plumbagin of Plumbago zeylanica (Plumbaginaceae) was evaluated for chemical constituent and antimalarial effect against the fourth instar larvae of Anopheles stephensi Liston (Diptera). In the chromatographic analyses of root compound with Rf value of 0.788 and NMR analyses also revealed that the effective compound contain naphthoquinone plumbagin were identified as the major chemical constituent. Larval mortality was observed after 3 h of exposure period. The plumbagin compound showed remarkable larvicidal activity against A. stephensi (LC50 32.65 and LC9072.27 ppm). Histopathological effects of compound was observed in the treated larvae. Based on the results, the plumbagin compound of P. zeylanica can be considered as a new source of natural larvicide for the control of malarial vector.

Anti-dengue efficacy of bioactive andrographolide from Andrographis paniculata (Lamiales: Acanthaceae) against the primary dengue vector Aedes aegypti (Diptera: Culicidae).

The current study investigated the toxic effect of the leaf extract compound andrographolide from Andrographis paniculata (Burm.f) against the dengue vector Ae. aegypti. GC-MS analysis revealed that andrographolide was recognized as the major chemical constituent with the prominent peak area compared with other compounds. All isolated toxic compounds were purified and confirmed through RP-HPLC against chemical standards. The larvicidal assays established at 25ppm of bioactive compound against the treated instars of Ae. Aegypti showed prominent mortality compared to other treated concentrations. The percent mortality of larvae was directly proportional to concentration. The lethal concentration (LC50) was observed at 12ppm treatment concentration. The bioactive andrographolide considerably reduced the detoxifying enzyme regulations of α- and ß- carboxylesterases. In contrast, the levels of GST and CYP450 significantly increase in a dose dependent manner. The andrographolide also showed strong oviposition deterrence effects at the sub-lethal dose of 12ppm. Similarly, the mean number of eggs were also significantly reduced in a dose dependent manner. At the concentration of 12ppm the effective percentage of repellency was greater than 90% with a protection time of 15-210min, compared with control. The histopathology study displayed that larvae treated with bioactive andrographolide had cytopathic effects in the midgut epithelium compared with the control. The present study established that bioactive andrographolide served as a potential useful for dengue vector management.

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.