Nanotechnology-based approaches for emerging and re-emerging viruses: Special emphasis on COVID-19.

In recent decades, the major concern of emerging and re-emerging viral diseases has become an increasingly important area of public health concern, and it is of significance to anticipate future pandemic that would inevitably threaten human lives. The novel severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a newly emerged virus that causes mild to severe pneumonia. Coronavirus disease (COVID-19) became a very much concerned issue worldwide after its super-spread across the globe and emerging viral diseases have not got specific and reliable diagnostic and treatments. As the COVID-19 pandemic brings about a massive life-loss across the globe, there is an unmet need to discover a promising and typically effective diagnosis and treatment to prevent super-spreading and mortality from being decreased or even eliminated. This study was carried out to overview nanotechnology-based diagnostic and treatment approaches for emerging and re-emerging viruses with the current treatment of the disease and shed light on nanotechnology's remarkable potential to provide more effective treatment and prevention to a special focus on recently emerged coronavirus.

Molecular Docking Studies on the Anti-viral Effects of Compounds From Kabasura Kudineer on SARS-CoV-2 3CLpro.

The COVID-19 has now been declared a global pandemic by the World Health Organization. No approved drug is currently available; therefore, an urgent need has been developed for any antiviral therapy for COVID-19. Main protease 3CLpro of this novel Coronavirus (SARS-CoV-2) play a critical role in the disease propagation, and hence represent a crucial target for the drug discovery. Herein, we have applied a bioinformatics approach for drug repurposing to identify the possible potent inhibitors of SARS-CoV-2 main proteases 3CLpro (6LU7). In search of the anti-COVID-19 compound, we selected 145 phyto-compounds from Kabasura kudineer (KK), a poly-herbal formulation recommended by AYUSH for COVID-19 which are effective against fever, cough, sore throat, shortness of breath (similar to SARS-CoV2-like symptoms). The present study aims to identify molecules from natural products which may inhibit COVID-19 by acting on the main protease (3CLpro). Obtained results by molecular docking showed that Acetoside (-153.06), Luteolin 7 -rutinoside (-134.6) rutin (-133.06), Chebulagic acid (-124.3), Syrigaresinol (-120.03), Acanthoside (-122.21), Violanthin (-114.9), Andrographidine C (-101.8), myricetin (-99.96), Gingerenone -A (-93.9), Tinosporinone (-83.42), Geraniol (-62.87), Nootkatone (-62.4), Asarianin (-79.94), and Gamma sitosterol (-81.94) are main compounds from KK plants which may inhibit COVID-19 giving the better energy score compared to synthetic drugs. Based on the binding energy score, we suggest that these compounds can be tested against Coronavirus and used to develop effective antiviral drugs.

Phenotypic Screening Identifies Synergistically Acting Natural Product Enhancing the Performance of Biomaterial Based Wound Healing.

The potential of multifunctional wound heal biomaterial relies on the optimal content of therapeutic constituents as well as the desirable physical, chemical, and biological properties to accelerate the healing process. Formulating biomaterials such as amnion or collagen based scaffolds with natural products offer an affordable strategy to develop dressing material with high efficiency in healing wounds. Using image based phenotyping and quantification, we screened natural product derived bioactive compounds for modulators of types I and III collagen production from human foreskin derived fibroblast cells. The identified hit was then formulated with amnion to develop a biomaterial, and its biophysical properties, in vitro and in vivo effects were characterized. In addition, we performed functional profiling analyses by PCR array to understand the effect of individual components of these materials on various genes such as inflammatory mediators including chemokines and cytokines, growth factors, fibroblast stimulating markers for collagen secretion, matrix metalloproteinases, etc., associated with wound healing. FACS based cell cycle analyses were carried out to evaluate the potential of biomaterials for induction of proliferation of fibroblasts. Western blot analyses was done to examine the effect of biomaterial on collagen synthesis by cells and compared to cells grown in the presence of growth factors. This work demonstrated an uncomplicated way of identifying components that synergistically promote healing. Besides, we demonstrated that modulating local wound environment using biomaterials with bioactive compounds could enhance healing. This study finds that the developed biomaterials offer immense scope for healing wounds by means of their skin regenerative features such as anti-inflammatory, fibroblast stimulation for collagen secretion as well as inhibition of enzymes and markers impeding the healing, hydrodynamic properties complemented with other features including non-toxicity, biocompatibility, and safety.

Bioinformatics approach to prioritize known drugs towards repurposing for tuberculosis.

New drugs are urgently needed to cure tuberculosis (TB) in a short period of time without causing any adverse effects since currently used drugs for the treatment of multi drug-resistant TB cause several adverse effects with poor success rate. Therefore, we aimed to prioritize known drugs towards repurposing for TB by employing bioinformatics approach in the present study. A total of 1554 FDA approved drugs were obtained from DrugBank. Serine/threonine-protein kinase, pknB (Rv0014c) of Mycobacterium tuberculosis (Mtb) was selected as the drug target since it involves in several vital functions of the Mtb. All of the 1554 drugs were subjected to molecular docking with pknB. Glide and AutoDock Vina were employed using rigid docking followed by induced fit docking protocol for prioritization of drugs. Out of 14 drugs prioritized, six are suggested as high-confident drugs towards repurposing for TB as they were consistently found within top 10 ranks of both methods, and strongly binding in the active site of the pknB. We also found atorvastatin as one of the high-confident drugs, which has already been demonstrated to be active against Mtb under in vitro conditions by other researchers. Therefore, we propose that the prioritized six high-confident drugs as potential candidates for repurposing for TB and suggest for further experimental studies. We also suggest that the bioinformatics procedure we have employed in this study could be effectively applied for prioritization of drugs for other diseases.

Green synthesis of silver nanoparticles using Rheum palmatum root extract and their antibacterial activity against Staphylococcus aureus and Pseudomonas aeruginosa.

In the present study, silver nanoparticles (AgNPs) were synthesized by using aqueous root extracts of Rheum palmatum and characterized by various spectroscopic methods. The nanoparticles were found to be in hexagonal and spherical shapes. The average particle size was found to be 121 ± 2 nm with zeta potential values of -21.6 mv by dynamic light scattering (DLS) method. Gas chromatography-mass spectrometry (GC-MS) analysis of R. palmatum revealed 35 compounds. The synthesized AgNPs showed significant activity against Staphylococcus aureus and Pseudomonas aeruginosa with IC90 values of 15 µg/ml and IC50 values of 7.5 µg/ml, respectively. The protein leakage level was high and morphological changes occurred in bacteria treated with AgNPs.

HER2 Targeted Breast Cancer Therapy with Switchable "Off/On" Multifunctional "Smart" Magnetic Polymer Core-Shell Nanocomposites.

Multifunctional magnetic polymer nanocombinations are gaining importance in cancer nanotheranostics due to their safety and their potential in delivering targeted functions. Herein, we report a novel multifunctional core-shell magnetic polymer therapeutic nanocomposites (NCs) exhibiting pH dependent "Off-On" release of drug against breast cancer cells. The NCs are intact in blood circulation ("Off" state), i.e., at physiological pH, whereas activated ("On" state) at intracellular acidic pH environment of the targeted breast cancer cells. The NCs are prepared by coating the cannonball (iron nanocore) with hydrophobic nanopockets of pH-responsive poly(d,l-lactic-co-glycolic acid) (PLGA) polymer nanoshell that allows efficient loading of therapeutics. Further, the nanocore-polymer shell is stabilized by poly(vinylpyrrolidone) (PVP) and functionalized with a targeting HER2 ligand. The prepared Her-Fe3O4@PLGA-PVP nanocomposites facilitate packing of anticancer drug (Tamoxifen) without premature release in the bloodstream, recognizing the target cells through binding of Herceptin antibody to HER2, a cell surface receptor expressed by breast cancer cells to promote HER2 receptor mediated endocytosis and finally releasing the drug at the intracellular site of tumor cells ("On" state) to induce apoptosis. The therapeutic efficiency of hemo/cytocompatible NCs drug delivery system (DDS) in terms of targeted delivery and sustained release of therapeutic agent against breast cancer cells was substantiated by in vitro and in vivo studies. The multifunctional properties of Her-Tam-Fe3O4@PLGA-PVP NCs may open up new avenues in cancer therapy through overcoming the limitations of conventional cancer therapy.

Docking-based virtual screening of known drugs against murE of Mycobacterium tuberculosis towards repurposing for TB.

Repurposing has gained momentum globally and become an alternative avenue for drug discovery because of its better success rate, and reduced cost, time and issues related to safety than the conventional drug discovery process. Several drugs have already been successfully repurposed for other clinical conditions including drug resistant tuberculosis (DR-TB). Though TB can be cured completely with the use of currently available anti-tubercular drugs, emergence of drug resistant strains of Mycobacterium tuberculosis and the huge death toll globally, together necessitate urgently newer and effective drugs for TB. Therefore, we performed virtual screening of 1554 FDA approved drugs against murE, which is essential for peptidoglycan biosynthesis of M. tuberculosis. We used Glide and AutoDock Vina for virtual screening and applied rigid docking algorithm followed by induced fit docking algorithm in order to enhance the quality of the docking prediction and to prioritize drugs for repurposing. We found 17 drugs binding strongly with murE and three of them, namely, lymecycline, acarbose and desmopressin were consistently present within top 10 ranks by both Glide and AutoDock Vina in the induced fit docking algorithm, which strongly indicates that these three drugs are potential candidates for further studies towards repurposing for TB.

Anti-chikungunya activity of luteolin and apigenin rich fraction from Cynodon dactylon.

OBJECTIVE: To obtain luteolin and apigenin rich fraction from the ethanolic extract of Cynodon dactylon (L.) (C. dactylon) Pers and evaluate the fraction's cytotoxicity and anti-Chikungunya potential using Vero cells. METHODS: The ethanolic extract of C. dactylon was subjected to silica gel column chromatography to obtain anti-chikungunya virus (CHIKV) fraction. Reverse phase-HPLC and GC-MS studies were carried out to identify the major phytochemicals in the fraction using phytochemical standards. Cytotoxicity and the potential of the fraction against CHIKV were evaluated in vitro using Vero cells. Reduction in viral replication was assessed by reverse transcriptase-polymerase chain reaction (RT-PCR) after treating the viral infected Vero cells with the fraction. RESULTS: Reverse Phase-HPLC and GC-MS studies confirmed the presence of flavonoids, luteolin and apigenin as major phytochemicals in the anti-CHIKV ethanolic fraction of C. dactylon. The fraction was found to exhibit potent viral inhibitory activity (about 98%) at the concentration of 50 µg/mL as observed by reduction in cytopathic effect, and the cytotoxic concentration of the fraction was found to be 250 µg/mL. RT-PCR analyses indicated that the reduction in viral mRNA synthesis in fraction treated infected cells was much higher than the viral infected control cells. CONCLUSIONS: Luteolin and apigenin rich ethanolic fraction from C. dactylon can be utilized as a potential therapeutic agent against CHIKV infection as the fraction does not show cytotoxicity while inhibiting the virus.

Biosynthesized silver nanoparticles using floral extract of Chrysanthemum indicum L.--potential for malaria vector control.

Mosquitoes transmit serious human diseases, causing millions of deaths every year. The use of synthetic insecticides to control vector mosquitoes has caused physiological resistance and adverse environmental effects in addition to high operational cost. Insecticides synthesized of natural products for vector control have been a priority in this area. In the present study, silver nanoparticles (Ag NPs) were green-synthesized using a floral extract of Chrysanthemum indicum screened for larvicidal and pupicidal activity against the first to fourth instar larvae and pupae of the malaria vector Anopheles stephensi mosquitoes. The synthesized Ag NPs were characterized by using UV-vis absorption, X-ray diffraction, transmission electron microscopy, and energy-dispersive X-ray spectroscopy techniques. The textures of the yielded Ag NPs were found to be spherical and polydispersed with a mean size in the range of 25-59 nm. Larvae and pupae were exposed to various concentrations of aqueous extract of C. indicum and synthesized Ag NPs for 24 h, and the maximum mortality was observed from the synthesized Ag NPs against the vector A. stephensi (LC50 = 5.07, 10.35, 14.19, 22.81, and 35.05 ppm; LC90 = 29.18, 47.15, 65.53, 87.96, and 115.05 ppm). These results suggest that the synthesized Ag NPs have the potential to be used as an ideal eco-friendly approach for the control of A. stephensi. Additionally, this study provides the larvicidal and pupicidal properties of green-synthesized Ag NPs with the floral extract of C. indicum against vector mosquito species from the geographical location of India.

Anti-chikungunya activity of luteolin and apigenin rich fraction from Cynodon dactylon

Objective:To obtain luteolin and apigenin rich fraction from the ethanolic extract ofCynodon dactylon (L.) (C. dactylon) Pers and evaluate the fraction’s cytotoxicity and anti-Chikungunya potential using Vero cells.Methods:The ethanolic extract ofC. dactylon was subjected to silica gel column chromatography to obtain anti-chikungunya virus (CHIKV) fraction. Reverse phase-HPLC and GC-MS studies were carried out to identify the major phytochemicals in the fraction using phytochemical standards. Cytotoxicity and the potential of the fraction against CHIKV were evaluatedin vitrousing Vero cells. Reduction in viral replication was assessed by reverse transcriptase-polymerase chain reaction (RT-PCR) after treating the viral infected Vero cells with the fraction.Results:Reverse Phase-HPLC and GC-MS studies confirmed the presence of flavonoids, luteolin and apigenin as major phytochemicals in the anti-CHIKV ethanolic fraction ofC. dactylon. The fraction was found to exhibit potent viral inhibitory activity (about 98%) at the concentration of 50 μg/mL as observed by reduction in cytopathic effect, and the cytotoxic concentration of the fraction was found to be 250 μg/mL. RT-PCR analyses indicated that the reduction in viral mRNA synthesis in fraction treated infected cells was much higher than the viral infected control cells.Conclusions:Luteolin and apigenin rich ethanolic fraction fromC. dactylon can be utilized as a potential therapeutic agent against CHIKV infection as the fraction does not show cytotoxicity while inhibiting the virus.

Rapid green synthesis of silver nanoparticles from Chrysanthemum indicum L and its antibacterial and cytotoxic effects: an in vitro study.

The present work reports a simple, cost-effective, and ecofriendly method for the synthesis of silver nanoparticles (AgNPs) using Chrysanthemum indicum and its antibacterial and cytotoxic effects. The formation of AgNPs was confirmed by color change, and it was further characterized by ultraviolet-visible spectroscopy (435 nm). The phytochemical screening of C. indicum revealed the presence of flavonoids, terpenoids, and glycosides, suggesting that these compounds act as reducing and stabilizing agents. The crystalline nature of the synthesized particles was confirmed by X-ray diffraction, as they exhibited face-centered cubic symmetry. The size and morphology of the particles were characterized by transmission electron microscopy, which showed spherical shapes and sizes that ranged between 37.71-71.99 nm. Energy-dispersive X-ray spectroscopy documented the presence of silver. The antimicrobial effect of the synthesized AgNPs revealed a significant effect against the bacteria Klebsiella pneumonia, Escherichia coli, and Pseudomonas aeruginosa. Additionally, cytotoxic assays showed no toxicity of AgNPs toward 3T3 mouse embryo fibroblast cells (25 µg/mL); hence, these particles were safe to use.

Characterization of ambrette seed oil and its mode of action in bacteria.

In the present study, chemical composition and the antibacterial mechanism of ambrette seed oil are investigated. Chemical composition of the oil was analysed by gas chromatography-mass spectrometry (GC-MS). Thirty-five compounds were identified and the major compounds were found to be farnesol acetate (51.45%) and ambrettolide (12.96%). The antibacterial activity was performed by well diffusion assay and the mechanisms were studied by measuring the alkaline phosphatase (ALP), lactate dehydrogenase (LDH) and protein leakage assays. The antibacterial effect of the ambrette seed oil showed inhibitory effect against Bacillus subtilis, Staphylococcus aureus and Enterococcus faecalis. The LDH activity was high in all tested bacteria compared with control, whereas the ALP and protein concentrations were also increased in E. faecalis. Molecular docking revealed the ligands farnesol acetate and ambrettolide had satisfactory binding energy towards the beta lactamase TEM-72 and dihydrofolate reductase (DHFR) protein. Due to its better antibacterial properties, the ambrette seed oil could be used as a source of antibacterial agents.

Larvicidal activity of Morinda citrifolia L. (Noni) (Family: Rubiaceae) leaf extract against Anopheles stephensi, Culex quinquefasciatus, and Aedes aegypti.

Morinda citrifolia leaf extract was tested for larvicidal activity against three medically important mosquito vectors such as malarial vector Anopheles stephensi, dengue vector Aedes aegypti, and filarial vector Culex quinquefasciatus. The plant material was shade dried at room temperature and powdered coarsely. From the leaf, 1-kg powder was macerated with 3.0 L of hexane, chloroform, acetone, methanol, and water sequentially for a period of 72 h each and filtered. The yield of extracts was hexane (13.56 g), chloroform (15.21 g), acetone (12.85 g), methanol (14.76 g), and water (12.92 g), respectively. The extracts were concentrated at reduced temperature on a rotary vacuum evaporator and stored at a temperature of 4°C. The M. citrifolia leaf extract at 200, 300, 400, 500, and 600 ppm caused a significant mortality of three mosquito species. Hexane, chloroform, acetone, and water caused moderate considerable mortality; however, the highest larval mortality was methanolic extract, observed in three mosquito vectors. The larval mortality was observed after 24-h exposure. No mortality was observed in the control. The third larvae of Anopheles stephensi had values of LC(50) = 345.10, 324.26, 299.97, 261.96, and 284.59 ppm and LC(90) = 653.00, 626.58, 571.89, 505.06, and 549.51 ppm, respectively. The Aedes aegypti had values of LC(50) = 361.75, 343.22, 315.40, 277.92, and 306.98 ppm and LC(90) = 687.39, 659.02, 611.35, 568.18, and 613.25 ppm, respectively. The Culex quinquefasciatus had values of LC(50) = 382.96, 369.85, 344.34, 330.42, and 324.64 ppm and LC(90) = 726.18, 706.57, 669.28, 619.63, and 644.47 ppm, respectively. The results of the leaf extract of M. citrifolia are promising as good larvicidal activity against the mosquito vector Anopheles stephensi, Aedes aegypti, and Culex quinquefasciatus. This is a new eco-friendly approach for the control of vector control programs. Therefore, this study provides first report on the larvicidal activities against three species of mosquito vectors of this plant extracts from India.

Larvicidal efficacy of Sphaeranthus indicus, Cleistanthus collinus and Murraya koenigii leaf extracts against filarial vector, Culex quinquefasciatus Say (Diptera: Culicidae).

Sphaeranthus indicus, Cleistanthus collinus and Murraya koenigii leaf extracts were tested against the third instar larvae of Culex quinquefasciatus. The plant material was shade dried at room temperature and powdered coarsely. From each plant, 500 g powder was macerated with 1.5 L of hexane, chloroform and ethyl acetate sequentially for a period of 72 h each and filtered. The yield of the S. indicus, C. collinus and M. koenigii crude extracts by hexane, chloroform and ethyl acetate was 9.16, 11.71 and 10.83 g for S. indicus; 8.17, 10.69 and 9.85 g for C. collinus; and 10.11, 11.92 and 9.87 g for M. koenigii, respectively. The extracts were concentrated at reduced temperature on a rotary vacuum evaporator and stored at a temperature of 4°C. The S. indicus, C. collinus and M. koenigii leaf extracts at 250, 500, 750 and 1,000 ppm caused a significant mortality of C. quinquefasciatus. The LC(50) and LC(90) values of S. indicus, C. collinus and M. koenigii against third instar larvae at 24, 48 and 72 h (hexane, chloroform and ethyl acetate) were the following: S. indicus LC(50) values were 544.93, 377.86 and 274.79 ppm and LC(90) values were 1,325.32, 1,572.55 and 1,081.29 ppm at 24 h; C. collinus LC(50) values were 375.34, 318.29 and 226.10 ppm and LC(90) values were 699.65, 1,577.62 and 1,024.92 ppm at 24 h; and M. koenigii LC(50) values were 963.53, 924.85 and 857.62 ppm and LC(90) values were 1,665.12, 1,624.68 and 1,564.37 ppm at 24 h, respectively. However, the highest larval mortality was observed in C. collinus followed by S. indicus and M. koenigii of various concentrations at 24, 48 and 72 h. The study proved that S. indicus, C. collinus and M. koenigii leaf extracts had larvicidal property against species of C. quinquefasciatus. This is an ideal ecofriendly approach for the control of vector control programmes.

Mosquitocidal properties of Calotropis gigantea (Family: Asclepiadaceae) leaf extract and bacterial insecticide, Bacillus thuringiensis, against the mosquito vectors.

Calotropis gigantea leaf extract and Bacillus thuringiensis were tested first to fourth-instar larvae and pupae of Anopheles stephensi, Aedes aegypti, and Culex quinquefasciatus. The medicinal plants were collected from the area around Bharathiar University, Coimbatore, India. Calotropis gigantea leaf was washed with tap water and shade-dried at room temperature. An electrical blender powdered the dried plant materials (leaves). The powder 500 g of the leaf was extracted with 1.5 L of organic solvents of methanol for 8 h using a Soxhlet apparatus and filtered. The crude leaf extracts were evaporated to dryness in a rotary vacuum evaporator. The plant extract showed larvicidal and pupicidal effects after 24 h of exposure; no mortality was observed in the control group. For Calotropis gigantea, the median lethal concentration values (LC(50)) observed for the larvicidal and pupicidal activities against mosquito vector species Anopheles stephensi I to IV larval instars and pupae were 73.77, 89.64, 121.69, 155.49, and 213.79 ppm; Aedes aegypti values were 92.27, 106.60, 136.48, 164.01, and 202.56 ppm; and Culex quinquefasciatus values were 104.66, 127.71, 173.75, 251.65, and 314.70 ppm, respectively. For B. thuringiensis, the LC(50) values of I to IV larval instars and pupae of Anopheles stephensi were 37.24, 45.41, 57.82, 80.09, and 98.34 ppm; Aedes aegypti values were 42.38, 51.90, 71.02, 96.17, and 121.59 ppm; and Culex quinquefasciatus values were 55.85, 68.07, 94.11, 113.35, and 133.87 ppm, respectively. The study proved that the methanol leaf extract of Calotropis gigantea and bacterial insecticide B. thuringiensis has mosquitocidal property and was evaluated as target species of mosquito vectors. This is an ideal ecofriendly approach for the control of vector control programs.

Bioefficacy of larvicdial and pupicidal properties of Carica papaya (Caricaceae) leaf extract and bacterial insecticide, spinosad, against chikungunya vector, Aedes aegypti (Diptera: Culicidae).

The present study was carried out to establish the properties of Carica papaya leaf extract and bacterial insecticide, spinosad on larvicidal and pupicidal activity against the chikungunya vector, Aedes aegypti. The medicinal plants were collected from the area around Bharathiar University, Coimbatore, India. C. papaya leaf was washed with tap water and shade-dried at room temperature. An electrical blender powdered the dried plant materials (leaves). The powder (500 g) of the leaf was extracted with 1.5 l of organic solvents of methanol for 8 h using a Soxhlet apparatus and then filtered. The crude leaf extracts were evaporated to dryness in a rotary vacuum evaporator. The plant extract showed larvicidal and pupicidal effects after 24 h of exposure; however, the highest larval and pupal mortality was found in the leaf extract of methanol C. papaya against the first- to fourth-instar larvae and pupae of values LC(50) = I instar was 51.76 ppm, II instar was 61.87 ppm, III instar was 74.07 ppm, and IV instar was 82.18 ppm, and pupae was 440.65 ppm, respectively, and bacterial insecticide, spinosad against the first to fourth instar larvae and pupae of values LC(50) = I instar was 51.76 ppm, II instar was 61.87 ppm, III instar was 74.07 ppm, and IV instar was 82.18 ppm, and pupae was 93.44 ppm, respectively. Moreover, combined treatment of values of LC(50) = I instar was 55.77 ppm, II instar was 65.77 ppm, III instar was 76.36 ppm, and IV instar was 92.78 ppm, and pupae was 107.62 ppm, respectively. No mortality was observed in the control. The results that the leaves extract of C. papaya and bacterial insecticide, Spinosad is promising as good larvicidal and pupicidal properties of against chikungunya vector, A. aegypti. This is an ideal eco-friendly approach for the control of chikungunya vector, A. aegypti as target species of vector control programs.

Studies on larvicidal and pupicidal activity of Leucas aspera Willd. (Lamiaceae) and bacterial insecticide, Bacillus sphaericus, against malarial vector, Anopheles stephensi Liston. (Diptera: Culicidae).

The efficacy of whole plant extracts of Leucas aspera and Bacillus sphaericus has been proven against larvicidal and pupicidal activities of the malarial vector, Anopheles stephensi. The present study investigated the larvicidal and pupicidal activity against the first to fourth instar lavae and pupae of the laboratory-reared mosquitoes, A. stephensi. The medicinal plants were collected from the area around Maruthamalai hills, Coimbatore, Tamil Nadu, India. L. aspera whole plant was washed with tap water and shade dried at room temperature. The dried plant materials were powdered by an electric blender. From the powder, 100 g of the plant materials was extracted with 300 ml of organic solvents of ethanol for 8 h using a Soxhlet apparatus. The extracts were filtered through a Buchner funnel with Whatman number 1 filter paper. The crude plant extracts were evaporated to dryness in a rotary vacuum evaporator. The plant extract showed larvicidal and pupicidal effects after 24 h of exposure. All larval instars and pupae have considerably moderate mortality; however, the highest larval mortality was the ethanolic extract of whole plant L. aspera against the first to fourth instar larvae and pupae values of LC(50) = I instar was 9.695%, II instar was 10.272%, III instar was 10.823%, and IV instar was 11.303%, and pupae was 12.732%. B. spaericus against the first to fouth instar larvae and pupae had the following values: I instar was 0.051%, II instar was 0.057%, III instar was 0.062%, IV instar was 0.066%, and for the pupae was 0.073%. No mortality was observed in the control. The present results suggest that the ethanolic extracts of L. aspera and B. sphaericus provided an excellent potential for controlling of malarial vector, A. stephensi.