Mosquitoes are the most important single group ofinsects in terms of public health importance, whichtransmit a number of diseases, such as malaria, filariasis,dengue, Japanese encephalitis, etc. causing millionsof deaths every year. Repeated use of syntheticinsecticides for mosquito control has disrupted naturalbiological control systems and led to resurgencesin mosquito populations. It has also resulted in thedevelopment of resistance1, undesirable effects onnon-target organisms and fostered environmental andhuman health concern2, which initiated a search foralternative control measures. Plants are considered asa rich source of bioactive chemicals3 and they may bean alternative source of mosquito control agents.Natural products of plant origin with insecticidalproperties have been tried in the recent past for controlof variety of insect pests and vectors. Essentialoils of leaf and bark of Cryptomeria japonica demonstratedhigh larvicidal activity against Aedesaegypti (Diptera: Culicidae) larvae4. Insecticidalactivity of plant essential oils has been well-describedby Isman5. Azadiractin, the active ingredient of neemhas long been recognised for its mosquito larvicidalcapability. The extracts of Murraya koenigii, Coriandrumsativam, Ferula asafetida and Trigonella foenumgraceum were found to be effective and showedencouraging results against Ae. aegypti6 and Culex(Diptera: Culicidae) mosquito larvae7. It is also reportedthat many compounds with insecticidal potentialhave been isolated from the genus Piper—Pipercide,isolated from Piper negrum (black piper) hasbeen found to be just as active against adjuki beanweevils as the pyrethroides8. Phytochemicals derivedfrom plant sources can act as larvicide, insect growthregulators, repellent and ovipositor attractant andhave different activities observed by many researchers9–11. However, insecticides of plant origin havebeen extensively used on agricultural pests and to avery limited extent, against insect vectors of publichealth importance.Northeastern region of India is considered as a majorbiodiversity hot spot. The eastern Himalayas range,which extends all through the northern border ofAssam, is a rich treasure house of many promisingmedicinal and aromatic plants. In the present communication,an attempt has been made to evaluate themosquito larvicidal efficacy of methanol and ethanolextracts of different parts of five indigenous plantsagainst Ae. albopictus (Diptera: Culicidae) and Culexquinquefasciatus larvae in laboratory conditions.Plant materials were collected from the foothill forestsof Sonitpur district, Assam bordering ArunachalPradesh during April and May 2005. They were segregatedas leaf, stem, bark, root and fruit/pericarp andair-dried in a shady place. Dried materials wereground in a table model grinder. The ground plant materialswere dipped in solvents (methanol and ethanol)Short Research Communications146 J VECT BORNE DIS 44, JUNE 2007in tightly capped jars separately for 48 h. The solventsalong with extracts were drained out, filtered andsemisolid extracts were obtained in vacuum usingrotary evaporator. The semisolid extracts were lyophilisedto obtain solid extracts. Stock solutions of desiredconcentration were prepared in distilled waterusing 1 ppm teepol as emulsifying agent and subsequentdilutions were made as per requirement. Larvicidalbioassay was carried out as per standard WHOtechniques in 500 ml glass beakers containing 250 mlof water and 25 numbers of late III or early IV instarmosquito larvae for various concentrations. Threedifferent concentrations of each extract were tried outat a time with six replicates. One control was kept witheach set of experiment and mortality was recordedafter 24 h. Five sets of experiments were conductedfor each extract. Tests were carried out under controlledlaboratory conditions (temperature 27 ± 2oC)against laboratory reared Ae. albopictus and Cx. quinquefasciatus(Diptera: Culicidae) larvae. Values obtainedwere subjected to log probit regression analysisto obtain LC50 and LC90 values with 95% confidencelimit12.The results showed that the larvicidal activity ofmethanol and ethanol extracts of five aromatic plantspecies against Ae. albopictus and Cx. quinquefasciatuslarvae varied according to plant species (Tables 1& 2). Methanol extract of Aristolochia saccata rootswas found to be the most effective against Ae. albopictuslarvae followed by ethanol extracts of A. saccata,Annona squamosa leaf and methanol extract ofA. squamosa leaf respectively. LC90 values of methanolextract of fruit/pericarp of Gymnopetelumcochinchinensis, bark of Caesalpinea species andethanol extract of stem of Piper species were obtainedat <200 ppm but methanol extract of seeds of G.cochinchinensis and stem of Piper species gave at<358 ppm against Ae. albopictus larvae (Table 1).Ethanol extract of leaf of A. squamosa was found tohave the most promising larvicidal activity againstCx. quinquefasciatus larvae. Methanol and ethanolextracts of A. saccata (root), methanol extract of A.squamosa (leaf) showed LC90 values at <100 ppmwhile methanol extract of G. cochinchinensis (fruit/pericarp), methanol and ethanol extract of Piper speciesshowed at <200 ppm and methanol extract of G.cochinchinensis (seed) showed at >302 ppm againstCx. quinquefasciatus larvae (Table 2).Long before the advent of synthetic insecticides,plants and their derivatives were used to kill pest ofagriculture, veterinary and public health. Sosan et al13reported larvicidal activities of essential oils of Ocimumgratissium, Cymbopogon citrus and Ageratumconyzoides against Ae. aegypti and achieved 100%mortality at 120, 200 and 300 ppm concentrationsrespectively. Similarly, it was reported that the essentialoil of Ipomoea cairica Linn. possesses remark-Table 1. Larvicidal efficacy of plant extracts against Ae. albopictus larvaeName of plant Part used Solvent used LC50 LC90 Regression equationAristolochia saccata Root Methanol 14.52 42.68 Y = 2.5683 *X+2.0164-do- Root Ethanol 17.30 58.51 Y = 2.3633 *X+2.0721Annona squamosa Leaf Methanol 20.26 86.59 Y = 1.9392 *X+2.4637-do- Leaf Ethanol 20.70 76.73 Y = 2.1991 *X+2.1020Gymnopetelum cochinchinensis Fruit/Pericarp Methanol 50.67 155.12 Y = 2.5821 *X+0.5927-do- Seed Methanol 100.42 312.45 Y = 2.3014 *X+0.3481Caesalpinea species Bark Methanol 53.66 169.41 Y = 2.3429*X+0.8638Piper species Stem Methanol 144.22 357.32 Y = 3.1826 *X–1.9688-do- Stem Ethanol 76.35 180.42 Y = 3.2525 *X–1.1333DAS et al : HERBAL MOSQUITO LARVICIDES 147able larvicidal properties as it could produce 100%mortality in the larvae of Cx. tritaeniorhynchus, Ae.aegypti, An. stephensi and Cx. quinquefasciatus mosquitoesat concentrations ranging from 100 to 170ppm14. Dwivedi & Kawasara15 found acetone extractof Lantana camara to be most effective against Cx.quinquefasciatus larvae at the dose of 1 ml/100 ml.Latha et al16 reported Piper longum and Zingiberwightianum extracts at 80 mg/l causing completemortality in Cx. quinquefasciatus and 60 mg/l for Cx.sitiens. In the present investigation LC90 values ofmethanol and ethanol extracts of roots of A. saccata,leaf of A. squamosa and fruits/pericarp of G.cochinchinensis against Ae. albopictus and Cx. quinquefasciatuslarvae ranged between 31.80 and 155ppm. Studies with essential oil of Ocimum americansand O. gratissium showed LC50 at 67 and 60 ppmrespectively against Ae. aegypti larvae17. In contrast,in the present study methanol and ethanol extracts ofroots of A. saccata, leaf of A. squamosa and fruits/pericarp of G. cochinchinensis against Ae. albopictusand Cx. quinquefasciatus larvae showed LC50 valuesbetween 6.96 and 57.4 ppm. Larvicidal activities ofthe plant extracts vary according to the plant species,the parts of the plant, the geographical location wherethe plants were grown and the application method.Plant could be an alternative source for mosquitolarvicides because they constitute a potential sourceof bioactive chemicals and generally free from harmfuleffects. Use of these botanical derivatives inmosquito control instead of synthetic insecticidescould reduce the cost and environmental pollution.Further studies on identification of active compounds,toxicity and field trials are needed to recommend theactive fraction of these plant extracts for developmentof eco-friendly chemicals for control of insect vectors.AcknowledgementThe authors are thankful to Dr. S.N. Dube, Director,Defence Research Laboratory, Tezpur, Assam, Indiafor extending necessary facilities, continuous supportand guidance in the study.References1. Brown AWA. Insecticide resistance in mosquitoes:pragmatic review. J Am Mosq Control Assoc 1986; 2: 123–40.2. Hayes JB Jr, Laws ER Jr. Handbook of pesticidetoxicology, v. 1. San Diego, CA: Academic Press 1991.3. Wink M. Production and application of pytochemicalsfrom an agricultural perspective. In: Van Beek TA, BretelerH, editors. Phytochemistry and agriculture. Oxford, UK:Clerendon Press 1993; p. 171–213.Table 2. Larvicidal efficacy of plant extracts against Cx. quinquefasciatus larvaeName of plant Part used Solvent used LC50 LC90 Regression equationAristolochia saccata Root Methanol 31.91 81.06 Y = 3.3086 *X-0.0062-do- Root Ethanol 19.83 60.44 Y = 2.5791 *X-1.6605Annona squamosa Leaf Methanol 17.70 64.29 Y = 2.1180 *X+2.3457-do- Leaf Ethanol 6.96 31.80 Y = 1.9441 *X+3.3592Gymnopetelum cochinchinensis Fruit/pericarp Methanol 57.4 108.3 Y = 4.1627 *X+2.3501-do- Seed Methanol 199.0 301.6 Y = 1.7586 *X+11.0557Caesalpinea species Bark Methanol 42.27 207.13 Y = 1.7586 *X+2.0573Piper species Stem Methanol 70.10 113.90 Y = 5.669 *X+5.4992-do- Stem Ethanol 57.4 108.3 Y = 4.1627 *X+2.3501148 J VECT BORNE DIS 44, JUNE 20074. Cheng SS, Chang HT, Chang ST, Tsai KH, Chen WJ.Bioactivity of selected plant essential oils against theyellow fever mosquito Aedes aegypti larvae. BioresTechnol 2003; 89(1): 99–102.5. Isman MB. Pesticides based on plant essential oils.Pesticide Outlook 1999; p. 68–72.6. Harve G, Kamath V. Larvicidal activity of plant extractsused alone and in combination with known syntheticlarvicidal agents against Aedes aegypti. Indian J ExptlBiol 2004; 42: 1216–9.7. Desai ST. Potency of larvicidal properties of plantextracts against mosquito larvae under laboratoryconditions (M.Sc. Dissertation submitted to MumbaiUniversity Mumbai, India 2002).8. Mwangi RW, Mukiama TK. Evaluation of Melia volkensiextract fractions as mosquito larvicides. J Am MosqControl Assoc 1988; 4: 442–7.9. Babu R, Murugan K. Interactive effect of neem seed kernaand neem gum extract on the control of Culex quinquefasciatusSay. Neem Newsletter 1998; 15(2): 9–11.10. Venketachalam MR, Jebasan A. Repellent activity ofFerronia elephantum Corr. (Rutaceae) leaf extract againstAedes aegypti. Biores Technol 2001; 76(3): 287–8.11. Venketachalam MR, Jebasan A. Larvicidal activity ofHydrocotyl javanica Thunb (Apiaceae) extract against Cx.quinquefasciatus. J Exptl Zool India 2001; 4(1): 99–101.12. Finney DJ. Probit analysis, III edn. Cambridge: CambridgeUniversity Press 1971.13. Sosan MB, Adewoyin FB, Adewunmi CO. Larvicidalproperties of three indigenous plant oils on the mosquitoAedes aegypti. Nigerian J Natl Prod Med 2001; 5: 30–3.14. Thomas TG, Rao S, Lal S. Mosquito larvicidal propertiesof an indigenous plant, Ipomoea cairica Linn. JapaneseJ Infect Dis 2004; 57: 176–7.15. Dwivedi SC, Karwasara K. Larvicidal activity of fiveplants extracts against Culex quinquefasciatus. Indian JEntomol 2003; 65(3): 335–8.16. Latha C, Vijhayakumar PD, Velayudhan S, Joseph A.Biological activity of indigenous plant extracts as mosquitolarvicides. Indian J Exptl Biol 1999; 37: 206–8.17. Cavalcanti ESB, Morais SM, Lima MAA, Santana EWP.Larvicidal activity of essential oils from Brazilian plantsagainst Aedes aegypti L. Mem Inst Oswaldo Cruz 2004;99(5): 541–4.Corresponding author: Dr. N.G. Das, Medical Entomology Division, Defence Researc
