Environmental Science and Pollution Research

, Volume 25, Issue 11, pp 10317–10327 | Cite as

Eco-friendly and cost-effective Ag nanocrystals fabricated using the leaf extract of Habenaria plantaginea: toxicity on six mosquito vectors and four non-target species

  • Chinnadurai Aarthi
  • Marimuthu Govindarajan
  • Pichaimuthu Rajaraman
  • Naiyf S. Alharbi
  • Shine Kadaikunnan
  • Jamal M. Khaled
  • Ramzi A. Mothana
  • Nasir A. Siddiqui
  • Giovanni Benelli
Plant-borne compounds and nanoparticles: challenges for medicine, parasitology and entomology


Recently, the biofabrication of metal nanoparticles has gained wide interest owing to its inherent features such as swift, simplicity, eco-friendliness, and cheaper costs. Different green-reducing agents led to the production of nanoparticles with varying toxicity on insects. In the current study, silver nanoparticles (AgNPs) were successfully synthesized using Habenaria plantaginea leaf extract. Ag nanoparticles were studied by UV–Vis spectroscopy (UV-Vis), X-ray diffraction (XRD), Fourier transform infrared spectroscopy (FTIR), atomic force microscopy (AFM), scanning electron microscopy (SEM) coupled with energy-dispersive spectroscopy (EDS), and transmission electron microscopy (TEM). H. plantaginea extract and AgNPs were tested for mosquito larvicidal activity on Anopheles stephensi, Aedes aegypti, Culex quinquefasciatus, An. subpictus, Ae. albopictus, and Cx. tritaeniorhynchus. LC50 values were 102.51, 111.99, 123.47, 123.96, 136.56, 149.42 μg/ml and 12.23, 13.38, 14.78, 14.37, 15.39, 16.89 μg/ml, respectively. Moreover, H. plantaginea aqueous extract and AgNPs were tested against the non-target species Anisops bouvieri, Diplonychus indicus, Poecilia reticulata, and Gambusia affinis obtaining LC50 values ranging from 831.82 to 36,212.67 μg/ml. Overall, this study showed the effectiveness of H. plantaginea-fabricated nanoparticles on a wide range of important mosquito vectors, highlighting their scarce toxicity on four natural enemies predating mosquito larvae and pupae.


Biofabrication Biosafety Biopesticide AFM, SEM, TEM Zika virus 



The authors extend their sincere appreciations to the Deanship of Scientific Research at King Saud University for funding the work through the research group project no. (RGP-073). The authors would like to thank the Principal and Head of the Department of Zoology, Thiru. Vi. Ka Government Arts College and the Professor and Head, Department of Zoology, Annamalai University for the laboratory facilities provided.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.


  1. Ajitha B, Reddy YAK, Reddy PS (2014) Biogenic nano-scale silver particles by Tephrosia purpurea leaf extract and their inborn antimicrobial activity. Spectrochim Acta Part A Mol Biomol Spectrosc 121:164–172CrossRefGoogle Scholar
  2. AlQahtani FS, AlShebly MM, Govindarajan M, Senthilmurugan S, Vijayan P, Benelli G (2017) Green and facile biosynthesis of silver nanocomposites using the aqueous extract of Rubus ellipticus leaves: toxicity and oviposition deterrent activity against Zika virus, malaria and filariasis mosquito vectors. J Asia Pac Entomol 20:157–164Google Scholar
  3. Antony JJ, Sithika MAA, Joseph TA, Suriyakalaa U, Sankarganesh A, Siva D, Kalaiselvi S, Achiraman S (2013) In vivo antitumor activity of biosynthesised silver nanoparticles using Ficus religiosa as a nano factory in DAL induced mice model. Colloids Surf B Biointerfaces 108:185–190CrossRefGoogle Scholar
  4. Ashokkumar S, Ravi S, Kathiravan V, Velmurugan S (2014) Synthesis, characterization and catalytic activity of silver nanoparticles using Tribulus terrestris leaf extract. Spectrochim Acta Part A Mol Biomol Spectrosc 121:88–93CrossRefGoogle Scholar
  5. Balavigneswaran CK, Sujin Jeba Kumar T, Moses Packiaraj R, Prakash S (2014) Rapid detection of Cr (VI) by AgNPs probe produced by Anacardium occidentale fresh leaf extracts. Appl Nanosci 4:367–378CrossRefGoogle Scholar
  6. Banumathi B, Vaseeharan B, Suganya P, Citarasu T, Govindarajan M, Alharbi NS et al. (2017) Toxicity of camellia sinensis-fabricated silver nanoparticles on invertebrate and vertebrate organisms: morphological abnormalities and DNA damages. J Clust Sci. doi: 10.1007/s10876-017-1201-5
  7. Benelli G (2015a) Research in mosquito control: current challenges for a brighter future. Parasitol Res 114:2801–2805CrossRefGoogle Scholar
  8. Benelli G (2015b) Plant-borne ovicides in the fight against mosquito vectors of medical and veterinary importance: a systematic review. Parasitol Res 114:3201–3212CrossRefGoogle Scholar
  9. Benelli G (2016a) Plant-mediated biosynthesis of nanoparticles as an emerging tool against mosquitoes of medical and veterinary importance: a review. Parasitol Res 115:23–34CrossRefGoogle Scholar
  10. Benelli G (2016b) Green synthesized nanoparticles in the fight against mosquito-borne diseases and cancer—a brief review. Enzyme Microbial Technol 95:58–68CrossRefGoogle Scholar
  11. Benelli G (2017) Commentary: data analysis in bionanoscience—issues to watch for. J Clust Sci 28:11–14Google Scholar
  12. Benelli G, Govindarajan M (2017) Green-synthesized mosquito oviposition attractants and ovicides: towards a nanoparticle-based “lure and kill” approach?. J Clust Sci 28:287–308Google Scholar
  13. Benelli G, Kadaikunnan S, Alharbi NS, Govindarajan M (2017d) Biophysical characterization of Acacia caesia-fabricated silver nanoparticles: effectiveness on mosquito vectors of public health relevance and impact on non-target aquatic biocontrol agents. Environ Sci Pollut Res DOI  10.1007/s11356-017-8482-y
  14. Benelli G, Lo Iacono A, Canale A, Mehlhorn H (2016) Mosquito vectors and the spread of cancer: an overlooked connection? Parasitol Res 115:2131–2137CrossRefGoogle Scholar
  15. Benelli G, Lukehart CM (2017) Special issue: applications of green-synthesized nanoparticles in pharmacology, parasitology and entomology. J Clust Sci 28:1–2Google Scholar
  16. Benelli G, Mehlhorn H (2016) Declining malaria, rising dengue and Zika virus: insights for mosquito vector control. Parasitol Res 115:1747–1754CrossRefGoogle Scholar
  17. Benelli G, Pavela R, Iannarelli R, Petrelli R, Cappellacci L, Cianfaglione K, Afshar FH, Nicoletti M, Canale A, Maggi F (2017b) Synergized mixtures of Apiaceae essential oils and related plant-borne compounds: larvicidal effectiveness on the filariasis vector Culex quinquefasciatus say. Ind Crop Prod 96:186–195CrossRefGoogle Scholar
  18. Benelli G, Pavela R, Maggi F, Petrelli R, Nicoletti M (2017a) Commentary: making green pesticides greener? The potential of plant products for nanosynthesis and pest control. J Clust Sci 28:3–10Google Scholar
  19. Benelli G, Rajeswary M, Govindarajan M (2017c) Towards green oviposition deterrents? Effectiveness of Syzygium lanceolatum (Myrtaceae) essential oil against six mosquito vectors and impact on four aquatic biological control agents. Environ Sci Poll Res doi:  10.1007/s11356-016-8146-3
  20. Chowdhery HJ (2009) Orchid diversity in north–eastern states of India. J Orchid Soc India 23(1–2):19–42Google Scholar
  21. Deo PG, Hasan SB, Majumdar SK (1988) Toxicity and suitability of some insecticides for household use. Int Pest Control 30:118–129Google Scholar
  22. Dhanasekaran D, Thangaraj R (2013) Evaluation of larvicidal activity of biogenic nanoparticles against filariasis causing Culex mosquito vector. Asian Pac J Trop Dis 31:74–179Google Scholar
  23. Dhiman RC, Pahwa S, Dhillon GPS, Dash AP (2010) Climate change and threat of vector-borne diseases in India: are we prepared? Parasitol Res 106(4):763–773CrossRefGoogle Scholar
  24. Edison TJI, Sethuraman MG (2012) Instant green synthesis of silver nanoparticles using Terminalia chebula fruit extract and evaluation of their catalytic activity on reduction of methylene blue. Process Biochem 47:1351CrossRefGoogle Scholar
  25. Finney DJ (1971) Probit analysis. Cambridge University Press, London, pp 68–72Google Scholar
  26. Geethalakshmi R, Sarada DVL (2012) Gold and silver nanoparticles from Trianthema decandra: synthesis, characterization, and antimicrobial properties. Int J Nanomedicine 7:5375–5384CrossRefGoogle Scholar
  27. Govindarajan M (2009) Bioefficacy of Cassia fistula Linn. (Leguminosae) leaf extract against chikungunya vector, Aedes aegypti (Diptera: Culicidae). Eur Rev Med Pharmacol Sci l13:99–103Google Scholar
  28. Govindarajan M (2010) Larvicidal efficacy of Ficus benghalensis L. plant leaf extract against Culex quinquefasciatus say. Aedes aegypti L. and Anopheles stephensi L. (Diptera: Culicidae). Eur Rev Med Pharmacol Sci 14:107–111Google Scholar
  29. Govindarajan M, Benelli G (2016) Artemisia absinthium-borne compounds as novel larvicides: effectiveness against six mosquito vectors and acute toxicity on non-target aquatic organisms. Parasitol Res 115:4649–4661CrossRefGoogle Scholar
  30. Govindarajan M, Jebanesan A, Pushpanathan T, Samidurai K (2008) Studies on effect of Acalypha indica L. (Euphorbiaceae) leaf extracts on the malarial vector, Anopheles stephensi Liston (Diptera: Culicidae). Parasitol Res 103:691–695CrossRefGoogle Scholar
  31. Govindarajan M, Mathivanan T, Elumalai K, Krishnappa K, Anandan A (2011) Mosquito larvicidal, ovicidal and repellent properties of botanical extracts against Anopheles stephensi, Aedes aegypti and Culex quinquefasciatus (Diptera: Culicidae). Parasitol Res 109:353–367CrossRefGoogle Scholar
  32. Govindarajan M, Sivakumar R, Amsath A, Niraimathi S (2012) Larvicidal efficacy of botanical extracts against two important vector mosquitoes. Eur Rev Med Pharmacol Sci 16(3):386–392Google Scholar
  33. Govindarajan M, Sivakumar R (2014) Larvicidal, ovicidal, and adulticidal efficacy of Erythrina indica (lam.) (family: Fabaceae) against Anopheles stephensi, Aedes aegypti, and Culex quinquefasciatus (Diptera: Culicidae). Parasitol Res 113:777–791CrossRefGoogle Scholar
  34. Govindarajan M, Rajeswary M, Senthilmurugan S, Vijayan P, Alharbi NS, Kadaikunnan S, Khaled J M, Benelli G (2017) Larvicidal activity of the essential oil from Amomum subulatum Roxb. (Zingiberaceae) against Anopheles subpictus, Aedes albopictus and Culex tritaeniorhynchus (Diptera: Culicidae), and non-target impact on four mosquito natural enemies. Physiol Mol Plant Path doi: 10.1016/j.pmpp.2017.01.003
  35. Kathiravan V, Ravi S, Ashokkumar S (2014) Synthesis of silver nanoparticles from Melia dubia leaf extract and their in vitro anticancer activity. Spectrochim Acta Part A Mol Biomol Spectrosc 130:116–121CrossRefGoogle Scholar
  36. Khan M, Khan ST, Khan M, Adil SF, Musarrat J, AlKhedhairy AA, Alkhathlan HZ (2014) Antibacterial properties of silver nanoparticles synthesized using Pulicaria glutinosa plant extract as a green bioreductant. Int J Nanomedicine 9:3551–3565Google Scholar
  37. Krishnaraj C, Jagan EG, Rajasekar S, Selvakumar P, Kalaichelvan PT, Mohan N (2010) Synthesis of silver nanoparticles using Acalypha indica leaf extracts and its antibacterial activity against water borne pathogens. Colloids Surf B Biointerfaces 76:50–56CrossRefGoogle Scholar
  38. Kumar PPNV, Pammi SVN, Kollu P, Satyanarayana KVV, Shameem U (2014) Green synthesis and characterization of silver nanoparticles using Boerhaavia diffusa plant extract and their antibacterial activity. Ind Crop Prod 52:562–566CrossRefGoogle Scholar
  39. Kumar R, Roopan SM, Prabhakarn A, Khanna VG, Chakroborty S (2012) Agricultural waste Annona squamosa peel extract: biosynthesis of silver nanoparticles. Spectrochim Acta A 90:173–176CrossRefGoogle Scholar
  40. Mabberley DJ (2008) Mabberley’s plant book: a portable dictionary of plants, their classification and uses, vol 3. Cambridge University Press, Cambridge, p 606Google Scholar
  41. Madhumitha G, Elango G, Roopan SM (2015) Bio-functionalized doped silver nanoparticles and its antimicrobial studies. J Sol-Gel Sci Technol 73:476–483CrossRefGoogle Scholar
  42. Maity D, Bain MK, Bhowmick B, Sarkar J, Saha S, Acharya K, Chakraborty M, Chattopadhyay D (2011) In situ synthesis, characterization, and antimicrobial activity of silver nanoparticles using water soluble polymer. J Appl Polym Sci 122:2189–2196CrossRefGoogle Scholar
  43. Maity D, Mollick MMR, Mondal D, Bhowmick B, Bain MK, Bankura K, Sarkar J, Acharya K, Chattopadhyay D (2012) Synthesis of methylcellulose–silver nanocomposite and investigation of mechanical and antimicrobial properties. Carbohydr Polym 90:1818–1825CrossRefGoogle Scholar
  44. Maridass M, Zahir Hussain MI, Raju G (2008) Phytochemical survey of orchids in the Tirunelveli Hills of South India. Ethnobotanical Leaflets 12:705–712Google Scholar
  45. Medhi RP, Chakrabarti S (2009) Traditional knowledge of NE people on conservation of wild orchids. Indian J Tradn Knowl 8:11–16Google Scholar
  46. Mishra A, Sardar M (2012) Alpha-amylase mediated synthesis of silver nanoparticles. Sci Adv Mater 4:143–146CrossRefGoogle Scholar
  47. Mohammad HM (2011) Therapeutic orchids: traditional uses and recent advances—an overview. Fitoterapia 82:102–140CrossRefGoogle Scholar
  48. Murugan K, Benelli G, Ayyappan S, Dinesh D, Panneerselvam C, Nicoletti M, Hwang J-S, Kumar PM, Subramaniam J, Suresh U (2015) Toxicity of seaweed-synthesized silver nanoparticles against the filariasis vector Culex quinquefasciatus and its impact on predation efficiency of the cyclopoid crustacean Mesocyclops longisetus. Parasitol Res 114(6):2243–2253Google Scholar
  49. Murugan K, Panneerselvam C, Samidoss CM, Madhiyazhagan P, Roni M, Subramaniam J, Dinesh D, Rajaganesh R, Paulpandi M, Wei H, Aziz AT, Alsalhi MS, Devanesan S, Nicoletti M, Pavela R, Canale A, Benelli G (2016) In vivo and in vitro effectiveness of Azadirachta indica-synthesized silver nanocrystals against Plasmodium berghei and Plasmodium falciparum, and their potential against malaria mosquitoes. Res Vet Sci 106:14–22CrossRefGoogle Scholar
  50. Naik BR, Gowreeswari GS, Singh Y, Satyavathi R, Daravath RR, Reddy PR (2014) Bio-synthesis of silver nanoparticles from leaf extract of Pongamia pinnata as an effective larvicide on dengue vector Aedes albopictus (Skuse) (Diptera: Culicidae). Adv Entomol:293–101Google Scholar
  51. Naqqash MN, Gökçe A, Bakhsh A, Salim M (2016) Insecticide resistance and its molecular basis in urban insect pests. Parasitol Res 115:1363–1373CrossRefGoogle Scholar
  52. Nazeruddin GM, Prasad NR, Waghmare SR, Garadkar KM, Mulla IS (2014) Extracellular biosynthesis of silver nanoparticle using Azadirachta indica leaf extract and its anti-microbial activity. J Alloys Compd 583:272–277CrossRefGoogle Scholar
  53. Pavela R (2015) Essential oils for the development of eco-friendly mosquito larvicides: a review. Ind Crop Prod 76:174–187CrossRefGoogle Scholar
  54. Pavela R, Benelli G (2016a) Ethnobotanical knowledge on botanical repellents employed in the African region against mosquito vectors—a review. Exp Parasitol 167:103–108CrossRefGoogle Scholar
  55. Pavela R, Benelli G (2016b) Essential oils as eco-friendly biopesticides? Challenges and constraints. Tr Plant Sci 21(12):1000–1007CrossRefGoogle Scholar
  56. Perni S, Hakala V, Prokopovich P (2014) Biogenic synthesis of antimicrobial silver nanoparticles capped with L-cysteine. Colloids Surf A Physicochem Eng Asp 460:219–224CrossRefGoogle Scholar
  57. Prakash P, Gnanaprakasama P, Emmanuel R, Arokiyaraj S, Saravanan M (2013) Green synthesis of silver nanoparticles from leaf extract of Mimusops elengi, Linn. for enhanced antibacterial activity against multi drug resistant clinical isolates. Colloids Surf B Biointerfaces 108:255–259CrossRefGoogle Scholar
  58. Prathna TC, Chandrasekaran N, Raichur AM, Mukherjee A (2011) Biomimetic synthesis of silver nanoparticles by Citrus limon (lemon) aqueous extract and theoretical prediction of particle size. Colloids Surf B Biointerfaces 82:152–159CrossRefGoogle Scholar
  59. Rafiuddin ZZ (2013) Bio-conjugated silver nanoparticles from Ocimum sanctum and role of cetyltrimethyl ammonium bromide. Colloids Surf B Biointerfaces 108:90–94CrossRefGoogle Scholar
  60. Rajan R, Chandran K, Harper SL, Yun SI, Kalaichelvan PT (2015) Plant extract synthesized nanoparticles: an ongoing source of novel biocompatible materials. Ind Crop Prod 70:356–373CrossRefGoogle Scholar
  61. Rajeswary M, Govindarajan M (2013) Mosquito larvicidal and phytochemical properties of Ageratina adenophora (Asteraceae) against three important mosquitoes. J Vector Borne Dis:50141–50143Google Scholar
  62. Raman N, Sudharsan S, Veerakumar V, Pravin N, Vithiya K (2012) Pithecellobium dulce mediated extra-cellular green synthesis of larvicidal silver nanoparticles. Spectrochim Acta Mol Biomol Spectrosc 96:1031–1037CrossRefGoogle Scholar
  63. Rastogi L, Arunachalam J (2011) Sunlight based irradiation strategy for rapid green synthesis of highly stable silver nanoparticles using aqueous garlic (Allium sativum) extract and their antibacterial potential. Mater Chem Phys 129:558–563CrossRefGoogle Scholar
  64. Roni M, Murugan K, Panneerselvam C, Subramaniam J, Hwang JS (2013) Evaluation of leaf aqueous extract and synthesized silver nanoparticles using Nerium oleander against Anopheles stephensi (Diptera: Culicidae). Parasitol Res 112:981–990CrossRefGoogle Scholar
  65. Roopan RSM, Madhumitha G, Rahuman AA, Kamaraj C, Bharathi A, Surendra TV (2013) Low-cost and eco-friendly phyto-synthesis of silver nanoparticles using Cocos nucifera coir extract and its larvicidal activity. Ind Crop Prod 43:631–635CrossRefGoogle Scholar
  66. Sathiya CK, Akilandeswari S (2014) Fabrication and characterization of silver nanoparticles using Delonix elata leaf broth. Spectrochim Acta Mol Biomol Spectrosc 128:337–341CrossRefGoogle Scholar
  67. Singh A, Sanjiv D (2009) Medicinal orchids—an overview. Ethnobotanical Leaflets 13:399–412Google Scholar
  68. Sivagnaname N, Kalyanasundaram M (2004) Laboratory evaluation of methanolic extract of Atlantia monophylla (family: Rutaceae) against immature stages of mosquitoes and non-target organisms. Mem Inst Oswaldo Cruz 99:115–118CrossRefGoogle Scholar
  69. Suganya A, Murugan K, Kovendan K, Mahesh Kumar P, Hwang JS (2013) Green synthesis of silver nanoparticles using Murraya koenigii leaf extract against Anopheles stephensi and Aedes aegypti. Parasitol Res 112:1385–1397CrossRefGoogle Scholar
  70. Suganya G, Karthi S, Shivakumar MS (2014) Larvicidal potential of silver nanoparticles synthesized from Lucas aspera leaf extracts against dengue vector Aedes aegypti. Parasitol Res 113:1673–1679CrossRefGoogle Scholar
  71. Suman TY, Rajasree SRR, Kanchana A, Elizabeth SB (2013) Biosynthesis, characterization and cytotoxic effect of plant mediated silver nanoparticles using Morinda citrifolia root extract. Colloids Surf B Biointerfaces 106:74–78CrossRefGoogle Scholar
  72. Tamboli DP, Lee DS (2013) Mechanistic antimicrobial approach of extracellularly synthesised silver nanoparticles against gram positive and gram negative bacteria. J Hazard Mater 260:878–884CrossRefGoogle Scholar
  73. Tamuly C, Hazarikaa M, Borah SCH, Das MR, Boruah MP (2013) In situ biosynthesis of Ag, Au and bimetallic nanoparticles using Piper pedicellatum C.DC: green chemistry approach. Colloids Surf B Biointerfaces 102:627–634CrossRefGoogle Scholar
  74. Vijayakumar M, Priya K, Nancy FT, Noorlidah A, Ahmed ABA (2013) Biosynthesis, characterisation and anti-bacterial effect of plant-mediated silver nanoparticles using Artemisia nilagirica. Ind Crop Prod 41:235–240CrossRefGoogle Scholar
  75. WHO (1980) Expert committee on diabetes mellitus. Second report. Technical report series 646. World Health Organization, Geneva, pp 12–15Google Scholar
  76. World Health Organization (2005) Guidelines for laboratory and field testing of mosquito larvicides. Communicable disease control, prevention and eradication, WHO pesticide evaluation scheme. WHO, Geneva, WHO/CDS/WHOPES/GCDPP/1.3Google Scholar
  77. World Health Organization (2014) A global brief on vector-borne diseases. WHO/DCO/WHD/2014.1Google Scholar

Copyright information

© Springer-Verlag Berlin Heidelberg 2017

Authors and Affiliations

  1. 1.Department of ZoologyThiru. Vi. Ka. Government Arts CollegeTiruvarurIndia
  2. 2.Unit of Vector Control, Phytochemistry and Nanotechnology, Department of ZoologyAnnamalai UniversityAnnamalainagarIndia
  3. 3.Department of Botany and Microbiology, College of ScienceKing Saud UniversityRiyadhSaudi Arabia
  4. 4.Department of Pharmacognosy, College of PharmacyKing Saud UniversityRiyadhSaudi Arabia
  5. 5.Department of Agriculture, Food and EnvironmentUniversity of PisaPisaItaly

Personalised recommendations