Budiman B, Ishak H, Stang S, Ibrahim E, Yudhastuti R, Maidin A, Naiem F, Wahid I, Mallongi A. Effectiveness of Clove Oil (Syzigium Aromaticum) as Biolarvacide of Aedes Aegypti. Biomed Pharmacol J 2022;15(4).

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Manuscript received on :15-10-2022
Manuscript accepted on :22-11-2022
Published online on: 07-12-2022

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Plagiarism Check: Yes
Reviewed by: Dr. Anand Ramasamy
Second Review by: Dr. Abeer Gatea
Final Approval by: Dr. H Fai Poon

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Budiman^1,2*, Hasanuddin Ishak², Stang², Erniwati Ibrahim², Ririh Yudhastuti³, Alimin Maidin², Furqaan Naiem², Isra Wahid⁴ and Anwar Mallongi²

¹Faculty of Public Health Muhammadiyah University, Palu, Indonesia

²Faculty of Public Health Hasanuddin University, Makassar, Indonesia

³Faculty of Public Health Airlangga University, Surabaya, Indonesia

⁴Faculty of Medicine Hasanuddin University, Makassar, Indonesia

Corresponding Author E-mail: budiman19k@student.unhas.ac.id

DOI : https://dx.doi.org/10.13005/bpj/2566

Abstract

Background: The vector control that is often done so far is by chemical control, which will continually have a negative impact. Therefore, it is necessary to control methods that are more friendly to the environment, effective and efficient, and safe for health, one of which is the use of essential oils from plants. Clove (Syzigium aromaticum) is one of the plants that can be used as an alternative larvicide to kill mosquito vectors because it contains many chemical compounds, including eugenol compounds, eugenol acetate, methyl eugenol, b-caryophyllene, methyl eugenol, saponins, flavonoids and larvicidal tannins.  Objective: This study aims to determine the effectiveness of clove leaf waste oil (Syzygium ) as a biolarvicide for the Aedes aegypti mosquito. Methods: The research method used is experimental with a Quasi-Experimental Design approach and Post-test Control Group Design. The object of this research is clove leaf waste oil (Syzigium aromaticum) that has been obtained from the distillation of dried clove leaf waste. The object of this research is the third-stage larvae and adult Aedes aegypti mosquitoes obtained by rearing or self-reproducing mosquitoes in the laboratory.  The bioassay test method used a glass test vessel with a volume of 200 ml of solution consisting of a mixture of water (aquades) with clove leaf waste oil. This study used a treatment concentration of 0.006%; 0.007%; 0.008%; 0.009% and 0.01%. The control used aquades (0%). Then put 25 Aedes aegypti larvae in the test container and then observe the number of mortality larvae during 24 hours of observation. Results: The results showed that clove (Syzygium aromaticum) leaf waste oil was effective as a biolarvicide with LC₅₀ at a concentration of 0.005% and Kruskal Wallis test value of 0.000 (P<0.05) which means that there is significant difference in the number of deaths of Aedes aegypti mosquito larvae at each concentration. Conclusion: Clove leaf waste oil (Syzygium aromaticum) is effective as a biolarvicide for Aedes aegypti mosquitoes.

Keywords

Aedes aegypti; Biolarvicides; Effectiveness; Syzygium aromaticum

Introduction

Vector disease is one of the diseases that pose a threat to public health worldwide ^1,2. Mosquitoes are an important and primary vector in the spread of disease and are highly contagious to humans³.

Each year around 50 million cases of dengue occur and around 500,000 patients are hospitalized, predominantly children^4,5,6. In 2019, regions of the Americas and 16 countries reported an increase in dengue cases at the national level, such as Antigua and Barbuda, Argentina, Brazil, Chile, Colombia, El Salvador, Guatemala, Guadeloupe, Guyana, Honduras, Jamaica, Martinique, Mexico, Paraguay, San Martin, and Venezuela. 99,998 cases of dengue were reported (incidence rate of 10.2 per 100,000 population), including 28 deaths. Of the total number of reported cases, 25,333 were examined in the laboratory and 632 were categorized as severe cases of dengue (0.63%)^7,8. Overall, all regions of Indonesia have high dengue cases (incidence rate of 78.0) and an increase in dengue cases spreading to cities and regencies in 34 provinces of Indonesia ⁹.

Vector mosquito control can be carried out mechanically, physically, biologically and environmentally, both in developing mosquitoes and in adult mosquitoes ^(10)(11)(12). Today, the control that is frequently carried out is chemical control using chemicals, including organophosphorus, organochlorine, carbamate and pyrethroid group compounds. However, the continued use of these chemicals will have negative impacts, such as the death of non-target organisms, environmental contamination, and danger to public health. In addition, this method is also costly and may cause resistance in mosquitoes ^10,11,12.

Various efforts have been made to prevent the emergence of resistance in mosquitoes to various chemicals. Therefore, it is necessary to control the methods, especially the insecticides that are more friendly to the environment, effective and efficient, and safe for health ¹³. Biological control can be an alternative to mosquito vector control by reducing the mosquito vector population and using natural materials ^14,15.

Natural larvicides/insecticides have been shown to make a significant contribution as a new alternative in an effort to reduce the number of diseases caused by mosquito vectors. The content of eugenol, alkaloids, flavonoids, saponins and other active principles of plants can be toxic for Aedes aegypti larvae. Plant essential oils can interfere with the metabolic, biochemical, physiological and behavioral processes of insects ¹⁶.

Clove plants (Syzigium aromaticum), which are widely cultivated in Indonesia, have potential to be used as alternative larvicides/insecticides to kill vector mosquitoes because they contain many chemical compounds reaching up to 40 chemical compounds, including eugenol compounds (87.24%), eugenyl acetate (5.8%), b-caryophyllene (3.85%), α-cadinol(2.43%), myrcene (1.84%), methyl eugenol (1.8% ) ^{17, 18,19, 20, 21, 22}.

Expert research shows that cloves (Syzigium aromaticum) have compounds that have the potential to be used as larvicides for Aedes aegypti and Culex quinquefasciatus mosquitoes with mortality rates greater than 85% ¹⁸. This study aims to determine the effectiveness of clove leaf waste oil (Syzygium aromaticum) as a bio larvicide for the Aedes aegypti mosquito.

Materials and methods

Types of Research

The research method used is experimental with a Quasi-Experimental Design approach and Post-test Control Group Design. This research was carried out for 3 months starting from the preparation of tools and materials including the process of rearing Aedes aegypti larvae and the process of making clove leaf waste oil (Syzigium aromaticum).

Acquisition of clove leaf waste oil

The clove leaf waste oil procurement process is carried out at the clove leaf waste oil refining factory located in Donggala regency, Central Sulawesi province. The distillation process requires 20 kg of dried clove leaf residue with a distillation time of 8 hours.

Collection of mosquito larvae

The process of collecting Aedes aegypti mosquito larvae is carried out by self-breeding (rearing) at the Donggala Health Research and Development Center Entomology Laboratory located in Labuan Village, Labuan Subdistrict, Donggala District, Central Sulawesi province. The stage used is stage III. The larvae used for the test should be normal and not exhibit a different body color from normal larvae. Larvae are classified as dead if they do not move after being touched with a sexy needle at the siphon (cervix), or continue to move but cannot reach the surface of the water or do not show a typical diving reaction when agitated Water.

Biolarvicide tests

Observation and laboratory test methods for biolarvicides refer to the WHO Standard Guidelines for Larvicide Testing ⁽²³) on mosquito larvicide testing. This test used 20 mg of abate powder (themefos) as a positive control and 200 ml of aquadest as a negative control. The number of test larvae used was 25 stage III and they were placed in a container with a volume of 200 ml of solution. The treatment used a dose of 0.006%; 0.007%; 0.008%; 0.009% and 0.01% and observations were made for 24 hours with 4 replications.

Statistic analysis

The results of the larval susceptibility test were grouped with the following larval mortality criteria ⁽²⁰⁾:

Mortality > 98% indicates a vulnerable species

80 – 98% mortality indicates a tolerant species

Mortality < 80% indicates resistant species

Data from all replicas must be collected for analysis. The results of the observation of dead larvae were analyzed by Probit analysis to determine the LC₅₀ value and Kruskal Wallis analysis using the SPSS software.

Ethical approval

This study was approved by the Health Research Ethics Committee of the Faculty of Public Health, Hasanuddin University, Makassar, Indonesia. Number: 3106/UN4.14.1/TP.02.02/2021.

Results

The results of testing the mortality rate (mortality) of Aedes aegypti larvae during 24 hours of observation can be seen in the following table and graph.

Table 1: Mortality Percentage of Aedes aegypti Larvae for 24 hours

Graph 1: Mortality Percentage of Aedes aegypti Larvae for 24 hours. Click here to view graph

Table 2:  Krusskall wallis test results Aedes aegypti larvae.

The table and graph above shows that the average mortality rate of Aedes aegypti mosquito larvae using clove leaf oil (Syzigium aromaticum) at a concentration of 0.006% was 77% (19.3 larvae), at a concentration of 0.007% it was 76% (19 larvae). Whereas at a concentration of 0.008% to 0.01%, all larvae tested experienced 100% kill.

The results of the probit analysis of the clove leaf (Syzigium aromaticum) used oil test on the mortality of Aedes aegypti mosquito larvae obtained an estimated LC₅₀ value of 0.005%. Clove (Syzigium aromaticum) leaf waste oil can kill 50% of Aedes aegypti larvae from concentrations of 0.004% to 0.006%.

The value obtained in the Kruskal Wallis test was P = 0.000, (P <0.05), so it can be concluded that there was a significant difference between the number of deaths of Aedes aegypti larvae and an increase in the total concentration of clove leaf waste oil (Syzigium aromaticum).

Discussion

According to the results of laboratory research, clove leaf waste oil (Syzigium aromaticum) has the ability to biolarvicide against the larvae of the Aedes aegypti mosquito. The test results showed an increase in mortality (mortality) of Aedes aegypti mosquito larvae along with increasing concentrations.

This study showed that the mortality rate of 50 percent of Aedes aegypti larvae (LC₅₀) at a concentration of 0.005% so the results of this study were more effective compared to previous studies where other studies ¹⁵ stated that essential oil from Syzigium aromaticum had an LC₅₀ at a concentration of 92.56 mg/ l (0.009256%) for Aedes aegypti larvae and LC₅₀ at a concentration of 124.42 mg/l (0.012442%) for Culex quinquefasciatus larvae for 24 hours. Furthermore, the research ²⁴ also found that the essential oil of Syzigium aromaticum can kill the larvae of Anopheles stephensi by 86.96%.

In principle, essential oils always float on the surface of the water (specific gravity lower than water) so they are effective as mosquito larvae larvicides, because mosquito larvae live in water and have the behavior of approaching or hang from the surface of the water to breathe²⁵.

The mortality of Aedes aegypti mosquito larvae is influenced by 2 factors, namely internal factors and external factors. Internal factors are the ability of the body of mosquito larvae or the resistance of the body to resist the influence of the given essential oil. While the external influence is the substance contained in the clove blade itself. The difference in the level of toxicity of a compound is determined by the chemical variation of the oil composition that will determine the bioactivity of the plant against mosquito larvae.

Clove (Syzigium aromaticum) has eugenol as its main component, which gives rise to a distinctive aroma. It has been reported to have high insecticidal and antimicrobial properties that have been included in many formulations to control insect pests and pathogens ^(25–27). Eugenol acts by affecting the nervous system of insects and can act as a contact poison, a stomach poison, a respiratory poison, and a neurotoxin that can cause death in mosquitoes ²⁶.

Conclusion

The results of this study indicate that clove (Syzigium aromaticum) leaf waste oil is effectively used as a biolarvicide for Aedes aegypti mosquito larvae. It is recommended that further research can examine the content of clove leaf waste oil to know which active substance can kill Aedes aegypti larvae.

Conflict of Interest 

There are no conflict of interest.

Funding Sources

There is no funding sources.

References

1. World Health Organization. Global Vector Control Response 2017-2030. Geneva; 2017. 64 p.
2. Kraemer MUG, Reiner RC, Brady OJ, Messina JP, Gilbert M, Pigott DM, et al. Past and future spread of the arbovirus vectors Aedes aegypti and Aedes albopictus. Nat Microbiol [Internet]. 2019;4(5):854–63. Available from: http://dx.doi.org/10.1038/s41564-019-0376-y
   CrossRef
3. Medeiros ES, Rodrigues IB, Litaiff-abreu E, Pinto ACS, Tadei WP. Larvicidal activity of clove ( Eugenia caryophyllata ) extracts and eugenol against Aedes aegypti and Anopheles darlingi. African J Biotechnol. 2013;12(8):836–40.
4. Guzman MG, Halstead SB, Artsob H, Buchy P, Farrar J, Nathan MB, et al. Europe PMC Funders Group Dengue : a continuing global threat Europe PMC Funders Author Manuscripts. Nat Rev Microbiol. 2010;8(12 0):7–16.
   CrossRef
5. Simmons CP, Farrar J, Chau VN van, Wills Bridget. Current Consepts Dengue. N Engl J Med. 2012;18(4):487–94.
6. Anderson KB, Chunsuttiwat S, Nisalak A, Mammen MP, Libraty DH, Rothman AL, et al. Burden of symptomatic dengue infection in children at primary school in Thailand: a prospective study. Lancet. 2007;369(9571):1452–9.
   CrossRef
7. Pan America Health Organization. Guidelines for Patient Care in The Region Americas. 2016.
8. Pan American Health Organization. Integrated Management Strategy for Dengue Prevention and Control in the Region of the Americas. Pan Am Heal Organ [Internet]. 2018; Available from: http://dx.doi.org/10.1590/S1020-49892007000100011
   CrossRef
9. CDC. The Centers for Disease Control and Prevention (CDC) in Indonesia. CDC. 2020;(Cdc).
10. Oduola AO, Olojede JB, Ashiegbu CO, Olufemi A, Otubanjo OA, Awolola TS. High level of DDT resistance in the malaria mosquito: Anopheles gambiae s.l. from rural, semi urban and urban communities in Nigeria. J Rural Trop Public Heal. 2010;9(Figure 1):114–120.
11. Roiz D, Wilson AL, Scott TW, Fonseca DM, Jourdain F, Müller P, et al. Integrated Aedes management for the control of Aedes-borne diseases. PLoS Negl Trop Dis. 2018;12(12):1–21.
    CrossRef
12. Idowu OA, Adeleke MA, Aina, T M. Evaluation of Indoor breeding activities of mosquitos during the dry season in abeokuta, Southwestern Nigeria. J Environ Heal Res. 2012;12(01):25–8.
13. Cavalcanti ESB, Morais SM de, Lima MAA, Santana EWP. Larvicidal activity of essential oils from Brazilian plants against Aedes aegypti L. Mem Inst Oswaldo Cruz. 2004;99(5):541–4.
    CrossRef
14. Krishnappa K, Elumalai K, Dhanasekaran S, Gokulakrishnan J. Larvicidal and repellent properties of adansonia digitata against medically important human malarial vector mosquito anopheles stephensi (Diptera: Culicidae). J Vector Borne Dis. 2012;49(2):86–90.
15. Budiman, Ishak H, Stang, Ibrahim E, Daud A, Amiruddin R. Essential oil as a new tool for larvicidal Aedes aegypti: A systematic review. Gac Sanit [Internet]. 2021;35:S459–62. Available from: https://doi.org/10.1016/j.gaceta.2021.10.074
    CrossRef
16. Maia MF, Moore SJ. Plant-based insect repellents: A review of their efficacy, development and testing. Malar J. 2011;10(SUPPL. 1):1–15.
    CrossRef
17. Mohammad Nazrul Islam Bhuiyan. Constituents of the essential oil from leaves and buds of clove (Syzigium caryophyllatum (L.) Alston). African J Pharm Pharmacol. 2012;6(16).
    CrossRef
18. Fayemiwo KA, Adeleke MA, Okoro OP, Awojide SH, Awoniyi IO. Larvicidal efficacies and chemical composition of essential oils of Pinus sylvestris and Syzygium aromaticum against mosquitoes. Asian Pac J Trop Biomed. 2014;4(1):30–4.
    CrossRef
19. Costa JGM, Rodriuges FFG, Silva MR, Mota ML, Santos NKA, Cardos ALH, et al. Artigo Aedes aegypti. 2005;15(4):304–9.
    CrossRef
20. Elzayyat E, Elleboudy N, Moustafa A, Ammar A. Insecticidal, Oxidative, and Genotoxic Activities of Syzygium aromaticum and Eucalyptus globulus on Culex pipiens Adults and Larvae. Turkiye parazitolojii Derg. 2018;42(3):213–22.
    CrossRef
21. Thomas A, Mazigo HD, Manjurano A, Morona D, Kweka EJ. Evaluation of active ingredients and larvicidal activity of clove and cinnamon essential oils against Anopheles gambiae (sensu lato). Parasites and Vectors. 2017;10(1):1–7.
    CrossRef
22. Faraco A, Ribeiro-Paes JT, de Deus JT, de Holanda Cavalcanti SC, Nunes R de S, Alves PB, et al. Larvicidal activity of Syzygium aromaticum (L.) Merr and Citrus sinensis (L.) Osbeck essential oils and their antagonistic effects with temephos in resistant populations of Aedes aegypti. Mem Inst Oswaldo Cruz. 2016;111(7):443–9.
    CrossRef
23. World Health Organization. Guidelines for laboratory and field testing of mosquito larvicides [Internet]. World Health Organization. 2005. 1–41 p. Available from: http://whqlibdoc.who.int/hq/2005/WHO_CDS_WHOPES_GCDPP_2005.13.pdf?ua=1
24. Osanloo M, Mehdi Sedaghat M, Esmaeili F, Amani A. Larvicidal Activity of Essential Oil of Syzygium aromaticum (Clove) in Com-parison with Its Major Constituent, Eugenol, against Anopheles stephensi. J Arthropod Borne Dis. 2019;12(December):361–9.
    CrossRef
25. Ansari MA, Mittal PK, Razdan RK, Sreehari U. Larvicidal and mosquito repellent activities of Pine (Pinus longifolia, Family: Pinaceae) oil. J Vector Borne Dis. 2005;42(3):95–9.
26. Yoo C Bin, Han KT, Cho KS, Ha J, Park HJ, Nam JH, et al. Eugenol isolated from the essential oil of Eugenia caryophyllata induces a reactive oxygen species-mediated apoptosis in HL-60 human promyelocytic leukemia cells. Cancer Lett. 2005;225(1):41–52.
    CrossRef
27. Bhat SK, Kempraj V. Biocidal potential of clove oils against Aedes albopictus – A comparative study. African J Biotechnol. 2009;8(24):6933–7.
28. Campbell, Neil A. 2004. Biologi. Edisi Kelima Jilid 3. Jakarta : Erlangga.
29. Pamungkas, Syafei, N.S. and Soeroto, A.Y. (2016) ‘Perbandingan Efek Larvasida Minyak Atsiri Daun Cengkeh (Syzygium aromaticum L.) Varietas Zanzibar dengan Temephos terhadap Larva Nyamuk Aedes aegypti’, Pharmaceutical Sciences and Research, 3(3), pp. 139–144. Available at: https://doi.org/10.7454/psr.v3i3.3566.
    CrossRef