CONTACT : P.S LAMA patienncesunday@gmail.com 179 Abstract Over the years, the use of agrochemicals has been one of the reasons for resistance of mosquitoes to insecticides, a nega tive trend that has bee n on the increase. This study was carried out to determine the effect of Azadirachta indica (neem) see d extract on the larvae and adult Anopheles mosquitoes in Nasarawa Local Government Area of Nasarawa state as a possible alternative to already existing insecticide against the parasite. The research was carried out at the insectary laboratory of Nasarawa state university Keffi. Impregnated papers of neem seed oil were used for adult Anopheles mosquitoe s and the larviciding effect was also determine d. Adult Anopheles mosquitoe s expose d to impregnated papers of neem seed extract at differe nt conce ntrations of 20%, 40%, 80% and 100% showed 5%, 29%, 66% and 91% respectively. M ortality rate was recorded at 24h of e xposure w ith LC50 (Lethal conce ntration) of 4.45ml. The larvae were also exposed to different conce ntration of neem see d extract of 1%, 4%, 8% and 10% for 72h and the mortality rate was 100% with LC50 value (Lethal conce ntration) of 4.16ml. This implie d that adult Anopheles mosquitoe s will required more of the extract to achieve 100% mortality than larvae. Using Azadirachta indica (neem) seed extract as an anti-mosquito agent was more effective on the larvae of Anophe les mosquitoe s than the adult Anopheles mosquitoes. ISSN : 2580-2410 eISSN : 2580-2119 Effects of Azadirachta indica (Neem) seed extract on larvae and adult Anopheles mosquitoes in Nasarawa Local Government Area, Nasarawa State Nigeria. P.S. Lama1*, M.D. Olayinka1, A. Ezekiel1, J.D.C Tongjura1, R.J Ombugadu1, O Akyengo2 1 Department of Zool ogy, Faculty of Natural and Applied Sciences, Nasarawa State University Keffi, Nasarawa state Nigeria. 2 Department of Science Laboratory Technology, Nigerian Institute of Leather and Science Technology, Samaru, Zaria, Nigeria. Introduction Malaria is one of the major causes of mortality and morbidity in Nigeria ( Morakinyo et al., 2018; WHO 2018). WHO has rated Nigeria 92% in African Region where Nigeria accounted for 25% of the global burden (WHO2018). Mosquitoes are not just the vectors of malaria but the vectors of several diseases (An et al., 2020) with Anopheles gambiae giles also known as Africa mosquito been the most common vector of human malaria in Afro Tropical Region (CDC, 2010). In recent years, research has been going on where botanical metabolites are increasingly realized as potential substitute for chemical insecticides (Vivekanandhan et al., 2018). OPEN ACCESS International Journal of Applied Biology Keyword Anopheles mosquitoes; Azadirachta indica; Lethal concentration; Nasarawa. Article History Received August 24, 2022 Accepted December 14, 2022 International Journal of Applied Biology is licensed under a Creative Commons Attribution 4.0 International License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly c ited. mailto:patienncesunday@gmail.com International Journal of Applied Biology, 6(2), 2022 180 The neem plants belong to the family of Meliaceae plants which contain so many varieties of compound tha t shows insecticidal, antifeedant, growth-regulating, and development-modifying properties (Nugroho et al., 1999; Greger et al., 2001; D'Ambrosio et al.,, 2002; Nakatani et al., 2004). Meliaazedarach L. and Azadirachtin indica (Sapindales: Meliaceae), commonly known as Chinaberry or Persian lilac tree, are deciduous trees that are native to north western India; and have long been recognized f or their insecticidal properties. These trees grow typically in tropical and subtropical parts of Asia, but nowadays they are also cultivated in other warm regions of the world because of their considerable climatic tolerance. Fruit extracts of Meliaazedarach and Azadirachta indica elicit a variety of effects in insects such as anti feedant, growth retardation, reduced fecundity, moulting disorders, morphogenetic defects, and changes of behaviour (Schmidt et al., 1998; Abou et al., 2001; Gajmer et al., 2002; Banchio et al., 2003; Wandscheer et al., 2004). Larval of Anopheles mosquitoes were exposed to undiluted extract of seed oil, leaf and bark of Aza dirachtin (neem) for 12 hours, with a high mortality rate to seed oil, leaf extract and less mortality to bark of Azadirachtin at Usman Danfodio University Sokoto (Aleiro,2003). Larvae of Anopheles mosquitoes were susceptible to the seed and leaf of Azadirachtin in Kogi state (Adobu,et al.,2018). Fathelrahman,(2017) also observed in Sinnar State -Sudan that Anopheles arabiensis were susceptible to the extract of leaf and seed of Azadirachtin with high susceptibility to the kernel seed. When applied to artificial water bodies in every two weeks within period of three months, emulsified neem oil have the same effect on larval mortality and adult density as the commonly synthetic insecticides used (Awad,et al.,2003). A study was also carried out using a neem oil formulation on third and fourth larvae stage of Anopheles gambiae and showed 50%inhibition of adult emergence at a concentration of 6ppm (Okumu, 2007).In a study using emulsified neem oil, it shows that within a three months, Anopheles larvae failed to develop resistance or change their susceptibility to the oil (Awad et al., 2003). Materials and Methods Study Area The study was carried out in Nasarawa Local Government Area of Nasarawa state which is located in the western senatorial zone with latitude 08°41ʹ20ʺ to 08°32ʹ10ʺ N and longitude 07°48ʹ43ʺ to 07°50ʹ00ʺ E. It has two seasons rainy seasons starts from April to October while dry season start from October to March. Sample collection and procedures The Anopheles mosquitoes larva were collected from their natural water habitat in some communities of Nasarawa Local Government Area using Entomological ladles into a well labelled container, they were transported the insectary laboratory, Department of Zoology Nasarawa State University Keffi. The Anopheles mosquitoes were reared to adult in a container in the laboratory, the larva were fed with biscuit and yeast while the adult were fed with sugar solution according to WHO, (1998) Procedure for WHO Method for Susceptibility of Neem Oil extract The susceptibility test was carried out using four tubes for each of the concentration (neem seed extract) used with two control tubes each. An aspirator was used to introduce 25 Anopheles mosquitoes into the impregnated tubes. A timer was used to record both the International Journal of Applied Biology, 6(2), 2022 181 number of dead a nd alive mosquitoes after 10 minutes for 1hour. Af ter which the mosquitoes were transfer into a recovery tubes and were fed for 24 hour and the mortality rate was also recorded. The mortality in the control tubes was recorded as well. (WHO, 1998). Procedure for Larviciding Distilled water was introduced into four different containers for each of the concentration of the oil used with two controls. A pipette was used to introduce 25 Anopheles larvae into the containers and different concentration of neem seed oil was introduce into the containers. Mortality was recorded at 60 minutes, 24hour, 48hour, and 72hour. Mortality in the control was also recorded. (WHO, 1998). Results Effects of Azadirachtin indica oil extract on adult Anopheles mosquitoes Result showed that a total of 400 adult Anopheles exposed to A. indica oil extract at different concentration f or 24h. Mortali ty was recorded at 10 minutes, 60 minutes and 24h post exposure as shown in Table 1. The percentage of adult Anopheles mosquitoes to 20%, 40%, 80% and 100% concentration of A. indica oil extract at 60 mins of exposure were 0, 20, 47 and 49 percentage mortality. While the mortality recorded for 24h post exposure were 5, 29, 66 and 91 respectively. Table 1. Effects of Azadirachtin indica oil extract on adult Anopheles mosquitoes Concentration of oil extract (%) No. of Anopheles mosquitoes exposed Mortality (%) 60(mins) 24h 20.0 100 0(0.0) 5(5.0) 40.0 100 20(20.0) 29(29.0) 80.0 100 47(47.0) 66(66.0) 100.0 100 49(49.0) 91(91.0) Control R1 25 0(0.0) 1(0.25) R2 25 0(0.0) 1(0.25) The larviciding effect of A. indica oil extract The larviciding effect of A. indica oil extract is as shown in Table 2 A total of 400 Anopheles larvae were exposed to different concentration of neem seed extract ( 1 %, 4%, 8% and 10%) for 24h, 48h and 72h. The percentage mortality of neem seed extract to larvae showed 86.5%, 98.8% and 100% mortality. While 2(1%) mortality in the control was also recorded. International Journal of Applied Biology, 6(2), 2022 182 Table 2: Larviciding effect of Azadirachta indica seed oil extract Concentration of oil extract (%) No. of Anopheles larval exposed Mortality % 24h 48h 72h 1 100 78(88.0) 95(95.0) 100(100.0) 4 100 85(85.0) 100(100.0) 100(100.0) 8 100 88(88.0) 100(100.0) 100(100.0) 10 100 95(95.0) 100(100.0) 100(100.0) Control R1 25 0(0.0) 0(0.0) 1(0.25) R2 25 0(0.0) 0(0.0) 1(0.25) Discussion Adult Anopheles mosquitoes exposed to different concentrati on of neem seed oil at 20%, 40%, 80% and 100% f or 60 mi ns and 24h post exposure mosquitoes were highly resistant at different dosage in all the concentra tion for 24h and at 100% they were less resistant. This may be as a result of the effect of agricultural chemicals that were used to control pests in which the neem oil will not be effective enough in the knockdown. This present study shows that adult Anopheles mosquitoes were resistant to neem seed oil and in previous study in Sudan, Fatherahman (2017) observed that A. aranbiasis were susceptible to neem seed oil. Larval of Anopheles mosquitoes were exposed to different concentration of neem seed oil at 1%, 4%, 8% and 10% for 60 mins and 24h post exposure. In this research, the larval of Anopheles mosquitoes were susceptible to neem seed oil, this is because neem seed oil can cause deterrence in insect, repellency, growth disruption, reduced fitness and sterility activities. This confirmed the previous study in Sokoto by Aleiro (2003) where larval of Anopheles mosquitoes were exposed to undilute d extract of neem s eed oil and susceptibility was obtained. It was also observed by Adobu et al.,(2018) in Kogi State where larval of Anopheles mosquitoes were 100% susceptible to neem seed oil. (Kela et al., 2019) uses neem oil water dispersible tablet against the third instar larvae of Anopheles mosquitoes and its showed 98% mortality. (Assalif et al., 2016) also observed that neem powder applied on 1st, 2nd and 3rd instar larvae showed88.9%, 87.9% and 79.4% mortality. The mortality was very high at the first instar larval stage. Conclusion From this present study, neem seed extract showed efficacy as an anti -malaria agent by killing the mosquitoes subjected to it. Its efficacy was however more pronounced on the Anopheles larvae than the adult Anopheles mosquitoes. Therefore the use of neem seed extract should be encouraged in the control of mosquitoes but for a more appreciable result, the mosquito larvae should be targeted when using the neem seed extract. Competing interests The authors wish to declare that there are no competing interests. Funding No external funding was received. 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