Cowpea (Vigna unguiculata (L.) Walp.) fits into the family of crops known as Fabaceae from the genus referred to as Vigna notably has a black eye peas and institutes one of the most significant food legumes especially in the subtropics and ropical countries of the world[1] which are ordinarilyeaten in the form of dry grains [2]. Dried grains are primed into moi-moi, akara or eaten in combination with other different crops such as yam or even rice. They can as well be rummage-sale as silage[3]. Approximately 200 million people of Africa consume the crop [4]. Furthermore, cowpea is a relatively cheap source of protein and invariably obliges as the source of plant protein to peasant farmers that cannot have enough money tobuy meat and fish as animal protein for consumption[4].

Cowpea is cultivated in North and South America, tropical Africa and Asia particularly as grain, but also as a fodder crop or even vegetable. The crop can adapt widely and tolerance several farm stresses which makes it a very imperative food source for both man and animal. The estimation of the crop serving as a major protein source for more than 200 million people in sub-Saharan Africa which brands it the ten fresh vegetables in the People’s Republic of China Organization for Economic Co-operation and Development [5].

Unfortunately, the crop is extensivelyre-counted to be confronted by an array of insect pests and diseases that challenges plants usuability as reported by [6 and 7]. Cowpea cultivation has equally received a report that more than 130 species exists which causes economic down turn in cowpea yield as reported by [8]. Thisis due to the presence of diverse pests in Northern Nigeria in all stages of plant growth, and losses due to pest attacks or diseases can be as high as 90 percent in broad-spectrum. Correspondingly, Callosobruchus maculatus causes a considerable amount of loss on stored cowpeasboth especially in the Sub-tropics and the tropics as reported by [9]. Additionally, up to 100% of seeds may be infested and damaged by this pest in 3-4 months of storage[6]. The cowpea weevil, C. maculatuswhich is a major pest of cowpea lays their eggs on cowpea pods or on avenues left by other biting and chewing insects likely (Mylabris spp). C. maculatus remains the most destructive amongst the insect pests of stored cowpea seeds which is equally known as cowpea seed bruchid or pulse beetle [6]. C.maculatus has been reported to be the most serious pest of stored legumes in most of tropical countries, thus a serious pest of many crops [10]. Callosobruchus also spasms wide range of leguminous crops that include but not limited to cowpea, chick pea, and many other legumeinious plants[5]. The larva of this pest feeds on the seed thereby cause a serious damage to them [11]. Whereas the egg and adult stage are found on the grain, the larval and pupal which lives inside the grain causes the damages therein.The use of chemical insecticides which could be by fumigation is an appropriate way of controlling C. maculatus and other insects [10]. Insecticides which have quick knockdown action are efficient persistent and effective means of control and management of the insect but some of them haveundesirableeffects to man and animal. These chemicals pose hazards to the health of man and livestock, they also result in pest resistance and possible resurgence. They could also penetrate into grains, become toxic to the consumer and exhibit casinogenic effects in mammals [3]. Due to the outlined consequences, there is need to develop an economical, harmless and tranquil method of preserving stored products especially cowpea against C. maculatus. Hence, there isan urgentneed for alternateways and means of control that is both parsimoniouslypracticable and ecologicallypleasant to the populace.Several researchers and canvassers haveindependently documented the use of plant constituents as biopesticides as an unconventional to usage of chemicals[3, 12, 13, 14, 15 and 16]. Apart from the fact that biospesticide usage are readily available, they are equally biodegradable and had non toxic effects to non-target organisms. They are correspondingly selective in actionand capable of retarding the development of resistance over long time and duration of usage. Leaves, seeds, root and stems are different parts of plants that are effective for this purpose as reported by [17]. Theseplant fragments have to be washed and one or the other air or shade-dried, ground and subsequently sieved into powderbeforehand being put into treatment.The use of these biopesticides would go a long way in helping farmers to sustain their livelihood, reduceover-dependence on insecticides and maintain a polluted-free environment thereby promoting healthy living[18] which is the aim of this study.

MATERIALS AND METHODS

3.1 Study Area

The study was undertaken in the Laboratory of the Department of Parasitology and Entomology, Faculty of Biological Sciences of Nnamdi Azikiwe University Awkawith geographical co-ordinates that lies between  Longitude 6⁰14’N, 6⁰14.5’N and Latitude 7⁰8.6’E, 7⁰9’EAnambra State, Southeastern region of Nigeria.

3.2 Study Design

The research study design used was a Completely Randomized Design (CRD) involving four treatments and a control with each treatment replicated three times including the control on the laboratory bench.

3.3 Experimental Cowpea Seed

Cowpea grains infested with Callosobruchus maculatuswere procured from the prominent Eke Awka market in Anambra State, Southeast Nigeria which was used to set up the experimental insect culture in a plastic jar. Thepurchased grains were sterilized in the oven at the laboratory for three hours to ensure that the grains were devoid of infestation prior to use. The jar was perforated at the top to allow air, covered with muslin cloth, and kept under ambient temperature and relative humidity for proper development and reproduction of insects.

3.4 Collection, Identification, and processing of Moringa oliefera

The Moringa leaves and seeds were collected from a local household Moringa plant stand. The plants were taken to the Botany Department Laboratory of Nnamdi Azikiwe University to obtain the identity and further substantiation by a plant taxonomist.They were air dried and then ground to fine powder using an electric blender andthen filtered through a 0.5mm mesh before the powders were collected into a clean container.

3.5 Preparation of Plant Extracts

The pulverized plant samples (100 g each) were dissolved in 800 ml of 70% methanol and allowed to stand for 24hrs. The dissolved solute was sieved through a muslin fabric with Whatman No. 1 filter paper. Whereas the methanol filtrate obtained was concentrated using a water bath at 50°C, the solution was stockpiled at 4°C after amassing into apasteurised bottle until its necessitated. Serial dilution of 5 %, 10%, 25%, 50% of each plant extract were prepared and used for the bioassay. Control treatments were also prepared using acetone and these were kept under ambient conditions in the laboratory.

3.6 Contact toxicity response of C. maculatus on Moringa oleifera leaf and seed extract.

No. 1 Whatman filter paper (9 cm in diameter) was positioned in each of the petri dishes used for the experimentation. The numerous dosage echelons of the plant extracts rummage-scaleencompassd 5%, 10%, 25%, 50% and every one was simulatedthrice including the control. Aliquots of 2 ml of every dosage was uniformly distributed in the filter paper and leftwardfor an hour to guaranteeappropriatedissemination of the mixture. Controls with acetone only were included which was allowed to evaporate. Subsequently, 10 unsexedC.maculatusadults were presented into each petri dish containing the pickled filter paper and the control respectively. The petri dishes were enclosedby means of lid to avert the flight of the insects. Afterwards, adult mortalitywas taken every 24 hours for four days.

3.7 Analysis of Data

All assembled data were imperilled to analysis of variance (ANOVA) by means of SPSS version 20. Mortality data was analyzed using log-probit regression for determining LD₅₀ and LD₉₀ (LT₅₀) and LT₉₀). Treatments with significant differences were measured up at 5% level of significance (P<0.05) withthe Duncan multiple range test. The mortality data obtained were corrected using [19] formula.

Where Po= observed percentage mortality

Pc=control percentage mortality

Pm= corrected percentage mortality

The corrected percentage mortality values were transformed into probit. Dosages were also transformed into log dose. The probit values were plotted against log dose to determine LC₅₀ andLC₉₀as well as LT₅₀ and LT₉₀.

Results

4.1 Contact toxicity of Moringa oleifera seed and leaf extract on Callosobruchus maculatus

4.1.1. Mortality response of Moringa oleifera seed extracts on Callosobruchus maculatus

The contact toxicity result in (Table 1) showed there was dose-dependent mortalityresponse to the  Moringa oleifera seed extract. Mortality increasedin accordance with the increase in concentration of the extract. The highest and lowest meanmortality of Callosobruchus maculatus at 50% and 5% concentrations, mortalities of 53.33%  and 33.33% respectively. The statistical analysis showed thatdoses were significantly different from each other (P ≤ 0.05) .

Table 1: Toxicity effect of Moringa oleifera seed extract on C.maculatus after periods of exposure

4.1.2 Mortality response of Moringa oleifera leaf extract on Callosobruchus maculatus.

The contact toxicity result in (Table 2) showed there was dose dependent mortality response to the  Moringa oleifera leaf extract. Mortality increased in accordance with increase in concentration of the extract. The highest and lowest mean mortality of Callosobruchusmaculatus at 50% and 5% concentrations, caused mortalities of 100 % and 36.66% respectively. The statistical analysis showed that doses were significantly different from each other (P ≤ 0.05).

4.2 LC₅₀ of Moringa oleifera seed and leaf extract at different exposure times.

Table 3 shows the LC₅₀ of Moringa oleifera seed and leaf extract at different exposuretimes. It reveals that the highest toxicity occurs at 48.96 concentration in 48 hours and thelowest toxicity occurs at 30.62 concentration in 96 hours for the leaf extract while thehighest and lowest toxicity for the seed extract occurs at 19.40 concentration in 24 thoursand 10.63 concentration in 96 hours.

4.3 LC₉₀ of Moringa oleifera seed and leaf extract at different exposure times

Table 4 shows the LC of Moringa oleifera seed and leaf extract at different exposure umes. It reveals that the highest toxicity occurs at 34.79% concentration in 48 hours and the lowest toxicity occurs at 33.74% concentration in 96 hours for the leaf extract while the highest and lowest toxicity for the seed extract occurs at 88.95% concentration in 24 hours and 60.02%

shows that mortality increases with increase in exposure time. It reveals that at a lowest possible concentration (5%), the Moringa seed extract causes 50% mortality in 153 hours and 90% mortality within 265 hours. While, the leaf extract causes 50% mortality within 105 hours and 90% mortality within 187 hours

Discussion

The current study focused on enlightment of how bio-insecticides could be used as alternatives to synthetics in providing high level of control for Callosobruchus maculatus without environmental contamination endangeringman and animal. It has formally been recommended that an integrated pest management (IPM) strategy to safeguard cowpeas in storage systems could be accomplishedexploiting traditional methods as reported by [20]. For a stretchedwhile now, to safeguard their harvest from insects, African farmers have familiarized aromatic plants among stored seeds or pods. These plants dischargefickle compounds that are thought to possess insecticidal competencesas as reported by [21].

The study showed aconsiderable susceptibility of C. maculatus to two botanicals; Moringa oliefera seed extract and leaf extract. Mortalities of adult C.maculatus were observed in the direct method application of both plant extracts. The leaf extract was more effective, causing mortality of 100% within 24hrs, unlike the seed extracts which caused 46.67% after 24 hours. This is in line with the report of [22], whose results show that Moringa leaf powder was more effective than other extracts from different part of the plant apart from the flowers. The Moringa leaf extract caused 53.3% mortality after 72 hrs while that of the seed caused 50% mortality after 72 hours also. Both plant extracts also caused 36.67% and 23.33% mortality respectively at 48 hrs. This also was in concordance with the report from [23] which shows that mortality increases with an increase in exposure time. Invariably, it demonstrates the importance of timing in application of botanicals.

The results also showed that the toxicity of both the seed and the leaf extract of Moringa on C.maculatus is dependent on the level of concentration. It was observed that the higher the concentration the higher the toxicity and vice versa. The percentage mortality recorded at 24 hours at 5% and 25% concentration for the leaf extract were 6.67% and 46.67% respectively while that of the seed were 13% and 40% respectively.This is a pointer that the insecticidal properties of botanical extracts, such as pyrethrins and neem, can disrupt the life cycle of stored product pests. Neem-based formulations have been proven effective against C. maculatus larvae and their corresponding adults as reported by[24]. Moreover, biopesticides generally have low mammalian toxicity and pose minimal risks to human health. This is in stark contrast to synthetic pesticides, which often contain harmful chemicals. The use of biopesticides aligns with sustainable agricultural practices and reduces the environmental impact associated with chemical residues especially to man and animal. According to the reports of [25], seed treated with botanical oil extracts did not mislay their viability and also established the powder made from essential oil at different basics provided complete protection against C. maculatus and did not show a significant consequence on the intoxicating power of seed germination proportion. [26] reported that callosobruchus maculatus is one of the foremost pests of stored cowpea in the tropics whereas[23] narrated that when jute stackswere treated with dissimilar plant excerpts of Azaraditcha indica andVitex nemadowhich were then used for cowpea seed storage, it was established that the egg laying capacity by the C. Maculatusalongside the adult development and damage done to the seed were abridgedsignificantly.

Conclusion and Recommendation

Obviously,statistics from the studyillustrates that Moringa oleifera leaf and seed extract are effective in the control and management of Callasobruchus maculatus in stored cowpea. Fortunately, this extract can also be prepared easily by farmers and applied to grains both in the field and in the store. The level of its effectiveness shows that higher concentration is important to obtain better seed protection which implies that mortality increases with a corresponding increase in concentration. The use of these plant materials to a large extent reduces the rate of dependence on chemical pesticides which can be costly and as such not readily available and assessible. It also reduces the high level of pesticide residue, resistance, and toxicity to farmers and consumers of produce which can over time lead to health concerns.It is hereby recommended that further study should be carried out to find out theefficacy of Moringa oleiferaon the F1 generation of the test insect, including the loss and damage due to the insects. Excaustive study should also be carried out to ratifythe constituents of the plants and the active ingredient that makes it insecticidal,nonethelesslocal farmers should be informed on the necessityof botanicals usage as pesticides compared to chemical pesticides.

Author contributions: OEO, OIE and EOVwrote the first draft and edited the manuscript, ACO and EAC developed the protocol, and oversaw study implementation and monitoring. OEO,  and UJCdid the experimental design and OIE did the analysis of the data collected.

Acknowledgments: We are grateful and sincerely appreciate the market sellers for their assistance during the sample collection.

Funding: Not Applicable

Competing interest: The authors unanimous affirm that they do not have any skirmishes of concern.

Data availability: The statistics used to substantiate the discovieries of this study are readily accessible upon reasonable entreaty.

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