Introduction

Mosquitoes are seen as a menace in different climes as they are the vectors of several pathogens alongside with their annoying sounds and bites. These pathogens cause various public health diseases such as dengue fever, malaria, filariasis encephalitis, chikungunya fever, West Nile virus and Zika virus among other disease conditions (Anupam et al. 2012). Anopheles gambiae is responsible for the transmission of malaria fever and lymphatic filariasis (WHO 2007; CDC 2019). One of the efforts aimed at controlling mosquitoes is the use of synthetic insecticides which today is seen as an unpopular choice because they are perceived not to be eco-friendly. For instance, the use of Dichlorodiphenyltrichloroethane (DDT), organophosphates, organochlorines have shown to be non-biodegradable and toxic to humans based on their persistence in the ecosystem (Mahmood et al. 2016). Research has proven that the use of synthetic pesticides or insecticides as the case may be in terms of their effects, has a cognate effect on non-targeted organisms by altering their conditions through interference with natural processes leading to a system deficiency which may lead to their death (Aktar et al. 2009; Abdel-Tawab 2016). The World Health Organisation (WHO) estimates that over 200,000 people die globally through synthetic pesticide exposure annually due to synthetic pesticide overuse (Belmain et al. 2013). Aside from the environmental problem associated with the use of synthetic pesticides (insecticides) is the issue of resistance and pest resurgence (Ojo 2016). Pest resistance implies that a particular pest is resistant to a specific pesticide rendering it ineffective in the control of the pest while pest resurgence emanates from scenario whereby pests become more virulent and difficult to control. Hence, the need to turn to the use of plants as insecticides which have been shown by researchers over the years to effectively control mosquitoes without causing the environment high cost in terms of preservation of the biodiversity and huge financial outlay.

Several researchers have demonstrated through their various works the pesticidal efficacy of plant materials and such plants could be referred to as botanicals. Their works have led to the development of numerous mosquito control agents such as ocimenone, rotenone, capllin, quassin, thymol, eugenol, neolignans, arborine and goniothalamin (Shaalan et al. 2005). Several species of plants have been shown to possess some mosquitocidal properties (Bekele 2018). A great number of these plant species were seriously investigated leading to the eventual characterization of their active constituents. The most notable of these constituents are alkaloids, terpenoids, steroids, phenols, saponins and tannins (Shaalan et al. 2005; Alqasoumi et al. 2012; Adesina and Rajashekar 2018). Hence, plants containing them are said to have insecticidal properties. Common plants such as neem, tobacco, pawpaw inter alia have been used as insecticides (Habibullah et al. 2007).

Petiveria alliacea L. is a perennial shrub/herb from the family Phytolaccaceae (García-González et al. 2006). The plant is a straight herbaceous plant that can grow up to 1 m tall. It has small white flowers that are bisexual, zygomorphic, which are slightly imbricate to rather remote (Ferrer 2007). The sepals are white or greenish to pinkish, linear-lanceolate to linear-oblong, 3.5–6 mm long with superior ovary (Nienaber and Thieret 2003). Its fruit forms a cuneiform berry with narrow oblong achenes, 6–8 mm long with four hooks turned downwards and the seeds are solitary, erect and linear (Alegre and Clavo 2007). Phytochemicals present in P. alliacea include terpenoids (triterpenoids) saponins, phenols (polyphenols), coumarins, benzaldehyde, benzoic acid, flavonoids, fredelinol, pinitol and allantonin with varying concentrations in the root, stems and leaves (Kubec et al. 2002, 2003). The GC-MS analysis of the ethanolic extract of the leaves revealed Phenol,2,4-bis(1,1)-dimethylethyl, 3,7,11,15-Tetramethyl-2-hexadecen-1-ol; Hexadecanoic acid, methyl ester, ethyl ester; 9,12,15-octadecatrienoic acid, ethyl ester; Stigmasterol; and Campesterol, n-Hexadecanoic acid, and Phytol as the major compound present (Abdul-Raheem et al. 2018). Petivera alliacea has been identified as a plant with medicinal, insecticidal and acaricidal properties (Nienaber and Thieret 2003; Gomes et al. 2005; Kim et al. 2006; Pérez-Leal et al. 2006; Williams et al. 2007; García-Mateos et al. 2007; Schmelzer and Gurib-Fakim 2008; Hernández et al. 2014; Hartmann et al. 2018).

Clerodendrum volubile P. Beauv is an indigenous plant that belongs to the family Lamiaceae (Verbenaceae). The plant is a green climbing shrub which can grow up to 3 m in height characterised with numerous flowers which are averagely 1.5 cm in length (Burkill 1985; Erukainure et al. 2011). Clerodendrum volubile is rich in several phytochemicals which include phenols, saponins, alkaloids and tannins (Erukainure et al. 2011). The gas chromatography-mass spectrometry (GC–MS) of its ethanolic crude extract revealed that oxirane, methyl 2-octylcyclopropene-1-heptanoate and Hexadecanoic acid, methyl ester were the top three major compounds as they accounted for 50.51%, 12.41% and 9.96% respectively (Molehin et al. 2017). Plants of the genus Clerodendrum which have been proven to be of high medicinal value (Erukainure et al. 2011, 2014). Research works on its insecticidal efficacies some members of the genus have been published (Patil et al. 2014; Muthu et al. 2015; Lurdu and Thampi 2017; Ileke and Adesina 2018). However, the insecticidal efficacy of C. volubile is yet to be reported. This study is aimed at determining the potency of C. volubile and P. alliacea ethanolic leaf extracts against different life stages of An. gambiae with oviposition deterrence, larvicidal, pupicidal, adulticidal and repellency potentials examined.

Methodology

Collection of plant materials

Defect-free fresh leaves of P. alliacea and C. volubile that were obtained from Ondo town (7.10° N, 4.84° E) in Ondo State, Nigeria. They were taken to the Crop, Soil and Pest Management Laboratory, Federal University of Technology, Akure, Ondo State for authentication. The leaves of the two plants were thoroughly cleaned with distilled water, air-dried in the laboratory for one month then pulverized into a fine powder using pestle and mortar. The resulting powder was stored inside dark bottles and refrigerated for preservation purpose until extraction procedure was carried out.

Plant active ingredient extraction

The cold maceration extraction method described by Udo (2011) was adapted in the study. Five hundred grams (500 g) of P. alliacea and C. volubile powders were soaked separately in an extraction bottle containing 1.5 l of absolute ethanol. The mixture was stirred occasionally with a glass rod and extraction was terminated after 3 days. The liquid filtrate was then concentrated with the aid of a rotary evaporator at a speed of 3 to 6 rpm, at temperatures of 30 to 40 °C for 8 h. The final extract is refrigerated at 4 °C until required.

Mosquito baits

The mosquito baiting method used for this study follows the ovitrap method described by Rawlins et al. (1998) with slight modification. Containers with 1 l of rainwater were sprinkled with 10 g of bakers yeast and then placed under a partially shaded area on an open field to serve as an attractant to wild mosquitoes to lay their eggs i.e. simulates the natural breeding ground of mosquitoes. After 2–5 days, the baits containing eggs/larva were then transferred to the laboratory. The eggs/larva were identified into species level using the morphological keys described by Gillies and De Meillon (1968) with the use of an Olympus stereo-dissecting microscope with model number VMT-2S-W. The An. gambiae larvae were then separated from the mixed culture and transferred into another plastic container containing distilled water, fed with bakers yeast and subsequently used or nurtured to the desired life stages (pupae and adults) as required by the different bioassays.

Concentrations preparation and experimental design

Two (2) grams of the crude extracts were dissolved in 1 l of solvent. From these stock solutions, different concentrations of 50 mg/l, 100 mg/l, 200 mg/l, 400 mg/l and 800 mg/l. The extract concentrations for oviposition deterrent, larvicidal and pupicidal bioassay were prepared using sterile distilled water as a solvent while 70% of ethanol was used for adulticidal and repellency bioassays. The experiment was carried out using a completely randomized design (CRD) under laboratory conditions.

Oviposition deterrent bioassay

The procedure for the oviposition deterrent of the plant extracts followed the ones described by Xue et al. (2001) and Fatima et al. (2011). An equal number (50) of a day-old male and female adults were placed inside a screen cage (45 X 45 X 30 cm) for mating purpose. They were provided with 10% sucrose solution soaked in cotton wool and placed in a plastic cup. After 4 days, an albino rat (Rattus norvegicus) with a shaved dorsal side (of about 20cm2 area) was used to feed the female mosquito. The rat with the help of wire mesh restrainer was placed on the cage with the shaved dorsal side towards it and left for about an hour in the absence of lightning. This was done to serve as blood source needed for oviposition by the female mosquito. About 50 ml of any given concentration of the plant extract was placed inside a plastic container that has filter paper serving as ovipositional surface. Three (3) cups were allotted to each concentration (50, 100, 200, 400, 800 mg/l) and control (water). The containers allotted to each concentration were arranged alternatively inside another screen cage of the same dimension to cancel out any effect on oviposition that may arise due to positioning. Ten gravid blood-fed female mosquitoes were then separated from the males and released inside the new cage containing the treatments where oviposition was observed. The test was replicated three times. The mosquitoes were sustained in conditions of 27° ± 2 °C and 75 ± 5% humidity. Egg deposition was observed and recorded after 24 h of egg-laying using a stereo-dissecting microscope.

The effective repellency (ER) was calculated using the formula below:

$$ \mathrm{ER}=\frac{\mathrm{NC}-\mathrm{NT}}{\mathrm{NC}}\times 100 $$
(1)

Where ER is effective repellency, NC is the number of eggs in control and NT is the number of eggs in treatment.

The formula was used to calculate the oviposition active index (OAI) was described by Kramer and Mulla (1979) which is:

$$ \mathrm{OAI}=\frac{\mathrm{NC}-\mathrm{NT}}{\mathrm{NT}+\mathrm{NC}}\kern0.5em $$
(2)

Oviposition active index of any substance can be categorised as follows:

  • No oviposition deterrence or attractant substance when OAI > 0

  • Moderated oviposition inhibit substance (moderate oviposition deterrence) - 0.5 < OAI < 0

  • Strong oviposition inhibit substance (strong oviposition deterrence) -1 < OAI < -0.5

Larvicidal bioassay

The method used for the larvicidal bioassay follows the procedure described by the WHO (2005) with little modifications. Twenty (20), 3rd/4th larval instar obtained from the culture were introduced separately into the various concentration of plant extracts along with a set of control containing sterile distilled water. All the tested concentrations were replicated three times. An aliquot of 50 ml of 50, 100, 200, 400, 800 mg/l and water (control) was used. Larvae that did not respond when pricked with a needle were recorded as been dead. The number of dead larvae was counted and recorded accordingly after 24, 48 and 72 h of exposure using the formula below.

$$ \%\mathrm{Larval}\ \mathrm{mortality}=\frac{\mathrm{Number}\ \mathrm{of}\ \mathrm{dead}\ \mathrm{larvae}}{\mathrm{Number}\ \mathrm{of}\ \mathrm{larvae}\ \mathrm{introduced}}\times 100 $$
(3)

Afterwards, the percentage of mortality was observed and corrected using Abbott (1925) formula shown below:

$$ \%\mathrm{corrected}\ \mathrm{mortality}=\frac{\%\mathrm{test}\ \mathrm{mortality}-\%\mathrm{control}\ \mathrm{mortality}}{100-\%\mathrm{control}\ \mathrm{mortality}}\times 100 $$
(4)

Pupicidal bioassay

A similar approach used for the larvicidal bioassay was used for the pupicidal bioassay. Twenty (20) two-day-old pupae were introduced separately into the various concentration of plant extracts along with a set of control containing sterile distilled water. All tested concentrations were replicated three times. An aliquot of 50 ml of 50, 100, 200, 400, 800 mg/l and water (control) was used. Pupae that didn’t respond to any stimulus (needle pricks) were considered to be dead. The number of dead pupae was counted and recorded accordingly after 24, 48 and 72 h of exposure using the formula below:

$$ \%\mathrm{Pupal}\ \mathrm{mortality}=\frac{\mathrm{Number}\ \mathrm{of}\ \mathrm{dead}\ \mathrm{pupae}}{\mathrm{Number}\ \mathrm{of}\ \mathrm{pupae}\ \mathrm{introduced}}\times 100 $$
(5)

Afterwards, the mortality is corrected using the Abbot formula given in eq. 4.

Adulticidal bioassay

The method described by WHO (2006) for mosquito adulticidal testing was slightly modified and used for this study. The Twenty (20) sugar-fed (10% sucrose solution) adults were introduced into a 1500 ml container perforated at the top and plunged with cotton wool. Cotton wool of 0.5 g was used to soak an aliquot of 0.5 ml of 50, 100, 200, 400, 800 mg/l and 70% ethanol (control) separately. These soaked cotton wools were left for a period of 2 h so that the ethanol used in the formulation of the concentrations can escape leaving behind the extract itself. The cotton wool with the extracts was then introduced into the containers containing the mosquitoes. Mortality was recorded when the mosquitoes failed to respond to an external stimulus such as tapping of the containers. All tested concentrations were replicated three times. The number of dead mosquitoes was recorded after 30, 60, 90 and 120 min. Percentage mortality was calculated using the formula below:

$$ \%\mathrm{Adult}\ \mathrm{mortality}=\frac{\mathrm{Number}\ \mathrm{of}\ \mathrm{dead}\ \mathrm{adults}}{\mathrm{Number}\ \mathrm{of}\ \mathrm{adults}\ \mathrm{introduced}}\times 100 $$
(6)

The percentage mortality was then corrected using Abbot formula given in eq. 4.

Repellency test

The animal model procedures used by Oluyemi et al. (2018) was adapted for this study. Insect cages measuring 45 X 45 X 30 cm was used for this bioassay. The Albino rat was used as test animals. An area of 20cm2 was shaved on the rat which was then washed with unscented soap. An aliquot of 0.5 ml of each concentration including the control (ethanol) was used to rub the shaved part of the rat separately. The rat with any given concentration was then placed inside the insect cage containing about disease-free 20 starved gravid female mosquitoes. The experiments were carried out during night time (19:00–05:00) when female Anopheles mosquitoes are considered to feed actively. When the mosquito lands on the surface and stays there for more than 2–3 s, the time of landing and the number of mosquitoes is recorded. The tests for each concentration were replicated three times and the percentage repellency was calculated with a formula described by Govindarajan and Sivakumar (2011) which is given as:

$$ \%\mathrm{Repellency}=\frac{\mathrm{Ta}-\mathrm{Tb}}{\mathrm{Ta}}\times 100 $$
(7)

Where Ta is the number of mosquitoes in the control group and Tb is the number of mosquitoes in the treated group.

Data analysis

The results from the bioassays were subjected to one-way analysis of variance (ANOVA) and the means separated using Tukey’s Honest Significance Test. Probit analysis was used to calculate lethal concentrations (LC) required to cause 50 and 90% (LC50 and LC90) for bioassays that percentage mortalities were the values recorded (Finney 1971). Multivariate regression analysis was used to find the regression between concentration with oviposition (mean no. of eggs laid) and concentration with repellency time (minute). All level of significance was set at p < 0.05. The ANOVA and Probit analysis were done using Statistical Package for the Social Sciences (SPSS) version 20 while the multivariate regression analysis was done using Paleontological Statistics (PAST) version 4.01.

Results

Oviposition deterrent activities of C. volubile and P. alliacea against An. gambiae

All the different concentrations of both plant extracts tested yielded a negative oviposition activity index which ranged between −0.02 to −0.76 (Table 1). This goes to show that they all inhibited the oviposition of An. gambiae. When the concentration was high, the oviposition was also high which shows that there is a concentration-dependent relationship existing between the extracts and their oviposition deterrent ability. This also indicates that an increment in the concentration resulted in a corresponding increase in effective repellency percentage (Table 1). The total number of eggs laid in the control plastic cup was very different from the number of eggs laid in treated plastic cups (p < 0.05) for all the extracts. Treated cups containing 50 mg/l of C. volubile extract had the highest number of eggs laid which was 272.78 ± 6.77. However, cups with 800 mg/l of P. alliacea presented the lowest eggs laid which were 40.44 ± 3.64. The effective repellency percentages for the extracts ranged from 4.22–86.45%. C. volubile extract at 50 mg/l had the lowest percentage while at 800 mg/l, P. alliacea recorded the strongest oviposition deterrence. Strong oviposition deterrence was observed for C. volubile at only 800 mg/l but at 400 mg/l and 800 mg/l for P. alliacea. This, therefore suggests that P. alliacea extract was the most active of the two extracts (Table 1). Table 2 shows the relationship that exists between the concentration and the mean number of eggs laid (oviposition). The result showed that the two plant extract significantly affected the number of eggs laid. This was so because the different concentrations of the two plant extracts reduced oviposition with respect to their various control.

Table 1 Oviposition deterrent activities of C. volubile and P. alliacea against An. gambiae
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Table 2 Relationship between the concentration and mean number of eggs laid (oviposition)
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Larvicidal activities of C. volubile and P. alliacea against An. gambiae

Mortality of larvae of An. gambiae were recorded for all concentrations of the two plant extracts used. The percentage of mortality is presented in Table 3. Significant differences were observed between different concentrations of C. volubile and P. alliacea that had the same period of exposure at p < 0.05. Exposing the larvae to the lowest (50 mg/l) and highest (800 mg/l) concentrations for 24 h yielded 20.00 and 58.33% for C. volubile; 25.00 and 100.00% for P. alliacea extracts respectively. Forty-eight (48) hours of the exposure recorded 100% mortality for 800 mg/l of C. volubile, 400 and 800 mg/l of P. alliacea concentrations. At about 72 h of exposure, three concentrations of P. alliacea extracts namely 200, 400 and 800 mg/l had 100% mortality while 400 and 800 m/l of C. volubile extract had the same result. The overall result points to the fact that the extracts had increasing mortalities proportionate to increasing concentrations and time of exposure. This result suggests that P. alliacea extract was the most active of the two extracts tested.

Table 3 Larvicidal activities of C. volubile and P. alliacea against An. gambiae
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Pupicidal activities of C. volubile and P. alliacea against An. gambiae

Mortality of pupae of An. gambiae were recorded for all concentrations of the two plant extracts used. The percentage of mortality is presented in Table 4. Significant differences were observed between different concentrations of C. volubile and P. alliacea that had the same period of exposure at p < 0.05. When the pupae were exposed to the lowest (50 mg/l) and highest (800 mg/l) concentrations for 24 h yielded 6.67and 51.67% for C. volubile; 13.33 and 85.00% for P. alliacea extracts. After 48 h of exposure 800 mg/l concentrations of C. volubile and P. alliacea extracts, mortalities of 71.67% and 100.00% respectively. At about 72 h of exposure, 100% mortality was recorded for 800 mg/l of C. volubile and, 400 and 800 mg/l of P. alliacea extract. The overall result points to the fact that the extracts had increasing mortalities proportionate to increasing concentrations and time of exposure. This result suggests that P. alliacea extract was the slightly more active of the two extracts tested as pupicides.

Table 4 Pupicidal activities of C. volubile and P. alliacea against An. gambiae
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Adulticidal activities of C. volubile and P. alliacea against An. gambiae

The results for the adulticidal efficacies of C. volubile and P. alliacea are presented in Table 5. There was a significant difference (p < 0.05) in the mortalities recorded among the various concentrations used with respect to the period of exposure. After 30 min of exposure, the lowest concentration used (50 mg/l) did not produce any mortality but the highest concentration produced the same mortality of 16.67% for both plant extracts. At the maximum period of exposure (120 min) used for this study, 400 and 800 mg/l of P. alliacea recorded mortalities of 100% while 800 mg/l of C. volubile had a mortality of 75%. The percentage of mortality recorded seems to appear in a concentration and exposure period dependent manner.

Table 5 Adulticidal activities of C. volubile and P. alliacea against An. gambiae
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Repellent activities of C. volubile and P. alliacea against An. gambiae

The results of the repellent activities of C. volubile and P. alliacea is presented in Table 6. At a period of 15 min, all the concentrations of the two plant extracts showed a 100% repellency. 800 mg/l of C. volubile provided 100% repellency until 90 min while P. alliacea still had 100% repellency until 120 min of post-exposure. However, after 150 min of exposure, none of the plant extracts could boast of 100% with 800 mg/l of C. volubile and P. alliacea only managing a 64.44 and 93.75% respectively. Table 7 shows the relationship between the concentrations of the two plant extracts and repellency time. The result revealed that repellency by all concentration significantly reduces after 90 min for C. volubile and 120 min for P. alliacea.

Table 6 Repellency activities of C. volubile and P. alliacea against An. gambiae
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Table 7 Relationship between the concentrations of the plant extracts C. volubile and P. alliacea and repellency time
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Lethal concentrations (LC) of C. volubile and P. alliacea against An. gambiae

Lethal concentrations of C. volubile and P. alliacea against required to cause 50 and 90% mortality in different life stages of An. gambiae are shown in Table 8. The 50% lethal concentration (LC50) of C. volubile and P. alliacea extracts after 24 h was 471.24 and 142.76 mg/l respectively for the larva stage. However, these values were significantly reduced to 48.20 and 40.00 mg/l for C. volubile and P. alliacea extract after 72 h. The 90% lethal concentration (LC90) of C. volubile and P. alliacea was 13,572.54 and 612.94 mg/l for 24 h and after 72 h, 194.13 and 109.39 mg/l respectively. The pupicidal LC50 and values for C. volubile extract after 24 and 72 h were 677.00 and 105.73 mg/l, and LC90 of 6640.76 and 549.86 mg/l correspondingly. Nevertheless, the lethal concentrations values recorded for P. alliacea extract were lower compared to C. volubile extract. Its LC50 values were 274.56 and 57.37 mg/l after 24 and 72 h and LC90 values of 1485.35 and 241.22 mg/l respectively. Generally, the lethal concentrations tend to decrease as the period of post-exposure increases for the larva and pupa stages. The adulticidal LC50 for C. volubile and P. alliacea extracts after 120 min was 208.45 and 80.65 mg/l respectively. The adulticidal LC90 of P. alliacea was lower than that of C. volubile.

Table 8 Lethal Concentrations (LC) of C. volubile and P. alliacea against An. gambiae
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Discussion

The usefulness and the prospect in the use of botanical based insecticide as a substitute for their synthetic contemporaries in the control of mosquito are gradually gaining traction (Ranganatha et al. 2013; Ileke et al. 2017; Ileke and Olabimi 2019). This study showed that ethanolic extracts of P. alliacea and C. volubile had mosquitocidal effects in terms of oviposition deterrence, larvicidal, pupicidal, adulticidal and repellent capabilities on An. gambiae.

The oviposition active index (OAI) recorded in this study revealed that the extracts were active in the deterring the oviposition of An. gambiae because of their negative values. These results corroborate the work of Wangrawa et al. (2016) on the oviposition-deterrence activities of ethanol, acetone and hexane extracts of Lantana camara L., Hyptis suaveolens Poit., Ocimum canum Sims. and Hyptis spicigera Lam. on sensitive and local strains of An. gambiae s. l. Also, Prathibha et al. (2014) demonstrated the efficacies of petroleum ether and ethyl acetate extract of Eugenia jambolana, Solidago canadensis, Euodia ridleyi and Spilanthes mauritiana in inhibiting the egg-laying capabilities of Aedes aegypti, Culex quinquefasciatus and Anopheles stephensi. The submissions of Prathibha et al. (2014) and Wangrawa et al. (2016) showed that OAI and effective repellency percentage exhibited by the different plant extracts used in their studies presented themselves in a dose-dependent manner synonymous to results from this present study. In this study, only a concentration of C. volubile had very high OAI and effective repellency percentage while for P. alliacea was two. It then follows that they can be very useful when added into stagnant waters that could serve as oviposition sites of An. gambiae species to control them. Thus, effectively controlling the egg-laying habits of any species of mosquito will go a long way towards reducing the menace they constitute. This is so because oviposition is a vital part of their life cycle (Xue et al. 2001; Qiu et al. 2006; Prathibha et al. 2014).

The larvicidal activity of the ethanolic extract of P. alliacea showed the highest percentage mortality throughout the different period of exposure. The work of Hartmann et al. (2018) showed that different extract of P. alliacea showed that they were highly effective in controlling the larva of Ae. aegypti. Unlike P. alliacea, C. volubile extract caused absolute mortality on An. gambiae larva only after the second day of exposure. The larvicidal activity of C. volubile in this study is similar to the ones produced by some other species of the genus Clerodendrum (Patil et al. 2014; Muthu et al. 2015; Lurdu and Thampi 2017). For instance, Patil et al. (2014) showed that extracts of Clerodendrum inerme were able to control the larva of Ae. aegypti and Cx. quinquefasciatus. Also,Muthu et al. (2015) demonstrated the efficacy of Pectolinaringenin, a compound isolated from Clerodendrum phlomidis Linn. F. against An. stephensi and Earias vittella larvae. The result gotten from this study conforms with several other works that have tested the efficacy of botanicals as potential mosquito larvicides (Ileke et al. 2017; Ileke and Olabimi 2019; Ileke and Adesina 2019). The high larvicidal efficacies of the plants used in this study may be because the larvae will swim around ingesting the extracts alongside phytochemicals present in them (Ileke et al. 2014).

The pupicidal bioassay in this study revealed that none of the two plants extracts caused maximum mortality on the test organism used until after the second day of exposure. Nevertheless, both plant extracts controlled the pupa of An. gambiae after the third day of exposure. The outcomes of this study agree with previous works carried out in this field. Ileke and Adesina (2018) reported the efficacy of Clerodendrum capitatum and Bridelia machrantha leaves extracts against An. gambiae pupae. Also, Panneerselvam et al. (2012) and Kashte et al. (2015) in their respective research showed the pupicidal activities of various plant leave extract against different malaria vectors. The mortality rate recorded in the pupicidal bioassay for the same exposure period was a bit lower compared to those recorded in the larvicidal bioassay. This might be due to the morphological differences that exist between the larvae and the pupae of mosquito thereby suggesting that the mode of action of these plant products is either by choking or contact and not necessarily ingestion as did pupal stage does not ingest whatsoever (Raveen et al. 2017).

Petiveria alliacea however, had more potent adulticidal effects after the maximum exposure period causing absolute mortality of the tested mosquito species. Although at lower concentrations and period of exposure, C. volubile induced very close percentage mortality as P. alliacea extract. The findings of this present study are in line with the works of other authors who have tested the efficacies of plant extract on different mosquitoes adults. For instance, difference extracts of G. pentaphylla leaves were effective in controlling the adults of An. stephensi, Cx. quinquefasciatus and Ae. aegypti (Ramkumar et al. 2016). Kovendan et al. (2013) in their work found the extract of Acalypha alinifolia leaves to be potent against adults of three different mosquito species. Mavundza et al. (2014) reported the adulticidal activities of ethanolic extract of about ten different plant species against An. arabiensis with the conclusion of them being effective. Also, Ileke and Ogungbite (2015) demonstrated the potency of Alstonia boonei in the management of adult An. gambiae mosquitoes.

The protectant ability of the plant extracts used in this study showed higher concentrations of the extract offered absolute protection from An. gambiae bites for a longer time. This increase of the plant extracts protection time at higher extract concentrations could be due to increased concentration of the active ingredient present in the extracts (Afolabi et al. 2018). Petiveria alliacea was observed to have better repellency efficacy as it provided absolute protection for a longer time compared to C. volubile. The result gotten from this study is in synchronisation with previous submissions of several authors in this area of study. Govindarajan and Sivakumar (2011) stated that different solvent extracts of Eclipta alba and Andrographis paniculata were efficacious in repelling mosquitos from the skin for a period of 150–120 min. Tribulus terrestris extracts have been proven to be able to repel Ae. aegypti, the dengue fever mosquito (El-Sheikh et al. 2016). Afolabi et al. (2018) proved that different solvent extracts of Ocimum caninum, Ocimum gratissimum, Chromolaena odorata and Datura stramonium were effective as repellents of An. gambiae.

The LC50 and LC90 values recorded in this study was dependent on the period of exposure. For instance, the LC50 and LC90 values for the larvicidal and pupicidal were observed to significantly decrease as the period of exposure increased. This was in agreement with previous studies of whereby plant extracts were used to control different mosquito species (Dahchar et al. 2016; Asiry et al. 2017; Abok and Ombugadu 2018; Ullah et al. 2018). It, therefore, means that higher concentration of the tested plant extracts is required to effect desired levels of control within a short time and lower concentration for a longer period. It was also observed that the LC50 and LC90 values were lower for the larval stages as compared to pupa stages. This observation is in tandem with Ileke (2018) who reported that the LC50 and LC90 values of Monodora myristica and Conyza sumatrensis extracts against the larvae of two mosquito species to be lower than the pupae. Overall, P. alliacea recorded the lowest LC50 and LC90 for all the bioassays were mortality was examined in this study. In general terms, the mosquitocidal effects of the two plant extracts used in this study were concentration and time-dependent as higher mortalities were observed as the concentration and period of exposure increased. This was also the observations of Edwin et al. (2013) and Kashte et al. (2015) in similar researches.

The effectiveness exhibited by the two plants used in this study in controlling different life stages of An. gambiae might be linked to the phytochemicals present in them. This is so because plants that have been found out to be insecticidal are rich in phytochemicals such as alkaloids, flavonoids, steroids, tannins, terpenes and terpenoids (Alqasoumi et al. 2012; Adesina and Rajashekar 2018; Afolabi et al. 2018). The results from this current study sides with several previous studies that have highlighted the utilization of the plants with bio-active chemicals as potential, environment safe, alternatives to synthetic insecticides.

Conclusion

The results obtained from the use of ethanolic extracts of C. volubile and P. alliacea in the management and control of different life stages of An. gambiae proved that these plants were highly efficacious most especially P. alliacea. This was so because it had the lowest LC50 and LC90 values across all the various bioassays. All the life stages were susceptible to all the concentrations of the plant extracts used in this study but concentrations of 400 and 800 mg/l were more effective in controlling them. Hence, they can be used to replace chemical insecticides as a way of curbing and reducing the hazardous effects associated with their use.