Skip to main content
Beni-Suef University Journal of Basic and Applied Sciences
Download PDF

Insecticidal activity and possible modes of action of secondary metabolites of some fungal strains and wild plants as natural pesticides against Spodoptera frugiperda

Beni-Suef University Journal of Basic and Applied Sciences volume 13, Article number: 9 (2024) Cite this article

Abstract

Background

The fall armyworm (FAW), Spodoptera frugiperda, is an invasive and destructive pest to certain strategic crops, especially maize in Egypt. This research was conducted mainly to obtain secondary metabolites from some wild plants and fungal strains, use them in controlling FAW, and investigate their mode of action.

Results

The ethyl acetate extracts of Cladosporium cladosporioides and Verticillium lecanii, as well as the alkaloid extracts of Ricinus communis and Nicotiana glauca, were extracted to obtain their secondary metabolites. The secondary metabolite contents were identified by gas chromatography mass spectrometry and NMR. The toxicity of all extracts against the 3rd-instar larvae of FAW was evaluated. The possible mode of action of the extracts was studied via their effects on larval enzyme activities and larval tissue. The toxicity results illustrated that, the extract of C. cladosporioides was more effective with LC50 229 ppm, followed by the extract of V. lecanii with LC50 341 ppm and N. glauca with LC50 404 ppm, while the least effective extract was R. communis with LC50 1110 ppm after 72 h of treatment. While, the results of larval enzyme activities showed that C. cladosporioides, V. lecanii, and R. communis extracts led to significant activity of AST, ALT, ACP, and ALP enzymes, but GST and AchE were inhibited in treated larvae compared with control. While N. glauca alkaloid extract caused significant inhibition of AST, ALT, ACP, AchE, and GST enzymes, ALP was activated in the treated larvae compared with control. The results of larval tissue slides indicated that the most affected tissues were the cuticle layer and the membrane lining of the midgut, in addition to the fatty bodies.

Conclusion

Thus, natural pesticides would have a promising role in terms of controlling the FAW and according to this study, it was recommended that, alkaloid extracts of tested wild plants and ethyl acetate extracts of fungal strains be used as natural pesticides to control the fall armyworm, S. frugiperda.

1 Background

Fall armyworm (FAW), Spodoptera frugiperda (JE Smith) (Lepidoptera: Noctuidae), a destructive pest to certain strategic crops [1, 2]. It is native to North and South America and since 2016 started to invade the African continent causing a significant damage to certain cultivated crops particularly maize crops [3, 4]. Then, it has subsequently spread throughout the continent and across Asia. However, fall armyworm had a wide range of plants hosts and could feed on over 350 plant species. Damage of FAW has been estimated to cause up to 13 billion US dollars per annum in crop losses throughout sub-Saharan Africa. According to CIMMYT Annual Report [5], FAW had devastated almost 1.5 million hectares of cultivated corn crops in six African countries. In Egypt, FAW was firstly recorded in maize field in a village in Kom Ombo city of Aswan governorate, Upper Egypt 2019 season according to the Agricultural Pesticide Committee (APC) and Ministry of Agriculture and Land Reclamation. As the north coast of Africa are environmentally suitable for FAW infestations besides its wide range of host plants, destructive damage would be expected, and it is contingent to harm other strategic crops such as cotton, soybean, and rice. Therefore, the population of FAW S. frugiperda should be under control.

So, many interested in the plant protection field in Egypt started to study several methods to suppress the FAW S. frugiperda population and reduce its damages. Among of these methods, using entomopathogenic fungi and plant extracts. Many previous studies reported that the mode of action of most entomopathogenic fungi refer to their production from bioactive secondary metabolites or enzymes and toxic proteins [6, 7]. In this study, authors used secondary metabolites of entomopathogenic fungi Cladosporium sp. and Verticillium sp. as well as alkaloid extract of wild plants castor bean, Ricinus communis and tobacco, Nicotiana glauca.

The fungus of Cladosporium sp. belongs to Ascomycota division. It was used as an entomopathogenic fungus to control several of insects in many previous studies. Shaker et al. [8] and Elbanhawy et al. [9] evaluated the efficiency of secondary metabolites which were extracted from Cladosporium cladosporioides against cotton aphids, Aphis gossypii. They found high toxicity effect where LC50 values were 24 and 36 ppm against nymphs and adults of aphids, respectively. Also, Habashy et al. [10] mentioned that spores suspension of C. cladosporioides caused high mortality percentage of spotted spider mites, Tetranychus urticae. Abdullah [11] isolated Cladosporium sp., Verticillium sp., Epicoccum sp., Alternaria sp., Fusarium sp., Penicillium sp., and Aspergillus sp. during his M.S. thesis from aphid and whitefly. The toxicity of isolated fungi was tested against aphids and whitefly. He found that Cladosporium sp., Verticillium sp., and Epicoccum sp. were more toxic fungi compared with other tested fungi. Also, Bahar et al. [12] isolated Cladosporium sp. from eggs of Helicoverpa armigera (Lepidoptera: Noctuidae) and tested its pathogenicity against eggs and larvae of H. armigera, cotton aphids, Aphis gossypii and whitefly, Bemisia tabaci. Verticillium lecanii is an opportunistic Ascomycete’s fungus and widely distributed. V. lecanii (Zimm.) is commonly known as the "white holo.," and it causes mycosis of numerous insects belonging to the orders Lepidoptera, Coleoptera, and Homoptera [13]. Lecanicillium lecanii, L. longisporum, L. attenuatum, L. nodulosum, and L. muscarium are among the new taxonomic entities divided from the species V. lecanii. There are number of important species from the Verticillium spp. group, which are used as biocontrol agents to control insect pests and diseases in agriculture [14]. There are 15 productions of bioinsecticides belonged to Verticillium spp. commercialized worldwide [15, 16].

Castor bean (Ricinus communis L) is a species of perennial flowering plant that belongs to family Euphorbiaceae [17]. The phytochemical screening of R. communis indicated the presence of flavones, isoflavones, flavonols, chalcones, aurones, sterols, saponins, and leucoanthocyanidins [18]. In addition, six flavonol glycosides; kaempferol-3–0-β-D-xylopyranoside, kaempferol -3–0-β-D-glucopyranoside, quercetin3-0-β-D-xylopyranoside, quercetin-3–0-β-D-glucopyranoside, kaempferol-3–0-β-rutinoside, and quercetin-3-O-β-rutinoside as well as two alkaloids; ricinine and N-demethylricinine were screened by Kang et al. [19].

Several previous studies reported that the crud extract of secondary metabolites of castor plant R. communis found to has insecticidal activity against pests such as sugarcane aphid, Melanaphis sacchari [20], acaricidal and insecticidal activities against Haemaphysalis bispinosa Neumann adult and hematophagous fly Hippobosca maculata Leach [21], malaria vector Anopheles gambiae [22] and in controlling the termites which damage the wood of Mangifera indica and Pinus longifolia [23].

Tobacco plant (Nicotiana glauca) is a flowering plant in the tobacco genus Nicotiana of family Solanaceae. The alkaloid fraction of N. glauca was analysis by UPLC/MS and GC/MS, anabasine was found to be the major constituent (60%). In addition to, five compounds were identified as nicotine, nornicotine, ammodendrine, chlorogenic acid, and rutin [24]. The acute toxicity of N. glauca was due to the highly percentage of anabasine compound [25]. It was found that N. glauca plant had antioxidant activity as a result of its content of flavonoids and phenolic compounds, which recorded their highest values in the leaves and flowers [26]. As a result of the presence of a high percentage of alkaloids in N. glauca, great importance was given to its use as antimicrobial agents [27] and insecticides to control aphids [28]. The aim of this study is to extract the secondary metabolites from entomopathogenic fungi, C. cladosporioides and V. lecanii, as well as from wild plants, R. communis and N. glauca. In addition to evaluating the influence of their crud extracts on larval mortality, enzyme activity, and larval tissue of the fall armyworm, S. frugiperda.

2 Methods

2.1 Insect rearing procedure

The colony of fall armyworm was established initially from larvae collected during the summer season of 2022 from corn field located in Dakahliah governorate. Larvae were reared individually to avoid cannibalism and were fed on fresh leaves of castor oil plants Ricinus communis L. replaced every 2 days, depending on how long they remain green and fresh. Larvae were kept in climatic control chamber (24 ± 1°C, 70% RH, 14L: 10D photoperiod) till pupation. Pupae were collected and placed in PVC container filled with sand under same condition until adult emergence. Following emerging, moths were collected and held in 1 L. glass mason jars, 20–40 moth per jar were kept together to encourage mating and were fed on honey bee solution from a cotton wick hanged from the jar top. Jars were kept in a growth chamber under same previous conditions with pieces of zigzag shaped papers to provide dark arena for eggs oviposition. Once mated adult started oviposition, egg mass were collected and held in plastic containers 200 ml in climatic control chamber until hatching. In order to obtain larvae for the experiment, a number of five neonate larvae were kept in plastic container 100 ml and were provided with small pieces of corn cob. Once larvae reached third instar, we isolate and count the proper number of larvae for the experiment then they were subjected to treatments.

2.2 Fungal strains

Fungal strains Cladosporium cladosporioides and Verticillium lecanii were kindly obtained from the Microbiology Department, Faculty of Agriculture, Mansoura University, Egypt. Both fungal strains were cultured on potato dextrose agar medium (PDA) made from potato infusion 200 ml, dextrose 20 g, and agar 15 g per liter, then incubated at 25°C. Figure 1 shows the colonies growth on potato dextrose agar medium and spore shapes under light microscope of both fungal strains.

Fig. 1
figure 1

The colonies growth of C. cladosporioides and V. lecanii on potato dextrose agar medium and their spore shapes under a light microscope

Full size image

2.3 Extraction of fungal metabolites from their broth cultures

One hundred ml of potato dextrose broth medium (PDB) was prepared in a 250-ml conical flask and sterilized in an autoclave. One disk of solid fungal culture was used to inoculate 100 ml of sterilized PDB medium. The inoculated medium was incubated to 7 days at 25 °C. After incubation period, one litter of sterilized PDB medium was inoculated by 100 ml broth culture in 2 litter conical flask and incubated to 21 days at 25 °C. After incubation period, the broth culture was filtered to remove the mycelium and obtain to the filtrate by Whatman no1 filter paper, Sigma-Aldrich. The volume of filtrate was measured then the equal volume from ethyl acetate was added. The mixture was poured in separating funnel and moved well for homogenate then left to 2 h. The upper layer was taken then ethyl acetate was evaporated by rotary evaporator system to obtain the crud extract of fungal metabolites. This method was conducted as described by Abdullah [29].

2.4 Gas chromatography–mass spectrometry (GC–MS) analysis

Trace GC-TSQ mass spectrometer (Thermo Scientific, Austin, TX, USA) with a direct capillary column TG–5MS (30 m × 0.25 mm × 0.25 µm film thickness) was used to perform the chemical composition of fungal strains extracts. The temperature of the column oven was initially maintained at 50 °C then raised by 5 °C/min to 250 °C and maintained for two minutes then raised to final temperature 300 °C by 30 °C/min, where it was held for two minutes. The temperatures of injector and MS transfer line were maintained at 270, 260 °C, respectively. The carrier gas was Helium at a constant flow rate of 1 ml/min. Electron impact (EI) mass spectra were collected at 70 eV ionization voltages over the range of m/z 50–650 in full scan mode. The ion source temperature was set at 200 °C. By comparing the components' retention times and mass spectra to those in the WILEY 09 and NIST 14 mass spectral databases, the components were identified [29].

2.5 Plant material

Ricinus communis L "Euphorbiaceae" seeds were collected from plants in the villages of Shoha and Salamoun, beside to Mansoura city (31.065745° N–31.453642° E), Egypt in Mars 2022. Nicotiana glauca "Solanaceae" aerial parts were collected from Mansoura city (31.067849° N–31.415351° E), Egypt in June 2022. The collected plants were identified by prof. Ibrahim Mashaly, Botany department, Faculty of science, Mansoura University according to Boulos [30].

2.6 Extraction of secondary metabolites from plants

2.6.1 Processing of Ricinus communis L. (Seeds)

The dried seeds of R. communis L. (1 kg) were grinded and extracted with methanol (2L × 5). The methanol extract was concentrated by rotary evaporator to obtained 300 ml oil extract. A certain amount of oil was exposed to PTLC (Preparative Thin Layer Chromatography, silica gel Merck GF 254 precoated plates 20 × 20 cm on aluminum sheets) with eluent system CHCl3: MeOH (6:1) to isolated major compound (1), (25 mg, Rf 0.44), which gave positive result (orange color) with Dragendroff's reagent.

2.6.1.1 Characterization of isolated compound (1)

Compound (1): Slightly yellow to white needle crystal; 1H NMR (Bruker 400 MHz with tetramethylsilane, CD3OD/drops CD3Cl, δ, ppm, J, Hz): 7.85 (d, J = 7.7 Hz, H-6), 6.33 (d, J = 7.7 Hz, H-5), 4.02 (s, 3H-9), and 3.53 (s, 3H-7). 13C NMR (100 MHz, CD3OD/drops CD3Cl): 37.8 (N-CH3), 57.9 (O-CH3), 88.2 (C-3), 95.2 (C-5), 114.4 (CN), 145.9 (C-6), 163.0 (C-2), 173.9 (C-4).

2.6.2 Processing of Nicotiana glauca

The dried powder aerial parts of N. glauca (1 kg) extracted with methanol (1.8L × 5). The methanol extract was concentrated by rotary evaporator till dryness (83 gm). The methanol extract was dissolved in small amount of methanol and then added distilled water acidified by 2N HCl till pH 3. The acidified mixture was extracted by separating funnel with chloroform three times. The acidified aqueous extract was basified using ammonia solution 33% till pH 11 then extracted with chloroform (Alkaloid fraction 1.85 gm).

2.6.2.1 Characterization of identified compound (2)

Compound (2) pale yellow oil. GC/MS, m/z (rel. int.): 162 (49%) [C10H14N]+., 133 (70%) [C9H11N]+, 119 (58%) [C8H9N]+, 105 (75%) [C7H7N]+, 92 (20%) [C6H6N]+, 84 (100%) [C5H10N]+.

2.7 Bioassay experiment

Spray bioassay method was used to evaluate the toxicity of four crud extracts as natural pesticides against S. frugiperda under laboratory conditions. Five serial concentrations of each crud extract were prepared (500, 1000, 1500, 2000, 2500 ppm). Ten third-instar larvae of S. frugiperda were transferred to each jar (10D * 25H cm). Pieces of castor leaves were used to feed larvae. Two milliliter of each concentrate were sprayed on larvae and castor leaves in each jar. Two milliliter of water were used to treat control group. The number of live larvae was recorded every day. Mortality percentages were calculated and corrected by Abbot’s formula [31]. LC50, confidence limits and slope values were calculated as described by Finney method [32]. Also, toxicity index was calculated as described by Sun [33].

2.8 Estimation of the changes in insect enzyme activities

Fourth-instar larvae of S. frugiperda were used to investigate the effect of tested secondary metabolites on insect enzymes activities. The same method of bioassay experiment was repeated but LC50 of each crude extract was used to spraying the larvae. Also, water was used to spray control group. After four days, the live larvae were weighted and frozen in Eppendorf tube until analysis. All enzyme activities were estimated colorimetrically by UV–visible spectrophotometer, model V1200, China. The estimated insect enzymes were aspartate transaminase (AST), alanine transaminase (ALT) according to Reitman and Frankel [34] at 520 nm, acid phosphatase (ACP), alkaline phosphatase (ALP) according to Powell and Smith [35] at 510 nm, acetyl choline esterase (AchE) according to Simpson et al. [36] at 515 nm, and glutathione S-transferase (GST) according to Pan et al. [37] at 540 nm.

2.9 Determination of histological changes in insect tissue

Also the fourth-instar larvae of S. frugiperda were treated by LC50 of each extract using spray technique. Two live larvae of treated and untreated larvae were taken after five days of treatment and saved in 10% formalin solution. Tissue processing, sectioning, and staining were conducted according to Gaaboub et al. [38] in Faculty of Medicine, Mansoura University, Egypt.

3 Results

3.1 Toxicity effect of fungal and wild plants extracts against S. frugiperda

Table 1 shows the toxicity effect of ethyl acetate extracts of two fungal strains (C. cladosporioides & V. lecanii) and alkaloid fraction of N. glauca as well as methanol extract of R. communis) against S. frugiperda. The more effective extract after 48 h of treatment was the alkaloid extract of N. glauca (LC50 = 788 ppm) followed by the ethyl acetate extract of C. cladosporioides (LC50 = 886 ppm), while there were different results after 72 h of treatment where the extract of C. cladosporioides was more effective extract (LC50 = 229 ppm) followed by the extract of V. lecanii (LC50 = 341 ppm) and N. glauca (LC50 = 404 ppm). Also, there were different values of toxicity line’s slope between all tested extracts. This illustrate that could be there were different mode of actions of tested extracts against S. frugiperda. In addition, toxicity index showed that S. frugiperda was more susceptible to C. cladosporioides extract than other tested extracts. Figure 2 shows some abnormal symptoms of treated S. frugiperda larvae as necrosis and analysis of larvae body also the larva could not get rid of some parts of the cuticle layer, which led to the larva not growing normally.

Table 1 Toxicity of secondary metabolites of tested fungal strains and wild plants against S. frugiperda
Full size table
Fig. 2
figure 2

Abnormal symptoms of S. frugiperda larvae after treatment by crude extracts of fungal strains and wild plants under laboratory conditions

Full size image

3.2 The changes in insect enzymes activity

In this study, the changes of five insect enzymes activities were estimated in the treated larvae of S. frugiperda. The results in Table 2 show that C. cladosporioides, V. lecanii, and R. communis extracts led to significant activity of AST, ALT, ACP, and ALP enzymes, but GST and AchE were inhibited in treated larvae compared with control, while N. glauca alkaloid extract caused significant inhibition of AST, ALT, ACP, AchE, and GST enzymes, but ALP was activated in the treated larvae compared with control.

Table 2 Changes in the enzyme activities in the treated larvae of S. frugiperda caused by the extracts of fungal strains and plants
Full size table

3.3 Histological changes of treated larvae

Figures 3, 4, 5, 6, and 7 show the histological changes of treated 4th-instar larvae by C. cladosporioides, V. lecanii, N. glauca, and R. communis extracts as well as control treatment. The description of histological changes is under each cross section’s photo. The most affected tissues were the cuticle layer and the membrane lining of the midgut, in addition to the fatty bodies.

Fig. 3
figure 3

Cross section through untreated 4th-instar larvae of S. frugiperda, stained using hematoxyline, Eosin (H, E X200), showing normal fibrous and cellular layer of cuticle (CT) also normal basement and peritrophic membranes in the midgut (MG) as well as normal cells of fat bodies (FB)

Full size image
Fig. 4
figure 4

Cross section through treated 4th-instar larvae of S. frugiperda by ethyl acetate extract of C. cladosporioides and stained using hematoxyline, Eosin (H, E X200), showing weak and necrosis of cuticle layers (CT), severe necrosis and dissolved cells of fat bodies (FB) and ruptured columnar cells and destroyed basement membrane of midgut (MG)

Full size image
Fig. 5
figure 5

Cross section through treated 4th-instar larvae of S. frugiperda by ethyl acetate extract of V. lecanii and stained using hematoxyline, Eosin (H,E X200), showing necrosis of some parts in cuticle layers (CT), no effects in cells of fat bodies (FB) and severe necrosis, dissolved and ruptured columnar cells and destroyed basement membrane of midgut (MG)

Full size image
Fig. 6
figure 6

Cross section through treated 4th-instar larvae of S. frugiperda by ethyl acetate extracts of N. glauca and stained using hematoxyline, Eosin (H,E X200), showing no effects in cells of cuticle layers (CT), severe necrosis and dissolved cells of fat bodies (FB), severe necrosis, dissolved and ruptured columnar cells and destroyed basement membrane of midgut (MG)

Full size image
Fig. 7
figure 7

Cross section through treated 4th-instar larvae of S. frugiperda by ethyl acetate extract of R. communis and stained using hematoxyline, Eosin (H,E X200), showing dissolved cells in fat bodies (FB), severe necrosis, dissolved and ruptured columnar cells and destroyed basement membrane of midgut (MG), necrosis and separating in some parts of cuticle layers (CT)

Full size image

3.4 Discussion

The entomopathogenic fungi C. cladosporioides and V. lecanii will be active and promising agents to control the fall army worm (FAW) in Egypt. In this study, C. cladosporioides and V. lecanii were more toxic against FAW larvae with LC50s of 229 and 341 ppm after 72 h, respectively. On the other hand, both fungi were used as entomopathogens against many insect pests in other studies, such as Aphis gossypii [8, 9, 11, 42], Helicoverpa armigera, Bemisia argentifolii [12], Sitophilus granarius, S. oryzae, S. zeamais, Rhyzopertha dominica, and Trogoderma granarium [43], Plutella xylostella [44], Nilaparvata lugens [45, 46].

However, using entomopathogenic fungi to manage insect pests still has several flaws when it comes to field application. Entomopathogenic fungi are exposed to different weather stresses after being used in the field, including temperature [47, 48], humidity [48,49,50], and UV rays [51]. The entomopathogenic fungi produce a large number of bioactive secondary metabolites that have insecticidal properties [52]. Additionally, the entomopathogenic fungi can kill the pest more rapidly by producing some mycotoxins like beauvericin, cyclodepsipeptide, destruxin, and desmethyldestruxin in the early stages of infestation [53]. In this study, the tested fungi produced some compounds in the liquid culture that have insecticidal activities, like n-hexadecanoic acid, octadecadienoic acid, tetramic acid, selinane, and methyl ester of ricinoleic acid, as shown in Tables 1 and 2. Also, previous studies reported that these compounds had insecticidal activity: n-hexadecanoic acid and octadecadienoic acid [29, 54]; methyl ester of ricinoleic acid [55]; selinane [56]; and tetramic [57, 58].

Tables 3 and 4 show the identified compounds in the ethyl acetate extract of C. cladosporioides and V. lecanii broth cultures by GC–MS analysis. Thirteen compounds were identified in C. cladosporioides extract. The major compounds were n-hexadecanoic acid (Palmitic acid), hexadecanoic acid, methyl ester, 9, 12-Octadecadienoic acid (Z, Z)- (Linoleic acid) and 9,12-octadecadienoic acid, methyl ester, (E,E)- where their peak area percentages were 31.63%, 14.74%, 17.97%, and 13.14%, respectively. On the other hand twelve compounds were identified in V. lecanii extract. Three compounds had the more percentages of peak area compared with other compounds. These compounds were tetramic acid (13.22%), 7-Isopropyl-1,4a-dimethyldecahydronaphthalene # (Selinane) (16.09%) and methyl ester of ricinoleic acid (14.9%).

Table 3 GC–MS analysis of ethyl acetate extract of the liquid culture of C. cladosporioides
Full size table
Table 4 GC–MS analysis of ethyl acetate extract of the liquid culture of V. lecanii
Full size table

On the other hand, two wild plants (R. communis and N. glauca) were selected according to their alkaloids, which have a wide range of uses in many fields, especially as insecticides. Alkaloids are among the most important compounds that have a toxic effect on insects [59]. The analysis of R. communis extract indicated that ricinine (Compound 1) was the major constituent in the extract [39]. Ricinine alkaloid was used as a natural pesticide against Tetranychus urticae and two predatory phytoseiid mites [40]. As well, ricinine can be used as an insecticide against leaf-cutting ants [60]. The extracts of R. communis were used by [20] as botanical insecticides to control the sugarcane aphid, Melanaphis sacchari.

Compound (1) was isolated as a needle crystal and appeared as orange color with Dragendroff's reagent. Figure 8 shows the chemical structure of compound (1). 1H NMR and 13C NMR spectra (Table 5and Fig. 9 and 10) showed two aromatic doublet signals at δH 6.33 (d, J = 7.7 Hz, H-5), δC 95.2 ppm and δH 7.85 (d, J = 7.7 Hz, 1H), δC 145.9 ppm. As well as, two singlet signals at δH 3.53 (s, N-CH3), δC 37.8 ppm and δH 4.02 (s, O-CH3), δC 57.9 ppm. Also, 13C NMR spectrum indicated four quaternary carbon atoms at δC 88.2 (C-3), 114.4 (CN), 163.0 (C-2), 173.9 (C-4). Compound (1) was identified as ricinine by comparing its data with literature, [39, 40].

Fig. 8
figure 8

Chemical structure of ricinine (compound 1)

Full size image
Table 5 1H NMR data for ricinine (compound 1) in CH3OD/CD3Cl (δ) [ppm] (Multiplicity, J [Hz])
Full size table
Fig. 9
figure 9

1H NMR (400 MHz) spectrum of ricinine (compound 1) in CD3OD/CD3Cl

Full size image
Fig. 10
figure 10

13C NMR (100 MHz) spectrum of ricinine (compound 1) in CD3OD/CD3Cl

Full size image

In addition, the anabasine (Compound 2) was found to be the major constituent of the alkaloid fraction of N. glauca [24]. The anabasine alkaloid has been widely used as a pesticide. Its insecticidal effect is due to interaction with nicotinic acetylcholine receptors [61, 62]. Alkaloid fraction of N. glauca was analyzed by GC/MS technique. Compound (2) was identified as the major constituent in the fraction (Rt = 17.34, 65.52%) by comparing their mass spectra with those of their analogous reported by NIST library (Fig. 11). The GC/MS analysis indicated that the molecular ion peak at m/z 162 as [M]+. ([C10H14N2]+.) and base peak at m/z 84 for piperidine ring [C5H10N]+. The fragment ion peaks at m/z 133, 119, 105, and 92 due to fragment ions [C9H11N]+, [C8H9N]+, [C7H7N]+, and [C6H6N]+, respectively, were in agreement with anabasine [24, 41]. Figure 12 shows the chemical structure of anabasine (compound 2).

Fig. 11
figure 11

GC/MS analysis of anabasine (compound 2)

Full size image
Fig. 12
figure 12

Chemical structure of anabasine (compound 2)

Full size image

4 Conclusion

In conclusion, natural pesticides would have a promising role in terms of controlling the FAW where they avoid environmental damage, reduce synthetic pesticides, and overall reduce control cost. Thus according to this study, it was recommended that, alkaloid extracts of tested wild plants and ethyl acetate extracts of fungal strains be used as natural pesticides to control the fall armyworm, S. frugiperda.

Availability of data and materials

All data and materials are available if requested.

Abbreviations

FAW:

Fall armyworm

AST:

Aspartate transaminase

ALT:

Alanine transaminase

ACP:

Acid phosphatase

ALP:

Alkaline phosphatase

AchE:

Acetyl choline esterase

GST:

Glutathione S-transferase

GC–MS:

Gas chromatography–mass spectrometry

LC50 :

Median lethal concentration

Ppm:

Part per million

NMR:

Nuclear Magnetic Resonance

References

  1. Nagoshi RN, Adamczyk JJ, Meagher J, Gore RL, Jackson R (2007) Using stable isotope analysis to examine fall armyworm (Lepidoptera: Noctuidae) host strains in a cotton habitat. J Econ Entomol 100(5):1569–1576. https://doi.org/10.1093/jee/100.5.1569

    Article  PubMed  Google Scholar 

  2. Bueno RCOF, Carneiro TR, Bueno AF, Pratissoli D, Fernandes OA, Vieira SS (2010) Parasitism capacity of Telenomus remus Nixon (Hymenoptera: Scelionidae) on Spodoptera frugiperda (Smith) (Lepidoptera: Noctuidae) eggs. Braz Arch Biol Technol 53(1):133–139. https://doi.org/10.1590/S1516-89132010000100017

    Article  Google Scholar 

  3. Goergen G, Kumar PL, Sankung SB, Togola A, Tamo M (2016) First report of outbreaks of the fall armyworm Spodoptera frugiperda (JE Smith) (Lepidoptera: Noctuidae), a new alien invasive pest in west and Central Africa. PLoS ONE 11(10):e0165632

    Article  PubMed  PubMed Central  Google Scholar 

  4. Kumela T, Simiyu J, Sisay B, Likhayo P, Gohole L, Tefera T (2018) Farmers’ knowledge, perceptions, and management practices of the new invasive pest, fall armyworm (Spodoptera frugiperda) in Ethiopia and Kenya. International Journal of Pest Management 65(1):1–9. https://doi.org/10.1080/09670874.2017.1423129

    Article  Google Scholar 

  5. CIMMYT Annual Report (2017) International Maize and Wheat Improvement Center. One of fifteen CGIAR research centers helps to improve the lives in developing countries by encouraging more productive, sustainable maize and wheat farming. https://www.cimmyt.org/annual-reports/cimmyt-annual-report-2017/

  6. Isaka M, Kittakoop P, Kirtikara K, Hywel-Jones NL, Thebtaranonth Y (2005) Bioactive substances from insect pathogenic fungi. Acc Chem Res 38:813–823. https://doi.org/10.1021/ar040247r

    Article  CAS  PubMed  Google Scholar 

  7. Ortiz-Urquiza A, Keyhani NO (2013) Action on the surface: entomopathogenic fungi versus the insect cuticle. Insects 4(3):357–374. https://doi.org/10.3390/insects4030357

    Article  PubMed  PubMed Central  Google Scholar 

  8. Shaker NO, Ahmed GMM, Ibrahim HYE, El-Sawy MM, Mostafa ME, Ismail HNA (2019) Secondary metabolites of the entomopathogenic fungus, cladosporium cladosporioides and its relation to toxicity of cotton aphid, aphis gossypii (Glov). Int J Entomol Nematol 5(1):115–120

    Google Scholar 

  9. Elbanhawy AA, Elsherbiny AE, Abd El-Mageed AE, Abdel-Fattah GM (2019) Potential of fungal metabolites as a biocontrol agent against cotton aphid, Aphis gossypii Glover and the possible mechanisms of action. Pestic Biochem Physiol 159:34–40. https://doi.org/10.1016/j.pestbp.2019.05.013

    Article  CAS  PubMed  Google Scholar 

  10. Habashy MG, Al-Akhdar HH, Elsherbiny EA, Nofal AM (2016) Efficacy of entomopathogenic fungi Metarhizium anisopliae and Cladosporium cladosporioides as biocontrol agents against two Tetranychid mites (Acari: Tetranychidae). Egypt J Biol Pest Control 26:197–201

    Google Scholar 

  11. Abdullah RRH (2004) Studies on some bioagents against certain sucking insect pests. Thesis (M.S.) – Mansoura University, Faculty of Agriculture, Department of Microbiology, 2004, p. 142

  12. Bahar MH, Backhouse D, Gregg PC, Mensah R (2011) Efficacy of a Cladosporium sp fungus against Helicoverpa armigera (Lepidoptera: Noctuidae), other insect pests and beneficial insects of cotton. Biocontrol Sci Technol 21(12):1387–1397. https://doi.org/10.1080/09583157.2011.622036

    Article  Google Scholar 

  13. Upadhyay V, Rai D, Rana M, Mehra P, Pandey AK (2014) Verticillium lecani (Zimm.): a potential entomopathogenic fungus. Int J Agric Environ Biotechnol 7(4):719–727. https://doi.org/10.5958/2230-732X.2014.01380.1

    Article  Google Scholar 

  14. Goettel MS, Koike M, Kim JJ, Aiuchi D, Shinya R, Brodeur J (2008) Potential of Lecanicillium spp. For management of insects, nematodes and plant diseases. J Invertebr Pathol 98:256–261. https://doi.org/10.1016/j.jip.2008.01.009

    Article  CAS  PubMed  Google Scholar 

  15. Goettel MS, Eilenberg J, Glare TR (2005) Entomopathogenic fungi and their role in regulation of insect populations. In: Gilbert LI, Iatrou K, Gill S (eds) Comprehensive Molecular Insect Science 6. Elsevier, Oxford, pp 361–406

    Chapter  Google Scholar 

  16. Faria MR, Wraight SP (2007) Mycoinsecticides and mycoacaricides: a comprehensive list with worldwide coverage and international classification of formulation types. Biol Control 43:237–256. https://doi.org/10.1016/j.biocontrol.2007.08.001

    Article  CAS  Google Scholar 

  17. Wedin GP, Neal JS, Everson GW, Krenzelok EP (1986) Castor bean poisoning. Am J Emerg Med 4(3):259–261. https://doi.org/10.1016/0735-6757(86)90080-x

    Article  CAS  PubMed  Google Scholar 

  18. Barrales-Cureño HJ, Ruiz JEZ, Reyes CR, Hoyos PA, Cruz AL, Herrera LMS, Ruiz JAC, Caballero MCC, Salinas SC, Valdez LGL (2016) Chemical bio-directed determination of Ricinus communis and Argemone mexicana extracts. IOSR J Pharm 6(9):01–06

    Google Scholar 

  19. Kang SS, Cordell GA, Soejarto DD, Fong HHS (1985) Alkaloids and Flavonoids From Ricinus communis. J Nat Prod 48(1):155–156. https://doi.org/10.1021/np50037a041

    Article  CAS  Google Scholar 

  20. Sotelo-Leyva C, Salinas-Sánchez DO, Peña-Chora G, Trejo-Loyo AG, González-Cortázar M, Zamilpa A (2020) Insecticidal compounds in Ricinus communis L (Euphorbiaceae) to control Melanaphis sacchari Zehntner (Hemiptera: Aphididae). Florida Entomologist 103(1):91–95. https://doi.org/10.1653/024.103.0415

    Article  CAS  Google Scholar 

  21. Zahir AA, Rahuman AA, Bagavan A, Santhosh KT, Mohamed RR, Kamaraj C, Rajakumar G, Elango G, Jayaseelan C, Marimuthu S (2010) Evaluation of botanical extracts against Haemaphysalis bispinosa Neumann and Hippobosca maculate Leach. Parasitol Res 107(3):585–592. https://doi.org/10.1007/s00436-010-1898-7

    Article  PubMed  Google Scholar 

  22. Wachira SW, Omar S, Jacob JW, Wahome M, Alborn HT, Spring DR, Masiga DK, Torto B (2014) Toxicity of six plant extracts and two pyridine alkaloids from Ricinus communis against the malaria vector Anopheles gambiae. Parasit Vectors 7(312):1–8. https://doi.org/10.1186/1756-3305-7-312

    Article  CAS  Google Scholar 

  23. Sharma S, Vasudevan P, Madan M (1991) Insecticidal Value of Castor (Ricinus communis) against Termites. Int Biodeterior 27(3):249–254. https://doi.org/10.1016/0265-3036(91)90053-T

    Article  Google Scholar 

  24. Massadeh RK, El-Elimat T, Al-Gharaibeh M, Tawaha K, Alali FQ (2022) UPLC-HRESI-MS and GC-MS analysis of the leaves of Nicotiana glauca. Acta Pharm 72:97–108. https://doi.org/10.2478/acph-2022-0001

    Article  CAS  PubMed  Google Scholar 

  25. Schep LJ, Slaughter RJ, Beasley DMG (2009) Nicotinic plant poisoning. Clin Toxicol 47(8):771–781. https://doi.org/10.1080/15563650903252186

    Article  CAS  Google Scholar 

  26. Hassan HE, Abd El-Hameed TZ, Nasr EA (2014) Ecological and phytochemical studies on nicotiana glauca from Egypt. Egypt Soc Exp Biol 10(1):87–95

    Google Scholar 

  27. Alghamdi AA (2021) Phytoconstituents screening and antimicrobial activity of the invasive species Nicotiana glauca collected from Al-Baha region of Saudi Arabia. Saudi J Biol Sci 28:1544–1547. https://doi.org/10.1016/j.sjbs.2020.12.034

    Article  CAS  Google Scholar 

  28. El-Rokh AR (2007) Chemical Studies on Some Natural Extracts and Their Constituents to Control Some Aphid Species. M. Sc. Thesis, Faculty of Science, Mansoura University, Egypt

  29. Abdullah RRH (2019) Insecticidal activity of secondary metabolites of locally isolated fungal strains against some cotton insect pests. J Plant Prot Pathol Mansoura Univ 10(12):647–653. https://doi.org/10.21608/jppp.2019.79456

    Article  Google Scholar 

  30. Boulos L (2000) Flora of Egypt. Al Hadara Publishing Cairo, Egypt 2, 288. ISBN: 9789775429223, 9775429226.

  31. Abbott WS (1925) A method of computing the effectiveness of an insecticide. J Econ Entomol 18:265–267. https://doi.org/10.1093/jee/18.2.265a

    Article  CAS  Google Scholar 

  32. Finney DJ (1971) Probit analysis. Cambridge University Press

    Google Scholar 

  33. Sun YP (1950) Toxicity index on improved method of comparing the relative toxicity of insecticides. J Econ Entomol 43(1):45–53. https://doi.org/10.1093/jee/43.1.45

    Article  CAS  Google Scholar 

  34. Reitman S, Frankel S (1957) A Colorimetric Method for the Determination of Serum Glutamic Oxalacetic and Glutamic Pyruvic Transaminases. Am J Clin Pathol 28(1):56–63. https://doi.org/10.1093/ajcp/28.1.56

    Article  CAS  PubMed  Google Scholar 

  35. Powell MEA, Smith MJH (1954) The determination of serum acid and alkaline phosphatase activity with 4- amino antipyrine (AAP). J Clin pathol 7(3):245–248. https://doi.org/10.1136/jcp.7.3.245

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  36. Simpson DR, Bulland DL, Linquist DA (1964) A semi microtechnique for estimation of cholinesterase activity in boll weevils. Ann Entomol Soc Am 57:367–371

    Article  CAS  Google Scholar 

  37. Pan L, Ren L, Chen F, Feng Y, Luo Y (2016) Antifeedant activity of Ginkgo biloba secondary metabolites against Hyphantria cunea larvae: Mechanisms and applications. PLoS ONE 11(5):e0155682. https://doi.org/10.1371/journal.pone.0155682

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  38. Gaaboub IA, El-Kady HA, El-Khayat EF, El-Shewy AM (2012) Biochemical and histological effect of some plant extracts, insecticide (methomyl) and bio insecticide (protecto) against cotton leafworm, Spodoptera littoralis (Boisd.). In: 1st International Conference On Biotechnology Applications In Agriculture. Benha University, Moshtohor and Hurghada, 18–22, February 2012, Egypt

  39. El-Naggar MH, Elgaml A, Abdel Bar FM, Badr FA (2018) Antimicrobial and antiquorum-sensing activity of Ricinus communis extracts and ricinine derivatives. Nat Prod Res 33(11):1–7. https://doi.org/10.1080/14786419.2017.1423306

    Article  CAS  Google Scholar 

  40. Ellaithy AYM, Abdel-Khalek AA, Mohammed MA (2022) The potency of ricinine biopesticide from Ricinus communis leaves as an alternative host for mass rearing process of tetranychus urticae and two predatory phytoseiid mites. Egypt J Chem 65(6):535–549. https://doi.org/10.21608/EJCHEM.2021.107114.4922

    Article  Google Scholar 

  41. Ali EA (2004) Isolation, Purification and Biochemical Characterization of Acetyl cholinesterase (Ach E) Inhibitor Anabasine alkaloid toxin from Anabasis setifera Toxicity to rats. Ph.D. Thesis, Faculty of Science, Mansoura University

  42. Ebadollahi A, Davari M, Razmjou J, Naseri B (2017) Separate and combined effects of mentha piperata and mentha pulegium essential oils and a pathogenic fungus Lecanicillium muscarium against Aphis gossypii (hemiptera: aphididae). J Econ Entomol 110(3):1025–1030. https://doi.org/10.1093/jee/tox065

    Article  CAS  PubMed  Google Scholar 

  43. Mantzoukas S, Lagogiannis I, Kitsiou F, Eliopoulos PA (2023) Entomopathogenic action of wild fungal strains against stored product beetle pests. Insects 14(1):91. https://doi.org/10.3390/insects14010091

    Article  PubMed  PubMed Central  Google Scholar 

  44. Keppanan R, Sivaperumal S, Hussain M, Dash CK, Bamisile BS, Qasim M, Wang L (2018) Investigation and molecular docking studies of Bassianolide from Lecanicillium lecanii against Plutella xylostella (Lepidoptera: Plutellidae). Compar Biochem Physiol Part C 206–207:65–72. https://doi.org/10.1016/j.cbpc.2018.03.004

    Article  CAS  Google Scholar 

  45. Minarni EW, Soesanto L, Suyanto A, Rostaman R (2021) Effectiveness of Secondary Metabolites From Entomopathogenic Fungi For Control Nilaparvata lugens Stål. in the laboratory scale. J Perlindungan Tanaman Indonesia 25(1):86–97. https://doi.org/10.22146/jpti.62116

    Article  Google Scholar 

  46. Atta B, Rizwan M, Sabir AM, Golgi MD, Farooq MA, Batta YA (2020) Efficacy of Entomopathogenic Fungi against Brown Planthopper Nilaparvata lugens (Stål) (Homoptera: Delphacidae) Under Controlled Conditions. Gesunde Pflanzen 72(2):101–112. https://doi.org/10.1007/s10343-019-00490-6

    Article  CAS  Google Scholar 

  47. Saldarriaga AJJ, D’Alessandro CP, Conceschi MR, Mascarin GM, Delalibera JI (2017) Efficacy of entomopathogenic fungi against adult Diaphorina citri from laboratory to field applications. J Pest Sci 90(3):947–960. https://doi.org/10.1007/s10340-017-0846-z

    Article  Google Scholar 

  48. Zaman S, Hasan M, Ahmad F, Javed N (2020) Pathogenicity of Entomopathogenic Fungi against Sitophilus granarius (L.) (Coleoptera: Curculionidae) under Abiotic Factors. Pak J Agric Sci 57(1):79–86. https://doi.org/10.21162/PAKJAS/20.8661

    Article  Google Scholar 

  49. Hsia ICC, Islam MT, Ibrahim Y, How TY, Omar D (2014) Evaluation of conidial viability of entomopathogenic fungi as influenced by temperature and additive. Int J Agric Biol 16(1):146–152

    CAS  Google Scholar 

  50. Rai D, Updhyay V, Mehra P, Rana M, Pandey AK (2014) Potential of entomopathogenic fungi as biopesticides. Indian J Sci Res Technol 2(5):7–13

    Google Scholar 

  51. Kaiser D, Bacher S, Mène-Saffrané L, Grabenweger G (2018) Efficiency of Natural Substances to Protect Beauveria bassiana Conidia from UV Radiation. Pest Manag Sci 75(2):556–563. https://doi.org/10.1002/ps.5209

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  52. Molnar I, Gibson DM, Krasnoff SB (2010) Secondary metabolites from entomopathogenic Hypocrealean fungi. Nat Prod Rep 27(9):1241–1275. https://doi.org/10.1039/C001459C

    Article  CAS  PubMed  Google Scholar 

  53. Wang X, Gong X, Li P, Lai D, Zhou L (2018) Structural diversity and biological activities of cyclic depsipeptides from fungi. Molecules 23(1):169. https://doi.org/10.3390/molecules23010169

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  54. Yokeswari NP, Mary JKS, Mohan VR (2018) Assessment of bio-active components present in the whole plant extract of Catharanthus pusillus. Int J Creat Res Thoughts (IJCRT) 6(1):406–612

    Google Scholar 

  55. Sogan N, Kapoor N, Kala S, Patanjali PK, Nagpal BN, Vikram K, Valecha N (2018) Larvicidal activity of castor oil nanoemulsion against malaria vector anopheles culicifacies. Int J Mosq Res 5(3):1–6

    Google Scholar 

  56. Sosa A, Costa M, Salvatore A, Bardon A, Borkosky S, Vera N (2017) Insecticidal Effects of Eudesmanes from Pluchea sagittalis (Asteraceae) on Spodoptera frugiperda and Ceratitis capitata. Int J Environ Agric Biotechnol 2(1):361–369. https://doi.org/10.22161/ijeab/2.1.45

    Article  Google Scholar 

  57. Weichen Y, Xiaoyu L, Xiankun L, Yuanqing C, Xiaofei W, Pei L, Rimao H (2022) Synthesis and insecticidal activity of tenuazonic acid and derivatives. Chin J Pest Sci 24(2):260–271. https://doi.org/10.16801/j.issn.1008-7303.2022.0001

    Article  Google Scholar 

  58. Mikami Y, Nishijima Y, Suzuki A (1972) Tenuazonic acids, stereochemical properties and biological activities of tenuazonic acids (Note). J Agric Chem Soc Jpn 46:473–476

    CAS  Google Scholar 

  59. Yamamoto I, Casida JE (1999) Nicotinoid insecticides and the nicotinic acetylcholine receptor. Springer Japan. https://doi.org/10.1007/978-4-431-67933-2

    Article  Google Scholar 

  60. Cazal CM, Batalhão JR, Domingues VC, Bueno OC, Filho ER, Forim MR, da Silva MFGF, Vieira PC, Fernandes JB (2009) High-speed counter-current chromatographic isolation of ricinine, an insecticide from Ricinus communis. J Chromatogr A 1216:4290–4294. https://doi.org/10.1016/j.chroma.2009.02.008

    Article  CAS  PubMed  Google Scholar 

  61. Mizrachi N, Levy S, Goren Z (2000) Fatal poisoning from nicotiana glauca leaves: identification of anabasine by gas-chromatography/mass spectrometry. J Forensic Sci 45(3):736–741. https://doi.org/10.1520/JFS14761J

    Article  CAS  PubMed  Google Scholar 

  62. Zammit M, Shoemake C, Attard E, Azzopardi LM (2014) The Effects of Anabasine and the Alkaloid Extract of Nicotiana glauca on Lepidopterous Larvae. International Journal of Biology 6(3):46–53. https://doi.org/10.5539/ijb.v6n3p46

    Article  Google Scholar 

Download references

Acknowledgements

The authors thank Dr. Ahmed Ramadan El-Rokh, Plant Protection Research Institute, Mansoura Branch, Agriculture Research Center, for his help to the identification of extracted secondary metabolites from wild plants.

Funding

There is no fund for this research.

Author information

Authors and Affiliations

  1. Cotton Pesticides Evaluation Research Department, Plant Protection Research Institute, Agricultural Research Center, Dokki, Giza, Egypt

    Reda R. H. Abdullah

  2. Piercing and Sucking Insect Research Department, Plant Protection Research Institute, Agricultural Research Center, Dokki, Giza, Egypt

    A. H. Abd El-Wahab

  3. Horticultural Insects Research Department, Plant Protection Research Institute, Agricultural Research Center, Dokki, Giza, Egypt

    Samira A. Abd El-Salam

Authors
  1. Reda R. H. Abdullah
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. A. H. Abd El-Wahab
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Samira A. Abd El-Salam
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

RRHA performed the culturing of the fungal strains, extracted the ethyl acetate extract of the tested fungal cultures, performed the estimation of insect enzyme experiment, and prepared the larvae for histological processing. AHA performed the rearing of the fall armyworm. SAA performed by collecting the wild plants and extracting the alkaloid extract of the tested wild plants. All authors detected toxic effects from the extracts, wrote, read, and approved the final manuscript.

Corresponding author

Correspondence to Reda R. H. Abdullah.

Ethics declarations

Ethics approval and consent to participate

Not applicable.

Consent for publication

All authors have approved the manuscript and agree with its submission to Beni-Suef University Journal of Basic and Applied Sciences.

Competing interests

The authors declare that they have no competing interests.

Additional information

Publisher's Note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Rights and permissions

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/.

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Abdullah, R.R.H., Abd El-Wahab, A.H. & Abd El-Salam, S.A. Insecticidal activity and possible modes of action of secondary metabolites of some fungal strains and wild plants as natural pesticides against Spodoptera frugiperda. Beni-Suef Univ J Basic Appl Sci 13, 9 (2024). https://doi.org/10.1186/s43088-024-00467-z

Download citation

  • Received:

  • Accepted:

  • Published:

  • DOI: https://doi.org/10.1186/s43088-024-00467-z

Keywords