Indian Journal of Agricultural Research

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Indian Journal of Agricultural Research, volume 56 issue 6 (december 2022) : 741-745

The Bio-Insecticidal Activity of Papaya (Carica papaya L.) Leaves Extract against Spodoptera litura Fabr. (Lepidoptera: Noctuidae) Larval Growth

Sofia Ery Rahayu1,4,*, Amin Setyo Leksono2, Zulfaidah Penata Gama2, Hagus Tarno3
1Doctoral Program, Department of Biology, Faculty of Mathematics and Natural Sciences, Universitas Brawijaya, Malang-65145, East Java, Indonesia.
2Department of Biology, Faculty of Mathematics and Natural Sciences, Universitas Brawijaya, Malang-65145, East Java, Indonesia
3Department Plant and Disease, Faculty of Agriculture, Universitas Brawijaya, Malang-65145, East Java, Indonesia.
4Department of Biology, Faculty of Mathematics and Natural Sciences, Universitas Negeri Malang, Malang-65145, East Java, Indonesia.
Cite article:- Rahayu Ery Sofia, Leksono Setyo Amin, Gama Penata Zulfaidah, Tarno Hagus (2022). The Bio-Insecticidal Activity of Papaya (Carica papaya L.) Leaves Extract against Spodoptera litura Fabr. (Lepidoptera: Noctuidae) Larval Growth . Indian Journal of Agricultural Research. 56(6): 741-745. doi: 10.18805/IJARe.AF-747.
Background: Papaya leaves produce several metabolites that may be toxic to an insect pest. In East Java, it is easy to find two cultivars in the yard, plantation, and moors, such as Thailand and local papaya. However, information about their utilization as bio-pesticides remains limited. This study aimed to investigate the relationship between those papaya cultivars and the effect of their extracts on Spodoptera litura larvae.

Methods: The Thailand papaya cultivar (TPC) and Indonesian papaya cultivar (IPC) leaves were obtained from the medium and lowlands and were macerated with methanol until obtained a papaya leaves extract (PLE). The S. litura larvae on the instar phase 2 (n= 630) were used for treatment. The Ricinus communis leaves were sprayed with PLE concentrations of 0, 10, 20, 30, 40 and 50% and fed to S. litura larvae. 

Result: Based on the secondary metabolites, the papaya cultivars form two clusters associated with its origin. The 40 and 50% concentrations of PLE significantly affect the mortality and head-capsule width of S. litura indicated with broken pupae and failure to transform to imago. This result signifies the potential of papaya leaves extract as bioinsecticide for S. litura larvae.

Spodoptera litura is a notorious leaf-feeding insect pest of many economically valuable crops in the Asia-Pacific region. The S. litura larvae attack more than 122 host plant species, including cotton, corn and soybean. The S. litura attack may cause leaf and pod damage and lead to a 68% reduction in soybean production (Ayudya et al., 2019; Motaphale et al., 2017). In Indonesia, S. litura has been distributed in 22 provinces and is reported to attack 11,163 ha/year, including soybean and mung bean plants (Melani et al., 2016; Indiati et al., 2017).
       
Most farmers apply chemical insecticide to control S. litura. However, chemical insecticide carries negative impacts, such as inducing pest resistance and population outbreak (Nandhakumar et al., 2020). Consequently, natural pesticides from plants can be used as an alternative pest control because they are more environmentally friendly character than those of chemical sources.
       
Indonesia as a tropical country has highly diverse plants used as natural pesticides. One of those plants is papaya (Carica papaya L.), which is massively cultivated in medium and lowlands in East Java, Indonesia. Papaya has many cultivars, such as the IPC and TPC, with the IPC having the purple petiole as its main characteristic. The previous studies demonstrated that IPC and TPC have different active compounds, including phenols, alkaloids and flavonoids that might be beneficial for inhibiting the growth and development of insects (Matsuura and Fett-Neto 2015; Aihetasham et al., 2017; Rahayu et al., 2020).

In addition, investigating the variety between family Caricaceae members mainly focused on its genetic features and morphology. However, the variety and relationship between the TPC and IPC based on their active compounds from different elevations have not been examined. Besides, there is lack of evidence about comparing the effectiveness of both PLE on the development and growth of S. litura. Therefore, this study focuses on investigating the relationship between IPC and TPC based on their active  compounds, different elevations and PLE effectiveness in the development of S. litura larvae. Furthermore, this study aims to discover the variety and potential of that cultivar as a natural pesticide.

Plant materials
 
The extract was prepared from the IPC and TPC leaves obtained from Malang and Nganjuk, East Java, Indonesia. The lowland was selected at Nganjuk at 70 m above sea level (S 07036.560'E 112001.290'), while the medium land was selected at Malang at 414 m above sea level (S 08005. 551'E 112041.175'). The leaves were washed, dried and mashed using a dry blender.
 
Preparation of papaya leaf extract
 
Papaya leaf simplicial was extracted using the maceration method with methanol solvent at a 1:10 ratio. The homogenate obtained was filtered using Whatman paper, and the residue was macerated again for 24 hours. The filtered results were mixed and concentrated by rotary evaporator. The PLE was stored at 4°C. The phytochemical screening was carried out using liquid chromatography mass spectrometry (LCMS).
 
Development and growth test of S. litura larvae
 
The experiment was conducted in the Department of Biology, Universitas Negeri Malang during June 2018 until June 2019. S. litura instar 2nd larva was left starving for 4 hours and placed in the R. communis leaves for four days that have sprayed with 0% (control), 10%, 20%, 40% and 50% PLE of IPC and TPC. After that, the larva was given fresh R. communis leaves with no extract. Each extract concentration was tested into ten larvae three times, and each larva was maintained individually (n=630 larvae). The larvae mortality was examined from 24-96 hours post-treatment. Each of the larvae head-capsule widths was measured. The survived larvae were taken care of until they grew into a pupa. The S. litura pupa formed in 24 hours was weighted and observed (died or alive). The pupa was placed into a plastic glass filled with sterilized sand. They were investigated until they became imago. The hatched imago’s morphology was also examined and counted.
 
Data analysis
 
The active compounds of PLE were analyzed descriptively to construct a dendrogram using Bray-Curtis analysis. The larvae head-capsule width, pupa weight and condition were analyzed using ANOVA (p<0.05) followed by the Duncan test if a palpable different analysis result was obtained.

Dendrogram of TPC and IPC
 
The phytochemical screening using LCMS demonstrated that there were 62 compounds founds on TPC, while 60 were found on IPC. In comparison, two compounds were only detected on the TPC than IPC, such as p-coumaric acid and 5,7 Dimethoxycoumarin. In contrast, the IPC has four unique compounds found than TPC, including phellandrene, myrcene, linalool oxide, and caryophyllene. Similarly, a previous study reveals that the active compounds in those two cultivars differed (Rahayu et al., 2020). Carpaine, pseudocarpaine, dehydrocarpaine I and II, quercetin, kaempferol, luteolin-7-glucoside, isoquercitrin, gallic acid, and benzyl glucosinolate are found with a high abundance in the PLE. These similar and different compounds obtained were used to construct a dendrogram to investigate the relationship between those cultivars, as presented in Fig 1.
 

Fig 1: Dendrogram of two papaya cultivars from different height habitats based on their active compounds.

        

IPC and TPC obtained from medium areas form one cluster. The obtained similarity value from the two clusters is relatively high (95.25%). The results of Bay-Curtis analysis on the different and similar active compounds content of the PLE from IPC and TPC showed values closer to 100%, indicating their closer kinship. Further, a dendrogram constructed from the data shows that those two cultivars form two clusters based on its country’s origin. This high similarity was observed because they are from the same species of C. papaya, although there were from different cultivars. 

       
The Genus Carica is monotypic and only consists of one species, namely C. papaya, so the IPC and TPC formed one cluster in the dendrogram. Meanwhile, the morphology analysis shows that papaya has phenotypic variation in its morphological characteristics and plant stature (de Oliveira et al., 2010).
 
Larval mortality of S. litura
 
The results suggest that the treatment of PLE from IPC and TPC was not significantly different. However, the different concentrations affect the mortality of S. litura larvae. The PLE 40% and 50% have significantly different mortality percentages among other concentrations. More prolonged exposure to PLE results in more larvae deaths. Similarly, a higher concentration also presents a higher mortality rate of S. litura, as shown in Table 1.
 

Table 1: The mortality of S. litura larvae after being treated by the various Papaya leaves extract concentrations.


 
PLE from IPC and TPC affects the mortality of S. litura instar 2. The highest mortality rate of S. litura larvae obtained in this study is below 50% (Table 1). In contrast, antifeedant test results on the S. litura larvae produce a high antifeedant index, 97% and 94%, respectively. Meanwhile, PLE 50% has a 100% antifeedant index (Rahayu et al., 2020). This result implies that PLE from both cultivars may act as an antifeedant. An ethanolic PLE suggested a high repellent activity against German cockroaches (Rahayu et al., 2020) or Tribolium castaneum (Tatun et al., 2014). However, the PLE effect might depend on their active compounds’ abundance.
 
Development and growth of S. litura larvae
 
S. litura larvae were treated with PLE while their growth and development were observed. The growth and development indicator used was the instar larvae head-capsule width. The results showed that the larvae head-capsule width in the instar II and III treated with PLE differs from the control group (Table 2). Meanwhile, the instar IV larvae head-capsule width presents no substantial differences in all concentrations. However, in the instar V phase, the S. litura larvae with PLE 40% and 50% have tighter and significantly different head-capsule widths than the other concentration.
 

Table 2: The head-capsule width of S. litura larvae was treated by the various concentrations of Papaya leaves extract.


       
PLE of IPC and TPC contains active compounds that affect the growth and development of S. litura larvae. The larvae head-capsule width was used to indicate its growth and development. The larvae head-capsule width continuously grows with no expanding size in the same instar phase. Food quality substantially affects insect larval growth (Delbac et al., 2010), including the head-capsule width (Montezano et al., 2014).
       
The S. litura head-capsule width measurement indicates that the older larvae have bigger head sizes with huge cephalic and robust mandibular muscles, affecting the effectiveness of the eating strategy (Calvo and Molina 2008). PLE resulted in 2.6-2.7 mm head-capsule width on the last instar (instar 5) of S. litura. Similar to this result, the head-capsule width of Spodoptera larvae had 2.5-2.7 mm head-width (Montezano et al., 2014). However, the results showed that the PLE 40% and 50% generate significantly different results on the S. litura larvae head-capsule width. PLE produces larvae with varying head-capsule widths due to toxic active compounds, reducing the food quality for the S. litura larvae. 
       
The weight of S. litura pupae have no difference between the control group and other groups treated with different PLE concentrations (Table 3). The average weight of the produced S.litura pupa was 0.30-0.34 grams. Another study also discovered similar results revealing that Spodoptera pupa weight is 0.3-0.4 grams (Montezano et al., 2014).
 

Table 3: Weight of S. litura pupae after treated by various concentrations of Papaya leaves extract.


       
The average number of damaged pupae from the larvae treated with PLE 0% was substantially different from the other groups treated with different PLE concentrations (p<0.05). Similar results have been observed in the number of pupae that failed to hatch into imago from the control group treated with PLE 0%, as shown in Table 4. Active compounds possibly caused the inability of the pupa to hatch in PLE, which affects the metabolism of S. litura. PLE contained flavonoids, alkaloids, phenols, saponins, and tannins (Rahayu et al., 2020).
 

Table 4: The average number of pupae S. litura damaged and fails to hatch into an imago.


       
PLE carries antifeedant function toward S. litura larvae (Rahayu et al., 2020), which might interrupt the physiological function or become toxic for insects, inhibiting insect growth (Kilani-Morakchi et al., 2021). The active compounds in the PLE might work through the food to inhibit the growth and development of S. litura. Flavonoids disturb the essential enzymatic reaction for insect development. Besides, a flavonoid also decreases the plants’ tissue palatability toward herbivorous insects (Marques et al., 2016). A previous study showed that quercetin decreased the percentage of S. litura pupa formation (Rashwan and Hammad 2020). Meanwhile, the phenolic compound could damage the cell in its midgut. Meanwhile, tannin reduces food digestibility by binding the salivary proteins and digestive enzymes, including trypsin and chymotrypsin, to disrupt protein absorption (Rashwan and Hammad 2020).
       
In addition, the alkaloid compound is toxic for S. litura. Alkaloid has a bitter insect taste, decreasing the larvae appetite and leading to malnutrition(Matsuura and Fett-Neto 2015). Alkaloids disturbed the hormonal balance and chitin synthesis mechanism resulting in a smaller cuticle weight and an incomplete of their outer frame of the exoskeleton (Ge et al., 2015; Ileke and Ogungbite 2014). Alkaloids also hinder α-amylase and protease enzymes which in turn disturbing the metabolism of protein, resulting in a lack of amino acid production that is essential for the growth and development of the larvae (Zou et al., 2019; Sharma 2019).
       
The shape and size changes occur during the larvae growing process, involving the cell growth control factor and synthesis process. Ecdysteroids and insulin-like peptides (ILPs) are the growth generator in insects. Disruption of the insulin signaling causes cell size and number changes within the adult insect organs (Lin and Smagghe 2019). The observable indicators in this study are the bend and smaller wings detected in the treatment groups. Therefore, the results suggested that the PLE from the IPC and TPC might be a biopesticide toward the S. litura larvae by disrupting its growth and development process.

Based on its active compounds, papaya cultivars form two clusters associated with the country’s origin. PLE from IPC and TPC carries the same effects on the S. litura larvae mortality and development. The highest S. litura larvae mortality rate and the smallest head capsule width was observed from the treatment using PLE 40% and 50%. Besides, PLE has no effects on the S. litura pupa weight but affects pupal growth and development.
None.

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