Lethal toxicity of six natural and organic insecticidal products namely, Darekastra, Dashparni, fermented buttermilk, fermented cow urine, Tamarlassi and vermiwash to second instar nymphs of greenhouse whitefly, Trialeurodes vaporariorum was evaluated using leaf dip method of bioassay. The insecticidal products evaluated at six concentrations ranging from 1.25 to 40.00% proved to be progressively more lethal to second instar nymphs with an increase in concentration resulting in mortality varying from 1.67 to 7.33% and 43.33 to 62.96 at concentrations of 1.25 and 40.00%, respectively. Tamarlassi proved most toxic against the nymphs. The order of toxicity was Tamarlassi = fermented buttermilk ≥ fermented cow urine ≥ Dashparni ≥ Darekastra ≥ vermiwash.
Research Article
Toxicity of natural and organic insecticidal products to greenhouse whitefly, Trialeurodes vaporariorum (Westwood) (Aleyrodidae: Hemiptera)
https://doi.org/10.21203/rs.3.rs-2451274/v1
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Greenhouse whitefly, Trialeurodes vaporariorum (Westwood) (Hemiptera : Aleyrodidae) is a serious pest under protected environment in temperate regions and in field crops where the summers are warm enough (Byrne and Bellows 1991; Sood and David 2012). It is highly polyphagous and is widely distributed throughout the world. Currently, greenhouse whitefly is known to exist on 859 plant species belonging to 469 genera and 121 families (CABI 2021). The nymphs and adults of T. vaporariorum suck the phloem cell sap thereby affecting the plant vigour. Additionally, they secrete honeydew, which facilitates the development of sooty mould interfering with photosynthetic activity. Apart from the whitefly T. vaporariorum, aphid Myzus persicae (Sulzer), two spotted spider mite Tetranychus urticae Koch, russet mite Aculops lycopersicae (Massee) are of importance under protected environment in Himachal Pradesh (Kashyap et al. 2015; Ghongade and Sood 2019; Ghongade and Sood 2021; Sharma et al. 2021).
Trialeurodes vaporariorum breeds throughout the year and completes 13 generations under protected environment inflicting losses to the extent of 50 % in different crops (Sood et al. 2014; Sood et al. 2018). The greenhouse whitefly feeds on the abaxial surface of the leaves and remains sessile during immature stages that minimize its contact with the insecticides thereby resulting in frequent insecticidal applications, which leads to pesticide residues on crop produce, killing of non-target organisms and development of resistance in T. vaporariorum (Van Lenteren 2000; Sood et al. 2006; Pilkington et al. 2010; Pappas et al. 2013). Now a days more emphasis is being laid on the use of natural products in plant protection. Natural products are effective in enhancing crop productivity, suppressing growth of insects, are inexpensive, easily biodegradable, have different modes of action and are less harmful to non-target organisms and the environment (Lengai et al. 2020). However, research on evaluation of natural and organic insecticidal products is lacking against greenhouse whitefly. Therefore, considering the economic importance of greenhouse whitefly as well as that of natural products (cattle and plant-based) against insect pests, studies were made on evaluating toxicity of natural insecticidal products against second instar nymphs of greenhouse whitefly under laboratory conditions.
Raising stock culture of greenhouse whitefly
Trialeurodes vaporariorum was reared on potted French bean plants raised in a soil-less medium comprising cocopeat, perlite, vermiculite and vermicompost (3:1:1:1). Field-collected population of greenhouse whitefly was used to raise the stock culture under laboratory conditions. The adults were collected with the help of an aspirator and were released in large numbers on 15-day-old French bean plants kept in screened cages (45×45×55 cm) for 24 hours for egg deposition. Thereafter the adults were removed and the plants were shifted to rearing cages for raising to desired developmental stage.
Natural and organic insecticidal products evaluated: Six natural and organic products namely, Darekastra, Dashparni, fermented buttermilk, fermented cow urine, Tamarlassi and vermiwash were evaluated for their lethal toxic effects against 2nd instar nymphs of T. vaporariorum. The products were prepared following the standardized methodologies of Department of Organic Agriculture and Natural Farming (Rameshwar et al. 2014). The composition of products is being given in Table 1
Table 1. Natural and organic products evaluated for insecticidal activity
|
Natural/Organic Product |
Composition |
|
Darekastra |
Melia leaves (5 kg), cow urine (5 L), cow dung (2 kg) and water (20 L) |
|
Dashparni |
Melia leaves (5 kg), Ipomea leaves (2 kg), Polygonum leaves (2 kg), Juglans leaves (2 kg), Lantana leaves (2 kg), Datura leaves (2 kg), Vitex leaves (2 kg), Psidium leaves (2 kg), Eupatorium leaves (2 kg), wild Tagetes leaves (2 kg), garlic paste (500 gm), red chilli paste (500 gm), turmeric powder ( 500 gm), dry ginger powder (200 gm), cow urine (10 L), cow dung (2 kg) and water (10 L) |
|
Fermented buttermilk |
Prepared from 15-20 days old cultured milk |
|
Fermented cow urine |
15 days old cow urine of Indian cow |
|
Tamarlassi |
Prepared by keeping the fermented buttermilk in copper pot for 10 to 15 days |
|
Vermiwash |
Cow dung (4 kg), biomass (1 kg) and adult earthworms (200-300) |
Each insecticidal product was regarded as 100% stock solution and the test concentrations were further prepared by serial dilution method.
Method of Bioassay
Leaf dip method of bioassay was used for evaluating toxicity of natural and organic insecticidal products against second instar nymphs. For obtaining the cohort of second instar nymphs, French bean plants were exposed to adult whiteflies for 24 hours for egg-laying. The eggs were allowed to develop to 2nd instar nymphs. After 13-14 days, the eggs develop into 2nd instar nymphs. The leaves containing the nymphs were detached and examined under a microscope to validate the stage. The leaves containing the second instar nymphs (n > 30) were excised into discs (2.5 cm diameter). These leaf discs were dipped in test concentrations of different products ranging at six concentrations of 1.25, 2.50, 5.00, 10.00, 20.00 and 40.00 % for 30 seconds. An untreated check (UTC) was maintained by dipping the leaf discs in water. The discs were air-dried at room temperature and placed on an agar-agar bed (2.00 %) in petri plates (5 cm dia.) with abaxial surface facing upwards. Each experiment was replicated three times. These petri plates were placed under controlled conditions at 25±1℃ temperature and 70±5 % relative humidity. Observations on mortality of nymphs were recorded upto 72 hours after treatment. The nymphs that shrunk, dried and turned brown after treatment were considered dead. Mortality data recorded upto 72 hours after treatment was taken into consideration for working out per cent mortality. The per cent mortality was corrected as suggested by Abbott (1925) using the following formula:
Natural and organic products resulted in mortality of second instar nymphs of T. vaporariorum ranging from 2.08 to 47.78, 3.33 to 50.07, 6.00 to 61.11, 5.00 to 54.07, 7.33 to 62.96 and 1.67 to 43.33 % for Darekastra, Dashparni, fermented buttermilk, fermented cow urine, Tamarlassi and vermiwash, respectively, in the concentration range of 1.25 to 40.00 % (Table 2). Nymphal mortality increased significantly with an increase in evaluated concentrations. Also, a relationship was established by working out correlation (r) between concentration and nymphal mortality which resulted in the corresponding ‘r’ values 0.96, 0.95, 0.96, 0.96, 0.96 and 0.96, respectively, indicating a positive and significant (P=0.05) effect of concentration on bringing about the kill. Based on the mean mortality observed in different products, Tamarlassi resulted in maximum mortality and was at par to fermented buttermilk. It was followed by fermented cow urine. Vermiwash resulted in minimum mortality among the second instar nymphs and was preceded by Darekastra and Dashparni, all being at par to each other. However, the interaction effect among the concentration and natural and organic products was found to be non-significant.
Table 2. Toxicity of natural and organic products to second instar nymphs of Trialeurodes vaporariorum
|
Concentration (%) |
Corrected mortality (%) 72 hours after treatment |
Mean |
|||||
|
Darekastra |
Dashparni |
Fermented buttermilk |
Fermented cow urine |
Tamarlassi |
Vermiwash |
||
|
1.25 |
2.08 (4.82) |
3.33 (8.61) |
6.00 (14.09) |
5.00 (12.92) |
7.33 (15.36) |
1.67 (4.31) |
4.24 (10.02) |
|
2.50 |
5.59 (13.52) |
6.67 (14.79) |
9.33 (17.70) |
8.00 (16.07) |
10.67 (18.98) |
4.89 (12.68) |
7.52 (15.62) |
|
5.00 |
11.11 (19.16) |
12.22 (20.41) |
15.56 (23.19) |
14.44 (22.30) |
17.14 (24.35) |
10.00 (18.43) |
13.41 (21.31) |
|
10.00 |
21.33 (27.48) |
24.67 (29.69) |
29.33 (32.77) |
25.33 (30.19) |
32.00 (34.41) |
20.05 (26.51) |
25.45 (30.18) |
|
20.00 |
36.67 (37.24) |
40.00 (39.21) |
46.67 (43.07) |
42.22 (40.48) |
48.89 (44.35) |
32.22 (34.53) |
41.11 (39.81) |
|
40.00 |
47.78 (43.71) |
50.07 (45.02) |
61.11 (51.40) |
54.07 (47.32) |
62.96 (52.55) |
43.33 (41.15) |
53.22 (46.86) |
|
Mean
|
20.76 (24.32) |
22.83 (26.29) |
28.00 (30.37) |
24.85 (28.21) |
29.83 (31.67) |
18.69 (22.93) |
|
|
Figures in parentheses are the arc sine transformed values |
|
C.D. (P=0.05) |
|
Concentration (A) = (2.19) |
|
Product (B) = (2.19) |
|
A × B = NS |
Although the literature on the bioactivity of natural and organic products against insects is limited, there are a few references available on their potential bioactive components responsible for their activity against various insect pests. The insecticidal activity of Darekastra comprising Melia leaves can be attributed to the biologically active compounds, triterpenoids in Melia azedarach. The insecticidal properties of the triterpenoids have been demonstrated on coleopteran, dipteran, and lepidopteran pests under laboratory conditions. (Carpinella et al. 2003). Ayil-Gutierrez et al. (2018) demonstrated that secondary metabolites from M. azedarach, such as coumarin, sterols, and terpenes, reduced pupal viability, resulting in adult deformities in Spodoptera litura. Thakur and Sood (2022) found Darekastra to be the best-performing bioacaricide for maintaining spider mite population at lower levels when used both as preventive and curative spray under greenhouse conditions. Dashparni ark has been known to contain phytochemicals such as saponin, proteins, phenol, steroid and phytosterol having protection activity (Kasarkar et al. 2021). Chandel et al. (2011) observed that the extracts of Azadirachta indica and Vitex negundo were more effective against Bagrada hilaris, with mortality rates of 80.9 % in nymphs and 74.9 % in adults, respectively. Joshi et al. (2021) observed Dashparni ark to the be most effective bioformulation against Agrotis ipsilon causing maximum mortality among different larval instars.
Bioactive constituents in cow urine affect insect development and survival (Gahukar 2013). Cow urine contains high content of urea which is toxic to most insect pests and can be considered a good means for insect pest management (Kumawat et al. 2014). Onunkun (2014) reported cow urine + water (1:1) to be the most effective insecticidal solution for the management of brown stink bug, Aspavia armigera, green stink bug, Nezara virudula and the coreid bug, Clavigralla tomentosiccolis in Amaranthus. Enrichment of cow urine with various botanicals enhanced the insecticidal property of different plant extracts against sucking pests of cotton (Patel et al. 2003) and sesame (Ahirwar et al. 2010). Cow urine, Darekastra and fermented buttermilk proved to be potent acaricides with deterrent properties against Tetranychus urticae (Thakur and Sood 2021).
Cow milk-based fermented products like fermented buttermilk have acidic properties (Gebreselassie et al. 2016) and Tamarlassi contains acidic buttermilk enriched with copper (Sudeshna 2017). Karthikeyan et al. (2005) found buttermilk to cause a 60 % reduction in whitefly infestation in bhendi. Fermented buttermilk and Tamarlassi have also been reported to possess acaricidal activity against mites (Thakur and Sood 2019). The coelomic fluid of earthworms in vermiwash is reported to possess insecticidal, antifungal and pesticidal bioactive compounds (Nandana et al. 2020). Vermiwash is toxic to the survival of insects (Sayyad 2017). It possesses excellent biopesticidal properties (Nayak et al. 2019) and is effective against aphids (Venkateswarlu et al. 2011).
The study concludes that the natural and organic insecticidal products assessed had toxic effects against the second instar nymphs of Trialeurodes vaporariorum. The order of toxicity was Tamarlassi = fermented buttermilk ≥ fermented cow urine ≥ Dashparni ≥ Darekastra ≥ vermiwash. Hence, Tamarlassi and vermiwash resulted in the maximum and the minimum mortality of the second instar nymphs, respectively.
Acknowledgements
The authors would like to thank ICAR “National Agricultural Higher Education Project - Centre of Advanced Agricultural Science and Technology” (NAHEP-CAAST) for providing financial support to undertake this study.
Conflict of interest
The authors declare that there is no conflict of interest in this research paper.
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