Research Article |
Corresponding author: Pratik Doshi ( pratik.doshi159@gmail.com ) Academic editor: Katalin Szlavecz
© 2018 Pratik Doshi, Anett Mészárosné Póss, Ferenc Tóth, Márk Szalai, György Turóczi.
This is an open access article distributed under the terms of the Creative Commons Attribution License (CC BY 4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
Citation:
Doshi P, Póss AM, Tóth F, Szalai M, Turóczi G (2018) Effect of neem-derived plant protection products on the isopod species Porcellionides pruinosus (Brandt, 1833). In: Hornung E, Taiti S, Szlavecz K (Eds) Isopods in a Changing World. ZooKeys 801: 415-425. https://doi.org/10.3897/zookeys.801.25510
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Neem-based products have gained major attention over the last few years due to their wide range of applications in pest management, and have been in the focus of biological plant protection research in the past decade. Yet, there is limited information available to understand the side effects of these neem-derived pesticides on non-target species in soil. Therefore, Porcellionides pruinosus, a terrestrial isopod, was chosen as a non-target species to investigate such possible effects. Two different experiments were conducted to study two different neem-derived plant protection products, i.e., NeemAzal T/S (1% azadirachtin) which is a commercial product registered in the EU, and neem leaf extract from dried neem leaves (1%).The latter simulates the plant protection product, is domestically produced, and widely used by farmers in India and other tropical and subtropical countries. Findings are consistent with previous results obtained with other non-target organisms, i.e., neither of the tested neem products have adverse effects on the mortality of P. pruinosus. However, further research on a wider range of soil organisms is needed to prove the safety of neem-based products as biological control agents and to be part of integrated pest management.
azadirachtin, biological pest control, isopod, neem leaf extract, non-target organism
The safe use of neem (Azadirachta indica)-based biological insecticides requires more information about their possible side effects on non-target organisms. Such organisms are the woodlice species, important decomposers of organic material in agricultural areas. It was demonstrated that neither a commercial product of neem nor a domestic neem leaf extract had any adverse effect on a ubiquitous woodlice species, Porcellionides pruinosus.
Different environmental conditions and chemical stressors may interact and can have a negative impact on the soil biota (
The neem tree, Azadirachta indica is also referred to as Melia azadirachta L., Indian lilac or Margosa (
Azadirachtin is a tetranortriterpenoid plant limonoid which possess anti-feedant and growth-disrupting properties. It was first isolated from seeds of Azadirachta indica by Butterworth and Morgan in 1968 and its detailed structure was given by Broughton and his team in 1987 (
Neem extracts and products have been used against different orders of insects such as Coleoptera (for example
The effect of neem-derived products on non-target organisms has also been studied. However, information on the effect of neem-based pesticides and formulations on non-target organisms of soil biota are still limited.
Woodlice species (Isopoda, Oniscidae) are ubiquitous saprophagous members of the soil fauna (
In another study, single and combined toxicity of atrazine, dimethoate, lindane, zinc and cadmium were tested in Porcellionides pruinosus (Brandt, 1833) and Enchytraeus albidus (Henle, 1837) an annelid, commonly known as white worm, using avoidance as test parameter. For both the species, patterns of antagonism were found when exposed to dimethoate and atrazine, synergism for lindane, dimethoate, and atrazine, synergism for lindane and dimethoate exposures and concentration addition for cadmium and zinc occurred, while the exposure to cadmium and dimethoate showed dissimilar patterns (
This soil ’cleaning’ result can be defined as a positive ecosystem service (ES) (
A small-scale terrestrial ecosystem containing soil collected from an agricultural field in Central Portugal was used to evaluate the effects of the combination of the herbicide glyphosate and the insecticide dimethoate in another study. The application of dimethoate led to a decrease in feeding activity in all concentrations tested. The mortality of isopods exposed to dimethoate in single and binary exposures was high. Exposure to dimethoate decreased the acetylcholinesterase activity of isopods (
In this paper we present results on the side-effect of NeemAzal T/S and neem leaf extract on the terrestrial isopod species Porcellionides pruinosus. We selected P. pruinosus as a non-target organism, because it is ubiquitous and it occurs in anthropogenic environments, where pest control is applied. They play a vital role in the fragmentation and decomposition process of leaf litter, thereby causing changes in soil quality and soil services (
The methodology of
Collection of isopod species: Porcellionides pruinosus adults were collected from Regional Waste Management Center Pusztazámor, Hungary, by hand sorting. Isopods were bred and maintained at the Institute of Plant Protection of Szent István University, Gödöllő, Hungary. Species level identification was based on the taxonomic key developed by Brandt (1833) (
Preparation of neem leaf extract: For neem leaf extract, air-dried neem leaves were obtained from local growers in India, Maharashtra, Konkan Division, Mumbai Suburban area. A stock concentration of 1% was prepared by soaking 1g of crushed dried neem leaves in 100 ml distilled water overnight and then filtered using a non-sterile filter paper. Different working concentrations (0.05, 0.1, 0.25, 0.5, 0.75, and 1%) of neem leaf extract were prepared from 1% stock solution using distilled water in the laboratory and were used on the same day. Generally the applied dosage used by local growers in India has a maximum concentration of 1%. In this experiment, we tried to model the concentration used by the local growers in the field conditions.
Preparation of azadirachtin: NeemAzal T/S (Trifolio-M GmBH), a commercial product containing 1% azadirachtin, registered in the EU, was used. A stock concentration of 1% azadirachtin was prepared (from NeemAzal T/S which is 1% azadirachtin) by diluting 1 ml NeemAzal T/S in 100 ml of distilled water which equals to 0.01% azadirachtin. It was further diluted to get the 0.0005, 0.001, 0.0025, 0.005, 0.0075 and 0.01% azadirachtin concentrations respectively and was used on the same day. The registered dosage of azadirachtin ranges from 0.0025 to 0.005%, depending on the plant culture in the EU.
A control with only distilled water was used for both experiments. The working concentrations and distilled water were sprayed using a hand sprayer under laboratory conditions.
Experimental design: Five adults of Porcellionides pruinosus were placed in glass Petri dishes (13 cm in diameter), with 1 g of commercial horticultural soil (pH = 7.0) and approximately 1 g of fresh potato as a food source. Two milliliters of different working concentrations of neem leaf extract and azadirachtin were sprayed using a hand sprayer. After spraying, the Petri dishes were kept in the dark, checked after time periods of 1, 24, 48, 72, 96, and 120 hours post-application of neem leaf extract and azadirachtin respectively, and mortality data was recorded. The mortality data obtained after 120 hrs was subjected to statistical analysis using R software (R Core Team 2017). Logistic regressions were fitted (as the response was binary, i.e., the isopods were either dead or alive) to check the effect of the two different products on isopod mortality. To test whether the concentrations have significant effect on mortality, chi-squared tests were performed on model deviances. Prior to running the tests model diagnostic plots were investigated to assess homoscedasticity and residual normality (
The mortality of P. pruinosus was generally low in all treatments. In case of azadirachtin, even after 120 hours zero mortality was observed in seven replicates of 0.0005% concentration, eight replicates at 0.001%, nine replicates of 0.0025%, seven replicates of 0.005 and 0.0075% each, and four replicates of 0.01%.
The same was observed in the case of neem leaf extract, after the time period of 120 hours: zero mortality in case of five replicates of 0.05% concentration, nine replicates of 0.1%, six replicates of 0.25%, seven of 0.5%, four replicates of 0.75%, and five replicates of 1% (see Table
The mortality slightly increased with the concentration but this observed increment was not statistically significant (Table
Different concentrations of NeemAzal T/S and neem leaf extracts were compared to check their respective effects on the mortality of the isopods. Neither azadirachtin nor neem leaf extract affected the observed isopod mortality (p-values are 0.43 and 0.39 and McFadden’s pseudo R2: 0.04 for azadirachtin, 0.05 for neem leaf extract respectively; Figs
Mortality rate of the isopod Porcellionides pruinosus after 120 hours at different concentrations of NeemAzal T/S (1% azadirachtin). The vertically jittered circles (to avoid perfect overlapping) indicate the individual isopods whereas the line indicates the trend of the mortality with respect to increasing concentrations and the grey area represents the 95% confidence level.
Mortality rate of the isopod Porcellionides pruinosus after 120 hours to at different concentrations of neem leaf extract. The vertically jittered circles (to avoid perfect overlapping) indicate the individual isopods whereas the line indicates the trend of the mortality with respect to increasing concentrations and the grey area being the 95% confidence level.
Effect of NeemAzal on the mortality of Porcellionides pruinosus expressed as cumulative mean for different time intervals.
Treatment | conc (%) | Mean mortality rate after time interval | ||||||
---|---|---|---|---|---|---|---|---|
1 hr | 24 hrs | 48 hrs | 72 hrs | 96 hrs | 120 hrs | |||
azadirachtin (NeemAzal T/S) | 0 | mean | 0.1 | 0.1 | 0.1 | 0.1 | 0.1 | 0.2 |
SD | 0.32 | 0.32 | 0.32 | 0.32 | 0.32 | 0.42 | ||
0.0005 | mean | 0.2 | 0.3 | 0.3 | 0.3 | 0.3 | 0.5 | |
SD | 0.42 | 0.67 | 0.67 | 0.67 | 0.67 | 0.85 | ||
0.001 | mean | 0 | 0.3 | 0.3 | 0.3 | 0.4 | 0.4 | |
SD | 0 | 0.95 | 0.95 | 0.95 | 0.97 | 0.97 | ||
0.0025 | mean | 0 | 0 | 0 | 0 | 0 | 0.1 | |
SD | 0 | 0 | 0 | 0 | 0 | 0.32 | ||
0.005 | mean | 0.1 | 0.2 | 0.2 | 0.2 | 0.2 | 0.3 | |
SD | 0.32 | 0.42 | 0.42 | 0.42 | 0.42 | 0.48 | ||
0.0075 | mean | 0.2 | 0.2 | 0.2 | 0.2 | 0.2 | 0.3 | |
SD | 0.42 | 0.42 | 0.42 | 0.42 | 0.42 | 0.48 | ||
0.01 | mean | 0 | 0.1 | 0.5 | 0.5 | 0.5 | 0.7 | |
SD | 0 | 0.42 | 0.71 | 0.71 | 0.71 | 0.67 |
Effect of neem leaf extract on the mortality of Porcellionides pruinosus expressed as cumulative mean for different time intervals.
Treatment | conc (%) | Mean mortality rate after time interval | ||||||
1 hr | 24 hrs | 48 hrs | 72 hrs | 96 hrs | 120 hrs | |||
neem leaf extract | 0 | mean | 0 | 0 | 0 | 0 | 0.1 | 0.33 |
SD | 0 | 0 | 0 | 0 | 0.33 | 0.5 | ||
0.05 | mean | 0 | 0 | 0 | 0.1 | 0.2 | 0.527 | |
SD | 0 | 0 | 0 | 0.32 | 0.42 | 0.52 | ||
0.1 | mean | 0 | 0 | 0 | 0 | 0 | 0.1 | |
SD | 0 | 0 | 0 | 0 | 0 | 0.32 | ||
0.25 | mean | 0 | 0 | 0 | 0 | 0.2 | 0.4 | |
SD | 0 | 0 | 0 | 0 | 0.42 | 0.52 | ||
0.5 | mean | 0 | 0 | 0 | 0 | 0 | 0.4 | |
SD | 0 | 0 | 0 | 0 | 0 | 0.7 | ||
0.75 | mean | 0 | 0.1 | 0.2 | 0.2 | 0.2 | 0.7 | |
SD | 0 | 0.32 | 0.42 | 0.42 | 0.42 | 0.67 | ||
1 | mean | 0 | 0.1 | 0.2 | 0.3 | 0.4 | 0.7 | |
SD | 0 | 0.32 | 0.42 | 0.67 | 0.7 | 0.95 |
While there are numerous literature references available on the effect of neem and neem-derived products on target organisms, some of the studies reported data on non-target organisms as well. For instance,
From our results it can be concluded that neither NeemAzal T/S nor neem leaf extracts pose any risk to the terrestrial isopod species studied in the tested concentrations. However, further research is needed to test the possible effect of various neem products on the members of the soil fauna. Also, it can be concluded that NeemAzal T/S and domestic neem leaf extract do not differ in respect to their mortality effects on P. pruinosus.
PD and AMP designed and conducted the experiments. MS analyzed the data. PD wrote the manuscript. AMP, FT, GT, and MS gave their feedback. All authors read and approved the manuscript.
The first author wishes to thank Tempus Public Foundation, Government of Hungary for doctoral scholarship (Stipendium Hungaricum Scholarship Program Reference number 2016/India/80903). The authors also wish to thank the assistance of Szőcs Tündér Ilona for her help. Our work was supported by grants received from ÚNKP-17-3 New National Excellence Program of the Ministry of Human Capacities and by the EFOP-3.6.3-VEKOP-16-2017-00008 project (co-financed by the European Union and the European Social Fund). This research was supported by the Higher Education Institutional Excellence Program (1783-3/2018/FEKUTSTRAT) awarded by the Ministry of Human Capacities within the framework of plant breeding and plant protection researches of Szent István University.