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Introduction

A greater part of the natural aquatic environment is faced with the risk of a shrinking genetic base and biodiversity due to the haphazard use of pesticides.¹Pesticides have been used forcenturies in agriculture for increased food production by destroying undesirable bugs and to control disease vectors.²Pesticides that reach the water frame can accumulate in fish and molluscs which might be dangerous to humans whilst ingested.³Fishes are vulnerable totoxic substances and their bioaccumulation causes serious risk to life.⁴The pesticides affect the aquatic environment by disrupting the aquatic food chain leading to the loss/ shiftin enormous amount of natural invertebrate as well as vertebrate life forms in the water habitats.^{5, 6}The toxicity studies are extremely useful for vulnerable species of an ecosystem and can be used to track different forms of pollutants in assessing the indicator organisms. Toxicity tests are usually stated with respect to median lethal concentration LC50. Histopathological assessment has been increasingly recognized as an effective tool for evaluating the environmental effects.^{7, 8}所以叶子有机磷农药残留il and water after exposure for several days, posing a constant threat to non-target species, particularly fish.⁹Malathion is a major source of environmental poisoning in the developing countries.¹⁰Malathion (Diethyl 2-[(dimethoxyphosphorothioyl)sulfanyl]butanedioate) is an organophosphate insecticides widely used in agriculture and house to control variety of insects.^{11, 12}Neem extracts and pure compounds have been evaluated against more than 300 species of insect pests. Neem oil contains several triterpenoids like azadiractin, salanin, alkaloids, flavanoids, glycosides etc.¹³Neem oil was found to be very effective as antirepellent and insecticide.¹⁴It was selected as it is a biodegradable safe, efficient, versatile, ecofriendly, low cost and natural biopesticide.Oreochromis mossambicusa freshwater fish is tolerant to a broad range of environmental conditions. The purpose of this study was to assess whether neem oil can influence malathion and combat with the stress and toxic effects on the fishOreochromismossambicus并阐明不同的目标器官的变化ns like liver and kidney withreference to histopathological and biochemical studies.

Materials and Methods

Oreochromis mossambicuswere collected from the Hydrological Research Station,Chetpet, Chennai in a clean container of 10 litres capacity ensuring that they were not harmed either physically or physiologically during collection and transportation. The fishes were acclimatized to the laboratory condition for a week by keeping them in clean glass containers of 25 litres capacity containing dechlorinated and aerated tap water. Active fishes of almost same size 5-6 cm length were selected for the experiment , precautions were followed as per.¹⁵

In the present investigation, a biopesticide (Herb essential Pure Neem Oil) and an organophosphate pesticide (Malathion) were chosen. Malathion was obtained from International Institute of Biotechnology and Toxicology Research (IIBTR), Paddapai.

The bioassay procedure^{16, 17}followed in the present investigation to determine the LC50 for both malathion and neem oil are reported in literature.

Fishes weighing from 10-20g were used in the present study. The bioassay was carried out for malathion, neem oil and the combination of malathion and neem oil separately. After calculating the LC50 values, the fishes were segregated into 4 groups and each group contained 6 fishes. Group 1 fishes were maintained in tap water, Group II in neem oil, Group III fishes were maintained in malathion , Group IV fishes were maintained in both neem oil and malathion. The LC50 dose of neem oil was found to be 0.9 ml/L and that of malathion was found to be 3.52 mg/L.

The fishes were starved prior to the experiment for a period of 24 hours. After the said period of 96 hours (acute toxicity) and 21 days (chronic toxicity), the fishes both control and experimental were sacrificed. The tissues were excised immediately and used for the present study. The tissues selected for the study were the liver and kidney.

The biochemical constituents like total protein, total free sugar and lipids were analysed for median lethal toxicity in the tissues selected by using the following standard procedures. Total protein, total free sugar and lipids^{18, 19, 20}were estimated by the methods described in the literature.

For histopathological preparation the different organs of the fishes such as liver and kidney of both control and experimental groups were excised and immediately fixed in Bouin's fluid. The tissues were then processed as with Haematoxylin and Eosin stains.²¹as described in the literature. Photo-micrographs were taken to substantiate observation and the results were discussed. Statistical analysis was carried out with two-way analysis of variances (ANOVA) followed by LSD tests using SPSS computer package.

Result

Results for biochemical analysis of protein, total free sugar and lipids from acute and chronic toxicity studies were obtained in all groups. The total protein content in the Group II fishes was observed to have a slight decrease while in the Group III fishes it’s significantly decreased. However an increase in Group IV was recorded which is comparatively more than Group III and this increase was found to be statistically significant. Acute and chronic toxicity study for total protein did not show a significant change (Table 1 and Fig 1). The total free sugar showed a decrease in all the groups subjected to acute toxicity study. However the chronic toxicity study showed an increase (Table 2 and Fig 2). The lipid content showed a slight decrease in Group III fishes more than Group II. However an increase in Group IV was recorded which is comparatively more than Group III and the increase was found to be statistically significant. Lipid content in all groups of acute toxicity was less when compared to chronic toxicity (Table 3 and Fig 3). The results were observed to be significant at p<0.0001.

Table 1: Protein content in kidney and liver ofOreochromis mossambicusof acute and chronic toxicity studies (mg/100mg of wet tissue)

Figure 1: Protein content in kidney and liver ofOreochromis mossambicusof acute and chronic toxicity studies (mg/100mg of wet tissue) Click here to View Figure

Table 2: Total free sugars in kidney and liver ofOreocheomis mossambicusof acute and chronic toxicity studies (mg/100mg of wet tissue)

Table 3: Lipid content in kidney and liver ofOreochromis mossambicusof acute and chronic toxicity studies (mg/100mg of wet tissue)

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