Adverse Effect of Insecticides on Various Aspects of Fish’s Biology and Physiology

Today, water quality management faces greater problems than at any time in its history. In addition to natural pollutants, varied contaminants exist in surface waters including multiple chemical compounds and different products of industrial and agricultural revolution. The insecticides constitute one group of these pollutants, both synthetic and natural, which contribute to the environmental problems. At present, it seems that the problem is more conspicuous in developing countries, where lately there has been an increase in the use of insecticides as a means of increasing agricultural productivity, without much concern to the consequences of indiscriminate application. There are many pathways by which insecticides leave their sites of application and distribute throughout the environment and enter the aquatic ecosystem. The major route of insecticides to water ecosystems in urban areas is through rainfall runoff and atmospheric deposition. Another source of water contamination by insecticides is from municipal and industrial dischargers. Most insecticides ultimately find their way into rivers, lakes and ponds (Tarahi Tabrizi, 2001; Honarpajouh, 2003; Bagheri, 2007; Shayeghi et al., 2007; Vryzas et al., 2009; Werimo et al., 2009; Arjmandi et al., 2010) and have been found to be highly toxic to non-target organisms that inhabit natural environments close to agricultural fields. The contamination of surface waters by insecticides is known to have ill effects on the growth, survival and reproduction of aquatic animals. In the past few years, the increase of mortality among the fish in various streams, lakes and ponds of around the world has drawn scholars’ attention to the problems caused by insecticides and pesticides runoff associated with intense agricultural practices. Different concentrations of insecticides are present in many types of wastewater and numerous studies have found them to be toxic to aquatic organisms especially fish species (Talebi, 1998; Uner et al., 2006; Banaee et al., 2008). Fishes are particularly sensitive to the environmental contamination of water. Hence, pollutants such as insecticides may significantly damage certain physiological and biochemical processes when they enter into the organs of fishes (John, 2007; Banaee et al., 2011). Authors found out that different kinds of insecticides can cause serious impairment to physiological and health status of fishes (Begum, 2004; Monteiro et al., 2006; Siang et al., 2007; Banaee et al., 2009). Since fishes are important sources of proteins and lipids for humans and domestic animals, so health of fishes is very important for human beings. Recently, many studies have been conducted to determine the mechanisms of insecticides in fishes, with the ultimate goal of


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monitoring, controlling and possibly intervening in xenobiotics exposure and its effects on the aquatic ecosystem.This chapter presents further information concerning the toxic effects of different concentrations of insecticides on various aspects of fish's biology and physiology.In other words, this chapter depicts the effects of insecticides on the survival chance, blood biochemical parameters, tissues and organs, reproduction, development and growth, nervous system, behavior, genetic and immune system of fish.The information given in this part facilitates the evaluation of potential toxic hazard resulting from exposure to different levels of these compounds.

Biokinetics and bioteransformation
After exposure to difference concentrations of insecticides in water, the fish absorbs them in its gills, skin or gastrointestinal tract.In the other words, Due to their lipophilicity, most insecticides easily permeate the biological membranes and it increases the sensitivity of fish to aqueous insecticides.Then, these compounds are rapidly metabolized and extracted, and may be bio-concentrated in various tissues of fish.In other words, bio-accumulation occurs if the insecticides is slowly metabolized or excreted from the body.As the amount of insecticide increases, it becomes more harmful to the consumer or animal.Accumulated insecticide can cause death or long-term damage.Ballesteros et al. (2011) showed that during the initial 24 h of exposure, insecticides may be transformed in various tissues of fish.However, some differences exist among tissues relating to metabolism rates, leading to different distribution models of the original compounds and their metabolites.For example, the low biodegradation and the high lipid solubility of some insecticides such as organochlorine insecticides have led to problems with the bio-concentrations of these compounds in different tissues of fish.In addition, since some fish are lower on the food chain, bioaccumulation of insecticides may increase in tissues of their predators and consumers, such as humans and thus affecting their health and survival.So, the bioaccumulation of these contaminants in fish and the potential biomagnification in humans are perceived as threats (Favari et al., 2002).Bioaccumulation rate of insecticides in fish depends on the species, life stages, the amount of fat reservation in different tissues and diet of fish, chemical and physical properties of insecticides and the rate of water pollution.In order to facilitate the elimination and detoxification of toxic compounds, fishes have developed partly complex detoxification mechanisms including the release of several enzymes collectively termed xenobiotic metabolizing enzymes.Enzymatic biotransformation of insecticides can potentially alter their activity and toxicity.Enzymes participating in the biotransformation of insecticides are classified into phase I and phase II enzymes.The phase I enzymes, cytochrome P450 enzymes including CYP1A and CYP3A, are generally involved in the biotransformation of exogenous and endogenous compounds; thereby creating a more polar and water soluble compound.A great diversity of cytochrome P450 enzymes in fish has been recognized (Stegeman and Hahn, 1994), and CYP1A, CYP2B, CYP2E1, CYP2K1 and CYP3A have been recently identified in liver of some freshwater fish (Nabb et al., 2006) which play an important role in the detoxification of organophosphate and carbamate insecticides (Ferrari et al., 2007).The common pathways of biotransformation of different kinds of insecticides include three cytochrome P450 (CYP) mediated reactions: O-dealkylation, hydroxylation, and epoxidation of insecticides (Soldano et al., 1992;Keizer et al., 1995;Kitamura et al., 2000;Straus et al., 2000;Behrens & Segner, 2001;Nebbia, 2001).In phase II reactions, metabolites produced in phase I detoxification often conjugate with glutathione, uridyl-diphosphate glucose (UDPG), uridyl-diphosphate-glucuronic acid (UDPGA), amino acid derivatives and sulfate derivatives and can readily excrete from the fish body (Iannelli et al., 1994;Keizer et al., 1995;Kitamura et al., 2000;Straus et al., 2000;Behrens & Segner, 2001;Nebbia, 2001).In other words, final metabolites may also be excreted from the body of fish through the skin, gills, genital products, urine as sulphated and glucuronidated metabolites and stool as glutathione conjugated metabolites (Kitamura et al., 2000;Straus et al., 2000;Behrens & Segner, 2001;McKim & Lein, 2001;Nebbia, 2001).Since metabolites produced during detoxification process may be more dangerous than parental compounds, these metabolites can cause serious damage in fish.Furthermore, the production of reactive oxygen species (ROS) during detoxification process can induce oxidative damage and may be a mechanism of toxicity for aquatic organisms living in environments receiving insecticides (Monteiro et al., 2009).ROS can indiscriminately attack and react with susceptible vital macromolecules -lipids, proteins and DNA-in living cells, inducing cytotoxicity and can result in serious disturbances in physiological cell processes (Dogan et al., 2011;Jin et al., 2011).Lipid peroxidation, the major contributor to the loss of cell function, DNA damage, enzyme inactivation, and hormone oxidation are bio-indicators of oxidative cell damage and examples of toxic mechanisms of insecticide induced ROS being involved in pathological processes and in the etiology of many fish diseases (Üner et al., 2006;Dogan et al., 2011).

Acute toxicity
In acute toxicity, sudden and intense mortality may be observed in a fish population exposed to toxic materials.The most apparent symptoms of insecticides' acute poisoning in fishes include lethargy, forward extension fins, pallor or blur parts of body, severe reaction to external stimuli and muscle spasms and sudden fast swimming in circles.The main clinical internal sings that can lead to death of fishes include neurological disorder and disruption of nerve functions, respiratory dysfunction and suffocation (Banaee et al., 2011).Acute toxicity testing is widely used in order to identify the dose or exposure concentration and the time associated with death of 50 percent of the fish exposed to toxic materials which is expressed as LC 50 in parts per million (ppm) or milligrams per liter (mg/L).In addition, we can use the LC 50 value in the classification of insecticides based on potential toxicity for fishes.Furthermore, the relative acute toxicity of chemicals to fish can be categorized as follows: Toxicity rating 96 hr LC 50 Slightly toxic 10-100 ppm Moderately toxic 1-10 ppm Highly toxic 0.1-1.0ppm Extremely toxic Less than 0.1 ppm Our literature reviews demonstrate that different fish species, even from the same family, show differences in the sensitivity to high concentrations of insecticides in water.Acute toxicity of different insecticides is influenced by the age, sex, genetic properties and body size of fish, water quality and its physicochemical parameters, and purity and formulation of insecticides.The eight tables, which give relative acute toxicity of some insecticides to fishes, can be used to determine the potential toxicity to fish of using these compounds in farms around surface waters and to select products which less likely to cause problems.The data are derived from

Sub-lethal toxicity
Sub-lethal toxicity testing was planned based on one tenth or more of LC 50 dose in moderate periods.In sub-lethal toxicity, the organs or biological systems which may be affected at such exposure can be respiratory, hepatic, haematopoietic, nervous, cardiovascular, and reproductive and immune systems.Different biomarkers of fish exposed to insecticides are usually evaluated in these experiments.Insecticides may lead to changes in the blood biochemical parameters and haematological profile of fish which can be investigated as biomarker in pollution monitoring (Mushigeri & David, 2005;Banaee et al., 2008;Kavitha & Rao, 2009).In fact, these compounds may induce alterations in the activities or levels of a number of different enzyme systems, including those necessary for biochemical reactions in cells (Banaee et al., 2011).Decreased rate of growth, reproductive disorder, spinal deformities, histopathological changes (Benli & Özkul, 2010) in gills, liver, haematopoietic tissue such as spleen, head of kidney, and renal tubules, endocrine tissues as well as brain, neurological and behavioral disorder and genetic defects are other biological indicators of exposure to insecticides which are described in details in the following sections.

Chronic toxicity
Chronic toxicity tests commonly include the measurement of long term effects of low concentrations of insecticides on the survival, growth, reproduction, nervous system and other biological and physiological aspects of fishes.Type of injury to fish in chronic toxicity is similar to sub-lethal toxicity damage, but the frequency and intensity injury and lesion resulting from chronic toxicity may be more or even less than damage of sub-lethal toxicity.Therefore, this experiment is important in insecticides toxicology.

Alterations in blood biochemical parameters
Insecticides can cause serious impairment to physiological and health status of fish.Therefore, biochemical tests are routine laboratory tests useful in recognizing acute or chronic toxicity of insecticides (Banaee et al., 2008;Al-Kahtani, 2011) and can be a practical tool to diagnose toxicity effects in target organs and to determine the physiological status in fish.Blood biochemistry test gives indicates what is happening in the body of fish exposed to insecticides.When different tissues are injured, the damaged cells release specific enzymes into plasma and we can recognize their abnormality levels in blood.Then it helps locate the lesions.Also, if certain organs are not eliminating certain waste products or not synthesizing certain important materials, this can tell us they are not functioning properly.

Tissue and organ damage
Histopathological investigations on different tissues of fish are valuable tools for toxicology studies and monitoring water pollutions.In histopathology, we can provide information about the health and functionality of organs.Tissues injuries and damages in organs can result in the reduced survival, growth and fitness, the low reproductive success or increase of susceptibility to pathological agents.Frequency and intensity of tissue lesions depend on the concentrations of insecticides and the length of the period fish are exposed to toxins.Nevertheless, many insecticides cause specific or non-specific histopathological damage (Fanta et al., 2003).For example, histopathological lesions in the liver tissue of freshwater fish (Cirrhinus mrigala) (Velmurugan et al., 2009) and common carp carp (Cyprinus carpio) (Banaee et al., In press) were observed after 10 and 30 days exposure to sublethal concentrations of dichlorvos and diazinon insecticides, respectively.Other researchers reported the same histopathological alterations in different tissues of fish treated with diazinon (Duttaa & Meijer, 2003;Banaee et al., 2011), deltamethrin (Cengiz, 2006;Cengiz & Unlu, 2006), fenitrothion (Benli & Özkul, 2010).

Reproductive dysfunction
Reproduction guarantees the survival of fish population.Any changes in environmental parameters or physiological conditions of fish can affect its reproductive success.Since fish may be exposed to environmental pollutants, including insecticides, herbicides, heavy metals and other xenobiotics, disorders may occur in their natural reproductive process.
Recent researches showed the dysfunction in the reproductive systems of fishes exposed to insecticides.Insecticides' effects on reproductive biology of fishes are numerous, and include decreased fecundity, testicular and ovarian histological damage (Duttaa & Meijer, 2003;Banaee et al., 2009), vitellogenesis process impairment (Haider and Upadhyaya, 1985;), and disruption in steroidogenesis process (Zaheer Khan & Law, 2005), delay in gonads maturation (Skandhan et al., 2008), alter in reproductive and parental behavior (Jaensson et al., 2007), impairment in olfactory response and disorder in reproductive migrations (Scholz et al., 2000), as well as disruption in coordinating courtship behavior of male and female fish and time of spawning (Jaensson et al., 2007).Some insecticides are known as endocrine disrupting chemicals (EDC) which can interfere with the normal functioning of endocrine system in fish.Adverse effects of insecticides on the hypothalamus-pituitary-gonads axis can also play a significant role in causing reproductive failures in fish.In fishes, chronic toxicity of insecticides can change sex steroid hormone levels in plasma.While the mechanism is not exactly known, it is possible that insecticides and their metabolites disrupt reproductive systems through activation or inhibition of key enzymes which participated in the steroid hormone biosynthesis in fishes.For example, DDT, endosulfan, methoxychlor and some other insecticides possess estrogenic properties and are probably capable of disrupting functions of endocrine system and causing disorder in the reproductive system of fish (Arukwe, 2001).These compounds may directly or indirectly interact with natural hormones, changing the hormone functions and thus altering physiological cellular response or mutate the natural patterns of hormone synthesis and metabolism.Impact of organophosphate insecticides such as malathion, diazinon and fenitrothion on the hypothalamus-pituitary-gonads axis and also disturbance in hormones associated with reproductive systems were studied by Kapur & Toor, (1978); Singh and Singh, (1987); Maxwell & Dutta, (2005); Skandhan et al. (2008).
Insecticides can also cause adverse effects on gonad histology, morphology and its growth.
In addition, there are significant relationships between blood sex steroid hormone concentrations, sperm or oocytes quality, rate of fecundity and histopathological alterations in ovary and testis of fish exposed to different insecticides (Duttaa & Meijer, 2003;Maxwell & Dutta, 2005).Banaee et al., (2008) reported that diazinon inhibits steroidogenesis in testis of male carp by histopatological alterations.Research results showed that direct toxic effects of insecticides on seminiferous tubules or Leydig cells may be the most important parameter for the low quality of sperms in fish (Fadakar Masouleh et al., 2011).Similar results were reported in walking catfish (Clarias batrachus), freshwater eel (Monopterus albus), and Atlantic salmon (Salmo salar) that were exposed to different insecticides (Singh & Singh, 1987;Singh, 1989;Moore & Waring, 1996;2001).Exposure of fish eggs and milt to insecticides also reduced the level of fertilization, hatching rate and the larval survivability.Further studies on bluegills (Lepomis macrochirus), atlantic salmon (Salmo salar L.) demonstrated that the gametes and fertilized eggs were sensitive to the insecticides (Tanner & Knuth, 1996;Moore & Waring, 2001) suggesting a further toxic impact of these toxicants on the fish reproduction.In addition, the waste of energy in the fish exposed to insecticides reduces their reproductive ability.

Development disorders
Study of development disorders caused by insecticides is to emphasize the links between the concentrations of toxins and dysfunction in normal development from embryonic to puberty periods.So, impairment in the normal development and the growth may reduce the fish's survival chance.
Embryos and larvae may be directly exposed to insecticides, through the yolk or via parental transfer in viviparous fish (Viant et al., 2006).Spinal deformities, mostly scoliosis and lordosis, and morphological abnormalities were among the more adverse effects registered for insecticides toxicity.Other alterations in the embryo of fish exposed to insecticides also consist of yolk sac edema, pericardial edema and crooked body of larvae (Xu et al., 2010).Teratogenic effects of carbaryl insecticides on the embryo of fish have been proved (Todd and Leeuwen, 2002).Similar results were reported in silversides Menidia beryllina exposed to tebufos during embryogenesis (Middaugh et al., 1990;Hemmer et al., 1990).
The most important factors decreasing fish growth consist of disorder in feeding behaviors, decrease in feeding rate, dysfunction in metabolism process and waste of energy to overcome the stress caused by insecticide exposure (Tripathi et al., 2003).For example, disorder in the metabolism of carbohydrates, proteins and lipids in various tissues, particularly liver of fish exposed to insecticides, may reduce their growth rates.Begum (2004) found out that protein and carbohydrate metabolism in the liver and muscle tissue is disrupted on the exposure to a carbofuran insecticide.In addition, exposure during embryonic or larval stage can result in behavioral abnormalities, such as decreased ability to capture prey after hatching, functional deficiencies or slowing of growth and finally death (Kuster, 2005;Viant et al., 2006;Arufe et al., 2007).These changes were observed in larvae and embryo of zebra fish (Danio rerio) in contact with endosulfan (Velasco-Santamaria et al., 2011), beta-cyprmethrin (Xu et al., 2010); paraoxon-methyl (Küster, 2005) and sevin (carbaryl insecticide) (Todd and Leeuwen, 2002).

Neurotoxicity
The primary mechanism of organophosphate and carbamate insecticides toxicity is well known -they function as inhibitors of acetycholinestrase enzyme (AChE) or and butyrylcholinesterase (BChE), as well as disturbing the metabolism of other neurotransmitters such as -aminobutyrate (GABA).The synthetic pyrethroids change normal neuronal function by interfering in the function of ion channels in the nerve cell membrane, alterations in intracellular calcium ion concentrations and possibly by blocking GABA receptors.Organochlorine insecticides act primarily by changing the transport of ions across the nerve cell membranes, thus altering the ability of nerve to stimulate.Fish exposure to these insecticides is frequently assessed by determining the alterations in AChE in brain, muscle, plasma and other tissues or probably GABA activity in brain (Banaee, 2010).AChE is an enzyme responsible for inactivating the neurotransmitter acetylcholine (Fulton & Key, 2001).AChE inactivation results in the accumulation of the neurotransmitter acetylcholine in cholinergic synapses space, leading to synaptic blockage and disruption of signal transmission (Ferrari et al., 2004;2007a, b).Inhibition of AChE induces alteration in the swimming behavior, shaking palsy, spasms and other undesirable effects (Sharbide et al., 2011).Disturbances in AChE activity can also impair feeding, identification and avoidance and escaping from predators, spatial orientation of the species, and reproductive behavior (Bretaud et al., 2000).Thus, AChE inhibition is considered to be a specific biomarker of exposure to organophosphorus and carbamate insecticides like diazinon, chlorpyrifos, propoxur, isoprocarb, (Üner et al., 2006;Cong et al., 2008;2009;Wang et al., 2009;Banaee et al., 2011;).Similar results have been observed for pyrethroids insecticide toxicity (Koprucu et al., 2006).Disorder in -aminobutyrate (GABA) system in brain of rainbow trout exposed to sub-lethal lindane was reported by Aldegunde et al., (1999).GABA receptors inhibit the transmission of nerve impulses; thus disturbances in this receptor would also lead to an over stimulation of the nerves.
In addition, nervous tissue has weaker antioxidant defense system than other kinds of tissue.On the other hand, the brain as center of the nervous system in fish contains low levels of enzymatic and non-enzymatic antioxidant and higher levels of oxidizable unsaturated lipids and catecholamines.So, nerve tissue is very sensitive to oxidative stress damage induced insecticide toxicity compared with other tissues (Üner et al., 2006;Li et al., 2010).These results have been reported by other scientists (Senger et al., 2005).

Behavioral alterations
Behavioral changes are the most sensitive indicators of potential toxic effects.The behavioral and the swimming patterns of the fish exposed to different insecticides include changes in swimming behavior, feeding activities, predation, competition, reproduction and speciesspecies social interactions such as aggression (Cong et al., 2008;2009;Werner and Oram, 2008).Banaee et al. (2008;2011) reported similar behavioral responses in common carp and rainbow trout exposed to sub-lethal levels of diazinon.In fact, most insecticides influence the behavior patterns of fish by interfering with the nervous systems and sensory receptors (Keizer et al., 1995;Pan & Dutta, 1998;Cong et al., 2008;2009); and this incident may impair the identification of situation and development of appropriate response by the fish exposed to insecticide.The effect of certain insecticides on the activity of acetylcholinestrase may lead to a decreased mobility in fish (Bretaud et al., 2000).Researchers have reported the same alterations in Oryzias latipes, Cyprinus carpio, Labeo rohita, Oncorhynchus tshawytscha, O.latipes, Cirrhinus mrigala, Oreochromis niloticus, Clarias gariepinus treated with chlorpyrifos (Rice et al., 1997;Halappa & David, 2009), malathion (Patil & David, 2008), diazinon (Scholz et al., 2000), endosulfan (Gormley & Teather, 2003), Fenvalerate (Mushigeri & David, 2005), fenitrothion (Benli & Özkul, 2010), dimethoate (Auta et al., 2002), respectively.

Genotoxicity
In genetic-toxicology any heritable damage or DNA inactivation resulting from the animal's exposure to xenobiotics is studied.Genotoxic chemicals such as insecticides have common chemicals and physical properties that enable them interact with genetic materials (Campana et al., 1999;Çava & Ergene-Gözükara, 2003;Candioti et al., 2010;Dogan et al., 2011).The mutation that may result from an interaction between a chemical and the genetic material is a heritable change in the cell genotype, and thus the error may be transferred to the daughter cell or the next generation.Carcinogenesis and the formation of some tumors in different tissues of fish exposed to insecticides may also be caused by genotoxic properties of these xenobiotics.One of the ill effects of insecticides' arrival into surface waters may be an induction of chromosomal damage in eggs and larvae of fishes in different stages of development.Some insecticides that behave as endocrine active compounds can change the expression of vital genes resulting in unusual concentrations of plasma steroid hormones and reproductive dysfunction or immuosuppression (Jin et al., 2010).

Immunosuppression
The immune system of fish is important for defense against a variety of pathogens.The system is very sensitive to homeostatic adjustments via endocrine regulation and is influenced by the biochemical status of the nervous system.Thus, any impairment in the nervous system and disturbance in the biochemical homeostasis can weaken the immune system of fish.On the other hand, insecticides may alter the function of the immune system and result in immunodepression, uncontrolled cell proliferation, and alterations of the host defense mechanisms including innate immunity and acquire immunity against pathogens.Different insecticides at sub-lethal levels have been recognized as stressors causing immunesuppression in fish (Werner and Oram, 2008).In addition, some insecticides may exert immunotoxic effects by altering the transcription of important mediators of the fish's immune system (Eder et al., 2009).Effects of insecticides like P,P'-DDE, lindane, cypermethrin, chlorpyrifos, diazinon on the immune factors of fish such as Interleukin-1 (IL-1 ), IL-1 receptor (IL-1R1), Interferon gamma (IFN-2b), TNF , MHCI , MHCII , Mx, TLR9, I ML and C-reactive protein (CRP), TCR in head-kidney leucocytes, Lysozyme activity, chemiluminuscence (CL) response and immunocompetent cells population size, IgM levels, value of white blood cells (WBC) and respiratory burst activity, head kidney phagocytes and peripheral blood leucocytes, etc., have been reported by scholars (Betoulle et al., 2000;Khoshbavar-Rostami et al., 2006;Banaee et al., 2008;Cuesta et al., 2008;Girón-Pérez et al., 2009;Shelley et al., 2009;Ahmadi et al., 2011;Jin et al., 2011, Wang et al., 2011).The exposure to sub-lethal concentrations of insecticides is what probably makes fish vulnerable to infectious diseases because of their immune-depressive effects (Zelikoff et al., 2000).For example, the susceptibility of juvenile chinoock salmon (O.tshawytscha) to infectious hematopoietic necrosis virus was significantly increased in fish exposed to sub-lethal concentrations of esfenvalerate (Clifford et al., 2005).Similar results were reported in goldfish and common carp that were exposed to carbaryl and lindane respectively (Shea, 1983;Shea & Berry, 1984;Cossarini-dunier & Hattenberger, 1988).

Table 1 .
are given only as a guideline and not absolute data of the acute toxicity of the insecticides to different species of fish (Table1-11).Summary of acute toxicity.

Table 2 .
Summary of acute toxicity.

Table 3 .
Summary of acute toxicity.

Table 4 .
Summary of acute toxicity.
www.intechopen.comAdverseEffect of Insecticides on Various Aspects of Fish's Biology and Physiology 107

Table 5 .
Summary of acute toxicity.

Table 6 .
Summary of acute toxicity.

Table 7 .
Summary of acute toxicity.

Table 8 .
Summary of acute toxicity.

Table 9 .
Summary of acute toxicity.

Table 10 .
Summary of acute toxicity.

Table 11 .
Summary of acute toxicity.