Abstract

This review paper aimed to assess the Bioaccumulation and toxicity of chromium and its toxicological effects on freshwater fishes in terms of hematological, biochemical, cytotoxic and genetical parameters. Chromium is considered as the most common pollutant in the aquatic environment. Chromium enters the aquatic environment through different industrial effluents like tanneries, textiles, dyeing, electroplating, medical industries, printing-photographic, etc. Chromium is consumed either via the respiratory system or the digestive tract. Bioaccumulation of chromium depends on several biotic and abiotic factors viz. age, species, developmental phase of the organism, temperature, pH, salinity, and alkalinity of the aquatic medium. Chromium impaired the aquatic environment due to its toxic effect on aquatic life. Different research reviews indicated toxicity of chromium in freshwater fishes at a hematological level like a decrease in hemoglobin percentage, RBC, and WBC total count. At the biochemical level, the decline in the total content of lipids and proteins was detected. At the enzyme activity level, the decline in the ROS, SOD level, etc. Chromium is also carcinogenic proved by many studies. The main objective of this review is to provide a future guideline for the scientific society involved in chromium toxicity in freshwater fish research to ensure a better environment for fish health.

1. Introduction

Heavy metals pollution is a major challenge to aquatic life and has conveyed enormous alterations and damaging consequences to aquatic organisms. Heavy metals Pollution of the aquatic environment is a crucial global issue for the previous two decades. The effect of Heavy metals shows at low concentration in the environment. Atomic weight and specific gravity of the heavy metals are respectively between 63.5 & 200.6 and more than 5.0 ¹. Heavy metals contamination is a major aquatic environmental load due to its high accumulation power, non-biodegradable, and flexibility ^{31, 32}. In aquatic ecosystems, sources of non-biodegradable and toxic heavy metals are tanneries, leather industries, paper industries, metal plating industries, electroplating industries, pesticides, mining operations, textile industry, etc. Heavy metals viz. chromium, arsenic, cadmium, nickel, copper, zinc, antimony, cobalt, etc. have toxic effects on aquatic life. Heavy metals have a toxic beyond the permissible limit effect on aquatic life ³⁶.

Chromium is the seventh element on the earth’s crust. Industries like tanning, leather, textile, electroplating, etc. are the main sources of chromium. Trivalent chromium, Cr (III), and hexavalent chromium (VI) are the most available form of chromium which is found in industrial effluent ^{29, 106}. The hexavalent form of chromium is more toxic than the trivalent chromium for aquatic organisms ^{57, 60}. An important role of Cr (III) is a carbohydrate and fat metabolism ^{17, 15}. One of the major sources of chromium contamination in water is Tannery effluents. In India, tannery effluents contribute more or less 2000–3000 tones of chromium ^{2, 3}.

Contamination of heavy metals has a huge impact on the ecological stability of the aquatic ecosystem ^{4, 5}. The most simple biomarkers of pollution of the aquatic environment are fish. In the Highly toxic heavy metals contaminated aquatic environment, fish appear to take up heavy metals from their environment ⁶. The heavy metals have toxic effects on the aquatic environmental equilibrium as well as aquatic life ^{43, 48}. Accumulation of the heavy metals in the food chain of the aquatic ecosystem produces adverse effects on fish, so among other animals, fishes are used to determine the health status of the aquatic life ^{28, 55}. Fishes consume heavy metals from their diet and the bioaccumulation of the heavy metals in the fish is highest because fishes are the highest consumer of the food chain in the aquatic environment ⁷. Fish accumulates heavy metals by consuming food or by ion exchange of the heavy metals via the cell membranes which are deposited in tissues ³⁸. By passive diffusion, Cr (VI) readily enters gill membranes which accumulates in several tissues, and can exhibit its toxic effect internally ⁴⁷. In fish, sequence of heavy metal bioaccumulation in the gills and liver is Cd > Pb > Ni >Cr and Pb > Cd > Ni > Cr, Simultaneously in case of kidney and muscle is Pb > Cd > Cr > Ni and Pb > Cr> Cd > Ni. ²⁷.

Toxicity of hexavalent form of chromium induces hematological and biochemical ⁸, hepatic ⁹, mutagenic and genetic ¹⁰, reproductive ¹², and behavioral ¹¹ effects on fish. The toxicity of chromium may be related to alterations of the antioxidants and reactive oxygen species (ROS) production in fish ^{13, 14}. The Hexavalent chromium, Cr (VI) can cross the cell membrane after that reduces to trivalent, Cr (III) form which is bind to intracellular macromolecules and is produce a mutagenic effect in the cell ⁷. The toxicity of chromium on the hematology, biochemistry, and histopathology of a variety of fishes has been well documented ⁷², however, little information is available about the chromium-induced genotoxicological effect among freshwater fishes ⁷³. The present study is aimed at finding out the bioaccumulation and toxicity of chromium (Cr) in freshwater fishes.

2. Chromium in the Environment

The crystalline form of Chromium is a steel-gray, lustrous, hard metal characterized by an atomic number of 24, an atomic weight of 51.996, a melting point of 1900 °C, and a boiling point of 2642°C, and a density of 7.14 ⁷⁹. More or less all chromium accumulation occurred via magmatic dispersion in ultrabasic rocks. The chromite is deposited in the environment during the chilling of magma. lateritic soils develop containing as much as 2–4% chromium under favorable tropical and subtropical conditions ¹⁶. Chromium accumulates in influent from natural, industrial or domestic sources ¹⁸. Major sources of both chromium (VI) and chromium (III) effluents are metal plating, anodizing, ink manufacture, dyes, pigments, tanning, glass, glues, ceramics, textiles, wood preservatives, etc ¹⁹. In a natural water system, Chromium concentration ranges between 1 and 2 mg/l in the dissolved form ⁸⁰. Groundwaters in La Spezia province, Italy, have a Cr (VI) content ranging from 5 to 73 µg L^-1 exceeding the Italian limit for drinking water at 5 µg L^-1 as well as the WHO limit for drinking water of 50 µg L^{-1 95, 96}. In 1997, Central Pollution Control Board, India reported Cr (VI) concentration upto 250 times higher than the WHO permissible limit (0.05 ppm) in areas at Kanpur ⁹⁷. The high level of chromium deposition in sediments may cause Cr deposition in water above the maximum discharge limit in European Union ¹⁰⁴. Chromium pollution of China is universally reported in coastal environment ¹⁰⁵. Hexavalent and trivalent the most stable forms of chromium can coexist in an aquatic ecosystem with few organic matters ⁷⁹. Hexavalent chromium exists in the form of chromate (CrO4^-2), hydrochlorate (HCrO4^-1), or dichromate (Cr2O7^-2) in dissolved conditions ⁸¹. According to WHO the mean concentration of chromium ions in river water is 0.050 mg/l ¹². It is reported that the average acute value of Cr (VI) in freshwater fishes is ranged from 30mg/l to 139.9mg/l ²². According to FAD, the permissible limit of Cr (VI) in fish tissue is 12–13 mg/kg body weight ²³.

3. Bioaccumulation of Heavy Metals in Freshwater Fishes

Heavy metals contamination influences fish health in the aquatic environment. Many reports said that bioaccumulation of the heavy metals found in fishes ⁵⁴. Age, sex, size, reproductive cycle, feeding behavior, swimming patterns, living environment, etc influence heavy metal uptake in the fish ⁵⁸. Through absorption and adsorption, heavy metals enter the fish’s organs ⁵⁹. It is reported that the accumulation of heavy metals Significantly varied among the fish species inhabiting the same aquatic environment. Bioaccumulation of heavy metals in different tissue of fish varies. Generally, the liver and kidney showed higher accumulation than muscle, and gill ⁵⁶.

4. Bioaccumulation of Chromium in Freshwater Fishes

The concentration of chromium was comparatively more in fish tissue than in the surrounding water. Fish uptake chromium through the food chain because fish are the top consumer in the aquatic environment ³⁰. No chromium biomagnification has been shown in food chains in the aquatic environment ²⁹. Hexavalent Chromium (VI) enters the fish body via the gill membrane and it is highly concentrated quickly in other tissues (gastrointestinal tract, gall bladder, intestine, stomach, kidney, opercular bone, spleen, and, brain) than in the gills ^{20, 63, 64}. It is reported that mostly chromium is concentrated in the gills followed by the liver. Accumulation of Cr was found to be maximum in Catla catla and minimum in Puntius sarana. It was observed that Puntius ticto to accumulate the least amount of metals as compared to six other species of fish like Oreochromis nilotica, Hypophthalmichthys molitrix, Labio rohita, Channa marulius, Oreochromis mossambica, and Catla catla ⁹⁸.

Comparatively, Fishes are resistant to Cr ion but when exposed to Cr concentrations from 0.013 to 50 mg/l fishes can be sublethally affected and when exposed within 3.5 to 280 mg/l, can be lethally affected ⁶². It was reported that bioaccumulation of chromium is dependent on water pH value ^{45, 49}. Gills were the principal organ of the toxic effect of chromium at a pH of 6.5. Followed by gills, the liver was the second-highest chromium bioaccumulation and it was the site of detoxification and storage for chromium ⁵³.

The toxicity of chromium is dependent on the pH of the water. It has been reported that chromium toxicity was found to be greatly affected by slight changes in pH in Nuria Henricus (teleost fish) ⁹³. This experiment signifies that at pH 7.8 a significant amount of chromium accumulates in the internal organs while at pH 6.5 the gills retained an enormous amount of chromium ⁹⁴. Another study revealed that chromium toxicity was 50-200 times much higher at pH 6.4 to 7.4 on young rainbow trout.

5. Toxicity of Chromium

A previous report said that in aquatic organisms, acute toxicity of hexavalent chromium is more toxic to freshwater biota than acidic and soft waters. Another report said that younger aquatic organism is more sensitive than older organisms to acute toxicity of Cr (VI). The toxicity of Cr to fish are species-specific. One recent investigation designed to test the sensitivity of five freshwater fish, rainbow trout (Oncorhynchus mykiss), three-spined stickleback (Gasterosteus aculeatus), roach (Rutilus rutilus), perch (Perca flavescens), and dace (Leuciscus leuciscus) exposed to acute concentrations of Cr (VI) showed that rainbow trout is 1.16 to 2.52 times more sensitive than the other test species to the metal ¹⁰¹. The toxicity of chromium to aquatic life is based on a variety of abiotic and biological factors. The biotic factors are age, species, and developmental stage of the organism; and abiotic factors are pH, salinity, alkalinity, and temperature of the medium; exposure duration; contaminants with other heavy metals and chemical nature of Cr found. For sensitive freshwater organisms, LC-50 of 96-hour values of hexavalent chromium were from 445 to 2,000 ppb, and for trivalent chromium, was from 2,000 to 3,200 ppb ²⁶. Water pH influenced the toxicity of chromium to aquatic organisms. As the pH value decreased from 7.8 to 6.5, chromium ion concentration was badly affected ³⁷. At pH 7.8, chromium concentrations are high in gill. At high Cr (VI) concentration, the highest bioaccumulation found in rainbow fish’s gill and this transfer into the chromium-free aquatic environment, the highest concentration found in kidney and liver ²⁴. Chromium is carcinogenic, mutagenic, and teratogenic to aquatic organisms in laboratory conditions. At higher concentrations, Hexavalent chromium altered blood chemistry, hematological parameter alternation, abnormal enzyme activities, behavioral modifications, and histopathological modifications of freshwater fishes ²⁵.

In freshwater fishes, chromium may bring alterations in various aspects after acute exposure or Short term exposure.

Chromium has a potent cytotoxic effect on aquatic organisms when bioaccumulating in the food chain. Chromium induces cytotoxicity mainly measured using viability, cell death, metabolism, cell attachment, morphology, proliferation, cell membrane growth kinetics, and permeability ³³. Recent studies have revealed that sodium chromate, a soluble form of hexavalent chromium has cytotoxic effects in a cell line of Oryzias latipes. In this experiment, sodium chromate cytotoxicity is measured by clonogenic cytotoxicity assay and estimate clastogenicity using measuring chromosome damage and DNA double-strand breaks, using gamma-H2A.X immunofluorescence ³⁴. Due to exposure to 250μM of Cr (VI), the cell viability of goldfish is reduced and influences Reactive oxygen species (ROS) production of goldfish ³⁵. Another study shows that single-cell made from gill, liver tissues, and the blood of freshwater fishes at concentrations of Cr(I), Cr (III), and Cr (III), cell viability(percentage) is significantly reduced compared to control.

Tan et al. have studied the response of six fish cell lines introduced in four heavy metals. The cell lines have been indicated as EPC (epithelioma papulosum, cyprinid), GCF (grass carp fins CIK (Ctenopharyngodon ideas kidney), BB (brown bullhead caudal trunk), CCO (channel catfish ovary), and FHM (fathead minnow muscle). These cell lines are examined comparatively for their cytotoxic sensitivity to different metals viz., chromium (Cr), zinc (Zn), cadmium (Cd), and copper (Cu). The cell viability, cytomorphology, and proliferation after an exposure of 24 hours to heavy metals at different concentrations have been measured in that experiment ⁸². A recent study showed that long term exposure of chromium at a concentration of 2-200 µmol/L on Cyprinus carpio showed cytotoxicity, decreased lymphocyte activation, mitogen-induced and phagocyte functions ¹⁰².

Exposure of the freshwater Saccobranchus fossils to the sublethal condition of Chromium resulted in less antibody production, higher sensitivity to infections with the bacteria, Aeromonas hydrophila, and reduced proliferation of splenic lymphocytes ⁷⁰. Freshwater cichlid, Oreochromis mossambicus introduced to chromium from tannery effluents resulted in spleen atrophy, and decreased leukocyte counts ⁷¹. Nonspecific immune mechanisms have been illustrated in terms of the formation of reactive oxygen species (ROS) and serum lysozyme activity intermediates by peripheral blood leucocytes during laboratory studies ⁷⁴.

Several studies reported that chromium has an ill effect to alter the biochemical parameter and enzyme activity of fishes. The toxicity of chromium ions alters the glucose transportation pattern in the intestine ⁵². Another study said that depletion of glycogen levels in the tissues of Labeo rohita occurred due to increased utilization of heavy metals ⁴⁴. In Labeo rohita, tissue proteins and lipids were also decreased due to exposure to chromium ⁴⁶. Another important study showed that depletion of liver glycogen content was noticed in a freshwater teleost Colisa fasciatus, on chromium exposure ¹⁰³. Alanine transaminase (ALT) and Aspartate aminotransferase (AST) levels increased in fishes due to exposure to higher heavy metals concentrations. The status of the kidney due to exposure to heavy metals is determined by Creatine kinase (CK). Maximum activity of superoxide dismutase (SOD) was found in the liver followed by kidney of Wallago attu due to exposure to increasing concentrations of chromium. Also the highest mean value of peroxidase and catalase in the liver followed by the kidney, and gills of the C. marvelous due to exposure to high chromium concentrations ⁵⁰. Sub-lethally Trivalent chromium alters Na+, K+-ATPase activity of various fishes ⁶⁸. Chromium can change the glucose transport rate in epithelial cells of the intestine. One of the experiments on glucose uptake by epithelial cells in the intestine of freshwater fish, rainbow trout (Oncorhynchus mykiss) showed a diminished rate of glucose absorption ⁹⁹. Subsequent experiment on the effect of Cr on glucose intake in freshwater fish, Channa punctatus at different concentrations (10 mM, 1 mM, 0.1 mM, 0.01 mM and 0.001 mM) appeared increasing absorption of glucose at all Cr concentrations investigated ¹⁰⁰.

Endocrine disruption experiments are useful tools in toxicological studies. Due to chromium exposure endocrine functions like thyroid-stimulating hormone (TSH), free triiodothyronine (T3), free thyroxin (T4), and plasma cortisol, have been observed to fluctuate in freshwater fishes. It has been revealed that plasma T4 values in eels decrease after exposure to heavy metals ⁸⁴.

Several behavioral changes in fish are shown due to exposure to high chromium concentrations. Modified behaviors are uneven swimming, accelerated operculum, and suspending feeding behavior ^{65, 66}. The effect of trivalent and hexavalent forms of chromium toxicity on the behavior of zebrafish are opening mouths for gasping, mucus discharge, Erratic motion, irregular swimming, color, and shade alterations ⁶⁷. Another study reported that behavioral patterns of Channa punctatus are altered due to exposure to hexavalent chromium.

It has been reported that Labeo rohita loses its balance after exposure to a potassium dichromate (K₂Cr₂O₇) medium of 24 hours at 28.99 mg/l concentration. After 24 hours at 56.59 mg/l concentration of potassium dichromate (K₂Cr₂O₇) medium, the activeness and swimming rate of the Labeo rohita have been found to vary and found to be highly decreased balance and mucus secretion ⁷⁵. It has been reported that after exposure of 20 mg/l and 40 mg/l concentration of potassium dichromate (K₂Cr₂O₇) are found to be altered some characteristics like erratic swimming, hyperactivity, erratic swimming, increased rate of swimming, loss of balance, the tendency of convulsion, and increased opercular beat rate in Channa punctatus ⁷⁶. Erratic swimming and hyperactivity of freshwater fishes are most important while exposed to chromium polluted environment ^{75, 77, 78}.

Several studies have reported that hexavalent chromium induced several alterations in cytology, histology, behavior, physiology, and morphology. Decrease in lymphocyte count and antibody production ⁸⁵, increase in blood and muscle lactic acid ⁸⁶, DNA damage, decrease in Growth and survival rate ⁸⁷, reduction in protein level ⁸⁸ after long exposure or chronic exposure in freshwater fishes.

Blood is used as a potential biomarker of stress in fish due to exposure to heavy metals in the aquatic environment. Blood is also used as an index for measuring the metabolism activity of fish under heavy metal stress. Sublethally hexavalent chromium significantly altered the hematological parameter of Labeo rohita. In this study, Haemoglobin (Hb), Red blood corpuscles (RBC), MCH, Haematocrit (PCV), and MCHC values significantly declined and the MCV and WBC values significantly increased compared to the control group ⁵¹.

The activity of lactate dehydrogenase (LDH) on organs like the liver, and kidney of Channa punctatus decreases significantly, and the inhibition in enzymatic activity of pyruvate dehydrogenase (PDH) in different organs like the kidney, brain, liver, gill, intestine, and muscles of Channa punctatus after exposure of 2.6 mg/l of chromium concentration for 60–120 days. Chromium promotes reactive oxygen species (ROS) such as hydrogen peroxides which may cause cell membrane damage and thus destroy the cell ⁸⁹.

Several studies revealed that the histopathological changes occurred in different tissue like the gill, liver, kidney, intestine, brain, heart, and muscle of freshwater fishes after chronic exposure to sublethal concentrations of chromium ^{90, 91}. In Labeo rohita and Heteropneustes fossilis, some changes occurred like Necrosis and hyperplasia of liver cells after 60 days of exposure to 96 h 1/10th LC50 of chromium ⁹¹. In Labeo rohita and Heteropneustes fossilis, glomerular disorganization fenestrated Bowman’s capsule and narrow lumen of Renal tube shows after 60 days exposure to 96 h 1/10th LC50 of chromium ⁹². It has been reported that Necrosis in epithelial cells, lamellar degradation, and thickening of blood vessels occurred after 60 days of exposure to 96 h 1/10th LC50 in Labeo rohita and Heteropneustes fossilis ^{90, 91, 92}.

Higher DNA damage (tail DNA percentage) occurred in freshwater fishes due to exposure to hexavalent chromium. Liver tissues showed significantly more maximum DNA damage than the other tissues, so DNA damage is tissue-specific due to exposure to hexavalent chromium. The DNA damage revealed that alteration of chromosomal structure which is measured by micronucleus (MN) observation. The micronucleus test in fish revealed that chromium-induced environmental pollutants are aneugenic and clastogenic ⁴⁰.

DNA base alteration and deoxyribose phosphate backbone breakage result due to the formation of the hydroxyl radical, as a result of depletion of Hexavalent chromium to form tetraperoxo-chromate in fishes ³⁸. The Comet assay was also used to measure the genetic toxicity of freshwater fishes due to exposure to chromium. Comet assay is a potential mechanism to correlate the DNA damage and the exposure of aquatic organisms to chromium ³⁹. Comet assay and MN assay are used to measure respectively alkalilabel position along with measurement of DNA damaging capacity of chromium ⁴¹ and measurement of the chromosome breakage ⁴². Trivalent chromium can modify the gene expression of various fishes both in laboratory and Cr- contaminated water bodies ⁶⁹.

6. Conclusion

This review article has revealed the toxicity of chromium in fishes. Alteration of various structural and functional activities of tissues and organs of freshwater fishes is due to the toxicity of chromium. The various report revealed that Fish are badly affected on several levels viz. physiologic, biochemical, hematological, enzymatic, histologic, and genetic levels due to exposure to chromium. Hexavalent chromium is a carcinogen to fish. Chromium toxicity varies from fish species to species which means some species appear to show more toxicity than other species. Hexavalent chromium is more toxic to freshwater fish as compared to trivalent chromium. The toxicity of chromium on fish has depended on several factors viz. species, the physiological condition of fish, age, developmental conditions of fish, environmental condition, pH, temperature, TDS, chromium concentration, and exposure duration. This review provides more data about the ecotoxicology and risk of chromium toxicity for further research.

References