Chloride is added to municipal (tap) water supplies to kill microorganisms
(Noga, 1996). Chlorine reacts with the natural organic matter or contaminants
in surface waters and produces a complex mixture of Disinfection By-Products
(DBPs), some of which have been shown to be carcinogenic, mutagenic and/or
teratogenic in animal studies (Ferraris et al., 2005). Like many
other toxins in water chlorine is much more toxic to fish than humans
(Noga, 1996). Toxicity with discharges of chlorine is common because it
is used to disinfect effluents, to control fouling organisms in cooling
water system and in industrial process particularly in the food and paper
industries (Fisher et al., 2003). Although chlorine toxicity has
been frequently reported as a cause of mortality in fish, the pathology
changes related to this toxicity is treated rather briefly in literature.
Thus, the purpose of this study was to ascertain some of these pathological
changes in a naturally occurring toxicity. This case study also illustrates
the importance of good history taking and how interrelated fish are with
their watery environment.
Eight-month old rainbow trout (Oncorhynchus mykiss) were found
dead in a freshwater recirculation system in the key center of fish health
research center of University of Tasmania, Launceston, Australia. Ten
days prior to this incident, the system was shut down during a blackout.
Fish were found swimming in a swirling motion or died suddenly. Ten days
later mortality rates reached 60%. The fish were immediately fixed in
10% formalin buffer and were sent to DPIWE diagnostic laboratories in
Launceston. Investigations revealed that the tanks had been refilled with
town water that had variable levels of chlorine and even after sodium
thiosulfate was added, chlorine levels were still as high as 0.2 ppm.
The type of Chlorine used by the water department was Chloride dioxide
and CL 2 (Liquid).
Dead fish had enlarged gall bladders. Mild multifocal dermatitis was
observed in the skin. Their carcasses appeared pale and anemic. Wet preparations
of skin scrapes and gill biopsies were negative for parasites.
Histopathological examination revealed gill edematosis and foamy vacuolation
especially in the proximal part of the secondary lamellae. Gill epithelial
cells were swollen. This feature was more prominent in chloride cells.
Epithelial lifting and telangiectasis in the secondary lamellae were also
found (Fig. 1).
The histopathological features of the kidney consisted of increase in
golden pigments and azurophilic bodies within phagocytes and variable,
but sometimes extensive erythrophagia. Occasionally renal tubular degeneration
was noted. The lumen of degenerated tubules contained copious amounts
of sloughed cells.
The histopathological findings in the spleen were similar but with more
prominent erythrophagocytosis. The spleen featured depletion of haematopoietic
tissue (Fig. 2).
Rainbow trout. Gill: Chlorine exposed gills featured
inter-lamellar epithelial hyperplasia, vacuolation and hypertrophy
of chloride cells. Note the epithelial lifting and mild telangiectasis
in the secondary lamellae
||Rainbow trout. Spleen: Increase in golden pigments,
azurophilic bodies and erythrophagocytosis. Note haematopoietic tissue
Variable amounts of colorless cytoplasmic vacuolation and small hyaline
droplets were seen in the hepatocytes. Foci of phaged golden pigment were
scattered throughout the liver. In the skin mild epithelial erosion accompanied
with mild leukocytic transmigration and elevated numbers of melanocytes
were recorded. Mild mural reactions in the myocardium were the other histopathological
findings in these cases. There were no significant findings in the brain,
pyloric caecae and pancreas. The condition appeared to be primarily due
to red blood cells destruction and is consistent with the marked anemia
that was confirmed by paraclinical examination. History and findings were
consisted with met-haemoglobinaemia, which seemed to be related to excess
Chlorine toxicity can present as acute to subacute mortality associated
to a newly set-up tank or when fresh tap water is used for a water change
(Noga, 1996). Sensitivity of trout and other fish to chlorine has been
reported and the toxicity of chlorine to freshwater fish has been reviewed
by several workers (Yonkos et al., 2000; Heath, 1997). Mitz and
Giesy (1985) have reported a 100% mortality in channel catfish kept in
cage in a sewage effluent biomonitoring study and noted that the mortality
was probably due to excessive mean total residual chlorine concentration
(0.24 and 0.30 mg L-1). Histopathological study on this fish
revealed two or more moderate to severe histopathological damages to the
gills, including severe hyperplasia of the epithelial cells, clubbing
and fusion of the secondary lamellae, moderate to severe edema in the
secondary lamellae and multiple, blood-filled aneurysms and extremely
vacuolated livers in 60% of the fish.
Rainbow trout is very sensitive to chemical environmental changes and
even have been considered as a good experimental model as an indicator
for toxicity in aquatic animals (Ferraris et al., 2005; Kilemade
et al., 2002). According to substance toxicity classification accepted
for Lithuanian inland waters, chlorine dioxide and chlorite can be referred
to substances of `moderate` toxicity to fish. The acute and chronic toxicity
of chlorine dioxide and chlorite to larval and adult rainbow trout was
investigated in 96 h to 20 day laboratory exposures. Both chemical compounds
induced similar toxic effects in rainbow trout larvae during chronic tests,
but chlorine dioxide had a higher toxic potency than chlorite (Svecevicius
et al., 2005).
Yonkos et al. (2000) in an experimental study found that gill
epithelium was the primary target tissue affected by chlorine dioxide
exposure. Chlorine dioxide exposure produced dose-dependent gill pathology
including epithelial lifting, hypertrophy, hyperplasia, lamellar fusion
and necrosis in Fat head minnow (Pimephales promelas). Chlorite did not
produce gill pathology even at a lethal exposure level (304 mg L-1
for 96 h) but did elicit a chronic inflammatory response with a marked
increase in circulating and fixed phagocytes within hematopoietic and
vascular tissues (Yonkos et al., 2000).
The gill histopathology features in our study is same as those reported
by other workers and resembles a typical irritant response beginning with
epithelial lifting, hypertrophy and hyperplasia. Telangiectasis of secondary
lamella is associated with chemical pollution (Roberts, 2001). This can
result to lamellar fusion and necrosis that was not seen in our study
that can be due to the acute nature of the toxicity in this study Lamellar
edema of the gills is most frequent following exposure to chemical pollutants
such as heavy metals, red tides, certain pesticides and therapeutic formalin
or hydrogen peroxicide. In gills damaged by the effects of acidification
of the water supply due to acid rain and subsequent increase in solubility
of soil aluminum, secondary lamellar swelling occurs. This is associated
to a degree with lamellar edema and hypertrophy of individual epithelial
cells. There is alteration in underlying pillar cell architecture, but
the principal factor is significant increase in the numbers of chloride
cells. These extend on to the surface of the secondary lamellae and instead
of being located in sunken pits, bulge out in the surface (Roberts, 2001).
Toxic organic compounds like chlorinated hydrobons herbicides and organophosphate
insecticides can cause nephrotoxic lesions like desquamation of tubular
epithelium, dilation of lumina and tubular necrosis (Roberts, 2001). Toxic
causes of haemolytic anaemia are highly variable, but commonly include
chlorine exposure. Not only is there destruction of erythrocytes, but
in the presence of nitrogenous wastes chloramines is also formed. These
lead to methaemoglobin formation, thereby reducing the oxygen-carrying
capacity of the blood (Noga, 1996).
Fish liver is particularly susceptible to chemical damage. Free-radical
damage to hepatocytes cell membrane following toxic conditions can cause
widespread necrosis and ceroid build up in remaining hepatocytes. Lipid
infiltration often is disseminated throughout the liver. In haemolytic
anaemias, ferrous iron is stored in the melanomacrophages of the liver
(Noga, 1996). The vacuolation observed in our cases can be related to
the haemolytic anaemia caused by hypoxia but the other microscopic features
noted by other workers were not observed in this study.
Glumerular disease can result in protein leakage into the filtrate, recognized
histologically as homogenous eosinophilic deposits within tubular lumens
and presumably leading to proteinuria (Ferguson, 1989; Svecevicius et
al., 2005). Their significance is unknown, although several authors
have noted an association with toxicants and high ammonia levels.
Present findings in the spleen supports other workers finding who suggest
that, in acute hemolytic crises, most of the blood pigments accumulate
in the spleen rather than the kidney.