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Histopatholgical manifestations in kidney of Clarias batrachus induced

by experimental Procamallanus infection

Shashi Ruhela1, A.K. Pandey*2 and A.K. Khare1

1Department of Zoology, Meerut College, Meerut - 250 001, India2Central Institute of Freshwater Aquaculture, Kausalyaganga, Bhubaneswar - 751 002, India

(Received: November 15, 2006; Revised received: June 25, 2007; Accepted: July 02, 2007)

Abstract: Kidney of Clarias batrachus infected with Procamallanus showed varying degrees of histopathological alterations on 15, 30, 45 and 60 days post-

infection. The infected kidney showed variable sized glomeruli, cloudy swelling in tubules, vacuolar/atrophic degeneration, fibrosis, mild degenerative changes

in distal convoluted tubules, enlarged Bowmen’s capsule, necrotic changes as well as increased granulation and hyperplasia in proximal convoluted tubules

after 15 days. After 30 days of infection, the changes were rupture of Bowmen’s capsule wall, degenerative changes, edema, necrosis, pyknosis, karyorrhexis

and karyolysis in proximal and distal convoluted tubules, fibrosis, cloudy swelling and inflammatory lymphocytes, proliferation and shrinkage in glomeruli, and

vacuolization in proximal convoluted tubules as well as cloudy swelling. After 45 days, the infected kidney showed cloudy swelling in glomeruli as well as

variation in their size, infiltration of RBCs in intralobular vein and necrosis in proximal convoluted tubules, cloudy swelling in interstitium, vacuolization in the

epithelial lining cells, necrosis in haemopoietic tissue and inflammatory edema. After 60 days post-infection, the changes were rupture of intralobular vein,

cloudy swelling, necrosis in few proximal convoluted tubules, atrophy and shrinkage in glomeruli, distinct inflammatory edema, pyknosis, karyorrhexis and

karyolysis, aggregation of lymphocytes and dilation in blood vessels.

Key words: Procamallanus, Histopathology, Kidney, Clarias batrachus

PDF of full length paper is available with author (*akpandey_cifa@yahoo.co.in, cifa@ori,nic,in, cifa@hun1.nic)

Introduction

Procamallanus is a common nematode parasite inhabiting

intestine of freshwater, brackishwater and marine fishes (Yamaguti,

1961; Sood, 1988; Chandra, 1994; Bijukumar, 1996; Lakshmi, 2000;

Lakshmi and Kumari 2000; Moravec et al., 2004). It has been

frequently reported from intestine of the commercially important Indian

freshwater catfishes such as Clarias batrachus (Sinha, 1988, 2000)

and Heteropneustes fossilis (Chandra, 1994; De and Maity, 2000;

Katoch, 2002). Helminth infection causes deterioration in food value

of the flesh and may result in heavy mortality of fishes (Madhavi,

2003). Further, heavy parasitic worm burden in fish may reduce

reproductive potential or delay sexual maturity serving as a limiting

factor to population size of the species. Chinabut et al. (1991) reported

the occurrence of Procamallanus planolatus in stomach and intestine

of Clarias macrocephalus and observed cloudy swelling, fibrosis

and degeneration in kidney of the infected fish. Since parasites

secrete/excrete certain endotoxins in circulation of the host which

affect the other body tissues/systems including alterations in the blood,

enzymes, vitamins and hormonal activity (Poynter, 1966; Chinabut

et al., 1991; Roberts, 2001; Ekman and Norrgren, 2003), an attempt

was made to record the histopathological alterations in kidney of

Clarias batrachus induced by experimental Procamallanus infection.

Materials and Methods

Clarias batrachus used in the present study were collected

from local freshwater ponds and also purchased from fish markets of

Meerut and adjoining region in Uttar Pradesh, India. Fishes were

acclimatized to laboratory conditions for about a week before initiating

the experiment. The adult female Procamallanus were collected

from infected Clarias batrachus by cutting open the intestine. They

were kept in watch glass filled with saline solution for natural egg

laying at 24-27oC. The eggs were kept in Lock-Lewis solution for

healthy embryonation. The solution was changed periodically and

0.1% formalin added to the culture medium to avoid fungal

contamination of the eggs. 40 healthy catfish (total length range 24-

26 cm; weight range 138-160 g) were randomly selected and divided

into two equal groups. Catfishes of group 1 were not given any

treatment and served as control whereas in catfish of group 2

experimental infection was induced by forcefully pushing 500

embryonated eggs of the nematode into stomach of each catfish by

means of a long-nozzled dropper (De and Maity, 2000; Ruhela et

al., 2006). Catfishes from the experimental as well as control groups

were killed on day 15, 30, 45 and 60 and kidney were surgically

removed and fixed immediately in freshly prepared aqueous Bouin’s

solution. After routine processing in ascending series of alcohol and

clearing in xylene, the tissue were embedded in paraffin wax at

60oC. Serial sections were cut at 6 µm and stained in haematoxylin

and eosin for histopathological examinations.

Results and Discussion

Trunk kidney of the control catfish, Clarias batrachus

consisted mainly of haemopoeitic tissue, nephrons, convoluted tubules

and collecting ducts. Each nephron possessed a renal capsule, a

short neck and convoluted tubules. The renal capsule comprised

glomerulus and Bowman’s capsule. Glomerulus was formed by

central rounded compact mass of numerous mesengial cells which

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Fig. 6: Kidney of infected catfish on day 30 depicting ruptured Bowmen’s

capsule wall (�), degenerative changes in proximal convoluted tubule (�),

edema (�) and shrinkage in glomeruli (�). H and E × 100

Fig. 1: Kidney of control Clarias batrachus on day 15 showing normal

structure of renal cortex with Bowmen’s capsule (�), glomeruli (�), blood

vessels (�), proximal and distal convoluted tubules (). H and E × 100

Fig. 2: Kidney of control catfish on day 15 exhibiting normal structure of

cortex, glomeruli (�), blood vessels (�) and tubules (). H and E × 200

Fig. 5: Kidney of infected catfish on day 15 exhibiting mild degenerative

changes in distal convoluted tubule (�), proximal convoluted tubule ()

and enlarged Bowmen’s capsule (�). H and E × 200

Fig. 3: Kidney of infected catfish on day 15 depicting variable sized glomeruli

(�), cloudy swelling in tubules (�), edema (), vacuolar/hydrophic

degeneration and fibrosis (�). H and E × 100

Fig. 7: Kidney of infected catfish on day 30 exhibiting necrosis, pyknosis,

k a r y o r r h e x i s , k a r y o l y s i s i n p r o x i m a l c o n v o l u t e d t u b u l e (�), fibrosis (�)

and cloudy swelling (�). H and E × 200

Fig. 4: Kidney of infected catfish on day 15 showing cloudy swelling (�),

mild degenerative changes in tubules (�), edema (�), necrotic changes in

PCT (), increased granulation and hyperplasia in PCT (�). H and E × 200

Fig. 8: Kidney of infected catfish on day 45 depicting variable sized glomeruli

(�), cloudy swelling in glomeruli (), infiltration of RBCs in intralobular

vein (�) and necrosis in proximal convoluted tubule (�). H and E × 100

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Procamallanus induced histopathological manifestions in kidney

was surrounded by the tufts of glomerular capillaries. Bowman’s

capsule contained thin squamous epithelium with an outer peritoneal

and inner visceral layers. The proximal convoluted tubule consisted

of cuboidal epithelial cells with basal nuclei, luminal surface being

lined by well-developed brush border. The distal convoluted tubule

comprised cuboidal epithelial cells which occupied only one third of

the total tubule. The cytoplasm of the epithelial lining cells was slightly

granular (Fig. 1-2). The renal tissues of the catfish infected with

Procamallanus showed glomeruli of variable sizes, enlargement of

Bowmen’s capsule, cloudy swelling in tubules, edema, vacuolar/

atrophic degeneration, fibrosis, mild degenerative changes in proximal

and distal convoluted tubules, necrotic changes in proximal convoluted

tubules, increased granulation and hyperplasia in proximal

convoluted tubules after 15 days post-infection (Fig. 3-5). After 30

days of infection, the kidney exhibited rupture of Bowmen’s capsule

wall, proliferation and shrinkage in glomeruli, degenerative changes

such as vacuolization, necrosis, pyknosis, karyorrhexis and

karyolysis in proximal convoluted tubules. Furthermore, cloudy

swelling, edema and inflammatory lymphocytes were also noticed in

proximal convoluted tubules (Fig. 6-7). After 45 days post-infection,

pathological changes included variable sized cloudy swelling in

glomeruli, infiltration of red blood cells (RBCs) in intralobular vein,

necrosis in proximal convoluted tubules, degenerative changes,

cloudy swelling in interstitium, inflammatory edema, vacuolization in

the epithelial lining cells and necrotic haemopoietic tissue (Fig. 8-10).

After 60 days, these changes were more pronounced showing

dilation in blood vessels, cloudy swelling, necrosis, atrophy and

shrinkage in glomeruli, rupture of intralobular vein, edema,

vacuolization, pyknosis, karyorrhexis and karyolysis as well as

necrosis in few proximal convoluted tubules. A distinct inflammatory

edema and aggregation of lymphocytes were also observed after

60 days of Procamallanus infection (Fig. 11-13).

Anderson et al. (1976) observed tubules with flattened dark

staining epithelium and their lumen usually filled with amorphous

basophilic deposit containing occasional cellular elements in the wild

fishes. Sharma and Gautam (1978) observed necrosis in glomeruli

and tubules of sheep infected with experimental toxoplasmosis. The

glomeruli were thickended with increase in glomerular space and

condensation of Bowmen’s capsule. Haemopoietic tissue was largely

replaced by periglomerular and peritubular fibrous tissue and dilated

Fig. 13: Kidney of infected catfish on day 60 exhibiting dilation in blood

vessels (�) and necrosis (�). H and E × 200

Fig. 10: Kidney of infected catfish on day 45 exhibiting necrosis in PCT

(�), vacuolization epithelial lining cells (�), necrotic haemopoietic tissue

() and non-inflammatory edema (�). H and E × 200

Fig. 9: Kidney of infected catfish on day 45 showing degenerative changes

(�), cloudy swelling in interstitium (�) and edema (�). H and E × 200

Fig. 11: Kidney of infected catfish on day 60 depicting ruptured intralobular

vein (�), cloudy swelling (�), necrosis in few proximal convoluted tubule

(�), edema (�) and atrophy/shrinkage in glomeruli (�). H and E × 100

Fig. 12: Kidney of infected catfish on day 60 showing edema (�), cloudy

swelling (�), vacuolization (�), pyknosis, karyorrhexis and karyolysis

(�). H and E × 200

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Ruhela et al.

renal collecting duct (Harrison and Richards, 1979). The present

observations are in agreement with the observations of Klei and

Crowell (1981) in Brugia pahangi infected birds. George and Iyer

(1981) observed variable sizes of glomeruli, degenerative changes

in distal convoluted tubules of rats infected with Trichosomoides

crassicaudo. The kidney showed necrotic tubules after infection

which appears to represent some sort of water and protein

disturbances in the epithelial cells (George and Iyer, 1981). In the

present study in Clarias batrachus, congested blood vessels,

proliferated glomeruli, edema and cloudy swelling were observed

after 15, 30, 45 and 60 days post-infection. After 60 days, these

changes were more pronounced showing necrosis in few proximal

convoluted tubules, atrophy and shrinkage in glomeruli. A distinctive

acute tubular epithelial necrosis associated with interstitial

haemorrhages of the posterior kidney was originally described as

haemorrhagic kidney syndrome in Atlantic salmon (Byrne et al.,

1998).

The histopathological changes observed in kidney of Clarias

batrachus elicited by the experimental Procamallanus infection might

have been induced by excretory/secretory metabolites (exotoxins/

endotoxins) produced by the parasites in intestine (Chinabut et al.,

1991; Roberts, 2001; Ekman and Norrgren, 2003). Similar

pathological changes such as cloudy swelling, variable sized

glomeruli, degenerative changes in proximal convoluted tubules,

distal convoluted tubules and Bowmen’s capsule, necrotic changes

as well as edema were also observed in chick challenged with

experimental Ascaridia galli infection. Necrosis is usually associated

with an early loss of membrane integrity resulting in leakage of the

cell content into immediate tissue environment and the induction of

inflammation (Holle et al., 2003). Kobayashi et al. (2004) also

reported necrosis and edema in fish infected with Pseudomonas

plecoglossicida. Further, the necrotic cells showed karyorrhexis,

karyopyknosis and hyperchromatosis as well as cloudy swelling in

infected kidney of Plecoglossus altivelis.

Acknowledgments

We are thankful to the Head, Department of Zoology, Meerut

College, Meerut for providing laboratory facilities to carry out the

work.

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