Comp Clin Pathol (2010) 19:301–305
DOI 10.1007/s00580-009-0871-2
ORIGINAL ARTICLE
Comparison of serum enzyme activity in great sturgeon,
Huso huso, cultured in brackish and freshwater
earth ponds in Iran
Farhad Rajabipour & Davar Shahsavani & Ali Moghimi &
Shahla Jamili & Nassrin Mashaii
Received: 16 December 2008 / Accepted: 4 June 2009 / Published online: 23 June 2009
# Springer-Verlag London Limited 2009
Abstract Blood samples were collected from 60 great
sturgeons, Huso huso, to establish the following serum
enzyme activity: aspartate amino-transferase (AST), alanine
amino-transferase (ALT), lactic acid dehydrogenase (LDH),
creatine kinase (CK), and alkaline phosphatase (ALP) using
an autoanalyzer, and acid phosphatase (ACP) by manual
method. Thirty 5-year-old cultured fish were caught from
each of two sites; a brackish-water earth pond in Bafgh and
a freshwater pond in Gorgan in the centre and northeast of
Iran, during May 2006. Results of the serum enzymes
activity for H. huso samples from Bafgh and Gorgan were:
AST, 502.9±258.2 and 436.1±186.8; ALT, 104.4±35.1 and
53.1±38.7; LDH, 3094.2±1277.5 and 2486.3±1393.3; CK,
3632.9±2618.7 and 3967±5054.9; ALP, 281.2±112.7 and
762.2±600.2; ACP, 13.3±2.5 and 33±6.8 IU/L. Mean
values of ALT, ALP and ACP were significantly different
in the fish from the two sites (p<0.05). These results may
F. Rajabipour (*) : N. Mashaii
Iranian Fisheries Research Organization, Brackishwater Fisheries
Research Station,
P. O. Box: 89715/1123, Bafgh,
Yazd, Iran
e-mail: Farhadrajabipour@yahoo.com
D. Shahsavani
Faculty of Veterinary, Ferdowsi University,
Mashhad, Iran
A. Moghimi
Department of Biology, Faculty of Sciences, Ferdowsi University,
Mashhad, Iran
S. Jamili
Iranian Fisheries Research Organization,
# 297, West Fatemy Ave.,
Tehran, Iran
be used to understand some biological (e.g., serum enzyme
activity) and ecological characteristics of cultured H. huso.
Keywords Sturgeon . Huso huso . Culture .
Serum enzymes . Iran
Introduction
Sturgeons are very important commercial fish because of
their expensive caviar. The Caspian Sea contains large
stocks of these fish but some species are becoming extinct,
resulting in the culturing of different species of sturgeons in
countries neighbouring the Caspian Sea (Bahmani 1998;
Kim et al. 2006). The great sturgeon Huso huso (Linnaeus
1758) has the most expensive caviar and has been cultured
in Iran since 1991 (Yousof-pour 1992). The population
numbers of H. huso is dependent on several complex
factors, such as: conditions for reproduction, abundance
and quality of spawners and food supply. In the 1990s, a
decline in the spawning population of the Ural River was
recorded and by the year 2000, the population consisted of
2,660 individuals. The population within the Volga River
also showed an enormous reduction when compared with
previous periods; 1971–1975, 20,700 specimens, 2,000 tons
and 1991–1995, 8,000 specimens, 500 tons. In the 1980s,
the abundance of great sturgeon in the sea was over 18
million individuals, however, this has dropped to 8 million
specimens (Bahmani 1996; Kim et al. 2006).
Many investigations concerning the physiological
aspects of sturgeon species have been carried out (Slynko
1976; Metallov and Aksenov 1999; Shahsavani et al. 1999,
2001; Ghorbanianfar 2003; Falahatkar et al. 2005;
Shirazian 2005; Taghvaiimoghaddam 2005; Rajabipour
2006; Khoshbavar Rostami et al. 2006; Asadi et al. 2006).
302
Haematological tests and analyses of serum constituents
have proved useful in the detection and diagnosis of
metabolic disturbances and disease processes. The lack of
physiological values for various blood parameters prevents
their use in assessing the condition of the aquatic environments and ecosystems (Çelik 2004).
The purpose of this study was to establish serum enzyme
activity to provide baseline data to detect differences between
the great sturgeon, H. huso cultured in brackish and
freshwater as two different ecological conditions in Iran.
Materials and methods
Experimental design Sixty great sturgeon, H. huso fish
were collected during May 2006 using a gill net from earth
ponds at two stations: a fisheries research farm in Bafgh
(brackish water) and Shahid Marjani fish aquaculture centre
in Gorgan (freshwater), situated in the centre and northeast
of Iran. All fish were initially hatched in the same hatchery
at Shahid Marjani from eggs from a single individual, but
were cultured in two different areas for 5 years. Following
the catch, total length and body weight were measured and
recorded, as was water temperature, salinity, hardness, pH
level and dissolved oxygen.
Blood collection Thirty fish in each site were handled
carefully to reduce any possibilities of stress. Blood
sampling was performed immediately after the fish were
captured. Samples were collected from behind the anal fin
using a 5-mL plastic syringe and 22 gauge needle. The
blood samples, approximately 2.5 ml were placed into dry
plastic tubes and immediately centrifuged at 4,000 rpm for
10 min (Çelik 2004; Casillas et al. 1983).
Biochemical analyses Separated serum samples were analyzed for aspartate amino transferase (AST), alanine amino
transferase (ALT), lactic acid dehydrogenase (LDH),
creatine kinase (CK) and alkaline phosphatase (ALP) by
Cobas Mira 27-6764 auto-analyzer using enzymatic procedures with a diagnostic kit (Pars Azmoon Chemical Co.).
Serum enzyme activity for acid phosphatase (ACP) was
undertaken using the manual method. Biochemical measurements were carried out 1 h after sample collection.
Sample collection and biochemical analyses were performed using the methods derived from several different
researchers (Anderson and Anderson 2002; Çelik 2004;
Cech et al. 2000; Shalaby 2005; Pepys et al. 1978).
Comp Clin Pathol (2010) 19:301–305
statistical package. Leven’s test was used for scaling the
equality of variances. Mean values were compared by t test.
Two-tailed Pearson’s correlation between each biometric
and biochemical serum parameter against each other were
measured. Differences were considered to be significant for
p values less than 0.05.
Results
The water quality of the culture ponds in Bafgh and Gorgan
were as follows: temperature 26°C and 23°C, salinity 10 ppm
and 1 ppm, pH 8.2 and pH 8.0, dissolved oxygen 6.3 ppm and
6 ppm and hardness 15,530 μm/cm and 100 μm/cm,
respectively. Mean values of the total length for the great
sturgeon, H. huso from Bafgh and Gorgan was 121.7±9.3
(96–139)cm and 125.7±13.2 (100–150)cm, respectively,
their average body weight was 10,952.8±4,880.4 g (4,200–
20,380) and 12,350±3,436.9 g (8,000–19,000). There was
no significant difference in either total length (p=0.184) or
body weight (p=0.211) between the two sites.
Mean±S.D values of the serum enzyme activity for
H. huso samples from Bafgh and Gorgan were as follows:
AST, 502.9±258.2 (99–1,214) IU/L and 436.1±186.8
(183.9–800.7) IU/L; ALT, 104.4±35.1 (39–192) IU/L and
53.1 ± 38.7 (8.1–163.8) IU/L; LDH, 3,094.2 ± 1277.5
(1,106–5,773) IU/L and 2,486.3±1,393.3 (724–5,001) IU/L;
CK, 3,632.9±2,618.7 (470–10,589) IU/L and 3,967±5,054.9
(531–23,060) IU/L; ALP, 281.2±112.7 (100–535) IU/L and
762.2±600.2 (117–2097) IU/L; ACP, 13.3±2.5 (8–20.5) IU/L
and 33±6.8 (21–49.2) IU/L (see Table 1).
Mean values for ALT, ALP and ACP were significantly
different in the fish from the two sites (p<0.0001), but there
were no significant differences between AST (p=0.225),
LDH (p=0.083) and CK (p=0.749) between the two sites.
Two-tailed Pearson-correlation showed significant direct
correlation between total length with body weight of the
H. huso (p=0.0005); between ALT with AST (p=0.04),
LDH (p=0.0005) and CK (p=0.0005); between LDH with
CK (p=0.006) and ALP (p=0.003); CK with ALP (p=0.0005)
and ALP with ACP (p=0.0005). Significant indirect correlation was obtained between total fish length and ALT (p=
0.001), LDH (p=0.02), CK (p=0.0005) and ALP (p=0.04);
between body weight and ALT (p=0.0005), LDH (p=0.003)
and CK (p=0.0005); and between ALT with ALP (p=0.0005)
as well (Table 1).
Discussion
Statistical methods Mean±SD, variance, standard error,
median, range, quartile, minimum and maximum values
were determined for biometric indices and each of the
serum enzyme activities using a SPSS ver.11.5/2002 system
The mean level of ALT in blood serum in H. huso samples
from Bafgh showed a significant twofold increase when
compared with samples from Gorgan (p<0.0001), suggest-
Comp Clin Pathol (2010) 19:301–305
Table 1 Pearson correlation
and p value between total
length, body weight and serum
enzymes of H. huso
ACP
ALP
CK
LDH
ALT
AST
BW
303
Cor.
p
Cor.
p
Cor.
p
Cor.
p
Cor.
p
Cor.
p
Cor.
p
TL
BW
AST
ALT
LDH
CK
ALP
0.14
0.27
-0.27
0.04
-0.52
<0.01
-0.31
0.02
-0.41
<0.01
0.22
0.09
0.76
<0.01
0.22
0.10
-0.21
0.10
-0.47
<0.01
-0.38
<0.01
-0.48
<0.01
0.23
0.08
-0.05
0.69
-0.01
0.97
0.01
0.92
0.19
0.14
0.27
0.04
-0.56
<0.01
0.18
0.17
0.48
<0.01
0.63
<0.01
-0.16
0.24
0.38
<0.01
0.35
0.01
0.03
0.82
0.47
<0.01
0.45
<0.01
ing that liver disorders may be more noticeable in cultured
fish from this area. Imprecise feeding management may
affect the activity of ALT in serum (Cech et al. 2000), also
higher water salinity of pond culture in the Bafgh area may
lead to an increase in the serum ALT level as previously
shown by Shalaby (2005) in the Nile tilapia, Oreochromis
niloticus. Androgen hormone levels may be higher in great
sturgeons in Bafgh, as shown by Casillas et al. (1983) in the
English sole, Parophrys notopterus notopterus. Increased
maturation is highly favoured in commercial fisheries for
the acquisition of caviar. Indirect significant correlations
were observed between ALT and total length and body
weight for H. huso samples. Low ALT activity was
recorded in large H. huso sturgeon from the Caspian Sea
(Taghvaiimoghaddam 2005). ALT is the optimum serum
factor for the prognoses of liver disorders (Shalaby 2005).
High AST activity in samples from Bafgh and Gorgan,
suggests more investigation into the concentration of heavy
metals in the earth ponds water is required. Many metals,
whether organically complex or not, are known to accumulate in plant and animal tissues to a very high level,
posing a potential toxic hazard to the organisms themselves,
or organisms higher in the food chain, including humans
(Abel 1998). Earth ponds for culturing sturgeon are close to
different mines in Bafgh and near to rice fields and
habitable areas in Gorgan, indicating that the viral
infections of the fish may be similar at the two fish culture
centres (Kaneko1980) due to the similarity of AST activity
between the two sites.
The serum activity of ALT and AST obviously varies
depending on fish species (Shalaby 2005). The increase in
plasma AST and ALT may be connected to stress conditions,
hepatocellular damages or cellular degradation due to heavy
metals, perhaps in the liver, heart or muscle (Yokoyama et al.
2003; Alter and Tokars 2001; Yamawaki et al. 1986).
Infections cause an increase in both ALT and LDH
(Smith 1987). ALT was significantly different in the
examined fish from the two areas, however, LDH activity
in the fish from Bafgh was not much higher than Gorgan
(p>0.05), therefore, infection may not be present in these
samples. LDH levels correlate directly with growth rate
(Khajali and Qujeq 2005; Hart and Reynolds 2002). The
activity of this enzyme in muscle and liver tissue of
ascipenserids were recorded as being similar to other
teleosts (Metallov and Aksenov 1999).
Muscle density and damage are probably similar at the
two centres since no significant difference in CK levels (p>
0.05) was found but levels are probably less than the large
great sturgeons from the Caspian Sea (Taghvaiimoghaddam
2005). Together, the increase in CK and ALT/AST show a
probable decrease in protein catabolism (Cech et al.
2000).
There were significantly higher levels of ALP in fish from
both sites compared to fish from the sea (Taghvaiimoghaddam
2005) and with values close to the common carp (Peyghan et
al. 2003). The activity of this enzyme is normally higher in
young vertebrates (Cech et al. 2000), suggesting that ALP
activity may differ in fish by age and size (Trivedi et al.
2001; Das 1966). Although the examined fish were of the
same age, an indirect significant correlation was observed
between ALP activity and the size of the fish. Increase in
ALP may be caused by the obstruction of the biliary canal or
osteoblast activity and feeding conditions (Moraes et al.
2005; Cech et al. 2000). Higher levels of ALP and similar
levels of AST, LDH and CK were observed in H. huso
samples from Gorgan compared with Bafgh. Similar findings
have been previously reported in canal catfish compared
with pond-cultured samples (Warner and Williams 1977).
ACP was significantly higher in fish from Gorgan (p<
0.05), indicating that phosphatase esters may be hydrolyzed
304
faster in these samples. Heavy metal pollution especially,
cadmium (Shalaby 2006) and mercury (Hossain and Dutta
1986) may be noticeable as well as some viral infections,
for example Myxobolus cerebralis. Lysosomal proliferation
and cellular damage (Moraes et al. 2005), plus disorders of
calcium and calcitonin (Suzuki 2005) may also be present.
Increased water hardness in Bafgh (Mashaii 2006) may
cause a decrease in ACP and ALP following an increase in
plasma calcitonin levels (Mukerjee et al. 2004; Chen 2003).
Although high variances are expected in serum enzyme
measurements (Folmar et al. 1992, 1993), variations
between blood chemistry parameters among fish species
depend on the sampling technique, analysis methods, age,
habitat and diet (Çelik 2004). This study gives valuable
information on the health of an economically important fish
species; the great sturgeon. The result of this study may
allow us to understand biological and ecological characteristics of cultured H. huso.
Acknowledgements This study was supported in part by the Iranian
Fisheries Research Organization. Many thanks to Dr. Sharif-Rohani,
Dr. Hossein-Zadeh, Mr. Sarsangi and Mr. Mohamadi for cooperation.
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