A study for investigating the values
of plasma glucose, serum total protein and serum ions (calcium,
potassium, magnesium, sodium and chloride) was carried out in young Colossomoma macropomum Cuvier, 1818, Piaractus mesopotamicus Holmberg, 1887 and the hybrid tambacu (P. mesopotamicus x C. macropomum), kept in intensive system. Glucose concentrations were higher in tambacu than in P. mesopotamicus and C. macropomum. Total protein levels were higher in P. mesopotamicus than both C. macropomum and tambacu. C. macropomum presented higher concentrations of sodium and chloride, while P. mesopotamicus
had higher concentrations of potassium and magnesium. However, levels
of calcium were similar for the species studied. The hybrid tambacu
showed the smallest levels of total protein, and intermediate levels of
sodium, potassium, magnesium and chloride, compared to both P. mesopotamicus and C. macropomum.
Baseline values for healthy fish in aquaculture were established, and
they may be used for further comparisons in studies with wild
populations of P. mesopotamicus and C. macropomum.
KEYWORDS: Biochemistry, blood, Colossoma macropomum, freshwater fish, Piaractus mesopotamicus.
Estudaram-se os valores de glicose
plasmática, proteína total sérica e íons séricos (cálcio, potássio,
magnésio, sódio e cloreto) em jovens de Colossomoma macropomum Cuvier 1818, Piaractus mesopotamicus Holmberg 1887 e híbrido tambacu (P. mesopotamicus x C. macropomum), mantidos em cultivo intensivo. As concentrações plasmáticas de glicose no híbrido tambacu foram maiores que em P. mesopotamicus e C. macropomum. Os níveis de proteínas totais em P. mesopotamicus foram maiores em C. macropomum e híbrido tambacu. O C. macropomum apresentou maior concentração de sódio e cloreto, ao passo que o P. mesopotamicus
mostrou maior concentração de potássio e magnésio. Porém, os níveis de
cálcio mostraram-se similares nas três espécies estudadas. O híbrido
tambacu apresentou os menores níveis de proteínas totais e níveis
intermediários de potássio, sódio, magnésio e cloreto quando comparado
ao P. mesopotamicus e C. macropomum.
Foram estabelecidos valores basais para peixes sadios criados em
cativeiro, os quais poderão ser usados em estudos de comparação futura
em populações selvagens de P. mesopotamicus e C. macropomum.
PALAVRAS-CHAVES: Bioquímica, Colossoma macropomum, peixes de água doce, Piaractus mesopotamicus, sangue.
Interspecific crossing has been used
in several fish species to increase genetic and phenotypic variation
for both commercial and research purposes (CNAANI
et al., 2004). In Brazil,
Piaractus mesopotamicus Holmberg, 1887 and
Colossoma macropomum
Cuvier, 1818, both species of great economical importance originated
the hybrid tambacu. These fish are widely appreciated in fish farming
due to quick growth and weight gain, in addition to sportive and
pleasure fishing. However, there is little knowledge of the blood
biochemical profile of these fish.
The assessments of blood biochemical parameters are important to
evaluate the health of many vertebrates, including fish (CNAANI
et al., 2004; PEDRO
et al.,
2005; TAVARES-DIAS & MORAES, 2007). Since blood and biochemical
parameters in fish may vary with ambient and other factors
(BENTINCK-SMITH
et al., 1987; TAVARES-DIAS, 2004; TAVARES-DIAS & MORAES, 2004; TAVARES-DIAS
et al., 2004; PEDRO
et al.,
2005; TAVARES-DIAS & MORAES, 2007), they have been used by fish
biologists for a variety of purposes, such as detecting cellular damage
caused by toxicant exposure, infection by pathogenic agents, and
traumatic handling. In addition, the blood biochemical assessments can
also be used to evaluate the effects of the diet on liver function, and
the osmoregulatory and ionoregulatory functions, effects of sex and
maturation cycle and responses to stressors.
In freshwater fish, transmembrane ion gradients are
the key for several physiological processes. Gill cells are exposed to
different ion gradients across their apical and basolateral surfaces as
a result of their exposure to water and blood, respectively, requiring
different membrane transport mechanisms for intracellular and blood
ionic balance. Thus, when sodium concentrations are increased,
potassium is generally decreased in fish plasma or serum (FURIMSKY
et al.,
1999). The maintenance of a consistent internal environment is
essential for normal cells to function in multicellular organisms. The
ionic composition of this internal environment, the extracellular
fluid, must remain within narrow limits to maintain the transmembrane
electrical potentials of different cell types. Specialized organs and
organ systems have evolved in vertebrates to carryout the maintenance
of the homeostasis of fluid compartments. Most commonly, the kidneys
are the organs that play a major role in controlling fluid and
electrolytes (BALDISSEROTTO, 2002; BRAUN, 2003). However, in fish,
renal function may be considered of a relatively minor role depending
on the external environment. Thereby for many fishes, the gills,
bladder and intestine make large contributions to ionic and fluid
regulation (BRAUN, 2003).
Calcium taken up from the water in freshwater fish follows a
transcellular, hormone-controlled pathway located in the chloride cells
of the gills. Magnesium is an essential element for all
vertebrates, indispensable for proper functioning of all cells,
involved in a variety of enzymic reactions (FLIK
et al.,
1993) in intermediate metabolism, skeletal tissue metabolism,
osmoregulation, and neuromuscular transmission (LIM & KLESIUS,
2003). At least 80% of the magnesium required for growth and
homeostasis is absorbed from the food via intestine, while the gills
contribute with less than 20% (FLIK
et al.,
1993). In fish, the sodium and potassium are predominant electrolytes,
with the predominance of sodium in the serum and in other fluids and
potassium in extracellular fluids. The main function of proteins
and electrolytes, mainly the sodium and potassium, is to
regulate the acid-basic balance maintaining thereby an ionic adequacy
on the tissue functions (DAVIS, 2004; TAVARES-DIAS, 2004; TAVARES-DIAS
et al., 2008). The levels of glucose and osmoregulatory disturbances can be used as stress indicators in fish (BENTINCK-SMITH
et al., 1987; ZUIM
et al., 1988; CNAANI
et al.,
2004; DAVIS, 2004; TAVARES-DIAS & MORAES, 2007). Thus, many
parameters have been used as quantitative indicators of stress.
In the present study it was proposed to determine the baselines values of plasma glucose, total protein and serum ions for
P. mesopotamicus and
C. macropomum
and also for their hybrid, the tambacu, because biochemical profile of
these fish when kept in intensive system has not been reported.
MATERIALS AND METHODS
Fish
Forty clinically healthy specimens of young
Piaractus mesopotamicus (301.5-839.0g and 24.5-34.0cm),
Colossoma macropomum
(369.5-873.0 g and 26.0-37.0 cm) and hybrid tambacu (199.0-418.5g and
22.0-29.0 cm) were obtained from a commercial fish farm (21º 07’45”S,
48º03’57”W), and transported to the Aquaculture Center (CAUNESP),
Jaboticabal, São Paulo, Brazil. These fish were maintained in twelve
cement boxes (500 L) and acclimatized for 15 days, with controlled
flow-through water system. These fish were fed with pelleted diets (35%
of crude protein) once a day. During the acclimatization and
study periods the fish were observed daily for any clinical signs of
diseases, including lack of appetite, increased opercular movements, or
visible lesions of the skin, tail and fins. Signs of disease were not
apparent during either period of observation. The water temperature
varied from 25.1 to 29.8ºC; pH 6.5 to 7.0; dissolved oxygen 5.2 to 6.3
mg/L; electric conductivity 111.6 to 139.5 µS/cm.
At the fish farm, the three species lived in similar water conditions.
This fact prevented major differences in the maintenance of the fish.
The
P. mesopotamicus, C. macropomum
and hybrid tambacu fry received pelleted and extruded fish diets with
42% and 35% of crude protein, respectively. After that, fish were
stocked in ponds of 1,000 m2 that received extruded ration with 28% of
protein. During this period the water temperature was within
24.5-30.2ºC; pH 6.7-6.9; dissolved oxygen 5.4-6.2 mg/L; electric
conductivity 115.6-129.8 µS/cm and pluviometric index 0.0-342.0 mm³
Blood collection procedure and biochemistry analyses
Two blood samples were collected by puncture of the caudal vessel as
follows, one with syringe containing 10%-EDTA and another without any
anticoagulant. All these procedures occurred within 0.8-1.2 minutes, to
avoid sampling stress. Blood samples collected with anticoagulant were
used for the determination of plasma glucose by glucose oxidase. The
second blood sample, which was taken without anticoagulants, was
left for 10 minutes at room temperature and then centrifuged for 10
minutes at 750 G to obtain the serum that was collected and frozen at
-80ºC until analyses were done. Total serum protein concentration was
determined by the biuret reaction. Serum ion concentrations were
determined in flame photometer (Zeiss M4Q2) by emission (sodium and
potassium) and atomic absorption (calcium and magnesium). Serum
chloride concentration was determined by the thiocynate method using a
commercial kit (Sigma 461).
Statistical analysis
The parameters were statistically analyzed using ANOVA followed by
Tukey test, when the differences were significant (p<0.05).
RESULTS AND DISCUSSION
Biochemical parameters assessments of blood are important in evaluating the fish health (TAVARES-DIAS & MORAES, 2004; PEDRO
et al.,
2005; TAVARES-DIAS & MORAES, 2007), in captivity or wild species.
In intensive culture, osmoregulatory disturbances can occur due to
diseases and several other factors (BENTINCK-SMITH
et al.,
1987; STOSKOPF, 1993; TAVARES-DIAS, 2004). Hence, aquaculture needs
constant accompainment of fish health status to avoid compromising
performance parameters and the financial aspects of fish farming.
Biochemical baselines values established may allow important clinical
decisions about fish species (TRIPATHI
et al., 2003; TAVARES-DIAS & MORAES, 2007).
C. macropomum have higher concentration of sodium and chloride than
P. mesopotamicus and tambacu, but
P. mesopotamicus
presented higher potassium and magnesium concentration and smaller
sodium concentration, while calcium level was similar for these three
fishes (
Table 1).
Since these species are originated from the same environment and
similar conditions, such variation levels are specific-species.
Interspecific variation have been also reported for protein, calcium
and magnesium levels in
Oreochromis aureus,
O. mossambicus, O. niloticus and hybrid red tilapia (CNAAI
et al., 2004), for sodium, potassium and chloride levels in
Brycon amazonicus (= cephaus) and
B. orbignyanus (TAVARES-DIAS, 2004), as well as for sodium, calcium, magnesium and chloride in
Leporinus macrocephalus and
Prochilodus lineatus (TAVARES-DIAS
et al.,
2008). Furthermore, minerals in the water, adequate gill function, and
alterations in water intake influence biochemical parameters in fish
(TRIPATHI
et al., 2003).
It has been stated that the glucose concentration depends on the fish
life mode, and particularly on its locomotive capacity (CARNEIRO &
AMARAL, 1979). Moreover, glucose levels have been attributed to the
stress caused by the low oxygen rate, the manipulation and diet, and
also to brachial damage. The characids
C. macropomum (VILLACORTA-CORREA & SAINT-PAUL, 1999) and
P. mesopotamicus (VAZ
et al.,
2000) are omnivorous and migratory fishes, which when in natural
environment, develop as alimentary strategy the accumulation of
reservations during periods of high food offer, to survive thereby in
periods of shortage. BALDISSEROTO (2002) reported that fish with
omnivorous habits are often exposed to natural food with variable
quantity of carbohydrates. They need to be capable to regulate the
glucose absorption according to its available quantity in the food. The
glucose levels were higher in hybrid tambacu than in
P. mesopotamicus and the latter one presented higher levels of glucose than
C. macropomum (
Table 1).
Interspecific variation in glucose levels was also reported for fish
species of different ecological habitat (HRUBEC & SMITH, 1999;
RODRIGUES
et al., 1999). In contrast, in
L. macrocephalus and
P. lineatus, which have different ecological habitats, the values of plasma glucose were similar (TAVARES-DIAS
et al., 2008). However, in
Brycon amazonicus and
B. orbignyanus,
which have similar ecological habitat, the values of plasma glucose
were similar (TAVARES-DIAS, 2004), as expected. Biochemical parameters
diverge among individuals and among species, too (TAVARES-DIAS &
MORAES, 2007), and glucose seasonal changes (BENTINCK-SMITH
et al., 1987).
In the present study, total protein levels of tambacu were smaller than in
C. macropomum and
P. mesopotamicus, but in this latter it was higher than in
C. macropomum. Similar interspecific variation in total protein levels has been reported for
L. macrocephalus and
P. lineatus of intensive culture (TAVARES-DIAS
et al., 2008). However, total protein levels of
P. mesopotamicus were similar to this same wild specie when in captivity (RANZANI-PAIVA & GODINHO, 1988) and in natural habitat (ZUIM
et al., 1988). Hence, the values found here may be considered baselines for cultured
P. mesopotamicus. On the other hand, total protein levels of
C. macropomum were higher than of this same specie cultivated in net cages (CHAGAS
et al., 2005).
A low level of total protein may occur in consequence of alimentary restriction (CHAGAS
et al.,
2005), of protein synthesis reduction due to hepatic disfunction, and
also in consequence of a permeability increase for plasma proteins or
of proteins degradation by the proteolytic enzymes that are released
from endothelial cells destroyed by virus or bacteria (STOSKOPF, 1993).
On the other hand, increases of total protein levels may be associated
to hemolysis (HRUBEC & SMITH, 1999). Thus, it has been reported
that captive adults of
P. mesopotamicus
have higher levels of plasma total protein when compared to wild fish
due to the reduction in energy expenditure and the reproductive
blockage that occurs in lack of migration (ZUIM
et al.,
1988). Therefore, these results indicate how important it is to
establish baseline values, which can be used to further assessment of
fish health status.
CONCLUSION
Tambacu hybrid had the smallest levels for total protein and
intermediate levels for sodium, potassium, magnesium and chloride when
compared to
P. mesopotamicus and
C. macropomum.
Only calcium levels were similar for the fish studied. It is unlikely
that species different in taxonomic and ecological affinities have
identical metabolic patterns that result in uniform blood biochemistry
values. Since biochemical parameters may be affected by several
factors, to avoid these variations, hybrid tambacu,
P. mesopotamicus and
C. macropomum
were sampled from a defined reference population that was maintained
and managed under identical environmental conditions. Furthermore,
laboratory procedures to analyze the serum specimens were also
performed through standardized techniques and protocols. Hence, the
biochemical levels variation found were specific-species. Therefore,
the baseline values obtained here may provide general guidelines for
the interpretation of laboratory data for these species of fish, which
are very important for Brazilian aquaculture.
ACKNOWLEDGMENTS
The authors gratefully acknowledge research grants from Fundação de
Amparo à Pesquisa do Estado de São Paulo (FAPESP, Grant N.o
00/05676-2), Brazil.
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Protocolado em: 8 jul. 2007.
Aceito em: 3 nov. 2009.