Haematological parameters have been recognized

as valuable tools for monitoring fish health. Haematological

and serum biochemical parameters were studied and

compared different feeding behaviour of teleost fishes.

Three marine teleost fishes, Lates calcarifer (carnivores),

Mugil cephalus (omnivores) and Chanos chanos (herbivores),

were carried out in order to find out a normal range

of blood parameters which would serve as baseline data for

assessment of the health status of the fish as well as

reference point for future comparative surveys. Blood

parameters such as red blood cell count (RBC) and white

blood cells count (WBC), haemoglobin, haematocrit, mean

cell haemoglobin concentration (MCHC), mean cell volume

(MCV), mean cell haemoglobin, glucose, protein,

cholesterol and urea were estimated from teleost fishes of

different trophic level. Statistical analysis revealed that

differences in haematological parameters between marine

fish were significant (P<0.01)

Ananthan, G

Jagadeesan, L

Satheeshkumar, P

Senthil Kumar, D

CMFRI Digital Repository

ORIGINAL ARTICLEHaematology and biochemical parameters of differentfeeding behaviour of teleost fishes from Vellar estuary, IndiaP. Satheeshkumar & G. Ananthan & D. Senthil Kumar &L. JagadeesanReceived: 7 December 2010 /Accepted: 7 April 2011# Springer-Verlag London Limited 2011Abstract Haematological parameters have been recognizedas valuable tools for monitoring fish health. Haematologicaland serum biochemical parameters were studied andcompared different feeding behaviour of teleost fishes.Three marine teleost fishes, Lates calcarifer (carnivores),Mugil cephalus (omnivores) and Chanos chanos (herbi-vores), were carried out in order to find out a normal rangeof blood parameters which would serve as baseline data forassessment of the health status of the fish as well asreference point for future comparative surveys. Bloodparameters such as red blood cell count (RBC) and whiteblood cells count (WBC), haemoglobin, haematocrit, meancell haemoglobin concentration (MCHC), mean cell vol-ume (MCV), mean cell haemoglobin, glucose, protein,cholesterol and urea were estimated from teleost fishes ofdifferent trophic level. Statistical analysis revealed thatdifferences in haematological parameters between marinefish were significant (P<0.01). The result revealed thathaematological RBC/WBC ratio, MCV and MCHC weresignificantly correlated at P<0.05 level. The RBC/WBClevel increased due to the decrease in WBC during thestudy. Blood serum biochemical parameters can be used forconfirming the maturity and monitoring any changes in thequality of waters and related soils.Keywords Blood . Fish . Haematology . Erythrocyte .Serum biochemistryIntroductionKnowledge of the haematological characteristics is animportant tool that can be used as an effective and sensitiveindex to monitor physiological and pathological changes infishes. Normal ranges for various blood parameters in fishhave been established by different investigators in fishphysiology and pathology (Rambhaskar and Srinivasa Rao1986; Xiaoyun et al. 2009). The analysis of blood indiceshas proven to be a valuable approach for analysing thehealth status of farmed animals as these indices providereliable information on metabolic disorders, deficienciesand chronic stress status before they are present in a clinicalsetting (Bahmani et al. 2001). Blood biochemistry param-eters can be also used to detect the health of fish (De Pedroet al. 2005). Exogenous factors, such as management(Svobodova et al. 2008), diseases (Chen et al. 2005) andstress (Cnaani et al. 2004), always induce major changes inblood composition. For example, significant fluctuationswere detected in the concentrations of cortisol, glucose,cholesterol and other basic components in response tohandling and hypoxic stress (Skjervold et al. 2001). Thelevels of cortisol and glucose are considered to be specificindicators of sympathetic activation during stress conditions(Lermen et al. 2004). Basic ecological factors, such asfeeding regime and stocking density, also have a directinfluence on certain biochemistry parameters (Coz-RakovacP. Satheeshkumar (*) :G. Ananthan : L. JagadeesanFaculty of Marine Science,Centre of Advanced Study in Marine Biology,Annamalai University,Parangipettai, Tamilnadu 608502, Indiae-mail: revathysathish86@gmail.comP. SatheeshkumarCentral Marine Fisheries Research Institute,Kochi, Kerala 682018, IndiaD. S. KumarDepartment of Zoology,Kandaswamy Kandar Arts and Science College,Paramathy-Velur, Tamilnadu 638181, IndiaComp Clin PatholDOI 10.1007/s00580-011-1259-7et al. 2005). To date, however, data reported in the literaturerefer mainly to acute stress status in different farmed fishspecies and there have been an insufficient number ofstudies on the comparative haematological differencesbetween different feeding behaviour of fish that share thesame ecological zone. This present study would form abaseline data for assessment of health status of the fish aswell as reference point for future comparative study effectsof different feeding behaviour of fishes.Materials and methodsSamplingHealthy marine teleost fishes were collected from Vellarestuary (lat. 11°29 N and 79°46 E), Parangipettai of south-east coast of Tamilnadu, India. The estuary is seasonallybar-built and semi-diurnal type flows eastwards andempties into the Bay of Bengal at Parangipettai on south-east coast of India, carrying the wastes from the adjacentagriculture lands and industries in addition to domesticmunicipal and distillery effluents. Fish samples werecollected from 2007 to 2008. From the collection site, thewater quality dissolved oxygen was estimated by Winkler'smethods (Strickland and Parsons 1972), salinity by an Ermahand refractometer (Tokyo). The pH and water temperaturewere measured by using pH tester pen (Japan) andthermometer, respectively.Haematology and biochemical analysisA total of three species of marine teleost fishes namely Latescalcarifer (carnivores), Mugil cephalus (omnivores) andChanos chanos (herbivores) were collected and acclimatedin large fibreglass tanks. Fishes ranged from 235 to 410 g inweight and 25 to 30 cm in body length. Twenty-five fishesfrom each species were selected for this study. Blood wascollected by direct heart puncturing using sterile disposableplastic syringe with a 22-gauge needle (Molnar 1960).Heparine sodium (1%) was used as an anticoagulant(Svobodova et al. 2008). The collected blood samples wereimmediately subjected to haematological analysis. The bloodwas diluted with appropriated diluting fluids for RBC andWBC counts were determined using improved Neubauerhaemocytometer and calculated (Blaxhall and Daisley 1973).Replicated counts were made for each blood samples. Sahli'shaemoglobinometer was used to estimate haemoglobin (HB)percentage (HB%). Haematocrit (HCT) was determinedusing micro haematocrit capillaries filled with blood andcentrifuged at 8,700×g for 5 min and expressed aspercentage of total blood volume (Wintrobe 1974). Meancorpuscular volume (MCV), mean cell haemoglobin (MCH)and mean cell haemoglobin concentration (MCHC) werecalculated from the average values of HB% (Dacie andLewis 1984). For the determination of erythrocyte sedimen-tation rate (ESR), the blood was mixed well and 200 mmwas drawn into a Westurgen tube. The tubes were placedvertically and were left undisturbed for 60 min; after that, thelevel of the column of sediment was noted as ESR.Biochemical estimation of blood glucose, blood protein,blood cholesterol and blood urea were determined byfollowing standard methods (Folin and Wu 1920; Annino1976; Henry 1968; Fawcette and Scott 1960).Statistical studyDifferences in haematological parameters between thethree marine fish were statistically analysed by two-wayTable 1 Physico-chemical factors of the study siteMean±SD RangeTemperature (°C) 31.83±3.5 27.0–36.5Salinity (ppt) 26.16±12.0 3.14–35.8DO (mg/l) 4.48±0.53 3.93–5.41pH 7.80±0.43 7.12–8.31DO dissolved oxygenParameters L. calcarifer M. cephalus C. chanos P valueRBC (×106 μl) 2.96±0.25 2.52±0.21 2.6±0.51 <0.01*WBC (×103 μl) 21.6±1.45 28.6±1.02 26.33±1.56 <0.01*RBC/WBC (%) 0.107±0.001 0.082±0.01 0.085±0.00 <0.01*Haemoglobin (%) 53.23±2.35 37.6±1.42 52.33±1.72 <0.01*Haematocrit (%) 44.33±1.32 28.66±1.02 43.66±3.23 <0.01*ESR 0.5±0.03 0.43±0.01 0.56±0.02 <0.01*MCV (%) 13.38±2.34 11.37±1.0 16.81±3.1 <0.01*MCH (%) 156.55±5.45 149.4±3.78 201.26±11.4 <0.01*MCHC (%) 120.07±9.45 131.86±7.65 119.85±12.02 <0.01*Table 2 Haematological param-eters of marine teleost fishes(mean±SD)*P<0.01 (significance)RBC red blood cell count, WBCwhite blood cells count, ESRerythrocyte sedimentation rate,MCV mean cell volume, MCHmean cell haemoglobin, MCHCmean cell haemoglobinconcentrationComp Clin Patholanalysis of variance without replication. All these statisticalanalyses were performed using the SPSS statistical software(Version 7.5 for Windows XP, SPSS, and Chicago, IL, USA).Bray–Curtis similarity and multidimensional scale plot wereperformed using PAST statistical (Version 1.93 for WindowsXP).Results and discussionTable 1 shows the physico-chemical parameters of waterobtained from Vellar estuary. The water temperature (31.83±3.5) ranged from 27°C to 36.5°C, and salinity level(26.16±12.0) varied from 3.14 to 35.8 ppt. pH fluctuatedmonthly (7.80±0.43) and varied from 7.12 to 8.31.Dissolved oxygen concentration (4.48±0.53) ranged from3.93 to5.41 mg/l. Haematological and biochemical valuesof three marine teleost fishes are given in Tables 2 and 3.Definite variations between the species were detected in allhaematological and biochemical values. High level of RBCwas recorded in L. calcarifer, followed by M. cephalus andC. chanos. In addition RBC count in the fishes wassignificantly higher (P<0.01). The high erythrocyte numberwas associated with fast movement, predaceous nature andhigh activity, with streamlined body (Rambhaskar andSrinivasa Rao 1986). However, significant differences inthe values of erythrocyte volumes between the groups wereobserved in this study, suggesting that in the intensiveestuary environments, the elevated RBC counts and HBconcentration are a response to the higher metabolicdemand and have no impact on erythrocyte volume. Theincreased number of RBC indicates oxygen demand in thetropical region to meet the higher oxygen requirement athigher metabolic rates (Engel and Davis 1964). MaximumHB was recorded in L. calcarifer and C. chanos, least valuewas recorded in M. cephalus. HB value in the present studywas within the range 37.6–53.23%; low haemoglobin valuewas associated with the low active fishes. Similar resultswere already reported by Rambhaskar and Srinivasa Rao(1986). RBC and HB concentration tend to increase withlength and age of the fishes (Das 1965). HCT value showeda polynomial relationship, quadratic regression to thedifferent to the length of the fish [Ht=−8.33 (Ln(x)+57.003; R2=0.3931), P<0.01]. Blaxhall and Daisley (1973)have reported the possibility of using HCT as a tool in theaquaculture and fishery management for checking theanaemic condition. HCT value in the present study waswithin the range 28.66–44.33%; fish HCT values wereusually between 20% and 35% and rarely attain greater than50% (Clark et al. 1976). RBC/WBC ratio, MCV, MCHCand cholesterol was significantly correlated at P<0.05level. This may be due to the overall oxygen consumptionParameters L. calcarifer M. cephalus C. chanos P valueGlucose (mg/dl) 64.29±2.5 90.06±3.6 81.06±5.02 <0.01*Protein (mg/dl) 4.77±0.95 4.2±0.65 4.36±0.25 <0.01*Cholesterol (mg/dl) 291.69±10.2 208.04±2.01 253.16±8.25 <0.01*Urea (mg/dl) 4.56±1.3 6.53±3.21 5.56±0.65 <0.01*Table 3 Blood serum biochem-ical parameters of marine teleostfishes (mean±SD)*P<0.01 (significance)Fig. 1 Cluster analyses between the marine teleost fishes Fig. 2 Multidimensional scale plots between the marine teleost fishesComp Clin Patholrates and swimming performance under normal condition(Stillwell and Benfey 1995). The RBC/WBC level increaseddue to the decrease of WBC during this study. The highleucocyte volume was recorded in M. cephalus, C. chanosand L. calcarifer. There is an inverse relationship betweenWBC and RBC count observed in all the three fishes of thepresent investigation. High RBC count perhaps lessens therequirement for large number of WBC (Xiaoyun et al. 2009).An increased ESR level was recorded in L. calcarifer, C.chanos and lowest in M. cephalus. The ESR level may be aconsequence of alternation in blood plasma of different fishspecies or may be due to the stress condition (Joseph John2007).Blood biochemical parameters with significant variations(P<0.01) were observed for the concentrations of glucose,protein, cholesterol and urea. The ranges of serumbiochemistry vary from species to species and can beinfluenced by many biotic and abiotic factors such as watertemperature, seasonal pattern, food, age and sex of the fish(Jawad et al. 2004). An increased blood glucose and proteinlevel was recorded in L. calcarifer, this may probably bedue to an increased depletion of liver glycogen (Ojolick etal. 1995). Our findings confirm those of Coz-Rakovac et al.(2005) who reported a significantly elevated glucose levelin farmed Dicentrarchus labrax as compared to the wildecosystem. Strange (1980) also reported that blood glucoselevel may decrease with increasing water temperature inchannel catfish. Blood glucose level may vary according toseason and water temperature, and glucose concentration infish decreases with age and size (Coz-Rakovac et al. 2005).The increased plasma protein concentration can be causedby structural liver alternations that reduce aminotransferaseactivity, with concurrent reduction in deamination capacity(Kavadias et al. 2004). Hrubec et al. (2001) stated thatprotein level in striped bass increased with age. Cholesterolconcentration of L. calcarifer was higher due to highproportion of fat in the chemical composition of their food.Hill (1982) reported that cholesterol concentrations increaseas the fish size increased. Bano (1985) observed an increasein the serum cholesterol level after administration ofpesticides. A high blood urea concentration recorded inM. cephalus is likely to be a sign of stress associated withthe increase in the cortisol level (Borges et al. 2007).Bray–Curtis similarities were calculated (root trans-formed) between three species, and the resulting dendro-gram is shown in (Fig. 1). Two distinct fish communitygroupings could be distinguished that apparently reflecteddifferences in haematological/habitat types within the fishgroup. The carnivore fish L. calcarifer and herbivore fishC. chanos were one distinct community group; M. cephaluswas generating independently and makes one group. Fromthe resulting dendrogram, it was not possible to grade theresults according to haematological parameters, but it waspossible for species. Based on the multivariate analysis,carnivore and herbivore are active fishes, followed byomnivore as low active fish. From the multivariateordination (MDS) plot (Fig. 2), it was found that all theM. cephalus was ordinated separately from all other specieswhich confirms the dendrogram.Haematological and biochemical parameters have beenacknowledged as valuable tools for monitoring fish health,confirming maturation and monitoring any changes in thequality of water and related soil. The ranges of normalvalues of the key biochemical parameters are still undefinedfor different species in different aquaculture conditions. Theresults of this research provide the knowledge of thecharacteristics of haematological parameters of marineteleost fishes. We suggest that the haematological studieson fishes have assumed greater significance due to theincreasing emphasis on pisciculture and greater awarenessof the pollution in aquatic ecosystem.Acknowledgement The authors would like to thank Dr. G.Manimehalai, HOD, Department of Zoology, Kandaswamy KandarCollege, Paramathi-Velur, Tamil Nadu, India, for providing the labfacility.ReferencesAnnino JS (1976) Clinical chemistry principles and procedures, 4thedn. Little Brown and Company, BostonBahmani M, Kazemi R, Donskaya P (2001) A comparative study ofsome hematological features in young reared sturgeons (Aci-penser persicus and Huso huso). Fish Physiol Biochem 24:135–140Bano Y (1985) Sublethal stress of DDT on biochemical compositionof catfish Clarias batrachus. Indian J Environ Health 27:230–236Blaxhall PC, Daisley KW (1973) Routine haematological methods foruse with fish blood. 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Haematology and biochemical parameters of different

feeding behaviour of teleost fishes from Vellar estuary, India

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Haematology and biochemical parameters of different feeding behaviour of teleost fishes from Vellar estuary, India

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