Comparative Clinical Pathology

, Volume 23, Issue 2, pp 361–366

Haematological reference ranges of cultured Clarias gariepinus in the Lower Benue River Basin, Nigeria

Authors

    • Department of Veterinary PathologyFaculty of Veterinary Medicine, University of Abuja
  • Roslene Ada Obande
    • Department of Fisheries and AquacultureUniversity of Agriculture
  • Shola Gabriel Solomon
    • Department of Fisheries and AquacultureUniversity of Agriculture
Original Article

DOI: 10.1007/s00580-012-1624-1

Cite this article as:
Adeyemo, B.T., Obande, R.A. & Solomon, S.G. Comp Clin Pathol (2014) 23: 361. doi:10.1007/s00580-012-1624-1

Abstract

Blood samples were collected from 170 cultured African sharptooth catfish (Clarias gariepinus) to establish haematological baseline values of this important tropical pisciculture fish species in the Guinea savannah agro-ecological zone of Nigeria. The total red blood cell count and the total white blood cell count were obtained by haemocytometry, while the packed cell volume and haemoglobin were obtained by microhaematocrit and cyanomethmoglobin methods, respectively. The results obtained varied between gender and age and were as follows: total red blood cell count, 1.72 ± 0.34 × 106/mm3 in juveniles and 4.50 ± 0.57 × 106/mm3 in adults; total white blood cell count, 15.50 ± 1.15103/mm3in juvenile and 16.41 ± 1.21 × 103/mm3in adults; packed cell volume, 30.08 ± 7.78 % in juveniles and 39.59 ± 3.93 % in adults; haemoglobin, 9.43 ± 3.45 g/dl in juveniles and 10.99 ± 3.29 g/dl in the adults; mean corpuscular volume, 173.75 ± 41.93 fl in juveniles and 87.01 ± 14.37 fl in adults; mean corpuscular haemoglobin, 51.11 ± 13.10 pg in the juveniles and 26.81 ± 8.61 pg in the adults, while the mean corpuscular haemoglobin concentration values obtained are 32.61 ± 10.42 g/dl in the juveniles and 33.80 ± 10.0 g/dl in the adults.

Keywords

ErythrocytesLeucocytesClarias gariepinusHaematologyHaematocrit

Introduction

Clarias gariepinus, commonly called the African sharp tooth catfish, is amongst the most common freshwater aquaculture species in tropical and subtropical Africa and represents a major source of protein in Nigeria. C. gariepinus culture has been on a steady rise in Nigeria and its net production in 2006 was established to be 10–12, 8.3–15 and 14.2–14.3/ha/year under various culture systems (Federal Ministry of Agriculture and Rural Development of Nigeria 2003). C. gariepinus is a preferred aquaculture fish species because it is hardy and can tolerate high stocking densities.

C. gariepinus catfish lives in intimate contact with the environment and therefore is susceptible to physical and chemical changes which may be reflected in their haematological components. Blood is recognised as a potential index of fish health, as it can be used to ascertain the effect of pathogen challenge (Shimura et al. 1983; Ruane et al. 2000), quality of the diet and nutritional state of the fish (Tacon 1992; Chavez et al. 1994) and/or the effect of pollutants in the environment (VanVuren 1986; Adeyemo et al. 2003; Arnandova et al. 2009). Health evaluation and disease diagnosis in warm-blooded animals have been facilitated by the development of a large database of normal haematological values; these values have been critical to the growth of the various animal industries. Haematological analysis is a useful tool in determining the health status of both animals and humans; unfortunately, the usefulness of the values obtained from haematological testing is limited to the availability of species-specific normal reference ranges against which to compare values (Anver 2004; Gul et al. 2011).

Haematological parameters in fishes have been determined to depend on fish species, age, sex, the cycle of sexual maturity, condition of health and on geographical location (Gul et al. 2011; Gabriel et al. 2004; Noga 2010). Barton (2000) and Kori-Siakpere et al. (2005) observed that these parameters were also influenced by handling and transportation stress, and thus, it was averred that sustainable aquaculture requires that a detailed physiological knowledge of the species be acquired; this they suggested includes the determination of species-specific haematological values prior to their utilisation in disease diagnosis or as prognostic tools in health management. There is, however, a paucity of information on the haematological features of C. gariepinus farmed in the Lower Benue River Basin. It is therefore pertinent to establish the haematological reference ranges of cultured C. gariepinus sampled from this area with a view of monitoring their health and production performance.

The objective of this study, therefore, was to determine and document the haematological reference ranges of clinically normal cultured C. gariepinus in the Lower Benue River Basin. This information could serve as standard for the physiological, pathological and toxicological comparisons of C. gariepinus maintained under intensive production conditions.

Materials and methods

The Benue River extends from its origins in the western Cameroun Mountains and terminates in a confluence with the River Niger at Lokoja (Kogi State), North Central Nigeria (Fig. 1). It is a major catfish aquaculture production area in Nigeria; it encompasses areas drained by the Benue River and its various tributaries. The abundant unpolluted water from these rivers serves as water supply source for the several catfish-farming operations in the region.
https://static-content.springer.com/image/art%3A10.1007%2Fs00580-012-1624-1/MediaObjects/580_2012_1624_Fig1_HTML.gif
Fig. 1

Map of Nigeria showing Lower Benue River Basin (2010)

Fish sampling

One hundred and seventy C. gariepinus (270–1,000 g body weight and 24.72–74.81 cm total length) were obtained at random from three catfish farms selected at random from a list of catfish farms obtained from the area. Sampled fishes were transported (in water drums containing water obtained from the respective ponds) to the experimental ponds at the Department of Fisheries and Aquaculture University of Agriculture, Makurdi Benue State, Nigeria, where they were divided into three allotments as per source and supplied with water from the Benue River. The sampled fishes were screened for parasites as described by Post (1993) and for haemorrhagic skin lesions as described by Wagner and Congleton (2004) and (Tavarez-Dias and Moraes 2007) and acclimatised for 14 days prior to blood collection. Mortality during this depuration period was less than 2 %.

Blood collection

Collection of blood samples was done after anaesthetizing fish in a 5-l water bath containing 0.2 mg benzocaine dissolved in 5 ml acetone as described by Carrasco et al. (1984). Blood was obtained by posterior caudal venipuncture using a 22-G hypodermic needle and 2-ml syringes as described by Schmitt et al. (1999). Two millilitres of aspirated blood was carefully emptied into plastic tubes containing the sodium salt of ethylenediaminetetraacetic acid as an anticoagulant. Whole blood (50 μl) was stained for enumeration of red blood cells as described by Adedeji and Adegbile (2011). Blood smears were then air dried for 5 min, fixed in absolute methanol and stained for 60 s in Giemsa stain.

Haematological analysis

Total red blood cell count (RBC), total white blood cell counts (WBCs) and platelet enumeration were done using the new Neubauer haemocytometer. The packed cell volume (PCV) and haemoglobin concentration values were determined by the microhaematocrit capillary tube and cyanomethaemoglobin methods, respectively, as described by Schalm et al. (1975). The secondary haematological parameters like the mean corpuscular volume (MCV), the mean corpuscular haemoglobin (MCH) and the mean corpuscular haemoglobin concentration (MCHC) were determined from the primary data as described by Schalm et al. (1975). Blood smears made and stained with Giemsa were used to determine total WBC, platelet counts and differential WBC counts. WBC counts and platelets were counted until 200 WBCs were enumerated on blood smears, and the percentage of each WBC type and of platelets was multiplied by the total WBC and platelet counts to obtain the absolute differential cell counts as described by Stoskopf (1993), as fish RBCs are nucleated and nucleated RBCs prevent accurate enumeration using automated analysis (Huffman et al. 1997).

Statistical analysis

Results are presented as mean with standard deviation of the mean. Reference ranges or intervals were determined using the guidelines of the National Committee for Clinical Laboratory Standards (1992) and by ranking the data using the 25th and 75th percentile nonparametric analysis (Duncan et al. 1994; Linnet 2000). Gender and weight influences on haematological parameters were examined using Student's t test. Data were examined for outliers and any outlying results were excluded from the data set.

Results

The total RBC count ranges from 1.11 × 106 to 2.22 × 1106/mm3 and there were more RBCs in peripheral circulation compared to total white blood cell count which ranged from 13.19 × 103 to 17.62 × 103/mm3 of blood; also, the PCV of these fishes ranged from 20.15 to 32.50 % (Table 1). Amongst the juvenile female C. gariepinus, the total red blood cell counts ranged from 1.62 × 106 to 1.82 × 106/mm3 and were more numerous than those for total white blood cell counts. The PCV ranged from 18.70 to 40.10 %, while the total haemoglobin ranged from 6.80 to 13.50 g/dl. These results are depicted in Table 2.
Table 1

Haematological parameters of juvenile male Clarias gariepinus

Analyte

Mean ± SD

Reference interval

Sample size

L

H

N

RBC (×106/mm3)

1.72 ± 0.34

1.11

2.22

42

WBC (×103/mm3)

15.44 ± 1.67

13.19

17.62

42

PCV (%)

28.80 ± 2.93

20.15

32.50

42

Hb (g/dl)

9.43 ± 3.248

6.2

15.70

42

MCV (fl)

173.75 ± 41.93

110.71

240.54

42

MCH (pg)

51.11 ± 13.10

31.47

70.58

42

MCHC (g/dl)

32.61 ± 10.42

21.23

55.45

42

Weight (g)

97.67 ± 21.72

50.05

127.27

42

Hb haemoglobin, L lowest value obtained, H highest value obtained

Table 2

Haematological parameters of juvenile female Clarias gariepinus

Analyte

Mean ± SD

Reference interval

Sample size

L

H

N

RBC (×106/mm3)

1.72 ± 0.05

1.62

1.82

42

WBC (×103/mm3)

15.50 ± 1.15

13.83

17.11

42

PCV (%)

30.08 ± 7.78

18.70

40.10

42

Hb (g/dl)

9.12 ± 2.71

6.80

13.50

42

MCV (fl)

158.26 ± 42.40

111.59

225.44

42

MCH (pg)

44.37 ± 6.65

37.36

55.03

42

MCHC (g/dl)

30.69 ± 5.55

22.04

38.20

42

Weight (g)

73.24 ± 23.03

50.91

123.41

42

Hb haemoglobin, L lowest value obtained, H highest value obtained

Results obtained for adult male and female C. gariepinus also show that there are more erythrocytes in peripheral blood than white blood cells with values ranging from 3.11 × 106 to 5.72 × 106/mm3 for total red blood cell counts and 15. 13 × 103 to 20.09 × 103/mm3 for white blood cell counts (Tables 3 and 4). The haematological evaluation of these groups reveals that haemoglobin concentration ranged from 6.30 to 50 g/dl in the males (Table 3) and 6.90 to 126.50 g/dl in the adult females (Table 4).
Table 3

Haematological parameters of adult male Clarias gariepinus

Analyte

Mean ± SD

Reference interval

Sample size

L

H

N

RBC (×106/mm3)

3.73 ± 0.62

3.11

5.10

44

WBC (×103/mm3)

16.30 ± 1.01

15.13

17.19

44

PCV (%)

28.45 ± 3.85

15.30

31.05

44

Hb (g/dl)

9.4 ± 2.37

6.30

11.50

44

MCV (fl)

79.50 ± 16.69

36.96

95.88

44

MCH (pg)

26.81 ± 8.61

13.15

42.12

44

MCHC (g/dl)

33.80 ± 10.10

21.72

54.24

44

Weight (g)

494.41 ± 233.50

232.4

1,009.1

44

Hb haemoglobin, L lowest value obtained, H highest value obtained

Table 4

Haematological parameters of adult female Clarias gariepinus

Analyte

Mean ± SD

Reference interval

Sample size

L

H

N

RBC (×106/mm3)

4.5 ± 0.57

3.49

5.72

42

WBC (×103/mm3)

16.41 ± 1.21

15.38

20.09

42

PCV (%)

39.59 ± 3.93

29.9

45.10

42

Hb (g/dl)

10.99 ± 3.29

6.9

16.5

42

MCV (fl)

87.01 ± 14.37

58.51

119.31

42

MCH (pg)

23.08 ± 7.27

15.16

35.62

42

MCHC (g/dl)

28.23 ± 9.16

16.42

41.30

42

Weight (g)

818.08 ± 256.91

561.16

1,068.6

42

Hb haemoglobin, L lowest value obtained, H highest value obtained

Statistical evaluations of the results obtained from the haematological analysis of blood samples reveal that was no difference in the means of PCV, RBC, MCV, MCH and MCHC (Table 5). In the adult stages (males and females), statistical significant difference (p < 0.05) was observed in a sex- and/or developmental (weight)-based comparison in the PCV, RBC, MCV, MCH and MCHC, respectively, as depicted in Table 5. The results for the water quality parameters are as follows: temperature (in degrees Celsius) 27.5 ± 1.5, dissolved oxygen (in milligrams per litre) 6.4 ± 0.2, alkalinity (in milligrams per litre as CaCO3) 456 ± 3.5 and pH 7.5 ± 0.2.
Table 5

Statistical comparison (t test) of haematological parameters of Clarias gariepinus sampled from Lower Benue River Basin

Basis of comparison

PCV (%)

RBC (×106/mm3)

WBC (×103/mm3)

Hb (g/dl)

MCV (fl)

MCH (pg)

MCHC (g/dl)

Juv male

28.80 ± 2.94a

1.73 ± 0.34a

15.45 ± 1.67a

9.44 ± 3.25a

173.75 ± 41.93a

51.11 ± 13.11a

32.62 ± 10.42a

Juv female

30.09 ± 7.78a

1.72 ± 0.06a

15.51 ± 1.16a

9.13 ± 2.71a

158.26 ± 42.41a

44.37 ± 6.66a

30.69 ± 5.56a

Adt male

28.45 ± 3.85a

3.78 ± 0.62a

16.30 ± 1.01a

9.44 ± 2.37a

79.50 ± 16.69a

26.81 ± 8.61a

33.81 ± 10.11a

Adt femal

39.59 ± 3.39b

4.56 ± 0.57b

16.41 ± 1.21a

10.99 ± 3.3a

87.02 ± 14.37b

23.09 ± 7.28b

28.23 ± 9.17b

Juv male

28.08 ± 2.94a

1.73 ± 0.34a

15.45 ± 1.67a

9.44 ± 3.25a

173.75 ± 41.93a

51.11 ± 13.11a

32.62 ± 10.42a

Adt male

28.45 ± 3.85a

3.78 ± 0.62b

16.30 ± 1.01a

9.40 ± 2.37a

79.50 ± 16.69b

26.81 ± 8.61b

33.81 ± 10.11a

Juv female

30.09 ± 7.78a

1.72 ± 0.06a

15.51 ± 1.16a

9.13 ± 2.71a

158.26 ± 42.41a

44.37 ± 6.66a

30.69 ± 5.56a

Adt female

39.59 ± 3.93b

4.56 ± 0.57b

16.41 ± 1.21a

10.99 ± 3.30a

87.02 ± 14.37b

23.09 ± 7.28b

28.23 ± 9.17b

Statistical significance is set at p < 0.05. Rows with the same superscripts are not statistically significant. Rows with different superscripts are statistically significant

Juv male juvenile males, Juv female juvenile females, Adt male adult males, Adt females adult females

Discussion

The count of red blood cells is quite a stable index and the fish body tries to maintain this count within the limits of certain physiological standards using various physiological mechanisms of compensation. The range of haematological parameters measured in this study was quite broad. This is not surprising because the fish were not of the same age and were sampled from different environments though the water parameter conditions were favourable for catfish culture (Boyd and Tucker 1992). In practical terms, these values support the findings of Ellis (1977) and Angelica et al. (2010) that baseline parameters be established for identified fish populations to demonstrate specific changes in the blood picture resulting from disease, stress or toxic challenges.

There was no significant difference (p ≥ 0.05) in the PCV values for juvenile males and juvenile females. However, a comparison of the PCV values of the adult males and adult females of C. gariepinus revealed an extreme statistically significant difference (p < 0.0001) in the mean PCV. Also, an intra-sex comparison between juveniles and adult (juvenile male–adult male and juvenile female–adult female pairings) reveals that there is no statistical difference in the mean PCV of C. gariepinus (Table 5).

A weight- and gender-based comparison shows that there is no statistical difference (p > 0.05) between the mean RBC count of juvenile males and juvenile females of fishes sampled. However, a sex-dependent weight comparison reveals a strong statistical difference (p < 0.05) in the mean RBC of fishes sampled (p < 0.0001) amongst adults. Again, a weight (age-wise), intra-sex-dependent comparison of juveniles and adults (e.g. juvenile females against adult females or juvenile males against adult males) reveals a strong statistical significant difference (p < 0.05) in the RBC of the fishes sampled. This finding concurs with Tavares-Dias and Moraes (2007) and Arnandova et al. (2009) who had observed that adult fishes especially in their breeding seasons exhibited elevated levels of erythropoieisis and thus display elevated physiological RBC counts. It is worthy of note, however, to state that erythrocyte values obtained were similar to those reported by Sabiri et al. (2009), in adult feral C. gariepinus sampled from the Nile River, and Ezeri (2001) who reported a mean RBC value of 3.272 × 106/mm3 in adult feral C. gariepinus sampled from Southeastern Nigeria.

A comparison along weight and sex lines revealed that there is no statistical significance (p > 0.05) in the mean WBC of the fishes sampled. Haemoglobin concentration reflects the supply of oxygen in an organism and the fish tries to maintain them in as much as stable condition as possible. There was a wide variability with the haemoglobin concentrations obtained in the study and these concur with Angelica et al. (2010) and Rainza-Paiva et al. (2000) and are similar for the results obtained by Adeyemo et al. (2003), Gabriel et al. (2004) and Ezeri (2001). Comparisons along weight and sex lines reveal that there are no statistical significant differences (p > 0.05) in the mean haemoglobin amongst juveniles. A comparison of the haemoglobin concentration in adult males and adult females, however, reveals a significant difference in mean haemoglobin (p < 0.05). The reason for this could not be elucidated, but one plausible explanation may be in the influence of sexual maturity on the haemoglobin concentrations of the concerned females.

The other haematological parameters like the MCV, the MCH and the MCHC were mathematically derived from the primary haematological analysis, i.e. red blood cell count, packed cell volume and haemoglobin, thus these values showed some resemblance with the results obtained in a t test evaluation of the primary haematological parameters. Hence, amongst the fishes sampled, there was a significant difference (p < 0.05) in the MCV, MCH and MCHC only amongst the adults and in MCV and MCH in juvenile–adult comparisons (Table 5).

The result of this research provides clinical pathology data for cultured C. gariepinus sample from the Lower Benue River Basin of Nigeria, thus providing fish health specialists with a reference on which to base observations obtained from commercial catfish ponds. As further understanding develops with regards to the interactions of blood parameters and fish health, this information may prove useful as a part of an integrated management system with the goal of understanding changes that predict the onset of disease, allowing appropriate interventions and preventions of fish losses.

Copyright information

© Springer-Verlag London 2012