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Comparison of haematological values in restricted‐ and ad libitum‐fed domestic fowls: Red blood cell characteristics a
a
a
M. H. Maxwell , G. W. Robertson , S. Spence & C. C. McCorquodale
a
a
Agricultural and Food Research Council, Institute for Grassland and Animal Production, Poultry Department, Roslin, Midlothian, EH25 9PS, Scotland Version of record first published: 08 Nov 2007.
To cite this article: M. H. Maxwell , G. W. Robertson , S. Spence & C. C. McCorquodale (1990): Comparison of haematological values in restricted‐ and ad libitum‐fed domestic fowls: Red blood cell characteristics, British Poultry Science, 31:2, 407-413 To link to this article: http://dx.doi.org/10.1080/00071669008417271
PLEASE SCROLL DOWN FOR ARTICLE Full terms and conditions of use: http://www.tandfonline.com/page/terms-andconditions This article may be used for research, teaching, and private study purposes. Any substantial or systematic reproduction, redistribution, reselling, loan, sub-licensing, systematic supply, or distribution in any form to anyone is expressly forbidden. The publisher does not give any warranty express or implied or make any representation that the contents will be complete or accurate or up to date. The accuracy of any instructions, formulae, and drug doses should be independently verified with primary sources. The publisher shall not be liable for any loss, actions, claims, proceedings, demand, or costs or damages whatsoever or howsoever caused arising directly or indirectly in connection with or arising out of the use of this material.
British Poultry Science (1990) 31: 407-413
COMPARISON OF HAEMATOLOGICAL VALUES IN RESTRICTEDAND AD LIBITUM-FED DOMESTIC FOWLS: RED BLOOD CELL CHARACTERISTICS M. H. MAXWELL, G. W. ROBERTSON, S. SPENCE AND C. C. McCORQUODALE
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Agricultural and Food Research Council Institute for Grassland and Animal Production, Poultry Department, Roslin, Midlothian EH25 9PS, Scotland Received for publication 2nd May 1989
Abstract 1. A haematological study of the red blood cell characteristics in domestic fowls fed restricted and ad libitum diets was examined. The characteristics included Hb (haemoglobin), PCV (Packed Cell Volume), Rbc (Red blood cell count), MCH (Mean Cell Haemoglobin), MCV (Mean Cell Volume), MCHC (Mean Cell Haemoglobin Concentration) and reticulocytes. 2. The investigation showed that Hb, PCV, MCH and MCV were all reduced when broilers were fed a restricted diet. The number of Rbc's in these birds also increased significantly compared with control ad libitum-fed broilers. MCHC and reticulocyte numbers did not alter significantly. 3. It is proposed that some of the red cell characteristics may be restored to within "normal" ranges by food restriction. However, the results did show that there was a tendency to induce a microcytic normochromic anaemia in these restricted-fed birds.
INTRODUCTION
Haematological reference values of normal domestic fowls have been reported by several workers (Lucas and Jamroz, 1961; Bell and Freeman, 1971; Farner and King, 1972; Archer and Jeffcott, 1977; Kai and Franklin, 1984; Sturkie, 1986). Food restriction in poultry has been studied comprehensively (Lee et al., 1971; Blair, 1972; Blair et al., 1976; Pearson and Shannon, 1979; Whitehead et al., 1987) and is now an accepted procedure in commercial practice for improving biological and economic performance. However, there appears to be no detailed information in the literature regarding the effect food restriction in fowls may have on the haematology. In the present study we examined 7 haematological reference values of the red cell characteristics in broilers of each sex fed a restricted diet from 4 to 20 407
408
M. H. MAXWELL et al.
weeks of age, measured at weekly intervals. Blood profiles of broiler and layer strains fed ad libitum were examined for comparison.
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MATERIALS AND METHODS
Two commercial strains of domestic fowl (Gallus gallus domesticus) bought at 1-d-old and totalling 610 birds were used. A medium-weight egg-laying strain (ISA Brown) and a commercial broiler breeder strain were used; both were obtained from local hatcheries. The birds were divided into 6 groups (Table 1) and maintained in pens 2 X 3 m 3 on deep litter under a conventional lighting regimen of 14 h light and 10 h darkness. Both strains were vaccinated at 1-d and 3-weeks-old against Marek's disease and at 4-weeks-old against Newcastle disease. Mortality was less than 0-05% in all groups. The birds were maintained on a standard starter and grower diet. Food was available ad libitum for groups A, B, C and D (Table 1). Birds of groups E and F had their intake restricted from 4 weeks of age according to a commercial regimen (D. B. Marshall Ltd, Newbridge, Scotland) (Table 1). Water was available ad libitum for all groups. The weekly food-restriction programme is presented in Table 2. TABLE 1
Details of birds and diets used
Strain Layer (Isa Brown)
Group A
n
Sex
no
M
Diet 0-4 weeks—starter (230 g protein, 12-55 MJ
B
110
F
5-20 weeks—grower (190 g protein, 12-55 MJ
C
110
M
0-4 weeks—starter (230 g protein, 12-55 MJ
D
110
F
ME/kg) ad libitum ME/kg) ad libitum
Broiler (breeding strain)
ME/kg) ad libitum
5-20 weeks—grower (190 g protein, 12-55 MJ ME/kg) ad libitum
Broiler (breeding strain)
E
85
M
0-4 weeks—starter (230 g protein, 12-55 MJ
F
85
F
4-20 weeks—restricted grower diet (190 e protein, 12-55 MJ ME/kg) adjusted as for commercial regimen for breeding birds
ME/kg) ad libitum
Birds from groups A to D were bled at 1-d-old, 3 d and one week. Thereafter, they were bled at weekly intervals between 09.00 and 10.00 h until 20 weeks of age. The birds on the restricted diet were bled weekly in a similar manner but from 4 weeks of age. Each bird was bled only once. Five birds from each group were selected at random for bleeding each week and the mean result was presented. The chicks were killed by decapitation until 3 weeks of age and thereafter birds were bled from the basilic vein using sterile disposable plastic syringes and needles (21 gauge). The blood (1-0 to 1-5 ml) was dispelled immediately into collecting tubes containing sequestrene (dipotassium salt of ethylene diamine tetra-acetic acid (EDTA, 1-5 mg/ml of blood) anticoagulant.
ERYTHROCYTE CHANGES AND FOOD RESTRICTION
409
TABLE 2
Weekly food-restriction programme (g/bird d) Age (weeks) Female Male
0-3 ad lib. ad lib.
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Age (weeks) Female Male
4 41 50
5 41 50 13 63 73
6 45 50 14 68
77
7
8
50 54
50 54
9 54 63
10 54 63
11 59 68
12 59 68
18
19 86 90
20 91 90
15
16
73 77
77
17 77
82
81
81
86
Haemoglobin (Hb) was determined photometrically as cyanmethaemoglobin:blood was diluted 1:50,000 with Isoton II (Coulter Electronics Ltd, Northwell Drive, Luton, Bedfordshire). A few drops of Zapoglobin (Coulter Electronics Ltd) were added to lyse the red blood cells (Rbc's). The diluted blood sample was left at room temperature for 20 min before centrifuging at 300g for 2 to 3 min to remove red cell nuclei. Hb values were read directly using a LK 540 photometer (TEM sales Ltd, Gatwick Road, Crawley, West Sussex). The blood sample was also diluted 1:50,000 with Isoton II and the total number of cells was recorded using an electronic particle counter (Fn model: Coulter Electronics Ltd). Two separate dilutions were made with each sample. Each sample was read 5 times and the mean of each diluted sample was corrected with the coincidence correction chart (Coulter Electronics Ltd). The following variables were judged to give good 'plateaux' for fowl blood cells: orifice diameter 100 fim, attenuation 0-707, aperture 16, threshold 12. The counter was calibrated with cell control 4C (Coulter Electronics Ltd) and blood from human volunteers. The Rbc count was determined by counting 12 microscope fields in a blood film with a X100 objective and once the white blood cell/thrombocyte ratio has been established (Maxwell et al., 1990) the total number of Rbc's could be computed. The packed cell volume (PCV) was measured by the microhaematocrit method (Hawksley and Sons Ltd, Lancing, Sussex). The following indices were calculated (Dacie and Lewis, 1975); mean cell haemoglobin (MCH), mean cell haemoglobin concentration (MCHC), mean cell volume (MCV). Reticulocyte counts were performed according to Dacie and Lewis (1975). Whole blood (50 fil) was mixed with equal parts 1% brilliant cresyl blue solution, allowed to stand at room temperature for 1 h and the blood films were air-dried. Fifty microscope fields were counted using a cut-down eyepiece. The reticulocytes were assigned to 5 stages according to the degree of reticulation (Coates and March, 1966). Briefly, this reticulation ranged from a heavily-stained network in the cytoplasm of cell 1 to a few isolated dots in cell 5. All counts were carried out by the same investigators. The variance between observations within the individual groups of birds tended to increase with the mean. Taking square roots of the red blood cell and reticulocyte counts and logarithms of the other variates removed much of this heterogeneity of variance. Analyses of variance were therefore performed in the transformed scale to assess the statistical significance of the effects of sex, strain, food restriction and age.
410
M. H. MAXWELL et al.
RESULTS
A summary of the main effects of food restriction, strain, sex and age on the red blood cell characteristics is presented in Table 3, as means of counts between 4 and 20 weeks of age, their approximate standard errors and t statistic associated with the main effects in the transformed scale. Standard errors were calculated using the approximation for the standard error of a function of a random variable (Kendall and Stuart, 1963). The age range 4 to 20 weeks was divided into two periods 4 to 12 weeks and 13 to 20 weeks.
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Haemoglobin
The mean Hb value for the restricted-fed broilers (10-0) (g/100 ml) was significantly lower than for birds fed ad libitum (10-6). Similar degrees of significance in Hb values were seen in favour of broilers (10-6) compared with layers (9-6), males (10-3) compared with females (9-8) and older (10-7) compared with younger birds (9-5) (Table 3). Packed cell volume
The mean PCV value of the restricted-fed broilers (31-2%) was also significantly reduced compared with the birds fed ad libitum (32-9). Similar levels of significance were recorded between broilers (32-9) and layers (29-8), males (32-2) and females (30-4) and older (32-7) compared with younger birds (30-0) (Table 3). Red blood cells
Table 3 shows that there were significantly more Rbc's in the restricted-fed broilers (2-85 XlO6 mm3) than ad libitum-kd birds (2-81). Broilers also had more Rbc's (2-81) in circulation than layers (2-56). Furthermore, the Rbc count in males was 2*80 and 2-70 in females. Older birds had significantly more Rbc's (2-86) than younger birds (2-65). Mean cell haemoglobin
There was a significant reduction in MCH values in the broilers that received the restricted diet (35-1 pg) compared with those birds fed ad libitum (37-9) (Table 3). Significance of similar magnitude was also recorded in older birds (37-8) compared with younger ones (36-2). There were no significant main effects of strain or sex. Mean cell volume
Table 3 shows that the mean MCV value for the restricted-fed broilers (109-0 ftm3) was significantly lower than for ad libitum-fed birds (117-5). Males had significantly larger red cells (115-4) than females (113-7). There was a tendency for them to be larger in older (115-4) than in younger birds (113-8).
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ERYTH]
TABLE 3
Summary of red blood cell characteristics and their effects in birds fed adlibitum and restricted from 4 to 20 weeks—means (±SEM) 6
3
A R I
10 6 (007) 100(006) 5-79***
32-9 (0-2) 31-2(0-2) 6-48***
2-81 (002) 2-88(0-02) 2-78***
Rbc (10 /mm ) MCH (pg) 37-9 (0-3) 351 (0-2) 911***
MCV (/im3) 117-5 (0-6) 1090 (0-5) 10-82***
MCHC (%) 32-2 (01) 32-2 (01) 0-81
Reticulocytes (%) 101 (0-2) 9-5 (0-2) 113
Strain ad libitum
L B t
9-6 (006) 10-6 (007) 10 32***
29-8 (0-2) 32-9 (0-2) 12-36***
2-56 (002) 2-81 (019) g.42***
37-8 (0-2) 37-9 (0-2) 0-42
117-1 (0-6) 117-5 (0-6) 0-61
32-3 (01) 32-2 (01) 0-20
11-8 (0-2) 101 (0-2) 5 62***
Sex
M F t
10-3 (005) 9-8 (005) 5-97***
32-2 (01) 30-4 (01) 8-46***
2-80 (002) 2-70 (001) 4-63***
37-1 (0-2) 36-8 (0-2) 0-99
115-4(0-5) 113-7 (0-5) 2.70***
3 2 1 (01) 32-4 (01) 1-68
10-9 (0-2) 10-0 (01) 3-83***
Age
4-12 weeks 13-20 weeks t
9-5 (005) 10-7 (006) 15-85***
3 0 0 (01) 32-7 (01) 13-68***
2-65 (001) 2-86 (002) 9-76***
. 36-2 (0-2) 37-8 (0-2) 607***
113-8 (0-4) 115-4 (0-5) 1-73
31-7 (01) 32-8(0-1) 7.46***
10-8 (0-1) 101 (0-2) 300***
Effect Restriction Broilers
HB (g/100 ml) PCV (%)
A Ad libitum, R Restricted, L Layer, B Broiler, M Male, F Female. Student's t statistic with 488 DF in transformed scale. ***p 0 w
z0
s 0 0
1 0
z
412
M. H. MAXWELL et al.
However, there was no significant difference in MCV values between the two different strains. Mean cell haemoglobin concentration
Table 3 shows that the older group of birds had a significantly larger MCHC value (32-8%) than the younger group (31-7). There were no further significant differences in MCHC values with food restriction, strain or sex.
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Reticulocytes
There were significantly more reticulocytes in the layer strain (11*8%). than in the broilers (10-1) (Table 3). Males had more reticulocytes (10-9) than females and (10-0) and younger birds (10-8) more than older birds (10-1). There was no significant main effect of food restriction. DISCUSSION
When broilers were maintained on a food-restricted diet for up to 20 weeks of age, there was a significant reduction of Hb and PCV but a significant increase in the number of circulating red cells. Furthermore, the indices MCH and MCV were lower than in control broilers that received an ad libitum diet. These results may indicate that more red cells were released from the bone marrow to compensate for the lowered haemoglobin and that these cells, because of the reduced MCV, were also smaller. These food-restricted broilers may thus be suffering from a mild microcytic normochromic anaemia. Evidence from published work (Lee et al., 1971; Blair, 1972; Pearson and Shannon, 1979) has suggested that the effects of food restriction during the rearing period of broiler chickens can be beneficial in terms of their overall biological performance. Because food restriction delays body weight, fat deposition, sexual maturity and even mortality (Pearson and Shannon, 1979), it follows that the progress of the haematological profile will also be reduced in momentum. The changes in red blood cell values probably have a further beneficial advantage on such organs as the heart and lungs, in reducing their overall combined workload (Julian, 1987) and assisting in the control of cardiopulmonary disease in broilers. There is a danger, however, that too severe a restriction programme may cause an even greater anaemia or possibly induce further stress (Maxwell et al., 1990). Therefore, it is important that a correct dietary balance of food-restriction be achieved. This in turn should control haematological values to within a "normal" range, otherwise birds may suffer. EDITORS
NOTE
Tables of raw data, summarised in Table 3, may be obtained from the authors on request.
ERYTHROCYTE CHANGES AND FOOD RESTRICTION
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REFERENCES ARCHER, R.K. & JEFFCOTT, L.B. (1977) Comparative Clinical Haematology, p. 516 (Oxford, Blackwell Scientific Publications). BELL, D.J. & FREEMAN, B.M. (1971) Physiology and Biochemistry of the Domestic Fowl, Vol. 2, pp. 841-850 (London, Academic Press). BLAIR, R. (1972) Feed restriction in breeding birds. Feedstuffs (Minneapolis), 44(10): 36-39. BLAIR, R., MACCOWAN, M.M. & BOLTON, W. (1976) Effects of food regulation during the growing and laying stages on the productivity of broiler breeders. British Poultry Science, 17: 215-223. COATES, V. & MARCH, B.E. (1966) Reticulocyte counts in the chicken. Poultry Science, 45: 1302-1305. DACIE, J.V. & LEWIS, S.M. (1975) Practical Haematology, p. 81 (Edinburgh, Churchill Livingston). FARNER, D.S. & KING, J.R. (1972) Avian Biology, Vol. II, pp. 159-163 (New York, Academic Press). JULIAN, R.J. (1987) The effect of increased sodium in the drinking water on right ventricular hypertrophy, right ventricular failure and ascites in broiler chickens. Avian Pathology, 16: 61-71. KAI, O. & FRANKLIN, R.M. (1984) The age-dependent size distribution of chicken peripheral blood cells analysed by a novel method utilising a Coulter counter. Developmental and Comparative Immunology, 8: 375-386. KENDALL, M.G. & STUART, A. (1963) The Advanced Theory of Statistics, 2nd edn, Vol. 1, pp. 231-242 (London, Charles Griffin). LEE, P.J.W., GULLIVER, A.L. & MORRIS, T.R. (1971) A quantitative analysis of the literature concerning the restricted feeding of growing pullets. British Poultry Science, 12: 413-437. LUCAS, A.M. & JAMROZ, C. (1961) Atlas of Avian Hematology. United States Department of Agriculture, Washington, pp. 216-217. MAXWELL, M.H., ROBERTSON, G.W., SPENCE, S. & MCCORQUODALE, C.C. (1989) Comparison of
haematological values in restricted- and ad libitum-fed domestic fowls: white blood cells and thrombocytes. British Poultry Science, 31: 399-405. PEARSON, A.R. & SHANNON, D.W.F. (1979) Controlled feeding systems, in: K. N. BOORMAN & B. M. FREEMAN (Eds) Food Intake Regulation in Poultry, pp. 365-390 (Edinburgh, British Poultry Science Ltd). STURKIE, P.D. (1986) Avian Physiology, 4th edn, pp. 102-129 (New York, Cornell University Press). WHITEHEAD, C.C., HERRON, K.M. & WADDINGTON, D. (1987) Reproductive performance of dwarf
broiler breeders given different allowances of food during the rearing and breeding periods and two lighting patterns. British Poultry Science, 28: 415-427.
British Poultry Science Publication details, including instructions for authors and subscription information: http://www.tandfonline.com/loi/cbps20
Comparison of haematological values in restricted‐ and ad libitum‐fed domestic fowls: Red blood cell characteristics a
a
a
M. H. Maxwell , G. W. Robertson , S. Spence & C. C. McCorquodale
a
a
Agricultural and Food Research Council, Institute for Grassland and Animal Production, Poultry Department, Roslin, Midlothian, EH25 9PS, Scotland Version of record first published: 08 Nov 2007.
To cite this article: M. H. Maxwell , G. W. Robertson , S. Spence & C. C. McCorquodale (1990): Comparison of haematological values in restricted‐ and ad libitum‐fed domestic fowls: Red blood cell characteristics, British Poultry Science, 31:2, 407-413 To link to this article: http://dx.doi.org/10.1080/00071669008417271
PLEASE SCROLL DOWN FOR ARTICLE Full terms and conditions of use: http://www.tandfonline.com/page/terms-andconditions This article may be used for research, teaching, and private study purposes. Any substantial or systematic reproduction, redistribution, reselling, loan, sub-licensing, systematic supply, or distribution in any form to anyone is expressly forbidden. The publisher does not give any warranty express or implied or make any representation that the contents will be complete or accurate or up to date. The accuracy of any instructions, formulae, and drug doses should be independently verified with primary sources. The publisher shall not be liable for any loss, actions, claims, proceedings, demand, or costs or damages whatsoever or howsoever caused arising directly or indirectly in connection with or arising out of the use of this material.
British Poultry Science (1990) 31: 407-413
COMPARISON OF HAEMATOLOGICAL VALUES IN RESTRICTEDAND AD LIBITUM-FED DOMESTIC FOWLS: RED BLOOD CELL CHARACTERISTICS M. H. MAXWELL, G. W. ROBERTSON, S. SPENCE AND C. C. McCORQUODALE
Downloaded by [University of Guelph] at 05:30 24 February 2013
Agricultural and Food Research Council Institute for Grassland and Animal Production, Poultry Department, Roslin, Midlothian EH25 9PS, Scotland Received for publication 2nd May 1989
Abstract 1. A haematological study of the red blood cell characteristics in domestic fowls fed restricted and ad libitum diets was examined. The characteristics included Hb (haemoglobin), PCV (Packed Cell Volume), Rbc (Red blood cell count), MCH (Mean Cell Haemoglobin), MCV (Mean Cell Volume), MCHC (Mean Cell Haemoglobin Concentration) and reticulocytes. 2. The investigation showed that Hb, PCV, MCH and MCV were all reduced when broilers were fed a restricted diet. The number of Rbc's in these birds also increased significantly compared with control ad libitum-fed broilers. MCHC and reticulocyte numbers did not alter significantly. 3. It is proposed that some of the red cell characteristics may be restored to within "normal" ranges by food restriction. However, the results did show that there was a tendency to induce a microcytic normochromic anaemia in these restricted-fed birds.
INTRODUCTION
Haematological reference values of normal domestic fowls have been reported by several workers (Lucas and Jamroz, 1961; Bell and Freeman, 1971; Farner and King, 1972; Archer and Jeffcott, 1977; Kai and Franklin, 1984; Sturkie, 1986). Food restriction in poultry has been studied comprehensively (Lee et al., 1971; Blair, 1972; Blair et al., 1976; Pearson and Shannon, 1979; Whitehead et al., 1987) and is now an accepted procedure in commercial practice for improving biological and economic performance. However, there appears to be no detailed information in the literature regarding the effect food restriction in fowls may have on the haematology. In the present study we examined 7 haematological reference values of the red cell characteristics in broilers of each sex fed a restricted diet from 4 to 20 407
408
M. H. MAXWELL et al.
weeks of age, measured at weekly intervals. Blood profiles of broiler and layer strains fed ad libitum were examined for comparison.
Downloaded by [University of Guelph] at 05:30 24 February 2013
MATERIALS AND METHODS
Two commercial strains of domestic fowl (Gallus gallus domesticus) bought at 1-d-old and totalling 610 birds were used. A medium-weight egg-laying strain (ISA Brown) and a commercial broiler breeder strain were used; both were obtained from local hatcheries. The birds were divided into 6 groups (Table 1) and maintained in pens 2 X 3 m 3 on deep litter under a conventional lighting regimen of 14 h light and 10 h darkness. Both strains were vaccinated at 1-d and 3-weeks-old against Marek's disease and at 4-weeks-old against Newcastle disease. Mortality was less than 0-05% in all groups. The birds were maintained on a standard starter and grower diet. Food was available ad libitum for groups A, B, C and D (Table 1). Birds of groups E and F had their intake restricted from 4 weeks of age according to a commercial regimen (D. B. Marshall Ltd, Newbridge, Scotland) (Table 1). Water was available ad libitum for all groups. The weekly food-restriction programme is presented in Table 2. TABLE 1
Details of birds and diets used
Strain Layer (Isa Brown)
Group A
n
Sex
no
M
Diet 0-4 weeks—starter (230 g protein, 12-55 MJ
B
110
F
5-20 weeks—grower (190 g protein, 12-55 MJ
C
110
M
0-4 weeks—starter (230 g protein, 12-55 MJ
D
110
F
ME/kg) ad libitum ME/kg) ad libitum
Broiler (breeding strain)
ME/kg) ad libitum
5-20 weeks—grower (190 g protein, 12-55 MJ ME/kg) ad libitum
Broiler (breeding strain)
E
85
M
0-4 weeks—starter (230 g protein, 12-55 MJ
F
85
F
4-20 weeks—restricted grower diet (190 e protein, 12-55 MJ ME/kg) adjusted as for commercial regimen for breeding birds
ME/kg) ad libitum
Birds from groups A to D were bled at 1-d-old, 3 d and one week. Thereafter, they were bled at weekly intervals between 09.00 and 10.00 h until 20 weeks of age. The birds on the restricted diet were bled weekly in a similar manner but from 4 weeks of age. Each bird was bled only once. Five birds from each group were selected at random for bleeding each week and the mean result was presented. The chicks were killed by decapitation until 3 weeks of age and thereafter birds were bled from the basilic vein using sterile disposable plastic syringes and needles (21 gauge). The blood (1-0 to 1-5 ml) was dispelled immediately into collecting tubes containing sequestrene (dipotassium salt of ethylene diamine tetra-acetic acid (EDTA, 1-5 mg/ml of blood) anticoagulant.
ERYTHROCYTE CHANGES AND FOOD RESTRICTION
409
TABLE 2
Weekly food-restriction programme (g/bird d) Age (weeks) Female Male
0-3 ad lib. ad lib.
Downloaded by [University of Guelph] at 05:30 24 February 2013
Age (weeks) Female Male
4 41 50
5 41 50 13 63 73
6 45 50 14 68
77
7
8
50 54
50 54
9 54 63
10 54 63
11 59 68
12 59 68
18
19 86 90
20 91 90
15
16
73 77
77
17 77
82
81
81
86
Haemoglobin (Hb) was determined photometrically as cyanmethaemoglobin:blood was diluted 1:50,000 with Isoton II (Coulter Electronics Ltd, Northwell Drive, Luton, Bedfordshire). A few drops of Zapoglobin (Coulter Electronics Ltd) were added to lyse the red blood cells (Rbc's). The diluted blood sample was left at room temperature for 20 min before centrifuging at 300g for 2 to 3 min to remove red cell nuclei. Hb values were read directly using a LK 540 photometer (TEM sales Ltd, Gatwick Road, Crawley, West Sussex). The blood sample was also diluted 1:50,000 with Isoton II and the total number of cells was recorded using an electronic particle counter (Fn model: Coulter Electronics Ltd). Two separate dilutions were made with each sample. Each sample was read 5 times and the mean of each diluted sample was corrected with the coincidence correction chart (Coulter Electronics Ltd). The following variables were judged to give good 'plateaux' for fowl blood cells: orifice diameter 100 fim, attenuation 0-707, aperture 16, threshold 12. The counter was calibrated with cell control 4C (Coulter Electronics Ltd) and blood from human volunteers. The Rbc count was determined by counting 12 microscope fields in a blood film with a X100 objective and once the white blood cell/thrombocyte ratio has been established (Maxwell et al., 1990) the total number of Rbc's could be computed. The packed cell volume (PCV) was measured by the microhaematocrit method (Hawksley and Sons Ltd, Lancing, Sussex). The following indices were calculated (Dacie and Lewis, 1975); mean cell haemoglobin (MCH), mean cell haemoglobin concentration (MCHC), mean cell volume (MCV). Reticulocyte counts were performed according to Dacie and Lewis (1975). Whole blood (50 fil) was mixed with equal parts 1% brilliant cresyl blue solution, allowed to stand at room temperature for 1 h and the blood films were air-dried. Fifty microscope fields were counted using a cut-down eyepiece. The reticulocytes were assigned to 5 stages according to the degree of reticulation (Coates and March, 1966). Briefly, this reticulation ranged from a heavily-stained network in the cytoplasm of cell 1 to a few isolated dots in cell 5. All counts were carried out by the same investigators. The variance between observations within the individual groups of birds tended to increase with the mean. Taking square roots of the red blood cell and reticulocyte counts and logarithms of the other variates removed much of this heterogeneity of variance. Analyses of variance were therefore performed in the transformed scale to assess the statistical significance of the effects of sex, strain, food restriction and age.
410
M. H. MAXWELL et al.
RESULTS
A summary of the main effects of food restriction, strain, sex and age on the red blood cell characteristics is presented in Table 3, as means of counts between 4 and 20 weeks of age, their approximate standard errors and t statistic associated with the main effects in the transformed scale. Standard errors were calculated using the approximation for the standard error of a function of a random variable (Kendall and Stuart, 1963). The age range 4 to 20 weeks was divided into two periods 4 to 12 weeks and 13 to 20 weeks.
Downloaded by [University of Guelph] at 05:30 24 February 2013
Haemoglobin
The mean Hb value for the restricted-fed broilers (10-0) (g/100 ml) was significantly lower than for birds fed ad libitum (10-6). Similar degrees of significance in Hb values were seen in favour of broilers (10-6) compared with layers (9-6), males (10-3) compared with females (9-8) and older (10-7) compared with younger birds (9-5) (Table 3). Packed cell volume
The mean PCV value of the restricted-fed broilers (31-2%) was also significantly reduced compared with the birds fed ad libitum (32-9). Similar levels of significance were recorded between broilers (32-9) and layers (29-8), males (32-2) and females (30-4) and older (32-7) compared with younger birds (30-0) (Table 3). Red blood cells
Table 3 shows that there were significantly more Rbc's in the restricted-fed broilers (2-85 XlO6 mm3) than ad libitum-kd birds (2-81). Broilers also had more Rbc's (2-81) in circulation than layers (2-56). Furthermore, the Rbc count in males was 2*80 and 2-70 in females. Older birds had significantly more Rbc's (2-86) than younger birds (2-65). Mean cell haemoglobin
There was a significant reduction in MCH values in the broilers that received the restricted diet (35-1 pg) compared with those birds fed ad libitum (37-9) (Table 3). Significance of similar magnitude was also recorded in older birds (37-8) compared with younger ones (36-2). There were no significant main effects of strain or sex. Mean cell volume
Table 3 shows that the mean MCV value for the restricted-fed broilers (109-0 ftm3) was significantly lower than for ad libitum-fed birds (117-5). Males had significantly larger red cells (115-4) than females (113-7). There was a tendency for them to be larger in older (115-4) than in younger birds (113-8).
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ERYTH]
TABLE 3
Summary of red blood cell characteristics and their effects in birds fed adlibitum and restricted from 4 to 20 weeks—means (±SEM) 6
3
A R I
10 6 (007) 100(006) 5-79***
32-9 (0-2) 31-2(0-2) 6-48***
2-81 (002) 2-88(0-02) 2-78***
Rbc (10 /mm ) MCH (pg) 37-9 (0-3) 351 (0-2) 911***
MCV (/im3) 117-5 (0-6) 1090 (0-5) 10-82***
MCHC (%) 32-2 (01) 32-2 (01) 0-81
Reticulocytes (%) 101 (0-2) 9-5 (0-2) 113
Strain ad libitum
L B t
9-6 (006) 10-6 (007) 10 32***
29-8 (0-2) 32-9 (0-2) 12-36***
2-56 (002) 2-81 (019) g.42***
37-8 (0-2) 37-9 (0-2) 0-42
117-1 (0-6) 117-5 (0-6) 0-61
32-3 (01) 32-2 (01) 0-20
11-8 (0-2) 101 (0-2) 5 62***
Sex
M F t
10-3 (005) 9-8 (005) 5-97***
32-2 (01) 30-4 (01) 8-46***
2-80 (002) 2-70 (001) 4-63***
37-1 (0-2) 36-8 (0-2) 0-99
115-4(0-5) 113-7 (0-5) 2.70***
3 2 1 (01) 32-4 (01) 1-68
10-9 (0-2) 10-0 (01) 3-83***
Age
4-12 weeks 13-20 weeks t
9-5 (005) 10-7 (006) 15-85***
3 0 0 (01) 32-7 (01) 13-68***
2-65 (001) 2-86 (002) 9-76***
. 36-2 (0-2) 37-8 (0-2) 607***
113-8 (0-4) 115-4 (0-5) 1-73
31-7 (01) 32-8(0-1) 7.46***
10-8 (0-1) 101 (0-2) 300***
Effect Restriction Broilers
HB (g/100 ml) PCV (%)
A Ad libitum, R Restricted, L Layer, B Broiler, M Male, F Female. Student's t statistic with 488 DF in transformed scale. ***p 0 w
z0
s 0 0
1 0
z
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M. H. MAXWELL et al.
However, there was no significant difference in MCV values between the two different strains. Mean cell haemoglobin concentration
Table 3 shows that the older group of birds had a significantly larger MCHC value (32-8%) than the younger group (31-7). There were no further significant differences in MCHC values with food restriction, strain or sex.
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Reticulocytes
There were significantly more reticulocytes in the layer strain (11*8%). than in the broilers (10-1) (Table 3). Males had more reticulocytes (10-9) than females and (10-0) and younger birds (10-8) more than older birds (10-1). There was no significant main effect of food restriction. DISCUSSION
When broilers were maintained on a food-restricted diet for up to 20 weeks of age, there was a significant reduction of Hb and PCV but a significant increase in the number of circulating red cells. Furthermore, the indices MCH and MCV were lower than in control broilers that received an ad libitum diet. These results may indicate that more red cells were released from the bone marrow to compensate for the lowered haemoglobin and that these cells, because of the reduced MCV, were also smaller. These food-restricted broilers may thus be suffering from a mild microcytic normochromic anaemia. Evidence from published work (Lee et al., 1971; Blair, 1972; Pearson and Shannon, 1979) has suggested that the effects of food restriction during the rearing period of broiler chickens can be beneficial in terms of their overall biological performance. Because food restriction delays body weight, fat deposition, sexual maturity and even mortality (Pearson and Shannon, 1979), it follows that the progress of the haematological profile will also be reduced in momentum. The changes in red blood cell values probably have a further beneficial advantage on such organs as the heart and lungs, in reducing their overall combined workload (Julian, 1987) and assisting in the control of cardiopulmonary disease in broilers. There is a danger, however, that too severe a restriction programme may cause an even greater anaemia or possibly induce further stress (Maxwell et al., 1990). Therefore, it is important that a correct dietary balance of food-restriction be achieved. This in turn should control haematological values to within a "normal" range, otherwise birds may suffer. EDITORS
NOTE
Tables of raw data, summarised in Table 3, may be obtained from the authors on request.
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