250 Laboratory Animals (1988) 22, 250-254

Osmotic fragility of erythrocytes of two breeds of domestic fowl

in the warm humid tropics
J. O. OYEWALE & L. A. DUROTOYE
Department of Veterinary Physiology and Pharmacology, University of Ibadan, Ibadan, Nigeria
Summary
Erythrocyte (RBC) count, packed cell volume
(PCV) and haemoglobin (Hb) values were found
to be higher in Nigerian domestic fowls than
those in Hubbard fowls. PCV and Hb values
were higher in males than in females in both
breeds. Erythrocytes of the Nigerian fowl were
observed to be more susceptible to osmotic
haemolysis than were erythrocytes of the
Hubbard fowl. Erythrocytes of male fowls were
osmotically more fragile than were those of
female fowls in both breeds.
Keywords: Erythrocyte osmotic fragility;
Domestic jowl; Warm humid tropics
The resistance of erythrocytes to osmotic
haemolysis has been investigated in various
animal species (Perk et al., 1964; Soliman &
Amrousi, 1966; Jain, 1973). Even for the same
animal species, several authors have presented
varied values (Schalm et al., 1975), probably as
a result of differences in the factors known to
influence osmotic fragility, such as size, volume
and age of erythrocytes, temperature and pH of
the medium (Stone et al., 1953; Perk et al., 1964;
Schalm et al., 1975). Data for fowl are especially
rare. Perk et al. (1964) and Soliman and Amrousi
(1966) reported marked differences between the
osmotic behaviour of erythrocytes of fowl and
other domestic animals in a temperate
environment. Jaffe (1960) also in a temperate
area observed a difference in the resistance of
erythrocytes to osmotic pressure between inbred
Received 13 November 1986; accepted 23 December 1987
White Leghorn strains C and W. It may,
however, be necessary to extend the study to fowl
in the tropical environment, considering the
clinical importance of the values of the
erythrocyte osmotic fragility and the many blood
parasitic diseases which are prevalent in this
environment. It has been shown that the
resistance of erythrocytes to osmotic haemolysis
may increase or decrease in some haematological
disorders (Jain, 1973). In an earlier study,
Durotoye and Oyewale (1986) found the value
of erythrocyte osmotic fragility of the Nigerian
domestic fowl to be lower than that of the semi-
domesticated guinea-fowl in the same environ-
ment. The aim of the present study was to
compare the erythrocyte osmotic fragility of the
Nigerian domestic fowl with that of an exotic
domestic fowl, the Hubbard fowl, in the warm
humid tropics.
Materials and methods
Adult Nigerian domestic fowl and adult
Hubbard fowl which belonged to the University
of Ibadan Teaching and Research Farm were
used. The birds were housed in deep litter pens
and given a balanced poultry feed (Pfizer Ltd,
Ibadan, Nigeria) and water ad libitum. Twenty-
six clinically healthy Nigerian domestic fowls (13
males, 13 females) and 20 clinically healthy
Hubbard fowls (8 males, 12 females) were
sampled.
Blood was drawn from the jugular vein of each
fowl into bottles, using ethylene diamine
tetraacetic acid (EDT A, 2 mg/ml) as
anticoagulant, and was analysed within 2 h of
collection. From a 1fr/o phosphate buffered
(Na2HP04 l' 37 mg/ml,. and NaH
2
P0
4
.2H
2
0
Erythrocyte osmotic fragility of fowl
o 24 mg/ml) sodium chloride (NaCl 9 mg/ml)
solution 5 ml of each of various concentrations
of NaCl solution (0'85, 0'80, 0'75, 0'70, 0'65,
0'60,0'55,0'50,0'45,0'40,0'35,0'30,0'25,
020 and 0'10070)were prepared in 15centrifuge
tubes, while the 16th tube contained 5 ml of
distilled water. The pH of the distilled water
(7' 2) and the NaCl solutions (7 4- 7, 9) were
measured using a pH meter (Model 7050,
Electronic Instruments Ltd, Chertsey, Surrey,
UK). To each concentration of the NaCl solution
and the distilled water, O' 02 ml of blood was
added. The contents were mixed and allowed to
stand at room temperature (28-29C) for
30 min. The tubes were then centrifuged at
2000r.p.m. for 10min and about 3'5 ml of the
supernatant in each tube was later transferred to
another set of tubes. The optical density of each
solution was read on a CE404 Colorimeter (Cecil
Instruments, Cambridge, UK) at a wavelength
of 540J.tM,using distilled water as the blank. A
cumulative erythrocyte osmotic fragiligram was
obtained by plotting the percentage haemolysis
against the concentration of the NaCl solutions.
The derivative fragiligram was obtained from the
values of percentage haemolysis by using the
principle of 'haemolytic increaments' (Suess et
al., 1948).
251
Other haematological parameters analysed
included packed cell volume (PCV) using
the microhaematocrit method; haemoglobin
(Hb) content using the cyanmethaemoglobin
method; erythrocyte (RBC) count using the
haemocytometer and the mean corpuscular
volume (MCV) calcualted from the values of
RBC and PCV as described by Schalm et al.
(1975).
Results
The mean values of PCV, Hb, erythrocyte count,
MCV and the NaCI concentrations at which
minimum (less than 10070)and maximum (over
90070)haemolysis of the erythrocytes occurred in
Nigerian domestic fowl and Hubbard fowl are
shown in Table I. The PCV was significantly
higher (P<O'OI) in males than in females in the
two breeds of fowl. The mean PCV in Nigerian
domestic fowl was significantly higher (P<O'()()l)
than the value in Hubbard fowl. Although no
significant sex difference was found in the
erythrocyte count in either breed, the mean count
was significantly higher (P<O'OI) in Nigerian
domestic fowl than in Hubbard fowl. The Hb
value was significantly higher in males than in
females in Nigerian domestic fowl (P<OOl) and
in Hubbard fowl (P<O05). The former breed
Table 1. Haematological values (mean SD) and osmotic fragility of erythrocytes of Nigerian domestic fowl and Hubbard fowl
Parameters Sex Nigerian domestic fowls Hubbard fowls
PCV (070) Male 3577297 (13) 33, 25 3, 31 (8)
Fema]e 33'39 170 (13) 28'292'16 (12)
Male and female 34'082'95 (26) 30, 28 36] (20)
RBC (l06/mm) Male 2'550'42 (13) 2'330'17 (8)
Fema]e 2'420'38 (13) 2'140'25 (12)
Male and female 2'480'40 (26) 2'220'24 (20)
Hb (g/dl) Male 1]'31 106 (13) 10'060'95 (8)
Female 9'84 1'01 (13) 9'190'65 (12)
Male and female 10, 58 I' 27 (26) 9'540'89 (20)
MCV (fl) Male 14287 15 87 (13) 142 66 I ] .67 (8)
Fema]e 13723 22 15 (13) 133'08 1167 (12)
Male and female 140'05 1947 (26) 136'96 12'46 (20)
Minimum fragility Male 0'70 (31) 0,70 (8)
(OJoNaCI10070haemolysis) Fema]e 0'65 (13) 0'55 (12)
Male and female 0,70 (26) 0'60 (20)
Maximum fragility Male 020 (13) 0,10 (8)
(OJoNaCI900/0haemolysis) Female 0'10 (13) 0,10 (12)
Male and female 020 (26) 0,10 (20)
Number of birds shown in parentheses
252 Oyewale & Durotoye
PERCENT"C,E SOOIUM CHLORIDE SOLJTION
\,
PERCENT4GE SODIUM cHLORIDE SOLUTION
10
-20
0-8 0-7 0-6 D-S
r:\
'" .' ,
,..- ,
, '
, ,
" ' ,
, ,
I ~J' \'0
/'
,
,
./
0-2 03
~
-_ .~ .. _ "
" "
fI'
,.'4 \,
, '
, '
// '>
i/
" I ,
,
,

04 0-8
-20
6C
100
G O
\I>
II>
> -
0
~
W
.,
X
" ;'
~
2'
W
u
:r
w
..
Fig. 1. Cumulative (A) and derivative (B) erythrocyte osmotic
fragiligrsms for male (0---0 and female (.--.)
Nigerial domestic fowl.
Fig. 3. Cumulative (A) and derivative (B) erythrocyte osmotic
frsgiligrams for Hubbard fowl (0---0) and Nigerian
domestic fowl (....---.).
10
00
&0
w
"
.,
~ 20
z
"'
U
0:
w
..
os
PERCENT"GE SODIUM CHLORIDE SOLUT:"N
-20
Fig. 2. Cumulative (A) and derivative (B) erythrocyte osmotic
fragiligrsms for male (0---0) and female (. .)
Hubbard fowl.
showed a significantly higher (P<001) mean
Hb value than the latter. There were no
significant sex differences in the values of MeV
within and between the breeds.
In both breeds, the osmotic fragility of
erythrocytes of the male fowl was higher than
that of the female (Table 1). Haemolysis
commenced and was completed in a higher saline
concentration in Nigerian domestic fowl than in
Hubbard fowl, indicating a higher osmotic
fragility of erythrocytes of the former breed.
Figures 1-3 show the cumulative (A) and
derivative (B) erythrocyte osmotic fragiligrams
for male and female Nigerian domestic fowl,
male and female Hubbard fowl, and the two
breeds of fowl. The cumulative curves were all
sigmoid in shape, but steeper for the Nigerian
fowl than that for the Hubbard fowl. The
derivative curves showed differences in the peak
of haemolytic increments, being higher for males
than for females of each breed, and for Nigerian
domestic fowl than for Hubbard fowl. The latter
curves also showed one peak for the Nigerian
Erythrocyte osmotic fragility of fowl
domestic fowl and two or more peaks for the
Hubbard fowl.
Discussion
The higher PCY and Hb values in males than in
females in the two breeds of domestic fowl are
similar to observations in other avian species
(Domm et al., 1943; Newell & Saffner, 1950;
Tanaka & Rosenberg, 1954; Nirmalan &
Robinson, 1971). Testosterone has been shown
to increase the PCY and Hb values in the male
(Domm et al., 1943; Newell & Saffner, 1950;
Panda & Juhn, 1961; Fred et al., 1964). We
observed higher RBC count, PCY and Hb values
in the Nigerian fowl than in the Hubbard fowl.
This may suggest a better adaptation of the
Nigerian fowl than the Hubbard fowl to the
warm humid tropics, where metabolic activity
and oxygen requirement are likely to be higher
than in the temperate environment.
The mean PCY reported in the White Leghorn
fowl is 29'62'8OJo (Medway & Kare, 1959)
or ranges between 28 8 and 30% (Lucas &
Denington, 1957). This is closer to the mean
value in the Hubbard fowl (30'283'61%)
than that in the Nigerian fowl (34'08 295%).
The RBC counts in the present study are
lower than those reported in the Brown
Leghorn (2'7-3'6x 1Q6/mm
3
; Domm et al.,
1943) and in the White Leghorn breeds
(3'08-3'36x 106/mm
3
; Lucas & Denington,
1957). The mean Hb values are similar
to those reported in the New Hampshire
fowl (8'61-13'19g/dl; Tanaka & Rosenberg,
1954).
Although the MCY values were not sig-
nificantly different between the Nigerian fowl
and the Hubbard fowl, the values are higher than
those reported in the inbred White Leghorn
strains C and W by Jaffe (1960). The values of
mean corpuscular haemoglobin (MCH) and
mean corpuscular haemoglobin concentration
(MCHC) were also not significantly different
between the Nigerian fowl and the Hubbard
fowl, as observed in a previous study (Oyewale,
1987).
The erythrocytes of the Nigerian fowl
253
appeared more susceptible to osmotic haemolysis
than were those of the Hubbard fowl. Osmotic
fragility varies with the age of red cells (Perk et
al., 1964), the old cells being more fragile
(Prankerd, 1961 cited by Perk et al., 1964), and
the proportion of erythrocytes of different ages
in the blood may vary with the level of metabolic
activity (March et al., 1966). The difference in
the erythrocyte osmotic fragility between
Nigerian fowl and Hubbard fowl may therefore
be associated with a difference in the metabolic
rates of the birds, as suggested between White
Leghorn and New Hampshire breeds (March et
al., 1966). It is however doubtful whether this
may be responsible for the difference obtained
in the osmotic haemolysis between males and
females of both breeds. The fact that erythro-
cytes from male fowls were more susceptible to
osmotic lysis than were those from female fowl
is in accord with the report of March et al. (1966)
in White Leghorns and New Hampshires. These
workers observed that oestrogen increases the
resistance of erythrocytes to osmotic haemolysis,
while androgen- shows no effect on red cell
fragility in birds.
The NaCl concentrations at which the
minimum and maximum haemolysis of erythro-
cytes occurred, respectively, in this study
(0' 7-0' 6% and O 2-0' 1%) are different from
those reported in adult domestic fowl by Jaffe
(1960) (0' 495-0' 420% and O 300-0' 285%),
Perk et al. (1964)(0' 34% and 0'25%), Soliman
& Amrousi (1966) (0'39% and 0'28%) and
Swenson (1970) (041-0'40% and 0'32-0'28%).
All these workers did their studies on birds
in the temperate environment, whereas the
present study was done in a tropical environ-
ment. Also, the breeds used by them were
different. As reported in the present study
and in that of March et al. (1966), a breed
difference exists in erythrocyte osmotic fragility
in domestic fowl. It is hoped, however, that the
data presented in this study should be of help in
assessing the significance of the values that may
be obtained in the tropical environment in disease
conditions, at least in Nigerian and Hubbard
fowl.
254
Acknowledgments
The authors thank Messrs Akin Olowookorun,
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