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223
PHY 253
METHODS
Male white carneaux pigeons (Columba livia) weighing between 450 and 600 g
were used. Each pigeon was individually housed and given food (Washtenaw's
Check-R-Mix scratch bird feed, Ann Arbor, Michigan) and water ad libitum. The
daily photoperiod was 12 hr and ambient temperature was maintained at 22-23 C.
Room temperature and the body temperature of each bird were recorded about
every 30 sec on a multipoint recorder (Honeywell Electronik 112). Copperconstantan thermocouples (36 s.w.G.) were placed inside polyethylene tubing
(P.E. 205) which was sealed at the end and placed about 20 mm into the cloaca
of each bird. The thermocouple lead wire was then taped to the tail feathers and
allowed to drape freely from the cage. This allowed the birds unrestrained movement
in their cages during the course of the experiment. Each group of birds was allowed
one day to adjust to the handling and placement of the cloacal probes. The second
day generally served as a control period and the experimental manipulations were
carried out on the third day.
The bacterium Pa8teurella multocida was chosen because it is a gram-negative
bacterium that is known to cause fowl cholera (Heddleston, 1972). The stock
cultures of bacteria were supplied by Dr G. R. Carter of thq Department of
Microbiology at Michigan State University. The bacteria were grown on sheep
blood agar at 370 C, washed twice with sterile pyrogen-free saline, centrifuged
and re-suspended in pyrogen-free saline. The appropriate concentrations were
made by diluting this suspension until they matched the reference concentration
of McFarland barium sulphate standards. Dead bacteria were obtained by initially
washing the agar plates with 70 % ethyl alcohol. Confirmation that the bacteria
were dead was made by plating the final suspension of bacteria on sheep blood
agar plates and observing no growth. The various concentrations of dead or live
bacteria as well as sterile pyrogen-free saline (controls) were all injected i.P. For
each experimental situation the injections and administration of sodium salicylate
(as well as the handling of the birds) was controlled for by injection of pyrogen-free
saline or the administration of water by mouth. To study the effect of antipyretics
on the fever, we administered 0-67 ml. tap water or sodium salicylate (300 mg/ml.)
orally by inserting 20-30 mm polyethylene P.E. 160 tubing, connected by an
18 s.W.G. needle (1-2 mm o.d.) to a syringe, into the mouth of each bird.
RESULTS
225
EVOLUTION OF FEVER
ranged from 40-72 + 0.040 C s.E. of mean during the day (08.00-18.00 hr)
to 39 70 + 0.09 C s.E. of mean at night (18.00-04.00 hr). (See Table 1 and
Fig. 1.)
TABLE 1. Febrile responses in pigeons
Time
1st lOhr
(08.00-18.00)
A, control body temperature 40-72 + 004
B, 5 x 107 P. multocida
+0-150-07
C, 5 x 108 P. multocida
+0-45 0*11
D, 5 x 109 P. multocida
- 0.01 + 0-10
E, 5 x 109 P. multocida
-0-080 11
F, 5 x 1010 P. multocida
+ 0 19 0-13
- 0 45 + 0-10
G, 5 x 108 P. multocida
+ sodium salicylate
H, sodium salicylate
-0-500-10
2nd 10 hr
(18.00-04.00)
39-70 0 09
+ 0-06 0-02
24 hr
(08.00-08.00)
40-21 + 0-11
+ 178+0-12
-0-23+0-10
+1-060-17
-0-320-06
N
22
5
5
7
4
5
8
-0-21 0-08
- 0-32 + 0-12
+0-09+0-03
+0-800-10
+0-50+ 011
+1X210-07
+1-01+0-04
+1-030-08
+ 0-48 + 0-12
A, average body temperature of pigeons injected with 1 ml. sterile pyrogenfree saline.
B-F, average changes in body temperature in response to varying concentrations
of dead Pasteurella multocida (1 ml., i.P.).
E, pigeons were re-inoculated with the same dose (5 x 109) dead P. multocida
after 6 days.
G, average changes in body temperature in response to 5 x 108 P. multocida and
sodium salicylate (0-67 ml. 300 mg/ml. at 08.00 and 16.00 hr).
H, average changes in body temperature in response to sodium salicylate (0-67 ml.
300 mg/ml. at 08.00 and 16.00 hr).
+ s.E. of mean, n = number of pigeons.
41-0
. .40.01
CL
E
0
390
39-0
12.00
I,,| | i
18.00
24.00
II
06.00
Hours
226
44
0_1%~ ~ ~~1
U 43_
39~~~~~~~~~~~~~~I
40 _
12.00
a
18.00
24.00
Hours
06.00 10.00
227
EVOLUTION OF FEVER
tail of each bird. On day 2, at 08.00 hr, 1 ml. sterile pryrogen-free saline
was injected (i.r.) into each bird. On day 3, at 08.00 hr, 1 ml. Pasteurella
multocida in saline vehicle was injected (i.P.). The concentrations of
bacteria used were 5 x 107 (five birds), 5 x 108 (five birds), 5 x 109 (seven
birds) and 5 x 1010 (five birds) organisms/ml. The results of a typical
experiment in which 5 x 109 organisms/ml. were injected on the 3rd day
are compared to the saline control injection on the 2nd day in Fig. 3.
The average data for each concentration of bacteria are presented as the
differences in body temperature at each hour between the control and
the experimental observations (Fig. 4A and B). A concentration of
5 x 107 organisms/ml. led to a minimal change in body temperature when
42
U 41:14
%- I
~40
391.
J lS
12.00
Lghts off rb
I I
18.00
24.00
Hours
1.
06.00
228
20 r A
, 10
e
M
0.CL
E0
0
.0
.C
0
C
-1.0
2
1800
Hours
P. multocida
5 x 10' (n=5)
onto 5 x 10' (n=7)
-.
0IS..
0
'a
0
0.
E0
0
.0
C
0
C
12.00
18.00
24.00
06.00
Hours
229
EVOLUTION OF FEVER
temperature (Fig. 4B). After this initial drop and oscillation, a steady
fever developed with a magnitude similar to that in the group of animals
who were injected with 5 x 108 organisms/ml. The pattern of response to
5 x 1010 organisms/ml. showed a similar initial drop and oscillation before
the full development of the fever. The possible significance of these
oscillations will be developed in the discussion.
To determine whether the febrile response in birds was reduced in
response to a subsequent injection of dead Pa8teurella multocida, four
birds were injected a second time with 1 ml. 5 x 109 organisms/ml. Six
days were allowed for recovery between the first and second experimental
sessions. The protocol for this tolerance test was that used for the initial
2-0
1-5
01W
L-
1*0
0.
E
V
0-5
.0
C
80
C
-0.5
-1 *0
12.00
24.00
18.00
06.00
Hours
231
EVOLUTION OF FEVER
mechanism, with a somewhat longer latency, could be the development
of granulocytic endogenous pyrogen which would then produce the
observed elevation in body temperature. Regardless of the mechanism,
this cryogenic response at higher doses obscures the initiation of the
daytime fever at the higher doses of bacteria.
TABLE 2. Febrile responses in the terrestrial vertebrates
Reptiles
Birds
+*
+
+
+
?
+
+*
Mammals
+
+
+
?
This work was supported by NSF research Grant GB 42749X. We are most
grateful to G. Carter for providing us with the stock culture of Pa8teurella multocida.
We thank H. Bernheim, W. Dawson, D. Mouw and L. Vaughn for their critical
review of this manuscript.
Special thanks goes to M. Foster for her technical assistance in the microbiological
aspects of the study and to C. Rose for her assistance in data reduction and
analysis.
REFERENCES
DAwsoN, W. R. & HUDSON, J. W. (1970). Birds. In Comparative Physiology of
Thermoregulation, vol. 1, Invertebrates and Nonmammalian Vertebrate8, ed.
WHITTOW, G. C., pp. 223-310. New York and London: Academic Press.
GLATT, M., PESKAR, B. & BRUNE, K. (1974). Leucocytes and prostaglandins in
acute inflammation. Experientia 30, 1257-1259.
HEDDLESTON, K. L. (1972). Avian pasteurellosis. In Diseawes of Poultry. 6th edn.,
ed. HOFSTAD, M. S., pp. 219-251. Ames: Iowa State University Press.
KLUGER, M. J., RINGLER, D. H. & ANVER, M. R. (1975). Fever and survival. Science,
N.Y. 188, 166-168.
SEIBERT, F. B. (1925). The cause of many febrile reactions following intravenous
injections. I. Am. J. Phy8iol. 71, 621-651.
VAUGHN, L. K., BERNIHEIN, H. A. & KLUGER, M. J. (1974). Fever in the lizard.
Dipeo8aurue dormali. Nature, Lond. 252, 473-474.