HEMATOLOGICAL responses to Mycoplasma

Transcription

1 Hematological Response to Cold and gallisepticum in Turkeys 1 ' 2 Mycoplasma A. B. FEDDE 3 AND B. S. POMEROY Department of Veterinary Bacteriology & Public Health, College of Veterinary Medicine, University of Minnesota, St. Paul, Minnesota (Received for publication August 15, 1966) INTRODUCTION HEMATOLOGICAL responses to Mycoplasma gallisepticum (S 6 strain) under normal temperature conditions have been studied by Fedde (1964) and Fedde and Pomeroy (1967). They found that there was a consistent monocytosis, which started as early as the second day post-inoculation and reached a maximum around the tenth day. There was an increase in the total number of white blood cells, total serum proteins, and the absolute number of heterophils, while the Arneth index was decreased. The infectious sinusitis caused by this microorganism is most prevalent in early fall when there is considerable variation in weather conditions and the temperature starts decreasing. This study was designed to determine the response of various blood components in M. gallisepticum inoculated and uninoculated turkeys in a cold environment. MATERIALS AND METHODS Thirteen male and fifteen female Broad White turkeys, five weeks old, were ran- 1 Minnesota Agricultural Experiment Station, University of Minnesota, St. Paul, Scientific Journal Series Paper No This work was supported in part by grants from the Animal Disease and Parasite Research Division, United States Department of Agriculture. 3 This paper is a portion of a study conducted by the senior author in partial fulfillment of the requirements for the Master of Science degree, Department of Veterinary Bacteriology & Public Health, College of Veterinary Medicine, University of Minnesota. 503 domly divided at the start of the experiment into three replicate groups of ten, ten, and eight birds and put in Hudson batteries. Blood samples were drawn from the cutaneous ulnar vein on three different days prior to the eighth day after the start of the experiment at which time all but two birds from each group were placed into a cold environmental room. Two days later four, four and three birds were taken from the respective groups in the cold environment and put in another cold environmental room and inoculated with Mycoplasma gallisepticum (S 6 strain). Thus, at this time there were six birds in a normal temperature environment not inoculated, eleven birds in a cold environment not inoculated, and eleven birds in a cold environment that were inoculated. These birds were kept in the cold environments for thirteen additional days. Then, still keeping them separate, they were placed in a normal temperature environment (similar to that of the normal temperature controls) for nine days longer. The cold environment was produced by a twenty inch fan blowing over one ton of ice, in five blocks (replaced three times weekly), directly onto the turkeys in the batteries. The temperature ranged from 4 C. to 11 C. and the relative humidity ranged from 55 to 74%. Later, with no ice or fan, the temperature ranged from 21 C. to 28 C. and the relative humidity ranged from 32 to 72%. The control birds were maintained at a temperature of 17 C. to 26 C. and the relative humidity of 56 to %. The relative humidity was read di-

2 504 A. B. FEDDE AND B. S. POMEROY rectly in percent from a continuously running Bendix hygrothermograph. This was checked several times with a Taylor sling psychrometer, which gave slightly lower readings. The Bendix hygrothermograph also continuously recorded the temperature. The wind velocity was measured by a velometer (Illinois Testing Laboratories, Inc.). It gave readings of approximately 125 to 150 feet per minute. Blood samples were obtained from birds in each replicate every three days, except for the time immediately following the cold stressor when they were taken every two days. Individual body weights and the amount of feed consumed were recorded at each bleeding time. Post-mortem examinations were made on all birds to determine if uninoculated birds were free of lesions and the severity of the lesions in the inoculated birds. All of the blood parameters were measured by the methods used and explained by Fedde (1964) and Fedde and Pomeroy (1967), except the total serum proteins which were measured in a T-S meter. The statistical analysis of the data was conducted according to the methods of Bennett and Franklin (1954) as described by Fedde (1964) and Fedde and Pomeroy (1967). Confidence bands were drawn about the control mean of the different blood components studied. Any treatment mean is significantly different from the control mean when it appears outside of the confidence band. This presentation shows significant differences as well as the behavior of the treatment means. The hematological responses of the uninoculated birds in the cold environment were compared to those of the control birds in the normal temperature environment in order to determine if significant responses due to cold had occurred. The confidence band for this comparison is shown by the horizontal, straight, solid lines. The responses of the inoculated birds in the cold environment were compared to those of the uninoculated birds in the cold environment in order to determine if significant changes due to inoculation had occurred. The confidence band for this comparison is shown by the horizontal, straight, broken lines. RESULTS The effect of ambient temperature changes and M. gallisepticum (PPLO) infection on various hematological parameters is shown in the following figures. The average values of the hematological parameters studied of uninoculated turkeys at normal temperatures for the entire experiment are shown in Table 1. White blood cell count. The number of white blood cells of uninoculated birds in the cold environment decreased on the first day after they were put in the cold room to a minimum of 20,500 cells/cu. mm. of blood (Figure 1) on the third day. The values remained lower than the control birds throughout the experiment. The inoculated birds in the cold environment showed a slight increase only on the seventh and tenth days post-inoculation with a value of 36,500 cells/cu. mm. of blood. TABLE 1. Grand average blood values of uninoculated turkeys in normal temperatures (17 C.-26 C.) W.B.O Mono." Het.» Lym." Eos. a Bas. a Arneth Index T.S.P> 31,250 2,180 11,560 15, , Units of measurement: a =cells/cu. mm. of blood b = grams/100 cc. of blood (Total Serum Protein)

3 MYCOPLASMA AND COLD 505 FIG. 1. Effect of ambient temperature changes and M. gallisepticum infection (PPLO) on the total white blood cell count. Monocytes. The uninoculated birds stressed with cold showed no significant differences from the control birds in the absolute num- 1 «4 1 1 m s s Z*^*^&*" 4 00LD a m _. _. "* PPLO ber of monocytes (Figure 2). The birds in the cold environment that were inoculated had an increase in the number of monocytes Cold + PPLO Cold *< VAJW - ^ I U v... 0 L-l L. J 1 A > J a o k» It a BUI FIG. 2. Effect of ambient temperature changes and M. gallisepticum infection (PPLO) on the absolute number of monocytes.

4 S06 A. B. FEDDE AND B. S. POMEROY FIG. 3. Effect of ambient temperature changes and M. gattisepticum infection (PPLO) on the absolute number of heterophils. on the second day post-inoculation with a value of 3,600 cells/cu. mm. of blood. The maximum of 5,400 cells/cu. mm. of blood occurred on the seventh day post-inoculation and then gradually decreased. Heterophils. The uninoculated birds in the cold environment gave a slight response in the absolute number of heterophils (Figure 3). This decrease started on the third day and again on the seventh day after the birds were put in the cold environment, and went to 7,000 cells/cu. mm. of blood. The inoculated birds in the cold environment gave the opposite response, or an increase in the number of heterophils. The initial response occurred on the fourth day post-inoculation with a value of 17,000 cells/ cu. mm. of blood. A maximum of 20,500 cells/cu. mm. of blood was reached on the tenth day. Lymphocytes. The cold environment stimulated a significant decrease in the absolute number of lymphocytes (Figure 4). The first day after the birds were placed in the cold environment, a minimum of 11,000 cells/cu. mm. of blood was reached. This minimum was present five other times during the experiment. The lymphocyte count of the inoculated birds in the cold environment also decreased, but was significantly different from that of the uninoculated birds in the cold environment only at one point when the value was 9,500 cells/cu. mm. of blood on the tenth day. Eosinophils. There was a significant decrease in the absolute number of eosinophils with the uninoculated group of birds in the cold environment (Figure 5). It started on the fifth day after presentation of the cold environment with a value of 280 cells/cu. mm. of blood, and reached a minimum of 180 cells/cu. mm. of blood on the seventh and twenty-second days. There was a significant increase in the number of eosinophils in the inoculated birds in the cold environment as compared to the uninoculated group in the cold environment on the

5 MYCOPLASMA AND COLD 507 FIG. 4. Effect of ambient temperature changes and M. gallisepticum infection (PPLO) on the absolute number of lymphocytes. second, seventh, tenth, and sixteenth days Basophils. The absolute number of basopost-inoculation with values of 450, 450, phils of the uninoculated birds in the cold 400, and 380 cells/cu. mm. of blood, re- environment was significantly decreased spectively. from the control birds (Figure 6). The iniw.. Oratnl 0»M + mo FIG. 5. Effect of ambient temperature changes and M. gallisepticum infection (PPLO) on the absolute number of eosinophils.

6 508 A. B. FEDDE AND B. S. POMEROY *5 y 1 } FIG. 6. Effect of ambient temperature changes and M. gallisepticum infection (PPLO) on the absolute number of basophils. tial response was on the fifth day after the birds were put in the cold environment when there were 750 cells/cu. mm. of blood, and a minimum occurred on the 2.1 t tenth day when there were 600 cells/cu. mm. of blood. The inoculated birds in the cold environment had significantly more basophils post-inoculation throughout the CO ID FPU) VABt _,. Control Cold PPLO Cold sin FIG. 7. Effect of ambient temperature changes and M. gallisepticum infection (PPLO) on the Arneth Index.

7 MYCOPLASMA AND COLD oois ««mo ^ HUH 1«S! * fc.o i {,, 3.0. K,, Ofmtral _._, -u y» OoH mo FIG. 8. Effect of ambient temperature changes and M. galusepticum infection (PPLO) on the total serum proteins of blood. experiment. The initial increase was on the tenth day post-inoculation when there were 1,450 cells/cu. mm. of blood and a maximum of 1,625 cells/cu. mm. of blood occurred on the twenty-second day. Arneth index. The Arneth index (Figure 7) was significantly decreased starting on the fifth day after presentation of the cold environment with a value of The minimum of 1.77 occurred nineteen, twentytwo, and twenty-five days after the birds were made cold. The two stressors, cold plus infection with M. galusepticum, decreased the index significantly from the cold alone on the fourth day post-inoculation. The index was then 1.74 and reached a minimum of 1.70 on the nineteenth day. Total serum protein. The cold environment had very little effect on the quantity of total serum protein (Figure 8). The value fluctuated from 4.63 to 4.55 grams per 100 cc. of blood. The pre-inoculation value was 4.58 grams percent for the inoculated birds in the cold environment and reached 4.82 OoU grams percent in seven days and to 5.0 grams percent in twenty-two days after inoculation. Feed consumption. There was less feed eaten by birds in both stressed groups than in the control group (Figure 9). The maximum difference of 1.5 pounds of feed consumption per bird between the control birds and the inoculated birds in the cold environment, occurred twenty-two and twenty-five days after the latter were placed in the cold environment. Body weight. There was also less average body weight for the uninoculated birds in the cold environment and the inoculated birds in the cold environment, as compared to the control birds (Figure 10). The inoculated birds in the cold environment gained less than the uninoculated birds in the cold environment. Serology. There were positive results with the M. galusepticum tests as early as the fourth day post-inoculation and most of the

8 510 A. B. FEDDE AND B. S. POMEROY 10». 9 00LD "^ PPLO 8 7 i: 3 3 _. _. Control Sold + PPLO Ooli I I I 1 1 i I I I L. -I 1 I t~- 0 2 H nun FIG. 9. Effect of ambient temperature changes and M. gallisepticum infection on the average accumulative feed consumption. birds were positive by the seventh day. All of the birds were positive by the tenth day. The uninoculated birds remained negative, Post-mortem. There were no lesions of in H.5 "= ML ft i (PPLO) fectious sinusitis in any of the uninoculated birds when autopsied. The lesions of the respiratory system in the inoculated birds ranged from mild to severe, with the majority being moderate. 0»m»r»l _, _. Ooli PPLO _ OoU 1.5 I I I I it 19 FIG. 10. Effect of ambient temperature changes and M. gallisepticum infection (PPLO) on the average body weights.

9 MYCOPLASMA AND COLD 511 DISCUSSION An experiment was designed to show how different blood components of turkeys were affected by a cold environment, and a cold environment compounded with inoculation of Mycoplasma galusepticum (S 6 strain). The absolute number of monocytes did not differ between the control birds and the uninoculated birds in the cold environment. There was a significant increase in the monocytes of the inoculated birds in the cold environment post-inoculation as compared to the uninoculated birds in the cold environment. Fedde and Pomeroy (1967) reported a significant increase in the absolute number of monocytes after inoculation of the same infective agent to birds in a normal environment. There were greater values for the monocytes of inoculated birds under normal temperature conditions in their experiments than the inoculated birds under the stress of cold temperatures in the present studies. A monocytosis in chickens infected with pleuropneumonia-like organisms was reported by Olson et al. (1957). The values of the total serum proteins of the uninoculated birds in the cold environment showed no difference from the control birds. All of the other blood components of the uninoculated turkeys in the cold environment were significantly decreased from the control turkeys. Recently Lloyd and Glick (1960) reported that white blood cell counts were not consistent in young chickens bled after thirty minute exposures to 5 C. to 7 C. temperatures. This lack of consistency may have been due to insufficent exposure to cold, either in length of time or intensity of temperature. The Arneth index of the inoculated birds in the cold environment was significantly decreased from that of the uninoculated birds in the cold environment, while the absolute number of lymphocytes was not significantly different between the two groups. All of the other blood cell components studied in the inoculated birds in the cold environment were significantly increased from the uninoculated birds in the cold environment. The values of the total serum proteins were increased by the third week. The group of uninoculated birds in the cold environment and the group of inoculated birds in the cold environment consumed less feed and had lower average body weights than the control group of turkeys. Shelton and Olson (1960) report that Mycoplasma infected chickens had less gain in weight than the control group. The difference appeared on the fourth day postinoculation and persisted for a month. SUMMARY Different blood components were studied in control turkeys in a normal temperature environment, and in turkeys uninoculated and inoculated with Mycoplasma galusepticum (S 6 strain) subjected to a cold environment of 4 C. to 11 C. There was a general leucopenic response in the blood of uninoculated turkeys in the cold environment. In the uninoculated birds there were no significant differences in the absolute number of monocytes or the values of the total serum proteins of the birds in the cold environment as compared to the birds in normal temperatures. All of the other parameters measured showed significantly decreased values for the turkeys in the cold environment. Some of these parameters decreased immediately on the first day after the exposure and some reached a minimum from the third to sixteenth days. All of the blood components studied in the group of birds in the cold environment inoculated with the infective agent were significantly increased from the uninoculat-

10 512 A. B. FEDDE AND B. S. POMEROY ed group in the cold environment except the absolute number of lymphocytes which showed no difference and the Arneth index which was decreased. The hematological response of turkeys to a cold environment as compared to normal temperatures was a significant depression of the majority of the blood components studied. The blood values of birds with the Mycoplasma infection were similar when affected by a cold environment or under normal temperatures. The maximum blood counts were not as great when the inoculated birds were made cold as when they were in normal temperatures. ACKNOWLEDGMENTS The authors wish to express sincere gratitude to Dr. Charles E. Gates for his advice on statistical analysis methods, to Dr. Alfred M. Lucas with his assistance on cell morphology and blood techniques, and to INTRODUCTION THE study of the metabolism of exogenous C 14 -epinephrine was begun in the chicken with an investigation of the influence of iproniazid upon the inactivation and removal of epinephrine through its inhibition of a monoamine oxidase (Scott, 1962). The role of the kidney tubule of the chicken in the excretion of labeled epinephrine was the subject of investigation by Rennick et al. (1963). Meanwhile, during x This research was supported by the U.S. Public Health Service Grant H Dr. Richard E. Dierks for his technical assistance. REFERENCES Bennett, C. A., and N. L. Franklin, 19S4. Statistical Analysis in Chemistry and the Chemical Industry. John Wiley and Sons, Inc., New York. Fedde, A. B., and B. S. Pomeroy, Hematological response to Mycoplasma gallisepticum in turkeys. Poultry Sci. 46: 492-S02. Fedde, A. B., Hematological response to infectious sinusitis in turkeys. Master's Thesis. University of Minnesota. Lloyd, C, and B. Glick, Effect of temperature as a stressor on white blood cells, adrenals, and bursa of Fabricius of chicks. Am. J. Physiol. 198: Olson, N. O., D. C. Shelton, D. A. Munro and R. Bletner, Preliminary blood studies in chickens with a synovitis caused by the infectious synovitis agent, pleuropneumonia-like organisms and a combination of the two agents. Avian Dis. 1: Shelton, D. C, and N. O. Olson, Serum protein of chicks with infectious and Mycoplasma synovitis. Poultry Sci. 39: The Metabolism of Injected Epinephrine-C 14 Acetate by Leghorn Pullets 1 JOSEPH L. SCOTT Department of Zoology, University of Connecticut, Storrs, Connecticut (Received for publication August IS, 1966) the past decade, the metabolic pathway by which epinephrine is inactivated within the mammalian body has been established, tentatively, based upon evidence obtained from experimentation on man and augmented by the results from other forms of mammals. A recent paper by Goodall and Alton (1965) contains the details of urinary excretion of the metabolites of injected C 14 -epinephrine in man. Axelrod (1966) has reviewed primarily the methylation reactions with some reference to deamination reactions involved in the formation and metabolism of catecholamines. Making