THE UNIVERSITY OF ILLINOIS LIBRARY cop. ACRICUITURE LION Cli BHECK FOR jCIRCULATINf Inheritance of Resistance to Bacterial Infection in Animals A Genetic Study of Pullorum Disease By ELMER ROBERTS and L. E. CARD UNIVERSITY OF ILLINOIS AGRICULTURAL EXPERIMENT STATION Bulletin 419 CONTENTS PAGE INTRODUCTION 467 MATERIALS AND METHODS 468 EXPERIMENTAL RESULTS 469 Resistance of Selected Strains of Chickens to Inoculation 470 Effect of Inbreeding on Resistance to Infection 473 Study of Chinese Birds for Resistance to Disease 474 Resistance of Different Breeds to Salmonella fullorum 477 RESULTS OF CROSSES 479 Nature of Resistance and Susceptibility 483 Results of Agglutination Tests 485 Was Resistance Acquired or Natural 485 BIOLOGICAL ASPECTS OF METHODS OF DISEASE CONTROL... 486 SUMMARY AND CONCLUSIONS 488 BIBLIOGRAPHY 489 APPENDIX.. . 491 Urbana, Illinois October, 1935 Publications in the Bulletin series report the results of investigations made by or sponsored by the Experiment Station Inheritance of Resistance to Bacterial Infection in Animals A Genetic Study of Pullorum Disease BY ELMER ROBERTS, CHIEF IN ANIMAL GENETICS, AND L. E. CARD, CHIEF IN POULTRY HUSBANDRY* AN HAS always been engaged in a struggle with the invis- ible forms of life, but not until after the time of Pasteur was fundamental information available enabling him to attack with any degree of success the problem of disease control. The accepted approach to this problem has been environmental ; that is, thru the control of microorganisms or of environmental factors con- ducive to the health of the host. Some people have begun to ask what the role of the host, whether man, animal, or plant, may be in the phenomenon of disease. If an infectious disease is considered as the reaction between a parasite and its host, then the host as well as the parasite should receive consideration in disease control. A genetic study of such a disease is a study of the host in its relation to its re- sistance or susceptibility to infection. Disease has been defined as a departure of the organism from normal functioning or constitution. On the basis of this definition diseases may be classified into two groups: those due primarily to abnormal structure and functioning of certain organs or tissues of the body and those due to living organisms of various kinds. The hereditary nature of many diseases that are due to abnormal structure and functioning has been firmly established. The inheritance and, in many cases, the mode of inheritance, of such disorders as haemophilia, anhidrosis, absence of enamel of teeth, resistance and susceptibility to transplantable tumors, and of many defects of both hair and skin have been fairly well worked out during the past few years. Most of the diseases due to abnormal structure and functioning are relatively rare in comparison with the number of diseases caused by infecting organisms. It is a disease of the latter class with which this study is concerned. To L. C. Thomas, W. M. Dawson, and J. H. Quisenberry acknowledg- ment is due for valuable assistance at various times during the course of this investigation ; also to J. A. Hunter of Tunghsien, China, for generous assistance in the study of Chinese fowls during 1929-30. 467 468 BULLETIN No. 419 [October, That resistance or immunity to certain diseases caused by infecting organisms varies with different genera of animals is a matter of common knowledge. One genus may be entirely resistant to a disease to which another genus is very susceptible. Furthermore, one species or race may be immune to a disease to which another species or race may be highly susceptible.* Facts such as these, drawn from general observation, are valuable for certain purposes, but fundamental infor- mation concerning the relation of heredity to disease resistance can be obtained only thru carefully conducted investigations under con- trolled conditions. Furthermore, it is upon the resistance or immunity of individuals that attention must be centered if facts are to be devel- oped concerning the nature of disease resistance and its application to disease problems. Those who work with animals have frequently noted that some individuals appear to be resistant to disease; that is, that they escape infection or give no evidence of having contracted infection when they have had ample opportunity to do so, or if infected the results are much less severe than in the case of other individuals. The investi- gation reported herein was designed to throw some light on the ques- tion whether such qualities in farm animals may be inherited. The immediate object of the investigation was to study chickens with respect to variability in their resistance to Salmonella pullorum, the causative organism of pullorum disease and, if a sufficiently wide dif- ference was found among them, to attempt to ascertain the degree to which resistance or susceptibility to infection with this organism seemed to be inherited. Several preliminary reports of this work have been made. 4 ' 30 ' 38 * MATERIALS AND METHODS Pullorum disease of fowls was chosen for this study for four reasons: (1) it is a well-defined disease of young chicks usually running its course in a few days; (2) the organism is easily cultured in the laboratory; (3) the biological tests for detecting the infection are considered reliable and are used in breeding flocks in many parts of the country for the elimination of infected individuals; (4) more 'The horse is immune to diphtheria, whereas man is susceptible. Brahma cattle are resistant to Texas fever whereas our native cattle, belonging to a different species, are very susceptible. It is reported that Negroes and Indians are more susceptible to tuberculosis than are Caucasians. The Chinese seem to be highly resistant to B. tetanus. Orientals in general are said to be much more resistant to syphilitic infection than are peoples of other races. *These numbers refer to literature citations on pages 489 and 490. 1935] INHERITANCE OF RESISTANCE TO BACTERIAL INFECTION 469 individuals could be used in studying a disease of young chicks than could possibly have been used had adults been necessary. Materials. More than 29,000 birds of various breeds of the domestic fowl have been used in this investigation, which has extended, in its various phases, over a period of ten years. The foundation breeding stock for the experiment was obtained from several hundred day-old chicks which were inoculated with Salmonella pullorum. The few that survived were used as breeders. Others obtained in like manner were added from time to time. At first only chickens that had survived inoculation were used in the selected stocks, but later the progeny test was used as a basis of selection, those parents whose offspring had shown the highest per- centage of survival being selected for further use as breeders. Procedure. Chicks were inoculated the day after they were hatched, or the twenty-second day after the eggs had been placed in the incubator. The culture was administered orally thru a pipette. a The quantities administered ranged from 1/16 to 1/4 cc., the amount depending upon the virulence of the culture. Since no satisfactory method has yef been devised for standardizing the cultures, the amount to be used had to be arrived at by trial. Eggs from all the stocks to be tested were placed in the same incubator at the same time, thus insuring the same environmental conditions for all during the period of incubation. All chicks of a given hatch were inoculated from the same culture and were then put in the brooder for observation. Chicks of all the different kinds to be tested were placed in the same brooder. In this way environmental differences in temperature, time, space, feed, and differences in cultures were eliminated. EXPERIMENTAL RESULTS The investigation reported herein included three phases: (1) a study of the resistance of certain strains of chickens to inoculation with Salmonella pullorum; (2) a study of various crosses between selected and unselected strains with regard to the inheritance of resist- ance or susceptibility to inoculation; (3) a study of the possibility of the experimental birds having acquired resistance to the disease. The term resistance, as used in reporting the data in these experi- ments, does not imply that the individual was necessarily free from the "The Laboratory of Animal Pathology and Hygiene prepared the 24-hour broth cultures used for inoculation and also made the autopsies and biological tests of the birds. 470 BULLETIN No. 419 [October, infecting organism, but that it was able to survive exposure to the organism or to produce offspring able to survive exposure. Without question, varying degrees of resistance existed in the surviving stock, some chicks exhibiting definite symptoms of infection, while others were free from any visible signs. Because of its simplicity and the accuracy with which it could be applied, survival was used thruout these tests as the basis for selecting resistant breeding stock and for judging the reaction of individuals to inoculation with the infecting organism. Incidentally some data are presented concerning the effect of inoculation on growth retardation and the relative susceptibility of the progeny of growth-retarded dams and the progeny of dams which, judged by their larger growth, had been less affected by inoculation. Resistance of Selected Strains of Chickens to Inoculation The results of inoculating 128 hatches of strains of chickens selected for resistance to infection with S. pullorum and 128 unselected controls are given in Table 16, Appendix. Table 1 gives a summary of all the hatches for Strain I, including those with fewer than fifteen chicks, which are not included in Table 16. The selected stock obviously showed a much greater resistance to disease than did the controls. The same results were obtained with two other strains (Table 2). The mean survival of the chicks from the selected stock was 71.5 1.08 and of the controls 27.44 1.26. The difference was 44.06 1.66, or 26.5 times the probable error. The probability that 4927 this difference occurred by chance is only - , which means that 10 72 the difference between the control and selected stocks was genetic in nature and not due to chance. 8 In only one hatch of the 128 was the percentage of survival among the controls as great as or greater than the percentage of survival among selected stock. In that hatch the selected strain showed 70.6 percent survival and the control 77.4 per- cent. This high survival among the controls indicates that a relatively nonvirulent culture had been used. While these results show that selection is effective in establishing resistant strains, the data do not show what progress can be made by continuous selection thru successive generations. A standardized, or Because of the high degree of statistical significance of the results in the different phases of this study, the mathematical constants are not given except for the data in Table 16 of the Appendix. Individual hatches were used in the statistical determinations. In all tables with the exception of Table 16 individual hatches are combined. 79J5] INHERITANCE OF RESISTANCE TO BACTERIAL INFECTION 471 TABLE 1. RESULTS OF INOCULATING CHICKS FROM STRAIN I SELECTED FOR RESIST- ANCE TO S. PULLORUM AND CHICKS FROM UNSELECTED STOCKS* Year Selected Strain I Control Number inoculated Number surviving Percent surviving Number inoculated Number surviving Percent surviving 1925 . 336 478 1 615 414 188 481 374 160 384 1 142 345 96 395 231 186 2 939 47.6 80.3 70.7 83.3 51.1 82.1 61.8 73\S 71.0 446 596 820 293 559 1 188 764 1 030 5 696 115 61 384 154 118 432 46 290 1 600 25.8 10.2 46.8 52.6 21.1 36.4 6.0 28.2 28.1 1926 1927 1928 1929 1930 1931 1932 1933 252 4 138 Total Records of individual hatches are given in Table 16, Appendix. TABLE 2.- -RESULTS OF INOCULATING CHICKS FROM SELECTED STRAINS II AND III AND CONTROL CHICKS' Year Number inocu- lated Number surviv- , ing Percent surviv- ing Number inocu- lated Number surviv- ing Percent surviv- ing Number inocu- lated Number surviv- ing Percent surviv- ing Selected Strain II Selected Strain III Control 1930 408 1 100 297 801 2 606 330 631 241 489 1 691 80.9 57.4 81.1 61.4 64.9 301 620 216 533 1 670 255 403 186 375 1 219 84.7 65.0 86.1 70.4 73.0 1 188 764 594 1 030 3 576 432 46 322 290 1 090 36.4 6.0 54.2 28.2 30.5 1931 1932 1933 Total Strain II X Strain III Control 1934 540 470 87.0 123 25 20.3 Data on individual hatches are given in Table 16, Appendix. uniform, culture would be necessary for such work. The cultures used in this experiment were prepared in the same way during the entire time, yet variations in survival are so great that comparisons between results of two different years might be misleading. Not only does the possibility of a varying culture exist but there is also great probability of uncontrolled environmental factors which may vary from year to year and thus influence the host. While a part of the death losses in these hatches may not have been due to inoculation with 5\ pullorum, the greatest part without doubt was the result of such infection. Noninoculated chicks of selected strains, of unselected strains, and of various crosses between selected and unselected strains were 472 BULLETIN No. 419 [October, kept under observation at different times in order to determine their mortality compared with the mortality of inoculated chicks. When not inoculated, all these groups showed low mortality (Table 3). While survival has been used in this study as the basis for judging the resistance of individuals to infection, other criteria of resistance could be used, such, for instance, as the effect of infection on the vitality of those that survived. One might expect that the rate of TABLE 3. SURVIVAL OF NONINOCULATED CHICKS FROM RESISTANT AND SUSCEP- TIBLE PARENTS, AND FROM CROSSES BETWEEN THE Two, WHEN SUBJECTED TO THE SAME ENVIRONMENT AS THE INOCULATED CHICKS EXCEPT FOR EXPOSURE TO S. PULLORUM Mating Number hatched Number surviving Percent surviving 1928 Resistant cf cf X Resistant 99 193 179 92.7 Resistant cf cf X Susceptible 9 9 144 138 95.8 Susceptible cf cf X Resistant 99 141 136 96.5 Susceptible cf cf X Susceptible 99 89 81 90.5 Total 567 534 94.2 Hatch No. 1 ... 22 22 100.0 Hatch No . 2 35 35 100.0 Hatch No. 3 37 36 97.3 Hatch No. 4 94 91 96.8 Hatch No. 5 84 77 91.7 Hatch No. 6 91 88 96.7 Hatch No. 7 85 82 96.5 Hatch No. 8 119 103 86.6 Total 567 534 94.2 1929 Fi cf cf X Susceptible 99 38 33 86.8 Fi cf cf X Resistant 9 9 50 43 86.0 Susceptible cf cf XFi99 86 76 88.4 Fi cf cf X Fi 9 9 154 141 91.6 Total 328 293 89.4 growth would be retarded in proportion to the severity of the infec- tion which an individual had experienced. If this is so, the chickens that gained weight more rapidly would perhaps have a higher resist- ance to inoculation than would those which grew less rapidly. A test of the comparative resistance of the progeny of rapidly and slowly gaining dams that had survived inoculation would shed some light on the validity of this assumption. Weights of several hundred inoculated birds that had survived inoculation were therefore taken and the progeny of these dams inoculated. The progeny of the heavy dams showed a survival after inoculation of 75.4 percent and the progeny of the light dams a survival of 64.6 percent (Table 4). Records were also kept of the growth of inoculated and noninocu- lated chicks to the age of 163 days. The average weights of the 79J5] INHERITANCE OF RESISTANCE TO BACTERIAL INFECTION 473 TABLE 4. SURVIVAL OF PROGENY OF HEAVY AND LIGHT DAMS THAT HAD SURVIVED INOCULATION Heavy Light Pen Sire Number inoculated Number surviving Percent surviving Number inoculated Number surviving Percent surviving 13 H-1907 177 140 79.1 152 97 63.8 14 J-3743 195 154 79.0 217 148 68.2 15 J-5649 234 163 69.7 16 J-7688 167 101 60 5 Total 606 457 75.4 536 346 64.6 2400 2 1600 K O Z 1200 I- X U 600 *0r INOCULATED INOCULATED l 30 44 56 72 60 100 114 126 142 150 170 AGE IN DAYS FIG. 1. EFFECT OF INOCULATION ON GROWTH OF CHICKS inoculated chicks that survived were consistently less thruout the period than were those of the noninoculated chicks (Fig. 1). Evidently inoculation has a very definite effect in retarding the growth of individuals that survive infection. Effect of Inbreeding on Resistance to Infection A few comparative tests for resistance to infection with 5"o/moc//a pullorum of inbred and noninbred strains were made. An inbred strain of White Plymouth Rocks had a survival of 72.1 percent among 68 inoculated, while among 110 noninbreds only 3.6 percent survived. Another test with White Leghorns gave among 113 inbreds 52.2 percent survival and 31.6 percent among 462 noninbreds. This suggests that natural selection had occurred among the inbreds, elim- inating to some extent the less resistant individuals. 474 BULLETIN No. 419 [October, The results of inbreeding tests with two strains (T and M) from Strain I (Table 1), which had been selected for two years for its resistance to S. pullorum, are shown in Table 5. Strain M had not been inbred until 1927, the year the data were taken. In the previous year the percentages of the progeny of Strains T and M surviving inoculation were practically the same. In 1927 there was no great difference in the survival of progeny as between brother-sister matings TABLE 5. RESULTS OF INBREEDING WITHIN RESISTANT STOCK, 1927 Mating Strain T, highly inbred for several years* Strain M, first inbred in 192 7 Number inoculated Percent alive at 21 days Number inoculated Percent alive at 21 days B X S 129 100 229 76.7 74.0 75.5 185 1 105 1 290 45.4 72.4 69.3 Other matings Total These strains are branches of Strain I (Table 1); see pages 470-471. and other matings in Strain T; whereas in Strain M, inbred for the first time, the progeny of brother-sister matings had a survival much lower than that of the progeny of other matings. These results are what would be expected if resistance and susceptibility are hereditary, provided no preventive measures for the protection of the birds had been used. In this case no such protection had been given Strain T. Study of Chinese Birds for Resistance to Disease If resistance and susceptibility are hereditary, one would expect, in an old country where fowls have long been domesticated and where there has been no artificial control of disease, to find the animals more resistant than those in a country where domestication is recent, where the disease had been present, and where disease-control measures have been in general use. Thru natural selection, in the older country, susceptible individuals would tend to be eliminated from the popula- tion, only the more resistant ones surviving. Under such circumstances it would be expected that strains would be developed showing greater resistance than the original population and presumably greater resis- tance than shown by animals under protection in a newer country. In order to test this assumption a study of Chinese birds was INHERITANCE OF RESISTANCE TO BACTERIAL INFECTION 475 made by the senior author at Tunghsien, China, in the spring of 1930. Six different kinds of chickens were selected for the tests: 1. Chia Gi (small-type chicken) from the vicinity of Peiping, China 2. Cochins from the vicinity of Peiping 3. Shansi from Shansi Province 4. Langshans from Nantungchow, Central China 5. White Leghorns from stock imported from the United States about 1923 6. Rhode Island Reds imported from Canada specifically for the experiment In addition some chicks from a local Chinese hatchery were tested. The same methods of incubation, inoculation, and brooding were used as described on page 469 except for the chicks from the local hatchery. TABLE 6. STUDY OF DISEASE RESISTANCE IN CHINESE CHICKENS: NUMBER INOCU- LATED WITH S. PULLORUM AND NUMBER AND PERCENTAGE SURVIVING AT THREE WEEKS OF AGE Kind of chicken Number inoculated Number surviving Percent surviving All hatches Chinese Chia Gi (small type) 552 337 61.1 Cochin 397 152 38.3 Shansi 221 59 26.7 409 142 34.7 Hatchery* 298 132 44.3 Imported 306 208 68.0 Rhode Island Red" 442 259 58.6 Total 2 625 Excluding hatches with low mortality due to nonvirulent cultures' 1 Chinese Chia Gi (small type) 370 185 50.0 Cochin 253 73 28.9 158 25 15.8 Langshan 261 41 15.7 Hatchery* 298 132 44.3 Imported White Leghorn b 200 139 69.5 Rhode Island Red" 282 133 47.2 Total 1 822 Parentage unknown. b lmported from the United States about six years before, from Canada for the experiment. ^Judged by the low resulting mortality. "Imported Judged by the number of chicks surviving inoculation with S. pullorum, the White Leghorns were much more resistant than the other kinds tested (Table 6). The small type of Chinese chicken, Chia Gi, was more than three times as resistant as the Shansi and the Langshan chickens and 1.8 times as resistant as the Cochin chickens, 476 BULLETIN No. 419 \October, and the Rhode Island Reds used were almost as resistant as the Chia Gi. The low mortality of the chickens in the first hatches indicates that the cultures used were probably less virulent than those used in later inoculations in which, with the same kinds of chickens, a higher mortality resulted. The mortality in the later hatches, however, showed about the same relative differences between the different kinds of chickens in their resistance to the disease as were obtained in the first hatches. It is not known to what extent the Rhode Island Reds, the third most resistant of the kinds tested, may have been subjected to selec- tion previous to this study. The disease had been present in the White Leghorn flock from which samples were taken for this study, accord- ing to the people from whom they were obtained, and the agglutina- tion test showed several reacting individuals. The White Leghorns had also been inbred to some extent after their importation into China, some for as long as five generations. If resistance and susceptibility to disease are hereditary, this inbreeding would automatically produce some individuals tending to be pure for factors for resistance and others pure for factors for susceptibility ; and the operation of natural selection would tend to eliminate the susceptible and retain the resist- ant. There would be no more rapid way of producing a highly resistant strain than to inbreed and at the same time subject the individuals to the disease in question. Agglutination tests were made on the birds used in this experi- ment. Tests of commercial stock were made by Chen Ken of the University of Nanking, blood samples being obtained at the place of killing (Table 7). TABLE 7. RESULTS OF AGGLUTINATION TESTS FOR PULLORUM DISEASES IN STUDY OF CHINESE CHICKENS Kind of chicken Test negative Test positive Suspicious Chinese Chia Gi (small type) 22 5 Cochin 24 2 21 4 Langshan 28 Commercial stock . 678 25 28 Imported Rhode Island Reds 27 1 White Leghorns 17 8 Positive reactors were found among all the Chinese chickens except the Langshans, indicating that the disease is present in China, but how long it has been present is not known. It may have been -7935] INHERITANCE OF RESISTANCE TO BACTERIAL INFECTION 477 brought in with imported stock or it may have been in existence in China for a very long time. The Langshan chickens, which showed no reactors, were obtained from a region where no importation of foreign birds had been made. Mr. Yao, of the College of Agriculture, University of Nanking, who lived in Nantungchow for many years, said that no ailment similar to pullorum disease had been found, to his knowledge, among Langshan chickens. Altho several reactors were found among the Shanshi chickens, the survival among these chickens, when inoculated with S. pullorum, was as low as among the Langshans. The possibility exists that pullorum disease had been of recent introduction into the Shanshi chickens; if so, results similar to those obtained with nonreacting Langshan chickens would be expected. The small-type Chinese chicken, Chia Gi, which is very numerous in the vicinity of Peiping, under all conditions was more resistant to inoculation with S. pullorum than any of the other Chinese birds. The larger number of these chickens may be due partly to their greater resistance to pullorum disease. The resistance of the Cochin, found in the same region as the Chia Gi but not in such large numbers, was not nearly so high as that of the Chia Gi, yet it was higher than the resistance of either the Shanshi or the Langshan chickens. Stocks of Langshans and Chia Gi were shipped to the United States (Urbana, Illinois) for further work in connection with this investigation. In 1931 the percentage of Chia Gi chicks that survived inoculation was 50.8, of the Langshans 7.5, and of unselected Rhode Island Reds 6.2. The difference between the Chia Gi and the Lang- shan chickens exhibited under conditions in Urbana was practically the same as found in China in the spring of 1930, indicating that the differences were due to genetic and not to environmental factors. The high resistance to pullorum disease shown by the Chia Gi chicken of North China and the low resistance of the Langshan of Cen- tral China suggest the operation of natural selection. In North China the birds had been exposed to pullorum disease ; apparently the more susceptible had been eliminated and the more resistant had survived. In Central China the Langshan, so far as was known, had not been in contact with the disease, so that natural selection was impossible. Resistance of Different Breeds to Salmonella Pullorum The resistance to Salmonella pullorum exhibited by the different breeds tested in this study varied greatly. For example, Rhode Island 478 BULLETIN No. 419 [October, Reds were, in general, less resistant than the White Leghorns, White Plymouth Rocks, Barred Plymouth Rocks, and Chia Gi which were tested. The Langshans of Chinese origin were very susceptible (Table 8). The stock of Rhode Island Reds obtained in Canada for use in China was found to possess much more resistance than any other Rhode Island Red stock tested. Another group of Canadian Rhode Island Reds proved to be extremely susceptible to inoculation with 6\ pullorum. These limited tests are not sufficient, however, to warrant the conclusion that all Rhode Island Reds are more susceptible than other TABLE 8. SURVIVAL OF INOCULATED CHICKS OF DIFFERENT BREEDS AND VARIE- TIES: UNSELECTED STOCKS" Breed Strain Number of hatches Number of chicks inoculated Number surviving Percent surviving 1926 White Leghorn B 5 76 39 51.3 White Leghorn D 5 462 146 31.6 Rhode Island Red E 5 508 51 10 1927 White Leghorn D 10 499 194 38.9 Rhode Island Red E 10 S16 190 36.8 Rhode Island Red F 10 113 42 37.2 1930 White Leghorn A 7 301 255 84.7 White Leghorn c 7 167 148 88.6 Rhode Island Red E 7 647 177 27.4 1931 White Leghorn c 8 285 201 70.5 Rhode Island Red E 8 495 42 8.5 Langshan 8 173 13 7.5 Chia Gi H 8 193 98 50.8 1932 White Leghorn A 5 632 500 79.1 Rhode Island Red E 5 594 322 54.2 Chia Gi H S 631 500 79.2 1933 Rhode Island Red E 7 712 226 31.7 Rhode Island Red G 7 191 28 14.7 This table includes only those tests in which there were five or more hatches in a given year, and each hatch contained chicks of the different breeds to be compared. breeds. Extensive tests of samples of breeds from various parts of of the country would be necessary to provide conclusive information on breed differences. The degree of resistance would be greatly influ- enced by the extent to which pullorum disease had been present in the stock sampled and the birds consequently subjected to natural selec- tion on the basis of their resistance to this disease. That resistance can be significantly increased by selection was demonstrated by the change produced in a strain of Rhode Island Reds used in one phase 1935} INHERITANCE OK RESISTANCE TO BACTERIAL INFECTION 479 of this study. In a strain selected for one generation the survival following inoculation was 32.8 percent among a population of 346 chicks, while among 764 controls the survival was only 6.0 percent. RESULTS OF CROSSES One of the important sources of evidence concerning the inherit- ance of characters is from crosses involving the characters to be studied. Analysis of the F^ F 2 , and back-cross generation provided information concerning the hereditary nature of resistance and sus- ceptibility. A graphic presentation of the results of the various crosses is given in Fig. 2. F,xC F,xR FIG. 2. SURVIVAL OF CONTROLS, RESISTANT, Fi, F 2 , AND BACK-CROSSES WHEN EXPOSED TO PULLORUM DISEASE BY INOCULATIONS WITH SALMONELLA PULLORUM F l Generation. The survival of the F : progeny was as high as that of the resistant parents, indicating dominance of resistance (Table 9). The characters found in wild animals have, in general, a survival value and are usually dominant in expression. It is interest- ing that in this case resistance to disease appears to be dominant. Back-Crosses. The back-crosses and Fj X F t matings gave fur- ther evidence of the hereditary nature of resistance and susceptibility to pullorum disease. From reciprocal crosses no evidence of sex- linked factors was found (Table 9). Selection Among Back-Cross and F 2 Generations. A selection for low and high resistance was made in back-cross and in F 2 genera- tions. The basis of selection was the survival of the progeny. The selections were made in 1930. In this year 13.8 percent of the progeny in the stock selected for low resistance and 85.4 percent of the progeny 480 BULLETIN No. 419 [October, TABLE 9. SURVIVAL OF CHICKS HATCHED FROM VARIOUS RESISTANT AND CON- TROL STOCKS, WHEN INOCULATED WITH S. PULLORUM Ty pe of mating Number Number Percent (R = resistant, of chicks alive at alive at C = control) inoculated 3 weeks 3 weeks From purebreds and first crosses, 1928 and 1929 Rcfcf X R9 9 . . 602 441 73.3 CdV X C9 9 852 272 31.9 Rcfcf XC99 517 405 78.3 Ccfc? 1 XR99 467 346 74.1 From Fi matings inter se and back-cross matings of resistant and control stock, 1929 only RdV X Fi 9 9 ex Rd 1 X C 9 219 142 64.4 RdV X Fi99 ex Cd 1 X R9 184 118 64.1 R99 X FidV ex Re/ 1 X C9 93 60 64.6 R9 9 X Fid'd 1 ex Co" X R9 59 31 52.5 Ccfd 1 X Fi 9 9 ex Ro" X C9 175 56 32.0 Ccfcf X Fi9 9 ex Cd 1 X R9 161 76 47.2 C 9 9 X FidV ex Rd" XC9 69 25 36.2 C9 9 X FidV ex Co" X R9 96 34 35.4 FidV ex Rd 1 XC9 XFi99ex Rd 1 X C 9 .... 193 106 54.9 Fid'd 1 ex Rd" XC9 XFi99ex Co" X R9 203 105 51.2 Ficfcf ex Co" XR9 XFi99ex Rd" X C9 194 95 48.9 Fid"d" ex Cd 1 XR9 XFi99ex Cd 1 X R9 132 70 53.0 All matings 4 216 2 382 56.5 of those selected for high resistance survived exposure to the organ- ism. The next year the survival was 8.2 percent and 69.2 percent, respectively in the two groups. The difference between the percentage survival of the two selections was 61.6 percent in 1930 and 61.0 percent in 1931 (Table 10). The fact that successful selection can be made among individuals resulting from the original cross of susceptible and resistant stocks, and that these selections perform consistently in respect to suscepti- bility and resistance is evidence of the importance of heredity in relation to this disease. Results of Matings of One Male to Both Resistant and Susceptible Females. Further evidence of the hereditary character of resistance to disease was obtained from matings of susceptible and resistant females to the same male, either susceptible or resistant. In every mating with susceptible males the progeny from the resistant dam had a higher survival than did the progeny from the susceptible dam (Table 11). The average survival of the progeny of all susceptible dams mated with susceptible males was 45.7 percent and of all resistant dams so mated 73.8 percent. Since the males were the same for both 1935] INHERITANCE OF RESISTANCE TO BACTERIAL INFECTION 481 TABLE 10. SURVIVAL OF PROGENY OF F 2 AND BACK-CROSS FEMALES SELECTED FOR HIGH AND Low RESISTANCE TO S. PULLORUM: FEMALES MATED TO SUSCEPTIBLE MALES (From the 15 females whose progeny had a survival of less than 20 percent in 1930, nine were selected for a similar test in 1931. From 20 females whose progeny had a survival of 70 percent or more the first year, ten were selected for the second year's test.) Distribution of 1930 females on basis of progeny surviving inoculation Females tested both years (Nos.) 1930 progeny from females tested both years 1931 progeny from females tested both years Number of chicks inoculated Number of chicks surviving Percent surviving Number of chicks inoculated Number of chicks surviving Percent surviving Percentage of progeny sur- viving Number of females Records of progeny of females showing low resistance 0-9.9 5 10 10 13 9 47 J 7747 6705 6919 6769 5814 6785 6915 7709 6907 25 31 28 44 34 27 19 36 38 282 2 3 3 6 5 4 3 6 7 39 8.0 9.7 10.7 13.6 14.7 14.8 15.8 16.7 18.4 13.8 36 3 32 10 28 14 18 26 29 196 7 1 1 1 2 1 2 1 16 19.4 33.3 3.1 10.0 7.1 7.1 11.1 3.4 8.2 10-19.9 20-29 .9 30-39.9 40-49 9 Total (9) Records of progeny of females showing high resistance 50-59.9... 60-69.9 70-79.9 80-89.9. ... 17 22 12 6 2 59 J 5668 5722 6775 5690 5678 7529 6686 5710 5646 5650 (10) 30 38 43 27 27 40 17 45 37 39 343 23 30 34 22 23 35 15 40 34 37 293 76.7 78.9 79.1 81.5 85.2 87.5 88.2 88.4 91.9 94.9 85.4 6 31 34 22 19 32 28 21 13 21 227 3 17 12 14 17 27 21 20 10 16 157 50.0 54.8 35.3 63.6 89.5 84.4 75.0 95.2 76.9 76.2 69.2 90-99.9 ... Total sets of dams the results indicate a genetic difference between the dams. When resistant males were used, no significant difference was found between the survival of progeny from susceptible and resistant dams, the averages being 78.5 percent and 79.8 percent respectively. Crosses of Chinese and American Strains. To ascertain whether the susceptibility to pullorum disease found in a Chinese strain of chickens was produced by the same genetic factors as susceptibility in 482 BULLETIN No. 419 [October, TABLE 11. SURVIVAL OF CHICKS FROM RESISTANT AND SUSCEPTIBLE DAMS MATED TO THE SAME SIRE Sire No. Chicks from susceptible dams Chicks from resistant dams Number inoculated Number surviving Percent surviving Number inoculated Number surviving Percent surviving Susceptible F1289 115 54 106 69 54 398 28 39 39 67 57 99 43 47 26 67 512 30 34 51 41 26 182 17 28 31 55 33 86 40 44 22 46 402 26.1 63.0 48.1 59.4 48.2 45.7 60.7 71.8 79.5 82.1 57.9 86.9 93.0 93.6 84.6 68.7 78.5 113 55 134 87 61 450 61 15 18 24 28 34 30 69 44 34 357 61 45 102 76 48 332 50 12 12 21 12 32 28 52 40 26 285 54.0 81.8 76.1 87.4 78.7 73.8 82.0 80.0 66.7 87.5 42.9 94.1 93.3 75.4 90.9 76.5 79.8 F1290 G2768 G2831 G8203 Total Resistant 70 578 . .... 1639 2133 2134 . ... 2537 5013 5546 5681 8163 Total TABLE 12. RESISTANCE OF CROSSES OF CHINESE AND AMERICAN STRAINS OF CHICKENS INOCULATED WITH S. PULLORUM Cross Number inoculated Number surviving Percent surviving Susceptifr Langshan Resistant Langshan Canadian e X Langshan 31 22 884 198 191 2 16 643 21 28 6.5 72.5 72.7 10.6 14.7 X resistant Rhode Island Red an American strain, a susceptible American strain (G), which showed a survival of 14.7 percent was crossed with the Langshan, a suscept- ible strain from China which had shown a survival of 10.6 percent. The survival of the offspring from this cross when inoculated with S. pullorum was only 6.5 percent (Table 12). If these two strains had possessed different genetic factors for susceptibility, their progeny would probably have been more resistant than either one of the strains, rather than less resistant. Evidently they possessed the same genetic factors. When the susceptible Langshan strain was crossed with a resistant Chinese strain, 72.5 percent of the progeny survived inoculation. The survival of the resistant Chinese strain was 72.7 percent. The genetic factor or factors for resistance would thus seem to be dominant. The progeny from these parental stocks and crosses were obtained during the same season but not at the same time. 79J5] INHERITANCE OF RESISTANCE TO BACTERIAL INFECTION 483 Nature of Resistance and Susceptibility The results of the various crosses just described seem to indicate that resistance to inoculation with Salmonella pullorum is dominant to susceptibility. Of more importance, however, than the mere expression of resistance and susceptibility in the transmission of dis- ease is the cause of such resistance or susceptibility. About this little is known. Pullorum disease is largely, so far as mortality is con- cerned, a disease of young chicks. Even in a susceptible strain, unless infection occurs within a very few days after hatching, the mortality is not high. According to Hanks and Rettger, 10 * the disease in young chicks appears to be a septicemia. Apparently some profound change occurs early in the life of the young chick causing it to become more resistant to the organism responsible for this disease. A preliminary note on blood studies of resistant and susceptible strains by Quisen- berry, Roberts, and Card 29 * is of interest in this connection (Tables 13 and 14). Blood samples from the progeny of susceptible Rhode Island Reds and resistant White Leghorns in three different hatches were studied 3, 6, 7, and 9 days after hatching. These birds had not been inoculated with S. pullorum. In six out of seven examinations the number of erythrocytes was greater for the resistant than it was for the suscepti- ble strain (Table 13). In every hatch the leucocyte count was lower for the resistant than for the susceptible strain. Also the percentage of neutrophiles was lower in the resistant birds in six out of seven hatches. Hutt 12 * obtained the same general results except that he found a higher erythrocyte count in Rhode Island Reds than in White Leghorns. In similar studies made on blood from chicks inoculated by mouth with 1/4 cc. of a 24-hour broth culture of S. pullorum (Table 14), the general relations among the red cells, white cells, and neutro- philes of the resistant and susceptible inoculated birds were the same as among the resistant and susceptible noninoculated chicks, but the degree of difference among the percentages of neutrophiles was changed. The neutrophilic percentage among the inoculated resistant birds was somewhat higher than among the noninoculated resistants of the same age, yet the difference was much less than that between the inoculated and noninoculated susceptible birds. The percentage of neutrophiles was high in the 3-day-old chicks and decreased with age, the 9-day-old chicks having the lowest percentage. This was true of the noninoculated susceptibles, noninoculated resistants, and inoculated resistants. Among the inoculated susceptibles the percentage of neu- 484 BULLETIN No. 419 [October, TABLE 13. NUMBER OF ERYTHROCYTES AND LEUCOCYTES AND PERCENTAGE OF POLYMORPHONUCLEAR NEUTROPHILES IN THE BLOOD OF SUSCEPT1- BLE AND RESISTANT CHICKS NOT INOCULATED WITH S. PULLORUM Hatch No. Breed Number of chicks Age in days Erythrocytes per cubic millimeter Leucocytes per cubic millimeter Percent polymor- phonuclear neutro- philes 1 Rhode Island Red 6 3 2 910 000 45 000 59.8 5 3 3 160 000 40 000 54 2 2 Rhode Island Red 6 3 2 835 000 63 000 60.7 6 3 2 945 000 37 000 53 3 3 Rhode Island Red 6 3 2 560 000 31 000 67 9 White Leghorn 6 3 3 095 000 18 000 58.2 3 Rhode Island Red 6 6 2 630 000 29 000 37.6 White Leghorn 6 6 3 035 000 16 000 35.8 1 Rhode Island Red 6 7 2 815 000 40 000 31.6 6 7 2 730 000 37 000 36 8 2 Rhode Island Red 6 7 2 630 000 29 000 43.9 White Leghorn 6 7 2 885 000 19 000 38.6 3 Rhode Island Red 6 9 2 753 000 46 000 39.0 6 9 2 905 000 17 000 29 6 TABLE 14. NUMBER OF ERYTHROCYTES AND LEUCOCYTES AND PERCENTAGE OF POLYMORPHONUCLEAR NEUTROPHILES IN THE BLOOD OF SUSCEPTI- BLE AND RESISTANT CHICKS INOCULATED WITH S. PULLORUM Hatch No. Breed Number of chicks Age in days Erythrocytes per cubic millimeter Leucocytes per cubic millimeter Percent polymor- phonuclear neutro- philes 1 Rhode Island Red. 6 3 2 845 000 41 000 62.2 5 3 3 035 000 47 000 53.4 2 Rhode Island Red 6 3 3 065 000 46 000 66.1 6 3 3 620 000 35 000 67.9 3 Rhode Island Red 6 3 2 630 000 24 000 68.3 White Leghorn 6 3 2 830 000 23 000 76.8 3 Rhode Island Red 6 6 2 480 000 30 000 52.7 6 6 2 690 000 18 000 43.7 1 Rhode Island Red . 6 7 2 565 000 75 000 62.5 White Leghorn 6 7 2 410 000 49 000 39.7 2 Rhode Island Red 6 7 2 640 000 55 000 58.5 6 7 2 685 000 41 000 37.2 3 Rhode Island Red 6 9 2 300 000 51 000 55.8 White Leghorns 6 9 2 490 000 22 000 30.7 trophiles was much higher at the ages of 6, 7, and 9 days than was the percentage among the inoculated resistants of the same ages. This higher percentage of neutrophiles in the inoculated suscepti- bles compared with the inoculated resistants suggests that percentage of neutrophiles may be used as a measure of the relative resistance of different strains of chickens to S. pullorum. INHERITANCE OF RESISTANCE TO BACTERIAL INFECTION 485 Results of Agglutination Tests In the fall of 1934, the last year of the experiments reported herein, all the birds in the strains selected for their resistance to Salmonella pidlorum were tested by the whole-blood stained-antigen method for the presence of infection with this organism. Table 15 gives the results of these tests. Of 271 birds tested, only 9 showed a positive reaction to the test. This may indicate a relatively high resistance in the selected birds. TABLE 15. REACTION OF Two RESISTANT STRAINS OF CHICKENS TO AGGLUTINATION TE'STS, 1934 Sex Age Number Reaction Positive Negative Strain I .... Female Male Female Male Female Male Female Male 1 year 1 year 4- 1 year + 1 year 1 year 1 year + 1 year + 1 year 39 5 15 2 136 16 52 6 271 3 3 3 9 36 5 12 2 136 16 49 6 262 Strain III Total At no time during these tests was any attempt made to eliminate the reactors from the flocks, for the possibility of resistant individuals giving positive reactions to the test was recognized. However, in view of the results shown in Table 15 there is no reason to believe that resistant individuals, unless they are infected, give other than negative reactions to this test. The fact that so few positives were found in the stocks selected for resistance, especially since these individuals came from ancestry that had survived inoculation, indicates that this stock did possess a high degree of resistance to pullorum disease. Was Resistance Acquired or Natural One usual explanation of resistance to a disease is that the individual at some time in its life cycle, either prenatally or post- natally, has been infected with the disease, unknown to the observer, and has thus established an immunity which may be mistaken for natural resistance. In these experiments the chicks were inoculated so short a time 486 BULLETIN No. 419 [October, after hatching that it would have been impossible for them to acquire immunity between hatching and inoculation. If they did acquire immunity, they must have acquired it during embryonic development. This would mean that the pullorum organism w r as present in the egg. It is well established that the organism of pullorum disease can be found in some eggs produced by infected hens. The resistant parental stock in this experiment was tested by means of the agglutination test. If the resistance shown by the progeny was acquired as the result of previous infection and was not a natural resistance, the offspring of reacting (infected) mothers would have shown a survival value higher than that found among offspring from nonreacting (noninfected) mothers. However,- the survival among the progeny of reactors was no greater than among the progeny of nonreactors. In one of the resistant strains 67.6 percent of 142 progeny from 13 reactors survived inoculation. Among 1,341 progeny from 92 nonreactors 70.0 percent survived. Acquired immunity could not, therefore, have been involved in the results of these experiments. BIOLOGICAL ASPECTS OF METHODS OF DISEASE CONTROL If resistance and susceptibility to disease are hereditary, the genetic improvement of any animal population is retarded by any method of disease control which prevents natural selection. If hereditary factors are involved in resistance and susceptibility, then we must conclude that present methods of disease control* tend to prevent improvement in the hereditary constitutions of our animal populations, for under present methods susceptible individuals are saved and the percentage of resistant individuals in the population is not increased. 15 *No evidence is available to show that protection from disease which results from vaccination, sanitation, or the use of serums or bacterins improves in any way the quality of succeeding generations. So far as is known, acquired characters of this kind have no effect upon the germ plasm. The importance of hereditary constitution is likewise being recognized in the problem of human health. Faust** (1934) says: "Without any disparage- ment of the splendid endeavors of men who have risked their lives to protect their fellows from disease it is time we pause to consider the final results of artificial protection alone without the fight which the body as a mechanism is prepared to wage against parasite invaders. It is obvious that natural and acquired resistance, and immunity are by far the more important to the human race. Let us recognize the interdependence and relative values of both types of defense and by rational as well as altruistic programs perpetuate the race as well as the individual. By so doing we can more surely guarantee a continuity on a higher plane of the contributions which are constantly being made to human knowledge and happiness." 1935] INHERITANCE OF RESISTANCE TO BACTERIAL INFECTION 487 Improvement in our animal populations made by increasing their resistance to disease would be very desirable from the standpoint of disease prevention. The necessity of utilizing present methods of disease control should not be minimized, but the importance of perma- nently improving the hereditary constitutions of our animals, and the necessity for so doing must be recognized as one of the urgent problems in the field of animal breeding. When man displaces natural processes which are biologically bene- ficial, it is necessary for him to replace them by others of his own creation which are also biologically beneficial unless he is to suffer from the changes which he has instituted. The outcome of these biological situations depends largely upon the intelligence which man applies to them. The three general methods of combating disease are prevention, cure, and genetic control. Which of these methods should be used under given circumstances is in part a question of which is most economical. Definite methods of cure or prevention that have already been worked out may, in some situations, be more economical than the genetic production of a resistant strain of animals. It must be remembered, however, that when methods of prevention and cure are relied upon, they must be in operation continuously unless the infecting organism can be entirely eliminated, which is unlikely with most dis- eases. The genetic method of control is more likely to be economically feasible with small animals which reproduce rapidly and have a low per-capita value than it is with the larger animals. In all probability a combination of the genetic method and the preventive and curative methods now in use will produce better results than those produced by any one method alone. It should also be remembered that any progress toward hereditary resistance will mean easier and better functioning of the usual methods of disease control. The processes by which hereditarily resistant strains of animals can be produced are so complicated and require so much time that the work of developing such animals cannot be done economically by the average animal producer. Some institution such as the agricultural experiment station must undertake it. After resistance and suscepti- bility to a given disease are established as hereditary, then the char- acter resistance must be combined with other commercially desirable characters of the breed. And finally, such animals must be subjected to the conditions under which they would have to live on the farm, before being distributed to commercial breeders. 488 BULLETIN No. 419 [October, The genetic approach to disease control is of such recent origin that time has not yet permitted the formation of resistant strains of any farm animals for distribution as breeding stock. SUMMARY AND CONCLUSIONS From this experiment, ten years in duration and involving more than 29,000 birds, evidence has been obtained which clearly indicates that heredity is an important factor in resistance and susceptibility to infection with Salmonella pullorum. The existence of hereditary factors for resistance and susceptibility to pullorum disease is shown by the following results: 1. Selection was effective in producing strains of the domestic fowl more resistant than were unselected stocks in respect to infection by Salmonella pullorum. 2. The selected stocks were consistent in maintaining resistance thru successive generations. 3. The F! generation produced by crossing resistant and susceptible stock was as resistant as the resistant parents. 4. Progeny of the F x individuals mated to resistant were signif- icantly more resistant than were the progeny of the back-cross to susceptible. 5. In the F 2 generation susceptible and resistant strains were recovered by selection. 6. A susceptible male mated to susceptible females produced progeny which were much less resistant than were progeny of the same male mated to resistant females. 7. No significant difference was found between the progeny of susceptible and resistant females mated to the same resistant male. 8. Acquired immunity was not present in the experimental birds, the progeny of infected hens exhibiting no greater resistance to dis- ease than the progeny of noninfected hens, infection and freedom from infection being determined by the agglutination test. 9. Resistance is dominant to susceptibility, but probably more than one gene is involved. 10. In an examination of the blood of noninoculated young chicks the number of erythrocytes was found to be greater in the resistant (6 out of 7 cases) than in the susceptible strain. The number of leuco- cytes was greater in the susceptible strain. The percentage of neutro- philes was lower in the resistant individuals (6 out of 7 cases). In inoculated chicks, the percentage of neutrophiles was much higher among the susceptibles at 6, 7, and 9 days than among inoculated re- sistant individuals of the same ages. 1935] INHERITANCE OF RESISTANCE TO BACTERIAL INFECTION 489 BIBLIOGRAPHY 1. ASMUNDSON, V. S., and BIELY, JACOB. Inheritance of resistance to fowl paralysis (neuro-lymphomatosis gallinarum). I. Differences in suscepti- bility. Canad. Jour. Res. 6, 171-176. 1932. 2. BECKER, E. R., and HALL, P. R. The possible role of inheritance in the quantitative character of a coccidian infection of the rat. Parasitology 25, 397-401. 1933. 3. BIELY, JACOB, PALMER, E., and ASMUNDSON, V. S. Inheritance of resistance to fowl paralysis (neuro-lymphomatosis gallinarum). II. On a significant difference in the incidence of fowl paralysis in two groups of chicks. Canad. Jour. Res. 6, 374-380. 1932. 4. CARD, L. E., and ROBERTS, ELMER. Inheritance of resistance to pullorum dis- ease. Proc. Fourth World's Poultry Cong., 488-493. 1930. 5. DUBLIN, L. I., and LOTKA, A. J. The history of longevity in the United States. Human Biol. 6, 43-86. 1934. 6. FAUST, E. C. The human body's defense against disease. Sigma Xi Quart. 22, 53-62. 1934. 7. FORSYTH, C. H. Decline in the average length of life. Human Biol. 2, 199- 223. 1930. 8. FRATEUR, J. L. The hereditary resistance of the fowl to the bacillus of diphtheria. Proc. Second World's Poultry Cong., 68-71. 1924. 9. GOWEN, J. W., and SCHOTT, R. G. Genetic predisposition to Bacillus pili- f or mis infection among mice. Jour. Hyg. [London] 33, 370-378. 1933. 10. HANKS, J. H., and RETTGER, L. F. Bacterial endotoxin. Search for specific intracellular toxin in 5". pullorum. Jour. Immunol. 22, 283-314. 1932. 11. HUFF, C. G. The inheritance of natural immunity to Plasmodium cathe- meriurn in two species of Culex. Jour. Prev. Med. 5, 249-259. 1931. 12. HUTT, F. B. On the physiological basis of genetic resistance to Salmonella pullorum in the fowl. Amer. Nat. 69, 66-67. 1935. 13. IRWIN, M. R. Inheritance as a factor in resistance to an infectious disease. Jour. Immunol. 24, 285-342. 1933. 14. - The inheritance of resistance to the Danysz bacillus in the rat. Iowa State Col. Jour. Sci. 2, 213-218. 1928. 15. and HUGHES, T. P. Inheritance as a factor in resistance to an infectious disease. VI. The correlation between resistance and the bac- tericidal power of the whole blood. Jour. Immunol. 24, 343-348. 1933. 16. LAMBERT, W. V. Mortality in chickens following the feeding of massive doses of irrident fowl typhoid bacteria. Jour. Amer. Vet. Med. Assoc. 73, n.s. 26, 480-483. 1928. 17. - - Natural resistance to disease in the chicken. Jour. Immunol. 23, 229-260. 1932. 18. - and KNOX, C. W. The inheritance of resistance to fowl typhoid in chickens with reference to its inheritance. Iowa Agr. Exp. Sta. Res. Bui. 153, 262-295. 1932. 19. - The inheritance of resistance to fowl typhoid in chickens. Iowa State Col. Jour. Sci. 2, 179-187. 1928. 20. MACDOWELL, E. C., and RICHTER, M. N. Studies on mouse leukemia. II. Hereditary susceptibility to inoculated leukemia. Jour. Cancer Res. 14, 434-439. 1930. 21. - - Studies on mouse leukemia. IV. Specificity of susceptibility to different lines of inoculated leukemia. Soc. Exp. Biol. and Med. Proc. 28, 1012-1013. 1931. 22. - Studies on mouse leukemia. V. A genetic analysis of susceptibility to inoculated leukemia of line I. Biol. Zentbl. 52, 266-279. 1932. 490 BULLETIN No. 419 23. MANRESA, MIGUEL. Inheritance of resistance and susceptibility to infectious abortion. Jour. Infect. Diseases 51, 30-71. 1932. 24. ONO, TADANORI. Study on inheritance of immunity. Japan. Jour. Exp. Med. 10, 265-390. 1932. 25. OSSENT, H. P. Ein seuche-immunes wildfarbiges Hausschwein. Der Zuchter 4, 152-160. 1932. 26. PRITCHETT, I. W. Microbic virulence and host susceptibility in paratypho- identeritidis infection of white mice. VI. The relative susceptibility of different strains of mice to per os infection with the Type II bacillus of mouse typhoid (Bacillus pestis caviae). Jour. Exp. Med. 41, 195-208. 1925. 27. Microbic virulence and host susceptibility in paratyphoid-enteri- tidis infection of white mice. X. The relative susceptibility of different strains of mice to per os infection with the Type II bacillus of mouse typhoid (Bacillus pestis caviae). Jour. Exp. Med. 43, 161-171. 1926. 28. Microbic virulence and host susceptibility in paratyphoid-enteri- tidis infection of white mice. Further studies. Jour. Exp. Med. 46, 557- 570. 1927. 29. QUISENBERRY, J. H., ROBERTS, ELMER, and CARD, L. E. Blood studies of strains of the domestic fowl, differing in resistance to pullorum disease. Poultry Sci. 14, 63-64. 1935. 30. ROBERTS, ELMER. Inheritance of resistance to disease. Amer. Soc. Anim. Prod. Proc., 50-53. 1925. 31. Inheritance of resistance to bacterial infection. Sixth Internatl. Cong. Genetics Proc. 2, 276. 1932. 32. - Inheritance of resistance to disease in animals. Sixth Internatl. Cong. Genetics Proc. 2, 169-170. 1932. 33. and CARD, L. E. Resistance of chicks to bacillary white diarrhea. Amer. Soc. Anim. Prod. Proc., 113-115. 1924. 34. - The inheritance of resistance to bacillary white diarrhea. Poultry Sci. 6, 18-23. 1926. 35. Genetic studies on resistance to disease. Anat. Rec. 37, 175. 1927. 36. - - Further data on inheritance of resistance to dis- ease. Amer. Soc. Anim. Prod. Proc., 201-203. 1926. 37. - Genetic studies on resistance to bacterial infec- tion. Jour. Bact. 13, 31. 1927. 38. - - Further studies on the inheritance of resistance to bacterial infection. Anat. Rec. 51, 118. 1931. 39. SCHOTT, R. G. Preliminary studies of hereditary resistance in mice to a specific bacterial infection. Anat. Rec. 44, 283. 1929. 40. - The inheritance of resistance to Salmonella aertrycke in various strains of mice. Genetics 17, 203-229. 1932. 41. WEBSTER, L. T. Inherited and acquired factors in resistance to infection. .1. Development of resistant and susceptible lines of mice through selective breeding. Jour. Exp. Med. 57, 793-817. 1933. 42. - - Inherited and acquired factors in resistance to infection. II. A comparison of mice inherently resistant or susceptible to Bacillus en- teritidis infections with respect to fertility, weight, and susceptibility to various routes and types of infection. Jour. Exp. Med. 57, 819-843. 1933. 43. WRIGHT, SEWALL, and LEWIS, P. A. Factors in the resistance of guinea-pigs to tuberculosis, with especial regard to inbreeding and heredity. Amer. Nat. 55, 20-50. 1921. 44. Heredity of resistance to tuberculosis in guinea- pigs. Anat. Rec. 23, 93. 1922. APPENDIX TABLE 16. SURVIVAL OF CHICKS FROM SELECTED ANd CONTROL STOCKS AFTER INOCULATION WITH S. PULLORUM (All hatches with 15 or more chicks in each of the selected and control groups are included) Strain and i selected stock Control stock date of hatch Number inoculated Number surviving Percent surviving Number inoculated Number surviving Percent surviving Strain I 5-12-26... 51 22 43.1 65 1 1.5 5-19 51 29 56 9 74 5 6 8 5-26 58 25 43.1 187 11 5.8 6-2 70 58 82.9 133 28 21.0 6-9 60 25 41.7 47 6 12.2 6-16 60 37 61.7 88 10 11.4 3-9-27 33 27 81.8 36 10 27.8 3-16 35 16 45.7 34 4 11.8 3-23 60 33 55 15 2 13 3 3-30 94 50 53 2 32 4 12 5 4-6 133 88 66.2 64 9 14.1 4-13 53 32 60.4 124 57 46.0 4-20 111 68 61.3 72 25 34 7 4-27 134 103 76.9 92 48 52.2 5-4 138 118 85.5 79 49 62.0 5-11 185 140 75.7 81 24 29.6 5-18 126 89 70.6 62 48 77.4 5-25 137 99 72.3 54 33 61.1 6-1 144 111 77.1 61 34 55.7 6-8 159 110 69.2 75 45 60.0 6-15 75 58 77.3 57 36 63.2 4-25-28 25 17 68.0 28 15 53.6 5-2 33 29 87.9 18 10 55.6 5-9 . 34 27 79 4 19 10 52.6 5-16 61 48 78.7 43 22 51.2 5-23 60 56 93.3 38 24 63.2 5-30 . . 63 53 84.1 62 31 50.0 6-6 48 40 83.3 51 28 54.9 4-24-29... 29 18 62.1 59 7 11.9 5-1 43 19 44 2 18 3 16.7 5-8 34 15 44.1 17 6 35.3 5-15 23 18 78.3 68 23 33.8 5-22 24 8 33.3 180 34 18.9 5-29 29 16 55.2 139 30 21.6 3-20-30 24 17 70.8 100 13 13.0 3-27 41 31 75.6 77 23 29.9 4-3 45 33 73.3 112 34 30.4 4-10 46 40 87.0 68 13 19.1 4-17 36 32 88.9 118 30 25.4 4-24 45 44 97.8 82 44 53.7 5-1 30 24 80.0 90 20 22.2 5-8 30 24 80.0 91 51 56.0 5-15 54 50 92.6 125 96 76.8 5-22 65 47 72.3 116 45 38.8 5-29 54 47 87.0 123 57 46.3 Strain II 3-13-30... 48 35 72.9 86 6 7.0 3-20. . 60 46 76.7 100 13 13.0 3-27 71 55 77.5 77 23 29.9 4-3 87 69 79.3 112 34 30.4 4-10 83 81 97.6 68 13 19.1 4-17 78 64 82.1 118 30 25.4 4-24 82 75 91.5 82 44 53.7 5-1 73 56 76.7 90 20 22.2 (Continued on page 492) 491 492 BULLETIN No. 419 [October, TABLE 16. Continued Strain and c selected stock Control stock date of hatch Number inoculated Number surviving Percent surviving Number inoculated Number surviving Percent surviving Strain III 3-20-30 50 42 84.0 100 13 13.0 3-27 44 38 86.4 77 23 29.9 4-3 45 37 82.2 112 34 30.4 4-10 39 35 89.7 68 13 19.1 4-17 48 45 93.7 118 30 25.4 4-24 35 31 88.6 82 44 53.7 5-1 40 27 67.5 90 20 22.2 Strain I 4-22-31 33 19 57.6 61 1 1.6 4-29 30 7 23.3 54 3 5.6 5-6 39 14 35.9 77 2 2.6 5-13 48 29 60.4 57 2 3.5 5-20 53 32 62.3 54 1 1.9 5-27 : 42 34 81.0 49 4 8.2 6-3 34 27 79.4 66 10 15.2 6-10 26 20 76.9 73 5 6.8 6-17 40 38 95.0 65 15 23.1 Strain 11 4-1-31 46 20 43.5 61 2. 3.3 4-8 37 11 29.7 65 0.0 4-15 57 9 15.8 74 2 2.7 4-22 72 26 36.1 61 1 1.6 4-29 78 40 51.3 54 3 5.6 5-6 86 20 23.3 77 2 2.6 5-13 118 48 40.7 57 2 3.5 5-20 . . . 114 77 67.5 54 1 1.9 5-27 96 62 64.6 49 4 8.2 6-3 94 68 72.3 66 10 15.2 6-10 91 69 75.8 73 5 6.8 6-17 84 77 91.7 65 15 23.1 Strain III 4-1-31 39 15 38.5 61 2 3.3 4-8 31 21 67.7 65 0.0 4-15 41 16 39.0 74 2 2.7 4-22 47 24 51.0 61 1 1.6 4-29 48 26 54.2 54 3 5.6 5-6 48 16 33.3 77 2 2.6 5-13 68 43 63.2 57 2 3.5 5-20 68 49 72.1 54 1 1.9 5-27 49 39 79.6 49 4 8.2 6-3 43 36 83.7 66 10 15.2 6-10 36 32 88.9 73 5 6.8 6-17 57 49 86.0 65 15 23.1 1935] INHERITANCE OF RESISTANCE TO BACTERIAL INFECTION 493 TABLE 16. Concluded Strain and Selected stock ( Control stock date of hatch Number inoculated Number surviving Percent surviving Number inoculated Number surviving Percent surviving Strain II 5-18-32 41 41 100.0 143 98 68.5 5-25 . . 63 40 63.5 116 25 21.6 6-1 70 57 81.4 117 42 35.9 6-8 67 56 83.6 114 83 72.8 6-15 56 47 83.9 104 74 71.2 Strain III 5-18-32... 44 42 95.5 143 98 68.5 5-25 . . 50 40 80.0 116 25 21.6 6-1 44 33 75.0 117 42 35.9 6-8 . 48 45 93.8 114 83 72.8 6-15 . 30 26 86.7 104 74 71.2 Strain I 5-3-33... 47 30 63.8 81 2 2.5 5-10 . . 41 31 75.6 95 34 35.8 5-17 41 27 65.9 129 39 30.2 5-24 34 27 79.4 104 36 34.6 5-31 35 32 91.4 99 39 39.4 6-7 34 23 67.6 125 33 26.4 6-14 26 16 80.0 79 43 54.4 Strain II 5-3-33... 98 61 62.2 81 2 2.5 5-10 113 66 58.4 95 34 35.8 5-17 113 75 66.4 129 39 30.2 5-24 101 80 79.2 104 36 34.6 5-31 79 73 92.4 99 39 39.4 6-7 68 56 82.4 125 33 26.4 6-14 48 43 89.6 79 43 54.4 Strain III 5-3-33 81 59 72.8 81 2 2.5 5-10 63 47 74.0 95 34 35.8 5-17 74 59 79.7 129 39 30.2 5-24 57 47 82.5 104 36 34.6 5-31 57 51 89.5 99 39 39.4 6-7 39 30 76.9 125 33 26.4 6-14 22 20 90.9 79 43 54.4 Strain II X Strain III 5-9-34... 90 73 81.1 37 7 18.9 5-16 . . . 154 138 89.6 22 3 13.6 5-23 149 126 84.6 32 7 21.9 5-30 147 133 90.5 32 8 25.0 10-355,0508468