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The Artificial Immunisation of Cattle Against Tuberculosis.*
4
@riginal thticles. THE
ARTIFICIAL IMM U NISATION OF CATTLE AGAINST TUBERCULOSIS. ~' BY LEONARD PEARSON AN D S. H. G ILLILAND.
(Frollt the Laboratory of tlu State Live Stock Sa?,itary Boa" d
of Peltllsy lva,tia.)
A Pa per presented to th e Forty ·second Annual Meeting o f the American Veterinary Medical Association, at Cleveland, Ohi o, August 15-18, 1905. EFFORTS to produce artificial immunity against tuberculosis have been conducted for a number of .years a nd along ma ny lines. In 1890 Robert Koch announced the discovery of tuberculin . It was claimed at that time that by the u se of tuberc ulin resistance to tuberculosis could be increased in such a way as to assist in withstanding infec tion , and also that som e infections already established might under the use of tuberc ulin b e o vercome. But extensive experim enta tion has shown that these effects are not sufficiently uniform, lastin g or powerful to be of distinct practical value. Following this pioneer work of Koch came the wo rk of many investigators with modified tuberculin s and with extracts from tubercle bacilli made in a variety of w ays. Repeated ex periments with all these substa nces h ave shown that it has not been possible by their use to rende r a nimal s immune to tuberculosis perm a nently, or to a practical degree. Experiments have been made by severa l invest igators to test the im munising value of dead tubercle ba cilli a nd of bacillary pulp. The results have been a bout the same as have followed the use of extracts from tu bercle ba cilli . It became evi den t years ago that immunity aga inst tuberculosis, to be of value, must confer protection against the organism of tuberculosis as well as against its toxins. There must be bac terial as well as toxic immunity. Efforts to immuni se animals again st living and virulent tubercle bacilli by inoc ulatin g th em with living tubercl e bacilli of low virulence, were m ade as early as 1891 by Granchez and L edou xLebard. In 1892 and 1893 Trudeau t found that by inoculating rab bits subcutaneously with living cultures of avian tubercle bacilli, he was able to increase th eir resista nce to infection from mammalian tu bercle bacilli of a culture known to be virulent for ra bbits. D e Schweinitz,:;: in 1894, di scovered that it was possible to very greatly increase the resistance of g uin ea-pigs to inocu lati ons of tu bercul osis by inoculatin g them with at tenua ted tub ercle bacilli of hum a n orig in; the process of a ttenuation consi sting in prolonged culti va tion (twenty generations) on g lycerin beef broth of acid reac tion . Tubercle bacilli of hum a n origin g rown in this way lost th eir vi rttlence for gui nea-pigs, but g uinea-pigs inoculated w ith this non -virul ent culture developed so much resistance to tuberculosis, th a t when th ey were a ft erwards inoculated with tuberculous tissue frol11 a cow they remained hea lthy, while other g uinea pigs inoculated with the same material fr om the cow died of tuber-
• From the America" Veterillary Review. t New York }II[eJicaIJoltrual, J uly 23, 1893. : Medical N elliS, New Yo rk , December 8, 1804 .
A rtijic/al I mmunisatio1t
of Cattle.
5
culosis in seven weeks. De Schweinitz injected very large quantities of human tubercle bacilli into ca ttle subcutaneously, intravenously and intraperitoneally. He found that by gradually increasin g the dosage enormou s quantities could be tolerated without injury. He administered as much as 500 c. c. of a heavy suspension of tubercle bacilli at one time . McFadyean, ~' in I g0 1 and 1902, reported that he had found that the r esistance of cattle to tuberculosis may be very greatly increased by the use of successive inocula tions with tuberculous m a terial or tu bercle cultures of low virulence. In a paper of McFadyean's en titled" Further Experiments Regardin g the Immunisation of Cattle against Tuberculosis," t the author states this conclusion: " It appears to be justifiable to conclude that, whatever may have been the degree o f natural immunity possessed by these three experimental animals, it was much in creased by the successive intravenou s inoculations to which they were subjected. Th e immunity was not ab solute, but it may be doubted wh ether a degree of resistance that would merit that term is obtainable by any method in cattle." von Behring announced in December, in I g OI, that he was engaged in studying the immunisation of cattle against tuberculosi s, and he has since issued several report s upon his work. von Behring has made a very larg e number of experiments in this field ; he has endeavoured to produce immunity by the use of tuberculin s , by the use of other tuberculosis to xins, by anti· toxins, by the use of dead tubercle bacilli, of tubercle bacilli weakened by chemical agents, and by the use of tubercle cultures of low virulence. A method for the vaccination of cattle against tubercu losis ha s been formu lated by von Behring, and it is ba sed on the u se of tubercle bacilli of human origin th at are nonvirulent for cattle . In makin g this" vaccine" the tubercle bacilli a re dried and ground to a powde r. The vaccine ma terial is sent out in this dried sta te. It is necessary to make a suspension of it in normal salt solution, whereupon it may be administered by intravenous injection . A number of investigators, as Lorenz, Schlegel, Eber and Hutyra , have tested the resi stance of cattle treat ed by von Behring, or according to von Behring's m ethod. It has been shown that tbe resistance of these a nimal s to tuberculosis has, in most cases, been increased and sometimes considerably so. lt is to be observed, however, that many of the cattle reported upon by the above authors were not vaccinated according to the method that von Behring now recommends, a nd as Eber says : " . .. The tuberculin test is not a reliable means for determining the freedom of tuberculosis of an animal that has been treated with mild bovine or human tubercle bacilli unle ss a long time (generally more than a ha lf year) has elapsed since th e las t administration of in fectious material. . . . Sin ce neither of the treated cattle was vaccinated bv the two-vaccination method with attenuated human tubercle bacilli, according to the method now recomm ended by von Behring, th e conclusions that are reached as a result of my investigations have only a re lative bea rin g on an estimation of the value of the method now recommended by von Behring. " The results of the investi gations show, however, that it is possible .. J ournal of ComjJarative Pat/lOlogy and ThemjJeutics, June,
t ibid. , March,
1902 .
19 01,
and March,
1902.
6
The Veterilzary
'.1 ournal.
to confer upon cattle, by treating them with attenuated bovine ·o r human tubercle bacilli, a certain degree of resistance to artificial tuberculosis infection . "Whether this resistance, as produced by the two-vaccination method, with attenuated human tubercle bacilli, that is now in use, furnishes protection against natural infection, can on ly be determined by years of careful observation and the greatest possible number of animals immunised in this way, and controls at the ·time of slaug h ter. " Hutyra ~' has reported some work conducted by him in 1903 and 1904 at the Royal Veterinary College of Hungary, wherein he tested the immunity of calves vaccinated according to von Behring's method, with material furnished by yon Behring, and also with material prepared from cultures made by himself. Hutyra tested the resistance of his .vaccinated animals by inoculating -them with v irulent bovine tubercle bacilli, and their resistance was compared to that of unvaccinated animals. He found that nearly all of the vaccinated animals had more resistance than the unvaccinated animals. One animal treated with von Behring's vaccine (No.3) appeared to have received no immunity as a result of vaccination. Hutyra's own vaccine material from different sources appeared to give a noticeably higher degree .of immunity than that obtained from von Behring's. Recently, a large number of cattle, amounting to several thousand, have been vaccinated after the von Behring method in Germany, Austria and Hungary. The process does not appear to be free from danger in every instance, as is shown by the reports of Marks, Casper and others. It is as yet impossible to draw any conclusions whatever as to the efficacy of vaccinations from the results of this great number of vaccinations of cattle on farms. The only conclusion at which one may fairly arrive is that vaccination appears, in most cases, to be unattended by dan ger to the vaccinated animals. Whether immunity is conferred, and, if so, whether it is sufficiently powerful or lasting for practical purposes, is not yet shown by the practical application of von Behring's method. Of the animals that have been vaccinated on farms and afterwards exposed to infection, a number have been slaughtered and some have been found to be afflicted with tuberculosis, other.s have been found to be free from tuberculosis. The presence of lesions of tuberculosis is taken by some of von Behring's observers to mean that the animal was tubercular before it was vaccinated. Of course, this may be true. But it must be remembered that the absence of lesions of tuberculosis in vaccinated animals cannot fairly be taken to signify the existence of a serviceable degree of immunity, unless it is shown that the conditions under which these animals were kept w.er.e such as to lead to the infection of a considerable. proportion of unvaccinated animals; and this evidence is in most cases lacking. Klimmer has carried out some very useful investigations upon the immunisation of cattle against tuberculosis at the Royal Veterinary College at Dresden. t Klimmer protests againg sending out vaccine material in the dry, powdered state, on account of the great danger to the operator who attempts, in the field, to prepare this material for use. Klimmer's practice is to prepare suspensions of tubercle bacilli in the • Zeitschrift fur Thiermedicin, Band ix., Heft 3.-4 , 190 5. t Berliner Tierarzliche Wochensch,.ijt, July 5. 1905·
Artificial immunisation of Cattle.
7
laboratory, just as has been done for a number of years at the laboratory of the P ennsylvania State Live Stock Sanitary Boa rd. Th e vaccine material that has been prepared in Dresden appears to be harmless to the vaccinated animals, and indications thus far are t hat it confers a serviceable degree of immunity. but it is not yet possible to draw final co nclusions from thi s work for th e reason that the vaccin ated a nimals are still living, and it is im possible to determine posi ti vely whether th ey are free from tuberc ulosis. Klimmer proposes the use of a vaccine material made o f mammalian tubercle bacilli that have been rendered less virulent by passage throug h a cold-blooded animal. The advantage from such a c ulture wo uld lie in the total absence of danger to the operator. Work upon the vaccination of cattle again st tuberculosis at the la boratory of th e State Live Stock Sanitary Board of Pennsylvania co mm enced in the yea r I gO I. It h as, therefore , been under way for abo ut five years. We were a ble to show three years ago* that it was possible to in crease the resistance of animals to tuberculosis to a very high degree by treating them with several intrave nou s inoculation s of non-virulent tubercl e bacilli of hum a n type. Two cattle thus trea ted were inoculated intra tracheally with a suspension of bovine tubercle bacilli , and both of th e m wholly resisted the inoculation. Th e only lesions in either of th ese animals was a slig ht thickening upon the wall of the trachea at the place where th e hypodermic needle was insert ed. Two unvaccin a ted animals, inoculated a t the same time, in the sa me way, with th e same quantity of bovine tubercle bacilli from the same culture, became exten siv ely infected with tuberculos is, showing the lesions on the mucous membrane of the trachea and bronchi, in the lun gs, and in tbe bronchial, mediastina l, and postphary ngea l lymphatic g lands. The immunity that was obtained in th e case of th ese two animals res ulted from th e a dmini stration of seven intravenous injections of vaccin e m ateria l during a period of t en weeks. The doses of tubercle bacilli ran ged from 13 to 26 mg. While this ex periment, and other simila r ex periments, were sufficient to lead liS to th e conclusion tha t anim a ls may be rendered immune to tuberculosis , it was at the same ti me evident th a t a process depending upon the application of vaccine material seven different times, would be of co mpa ratively small value in ge neral practice. The refore we set oursel ves to the stud y of the effects of other methods of vaccination, and to the development of a simple and effec tive m ethod. During the past th ree years we have a ttempted to determine tbe immunising effects of severa l strains o f tu bercle bacilli. We have administered vaccines in di ffere nt doses, at long a nd short interval s, a nd w ith varyin g numbers of doses. For these experim ents we have been supplied by the State Li ve S tock Sanitary Board with a large number of animals. For two years we h ave had the use of a farm where about one hundred ca ttl e have been kept for experim ental purpose s. We have also had the use of stables a t the Veterinary Departm e nt of the U ni versity of Pen nsylvania , where from twe nty to tw enty·five cattle have been kept , toge ther with numerous goats and small labora tory animals. Throug h th e use of the farm we have been enabled to keep a considerable number of cattle under practic al farm conditions. * Philadelphia lIfedical J our1lal , November
29, 1902.
The Veterinary! ournal. During the time that they have been under observation in our experiments the resistance to tuberculosis of vaccinated animals has been tested through exposure to natural infection and by inoculation. When vaccinated animals have been exposed to infection, and when they have been inoculated, unvaccinated control animals have been equally exposed or inoculated. We regard this use of controls of the highest importance, because it is only in this way that a definite standard for comparison can be provided in order to measure the extent of resistance to a given amount of exposure. Moreover, it is only by the use of controls tqat one can ascertain whether exposure under natural causes has been sufficient to lead to the infection of unvaccinated animals. It is not enough to associate vaccinated cattle with tuberculosis cattle, and then to conclude that the vaccinated cattle were immune, because, when they were killed, they did not show lesions of tuberculosis. It might readily happen that the exposure was not sufficient to lead to the infection of unvaccinated cattle; but if, upon equal ex posure, we find that the unvaccinated cattle are tuberculous, and the vaccinated cattle are free from tuberculosis, then we may ~afely and justly conclude that the vaccinated cattle received immunity from their vaccination. Our proof is based upon such clear evidence as this, frequently repeated. Our experiments have shown that different strains of tubercle bacilli hav e different immunisin g values. Avian tubercle bacilli may produce an intoxication resulting in great emaciation, but avian cultures do not appear to confer upon animals so much immunity as results from the use of mammalian cultures that are non- virulent for the animals upon which they are used. In order to produce artificial immunity in mamm a ls it appears to be necessary to vaccinate with an attenuated mammalian culture. We have not found any advantag e either in respect to th e degree of immunity or economy of time in the use of several vaccines made from cultures of different and progressive degrees of virulence . The amount of immunity bears a rather definite proportion to the number of vaccinations and the amount of vaccine material used, provided, however, that the animal be not "over-vaccinated"; that is, that th e vaccinations are not too close to one another, and that an excessive amount of vaccine is not administered. By over-vaccination the resistance of an animal to tuberculosis may be [f~duced to a point below normal, or a fatal toxcemia may be caused. Experiments have been made wherein animals vaccinated with different doses and different numbers of doses have been inocula t ed at the same time, and in the same way. While the resistance of all of the vaccinated animals to the inoculation has been much greater than the resistance of unvaccinated animals, it has been possible to observe a marked differen ce between animals vaccinated in different ways . For example, an animal vaccinated five times has more resistance than an animal vaccinated four times, and, other conditions being equal, an animal vaccinated four times has more resi s tance than an animal vaccinated three times. From the practical standpoint, it is more important to ascertain the amount of vaccination that is n ecessary to protect animals under customary conditions of exposure and the shortest possible time during
A rfi/icial i11tJltUl'lisafz·01l
0/ Cattle.
9
wh ic h a s ufficient degree of immunity may b e conferred, than it is to d ete rmin e h ow g rea t a deg ree of immunity may be produced. Since it has been necessary to try a g reat number of different m ethods of vacci na tion, and since the tim e required for an ex periment of thi s sort is mu ch g rea ter than is usual in experiments conducted through labora tories of hygie ne, it has b ee n difficult and time-consumin g to a cc umlllat e evidence upon which to base a genera l method for t he vaccination of ca ttle . It is a lread y clearly evident that different deg rees of immunity, a nd serviceable degrees of immunity, may be obtained at will. Under nat ura l conditi ons, animals differ in respect to their inh erited or acquired r esis tan ce to tube rc ulosis. Conditions of exposu re differ, bein g very mu ch greater upon some farms and in some h erds than in o th e rs. Th erefore it seems to be reasonable to conc lude th at animals will require more artificial immunity to tu bercul os is un de r some conditions than un der othe rs . The case is s imil a r to th e protection of cattle again st a n t hrax by vaccinat io n. In Pe nn sylvania we have fo und by ex perien ce , extending over a series of years, that the imm unity conferred by vacc inatin g with th e first and second vaccine of Past eur usually is suffi cie nt. On a few farm s, how e ver, the exposure a ppears to be 1l 10re inten se , a nd it is fou nd th a t on s uch farms it is n ece ssary to u se the third vacci ne o f Pa st eur. Ex p erience may show that it may be necessary to vaccinate different con dit io ns in slightly different ways . The amou nt of immun ity that is confe rred by three vaccinations is rat h e r high, a ncj will probably be sufficient in most cases. Indeed, two vaccinations may s upply a suffic ien t a m ou nt of immunity under s om e cond itio ns. The subject of the durat ion of immunity is ver y important, and is o ne th at can be solved on ly by observations upon a large series of vaccin ated ca ttl e, some of which ma y be killed and examin ed post-mortem fro m time t o time during a t erm of years. O f cou r se it is necessary that animals k ept for this purpo se should con stant ly be ex posed to infec tion, and vaccin ated controls must be simil a rly exposed, and the post-lIlortem results must s how that th e amount of exposure was sufficient to cau se the infection of the unvaccinat ed a nim a ls . Our own ex pe riments indicate that artificial immunity m a y endure a t leas t t wo years, and th ere is eve ry r eason to expect that a dditional ex periments will show that immunity will be of lo nger duration than this. Perh aps it may la st throughout th e entire life of th e a nimal ; but conclu s ion s on this point a re entirely pre mature. The effect of tuberculosis vacci nati on u pon ca ttle a lready infected w ith tubercu losis has also recei ved our attention';' ; it h as been found tha t intra v enous injections of tllbercl e bacilli of hum a n type, nonv ir ul ent for ca ttl e, have a very marked influence not o n Iy in restraining th e progres s of e xisting lesions but also in causing them to become e ncaps uled and to recede. In o th e r wo rds, such treatment a ppears to have a marked curative effec t on some tuberc ulou s an imal s. B ut this sta te ment sho uld be accompanied by a n ote of warning. 'vVe do not for a moment wish to imply tha t it is pra cticabl e , or eve n possible, a t this tim e , to c ure tubercu lous cat tIe. The observations to be made • Uu iversity 0/ FCII1IS) llvall ia llfedical Bulletill . . \pril, 19 :>5. R eview , J 905.
Amcricall Veterilla,J'
10
The Veterinary
'.1 oUr1zal.
in this direction are ' regarded by us as of importance chiefly as tending to throw additi onal lig ht on the subject of immunity. It was shown tbat the treatment give n had the same immunising effect upon infected animals as upon hea lthy animals. The effect upon the immunisa tion in these cases was, first, to prevent the further dissemination of infection, and, second , to ca use the encapsulation and elimination of the existing lesions. It is ' probable that this method of treatm en t may prove to be of value with relation to young animals, or animals known to be but recently infec ted with tubercul osis . Such treatment will always have to be carried out with the most ex treme care and under co ndition s that can be very fully controlled . It is not to be recommended for general prac tice. As to the applicati on of vaccination against tuberculosi s: vaccination, properly applied, is effective, and we believe that it will prove to be of very g rea t practical va lue. It is not poss ible on most American farms to establish a sepa rate herd of infected cattl e, an d to keep the infected cattl e as separate and di st inct from the uninfected cattle as is necessary, if tubercul osis is to be treated according to the Bang system. The alternative, if tube rculosis is to be eradicated, is to destroy all of th e anim als that react to the tubercul in test. Th e latter plan involves a grea t deal of waste and loss, which he ret ofore has been unavoidable , and which has been amply justified because it has been less than that resulting from th e continuat ion o f the in fec tion in the herd . But it l S im por tant that a better, a less expensive, and, if possible , a more effec tive method shall be devised. It appears to be hig hly probable that vacc in a tion may supply this method. It is a dmittedly impossibl e to test with tuberculin all of th e tubercu lous herds of a state. Therefore, it is not possible to discover a nd to eith er destroy or sequester all of tbe cattle th a t m ay be dis tributers of tuberculosis. Herds freed from tuberculosis by the use of the tuberculin test are always exposed to reinfection, and must be g uarded by the double testin g of all r ecruits and by systematic reinspections of tbe herd. All of this may be, and is, carried through successfull y , and sho uld be practiced to a much greater extent than it is, unless a n equally effecti ve and c heape r method can be devised. It will be a grea t step in advance to be able, not only to discover that an animal is free from tuberculosi s, but also t o r ender that ' animal permanently immune to infection . It will be a g reat advantage to be able so t o treat yo un g cattle that they may resist tuberculosis even thoug h they are continu a lly ex posed. Th e advantage to the breeder of being able, not only to g row a herd free from tuberculosis, but immun e to tu berc ul osis, will be very grea t. We believe that all of this may be looked forward to; but until more experiments and obser vations are completed, vaccination should be applied only under such conditions a s will permit exact observations to be made, and each vaccination must, for the present, be regarded as a scientific experiment.
@riginal thticles. THE
ARTIFICIAL IMM U NISATION OF CATTLE AGAINST TUBERCULOSIS. ~' BY LEONARD PEARSON AN D S. H. G ILLILAND.
(Frollt the Laboratory of tlu State Live Stock Sa?,itary Boa" d
of Peltllsy lva,tia.)
A Pa per presented to th e Forty ·second Annual Meeting o f the American Veterinary Medical Association, at Cleveland, Ohi o, August 15-18, 1905. EFFORTS to produce artificial immunity against tuberculosis have been conducted for a number of .years a nd along ma ny lines. In 1890 Robert Koch announced the discovery of tuberculin . It was claimed at that time that by the u se of tuberc ulin resistance to tuberculosis could be increased in such a way as to assist in withstanding infec tion , and also that som e infections already established might under the use of tuberc ulin b e o vercome. But extensive experim enta tion has shown that these effects are not sufficiently uniform, lastin g or powerful to be of distinct practical value. Following this pioneer work of Koch came the wo rk of many investigators with modified tuberculin s and with extracts from tubercle bacilli made in a variety of w ays. Repeated ex periments with all these substa nces h ave shown that it has not been possible by their use to rende r a nimal s immune to tuberculosis perm a nently, or to a practical degree. Experiments have been made by severa l invest igators to test the im munising value of dead tubercle ba cilli a nd of bacillary pulp. The results have been a bout the same as have followed the use of extracts from tu bercle ba cilli . It became evi den t years ago that immunity aga inst tuberculosis, to be of value, must confer protection against the organism of tuberculosis as well as against its toxins. There must be bac terial as well as toxic immunity. Efforts to immuni se animals again st living and virulent tubercle bacilli by inoc ulatin g th em with living tubercl e bacilli of low virulence, were m ade as early as 1891 by Granchez and L edou xLebard. In 1892 and 1893 Trudeau t found that by inoculating rab bits subcutaneously with living cultures of avian tubercle bacilli, he was able to increase th eir resista nce to infection from mammalian tu bercle bacilli of a culture known to be virulent for ra bbits. D e Schweinitz,:;: in 1894, di scovered that it was possible to very greatly increase the resistance of g uin ea-pigs to inocu lati ons of tu bercul osis by inoculatin g them with at tenua ted tub ercle bacilli of hum a n orig in; the process of a ttenuation consi sting in prolonged culti va tion (twenty generations) on g lycerin beef broth of acid reac tion . Tubercle bacilli of hum a n origin g rown in this way lost th eir vi rttlence for gui nea-pigs, but g uinea-pigs inoculated w ith this non -virul ent culture developed so much resistance to tuberculosis, th a t when th ey were a ft erwards inoculated with tuberculous tissue frol11 a cow they remained hea lthy, while other g uinea pigs inoculated with the same material fr om the cow died of tuber-
• From the America" Veterillary Review. t New York }II[eJicaIJoltrual, J uly 23, 1893. : Medical N elliS, New Yo rk , December 8, 1804 .
A rtijic/al I mmunisatio1t
of Cattle.
5
culosis in seven weeks. De Schweinitz injected very large quantities of human tubercle bacilli into ca ttle subcutaneously, intravenously and intraperitoneally. He found that by gradually increasin g the dosage enormou s quantities could be tolerated without injury. He administered as much as 500 c. c. of a heavy suspension of tubercle bacilli at one time . McFadyean, ~' in I g0 1 and 1902, reported that he had found that the r esistance of cattle to tuberculosis may be very greatly increased by the use of successive inocula tions with tuberculous m a terial or tu bercle cultures of low virulence. In a paper of McFadyean's en titled" Further Experiments Regardin g the Immunisation of Cattle against Tuberculosis," t the author states this conclusion: " It appears to be justifiable to conclude that, whatever may have been the degree o f natural immunity possessed by these three experimental animals, it was much in creased by the successive intravenou s inoculations to which they were subjected. Th e immunity was not ab solute, but it may be doubted wh ether a degree of resistance that would merit that term is obtainable by any method in cattle." von Behring announced in December, in I g OI, that he was engaged in studying the immunisation of cattle against tuberculosi s, and he has since issued several report s upon his work. von Behring has made a very larg e number of experiments in this field ; he has endeavoured to produce immunity by the use of tuberculin s , by the use of other tuberculosis to xins, by anti· toxins, by the use of dead tubercle bacilli, of tubercle bacilli weakened by chemical agents, and by the use of tubercle cultures of low virulence. A method for the vaccination of cattle against tubercu losis ha s been formu lated by von Behring, and it is ba sed on the u se of tubercle bacilli of human origin th at are nonvirulent for cattle . In makin g this" vaccine" the tubercle bacilli a re dried and ground to a powde r. The vaccine ma terial is sent out in this dried sta te. It is necessary to make a suspension of it in normal salt solution, whereupon it may be administered by intravenous injection . A number of investigators, as Lorenz, Schlegel, Eber and Hutyra , have tested the resi stance of cattle treat ed by von Behring, or according to von Behring's m ethod. It has been shown that tbe resistance of these a nimal s to tuberculosis has, in most cases, been increased and sometimes considerably so. lt is to be observed, however, that many of the cattle reported upon by the above authors were not vaccinated according to the method that von Behring now recommends, a nd as Eber says : " . .. The tuberculin test is not a reliable means for determining the freedom of tuberculosis of an animal that has been treated with mild bovine or human tubercle bacilli unle ss a long time (generally more than a ha lf year) has elapsed since th e las t administration of in fectious material. . . . Sin ce neither of the treated cattle was vaccinated bv the two-vaccination method with attenuated human tubercle bacilli, according to the method now recomm ended by von Behring, th e conclusions that are reached as a result of my investigations have only a re lative bea rin g on an estimation of the value of the method now recommended by von Behring. " The results of the investi gations show, however, that it is possible .. J ournal of ComjJarative Pat/lOlogy and ThemjJeutics, June,
t ibid. , March,
1902 .
19 01,
and March,
1902.
6
The Veterilzary
'.1 ournal.
to confer upon cattle, by treating them with attenuated bovine ·o r human tubercle bacilli, a certain degree of resistance to artificial tuberculosis infection . "Whether this resistance, as produced by the two-vaccination method, with attenuated human tubercle bacilli, that is now in use, furnishes protection against natural infection, can on ly be determined by years of careful observation and the greatest possible number of animals immunised in this way, and controls at the ·time of slaug h ter. " Hutyra ~' has reported some work conducted by him in 1903 and 1904 at the Royal Veterinary College of Hungary, wherein he tested the immunity of calves vaccinated according to von Behring's method, with material furnished by yon Behring, and also with material prepared from cultures made by himself. Hutyra tested the resistance of his .vaccinated animals by inoculating -them with v irulent bovine tubercle bacilli, and their resistance was compared to that of unvaccinated animals. He found that nearly all of the vaccinated animals had more resistance than the unvaccinated animals. One animal treated with von Behring's vaccine (No.3) appeared to have received no immunity as a result of vaccination. Hutyra's own vaccine material from different sources appeared to give a noticeably higher degree .of immunity than that obtained from von Behring's. Recently, a large number of cattle, amounting to several thousand, have been vaccinated after the von Behring method in Germany, Austria and Hungary. The process does not appear to be free from danger in every instance, as is shown by the reports of Marks, Casper and others. It is as yet impossible to draw any conclusions whatever as to the efficacy of vaccinations from the results of this great number of vaccinations of cattle on farms. The only conclusion at which one may fairly arrive is that vaccination appears, in most cases, to be unattended by dan ger to the vaccinated animals. Whether immunity is conferred, and, if so, whether it is sufficiently powerful or lasting for practical purposes, is not yet shown by the practical application of von Behring's method. Of the animals that have been vaccinated on farms and afterwards exposed to infection, a number have been slaughtered and some have been found to be afflicted with tuberculosis, other.s have been found to be free from tuberculosis. The presence of lesions of tuberculosis is taken by some of von Behring's observers to mean that the animal was tubercular before it was vaccinated. Of course, this may be true. But it must be remembered that the absence of lesions of tuberculosis in vaccinated animals cannot fairly be taken to signify the existence of a serviceable degree of immunity, unless it is shown that the conditions under which these animals were kept w.er.e such as to lead to the infection of a considerable. proportion of unvaccinated animals; and this evidence is in most cases lacking. Klimmer has carried out some very useful investigations upon the immunisation of cattle against tuberculosis at the Royal Veterinary College at Dresden. t Klimmer protests againg sending out vaccine material in the dry, powdered state, on account of the great danger to the operator who attempts, in the field, to prepare this material for use. Klimmer's practice is to prepare suspensions of tubercle bacilli in the • Zeitschrift fur Thiermedicin, Band ix., Heft 3.-4 , 190 5. t Berliner Tierarzliche Wochensch,.ijt, July 5. 1905·
Artificial immunisation of Cattle.
7
laboratory, just as has been done for a number of years at the laboratory of the P ennsylvania State Live Stock Sanitary Boa rd. Th e vaccine material that has been prepared in Dresden appears to be harmless to the vaccinated animals, and indications thus far are t hat it confers a serviceable degree of immunity. but it is not yet possible to draw final co nclusions from thi s work for th e reason that the vaccin ated a nimals are still living, and it is im possible to determine posi ti vely whether th ey are free from tuberc ulosis. Klimmer proposes the use of a vaccine material made o f mammalian tubercle bacilli that have been rendered less virulent by passage throug h a cold-blooded animal. The advantage from such a c ulture wo uld lie in the total absence of danger to the operator. Work upon the vaccination of cattle again st tuberculosis at the la boratory of th e State Live Stock Sanitary Board of Pennsylvania co mm enced in the yea r I gO I. It h as, therefore , been under way for abo ut five years. We were a ble to show three years ago* that it was possible to in crease the resistance of animals to tuberculosis to a very high degree by treating them with several intrave nou s inoculation s of non-virulent tubercl e bacilli of hum a n type. Two cattle thus trea ted were inoculated intra tracheally with a suspension of bovine tubercle bacilli , and both of th e m wholly resisted the inoculation. Th e only lesions in either of th ese animals was a slig ht thickening upon the wall of the trachea at the place where th e hypodermic needle was insert ed. Two unvaccin a ted animals, inoculated a t the same time, in the sa me way, with th e same quantity of bovine tubercle bacilli from the same culture, became exten siv ely infected with tuberculos is, showing the lesions on the mucous membrane of the trachea and bronchi, in the lun gs, and in tbe bronchial, mediastina l, and postphary ngea l lymphatic g lands. The immunity that was obtained in th e case of th ese two animals res ulted from th e a dmini stration of seven intravenous injections of vaccin e m ateria l during a period of t en weeks. The doses of tubercle bacilli ran ged from 13 to 26 mg. While this ex periment, and other simila r ex periments, were sufficient to lead liS to th e conclusion tha t anim a ls may be rendered immune to tuberculosis , it was at the same ti me evident th a t a process depending upon the application of vaccine material seven different times, would be of co mpa ratively small value in ge neral practice. The refore we set oursel ves to the stud y of the effects of other methods of vaccination, and to the development of a simple and effec tive m ethod. During the past th ree years we have a ttempted to determine tbe immunising effects of severa l strains o f tu bercle bacilli. We have administered vaccines in di ffere nt doses, at long a nd short interval s, a nd w ith varyin g numbers of doses. For these experim ents we have been supplied by the State Li ve S tock Sanitary Board with a large number of animals. For two years we h ave had the use of a farm where about one hundred ca ttl e have been kept for experim ental purpose s. We have also had the use of stables a t the Veterinary Departm e nt of the U ni versity of Pen nsylvania , where from twe nty to tw enty·five cattle have been kept , toge ther with numerous goats and small labora tory animals. Throug h th e use of the farm we have been enabled to keep a considerable number of cattle under practic al farm conditions. * Philadelphia lIfedical J our1lal , November
29, 1902.
The Veterinary! ournal. During the time that they have been under observation in our experiments the resistance to tuberculosis of vaccinated animals has been tested through exposure to natural infection and by inoculation. When vaccinated animals have been exposed to infection, and when they have been inoculated, unvaccinated control animals have been equally exposed or inoculated. We regard this use of controls of the highest importance, because it is only in this way that a definite standard for comparison can be provided in order to measure the extent of resistance to a given amount of exposure. Moreover, it is only by the use of controls tqat one can ascertain whether exposure under natural causes has been sufficient to lead to the infection of unvaccinated animals. It is not enough to associate vaccinated cattle with tuberculosis cattle, and then to conclude that the vaccinated cattle were immune, because, when they were killed, they did not show lesions of tuberculosis. It might readily happen that the exposure was not sufficient to lead to the infection of unvaccinated cattle; but if, upon equal ex posure, we find that the unvaccinated cattle are tuberculous, and the vaccinated cattle are free from tuberculosis, then we may ~afely and justly conclude that the vaccinated cattle received immunity from their vaccination. Our proof is based upon such clear evidence as this, frequently repeated. Our experiments have shown that different strains of tubercle bacilli hav e different immunisin g values. Avian tubercle bacilli may produce an intoxication resulting in great emaciation, but avian cultures do not appear to confer upon animals so much immunity as results from the use of mammalian cultures that are non- virulent for the animals upon which they are used. In order to produce artificial immunity in mamm a ls it appears to be necessary to vaccinate with an attenuated mammalian culture. We have not found any advantag e either in respect to th e degree of immunity or economy of time in the use of several vaccines made from cultures of different and progressive degrees of virulence . The amount of immunity bears a rather definite proportion to the number of vaccinations and the amount of vaccine material used, provided, however, that the animal be not "over-vaccinated"; that is, that th e vaccinations are not too close to one another, and that an excessive amount of vaccine is not administered. By over-vaccination the resistance of an animal to tuberculosis may be [f~duced to a point below normal, or a fatal toxcemia may be caused. Experiments have been made wherein animals vaccinated with different doses and different numbers of doses have been inocula t ed at the same time, and in the same way. While the resistance of all of the vaccinated animals to the inoculation has been much greater than the resistance of unvaccinated animals, it has been possible to observe a marked differen ce between animals vaccinated in different ways . For example, an animal vaccinated five times has more resistance than an animal vaccinated four times, and, other conditions being equal, an animal vaccinated four times has more resi s tance than an animal vaccinated three times. From the practical standpoint, it is more important to ascertain the amount of vaccination that is n ecessary to protect animals under customary conditions of exposure and the shortest possible time during
A rfi/icial i11tJltUl'lisafz·01l
0/ Cattle.
9
wh ic h a s ufficient degree of immunity may b e conferred, than it is to d ete rmin e h ow g rea t a deg ree of immunity may be produced. Since it has been necessary to try a g reat number of different m ethods of vacci na tion, and since the tim e required for an ex periment of thi s sort is mu ch g rea ter than is usual in experiments conducted through labora tories of hygie ne, it has b ee n difficult and time-consumin g to a cc umlllat e evidence upon which to base a genera l method for t he vaccination of ca ttle . It is a lread y clearly evident that different deg rees of immunity, a nd serviceable degrees of immunity, may be obtained at will. Under nat ura l conditi ons, animals differ in respect to their inh erited or acquired r esis tan ce to tube rc ulosis. Conditions of exposu re differ, bein g very mu ch greater upon some farms and in some h erds than in o th e rs. Th erefore it seems to be reasonable to conc lude th at animals will require more artificial immunity to tu bercul os is un de r some conditions than un der othe rs . The case is s imil a r to th e protection of cattle again st a n t hrax by vaccinat io n. In Pe nn sylvania we have fo und by ex perien ce , extending over a series of years, that the imm unity conferred by vacc inatin g with th e first and second vaccine of Past eur usually is suffi cie nt. On a few farm s, how e ver, the exposure a ppears to be 1l 10re inten se , a nd it is fou nd th a t on s uch farms it is n ece ssary to u se the third vacci ne o f Pa st eur. Ex p erience may show that it may be necessary to vaccinate different con dit io ns in slightly different ways . The amou nt of immun ity that is confe rred by three vaccinations is rat h e r high, a ncj will probably be sufficient in most cases. Indeed, two vaccinations may s upply a suffic ien t a m ou nt of immunity under s om e cond itio ns. The subject of the durat ion of immunity is ver y important, and is o ne th at can be solved on ly by observations upon a large series of vaccin ated ca ttl e, some of which ma y be killed and examin ed post-mortem fro m time t o time during a t erm of years. O f cou r se it is necessary that animals k ept for this purpo se should con stant ly be ex posed to infec tion, and vaccin ated controls must be simil a rly exposed, and the post-lIlortem results must s how that th e amount of exposure was sufficient to cau se the infection of the unvaccinat ed a nim a ls . Our own ex pe riments indicate that artificial immunity m a y endure a t leas t t wo years, and th ere is eve ry r eason to expect that a dditional ex periments will show that immunity will be of lo nger duration than this. Perh aps it may la st throughout th e entire life of th e a nimal ; but conclu s ion s on this point a re entirely pre mature. The effect of tuberculosis vacci nati on u pon ca ttle a lready infected w ith tubercu losis has also recei ved our attention';' ; it h as been found tha t intra v enous injections of tllbercl e bacilli of hum a n type, nonv ir ul ent for ca ttl e, have a very marked influence not o n Iy in restraining th e progres s of e xisting lesions but also in causing them to become e ncaps uled and to recede. In o th e r wo rds, such treatment a ppears to have a marked curative effec t on some tuberc ulou s an imal s. B ut this sta te ment sho uld be accompanied by a n ote of warning. 'vVe do not for a moment wish to imply tha t it is pra cticabl e , or eve n possible, a t this tim e , to c ure tubercu lous cat tIe. The observations to be made • Uu iversity 0/ FCII1IS) llvall ia llfedical Bulletill . . \pril, 19 :>5. R eview , J 905.
Amcricall Veterilla,J'
10
The Veterinary
'.1 oUr1zal.
in this direction are ' regarded by us as of importance chiefly as tending to throw additi onal lig ht on the subject of immunity. It was shown tbat the treatment give n had the same immunising effect upon infected animals as upon hea lthy animals. The effect upon the immunisa tion in these cases was, first, to prevent the further dissemination of infection, and, second , to ca use the encapsulation and elimination of the existing lesions. It is ' probable that this method of treatm en t may prove to be of value with relation to young animals, or animals known to be but recently infec ted with tubercul osis . Such treatment will always have to be carried out with the most ex treme care and under co ndition s that can be very fully controlled . It is not to be recommended for general prac tice. As to the applicati on of vaccination against tuberculosi s: vaccination, properly applied, is effective, and we believe that it will prove to be of very g rea t practical va lue. It is not poss ible on most American farms to establish a sepa rate herd of infected cattl e, an d to keep the infected cattl e as separate and di st inct from the uninfected cattle as is necessary, if tubercul osis is to be treated according to the Bang system. The alternative, if tube rculosis is to be eradicated, is to destroy all of th e anim als that react to the tubercul in test. Th e latter plan involves a grea t deal of waste and loss, which he ret ofore has been unavoidable , and which has been amply justified because it has been less than that resulting from th e continuat ion o f the in fec tion in the herd . But it l S im por tant that a better, a less expensive, and, if possible , a more effec tive method shall be devised. It appears to be hig hly probable that vacc in a tion may supply this method. It is a dmittedly impossibl e to test with tuberculin all of th e tubercu lous herds of a state. Therefore, it is not possible to discover a nd to eith er destroy or sequester all of tbe cattle th a t m ay be dis tributers of tuberculosis. Herds freed from tuberculosis by the use of the tuberculin test are always exposed to reinfection, and must be g uarded by the double testin g of all r ecruits and by systematic reinspections of tbe herd. All of this may be, and is, carried through successfull y , and sho uld be practiced to a much greater extent than it is, unless a n equally effecti ve and c heape r method can be devised. It will be a grea t step in advance to be able, not only to discover that an animal is free from tuberculosi s, but also t o r ender that ' animal permanently immune to infection . It will be a g reat advantage to be able so t o treat yo un g cattle that they may resist tuberculosis even thoug h they are continu a lly ex posed. Th e advantage to the breeder of being able, not only to g row a herd free from tuberculosis, but immun e to tu berc ul osis, will be very grea t. We believe that all of this may be looked forward to; but until more experiments and obser vations are completed, vaccination should be applied only under such conditions a s will permit exact observations to be made, and each vaccination must, for the present, be regarded as a scientific experiment.