Part 10

When a sufficiently large dose of vaccine is given to an individual there is usually a transient rise in temperature for from 12 to 48 hours; the local focus of injection becomes sore and inflamed, and a white count often shows an actual increase in the number of polymorphonuclear leucocytes in the general circulation. A series of doses are usually given. If after a few days blood is withdrawn from the patient and immuniological tests made, it will generally be found that the patient’s leucocytes take up bacteria, and particularly the type of bacteria of which the vaccine was composed, more readily and in greater numbers than the leucocytes of the ordinary individual. Wright and Douglas (52) and Neufeld and Rimpau (53) have shown that this effect of increased phagocytosis is brought about by the vaccine through the production of substances which act specifically on the bacteria and render them more susceptible to inclusion within the white cells. These substances belong to the group of antibodies, and are known as “opsonins” or “bacteriotropins,” and are specific for any given bacteria. Moreover, the serum of the patient will, as a rule, be found to have developed the faculty of agglutinating and bacteriolysing suspensions of the specific organism injected and of fixing complement in the presence of an antigen prepared from that organism. In animal work it has been possible to go still farther, for it can be shown that the resistance of the animal can be raised until it is no longer possible to kill it with the same dose which is found to be fatal for the unimmunized animals. Not only has animal work made it possible to determine the protective powers of vaccines, but it has also served to show the specific nature of the protective power and the relative extent to which “group” or “crossed” protection can be conferred by vaccinating with closely allied organisms—as, for instance, paratyphoid bacilli in typhoid fever. The non-toxic nature of vaccines is also determined by animal experiment before such preparations are injected into humans.

The most successful prophylactic bacterial vaccine which has been developed so far is that for typhoid fever. A comparison of the occurrence of typhoid fever in the United States Army before and since the use of anti-typhoid vaccine is all that need be cited to convince one of its value. At the time of the Spanish War there was no vaccination against typhoid fever, and there were 20,738 cases, with 1,580 deaths, among 107,973 men who remained in the camps in the United States during the war (54).

During the summer of 1911, the maneuver division of the United States Army, having 12,801 men, all of whom had been vaccinated against typhoid fever, were stationed at San Antonio, Texas. Two cases of typhoid fever developed among them, and neither case died. Among the civilian population of the city, living under usual conditions during the same time, there were 49 cases of typhoid fever, with 19 deaths. Since 1912, typhoid vaccination has been compulsory in the United States Army, and the largest epidemic of typhoid fever which I have found reported so far during the late war was that at Camp Greene (55), Charlotte, N. C., where 18 cases developed. Only 12 of these men had received the complete series of immunizing doses. For a complete discussion of the value of typhoid vaccine the interested reader is referred to Gay’s Monograph (56) on typhoid fever.

_Prophylactic Vaccination Against Influenza_

The hope of finding an early solution to the vaccine problem in influenza appeared to be in the development of a prophylactic “bacterial vaccine” similar to that which proved so efficient for typhoid. In his discussion of the vaccine problem in pneumonia, Fennel pointed out that, theoretically, any disease of microbic origin in which spontaneous recovery is at all possible should yield to specific prophylactic measures. The difficulty, however, of preparing a bacterial vaccine for influenza comparable to that for typhoid fever is that the unquestioned cause of influenza has yet to be determined. The probable cause of influenza is the Pfeiffer bacillus, but its relationship has not been proven beyond question. On the other hand, the innocence has likewise not been proven, as Dr. Holman in his article of this series has ably shown. It is not my intention to go deeply into the question of etiology, but simply to bring out a few points which _a priori_ seemed to indicate that the reasonable solution of vaccine prophylaxis was in the preparation of a pure Pfeiffer bacillus suspension.

The experiments in man lead to very surprising results. Rosenau, Keegan, Goldberger and Lake, at Gallops Island, Boston, Mass., (1) inoculated volunteers with pure culture of B. Pfeiffer, with secretions of the upper air passages and with blood from typical cases of influenza. Sixteen men, of whom 13 were supposedly non-immune, had Pfeiffer bacilli installed into their nasal passages, and none of them developed the disease. Secretions filtered and unfiltered also gave negative results. Contact with well-developed early cases also failed. McCoy and Richey (1a) conducted similar experiments in San Francisco, with negative results. The men of the latter group had been vaccinated with a mixed streptococcic vaccine, which may have played some part. Had the experiments with the Pfeiffer bacillus been negative and the other experiments positive, they would have shown that the bacillus of Pfeiffer was not the cause of influenza; but since all attempts were negative, it merely brought out the fact that there had been a change, due probably to some immune factor, which seemed to have acted alike on the Pfeiffer bacillus and all other types of virus present, and to have made them all innocuous. These experiments still leave the cause of influenza in question.

Those who are opposed to the Pfeiffer bacillus being the cause of influenza in its epidemic form base their position on the points that the common finding of the bacillus might be accounted for on the grounds of its being a secondary rather than a primary invader; that while it is not so common at ordinary times, it does occur with other organisms in whooping cough and sometimes in chronic diseases of the air passages, and that the rules of Koch have not been complied with in that the organism has not been found in every case of the disease; that where it has been grown in pure culture and inoculated into man and animals, it has either produced no disease, or the lesions which followed have not been typical of epidemic influenza. On the side of those who believe that the Pfeiffer bacillus is the chief cause, or, at any rate, that it is partly responsible for epidemic influenza, are the facts of its fairly constant presence in the purulent bronchial secretion of patients suffering from epidemic influenza; its relatively uncommon occurrence at other times; its known pathogenicity in occasional cases of meningitis, and in the inflammation of the bony sinuses of the head and face; the relative immunity of nearly all common laboratory animals and the fact that the attempts to transfer epidemic influenza from man to man failed not only when Pfeiffer bacilli were used, but also when direct contact and direct coughing by the patient into the face of the volunteer were tried. The argument that many cantonment laboratories failed to find the organisms loses weight when we find that the percentage of positives increased where the material examined was removed directly from the lungs at autopsy, where special cultural methods were in use and where the laboratory personnel was large enough to devote a sufficient amount of time to each individual culture. All of these points indicate that the organism was overlooked in a great many instances. In our laboratory we found the examination of sputa very unsatisfactory because of the great amount of contamination, and because the bacillus seemed to lose its ability to grow after a relatively short time in the sputum in vitro. Moreover, I am convinced that the bacillus changes its morphology to such an extent under varying conditions as to make it impossible of identification when present among other organisms in sputum smears. The failure of animal inoculations is also not conclusive evidence against the Pfeiffer organism, because guinea pigs, rats and mice have a natural immunity for them. Rabbits are only slightly susceptible, and then only to intravenous injections. The mixture of the Pfeiffer bacillus with any one of several other pathogenic organisms will increase the pathogenicity of both. Monkeys inoculated intracranially develop a typical Pfeiffer bacillus meningitis.

Whatever the ultimate outcome of the investigations as to the parasitic cause of epidemic influenza, the Pfeiffer bacillus was the generally accepted cause at the beginning of the 1918 epidemic, though it was at once realized that most of the deaths were due to complicating pneumonias and to secondary infections with other organisms. Under the circumstances, one of two courses was open: (a) the acceptance of the Pfeiffer bacillus as the presumptive cause of influenza and the preparation of a specific prophylactic vaccine against infections with that organism; or (b) the use of a mixed bacterial vaccine containing the common and most deadly secondary infecting organisms, designed to increase the patient’s general resistance by decreasing his susceptibility to the allied, collateral and secondary infecting agents. Attempts were made along both lines, with more or less unsatisfactory results.

_The Attempt to Develop a Specific Prophylactic Vaccine by the Use of Pure Pfeiffer Strains_

By a specific prophylactic vaccine for any given disease, we mean a material which when inoculated into an individual will actively protect that individual against the given disease. In infectious diseases, the immunizing material is usually of microparasitic origin (in contrast to desensitizing substances used in pollen diseases and those due to unusual sensitiveness to foreign proteins), and is specific only for the disease caused by the microparasite from which the material was prepared. With the knowledge in hand during the epidemic, the logical plan seemed to be to prepare a pure Pfeiffer bacillus vaccine, the object of which was to eliminate primary infection with that organism and thus prevent the secondary invaders from obtaining a fertile soil.

While specific Pfeiffer bacillus vaccines had been tried in treatment, the field was a comparatively new one so far as prevention was concerned. Many of the biological products companies had so-called influenza vaccines on the market for treatment purposes, and many of these contained Pfeiffer bacilli. A few preparations of pure strains of the bacilli were also available, but I was unable to find any records of their use for prophylaxis. Lacy (2) reported two cases of sinusitis treated with autogenous vaccines made from pure Pfeiffer strains—one patient improved rapidly and the other showed no change. Investigation of several of the other references on influenza vaccines showed that mixed vaccines had been used in each instance. The work of Flexner and Wolstein (3, 4 and 5) indicated that active immunizing substances could be prepared from the Pfeiffer bacillus, although they worked with serum instead of vaccines. They prepared an anti-influenza-meningitis serum by immunizing goats and horses. These sera cured monkeys of experimentally produced influenzal meningitis. The sera showed agglutinins and bacteriotropins for Pfeiffer bacilli, as well as positive fixation tests in dilutions of 1 in 100, but they contained no lysins. The serum was offered for intradural use in treating influenzal meningitis, but was found to have no value when used in human cases.

The first references which we have found on the use of pure Pfeiffer bacillus vaccines for the prevention of epidemic influenza were those of Leary (6), (7), and of Rosenau (8). Shortly after the appearance of the first influenza cases in Boston, Leary used a vaccine prepared from several strains of Pfeiffer bacilli both for the treatment of influenza and for its prevention. The vaccine for the latter purpose was given to medical students and nurses, and the first results were apparently very encouraging. Continued use has not been convincing. Barnes (9) reported an attempt to protect the employees and patients of an institution near Woonsocket. On October 9 a case of influenza developed in the female ward, and was followed five days later by another. On October 22 the disease appeared in the male ward, and the same day 172 employees and patients were given their first inoculation with Leary’s vaccine. Doses of 400, 800 and 1,200 million bacilli were given at 24–hour intervals. All persons who had developed influenza before the three doses had been completed were excluded from the computation of the disease incidence, which was found to be 20 per cent. both among vaccinated and unvaccinated individuals. The mortality rate was 16 per cent. for the 25 cases among the vaccinated, and 15.8 per cent. among 57 unvaccinated patients. The result failed to show any protective qualities for the vaccine.

The best controlled vaccine experiment in which Leary’s vaccine was used was that reported by Hinton and Kane (10), and was carried out at the Monson State Hospital for epileptics. The hospital had a population of 979 inmates, ranging from 4 years of age to senility; of these 461 were vaccinated and 518 were not. Vaccination was begun on October 6, and three doses of 400, 800 and 1,200 million were given at 24–hour intervals. The first case of influenza developed a few hours after vaccination was completed, but there were no more cases before October 12, when five cases developed. The table shows the result of the work, and that the vaccine failed to protect.

Population. No. of % of No. of % of Cases. Cases. Deaths. Deaths. Vaccinated 461 163 35.4% 28 17.1% Unvaccinated 518 178 32.4% 24 13.4%

Attempts to protect by the use of Leary’s influenza vaccine were made in 11 other Massachusetts institutions, but the results cannot be used to compare the incidence and mortality rates between the vaccinated and unvaccinated, because the epidemic was either on the wane, or at least well advanced when the vaccinations were begun. The reports are of great interest in showing the large number of vaccinations which failed to protect.

In the Taunton State Hospital about 800 were vaccinated, and among them there were 81 cases of influenza and 17 deaths from pneumonia, even though the epidemic was on the wane when vaccinations were begun.

In the Gardner State Colony 834 were vaccinated after the peak of the epidemic had passed. This number included all but 15 of the inmates who had not contracted influenza up to that time. Out of this group, 62 vaccinated individuals developed the disease.

At the Massachusetts School for Feeble-Minded 457 inmates were selected for vaccination and controls. Of the 234 vaccinated, 56 developed influenza. Of the 223 unvaccinated, 185 developed influenza, with 16 pneumonias and 12 deaths. The vaccinated group, however, were a more vigorous group of individuals to begin with, and represented a higher mental grade than the unvaccinated group, so that the evidence was considered of questionable value.

At the Wrentham State School the influenza epidemic was well under way before vaccinations were begun, and hence the susceptible individuals were in a large part either affected or infected with the disease. Of 1,198 unvaccinated persons, 758 developed influenza, giving a morbidity rate of 63 per cent. Of 128 vaccinated, 13 developed influenza and 1 died. Physicians in this institution believe that the vaccinated were not as ill as the unvaccinated patients.

In the Medfield State Hospital, having a total population of 1,940,421 cases of influenza, with 63 deaths, had occurred before vaccinations were begun. Of the remaining unattacked inmates 902 were vaccinated. After the completion of vaccination one new case appeared among the unvaccinated, and there were none among the vaccinated.

At the North Hampton State Hospital there were 9 cases of influenza, 4 of whom died, among 444 unvaccinated individuals, and 9 cases, with 1 death, among 563 vaccinated patients.

Among 506 patients vaccinated at the Westborough State Hospital there developed 15 cases of influenza, 2 of which terminated fatally. Of the 415 unvaccinated controls, 25 developed influenza and there were no deaths. At the time vaccinations were completed only 13 had developed influenza.

In the Worcester State Hospital vaccination was carried out after the epidemic had entirely subsided.

At the Bridgewater State Hospital no vaccines were used, but the morbidity rate was 29.9 per cent., as contrasted with 32.9 per cent. among the unvaccinated at Monson.

At the Danvers State Hospital the population of 853 adults was divided into three sections. One section was vaccinated with the Leary vaccine, one section with an unheated influenza vaccine prepared by Dr. Rosenau at the Chelsea Naval Hospital, and one section held as controls. The epidemic had, however, reached its height before vaccination was begun, and no information as to the relative value of the vaccines could be determined.

In Hinton’s (11) report the analysis covered the studies on about 6,000 vaccinated individuals, which represented slightly less than half of the population of 12 Massachusetts State institutions. Hinton’s conclusions were as follows: “The heated suspension of influenza bacilli used as a prophylactic vaccine did not prevent influenza, lessen its severity nor its complications, and, as far as could be ascertained, resulted in no harm.”

About the same time that Leary was working on his vaccine, Rosenau prepared an unheated suspension of Pfeiffer bacilli, isolated from cases of influenza of the existing epidemic, which he used at the Chelsea Naval Hospital and in an experiment at the Pelham Bay Naval Training Station. The writer is indebted to Surgeon-General of the Navy W. C. Braisted for the data from which this report was compiled—the report of the Sanitary Officer of the station not having been completed at the time the information was furnished. The vaccine experiment was made in the isolation regiment, which had remained practically free of influenza. Inoculations were begun on September 30, when 638 men were given the first dose of vaccine, 833 men being held as controls. On October 4 the second dose was given to 589 men, and vaccination was completed on October 8, when 565 men were inoculated. This group comprised the total number who received three inoculations. On October 14 practically all of these men were transferred, so that it was very difficult to get a complete record. Those cases which developed influenza prior to October 10 have been omitted by the writer, both from the control and vaccinated groups, because it is unfair to consider the incidence of influenza among controls which developed prior to the time the inoculations were completed in the vaccinated group. Between October 10 and October 24 there were 27 cases of influenza which developed among the vaccinated, and 30 among the controls, giving a morbidity rate of 3.6 per cent. among the 833 controls, as compared to 4.7 per cent. among the 565 vaccinated men. Emphasis is laid on the fact that these morbidity rates were calculated for both groups on the number of cases that appeared after vaccination had been completed. The result failed to show protective qualities in the vaccine.

Influenza vaccines for prophylaxis were also prepared in great quantities by the New York City Board of Health, and were made under the direction of W. H. Parke. No reports on the value of their vaccines have as yet appeared, and the writer has been unsuccessful in obtaining any data on the matter. The Parke vaccine was made in the following way: A large number of strains of Pfeiffer bacilli were isolated from cases of influenza during the epidemic. These were grown on a veal infusion agar containing 1 per cent. peptone, 0.5 per cent. of sodium chloride, 5 per cent. chemically pure glycerin, and the reaction of which was made neutral to phenolthalein in the cold. The agar was melted, and from 3 per cent. to 5 per cent. of citrated horse blood was added to it at a temperature above 95° C. The media was then slanted and cooled in 6 × 1 inch test tubes. Most of the vaccines contained about 17 different strains of Pfeiffer bacilli. The strains were inoculated separately on a series of slants, and at the end of 24 hours the cultures were washed off with sterile water and the washings from each series were placed in a separate bottle. Smears were then made to determine whether or not gram positive organisms were present, and as soon as each bottle was found to be free from contamination the contents were pipetted off into a 1,000 c.c. flask, and the dilution with sterile salt solution containing 0.25 per cent. phenol made. All of the strains were mixed together in the large flask. A sample was then removed for standardization by Wright’s method, and the flask was submerged for one hour in water at 53° C. Transplants for sterility were made and watched for 48 hours. The vaccine was then diluted so that each cubic centimeter contained 1,000,000,000 Pfeiffer bacilli. Prophylactic vaccination was carried out by giving ½ c.c., 1 c.c. and 1½ c.c. doses at seven-day intervals.

_Author’s Vaccine_

At the request of the Department of Public Health of the city of Pittsburgh, the writer undertook to prepare Parke’s vaccine in large quantities. The vaccine was to be prepared under the direction of a committee consisting of Drs. Oskar Klotz, W. L. Holman, E. W. Willetts, George L. Hoffman and the writer, and the vaccine was to be turned over to the City Health authorities for distribution in the community. The work was carried out at the Singer Memorial Laboratory, and was begun the same day that the committee was appointed. Thirteen strains of Pfeiffer bacilli were used. Holman contributed six strains, isolated at autopsies done by Klotz at the Magee Hospital. Other fresh cultures were furnished by Willetts; Wiese, of the City Laboratory, and by the Singer Laboratory. The media used was that recommended by the New York Board of Health, save that sheep’s blood was used instead of horse blood because of convenience. The same technique was employed, with the exception that a modification of the Hopkins method of standardization was used instead of the Wright method. This was done because Pfeiffer bacilli are extremely small, tend to form unbreakable clumps and tangles, and so increase the difficulties of making satisfactory counts, either by means of the Wright method or with the Helber-Glynn counting chamber, that the methods are independable. Opalescent standards permit of such enormous variations that it was decided to use the Hopkins method, or a slight modification which we found so satisfactory that we will give our method here in detail.

_Method of Standardization_