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
When the sample was removed for standardization it contained not only a thick suspension of Pfeiffer bacilli, but also bits of agar and blood-stained debris. It was necessary to rid the suspension of the gross contamination, and this was done at first by filtering it through sterile glass wool filters, and later by centrifuging it at slow speed for about 10 minutes. The suspension then contained little but the Pfeiffer bacilli, and was placed in the Hopkins tube and centrifuged for ½ hour on the sixth contact of the rheostat. This gave the per cent. of Pfeiffer bacilli in the suspension, and the necessary dilutions to make 1,000,000,000 per cubic centimeter were readily determined. The Hopkins tube consists of a centrifuge tube, with a capillary tube sealed on at the smaller end. The centrifuge tube is graduated in 10 c.c., 5 c.c. and 1 c.c. amounts, and the capillary portion is graduated in 0.01, 0.02, 0.03, 0.04 and 0.05 c.c. amounts. To standardize the vaccine, 10 c.c. of the sample was centrifuged in the tube and the amount of sediment read on the capillary scale. If the amount of bacilli fell between the graduations, an additional amount of sample was added, so that the sediment reached one of the graduated lines, the exact amount of sample added being noted. The percentage of the suspension could thus be determined by dividing the number of c.c. of sample used into the amount of the sediment obtained, and the number of bacteria calculated according to Hopkins table. The table available to us did not list the Pfeiffer bacillus, but according to it a 1 per cent. suspension of staphylococcus contains 10 billion organisms to the cubic centimeter, and we estimated that Pfeiffer bacilli were about half the size of staphylococci. This assumption was borne out by a number of Wright’s method counts on standardized suspension of bacilli. We, therefore, calculated that a 1 per cent. suspension of Pfeiffer bacilli should contain about 20 million organisms. Then, if 10 c.c. contain 0.02 c.c. of bacterial sediment, the per cent. was calculated by taking 0.02
10 = 0.2 per cent., the strength of the suspension. If 1 per cent. contains 20 billion, then 0.2 per cent. contains 4 billion per c.c. In order to get a 100 million per c.c. suspension, it would be necessary to dilute the original suspension 40 times.
Every method of standardization is more or less inaccurate, but the above described method gave a fairly uniform product. Drying and weighing is claimed by many to be more accurate, but even with this procedure a fair amount of non-bacterial sediment is present in the material to be weighed.