SYNOPSIS FOR PROTECTIVE INOCULATION OF CATTLE IN AGRICULTURAL PRACTICE.

GENERAL CONSIDERATIONS.

For the protective inoculation carried out by us in practice we make use of “Mxt. Tb.” in the form of our “Culture No. 1,” which has been studied by us for years and which we have minutely described in the Beiträge, Nos. 5 and 6. The accumulation of orders for this virus on the one hand and the multitudinous experimental labors in the institute here on the other, compelled us to rid ourselves of the technical labor of manufacture, weighing off and shipping of the virus, which accordingly has been undertaken by the local firm of Drs. Siebert & Ziegenbein. The virus, however, continues to be tested as to purity, virulence, etc., in the scientific department of the institute at Marburg.

The virus is accompanied by directions for use which have already appeared in the Zeitschrift für Thier-medicin. These are as follows:

I. Selection of the Animals to be Inoculated.—As a rule only animals without external evidences of disease, from three weeks to three months (for the first inoculation) should be inoculated. In healthy animals of this age a previous tuberculin test is unnecessary, even if the animal comes from a notoriously tubercular herd.

II. Numbering the Inoculated Animals.—Every inoculated animal must be marked with a running number. The marking must be of such a character as to be distinct, not liable to be mistaken for some other, and to last the lifetime of the animal. (Ear-marking, tattooing, etc.) If necessary, the marking is to be repeated should the first mark become indistinct.

III. Keeping of Records.—See under “Conditions governing the distribution of the virus, etc.,” [p. 75.]

IV. Technique of Taking the Temperature.—The body temperature is determined by means of a self-registering thermometer completely inserted into the rectum. Before introducing it, the rectum is to be cleared of any hard fecal masses. A tape about a foot long, having a clamp at its end, is tied to the neck of the instrument. The thermometer is left in the rectum for four minutes, the clamp meanwhile being fastened to the hairs at the root of the tail. In order to save time, thermometers are introduced into a number of animals (about six) consecutively. When the last thermometer has been inserted it will usually be time to extract and read the first. In this way one can take the temperature of fifty head of cattle in 1-1½ hours.

V. The Virus.—The inoculating virus consists of living tubercle bacilli whose action has been accurately tested in the Marburg Institute for Experimental Therapy. The tubercle bacilli have been dried without losing their vital powers in any way. These dry tubercle bacilli (Trocken Tb.), kept in sealed glass tubes, will retain their action on cattle unchanged for a period of thirty days. If, therefore, a tube of Tb. bears the date VII-1-02, the contents can be used for cattle immunization until VIII-1-02. After thirty days, although the immunizing power is not entirely lost, it is so far decreased as to render it ineffective in the dosage recommended.

VI. Dosage of the Virus.—For the first inoculation, one immunizing unit, 1 I. E. [= 1 Immun Einheit] is used for each calf; for the second inoculation, which is not to be undertaken until at least twelve weeks after the first, five units (5 I. E.) are used for each calf. As a rule the dose of 1 I. E. is 0.004 gramme dry Tb., that of 5 I. E. for the second inoculation is therefore 0.02 dry Tb.

If the tube contains the quantity requisite for the first inoculation of twenty cattle, it will bear the label 20 I. E. In order at all times to control the manner of production of the dry Tb. each tube also bears in Roman numerals the consecutive laboratory number thus:

Op. No. IV.
20 I. E.
VII-17-02

This label is interpreted as follows:

Dry Tb., which in the records of the Marburg Institute bears the laboratory number IV, contain on the day of delivery, i.e., on VII-17-02, twenty immunizing units, and retain this strength until VIII-17-02.

For the inoculation the virus is uniformly mixed with 1% salt solution which has previously been boiled and cooled. The procedure is as follows:

The entire contents of the tube are placed in a small mortar and crushed with the pestle. Then 2 to 3 c.c. salt solution are added and the whole rubbed into a uniform mixture or emulsion, after which it is poured into a graduated cylinder holding 50 c.c. A little more salt solution is then added to the mortar, and thus the remaining particles of virus are added to the previous mixture. The fluid in the graduate is then made up to 30 c.c. with salt solution, and then poured into a sterile wide-mouthed bottle holding 100 c.c. Any remaining emulsion is then washed out of the graduate with an additional 10 c.c. salt solution and added to the 30 c.c. in the bottle. The bottle thus contains virus ready to inoculate, and each 2 c.c. will be the dose for the first inoculation of a calf, provided that the tube originally contained 20 I. E. The dose for a second inoculation would then be contained in 10 c.c. of this fluid.

VII. Instrument Case.—In order to carry out the inoculations in agricultural practice, the instrument case constructed by W. Holzhauer, Marburg, is to be recommended. It contains the following:

(a) Two bottles for the virus, each 100 c.c., made of colored glass.

(b) One bottle lysol.

(c) One bottle alcohol.

(d) One Erlenmeyer flask (for 1% salt solution).

(e) One mortar and pestle.

(f) One graduated cylinder 50 c.c.

(g) One graduated cylinder of 10 c.c.

(h) One basin to hold disinfectant solutions.

(i) Six thermometers with tapes and clamps. (Price 10.50 marks or 1.75 marks each.)

(k) Two virus syringes with two strong canulas. (Price 11 marks.)

(l) One small hand-scale and weights.

(m) One alcohol stove and cover.

(n) Sterile cotton.

(o) One wire basket.

The entire box is supplied with a canvas cover and a leather handle. The lid, which bears four movable legs, can, when the same is removed from the box, be set up like a saucepan and used as a sterilizer. For this purpose the vessels and instruments are placed into the wire basket, the sterilizer half-filled with 2% lysol solution, and the whole heated just to boiling by means of the alcohol stove.

The price of the complete outfit is 85 marks. By omitting the thermometers and syringes the price is reduced to 63.50 marks.

VIII. Method of Making the Intravenous Injection.—The injection is best made by means of a 5-c.c. glass syringe with an asbestos plunger (paragraph VII, k), which is cleaned by means of the lysol solution and rinsed in sterile salt solution. The canula attached to the syringe is cleaned in the same manner. One immunizing unit (1 I. E.) is then drawn up into the syringe and any air-bubbles are expelled, care being taken that any drops of fluid expelled in this manner do not fall on the floor but are caught up in the basin. The filled syringe, together with the canula, which is now detached, is then placed on the previously mentioned basin. The virus is injected into the left jugular vein. When possible the cattle should be inoculated, each in its own stall, so as to avoid exciting them. After washing the left side of the neck with 2% lysol solution, the operator, by pressing his thumb on the jugular vein, stops the circulation, causing a distinct, sausage-shaped, fluctuating swelling to appear. With his right hand he now takes the canula from the basin and thrusts it, just above the compressing thumb, upward into the vein at an angle of 45°. One recognizes that the vein has been entered, by the blood that at once flows through the canula. If no blood flows, the vein has not been entered. In that case the canula is withdrawn slightly but not completely, and again thrust into the fluctuating swelling. As soon as blood flows, the left hand ceases making pressure and grasps the canula, whilst the right hand fits the syringe thereto. Then the virus is slowly and uniformly injected into the vein. When the syringe is emptied, the skin at the site of injection is pressed together and the syringe and canula withdrawn. As a rule, bleeding ceases almost at once. The region is then rubbed over with 2% lysol solution and the inoculation is completed.[9]

CONDITIONS GOVERNING THE DISTRIBUTION OF THE VIRUS PROTECTIVE AGAINST TUBERCULOSIS.

1. The virus is supplied by the firm of Drs. Siebert & Ziegenbein in Marburg a. d. Lahn, Germany.

2. This firm supplies the virus in packages containing 5 I. E. and 20 I. E. The price for the former at present is 40 pf. per I. E. and for the latter 25 pf. per I. E. In ordering, the style of package is to be specified.

3. The recipients bind themselves to fill out and return the following records:

A. Temperature charts: The temperature is to be recorded as a curve.

B. Other data as per blank herewith enclosed (see next page) [(original illustration)].

A Consecutive Marburg Number.
B Consecutive Inoculation Number.
C Breed, Herd and Designating Number.
DNationality. I.Owner.
II.Location. AAt Birth.
BAt Time of Inoculation.
III.Breed.
IV.Sex.
EManner of Raising. I.Natural (Suckling Calf).
II.Artificial.aWith Nonsterilized Milk.
bWith Sterilized Milk.
FProtective Innoculation. I.Laboratory Number of the Virus.
II.Dose in Grammes or in c.c.
III.Date and Manner of Inoculation.
IV.Degree of Reaction.
GTuberculin Test.Date.
Degree of Reaction.
HWeights.
ISpecial Remarks.

Directions for filling this Blank.—Column A is to be left blank by the attending physician; column B contains the running number of the animal inoculated; C is to be recorded only in fitting cases; D, on the other hand, is never to be omitted; under E the fitting column is to be marked thus: “!”; in column F, IV, the degree of reaction following the inoculation is to be noted by signs, thus: O = Failure of any reaction whatsoever (R = O); I = short febrile reaction; II = 2-4 days of fever; III = 5-8 days of reaction associated with other disturbances of health (cough, loss of weight, diminution of appetite, diarrhœa, etc.).

In the same manner the degree of the reaction to a later tuberculin test is to be recorded (column G). In column H the weight is always to be accompanied by the date.

The returns are to be made as soon as the data for about 100 cattle have been secured.

C. General report: This includes a report on the manner of keeping the inoculated cattle, the hygienic conditions of the stables, previous occurrence of perlsucht in the cattle of the herd, previous use of other means of suppression of tuberculosis and their result, etc., etc.

4. The owners of the cattle must see to it that the inoculations are properly made and they themselves must bear all consequences resulting from defective inoculation. Drs. Siebert & Ziegenbein can in no way accept responsibility for this.

The preceding conditions are still adhered to in many large dairy farms. To a number of smaller dairies we have allowed easier conditions. An example is afforded in the following order of the Ministry of the Interior of the Grandduchy of Hesse.

MINISTRY OF THE INTERIOR OF THE GRANDDUCHY OF HESSE,

Department of Public Health.

To the Grandducal County Veterinarians:

Whereas, following the proclamation of the Ministry of the Interior dated Aug. 19, 1902, a large number of replies has been received from the farmers, it is ordered that the inoculations be hereafter made in accordance with the following principles:

1. The protective inoculation consists in the intravenous injection of a single immunizing unit of tubercle germs (derived from man), rubbed up with sterile salt solution; and in a second intravenous injection, three months later, of five immunizing units of the same germs.

2. The germs in question can be obtained in glass tubes containing 5 or 20 units.

3. As a rule only cattle without external manifestations of disease, aged from three weeks to four months (at the first inoculation) should be inoculated. With healthy animals of this age it is unnecessary to test them previously with tuberculin, even if they belong to a notoriously tubercular herd.

4. In exceptional cases older animals (from four months to two years) may be inoculated with the virus, but only when they are entirely free from disease, and when a tuberculin test made on them results without any reaction whatsoever.

5. The inoculated animals are to be kept in their stalls two days before and two weeks after the inoculation.

6. In all cases in which the owners can manage the rectal temperatures, the same are to be taken two days before the inoculation (morning and evening) and again on the morning of the inoculation. After this they are taken once more in the evening and then once daily for the next five days. If, after this, the animals still have temperatures above 39.2° C., the temperatures are to be taken daily until the same reach 39.2° C.

7. In animals not over four months old at the time of the first inoculation, the temperature may be omitted if there be any difficulties in taking it. In those, however, which are over that age at the time of the first inoculation, the temperature must invariably be recorded according to paragraph 6. Even when inoculating animals in infected herds the above-mentioned measurements should be taken whenever practicable.

8. If possible, the inoculated animals should be weighed every fourteen days on a cattle scale.

9. Owners of cattle are to keep record of all the temperatures and weights. You are to collect the records concerning temperatures as soon as they have been completed; the records of the weights, however, only after three months. The results noted in the temperature returns are to be entered by you on the charts herewith provided, and a temperature curve plotted. These charts are to be made in duplicate, one copy to be sent to our representative and the other to the Experimental Division of the Hygienic Institute, Marburg.

10. The inoculated animals are all to be permanently marked. Those on which temperature and weight observations have been made should also be numbered.

The method of thus marking and numbering the animals is left to your judgment.

11. Should you not be informed in regard to the previous notifications respecting the inoculations, kindly place yourself in communication with the grand-ducal “Kreis Amt.” Following this, as soon as you have informed yourself regarding the age and health of the animals to be inoculated, notify our representative as to the number of these animals suitable for inoculation. The reports for each village are to be made out separately.

12. The necessary virus and utensils, as well as explicit directions for use, will be furnished by us.

13. We shall see that your first inoculation is attended by our representative.

(Signed) ————

Darmstadt, Nov. 16, 1903.