IE48627B1 - Injectable composition for the treatment,of helminthiasis and clostridial diseases in animals - Google Patents

Abstract

Injectable compositions comprise tetramisole or its laevorotatory isomer and a vaccine preferably Clostridial vaccine or mixture of (i.e. a multivalent). Clostridial vaccines and are used for the treatment of sheep and cattle to combat helminthiasis and Clostridial diseases.

Description

This invention relates to a new composition comprising the imidazo/2,l-h7thiazole tetramisole and a vaccine and to a process for its use in non-human,warmblooded animals; in particular it relates to a stable composition suitable for administration by injection for use in the treatment of helminthiasis and Clostridial diseases in warm-blooded animals.

D,L-2,3,5,6-tetrahydro-6-phenylimidazo/2,1-b/thiazole, hereinafter referred to as D,L-tetramisole, and its pharmaceutically acceptable acid addition salts are potent anthelmintic agents and the laevorotatory isomer, hereinafter referred to as L-tetramisole, is responsible for all or most of the anthelmintic activity, at least in ruminants such as sheep or cattle. One of the pre15 ferred methods for administration of tetramisole is by subcutaneous injection of an aqueous solution and our Australian Patents No 440,746 and 450,036 disclose aqueous formulations suitable for administration by injection.

Vaccines are widely used to protect warm-blooded animals from a wide range of diseases. Anaerobic vaccines have proved to be of considerable importance in the prevention of a number of common diseases of domestic animals such as cattle, sheep, pigs and fowl. Of parti25 cular importance have been anaerobic vaccines for the prevention of Clostridial diseases such as, for example. Pulpy Kidney (Enterotoxaemia), Blackleg, Malignant Oedema (blood poisoning), Tetanus and Black disease, in sheep and cattle.

Although in the past both tetramisole and vaccines have been administered to animals by subcutaneous injection they have always been administered separately because it was hitherto believed to be impossible to combine a vaccine and tetramisole in a stable aqueous formulation suitable for injection because of the - 3 different stability requirements for aqueous tetramisole formulations and vaccine formulations.

Tetramisole readily undergoes base-catalysed hydrolysis to an inactive derivative and as a result aqueous tetramisole solutions are adjusted to an acid pH to prevent loss of the active ingredient. The aqueous formulations disclosed in Australian Patents No 440,746 and 450,036 are adjusted to a pH of less than 4, preferably approximately 3.5, to provide the formulations with the required storage stability.

In contrast, it is well known that in order to maintain their activity vaccines should not be subjected to a pH of less than 6.0 or more than 7.0 and that as a general rule vaccines are unstable at low pH conditions which promote the denaturing of proteins. For example, -J R Hepple in International Symposium on Adjuvants of Immunity, Utreckt 1966; Symp. Series Immunobiol. Standard, Vol 6 pp. 173-180, Karger, Basel/New York 1967, reports that with Clostridial vaccines it is important to maintain the pH in the range 6.1 to 6.4. Too high a pH results in desorption of the antigen from the carrier while at low pH's denaturing of the antigens can occur, Clostridium perfringens type B and Clostridium septicum being sensitive to pH values below 6.0.

Furthermore, when formulated as an aqueous injectable solution tetramisole is preferably in the form of the hydrochloride, citrate, tartrate or more preferably the dihydrogen phosphate salt and may be accompanied by water soluble therapeutically acceptable salts, particularly the sodium or potassium salts of citric, tartaric or phosphoric acid, in order to prevent or reduce adverse tissue reaction at the site of injection. Vaccines on the other hand are in general incompatible with certain anions such as citrate, phosphate and sulfate because these anions can cause the elution of the - 4 antigen from the carrier adjuvant to which it is reversibly bound, thereby inactivating the vaccine or reducing its storage stability.

The abovementioned apparently incompatible formu5 lation requirements for injectable compositions of tetramisole and vaccines has meant that tetramisole has not previously been combined with a vaccine in the one formulation suitable for administration to warm-blooded animals by subcutaneous injection. We have now surpris ingly found that tetramisole and vaccines may be combined in one formulation suitable for subcutaneous injection without adversely affecting either the efficacy or the stability of the tetramisole or the vaccine.

Accordingly, the invention provides an acidic aqueous composition, which is therapeutically acceptable to warm blooded animals by injection, said composition comprising a tetramisole salt or a laevorotatory tetramisole salt and a Clostridial vaccine.

Preferably the tetramisole is in the form of a salt of the laevorotatory isomer (L-tetramisole or levamisole) .

Suitable L-tetramisole salts include the hydrochloride, acetate, citrate, tartrate and phosphate salts Preferably the tetramisole is in the form of the L25 tetramisole dihydrogen phosphate salt.

Suitable vaccines include the Clostridial vaccines described in the caper by J R Hepple referred to above and the references cited therein. Such vaccines include, for example, those which contain antigens crecared from strains of Clostridia such as Clostridium welchii (Clostridium perfringens) types B, C and D), Clostridium septicum, Clostridium tetani, Clostridium chauvoei and - 5 Clostridium novyi (Clostridium oedematiens) type B which are used in the treatment of Lamb dysentery, Pulpy Kidney disease (enterotoxaemia), Malignant Oedema (blood poisoning), Tetanus, Blackleg disease and Black disease, and mixtures of two or more of those antigens.

As hereinbefore discussed, aqueous tetramisole solutions are preferably adjusted to an acid pH to prevent the hydrolysis of the tetramisole to an inactive derivative. Accordingly, the injectable compositions of the invention are preferably adjusted to an acid pH in the range from 2.0 to 6.0, and more preferably to a pH in the range from 3.0 to 4.0, by the addition of an acid having a therapeutically acceptable anion such as, for example, hydrochloric, tartaric, citric, or preferably phosphoric acid.

The injectable compositions of the invention may also comprise therapeutically acceptable salts such as, for example, the sodium salts of citric, tartaric or phosphoric acid or mixtures thereof in order to prevent or reduce the incidence of adverse tissue reaction. If present, the therapeutically acceptable salts are preferably at a concentration equivalent to from 0.1 to 0.15 moles per litre of solution.

Vaccines are normally prepared and stabilized in the presence of additives known as vaccine adjuvants.

Thus the injectable compositions of the invention preferably comprise pharmaceutically acceptable adjuvants and/or preservatives including antigen carriers.

Suitable adjuvants include potassium alum, protamine, aluminium phosphate, aluminium hydroxide, calcium phosphate, glycerol, sorbitol, propylene glycol, carboxyvinyl polymers available under the Trade Mark Carbopol and bearing the designation 934, 940 and 941, Freund’s universal adjuvant, soluble diethylaminoethyl (DEAE) dextran, saponin, Quil-A, sodium chloride - 6 solution, and the fixed oils and synthetic esters of higher fatty acids which are known to be effective adjuvants .

Suitable preservatives include phenol, formaldehyde, propylene glycol, glycerol, esters of ]3-hydroxybenzoic acid, benzoic acid and its sodium salt, hexachlorophene, quaternary germicides and thiomersal as such or in the form in which it is available under the Trade Mark Merthiolate.

In preparing the injectable compositions of the invention it has been found preferable to formulate the vaccine and tetramisole components separately in the normal manner and then to mix the two components and then to adjust the pH of the combined vaccine - tetrami15 sole composition before storage and subsequent use. In contrast to all expectations it has been found that the injectable compositions of the invention prepared in this fashion and adjusted to an acid pH do not precipitate vaccine toxoids and the activity of both the vaccine component and the tetramisole component is maintained on storage.

In view of the known incompatible requirements for the formulation on the one hand of stable aqueous compositions of tetramisole and on the other hand stable vaccine compositions the present invention of a stable combined tetramisole - vaccine composition is completely unexpected. Moreover, it should be noted that the compositions of the invention are not merely stable for a short period of time after preparation. The efficacy of the compositions is unimpaired after long storage under the conditions normally employed to store vaccines.

The injectable compositions of the invention comprising a (L-) tetramisole salt and a Clostridial vaccine have been found to be effective in killing helminths in - 7 warm-blooded animals and in vaccinating said animals against Clostridial diseases when administered parenterally to said animals. Accordingly in a further aspect the invention provides a process for combatting helminthiasis and Clostridial diseases in non-human, warm-blooded animals by the parenteral administration of a therapeutically effective amount of a composition comprising a tetramisole salt or a laevorotatory tetramisole salt and a Clostridial vaccine.

The term parenteral is used herein to mean intravenous, intramuscular and subcutaneous injection. Preferably the compositions are administered according to the process of the invention by subcutaneous injection.

The process of the invention is particularly suitable for the treatment of large farm animals such as sheep and cattle. However, the process may also be used to treat helminthiasis and Clostridial diseases in other domestic, farm and laboratory animals.

It will be evident to those skilled in the art that the process of the invention offers the advantage of being able to treat a warm-blooded animal with an anthelmintic and a vaccine in the one operation with important savings in labour costs. This advantage may be put to particular benefit in the treatment of pregnant ewes before lambing. In the past, the conventional procedure has been to treat pregnant ewes with an anthelmintic 4 to 6 weeks before lambing and then to treat the ewes with a Clostridial vaccine 2 weeks before lambing. It has now been found that these two operations can be combined by treating pregnant ewes with a composition of the invention comprising a (L-) tetramisole salt and a Clostridial vaccine 4 to 6 weeks before lambing.

When formulated into a composition of the invention no loss in activity has been observed in either - 8 the tetramisole component or the vaccine component. Therefore, the compositions are preferably formulated to contain, in a suitable dosage volume, the dose of (L-) tetramisole and the dose of vaccine usually employed in the treatment of that particular animal when the (L-) tetramisole and the vaccine are parenterally administered separately, as single therapeutic agents.

Such dose rates vary with the animal being treated and the specific (L-) tetramisole salt and vaccine being used. However, in general L-tetramisole is administered at a dose rate of approximately 5 to 10 mg (calculated as the free base) per kilogram of animal bodyweight, D,life tramisole is administered at a dose rate of approximately 10 to 17 mg (calculated as the free base) per kilogram of animal body weight and in general vaccine preparations have been standardized to a dose volume of 2 ml for sheep and 4 ml for cattle for mono-, di-, tritetra- and multivalent vaccines.

In combatting diseases by vaccination it is usual to administer two doses of vaccine the second dose being administered at least four weeks after the first dose. Thus in order to optimize the protection afforded by the vaccine component of the composition of the invention it is preferable to repeat the parenteral administration of a therapeutcially effective amount of the composition at least four weeks later.

The compositions of the invention may comprise, in addition to the components hereinbefore defined: other pharmaceutically therapeutic agents such as, for example, flukicides, selenium (to combat white muscle disease) and systemically active pesticides; additives to improve the shelf life of the composition; buffering agents; preservatives; and/or additives to prevent or to reduce adverse tissue reaction at the site of the injection. - 9 The invention is now illustrated by, but by no means limited to, the following Examples.

Example 1 In order to evaluate the stability and the efficacy 5 of the tetramisole-clostridial vaccine compositions of the invention injectable compositions Aj, Aj and A^ were prepared by admixture of a 7 component Clostridial vaccine comprising antigens prepared from Clostridium welchii Type B, Clostridium welchii Type C, Clostridium welchii, Type D, Clostridium septicum, Clostridium tetani, Clostridium chauvoei and Clostridium novyi Type B /available from ICI Tasman Limited under the name Tasvax 7 (Tasvax is a Trade Mark// and an aqueous sterile filtered solution of L-tetramisole dihydrogen phosphate (17.6% w/v free base) and adjustment of the pH of the -composition to the required level before storage and subsequent use. The make up of the test compositions and the control compositions are detailed in Table 1 below: TABLE 1 Composition* No Vaccine Component Volume (ml); pH Tetramisole Component Volume (ml); PH pH of CompositionA1 500; 3.75 425; 3.5 3.55 A2 500; 5.35 425,- 5.0 5.3A3 500 6.05 425; 5.9 6.0 Control 1 500 6.3 - - Control 2 - 500; 3.45 - + After formulation each composition was stored at 4° to 6°C in a glass bottle * L-tetramisole dihydrogen phosphate of free base concentration 6.8% w/v. - 10 Example 2 Each of the compositions and control compositions detailed in Table 1 above was tested for efficacy by injection into 2 sheep following the dosing schedule detailed in Table 2 below.

TABLE 2 Day Operation 0 4 43 46 57 Compositions formulated a) Sheep tagged; Blood serum sample taken; Faecal egg count made b) Sheep injected as follows; Composition A^ 3.5 ml Composition A2 3.5 ml Composition A^ 3.5 ml Control composition 1 2.0 ml Control composition 2 4.0 ml Faecal egg count made a) Faecal egg count made b) Injections given on Day 1 repeated Faecal egg count made Blood serum sample taken 48637 - 11 The anthelmintic activity of the test compositions was evaluated by measuring the faecal egg count of the sheep before and after treatment and the results are presented in Table 3 below. It should be noted that the sheep were reinfected by helminths between treatments. TABLE 3 Test Composition Faecal Egg Count Day 1 Day 4 Day 43 Day 46A1 500 0 ·* 1200 0A1 200 0 1600 0A2 350 0 1100 0A2 100 0 800 0A3 50 0 1900 0A3 150 0 3350 0 Control 1 100 200 1350 700 Control 1 750 1100 850 1100 Control 2 750 0 900 0 Control 2 350 0 2450 0 - 12 The antigenicity of the vaccines was evaluated by assaying the blood serum samples taken at day 57 for Clostridium perfringens Type C (Clostridium welchii Type C; common name - lamb dysentery); Clostridium perfringens Type D (Clostridium welchii Type D; common name - pulpy kidney); Clostridium novyi Type B (Clostridium oedematiens Type B; common name - Black disease) and Clostridium tetani (common name - tetanus) antitoxins using conventional assay methods using mice.

The results are presented in Table 4 below.

TABLE 4 Test Compositions Antitoxin Titre+ (units/ml) LD PK BD TETA1 >27 13-16 >13.5 >13.5A1 13-27 32-40 8-13.5 8-13A2 13-27 13-16 >13.5 >13A2 <5 3.2 3-8 <3A3 13-27 20-28 8-13 3-8A3 5-13 6.6 3-8 3-8 Control 1 <5 20-27 8-13.5 8 Control 1 5-13 20-27 8-13.5 8-13 Control 2 <5 <0.7 <3.3 - Control 2 <5 2-3 <3.3 — + LD - lamb dysentery PK - pulpy kidney BD - Black disease TET - tetanus Example 3 After storage for 6 months at a temperature of 4 °C the compositions A^ and Aj and control compositions Control 1 (cp and Control 2 (C2)» prepared as detailed in Example 1, were tested for efficacy by injection into sheep following the dosing schedule detailed in Table 5 below. Each composition was tested on 40 sheep with a control group of six sheep not being treated.

TABLE 5 Day Operation a) Blood serum sample taken b) Group A^ sheep injected with 3.5 ml of Composition A^ Group Aj sheep injected with 3.5 ml of Composition A^ Group C1 sheep injected with 2.0 ml of Composition C^ Group C2 sheep injected with 4.0 ml of Composition C2 a) Blood serum sample taken b) Injections given on Day 1 were repeated Blood serum sample taken Blood serum sample taken The antigenicity of the vaccines was evaluated by assaying the blood serum samples taken at days 56 and 98 for Clostridium perfringens Type D (Clostridium welchii - 14 Type D; common name - pulpy kidney), Clostridium septicum (common name - malignant oedemia), Clostridium novyi Type B (Clostridium oedematiens Type B; common name - Black disease), Clostridium tetani (common name 5 tetanus) and Clostridium perfringens Type C (Clostridium welchii Type C; common name - lamb dysentery). The antitoxin titres on the pooled serum samples from the 40 sheep in each group were determined by conventional assay methods using mice. The results are presented in Table 6 TABLE 6 Test Compo- sition Day of Bleed 4. Antitoxin Titre (units/ml) PK MO BD TET LDA1 56 16-20 13.3-16 27-32 20-26.7 26-32A1 98 3.2-4.0 2-4 4-5.3 3.2-4 2-4A3 56 6.65-8.0 5.3 10-13.3 13.3-16 10-13.3A3 98 1-2 0.67-1.0 1-2 1-2 0.67C1 56 5-6.65 4 20-27 13.3 8-10C1 98 0.67-1.0 0.67-1.0 2 1-2 Q.67-1.0C2 56 <0.67 <0.67 <0.67 <0.67 <0.67 On- treate 56 d <0.67 <0.67 <0.67 <0.67 <0.67 + PK - pulpy kidney MO - malignant oedemia BD - Black disease TET - tetanus LD - lamb dysentery 4863? - 15 The only meaningful test available to determine the effectiveness of the vaccination against Clostridium chauvoei (blackleg) is by direct challenge with a living culture or spore suspension.

After completion of the dosing and bleeding schedule in Table 5 above five sheep from each of the groups injected with Test Compositions A^ and were injected intramuscularly with 2 ml of an 18 hour culture of Clostridium chauvoei (strain F6028) containing 2.5% w/v calcium chloride solution. Two of the untreated controls were injected with one tenth, and one of the untreated controls was injected with one hundredth, of the dose injected into the vaccinated sheep.

Within 24 hours of the challenge with Clostridium chauvoei the three unvaccinated controls had died while the ten vaccinated sheep survived the direct challenge. 'Example 4 This Example demonstrates the preparation of compositions of the invention comprising a 5 component Clostridial vaccine and L-tetramisole dihydrogen phosphate.

A pentavalent Clostridial vaccine (Control 3) comprising antigens prepared from Clostridium welchii type D, Clostridium chauvoei, Clostridium septicum, Clostridium oedanatiens and Clostridium tetani (500 carts by volume containing 0.53 standard dose units per part) was combined with an aqueous solution of L-tetramisole dihydrogen phosphate (400 parts by volume containing 15.3% w/v L-tetramisole calculated as the free base) and the pH of the resulting composition was adjusted to 3.5 by the addition of phosphoric acid. The composition (code number A^) contained 0.29 standard dose units of vaccine per ml and 68 mg per ml of L-tetramisole (calculated as the free base).

The above procedure was repeated by combining the pentavalent Clostridial vaccine (Control 3; 500 narts by volume - 16 containing 0.53 standard dose units per ml) with an aqueous solution of L-tetramisole dihydrogen phosphate (350 parts by volume containing 18.2% w/v L-tetramisole calculated as the free base) and the pH of the resulting composition was adjusted to 3.5 by the addition of phosphoric acid.

The composition (code number Ag) contained 0.31 standard dose unit of vaccine per ml and 75 mg/ml of L-tetramisole (calculated as the free base).

The above procedure was repeated by combining the 10 pentavalent Clostridial vaccine (Control 3; 500 parts by volume containing 0.53 standard dose units per ml) with an aqueous solution of L-tetramisole dihydrogen phosphate (560 parts containing 11.4% w/v L-tetramisole calculated as the free base) and the pH of the solution was adjusted to 3.5 by the addition of phosphoric acid. The com.position (code number Ag) contained 0.25 standard dose units of vaccine per ml and 60.2 mg/ml of L-tetramisole (calculated as the free base).

Example 5 This Example demonstrates the preparation of a composition of the invention comprising a 5 component Clostridial vaccine and the hydrochloride salt of Ltetramisole.

A pentavalent Clostridial vaccine (Control 3) com25 prising antigens prepared from Clostridium welchii type D, Clostridium chauvoei, Clostridium septicum, Clostridium oedematiens and Clostridium tetani (500 parts containing 0.53 standard dose units per part) was combined with an aqueous solution of the hydrochloride salt of L30 tetramisole (560 parts by volume containing 11.4% w/v L-tetramisole calculated as the free base) and the pH of the solution was adjusted to 3.5 by the addition of hydrochloric acid. The composition (code number Ay) contained 0.25 standard dose units of vaccine per ml and 60.2 mg/ml of L35 tetramisole (calculated as the free base). - 17 Example 6 This Example demonstrates the antigenic efficacy of the compositions of the invention after prolonged storage.

Immediately after preparation the antigenicity of composition A^, prepared as described in Example 1, was tested in laboratory rabbits (Test No 1). Each rabbit was injected on day 0 with 2.0 ml of composition A^ and the injection was repeated 42 days later. After 2 weeks (day 56) a blood sample was taken from each rabbit and the antitoxin titres of the pooled samples were determined by conventional assay methods.

After storage for a period of 9 months at a temperature of approximately 4°C the antigenicity of composition A^ was retested (Test No 2) in laboratory rabbits following the procedure described above with the exception that the amount of composition A·^ injected on each of days 0 and 42 was reduced from 2.0 to 1.4 ml (30% reduction in dose) .

The results are presented in Table 7, the code for the antitoxins assayed being the same as that used in Example 3 Table 6.

TABLE 7 Test No Dose Size Antitoxin Titre (units/ml) PK MO BD TET LD 1 2x2 ml 6.6 6.4-8.0 5.3-6.4 6.6-8.0 20-27 2 2x1.4ml 5-6.6 5-6.6 4-5 6.6 6.6-8.0 Example 7 This Example demonstrates the anthelmintic efficacy of the compositions of the invention. - 18 10 To ensure that the compositions were tested in heavily infected animals Merino weaners harbouring a heavy naturally acquired parasitic infection were chosen and further infected with larvae of the species Ostertagia and Trichostrongylus. The nematode infections were allowed to reach maturity and the sheep were divided into four groups of eight animals each, the division being made on the basis of faecal egg count to ensure that the groups had a similar mean infection. The sheep were weighed and treated, on the basis of their weight, with sufficient test composition to ensure a dose rate of 6.0 mg per kg of animal body weight of L-tetramisole base.

The four groups were treated as follows: Group 1 - Control (no treatment) Group 2 - L-tetramisole dihydrogen phosphate (6.0% w/v base) Group 3 - Composition (Example 4) Group 4 - Composition A,- (Example 4) which had been stored for 6 months at a temperature of approximately 4°C.

The compositions were administered by subcutaneous injection into the neck. The animals were slaughtered 4 to 5 days after treatment and the parasites in the abomasum,small intestine, large intestine and lungs were counted.

The total and the mean parasite counts for each Group are presented in Table 8 in which the parasites are coded as follows: H - Haemonchus - Ostertagia TA - Trichostrongylus axei I - Immature parasites TR - Trichostrongylus spp N - Nematodirus spp C - Cooperia oncophora - 19 CH - Chabertia OE - Ocsphogostomum venulosum D - Dictyocaulus (lungworm) TABLE 8a Parasite Count - Abomasum Group Count Parasite H 0 TA I Group 1 Total 1260 26070 16100 9500 Mean 157 3260 2012 1187 % Efficiency - - - - Group 2 Total 0 390 170 1100 Mean 0 98 21 137 * % Efficiency 100 97.1 99.9 88.5 Group 3 Total 0 750 360 950 Mean 0 94 45 119 % Efficiency 100 97.2 97.8 90.0 Group 4 Total 0 800 330 300 Mean 0 100 41 37 % Efficiency 100 1 97.0 98.0 96.9 - 20 TABLE 8b Parasite Count - Small Intestine Group Count Parasite TR N C Group 1 Total 31600 10360 19960 Mean 3950 1295 2495 % Efficiency - - Group 2 Total 30 0 0 Mean 4 0 0 % Efficiency 99.9 100 100 Group 3 Total 30 no 0 Mean 4 14 0 % Efficiency 99.9 99.9 100 Group 4 Total 40 40 0 Mean 5 5 0 % Efficiency 99.9 99.6 100 - 2i TABLE 8c Parasite Count-Large Intestine and Lung Group Count 1 Parasite - Large Parasite - Intestine Lung CH OE D Group 1 Total 0 1364 70 Mean 0 170 9 % Efficiency - - - Group 2 Total 0 1 2 Mean 0 0.12 0.25 % Efficiency - 99.9 97.3 Group 3 Total 0 6 0 Mean 0 0.75 0 % Efficiency - 99.6 100 Group 4 Total 0 2 0 Mean 0 0.25 0 % Efficiency - 99.9 100 1; 48627 - 22 Ex.jjiijj lc 8 'fill:; Example demonstrates the antigenic efficacy of the compositions of the invention in cattle.

Three months old cattle which had not previously 5 been vaccinated were selected and divided into four groups. The cattle in each group were treated on day 0 and again on day 28 by subcutaneous injection with composition Ag (Example 4) or the standard pentavalent vaccine (Control 3) used in the preparation of com10 position Ag (Example 4) as follows: Group 1 - Standard vaccine Control 3; (normal cattle dose) dose 4.0 ml (half normal Group 2 Composition Ag; cattle dose) dose 4.0 ml 15 Group 3 - Composition Ag; dose) dose 8.0 ml (normal cattle Group 4 Composition Ag; dose 16.0 ml (twice normal cattle dose).

Two weeks after the final injection (day 42) a 20 blood sample was taken from each of the animals and the antitoxin titres of the pooled sera samples from each Group were determined by conventional assay methods.

The results are presented in Table 9, the code for the antitoxins assayed being the same as that used in Example 3 Table 6. - 23 TABLE 9 Group Antitoxin Titre (units/ml) PK MO BD TET 1 1.0 2.0 10-13 10 2 4-5.3 4 0.67 6-8 8-10 3 8-10 1-2 20-26.7 20-26.7 4 10-13 2-4 26-32 53-64

Claims (19)

CLAIMS:

1. An acidic aqueous composition which Is therapeutically acceptable to warm blooded animals by injection said composition comprising a tetramisole salt or a laevorotatory tetrami 5 sole salt and a Clostridial vaccine.

2. A composition according to claim 1 wherein said vaccine is a Clostridial vaccine which comprises antigens prepared from Clostridia chosen from Clostridium welchii type B, C lostridium welchii type C, Clostridium welchii type D, 10 Clostridium septicum, Clostridium tetani, Clostridium chauvoei and Clostridium novyi type B or a mixture of two or more of said antigens.

3. A composition according to any one of claims 1 and 2 wherein said tetramisole salt or laevorotatory tetramisole 15 salt is chosen from the hydrochloride, acetate, citrate, tartrate and phosphate salts of laevorotatory tetramisole. - 25

4. A composition according to claim 3 wherein said salt is the hydrochloride salt of laevorotatory tetramisole.

5. A composition according to claim 3 wherein said salt is the dihydrogen phosphate salt of laevorotatory tetramisole.

6. A composition according to any one of claims 1 to 5 inclusive wherein the pH of said composition is in the range from 2.0 to 6.0.

7. A composition according to any one of claims 1 to 6 inclusive wherein the pH of said composition is in the range from 3.0 to 4.0.

8. A composition according to any one of claims 1 to 7 inclusive having a pH in the range of from 3 to 4 and comprising the hydrochloride salt of L-tetramisole and vaccine antigens prepared from Clostridium welchii type B, Clostridium welchii type C, Clostridium welchii type D, Clostridium septicum, Clostridium tetani, Clostridium chauvoei and Clostridium novyi type B.

9. A composition according to any one of claims 1 to 7 inclusive having a pH in the range of from 3 to 4 and comprising the dihydrogenphosphate salt of L-tetramisole and vaccine antigens prepared from Clostridium welchii type B, Clostridium welchii type C, Clostridium welchii type D, Clostridium septicum, Clostridium tetani, Clostridium chauvoei and Clostridium novyi type B.

10. A composition according to any one of claims 1 to 9 inclusive comprising therapeutically acceptable salts chosen from the sodium salts of citric, tartaric and phosphoric acid or mixtures thereof at a concentration equivalent to from 0.1 to 0.15 moles per litre of - 26 solution.

11. A composition according to any one of claims 1 to 10 inclusive comprising pharmaceutically acceptable vaccine adjuvants and/or preservatives. 5

12. A composition according to any one of claims 1 to 11 comprising one or more additional medicinal or therapeutic agents.

13. A process for combatting helminthiasis and Clostridial diseases in noil-human, warm-blooded animals which process 10 comprises the parenteral administration of a therapeutically effective amount of a composition comprising a tetramisole salt or a laevorotatory tetramisole salt and a Clostridial vaccine as defined according to any one of claims 1 to 12 inclusive. 15

14. A process according to claim 13 wherein said warm-blooded animals are sheep.

15. A process according to claim 13 wherein said warm-blooded animals are cattle.

16. A process according to claim 14 wherein said com2o position is administered to pregnant ewes 4 to 6 weeks before lambing.

17. A process according to any one of claims 13 to 16 inclusive wherein said composition is administered by subcutaneous injection. 25 18. A composition according to any one of claims 1 to

12. Inclusive substantially as herein described with reference to any one of Examples 1, 4 or 5.

19. A process according to any one of claims 13 to 17 inclusive substantially as herein described with 30 reference to any one of Examples 2, 3 or 6 to 8 inclusive.