Composition preventing necrotic enteritis in galloanserans

Abstract

The present invention refers to the use of a glycerol ester composition of at least one short chain fatty acid for preventing and/or alleviating necrotic enteritis in the gastric tract of galloanserans. The glycerol ester composition comprises at least 75% by weight of glyceryl tributyrate, below 25% by weight of glyceryl dibutyrate and below 8% by weight of glyceryl monobutyrate. The present invention also refers to the use of the glycerol ester composition for modulating the gut flora of galloanserans.

Claims

1. A therapeutic method for preventing and/or alleviating necrotic enteritis in the gastric tract of galloanserans, the method comprising administering a glycerol ester composition to the galloanserans in dry form, wherein the glycerol ester composition is present on a carrier and the glycerol ester composition comprises: glyceryl tributyrate being present in an amount of at least 75% by weight of the total glycerol ester composition but less than 100% by weight of the total glycerol ester composition, glyceryl monobutyrate being present in an amount of more than 0% and below 8% by weight of the total glycerol ester compositions, glyceryl dibutyrate being present in an amount of more than 0% and below 25% by weight of the total glycerol ester composition, and free butyric acid being present in amount of more than 0% and below 0.5% by weight of the total glycerol ester composition.

2. The method according to claim 1, wherein the necrotic enteritis is caused by Clostridium perfringens.

3. The method according to claim 1, wherein the galloanserans are galliformes.

4. The method according to claim 1, wherein the galloanserans are anseriforms.

5. The method according to claim 1, wherein the glycerol ester composition comprises glyceryl tributyrate being present in an amount of at least 80% by weight of the total glycerol ester composition but less than 100% of the total glycerol ester composition, glyceryl monobutyrate being present in an amount not exceeding 5% by weight of the total glycerol ester composition.

6. The method according to claim 1, wherein the glycerol ester composition comprises glyceryl tributyrate being present in an amount of at least 85% by weight of the total glycerol ester composition but less than 100% of the total glycerol ester composition, and glyceryl monobutyrate being present in an amount not exceeding 4% by weight of the total glycerol ester composition.

7. The method according to claim 1, wherein the glycerol ester composition comprises glyceryl tributyrate being present in an amount of at least 90% by weight of the total glycerol ester composition but less than 100% by weight of the total glycerol ester composition, glyceryl monobutyrate being present in an amount not exceeding 2.5% by weight of the total glycerol ester composition.

8. The method according to claim 1, wherein the amount of free butyric acid in the glycerol ester composition is below 0.2% by weight of the total glycerol ester composition.

9. The method according to claim 1, wherein the carrier comprises porous silica particles.

10. The method according to claim 9, wherein the glycerol ester composition is adsorbed on the silica particles in a weight ratio of 50-80% glycerol ester and 20-50% silica particles.

11. The method according to claim 1, wherein the glycerol ester composition is fed to the galloanserans in an amount of between 0.05 to 1.5% by weight of the galloanserans' daily feed ration.

12. The method according to claim 1, wherein the glycerol ester composition is fed to the galloanserans in an amount of between 0.1 to 1.0% by weight of the galloanserans' daily feed ration.

13. The method according to claim 11, wherein the amount of the glycerol ester composition is administered over time intervals of 5-10 days and then the amount is subsequently decreased over a 1-5 week period.

14. The method according to claim 13, wherein the decrease of the amount is 20-50% of the amount given during the previous interval.

15. The method according to claim 12, wherein the amount of the glycerol ester composition is administered over time intervals of 5-10 days and then the amount is subsequently decreased over a 1-5 week period.

16. The method according to claim 3, wherein the galloanserans are galliformes selected from the group consisting of chicken, turkey, grouse and pheasant.

17. The method according to claim 4, wherein the galloanserans are anseriformes selected from the group consisting of ducks, goose and swan.

18. A therapeutic method for preventing and/or alleviating necrotic enteritis caused by Clostridium perfringens in the gastric tract of galloanserans, the method comprising administering a glycerol ester composition to the galloanserans in dry form, wherein the glycerol ester composition is present on a carrier and the glycerol ester composition comprises: glyceryl tributyrate being present in an amount of at least 75% by weight of the total glycerol ester composition, but less than 100% by weight of the total glycerol ester composition, glyceryl monobutyrate being present in an amount of more than 0% and below 8% by weight of the total glycerol ester composition, glyceryl dibutyrate, and free butyric acid being present in amount of more than 0% and below 0.5% by weight of the total glycerol ester composition.

19. The therapeutic method of claim 18, wherein the glyceryl monobutyrate is present in an amount not exceeding 2.5% by weight of the total glycerol ester composition.

20. The therapeutic method of claim 19, wherein the glycerol ester composition is distributed in a feed to the galloanserans.

21. The therapeutic method of claim 20, wherein the amount of free butyric acid in the glycerol ester composition in the feed is in the range of 0.2% to 0.5% by weight.

22. The therapeutic method of claim 9, wherein the porous silica particles have an average particle size of 20-70 nm.

Description

BRIEF DESCRIPTION OF DRAWINGS

(1) FIG. 1 is a graphic representation of positive chickens (with macroscopic lesion score 2) comparing glyceryl MB low; glyceryl MB high and glyceryl TB to a Control.

EMBODIMENT EXAMPLE

(2) In vivo trial on the efficacy of glyceryl butyrates to control Clostridium perfringens-induced necrotic enteritis in broiler chicken.

(3) Bacterial Strains and Vaccines

(4) The challenge strain used in the in vivo trials, C. perfringens strain 56, was isolated from the gut of a broiler chicken with necrotic lesions from a flock with weight gain problems and has been shown to be highly virulent in earlier in vivo trials. The strain was classified as a type A strain (netB positive, beta-2 and enterotoxin negative) and produces moderate amounts of alpha toxin in vitro (Gholamiandehkordi et al., 2006).

(5) For inoculation, the strain was grown for 24 h in Brain Heart Infusion broth (BHI, Oxoid, Basingstoke, England).

(6) A ten-fold dose of the anticoccidial vaccine Paracox-8 (Schering-Plough Animal Health, Brussels, Belgium), containing live, attenuated oocysts of Eimeria (E.) acervulina (two lines), E. brunetti, B. maxima, E. necatrix, E. praecox, E. mitis and E. tenella was used in this study. Nobilis Gumboro D 78 vaccine (Schering-Plough Animal Health, Brussels, Belgium) was given in the drinking water.

(7) Animals and Housing

(8) In this experiment, 114 broilers of mixed sex were used. They were obtained at 1 day old from a commercial hatchery. Before the trial, all rooms were decontaminated with Metatectyl (Clim'oMedic) and with a commercial anticoccidial disinfectant (Bi-OO-Cyst; Biolink Ltd, York, UK). The birds were divided in 4 cages of 1.5 m.sup.2, on wood shavings. They were given drinking water and feed ad libitum. A 23 h/1 h light darkness program was applied.

(9) Experimental Study Design:

(10) The first 7 days, the chickens were fed a starter diet and from day 8 until 15, a grower diet. Both the starter and the grower diet were a wheat/rye (43%/7.5%) based diet, with soybean meal as protein source. From day 17 onwards, the same diet was used with the exception that fishmeal (30%) was used as a protein source. The feed was provided by the Institute for Agricultural and Fisheries Research (ILVO). The tested products were mixed in the feed. The Gumboro vaccine was given in the drinking water at day 16 in all groups. All groups were challenged orally one time a day with approximately 4.10.sup.8 cfu C. perfringens strain 56 at days 17, 18, 19 and 20. At day 18 all birds were orally inoculated with a ten-fold dose of Paracox-8. At days 21, 22 and 23, 9 animals of each group were euthanized by intravenous sodium pentobarbital injection.

(11) Model:

(12) TABLE-US-00001 TABLE 1 Time schedule for the in vivo study d16 d17 d18 d19 d20 d21 d22 d23 Gumboro x Feed + fishmeal x x x x x x x Inoculation C. perfringens x x x x Paracox 10 x Scoring x x x

(13) Products Tested: Pen 1: positive control Pen 2: glyceryl monobutyrate (adsorbed on a silica carrier), added to the feed at a lower concentration Pen 3: glyceryl monobutyrate (adsorbed on a silica carrier), added to the feed at a higher concentration Pen 4: glyceryl tributyrate (adsorbed on a silica carrier)

(14) The products were added to the feed at the concentrations shown in Table 2 below:

(15) TABLE-US-00002 TABLE 2 Concentrations of products added to the feed (kg/ton) Glyceryl Glyceryl monobutyrate, monobutyrate, Glyceryl on silica carrier on silica carrier tributyrate, low conc high conc on silica carrier Week 1 5 7.5 5 Week 2 2.5 6 2.5 Week 3 2 5.5 2

(16) Intestinal lesions in the small intestine (duodenum to ileum) were scored blinded as follows;

(17) 0: no gross lesions

(18) 1: congested intestinal mucosa

(19) 2: small focal necrosis or ulceration (1-5 foci)

(20) 3: focal necrosis or ulceration (5-16 foci)

(21) 4: focal necrosis or ulceration (16 or more foci)

(22) 5: patches of necrosis 2-3 cm long

(23) 6: diffuse necrosis typical of field cases

(24) Lesion scores of 2 or more were classified as necrotic enteritis positive

(25) Statistical Analysis

(26) The GraphPad Prism Software, Inc was used to determine whether there were significant differences between groups. Statistical significance was determined at a P value of <0.05.

(27) Clinical Observations

(28) No abnormal clinical observations were observed.

(29) 3 chickens died during the trial. 1 chicken died in pen 1 2 chickens died in pen 3

(30) Table 3 shows the number of birds with necrotic enteritis lesions for each group, at day 21, day 22 and day 23. Also, the total number of birds with lesions per group is shown.

(31) TABLE-US-00003 TABLE 3 Number of birds with macroscopic necrotic enteritis lesions on the three sampling days. glyceryl glyceryl pos monobutyrate, monobutyrate, glyceryl control low high tributyrate day 21 9/9 5/9 4/9 3/9 day 22 4/9 3/9 5/9 2/9 day 23 4/10 2/10 3/8 2/11 total 17/28 10/28 12/26 7/29 Total (%) 61 36 46 24

(32) The percentage of positive chickens (with macroscopic lesion score=2) is shown in FIG. 1.

(33) The results in FIG. 1 states that adding glyceryl tributyrate to the feed gives a significantly better result, with respect to preventing Clostridium perfringens-induced necrotic enteritis in broiler chicken, than adding glyceryl monobutyrate at either one of the two concentrations tested.

(34) If the addition of glyceryl monobutyrate at the higher concentration is compared to the addition of glyceryl tributyrate, it is about the same weight of butyric acid that is added (making the assumption that glycerol and butyric acid have about the same molar weights). Despite this fact, glyceryl tributyrate gives a much better result (24% positive chicken in total for glyceryl tributyrate compared to 46% positive chicken in total for glyceryl monobutyrate at the higher concentration).

(35) It is also remarkable that adding a lower concentration of glyceryl monobutyrate gives a better result than adding a higher concentration (36% positive chicken in total for glyceryl monobutyrate at the lower concentration compared to 46% positive chicken in total for glyceryl monobutyrate at the higher concentration).

(36) The results show that using a glycerol ester composition according to the present invention is an effective way of preventing Clostridium perfringens-induced necrotic enteritis in galloanserans.

(37) The embodiment example is to be construed as illustrative and not limiting in any way. The glycerol ester composition may for example comprise a blend of glycerol esters of different short chain fatty acids, such as butyric acid, propionic acid and valeric acid. The dosage levels will then have to be calculated with respect to the total amount of glycerol esters.

(38) It may further prove to be useful to feed the glycerol ester composition to the animal for a shorter or longer period of time and to extrapolate the dosage levels accordingly.