Field

The present invention relates to methods and means for reducing and/or inhibiting the occurrence of footpad dermatitis in birds.

Background

The problem of footpad dermatitis (FPD) has long been recognized in the field. Large scale production of poultry inevitably leads to birds suffering from FPD. FPD is a type of contact dermatitis which primarily affects the footpad of the feet and the skin on the hock joint. Birds acquire FPD when they are standing on litter that is not exchanged for weeks at a time. The litter becomes increasingly wet as the bird droppings are not removed but accumulate in the bedding. Over time, the alkaline moisture harms the birds' feet and lesions of various severity accumulate. FPD in an early stage results in hyperkeratosis, erosions and discoloration of the skin. The erosions can develop into ulcers. In severe cases, thefootpad lesions may cause pain which together with a deteriorated state of health constitutes a welfare issue. It has also been indicated that broilers with severefootpad dermatitis show slower weight gain.

At the time of slaughter, the severity of the dermatitis is measured on a scale of 0 to 7 (Mayne et al., 2007a). In slaughterhouse there is general practice to divide the footpads into two groups: ones with some necrosis or lesions and footpad without any lesions (score 0-3). From score 4-5 (using score system 0-7) paws cannot be sold.

Paw scoring depends on consumer specifications. Some slaughterhouses use an A, B, C, or condemn scale, whereas others use A, B, or condemn grade. Grading is based on the size of the lesion on the pad from FPD, discoloration, mutilations from processing, and also trauma injuries that may occur during catching and live haul such as broken toes. Roughly 99% of the downgrades come from FPD lesions, with the other 1% encompassing both catching and live haul injuries and processing mutilations.

Recently, chicken paw prices have escalated due to an insatiable demand for high-quality paws in export markets. This demand has turned paws into the third most important economic part of a chicken, behind the breast and wings, with paws accounting for approximately $280 million a year.

Solutions to prevent FPD

Drinker design can play an important role in the overall moisture of the litter and thus the occurrence of FPD. Turkeys that are provided with small water cups have been shown to have a lower occurrence of FPD than bell drinkers (Ekstrand and Algers, 1997). Bray and Lynn (1986) found that nipple drinkers with drip cups were most efficient and resulted in better litter conditions than nipple drinkers alone and bell drinkers. Drinkers that are too low or have the water pressure set too high tend to result in wetter floors. Water lines that may have a biofilm or other particulates can result in leaky drinkers, which will result in increased litter moisture. Regular flushing and sanitizing of the water lines will reduce water leakage. This will keep litter drier and improve its quality, subsequently resulting in better paw and hock quality (Tucker and Walker, 1999; Mayne et al., 2007b).

Further known measurements are directed at reducing the moisture of the litter, and include the ventilation rate, stocking rate and feed composition such as reduced protein content and reduced levels of minerals such as sodium, potassium and magnesium. McGinnis and Carver (1947) found that riboflavin supplementation into turkey diets prevented dermatitis in poults. Footpad dermatitis in young poults has been associated with methionine deficiency. Extra methionine and zinc are known to reduce FPD, while biotin and riboflavin deficiencies increase FPD.

Zinc compounds, including chelated zinc compounds, methionine in combination with biotin are known in the field to have beneficial effects on skin. Youssef et al. (2011) described the effects of high dietary biotin or zinc on FPD. In this article, it was found that high dietary biotin and high dietary zinc independently reduced the incidence of FPD when turkeys were housed on dry litter, but not when they were housed on wet litter. In a study by Harms and Simpson (1977), supplemental biotin resulted in significantly reduced footpad scores when given to poults grown on dry litter but not when given to poults grown on wet litter.

There is thus a clear need for methods and means that reduce and/or prevent the occurrence of food pad dermatitis in poultry such as turkey and chicken, especially when the birds are housed on wet litter.

The present inventors surprisingly found that the addition of zinc bismethionine (1:2 zinc methionine complex) at between 50 and 150 mg of Zn/kg complete feed with a moisture content of 12 %, preferably at about 150 mg of Zn/kg complete feed with a moisture content of 12 %, led to a strong reduction of the incidence of FPD in poultry when the birds were housed on wet litter. Thus, in contrast to results obtained with zinc monomethionine (Youssef et al. 2011), zinc

bismethionine leads to a strong reduction of the incidence of FPD in poultry even when the birds are housed on wet litter. This reduction was evident in the presence of normal dietary levels of biotin (for chicken 300 μg/kg complete feed, for turkey 800 ug/kg) (moisture content 12%), and was enhanced in the presence of high dietary levels of biotin (2000 μg/kg complete feed (moisture content 12%)). Therefore, the invention provides an animal feed having between 50 and 150 mg of Zn/kg complete animal feed with a moisture content of 12 %, comprising zinc bismethionine. It is preferred that at least 80% of the total amount of zinc in the animal feed is provided in the form of zinc bismethionine, more preferred at least 90%, more preferred at least 95%, most preferred at least 99%.

The term "zinc bismethionine" refers to a chelate complex of zinc and

methioninate in a 1:2 molar ratio (C5H10NO2S)2Zn), having a molecular weight of 361.8 g/mol. The present inventors further found that high dietary zinc in the form of zinc bismethionine in combination with high dietary biotin leads to a reduction of the incidence of FPD in poultry by 30 to 50% when the birds were housed on wet litter. Especially the two highest levels of a standard scale of FPD (Mayne et al., 2007a) were strongly reduced or even absent when animals were fed with high dietary zinc in combination with high dietary biotin. In two separate trials, the incidence of animals with severe footpad lesions having the most severe scores of 6 and 7 was reduced from a total of 19/99 chicken to 1/97 chicken by feeding the birds with animal food comprising high zinc (150 mg Zn/kg complete feed

(moisture content 12%)) and high biotin (2000 μg biotin/kg complete feed

(moisture content 12%)).

The invention therefore provides an animal feed, preferably a bird feed, having between 50 and 150 mg Zn/kg complete feed (moisture content 12%), comprising zinc bismethionine, further comprising between 1000 and 3000 μg biotin/kg complete feed (moisture content 12%)). An animal feed according to the invention preferably comprises about 2000 μg biotin/kg complete feed (moisture content 12%)).

The zinc in the animal feed of the invention preferably comprises about 150 mg Zn/kg complete feed (moisture content 12%), such as about 120 mg Zn/kg complete feed (moisture content 12%), about 130 mg Zn/kg complete feed

(moisture content 12%), about 140 mg Zn/kg complete feed (moisture content 12%), and 150 mg Zn/kg complete feed (moisture content 12%). Methods for determining the relative amounts of zinc in an animal feed are known in the art and include DIN EN ISO 11885:2009-09.

It was found that high dietary intake of zinc as zinc-bismethionine efficiently reduces the incidence of FPD in poultry, especially when combined with high dietary biotin. Methods for producing zinc-bismethionine are known in the art. For example, DL-methionine and zinc oxide are dissolved in water at a molecular ratio of 2:1 by adding diluted hydrochloric acid (25 % w/v). After all raw materials have been dissolved completely, the pH of the solution is adjusted to 7.2 by adding a solution of sodium hydroxide in water (20 % w/v) while constantly stirring. The physical complex of methionine-zinc precipitates and is separated from the solution by filtering. Technically pure zinc-bismethionine is obtained after drying at 105 °C and contains 18.0% zinc and 78.0% methionine. Zinc- bismethionine is not classified as dangerous according to Directive 67/548/EEC.

The term "high dietary zinc" refers to the amount of zinc bismethionine that is provided to an animal, preferably a chicken or a turkey, when the animal is fed with an animal feed comprising between 50 and 150 mg Zn/kg complete feed (moisture content 12%), preferably about 150 mg Zn/kg complete feed (moisture content 12%). It is preferred that at least 80% of the total amount of zinc in the animal feed is provided in the form of zinc bismethionine, more preferred at least 90%, more preferred at least 95%, most preferred at least 99%.

The term "high dietary biotin" refers to the amount of biotin that is provided to an animal, preferably a chicken or a turkey, when the animal is fed with an animal feed comprising between 1000 and 3000 μg biotin/kg complete feed (moisture content 12%), preferably about 2000 μg biotin/kg complete feed

(moisture content 12%), or when the animals are allowed to drink water comprising between 500 and 1500 μg biotin/kg, preferably about 750 mg biotin/l.

The term "about 2000 μg biotin/kg complete feed (moisture content 12%)" refers to an animal feed comprising about 1600 μg biotin/kg complete feed (moisture content 12%), about 1700 μg biotin/kg complete feed (moisture content 12%), about 1800 μg biotin/kg complete feed (moisture content 12%), about 1900 μg biotin/kg complete feed (moisture content 12%), about 2100 μg biotin/kg complete feed (moisture content 12%), about 2200 μg biotin/kg complete feed (moisture content 12%), about 2300 μg biotin/kg complete feed (moisture content 12%), and about 2400 μg biotin/kg complete feed (moisture content 12%). Methods for determining the relative amounts of biotin in an animal feed are known in the art and include DIN EN 15607. The term "animal", as used herein, refers to domesticated animals such as cattle, pigs, birds, sheep and goats. The term "birds", as used herein, refers to poultry and includes chicken, duck, emu, goose, peafowl and turkey.

The term "poult", as is used herein, refers to a bird, especially a young bird such as a chicken or a turkey of less than 12 months.

The term "animal feed", as used herein, refers to fodder and, preferably, compound feed for domesticated animals. The term "animal feed" preferably refers to compound feed for birds, such as chicken and turkey.

An animal feed according to the invention is preferably provided to an animal such as chicken and turkey. These birds are routinely kept under conditions that their litter becomes wet. Birds that are kept on litter that is wettened during the keeping of the birds often develop food pad dermatitis, which can be reduced or prevented by providing the birds with high dietary zinc bismethionine, or a combination of high dietary zinc bismethionine and high dietary biotin.

Therefore, the invention provides an animal feed according to the invention, for use in a method to reduce and/or prevent the occurrence of FDP in birds. The invention also provides the use of an animal feed according to the invention, for the manufacture of a medicament for the reduction and/or prevention of the occurrence of FDP in birds. The invention also provides a method for reducing and/or preventing the occurrence of FDP in birds, comprising providing an animal feed according to the invention to the birds.

The incidence of FDP increases when birds are housed on wet litter. The materials that are routinely used as bedding material for birds comprise wood shavings, sawdust, peanut hulls, shredded sugar cane, straw, and/or other dry, absorbent, low-cost organic materials, including sand. Typically, a layer of new bedding material is deposited when housing is filled with a new batch of birds. Birds such as chicken broilers are usually slaughtered between 28 and 50 days of age, with a mean of approximately 35 days. Birds such as turkey are usually kept for a total of 16-22 weeks. In this period, the bedding material becomes wet because the rate of water addition (urine/faeces/spillage) exceeds the rate of removal (evaporation). The term "wet litter" preferably refers to a moisture content of the bedding material of at least 25%, preferably at least 30%. Methods for determining the moisture content of litter are known in the art and include determining the weight difference of litter before and after oven-drying fresh litter matter at 103°C for approx. 48 hours.

Therefore, the invention provides an animal feed according to the invention, for use in a method to reduce and/or prevent the occurrence of FDP in birds when they are housed on wet litter.

The invention additionally provides a feed supplement comprising zinc bismethionine or comprising zinc bismethionine and biotin that, when mixed with animal feed, provides between 50 and 150 mg/kg complete feed (moisture content 12%) of zinc, or between 50 and 150 mg/kg complete feed (moisture content 12%) of zinc and between 1000 and 3000 μg/kg complete feed (moisture content 12%) of biotin, respectively. The feed supplement may conveniently be provided to farmers, to mix it with normal animal feed to arrive at the indicated levels of zinc or of zinc and biotin. Said feed supplement preferably comprises instructions for mixing or diluting the supplement into the animal feed to arrive at levels of between 50 and 150 mg/kg complete feed (moisture content 12%) of zinc, preferably about 150 mg Zn/ kg complete feed (moisture content 12%), or of between 50 and 150 mg/kg complete feed (moisture content 12%) of zinc, preferably about 150 mg Zn/ kg dry feed matter, and between 1000 and 3000 μg/kg complete feed (moisture content 12%) of biotin, preferably about 2000 μg biotin/ kg complete feed (moisture content 12%), respectively.

Said feed supplement comprising zinc and biotin can be provided in one container, or in two separated containers. Therefore, the invention also provides feed supplement as a kit of parts, comprising separate containers for zinc and biotin, further comprising instructions to mix the zinc and biotin into the animal food to provide between 50 and 150 mg Zn/kg complete feed (moisture content 12%), preferably about 150 mg Zn/ kg complete feed (moisture content 12%), and between 1000 and 3000 μg/kg complete feed (moisture content 12%) of biotin, preferably about 2000 μg biotin/ kg complete feed (moisture content 12%).

List of references

Bray, T.S., and N.J. Lynn 1986. Effects of nutrition and drinker design on litter condition and broiler performance. Br. Poult.Sci, 27:151-156

Harms, R.H, B.L. Damron, and C.F. Simpson. 1977, Effect of wet litter and supplemental biotin and /or whey on the production of foot pad dermatitis in broilers. Poult.Sci. 56:291-296

Ekstrand, C.,and B. Algers, 1997. Rearing conditions and foot-pad dermatitis in Swedish turkey poults. Acta Vet. Scand. 38:167-174

Mayne, R.W., Else, and P.M. Hocking. 2007a High dietary concentration of biotin did not prevent foot pad dermatitis in growing turkeys and external scores were poor indicators of histoptahological lesions. Br. Poult. Sci.48:291-298 Mayne, R.K., R.W. Else, and P.M. Hocking. 2007b. High litter moisture alone is sufficient to cause footpad dermatitis in growing turkeys. Br. Poult. Sci. 48:538- 545

Tucker, S.A., and A.W. Walker. 1999. Hock burn in broilers. Pages 33-50 in Recent Developments in Poultry Nutrition 2. J. Wiseman and P.C. Garnsworthy, ed. Nottingham University Press, Thrumpton, UK.

McGinnis, J., and J.S Carver. 1947. The effect of riboflavin and biotin in the prevention of dermatitis and perosis in turkey poults. Poult.Sci. 26:364-371

Youssef IM, Beineke A, Rohn K, Kamphues J.2011. Influences of increased levels of biotin, zinc or mannan-oligosaccharides in the diet on foot pad dermatitis in growing turkeys housed on dry and wet litter. J Anim Physiol Anim Nutr (Berl). Article first published online: 4 January 2011

Examples

Example 1

Performed trials

Two trials were performed at Institute of Animal Nutrition, University of

Veterinary Medicine, Hannover

Trial design

In each trial 110 chicken poults of Ross 708 genetic were divided into four groups. Each group was kept in a separate box with a size of 1.51 m x 1.32 m (approx.2 m ² ). Wood shavings served as litter material. During the whole trial no litter refreshment was done. A "critical moisture" value of 35% was achieved by repeated adding of distilled water. In the second trial, animals were placed in different boxes compared to the first trial in order to avoid any environmental effects (Table 1).

Each group received a complete feed ad libitum (day 7-40) which varied only in zinc source and biotin inclusion rate.

Group 1: Zinc 150 mg/kg feed (120 mg as zinc oxide) + biotin 300 μg/kg feed (diet

1)

Group 2: Zinc 150 mg/kg feed (120 mg as zinc oxide) + biotin 2000 μg/kg feed (diet

2) Group 3: Zinc 150 mg/kg feed (120 mg zinc bismethionine complex) + biotin 300 μg/kg feed (diet 3)

Group 4: Zinc 150 mg/kg feed (120 mg zinc bismethionine complex) + biotin 2000 μg/kg feed (diet 4).

Botanical composition and chemical analysis are shown in tables 2 and 3, respectively.

Evaluation of footpad health was done weekly at day 7, 14, 21, 28, 35 and 40. During evaluation scoring system by Mayne et al., 2007a was used. Results of evaluation of footpad health are shown in table 4, table 5 and table 6

Example 2

Performed trials

A trial with turkeys was performed at the Institute of Animal Nutrition,

University of Veterinary Medicine, Hannover on experimental and research Farm Ruthe.

Trial design

In the trial, 2900 one-day old turkeys were divided into four groups. Each group was kept in a separate box with a size of approximately 236 m2. Wood shavings served as litter material (at 7.79 kg/ m2 and at a height of 5-6 cm) in two of the boxes, while straw pellets (at 8 kg/m2 and at a height of 2.5 cm) served as litter material in the remaining two boxes. During the whole trial no litter refreshment was done, and no fresh litter material was added until day 37.

Each group received a complete mixed feed ad libitum (until week 11) which varied only in zinc source and biotin inclusion rate.

The mixed feed composition was based on normal mast practice-related values for different fattening stages and was in principle nearly identical for the control and experimental groups. Only the tested ingredients Zn and Biotin differed: In the control diet, unfortunately consistently slightly lower Zn levels were reached; when using ZnO as the Zn source. The Zn-content varied in the mixed feed of the actual fattening stages (P3-P6) around values of about 110 mg / dry feed, while in the experimental diet (Zn-Met) in general somewhat higher values were found (about 160-180 mg Zn). The biotin concentrations differed - as planned - very significantly: while the analysed values in the control diet varied between 485 to 880 μg / kg dry feed, in the experimental diet a much higher level (1400 to 2600 μg / kg dry feed) was recorded.

The chemical analysis of the mixed feed for the control groups 1 and 2 is shown in table 2.

The chemical analysis of the mixed feed for the experimental groups 3 and 4 is shown in table 3.

Regardless of the food and the type of litter, the litter remained relatively dry in the first weeks (up to 35 days of age). Around the 35th day of age, the determined dryness was quite low for the first time. At the end of the rearing, a quite

"critical" moisture content (i.e. 35% humidity, and more) was measured in all compartments. In all compartments fresh litter was added between day 35 and day 77, which was very effective in relation to the moisture content. From the 80th day onwards, only straw was used as added litter, in all compartments, so that the feed was the only difference between the groups, but not the different litter. The total moisture content of the accumulated litter (excrement litter mixture) at the end of the mast period was determined in all 4 compartments and was found to be nearly identical between 51.4 and 50.1 moisture content.

NH3 concentrations were detectable from the 8th week onwards. Significantly better results were obtained using straw pellets compared to the use of wood shavings (on average 3.63 ppm for straw pellets vs. 7.56 ppm for shavings). Footpad health was determined at days 39, 81 and 142 (see Tables 4-6). The litter remained very dry until the 39th day of life without any addition of litter.

Evaluation of footpad health at day 39 showed a comparable low score of 1.5 for all groups. A moderate change at that time was still the exception and observed in only 3 of 320 animals. At normal ZnO and biotin supplementation, mean score values were 1.70 (n = 160), whereas the Zn-Met and high biotin mean score values were 1.52, corresponding to a reduction of the score of about 10%.

The mean scores were on day 81 were significantly higher (~ 3.8). In the "wood shavings" compartments, mean scores were about 3.77, while the mean score in the "straw pellets" compartments was about 3.93.

When differentiated according to the nutritional supplements, the scores varied between about 3.8 for control animals that had received the control diet, and about 3.9 for animals in the experimental groups whose feed was supplemented with high Zn-Met and biotin -additive. It is worth mentioning that at this age the indicated scores are considered "normal". Severe lesions (score 6 + 7) were not observed.

Between day 81 and the end of the fattening period (day 142) only straw was added to the litter so that all animals can be viewed with respect to the litter management as an identical population, although they were still housed in "their" compartments. Evaluation of footpath health was in this mast phase based on the number of individual legs (300 per feed variant), without

mentioning the exact number of animals (between 150 and 300 animals)

Two main observations were made at the end of the mast period (see Table 4): the use of the control diet (whole mast period only ZnO and conventional biotin supplementation) resulted in mean scores of 5.63 ± 1.23, while the use of a high biotin-feed (2000 mg / kg) and the use of Zn-Met resulted in lower values of 5.06 ± 1.24. This is a decline by 10%. More important, however, is the change in the frequencies of severe footpad lesions. Using the control diet about 58% of the footpads had high grade changes, while such scores were less frequent after using the experimental diet (~ 37% of the footpads).

At slaughter, the mean body mass of the animals was:

- 20.7 kg (control diet with ZnO and normal biotin intake) or

- 21.6 kg (experimental diet with Zn-Met and high biotin intake), which corresponds with a good mast result.

Throughout the feeding trial, the rate of loss of animals amounted to 10.8% (from 1 to 142 days), as is shown in Table 7.

A statistically significant, favorable impact of the special nutritional supplementation resulted from the experimental diet. The mean score was significantly reduced the experimental diet, about 10%. From the aspects of animal welfare, animal health and product image, however, another change is much more significant: the experimental diet reduced the frequency of severe alterations in the footpad from over 58% in the control group to about 37%.

Example 3

Skin permeation testing by high throughput Franz-cell technology in ex-vivo model

Broiler chicks (25 birds per group) were divided into four groups. From 8 days of age, the chicken were fed with diets with different amounts of zinc from different sources (ZnO 80 mg/kg (group 1); ZnMet 1:2 40 mg/kg (group 2); ZnMet 1:2 80 mg/kg (group 3); and ZnMet 1:2 120mg/kg (group 4); whereby the amounts refer to the Zn/kg complete feed) and high biotin inclusion rate (2000 μg/kg).

No litter refreshment was done during the trial. At the end of trial (day 35), the following test was performed for chicks from groups 2 and 4, according to the industrial standard of Franz-cells principle operating in a high-throughput format (Zyleris Pharmatech) to assess skin absorption and penetration of active ingredients in topical formulations. Core of the method is a 108-cell screening platform with 0.5 cm2 diffusion area in each cell and precision mixing and temperature control. During this trial, permeation of flufenamic acid (Sigma-Aldrich; Taufkirchen near Munich) through skin (breast) of broiler chicks was measured after 24 hours (Table 8).

The reduced permeation of breast skin from broiler chicks that were fed with the higher dose of zinc-bismethionine is indicative of the improved skin quality under these conditions.

In addition, a significant improvement of footpad health status was observed at days 14 and 21 for broiler chicks that were fed with the higher doses of zinc- bismethionine, especially 80 and 120 mg/kg of ZnMet; as is shown in Table 9. This is further indicative of the improved skin quality under these conditions.