System for the control of enteropathogenic bacteria in the crops of poultry

The invention provides a method and compositions for controlling food borne enteric bacterial pathogens in poultry populations. The incidence of the colonization of poultry by enteropathogenic bacteria, and/or the populations of enteropathogenic bacteria within colonized poultry, may by substantially reduced by providing particles of an expanded matrix material to the locus or vicinity of the animals, particularly during the period of feed removal prior to slaughter. The method and compositions are particularly useful for the control of Salmonella species, enteropathogenic Escherichia coli, and Campylobacter species.

BACKGROUND OF THE INVENTION
 1. Field of the Invention
 This invention relates to a process for the control of enteric bacterial
 pathogens in animals using expanded matrix materials.
 2. Description of the Prior Art
 Despite the efforts of researchers and public health agencies, the
 incidence of human infections from enteropathogenic bacteria such as
 Salmonella, E. coli 0157:H7, and Campylobacter has increased over the past
 20 years. For example, the number of actual reported cases of human
 Salmonella infection exceeds 40,000 per year. However, the Communicable
 Disease Center estimates that the true incidence of human Salmonella
 infections in the United States each year may be as high as 2 to 4
 million. The USDA Economic Research Service has recently reported that the
 annual cost of the food borne illnesses caused by six common bacterial
 pathogens, Campylobacter spp., Clostridium perfringens, Escherichia coli
 0157:H7, Listeria monocytogenes, Salmonella spp., and Staphylococcus
 aureus, ranges from 2.9 billion to 6.7 billion dollars (Food Institute
 Report, USDA, AER, December, 1996). In addition to the impact of enteric
 pathogens on human health, many of these bacteria also cause significant
 infections in animals. For example, Salmonella infections in swine alone
 cost the United States swine industry more than 100 million dollars
 annually (Schwartz, 1990, "Salmonellosis in Midwestern Swine", In:
 Proceedings of the United States Animal Health Assoc., pp. 443-449).
 Animal food products remain a significant source of human infection by
 these pathogens. Contamination of meat and poultry products with many
 bacterial food-borne pathogens, including the particularly onerous
 pathogens Campylobacter spp., Escherichia coli 0157:H7, and Salmonella
 spp., often occurs as a result of exposure of the animal carcass to
 ingesta and/or fecal material during or after slaughter. Any of the
 above-mentioned pathogens can then be transmitted to humans by consumption
 of meat and poultry contaminated in this manner.
 Preharvest control of enteropathogenic bacteria is a high priority to the
 food industry. However, few products have been developed to facilitate
 such efforts. Currently, preharvest pathogen control within the poultry
 industry is accomplished through use of competitive exclusion cultures or
 probiotics. In fact, at this time, only one such product, developed by
 Nisbet et al. of the USDA Agricultural Research Service (U.S. Pat. No.
 5,478,557) and sold under the trademark PREEMPT (Milk Specialties
 Biosciences, Dundee, Ill.), is available commercially in the United
 States. Moreover, the administration of competitive exclusion cultures is
 preferably targeted to very young animals. Immune lymphokines (ILK) have
 also been recently developed for protecting poultry from colonization with
 enteric pathogens as described by Ziprin et al. (1989, Poult. Sci.,
 68:1637-1642), McGruder et al. (1993, Poult. Sci., 72:2264-2271), Ziprin
 et al. (1996, Avian Dis., 40:186-192), and Tellez et al. (1993, Avian
 Dis., 37:1062-1070), and more recently by Kogut et al. (U.S. Pat. Nos.
 5,891,443 and 5,691,200).
 Despite these advances, the need persists for technologies for controlling
 enteric pathogens in animals, and particularly for the treatment of
 animals immediately prior to slaughter.
 SUMMARY OF THE INVENTION
 We have now discovered a method and compositions for controlling the spread
 of food borne enteric bacterial pathogens in poultry populations. The
 incidence of the colonization of poultry by enteropathogenic bacteria,
 and/or the populations of enteropathogenic bacteria within colonized
 poultry, may by substantially reduced, by providing particles of an
 expanded matrix material to the locus or vicinity of the animals,
 particularly during the period of feed removal prior to slaughter. The
 method and compositions are particularly useful for the control of
 Salmonella species, enteropathogenic Escherichia coli, and Campylobacter
 species.
 In accordance with this discovery, it is an object of this invention to
 provide a method for controlling food borne enteropathogenic bacteria in
 animals.
 Another object of this invention is to provide a method for controlling the
 spread of enteropathogenic bacteria between animals in a population of
 poultry.
 Yet another object of this invention is to provide a method for
 significantly reducing the populations of enteropathogenic bacteria in
 meat producing animals prior to slaughter.
 Other objects and advantages of this invention will become readily apparent
 from the ensuing description.
 DETAILED DESCRIPTION OF THE INVENTION
 During the processing of poultry in slaughterhouses, carcasses may become
 inadvertently exposed to ingesta and/or fecal material. Consequently,
 contamination of the poultry product may occur if the ingesta or fecal
 material contain bacterial food-borne pathogens such as Campylobacter
 spp., Escherichia coli 0157:H7, and Salmonella species. The contamination
 of carcasses with feces or ingesta in this manner is the primary source of
 contamination of poultry with these pathogens.
 In an effort to reduce bacterial/fecal carcass contamination in the United
 States, commercially produced poultry are now deprived of feed prior to
 their transport to the slaughterhouse. However, while this practice
 successfully reduces the amount of ingesta and fecal material in the
 bird's gut and hence decreases fecal contamination levels during
 processing, bacterial levels in the upper gastrointestinal tract (crop)
 are increased. In fact, the number of birds with crops contaminated with
 Salmonella or other pathogens increases from as low as 1-2% prior to feed
 withdrawal, to as much as 60% or more by the time of slaughter. This
 increase is primarily due to the birds' consumption of feces and litter
 lying in their vicinity; once feed has been withdrawn, the birds
 characteristically tend to peck at and consume feces and litter on the
 floor of their enclosure. Thus, bacterial pathogens may be quickly spread
 throughout the population. Moreover, feed withdrawal also effects an
 increase in the pH of the crop, providing an environment in the crop more
 conducive to the growth of the pathogens.
 In accordance with this invention, we have discovered that the
 contamination of poultry with bacterial pathogens may be controlled by
 providing particles of an expanded matrix material to the locus or
 vicinity of the poultry population. Interestingly, the poultry will
 preferentially consume the particulate expanded matrix material rather
 than the feces and litter, thereby reducing the uptake of the pathogenic
 bacteria. Either or both of the spread of the bacterial pathogens among
 the population (i.e., the incidence or number of birds contaminated or
 colonized with the pathogens), as well as the concentration of the
 pathogens in the birds, may be significantly reduced in comparison to
 untreated controls. Moreover, in a preferred embodiment, even greater
 control of the pathogens may be achieved by incorporating one or more
 disinfectants or bactericidal agents in the particles.
 Virtually any enteropathogenic bacterium may be controlled in accordance
 with this technique, including but not limited to Salmonella species,
 enteropathogenic Escherichia coli 0157:H7, and Campylobacter species.
 While the process may be used for the treatment of a variety of poultry,
 it is preferred for use with meat producing poultry such as ducks, geese,
 and particularly chickens and turkeys where large populations are commonly
 reared in confined pens or enclosures.
 In the preferred embodiment, the particles of expanded matrix material are
 provided to the poultry during any period of feed removal or restriction.
 In the particularly preferred embodiment, the particles are provided
 during the period of feed removal which is instituted prior to slaughter
 (also referred to as processing) and/or transport to the slaughterhouse.
 Although the precise time of the feed withdrawal may vary between
 producers, it will typically be within about 1 day prior to slaughter of
 the poultry, most often within about 12 to 24 hours prior to slaughter,
 and may be less than or equal to about 10 hours prior to slaughter. As a
 practical matter, the particles are provided at or shortly before the time
 that feed is removed to minimize consumption of fecal material and litter
 by the poultry. It is also understood that the use of the particles of
 expanded matrix material is not limited to the period prior to slaughter.
 For instance, the particles may be provided to the poultry to prevent or
 treat illness within the poultry population.
 The particles of the expanded matrix material are provided in the vicinity
 of the poultry in an amount or density effective to significantly reduce
 or eliminate their consumption of fecal material and litter relative to an
 untreated population. Thus, the amount should be sufficient that the
 particles are readily available for consumption by the birds over the
 course of the feed withdrawal period, substantially without their
 reverting to consumption of fecal material and litter. The specific amount
 may vary with the extent of the period of feed withdrawal, the type floor
 litter used, and the population density of the birds. However, in the
 preferred embodiment, the particles are distributed or spread on the floor
 or ground within the enclosure in which the poultry are maintained in an
 amount effective to cover approximately 10% or more of the floor area. In
 a particularly preferred embodiment, a sufficient amount of the particles
 are provided to cover approximately 25% or more of the floor area.
 The particles for use in this invention may be prepared from a wide variety
 of expanded matrix materials, including but not limited to polymer foams
 (which may also be referred to as expanded polymers or cellular polymers),
 syntactic foams, or extruded and expanded grains. As defined herein, the
 expanded matrices are solid structures comprising voids filled with a
 gaseous phase derived from a blowing agent or hollow microparticles, which
 are surrounded or partially surrounded by a solid polymer matrix. The
 blowing agents may be introduced chemically, physically, or mechanically.
 A variety of methods for preparing expanded matrix materials from numerous
 base materials are known in the art and the selection of the particular
 method and material is not critical. Furthermore, as described in greater
 detail hereinbelow, other optional solid and/or liquid phase materials may
 also be included within or on the matrix.
 In a first preferred embodiment, the expanded matrix is a polymer foam,
 prepared from a single polymer or a mix or blend of different polymers,
 which may be naturally occurring or synthetic polymers or copolymers.
 Without being limited thereto, examples of naturally occurring polymers
 which are suitable for use herein include carbohydrates, polysaccharides,
 celluloses, and starches. Starch derivatives such as modified starches and
 starch-graft copolymers may also be used. In addition, polystyrene,
 polyurethanes and other isocyanurate based polymers, polycaprolactams,
 polycyclopentadienes, polyolefins, polyvinylchlorides (PVC), epoxy resins,
 urea-formaldehyde resins, latex, silicones, fluoropolymers, and other
 synthetic polymers may also be suitable for use.
 Generally, biodegradable polymers are preferred, particularly starch,
 modified starch, and starch-graft copolymers, to minimize the accumulation
 of waste in the poultry house and simplify its disposal. Such
 biodegradable materials will bioerode relatively quickly in gut, reducing
 the volume of ingesta and fecal material. Starches may be derived from any
 available source. However, even though such biodegradable starch-based
 polymers are generally preferred, their combination with synthetic
 polymers may provide enhanced strength, structural stability, or other
 desired properties.
 Many techniques have been previously described for the preparation of
 polymer foams which are suitable for use herein, and particularly for the
 well-known particulate polymer foams such as loose fill packing or
 "peanuts". The selection of the particular technique is not critical and
 is a matter of choice. Representative non-limiting examples of synthetic
 polymers and techniques for producing polymer foams which may be used
 herein are described by Klempner and Frisch (editors, Handbook of
 Polymeric Foams and Foam Technology, Hanser Publishers, Munich, 1991),
 Throne (Thermoplastic Foams, Sherwood Publishers, Hinckley, Ohio, 1996),
 Stober (U.S. Pat. No. 2,576,977), Ropiequet (U.S. Pat. No. 3,961,000),
 Stochdopole et al. (U.S. Pat. No. 3,723,240) and Holden (U.S. Pat. No.
 3,188,264). Examples of several techniques which may be used for producing
 polymeric foams from starch, starch containing materials, modified starch,
 or starch-graft copolymers, include but are not limited to those described
 by Altieri (U.S. Pat. No. 5,153,037), Protzman (U.S. Pat. No. 3,137,592),
 Fisk (U.S. Pat. No. 5,853,848), Lacourse (U.S. Pat. Nos. 5,043,196 and
 5,863,655), Neuman et al. (U.S. Pat. No. 5,208,267) and Ernst
 (International application WO8302955 or European patent 0087847). The
 contents of each of the above mentioned publications and patents are
 incorporated by reference herein.
 In an alternative embodiment, the expanded matrix material may be prepared
 from syntactic foams. Although closely related to polymer foams, syntactic
 foams are produced by forming the polymer matrix around hollow or gas
 filled microparticles or microspheres. Techniques for the preparation of
 syntactic foams are also known in the art and are described by Klempner
 and Frisch (ibid).
 In another preferred embodiment, the expanded matrix may also be prepared
 from extruded and expanded grains. Briefly, one or more of corn, wheat,
 rice or other grain are mixed with a binding agent, a blowing agent or
 water is incorporated therein, and the mixture is then extruded under heat
 and pressure. Preferred techniques for producing the expanded and extruded
 grain are described, for example, in Starcevich (U.S. Pat. No. 5,186,990,
 the contents of which are incorporated by reference herein).
 The physical properties of the expanded matrix material are not critical.
 The expanded matrix may be open or closed cell, or flexible or rigid,
 although rigid to semi-rigid matrices are preferred. The particle size and
 shape are also variable, although the particles should be large enough to
 be readily distinguished by the birds when they are distributed in the
 holding facility. Accordingly, if the particles are to be spread on a
 floor which is covered with litter, the particles should be larger than
 the litter and droppings, preferably greater than or equal to about 0.5 cm
 in each dimension, with dimensions between about 0.5 to 5.0 cm being
 preferred (e.g. diameter for substantially cylindrical or spherical
 particles).
 Coloring agents may be incorporated into the matrix or provided on the
 surface hereof to impart color to the particulate material. Interestingly,
 poultry typically exhibit a preference for particles which are natural or
 white (reflecting substantially all wavelengths of light within the
 visible spectrum at substantially the same reflectance, i.e. the ratio of
 intensity of incident to reflected light), green (reflecting wavelengths
 within the visible spectrum only at approximately 530 nm), or black
 (absorbing substantially all wavelengths of light within the visible
 spectrum at substantially the same reflectance).
 The particles of the expanded matrix material may also function as carriers
 for delivery of the active agents to the poultry. When used in combination
 with biodegradable particles, the matrix will bioerode in the upper
 gastrointestinal tract and crop of the bird, releasing the agents. Thus,
 in a preferred embodiment, improved control of the enteropathogenic
 bacteria may be effected by incorporating one or more bactericidal or
 bacteriostatic agents (compounds) effective against the bacteria into the
 particles. Without being limited thereto, preferred agents include organic
 acids, alcohols, organic solvents, and cationic detergents (including
 cationic bisiguanides such as chlorhexidine and cyclohexidine), iodine,
 iodophores (i.e. povidoneiodine). Particularly preferred agents include
 citric acid, lactic acid, and/or limonene. It is also understood that
 other adjuvants conventional in the art for the treatment of the animals
 may also be formulated in the particles, such as antitoxins, deworming
 agents, therapeutic antibiotics, and/or non-therapeutic levels of
 antibiotics.
 The particles used herein are not to be confused with a nutritive feed for
 the poultry. Rather, the particles are provided to poultry during periods
 when their access to feed is to be removed or restricted. Thus,
 substantial amounts of nutrient additives typically found in commercial
 feeds, particularly supplements which are provided in addition to grains,
 such as one or more of vitamins, minerals (particularly sodium, calcium,
 and phosphorous), proteins, and amino acids (particularly lysine and
 tryptophan), are preferably not incorporated into the particles of this
 invention. In those embodiments whereon the expanded matrix is prepared
 from starch or a starch containing material such as flour, or from grains,
 the amounts of these vitamins, minerals, proteins, and amino acids would
 be substantially the same as normally present therein without added
 supplementation.

The following examples are intended only to further illustrate the
 invention and are not intended to limit the scope of the invention which
 is defined by the claims.
 EXAMPLE 1
 Particles of extruded and expanded grain (corn) were prepared using the
 procedure described by Starcevich (U.S. Pat. No. 5,186,990) except that
 corn starch and polyvinyl alcohol (PVA) were used rather than corn grits
 and guar gum. The average size of the final extruded material varied
 between 1-2 cm.
 To determine the effect of the particles alone in comparison with particles
 containing different adjuvants, particles were prepared with no added
 adjuvants, with 2% lactic acid (LA), with 0.5% D-limonene (DL) plus 2%
 citric acid (CA), and with 0.5% D-Limonene plus 2% citric acid plus 0.5%
 dioctyl sulfosuccinate (DSS).
 Populations of adult broiler chickens were deprived of feed and divided
 into one of five groups: 1, an untreated control; 2, chickens fed
 particles of extruded and expanded grain (BP) without adjuvants; 3,
 chickens fed particles of extruded and expanded grain containing lactic
 acid (BP+LA); 4, chickens fed particles of extruded and expanded grain
 containing D-limonene and citric acid (BP+DL+CA); and 5, chickens fed
 particles of extruded and expanded grain containing D-limonene, citric
 acid, and DSS (BP+DL+CA+DSS).
 In a first trial, 8 hours after feed withdrawal, the crops of the chickens
 were examined for pH and Salmonella concentration. The results are shown
 in Table 1.
 TABLE 1
 group pH Salmonella conc (log 10) ST Sig
 control 5.43 0.73 A
 BP 6.10 0.62 AB
 BP + LA 5.62 0.65 B
 BP + LA + CA 5.54 0.59 AB
 BP + LA + CA + DSS 5.98 0.57 AB
 In a second trial, the broilers were treated as in the first trial except
 that they were challenged with 10.sup.8 cfu of Salmonella typhimurium at 1
 and 6 days prior to the termination of the experiment. Feed was removed on
 the last day, 8 hours prior to termination of the experiment, and
 particles were provided to the appropriate test groups, as in trial 1.
 Challenge of the poultry with Salmonella at an earlier time ensured that
 Salmonella would be excreted and present in the environment and in the
 litter prior to the feed withdrawal and provision of particles. The
 results are shown in Table 2.
 TABLE 2
 group S. typhimurium conc (log 10/ml) ST Sig
 control 1.87 .+-. 1.66 A
 BP 1.56 .+-. 1.29 AB
 BP + LA 0.84 .+-. 1.22 AB
 BP + LA + CA 1.69 .+-. 1.57 B
 BP + LA + CA + DSS 1.21 .+-. 1.29 AB
 means with different letters indicate significant differences (P &lt; 0.05)
 data presented as mean + standard deviation, n = 20
 It is understood that the foregoing detailed description is given merely by
 way of illustration and that modifications and variations may be made
 therein without departing from the spirit and scope of the invention.