Salmonella Control

Salmonella monitoring and control is a priority in the EU. Control strategies have been implemented on a voluntary basis but are also imposed by various authorities. Obligatory and voluntary monitoring programs provide valuable information on the present status of Salmonella contamination at the different levels of the poultry production chain. In this context, information regarding the relative efficacy of different control methods is important for the poultry industry when setting up an integrated control strategy. By now, there is ample experience with vaccination of the layers, underlining the relatively high efficacy at reducing the egg contamination rate. 

Reducing the contamination rates of broiler carcasses seems to be much more of a challenge, as decontamination of the carcasses still is a matter of debate in Europe. Nutritional strategies certainly have their role to play, be it through formulation or additives or processing protocols. Hygienic measures and other management procedures such as rodent control and insect control are equally important. Future developments should aim at reducing the risk of early post hatch colonization with persistent serotypes such as Salmonella Typhimurium or Paratyphi B.

The European Union is in the forefront regarding Salmonella control in farm animals in general and in poultry in particular, because consumer health, well being and protection constitute a major focus of European policy. This was first laid down in Council Directive 92/117/EEC which aimed to establish monitoring systems for a wide range of zoonoses and to eradicate Salmonella Enteritidis and Salmonella Typhimurium from parent chicken flocks. This directive was based on two principles, namely a top-down approach and a zero tolerance. In spite of the very ambitious objectives of the directive, already on april 12, 2000, the European scientific committee on veterinary measures relating to public health stated that the measures in place to control food-borne zoonoses were insufficient. As a consequence, a Commission Regulation (2160/2003/EC) was issued "on the control of Salmonella and other specified food-borne zoonotic agents". For most aspects this regulation no longer was based on the zero tolerance principle, but rather on the principle of "appropriate level of sanitary protection". Indeed, in this and subsequent complementary regulations, a stepwise reduction of the levels of Salmonella contamination is planned simultaneously at all levels of the production chain, with the notable exception of a zero tolerance in fresh chicken meat. In order to coordinate this action, baseline studies, evaluating the levels of contamination, have been carried out at the same time in all member states of the EU. Community targets have been set for the reduction of all Salmonella serotypes with public health significance in breeding flocks, laying flocks and broiler flocks. To reach these targets, certain control methods are made obligatory, while other methods, such as antibiotic treatment, are prohibited. Continuous monitoring of the progress is done through an obligatory official monitoring scheme. All of this puts considerable pressure on the producers, making Salmonella control a priority for the poultry industry.

In this report the focus is on the control of non-host specific Salmonella serotypes. It does not include control measures for the host specific Salmonella Gallinarum/Pullorum biotypes. 

Numerous studies have investigated the epidemiology of Salmonella infections in layers and in broilers. The overall conclusion is that infection routes are numerous. Non-host specific Salmonella can be considered as part of the ecosystem, as it can infect the intestinal tract of numerous different animal species and it can survive for a long time in the environment. Moreover, in poultry, some serotypes, especially Salmonella Enteritidis, can be transmitted vertically. 

Numerous products and techniques have been proposed for control of Salmonella. The present status of Salmonella control strategies in the broiler production chain has been reviewed recently (Van Immerseel et al., 2009). The tools available for Salmonella control can be divided into three categories: 1. Methods and techniques to increase the resistance of  the birds to Salmonella colonization; 2. Methods and techniques to prevent introduction of Salmonella from the environment; 3. Methods and techniques to reduce the concentration of Salmonella which the birds get exposed to; 4. Methods and techniques to kill the Salmonella once it has colonized the birds. The latter refers to the use of antibiotics in feed or in drinking water. In the third category fall a number of hygiene and disinfection measures, as well as feed additives and drinking water supplements other than antibiotics. Rodent and insect control, limiting the access to the barns, as well as purchasing certified Salmonella-free goods and animals, all can be considered to fall in the second category. Vaccination should increase the resistance of the birds and thus falls in category 1.

Commission Regulation 1177/2006/EC regulates the use of specific methods for control of Salmonella in poultry: The use of antimicrobials for control of Salmonella in poultry is prohibited. Live vaccines are prohibited if the strains used in the vaccines cannot be distinguished from wild Salmonella strains. Live vaccines are prohibited during lay unless they are documented to be harmless. In contrast, vaccination of layer flocks against Salmonella Enteritidis is mandatory from Januari 1, 2008 in all member states of the EU with a layer flock contamination rate of 10% or higher. 

In the light of all this, it is important for the poultry producers to be correctly informed about the relative efficacy of different methods and strategies to control Salmonella. In the following paragraphs, we shall not focus on procedures of general biosecurity, sanitation and decontamination aiming at reducing the environmental infection pressure, as these have been reviewed recently (Ducatelle and Van Immerseel, in press ; Van Immerseel et al., 2009). 

Ingredients used for poultry feed formulation are a potential source of Salmonella contamination, although there are considerable differences in the relative risk of different feed components. Moreover, compound feed can become (cross)contaminated during transport and storage. Therefore feed constitutes a serious threat, especially for the broilers, since these birds are highly susceptible to a wide range of different serotypes particularly in the first week after hatching. As a consequence, numerous efforts have been made to decontaminate finished poultry feeds. Several EU member states have made pelleting of poultry feed mandatory, as a simple straightforward measure to reduce this risk. As antibiotic medication, whether through feed or through drinking water, is strictly prohibited for control of Salmonella in the EU, a wealth of different other (non-antibiotic) feed additives have been launched for which a protective effect against Salmonella is claimed. Sometimes, these claims are merely based on in vitro testing. For only few of these products, a reproducible in vivo effect is documented. These products can be subdivided in different classes, including probiotics, prebiotics, bacteriophages, organic acids, and miscellaneous. 

Most probiotics that are on the market are Lactobacillus or Bifidobacterium species. Some of the metabolites of the lactobacilli, such as hydrogen peroxide, lactic acid and bacteriocins have been shown to inhibit growth of Salmonella in vitro. Some lactobacilli may express mannose-sensitive receptors (thus possibly competing with Salmonella for adhesion) or may possess immunomodulatory properties. Successful in vivo colonization inhibition has been documented a.o. with a Lactobacillus salivarius strain (Pascual et al., 1999).

Most of the available prebiotics are fermentable carbohydrates. Some of these, such as the mannanoligosaccharides, have immunomodulatory effects. The mannan residue is thought to possibly bind the mannose-sensitive fimbriae of Salmonella, thereby possibly blocking adhesion. In vivo studies concerning the ability of these prebiotics in controlling the intestinal colonization of Salmonella, however, showed inconsistent results (Rehman et al., 2009). We recently examined the effect of partially hydrolyzed arabinoxylans on Salmonella Enteritidis caecal colonization, shedding and invasion in broilers (Eeckhaut et al., 2009). Arabinoxylans from wheat bran, treated with an endoxylanase in order to obtain an arabinose/xylose ratio of 0.25 and an average polymerization of 9, added to the feed at a dose of 0.2 and 0.4%, resulted in a significant reduction of the percentage of positive cloacal swabs and of the Salmonella titer in the spleen. This effect, however, was only temporary. This  result is in remarkable contrast with a feeding experiment in which we had two groups of broilers. Both were on feeds formulated to achieve the same energy and protein content, and fulfilling all the requirements of a standard broiler ration, but with the energy in the one group coming from corn, and in the other group from wheat and rye. Experimental infection with Salmonella Enteritidis early after hatching resulted in significant differences on day 15 in the levels of colonization of the caeca and the spleens. These differences were of the magnitude of 1 or 2 logs (Teirlynck et al., 2009). These results suggest that the mere choice of ingredients for feed formulation may influence susceptibility of birds to Salmonella colonization.

The most common feed additives used to control Salmonella probably are the organic fatty acids. It is now commonly accepted that these additives cannot exert any effect in the dry environment of the feed. They require a moist environment and a metabolically active microorganism, conditions which are fulfilled after ingestion of the feed. Numerous studies have been published on the in vitro antimicrobial activity of short and medium chain fatty acids. These studies do not take into account the important effects of pH, enzymes and absorption taking place in the gastro-intestinal tract. Moreover, they do not take into account that the most important site of colonization of Salmonella is the caeca (Desmidt et al., 1997), a  segment that may never be reached by some of these products. We compared different short chain fatty acids in vivo and showed that propionic acid and especially butyric acid in coated form, thus possibly reaching the lower intestinal tract, reduce the level of caecal colonization and faecal shedding, whereas formic acid and acetic acid have the opposite effect (Van Immerseel et al., 2004b). The effect is a reduction of only one or 2 logs and it was only investigated shortly after experimental infection. We showed that this effect was due to a change in the invasiveness of the bacteria into the host epithelial cells (Van Immerseel et al., 2003). The molecular mechanism involved with butyrate is a direct suppressive effect on the expression of the Salmonella pathogenicity island 1 at non-bactericidal, biologically relevant concentrations (Gantois et al., 2006a). Medium chain fatty acids may have a similar mode of action (Van Immerseel et al., 2004a) 

Vaccination of chickens against non-host specific Salmonella serotypes is not meant to protect the birds, since the birds only very rarely develop clinical disease. These vaccines are meant to protect the consumers. In the laying hens, this means that vaccination should stop transmission of Salmonella in eggs. The most important serotype in laying hens, causing internal egg contamination, is Salmonella Enteritidis. Until recently, this was also by far the most important serotype causing disease in humans. It has been quite difficult to prove efficacy of vaccines at this point, for the simple reason that the number of contaminated eggs laid by a contaminated layer flock may be as low as 1/1000 or less. One way out of this is to use retrospective data from experience in the field, as has been documented by Feberwee et al. (2001). We used an intravenous infection model in which, in the second week after intravenous challenge with Salmonella Enteritidis, approximately 50% of contaminated eggs are laid. Using this model, we could show that, under the conditions of this experimental setup, a vaccine containing a live attenuated Salmonella enteritidis strain, significantly reduced internal egg contamination with wild type Salmonella Enteritidis. Dual vaccination with both live attenuated Salmonella Enteritidis and live attenuated Salmonella Typhimurium, however, completely blocked wild type Salmonella Enteritidis contamination of the eggs (Gantois et al., 2006b). Based on these results, Salmonella vaccination of layers has  been made mandatory in Belgium. Since then, a drastic reduction in the number of human cases of Salmonella Enteritidis infection has been observed in Belgium (Collard et al., 2008). It is, however, impossible to prove a causal relationship between the two phenomena. 

Salmonella control is a major public health issue since many years in the EU. For a long time, poultry products have been the most important source of Salmonella food poisoning in humans. According to the EU "Trends and Sources Report", this is situation is changing. The numerous efforts made in poultry production appear to be rewarding. 

From what is mentioned above, it appears that the different available tools are not all equally efficacious. Vaccination of laying hens does appear to be efficacious, in order to prevent egg contamination. Contamination of chicken meat, however, still is a matter of concern. The available tools for control are more limited. The use of antibiotics in the live birds is prohibited and decontamination of carcasses is a controversial issue in the EU. Taking into account the Salmonella risk when formulating poultry feeds or using feed additives may reduce the level of contamination by a few logs. Pelleting under optimal conditions may also contribute greatly to the safety of the feed. Classical vaccination strategies so far have not been successful in broilers because of their short life span, their extreme susceptibility early post hatch, and the wide spectrum of different serotypes involved.

Already in 1987, Barrow et al. observed that inoculation of young chicks with a given Salmonella strain prevented the colonization by a second Salmonella strain, particularly when the second strain belonged to the same serogroup. The phenomenon was called colonization inhibition. We extended these observations by showing that the inhibitory effect lasted at least up to 28 days and could suppress the colonization rate of the second strain below 1 log10/g in the caeca (Bohez et al., 2007). Using a seeder bird challenge model, we could show that the colonization inhibitory effect of one Salmonella Enteritidis strain against another Salmonella Enteritidis strain may last up to slaughter age, even when the first strain is defective in the expression of the pathogenicity island 1 (Bohez et al., 2008). Since the Salmonella pathogenicity island 1 is essential for virulence in humans, this approach may open new perspectives for safe Salmonella control in broilers. 

Considering the widespread presence of Salmonella in the environment and in various animal carrier hosts, eradication of Salmonella from production animals must be considered unrealistic. An acceptable level of protection against egg transmission, however, can be achieved in laying hens using vaccination in combination with stringent hygienic measures. In broilers, combining hygienic measures throughout the production chain together with nutritional strategies and, hopefully in the future, colonization inhibition, should allow to achieve an acceptable level of protection against carcass contamination. This requires followup, and as such, permanent monitoring will continue to be the cornerstone of the Salmonella control programs for many years to come. 

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Bohez, L., Dewulf, J., Ducatelle, R., Pasmans, F., Haesebrouck, F., and Van Immerseel, F. (2008) The effect of oral administration of a homologous hilA mutant strain on the long-term colonization and transmission of Salmonella Enteritidis in broiler chickens. Vaccine 26, 372-378. 

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Van Immerseel, F., De Buck, J., Boyen, F., Bohez, L., Pasmans, F., Volf, J., Sevcik, M., Rychlik, I., Haesebrouck, F., and Ducatelle, R. (2004a) Medium-chain fatty acids decrease colonization and invasion shortly after infection with Salmonella Enteritidis in chickens through hilA suppression. Appl Environ Microbiol 70, 3582-3587.

Van Immerseel, F., Fievez, V., De Buck, J., Pasmans, F., Martel, A., Haesebrouck, F., and Ducatelle, R. (2004b) Microencapsulated short-chain fatty acids in feed modify colonization and invasion early after infection with Salmonella Enteritidis in young chickens. Poultry Sci 83, 69-74 

Van Immerseel, F., De Zutter, L., Houf, K., Pasmans, F., Haesebrouck, F. and Ducatelle, R. (2009). Strategies to control Salmonella in the broiler production chain. Worlds Poult Sci J, 65, 365-391. 

This paper was presented at the XVII World Veterinary Poultry Association in Cancun, Mexico, August 14-18, 2011. Engormix.com thanks the authors and the organizing committee for this huge contribution.

Watch Dr. Ducatelle´s interview during the WVPA in Cancun: