**1. Introduction**

258 Salmonella – A Dangerous Foodborne Pathogen

Zhao, C., B. Ge, J. Villena, R.Sudler, E. Yeh, S. Zhao, D. White, D. Wagner, and J. Meng. 2001.

Zhao, S., S. Qaiyumi, S. Friedman, R. Singh, S. L. Foley, D. G. White, P. F. McDermott, T.

Zhao S, Fedorka-Cray PJ, Friedman S, McDermott PF, Walker RD, Qaiyumi S, Foley SL,

Zhao, S., P. F. McDermott , S. Friedman, J. Abbott, S. Ayers, A. Glenn, E. Hall-Robinson, S.

*Environ Microbiol.* 67:5431-5436.

*Clin Microbiol* 41 (12):5366-5371.

Summer;2(2):169-81.

Prevalence of *Campylobacter* spp., *Escherichia coli*, and *Salmonella* serovars in retail chicken, turkey, pork, and beef from the greater Washington, D.C. area. *Appl.* 

Donkar, C. Bolin, S. Munro, E. J. Baron, and R. D. Walker. 2003. Characterization of Salmonella enterica serotype newport isolated from humans and food animals. *J* 

Hubert SK, Ayers S, English L, Dargatz DA, Salamone B, White DG. 2005. Characterization of Salmonella Typhimurium of animal origin obtained from the National Antimicrobial Resistance Monitoring System. Foodborne Pathog Dis. 2005

K. Hubert, H. Harbottle, R. D. Walker, T. M. Chiller, D. G. White. 2006. Antimicrobial resistance and genetic relatedness among *Salmonella* from retail foods of animal origin: NARMS retail meat surveillance. Foodborne Pathog. Dis. 3:106-17.

*Salmonella enterica* serovars continue to be among the most important foodborne pathogens worldwide due to the considerable human rates of illness reported and the wide range of hosts that are colonized by members of this genus, which serve as vectors and reservoirs for spreading these agents to animal and human populations. Furthermore, public concern for the appearance of resistant strains to many antibiotics, particularly among zoonotic pathogens such as common *Salmonella* isolates, is also challenging the poultry industry to find alternative means of control (Boyle, Bishop, Grassl, & Finlay, 2007). For example, in January 2006 Europe implemented a complete ban on growth promoting antibiotics in animal feed (Anadon, Martinez-Larranaga, & Aranzazu Martinez, 2006). Thus, while attempting to control human foodborne pathogens poultry producers are simultaneously challenged to improve production in the face of increasing feed costs while using fewer antibiotics due to increased restriction of antimicrobial usage. These regulations were implemented because of export market restrictions and consumer or customer preferences in local markets. For these reasons continued research on sustainable alternatives to antibiotic growth promoters for animal production such as probiotics or direct fed microbials (DFM) consisting of live or dead organisms and spores (Patterson & Burkholder, 2003), non-traditional chemicals (Ko, Mendoncam, Ismail, & Ahn, 2009), bacteriophages (Andreatti Filho et al., 2007; Bielke, Higgins, Donoghue, Donoghue, & Hargis, 2007; J. P. Higgins et al., 2005; J. P. Higgins, Andreatti Filho et al., 2008), organic acids and other plant extracts and essential oils (Aengwanich & Suttajit, 2010; Allen-Hall, Arnason, Cano, & Lafrenie, 2010; Bagchi et al., 2000; Kubena, Byrd, Young, & Corrier, 2001; Over, Hettiarachchy, Johnson, & Davis, 2009; Van Immerseel et al., 2006), and vaccines (Kremer et al., 2011; O'Meara et al., 2010; Wolfenden et al., 2010; Van Immerseel et al., 2005; Dueger et al., 2001, 2003) are increasingly more important. These potential solutions have emerged in the last decade as tools that could be potentially useful in the near future for pathogen control and poultry performance improvement.

Probiosis, although not a new concept, has only recently begun to receive an increasing level of scientific interest. In agriculture, probiotics and DFMs used in animal feed are becoming accepted as potential alternatives to antibiotics for use as growth promoters, and in select cases, for control of specific enteric pathogens (Anadón, Rosa Martínez-Larrañaga, & Aranzazu Martínez, 2006; Boyle et al., 2007; Cartman, La Ragione, & Woodward, 2008; Vila et al., 2009; L. D. Williams, Burdock, Jimenez, & Castillo, 2009). For these reasons the

Alternative Strategies for *Salmonella* Control in Poultry 261

Probiotics are defined as live microorganisms that, when administered in adequate amounts, confer a health benefit on the host. Amongst the many benefits associated with the consumption of probiotics, modulation of the immune system has received considerable attention (Borchers, Keen, & Gershwin, 2002; Borchers, Selmi, Meyers, Keen,

Previously, it was thought that administration of bacteria such as probiotics to neonates directly reduced infection by pathogens due to competition amongst the bacteria for attachment sites and nutrients and, that beneficial bacteria would out-compete pathogens within the GIT. This competition, coined as "competitive exclusion" was first described in 1973 by Nurmi and Rantala (Nurmi& Rantala, 1973). Their data indicated that early administration of beneficial bacteria to chicks prevented infection by pathogens. Since Nurmi and Rantala proposed competitive exclusion could be used as a method to prevent *Salmonella* infection, numerous researchers have reported the ability of live bacterial cultures to also reduce colonization of opportunistic microorganisms in the gastrointestinal tract (Callaway et al., 2008; Wagner et al., 2003; Hollister et al., 1999; Corrier et al., 1998; Hume et al., 1998; Nisbet et al., 1998) and probiotic organisms (J. P. Higgins et al., 2010; S. E. Higgins et al., 2008; Vicente et al., 2008; J. P. Higgins et al., 2007; Bielke et al., 2003; Patterson & Burkholder, 2003). Yet, understanding of how probiotics mediate these health benefits,

Balanced gastrointestinal microflora and immune-stimulation are major functional effects attributed to the consumption of probiotics (Amit-Romach, Uni, & Reifen, 2010; Boirivant & Strober, 2007; Boirivant, Amendola, & Butera, 2008; Flint, O'Toole, & Walker, 2010; Flore, Francois, & Felicite, 2010; Ibrahim et al., 2010; Klein, Sanders, Duong, & Young, 2010; Nayak, 2010). Many probiotic effects are mediated through immune regulation, particularly through balance control of pro-inflammatory and anti-inflammatory cytokines (Di Giacinto, Marinaro, Sanchez, Strober, & Boirivant, 2005; Foligne et al., 2010; Hacini-Rachinel et al., 2009; Jobin, 2010; Li, Xia, & Li, 2009). However, several animal and human studies have provided unequivocal evidence that specific strains of probiotics are able to stimulate multiple aspects of innate immunity (Amit-Romach et al., 2010; Boirivant & Strober, 2007; Boirivant et al., 2008; Farnell et al., 2006; Romanin et al., 2010; Weiss et al., 2010) as well as to increase humoral immunity (Fang, Elina, Heikki, & Seppo, 2000; Galdeano, de Leblanc Ade, Carmuega, Weill, & Perdigon, 2009; Leblanc, Fliss, & Matar, 2004; Nermes, Kantele, Atosuo,

Using a *Salmonella* challenge model, an effective LAB probiotic, administered 2 hours after *Salmonella* challenge, had no effect during the first 12 hours on increasing cecal colonization by this pathogen, although marked and rapid decreases were observed between 12 and 24 hours post-challenge (J. P. Higgins et al., 2007; J. P. Higgins et al., 2010). Later, using the same model and microarray analysis of gut mRNA expression, gene expression differences in birds treated with a *Lactobacillus-*based probiotic were compared to saline treated birds. At 12h postprobiotic treatment, 170 genes were significantly different (P<0.05), but by 24h post treatment, the number of differentially regulated genes were 201. Pathway analysis revealed that at both time points, genes associated with the NFκB complex were significantly regulated, as well as genes involved in apoptosis. Probiotic-induced differential regulation of the genes *GAS2* and *CYR61* may result in increased apoptosis in the ceca of chicks. Because *Salmonella* is an intracellular pathogen, it was suggested that increased apoptosis may be a mechanism by

**3. Mechanism of action of probiotics against** *Salmonella* 

specifically reduction of *Salmonella* infection, is very limited.

which B11 reduces *Salmonella* infection (S. E. Higgins et al*.,* 2011).

& Gershwin, 2009).

Salminen, & Isolauri, 2011).

development of new and more effective probiotic products that can be licensed for animal use continues to receive considerable interest (Hong, Duc le, & Cutting, 2005; Hong, Huang, Khaneja, Hiep, Urdaci, & Cutting, 2008a; Jadamus, Vahjen, & Simon, 2001; Osipova, Makhailova, Sorokulova, Vasil'eva, & Gaiderov, 2003; P. Williams, 2007b; Wolken, Tramper, & van der Werf, 2003).

Currently, there is no universal class of probiotic bacterium. However, the most common types that have been indisputably effective involve LAB. These bacteria are found normally in the gastrointestinal tract (GIT) of vertebrates and invertebrates, and the use of some LAB cultures are able to restore the natural microflora within the gut (Shahani & Ayebo, 1980). Lactic acid bacteria include the genera *Lactobacillus*, *Pediococcus*, and others that have long been associated with health benefits and which have been used for fermentation of certain foods. While speciation of members of these genera is difficult and inconsistent, these organisms are considered uniformly safe and are not associated with disease in healthy animals or humans (Tellez et al., 2006).

A second classification of probiotic cultures are those microorganisms that are not normally found in the GIT (such as allochthonous flora). For example, *Saccharomyces boulardii*, a strain of yeast found on some tropical fruits, has been shown to be effective in preventing the recurrence of *Clostridium difficile* infections (Czerucka, Piche, & Rampal, 2007) and some colibacillosis in humans (Czerucka& Rampal, 2002). Otherallochthonous probiotic microbes are the spore-forming bacteria, normally members of the genus *Bacillus*.
