Immunopathology and Cytokine Responses in Broiler Chickens Coinfected with Eimeria maxima and Clostridium perfringens with the Use of an Animal Model of Necrotic Enteritis

Abstract

The incidence of necrotic enteritis (NE) due to Clostridium perfringens (CP) infection in commercial poultry has been increasing at an alarming rate. Although pre-exposure of chickens to coccidia infections is believed to be one of the major risk factors leading to NE, the underlying mechanisms of CP virulence remain undefined. The objectives of this study were to utilize an experimental model of NE produced by Eimeria maxima (EM) and CP coinfection to investigate the pathologic and immunologic parameters of the disease. Broilers coinfected with EM plus CP exhibited more severe gut pathology compared with animals given EM or CP alone. Additionally, EM/CP coinfection increased the numbers of intestinal CP bacteria compared with chickens exposed to an identical challenge of CP alone. Coinfection with EM and CP repressed nitric oxide synthase gene expression that was induced by EM alone, leading to lower plasma NO levels. Intestinal expression of a panel of cytokine and chemokine genes following EM/CP coinfection showed a mixed response depending on the transcript analyzed and the time following infection. In general, IFN-alpha, IFN-gamma, IL-1beta, IL-2, IL-12, IL-13, IL-17, and TGF-beta4 were repressed, whereas IL-8, IL-10, IL-15, and LITAF were increased during coinfection compared with challenge by EM or CP alone. These results are discussed in the context of EM and CP to act synergistically to create a more severe disease phenotype leading to an altered cytokine/chemokine response than that produced by infection with the individual pathogens.

Effect of <italic>Bacillus Subtilis</italic>-based Direct-fed Microbials on Immune Status in Broiler Chickens Raised on Fresh or Used Litter

“…Total RNA was extracted from intestine using TRIzol (Invitrogen) as described (Hong et al, 2006; Park et al, 2008). Five micrograms of total RNA were treated with 1.0 U of DNase I and 1.0 μL of 10× reaction buffer (Sigma), incubated for 15 min at room temperature, 1.0 μL of stop solution was added, and the mixture was heated at 70°C for 10 min.…”

“…Quantitative RT-PCR oligonucleotide primers for chicken cytokines, chemokines, and GAPDH are listed in Table 1. Amplification and detection were carried out using equivalent amounts of total RNA from intestine using the Mx3000P system and Brilliant SYBR Green QPCR master mix (Stratagene) as described (Hong et al, 2006; Park et al, 2008). Standard curves were generated using log 10 diluted standard RNA and the levels of individual transcripts were normalized to those of GAPDH using the Q-gene program (Muller et al, 2002).…”

Effect of Dietary Antimicrobials on Immune Status in Broiler Chickens

“…Quantitative (q)RT-PCR oligonucleotide primers for chicken cytokines and glyceraldehyde 3-phosphate dehydrogenase (GAPDH) internal control are listed in Table 2. Amplification and detection were carried out using equivalent amounts of total RNA using the Mx3000P system and Brilliant SYBR Green QPCR master mix (Stratagene) as described (Hong et al, 2006; Park et al, 2008). Standard curves were generated using log 10 diluted standard RNAs and the levels of individual transcripts were normalized to those of GAPDH using the Q-gene program (Muller et al, 2002).…”

Worms, flies and four-legged friends: the applicability of biological models to the understanding of intestinal inflammatory diseases

“…In addition to studies performed in mice and rats, NEC can also be induced in other vertebrate animals, including sheep (Wolfs et al, 2009), rabbits (Choi et al, 2010), piglets (Sangild et al, 2006) and chickens (Park et al, 2008), using animal-specific variations of the protocol described above. Although induction of the disease in large animals might resemble the human disease more closely than that induced in mice, disadvantages of using a large animal model include the increased cost of animals and their housing, and the limited availability of transgenic strains.…”