Rethinking our understanding of the pathogenesis of necrotic enteritis in chickens

doi:10.1016/j.tim.2008.09.005

Trends in Microbiology

Volume 17, Issue 1, January 2009, Pages 32-36

https://doi.org/10.1016/j.tim.2008.09.005 Get rights and content

For decades, low doses of antibiotics have been used widely in animal production to promote growth. However, there is a trend to reduce this use of antibiotics in feedstuffs, and legislation is now in place in Europe to prohibit their use in this way. As a consequence, economically important diseases, such as necrotic enteritis (NE) of chickens, that are caused by Clostridium perfringens have become more prevalent. Recent research is creating a paradigm shift in our understanding of the pathogenesis of NE and is now providing information that will be necessary to monitor and control the incidence of NE in poultry.

Section snippets

Removing antimicrobial growth promoters and the rise of necrotic enteritis

For decades, antibiotics have been used extensively in animal production worldwide [1] (Box 1). Added in low doses to the feed of farm animals, they have been shown to increase daily weight gain and conversion of feed into body mass, leading to economic advantages for farmers 2, 3, 4, 5. However, there is an increasing trend to reduce this use of antibiotics in feedstuffs. There are concerns that the use of antibiotics in the feed contributes to the spread of antibiotic-resistance genes by

Necrotic enteritis in broiler chickens

C. perfringens produces several myonecrotic and gastrointestinal diseases in humans, domesticated livestock and birds 14, 15. As a species, C. perfringens is capable of producing a wide range of toxins (Box 2), but individual strains produce only a subset of these toxins. The differential production of the four so-called ‘major toxins’ (α-, β-, ɛ- and ι-toxin) is used to classify strains into five toxinotypes (A, B, C, D and E) [16]. In poultry, both clinical and subclinical NE are typically

Histopathology of NE lesions

Early microscopic studies of NE lesions described clearly demarcated foci of advanced tissue degeneration with villi partially or fully denuded of epithelium, large sheets of disintegrating cells found in the lumen and congestion of blood vessels in the lamina propria and submucosa 28, 29. Recent studies of earlier stages of disease have provided valuable new insights into the pathogenesis of NE, indicating damage to the villi initially occurs in the basement membrane and lateral domain of the

Do Clostridium perfringens toxins play a part in NE?

For more than 20 years, α-toxin has been proposed to be the main virulence factor for NE in poultry. The origin of this assumption is unclear but seems to lie in the observations that crude supernatant from C. perfringens type A cultures induced necrotic lesions in broilers [32] and that antibodies to C. perfringens α-toxin prevented the development of lesions [33]. Maternal vaccination with a crude C. perfringens type A and C toxoid has been shown to induce antibodies against α-toxin in

NetB, the new toxin on the block

Recently, NetB, a novel toxin that is associated with broiler NE, has been described [43]. The toxin was identified using screens for proteins from the supernatant of C. perfringens cultures that were cytotoxic for chicken hepatocellular carcinoma cells (LMH) in vitro. The netB structural gene was identified by genomic sequencing of an NE isolate [43]. Both native and recombinant NetB were shown to be cytotoxic for LMH cells, and the mechanism of action seems to involve the formation of small

Vaccination with α-toxin

Until recently, most vaccination efforts directed against NE have concentrated on the use of culture supernatant toxoids in which α-toxin was the major component 34, 44, 45. These trials have been of limited success. More recently, vaccines using recombinant α-toxin and live delivery of α-toxin by attenuated Salmonella have been evaluated and shown to give partial protection against NE in experimental challenge models 31, 46, 47. With evidence accumulating that α-toxin might not play an

Identification of other vaccine antigens

Thompson et al. [40] have used attenuated strains of C. perfringens that no longer express α-toxin to vaccinate chickens. They found that the attenuated vaccine strains offered varying levels of protection. This study clearly demonstrated that C. perfringens carries other molecules that can provide some efficacy as vaccine antigens. Subsequent work by this group has identified several immunoreactive proteins in virulent C. perfringens strains that, in recombinant subunit form, offer some

Concluding remarks and future directions

The story of NE in intensively reared poultry is fascinating and highlights some of the pitfalls in both policy and research. The abolition of growth promoters in feedstuffs was initially accompanied by an increase in the incidence of disease in animals. In part, this change in the pattern of disease reflects the ways in which farming practices have become interdependent and the difficulties associated with isolated changes in these practices. The story of NE also highlights the dangers of

References (60)

S. Schwarz
Use of antimicrobial agents in veterinary medicine and food animal production
Int. J. Antimicrob. Agents
(2001)
A.L. Izat
Effects of different antibiotics on performance, processing characteristics, and parts yield of broiler chickens
Poult. Sci.
(1990)
T. Belay et al.
Virginiamycin and caloric density effects on live performance, blood serum metabolite concentration, and carcass composition of broilers reared in thermoneutral and cycling ambient temperatures
Poult. Sci.
(1996)
A.A. Pedroso
Intestinal bacterial community and growth performance of chickens fed diets containing antibiotics
Poult. Sci.
(2006)
J. Guban
Relationship of dietary antimicrobial drug administration with broiler performance, decreased population levels of Lactobacillus salivarius, and reduced bile sat deconjugation in the ileum of broiler chickens
Poult. Sci.
(2006)
J.J. Dibner et al.
Antibiotic growth promoters in agriculture: history and mode of action
Poult. Sci.
(2005)
J.I. Castanon
History of the use of antibiotic growth promoters in European poultry feeds
Poult. Sci.
(2007)
K. Grave
What has happened in Norway after the ban of avoparcin? Consumption of antimicrobials by poultry
Prev. Vet. Med.
(2004)
L. Petit
Clostridium perfringens: toxinotype and genotype
Trends Microbiol.
(1999)
J.G. Songer et al.
Genotyping of Clostridium perfringens by polymerase chain reaction is a useful adjunct to diagnosis of clostridial enteric disease in animals
Anaerobe
(1996)

M. Kaldhusdal et al.

Barley inclusion and avoparcin supplementation in broiler diets. 2. Clinical, pathological, and bacteriological findings in a mild form of necrotic enteritis

Poult. Sci.

(1992)

J. Brennan

Efficacy of in-feed tylosin phosphate for the treatment of necrotic enteritis in broiler chickens

Poult. Sci.

(2001)

J.P. Dahiya

Potential strategies for controlling necrotic enteritis in broiler chickens in post-antibiotic era

Anim. Feed Sci. Technol.

(2006)

T. Fukata

Influence of Clostridium perfringens and its toxin in germ-free chickens

Res. Vet. Sci.

(1988)

B. Nauerby

Analysis by pulsed-field gel electrophoresis of the genetic diversity among Clostridium perfringens isolates from chickens

Vet. Microbiol.

(2003)

A.R. Gholamiandekhordi

Molecular and phenotypical characterization of Clostridium perfringens isolates from poultry flocks with different disease status

Vet. Microbiol.

(2006)

D.R. Thompson

Live attenuated vaccine-based control of necrotic enteritis of broiler chickens

Vet. Microbiol.

(2006)

A.A. Olkowski

Responses of broiler chickens orally challenged with Clostridium perfringens isolated from field cases of necrotic enteritis

Res. Vet. Sci.

(2006)

R.R. Kulkarni

Oral immunization of broiler chickens against necrotic enteritis with an attenuated Salmonella vaccine vector expressing Clostridium perfringens antigens

Vaccine

(2008)

S. Brynestad et al.

Clostridium perfringens and foodborne infections

Int. J. Food Microbiol.

(2002)

M. Flores-Diaz

Role of Clostridium perfringens phosholipase C in the pathogenesis of gas gangrene

Toxicon

(2003)

M. Lebrun

The expression of Clostridium perfringens consensus β2 toxin is associated with bovine enterotoxaemia syndrome

Vet. Microbiol.

(2007)

Anon (2003) Regulation (EC) No 1831/2003 of the European Parliament and of the Council of 22 September 2003 on...

S.D. Feighner et al.

Subtherapeutic levels of antibiotics in poultry feeds and their effects on weight gain, feed efficiency, and bacterial cholyltaurine hydrolase activity

Appl. Environ. Microbiol.

(1987)

L. Hughes

Risk factors for the use of prescription antibiotics on UK broiler farms

J. Antimicrob. Chemother.

(2008)

W. Van Der Sluis

Clostridial enteritis is an often underestimated problem

World Poultry

(2000)

W. Van Der Sluis

Clostridial enteritis – a syndrome

World Poultry

(2000)

Rood, J.I. (2007) Clostridium perfringens and histotoxic disease. In The Prokaryotes: A handbook on the Biology of...

J.G. Songer

Clostridial enteric diseases of domestic animals

Clin. Microbiol. Rev.

(1996)

M.M. Awad

Virulence studies on chromosomal α-toxin and τ-toxin mutants constructed by allelic exchange provide genetic evidence for the essential role of α-toxin in Clostridium perfringens-mediated gas-gangrene

Mol. Microbiol.

(1995)

Cited by (261)

Treatment of chickens with lactobacilli prior to challenge with Clostridium perfringens modifies innate responses and gut morphology
2024, Research in Veterinary Science
Necrotic enteritis caused by Clostridium perfringens (CP), is a common enteric disease of poultry that has been previously controlled by in-feed antibiotics. However, due to the rapid emergence of antimicrobial resistance, alternatives to antibiotics such as probiotics have received considerable attention because of their immunomodulatory and intestinal health benefits. The present study investigated the effects of probiotic lactobacilli on gut histomorphology and intestinal innate responses in chickens. Day-old male broiler chickens were treated with 1 × 10⁷ or 1 × 10⁸ colony-forming units (CFU) of a lactobacilli cocktail on days 1, 7, 14, and 20 post-hatch, while control groups were not treated with lactobacilli. On day 21, birds in all groups (except the negative control) were challenged with 3 × 10⁸ CFU of CP for 3 days. Intestinal tissue samples were collected before and after the CP challenge to assess gene expression and for histomorphological analysis. Lactobacilli treatment at a dose of 1 × 10⁸ CFU conferred partial protection against NE by lowering lesion scores, increasing villus height in the ileum and reducing crypt depth in the jejunum. In addition, 1 × 10⁸ CFU of lactobacilli enhanced the expression of Toll-like receptor (TLR) 2, interferon-gamma (IFN-γ), interleukin (IL)-10, IL-12, and IL-13 in both the jejunum and ileum at different timepoints and subsequently decreased the expression of transforming growth factor beta (TGF-β) and IL-1β post-CP challenge. In conclusion, the results indicate that treatment with lactobacilli mitigated NE in a dose-dependent manner via improvement of intestinal morphology and modulation of innate immune response in chickens.
Comparison of necrotic enteritis effects on growth performance and intestinal health in two different meat-type chicken strains Athens Canadian Random Bred and Cobb 500
2024, Poultry Science
Chickens have undergone genetic improvements in the past few decades to maximize growth efficiency. However, necrotic enteritis (NE), an enteric disease primarily caused by C. perfringens, remains a significant problem in poultry production. A study investigated the differences in intestinal health between the nonselected meat-type chicken Athens Canadian Random Bred (ACRB) and the modern meat-type Cobb 500 broilers (Cobb) when challenged with experimental NE. The study utilized a 2 × 3 factorial arrangement, consisting of two main effects of chicken strain and NE challenge model (nonchallenged control, NC; NE challenge with 2,500/12,500 Eimeria maxima oocysts + 1 × 10⁹ C. perfringens, NE2.5/NE12.5). A total of 432 fourteen-day-old male ACRB and Cobb were used until 22 d (8 d postinoculation with E. maxima on d 14, dpi), and the chickens were euthanized on 6 and 8 dpi for the analysis. All data were statistically analyzed using a two-way ANOVA, and Student's t-test or Tukey's HSD test was applied when P < 0.05. The NE12.5 group showed significant decreases in growth performance and relative growth performance from d 14 to 20, regardless of chicken strain (P < 0.01). The ACRB group exhibited significant decreases in relative body weight and relative body weight gain compared to the Cobb group from d 14 to 22 (P < 0.01). On 6 and 8 dpi, both NE challenge groups showed significant decreases in intestinal villus height to crypt depth ratio, jejunal goblet cell count, and jejunal MUC2 and LEAP2 expression (P < 0.01). Additionally, the NE12.5 group had significantly higher intestinal NE lesion score, intestinal permeability, fecal E. maxima oocyst count, intestinal C. perfringens count, and jejunal IFNγ and CCL4 expression compared to the NC group (P < 0.05). In conclusion, NE negatively impacts growth performance and intestinal health in broilers, parameters regardless of the strain.
Hyperimmune egg yolk antibodies developed against Clostridium perfringens antigens protect against necrotic enteritis
2023, Poultry Science
Necrotic enteritis (NE) is a widespread infectious disease caused by Clostridium perfringens that inflicts major economic losses on the global poultry industry. Due to regulations on antibiotic use in poultry production, there is an urgent need for alternative strategies to mitigate the negative effects of NE. This paper presents a passive immunization technology that utilizes hyperimmune egg yolk immunoglobulin Y (IgY) specific to the major immunodominant antigens of C. perfringens. Egg yolk IgYs were generated by immunizing hens with 4 different recombinant C. perfringens antigens, and their protective effects against NE were evaluated in commercial broilers. Six different spray-dried egg powders were produced using recombinant C. perfringens antigens: α-toxin, NE B-like toxin (NetB; EB), elongation factor-Tu (ET), pyruvate:ferredoxin oxidoreductase, a mixture of 4 antigens (EM-1), and a nonimmunized control (EC). The challenged groups were either provided with different egg powders at a 1% level or no egg powders (EN). The NE challenge model based on Eimeria maxima and C. perfringens dual infection was used. In Experiments 1 and 2, the EB and ET groups exhibited increased body weight gain (BWG; P < 0.01), decreased NE lesion scores (P < 0.001), and reduced serum NetB levels (P < 0.01) compared to the EN and EC groups. IgY against NetB significantly reduced Leghorn male hepatocellular cytotoxicity in an in vitro test (P < 0.01). In Experiment 3, the protective effect of the IgYs mixture (EM-2) against C. perfringens antigens (NetB and EFTu) and Eimeria antigens (elongation factor-1-alpha: EF1α and Eimeria profilin: 3-1E) was tested. The EM-2 group showed similar body weight, BWG, and feed intake from d 7 to 22 compared to the NC group (P < 0.05). On d 20, the EM-2 group showed comparable intestinal permeability, NE lesion scores, and jejunal NetB and collagen adhesion protein levels to the NC group (P < 0.05). In conclusion, dietary mixture containing antibodies to NetB and EFTu provides protection against experimental NE in chickens through passive immunization.
Nigella sativa as an antibiotic alternative to promote growth and enhance health of broilers challenged with Eimeria maxima and Clostridium perfringens
2023, Poultry Science
The poultry industry has significant coccidiosis and necrotic enteritis (NE) challenges, leading to high mortality and unacceptable growth without antibiotic treatment. This research explored supplementing Nigella sativa (black cumin) seed oil in poultry feed to mitigate coccidiosis and prevent or lessen NE in broilers. In vivo studies consisted of 384 and 320 Cobb 500 male broiler chicks distributed in a randomized complete block experimental design for trials 1 and 2, respectively. The first trial compared 3 concentrations (1, 2, and 5 mL/kg) of black cumin seed oil (BCSO), and trial 2 compared 2 concentrations (2 and 5 mL/kg) BCSO, with birds challenged with Eimeria maxima and Clostridium perfringens (Cp) strains Cp#6 and Cp#4, respectively. Broiler live performance, NE disease outcomes, and Cp populations were measured for both trials. A commercially available BCSO oil product, determined in a preliminary in vitro study to have the highest anti-Cp activity, was selected for in vivo studies. Gas chromatography-mass spectrometry analysis indicated the major bioactive compounds p-cymene, thymoquinone, carvacrol, and thymol were present in the BCSO. In trial 1 with strain Cp#6, BCSO concentrations of 2 and 5 mL/kg reduced NE lesion score and mortality rate to 1.6% compared with 7.8% for positive control, with no adverse impact on live performance. In trial 2 with strain Cp#4, BCSO reduced NE lesion scores and mortality rate to 35.9% compared with 51.6% for positive control and also improved weight gain when there was a Cp infection in broiler chickens. The current study compared NE in broilers challenged with 2 different Cp strains producing different levels of NE. Following Cp infection, both the population of vegetative cells and spores of Cp in cecal contents decreased for all treatments in trial 2. In conclusion, BCSO at concentrations of 2 and 5 mL/kg enhanced broiler live performance and alleviated NE and has potential as a natural, non-medication antimicrobial nutritional supplement for use as a feed additive in chickens.
Collagen adhesin protein and necrotic enteritis B-like toxin as biomarkers for early diagnosis of necrotic enteritis in commercial broiler chickens
2023, Poultry Science
Mouse monoclonal antibodies (mAbs) reactive with Clostridium perfringens collagen adhesin protein (CNA) and necrotic enteritis B-like toxin (NetB) were developed. The best capture/detection mAb pairs for CNA and NetB were selected based on their affinity and specificity to develop sandwich enzyme-linked immunosorbent assays (ELISAs) to detect CNA and NetB proteins, respectively, in jejunal digesta samples from commercial broiler farms in the United States. Prior to the analysis of samples from commercial broiler flocks, the specificity and sensitivity of the CNA and NetB ELISAs were validated using sera, jejunal digesta, and fecal samples from chickens coinfected with Eimeria maxima and CNA⁺/NetB⁺ C. perfringens in an animal model of necrotic enteritis (NE). Subsequently, a total of 251 field samples were collected from 74 commercial poultry farms. Among these, 18 samples were from 6 broiler farms that used certified organics (CO), and 155 samples were from 42 farms with nonantibiotics (NA). In jejunal digesta samples, CNA levels ranged from 0.02 to 0.59 ng/mL and NetB levels ranged from 0.09 to 1.91 ng/mL. CNA and NetB levels showed a positive correlation with each other (Pearson correlation coefficient r = 0.772, P < 0.001). CNA and NetB levels in jejunal digesta were significantly decreased in CO farms compared with those from NA farms (P < 0.001). In conclusion, these new C. perfringens antigen-specific sandwich ELISAs offer a sensitive and specific means to detect C. perfringens CNA and NetB proteins as biomarkers of early NE occurrence in field samples from commercial broiler chickens.
Effect of benzoic acid, Enterococcus faecium, and essential oil complex on intestinal microbiota of laying hens under coccidia and Clostridium perfringens challenge
2023, Poultry Science
The objective of this study was to investigate whether dietary supplementation with benzoic acid, Enterococcus faecium, and essential oil complex (BEC) could help laying hens recover from coccidia and Clostridium perfringens type A challenge. A total of 60 (35-wk-old) Lohmann-laying hens were randomly assigned to 3 experimental groups (10 replicates with 2 hens per replicate): I) control group (CON), II) challenge group (CC), and III) BEC group (2,000 mg/kg BEC). The total experimental period was 8 wk. The results shown that the challenge layers had lower egg-laying rate and average daily feed intake (ADFI) (P < 0.05), and addition of BEC after challenge increased egg-laying rate (P < 0.05). The content of propionic acid (PA) and butyric acid (BA) in short-chain fatty acid (SCFA) was significantly decreased by challenge (P < 0.05). CC and BEC groups had lower villus height to crypt depth ratio (V/C) and higher pathological scores in duodenum (P < 0.05), whereas the BEC group had lower pathological scores in jejunum when compared with the CC group (P < 0.05). The challenge increased the concentration of proinflammatory cytokines (IL-1β and IL-6) (P < 0.05). An increase in the abundance of Bacteroidoes (genus), Bacteroidaceae (family), Bacteroidoes sp. Marseille P3166 (species), Bacteroidoes caecicola (species) was observed in the CC group, whereas the BEC group had higher abundance of Bacteroides caecigallinarum (species). The genera Faecalibacterium and Asterolplasma were positively correlated with egg-laying rate (r = 0.57, 0.60; P < 0.01); and the genera Bacteroides and Romboutsia were negatively correlated with egg-laying rate (r = −0.58, −0.74; P < 0.01). The genera Bacteroides, Lactobacillus, and Rombutzia were positively correlated with jejunal mucosa proinflammatory factor IL-1β level (r = 0.61, 0.60, 0.59; P < 0.01), which were negatively correlated with genera Rikenbacteriaceae RC9, Faecalibacterium, and Olsenlla (r = −0.56, −0.57, −0.61; P < 0.01). There genera UCG.005 was positively correlated with proinflammatory factor IL-6 level in jejunal mucosa (r = 0.58; P < 0.01), which was negatively correlated with Rikenbacteriaceae RC9 (r = −0.62; P < 0.01). The experiment results revealed that the addition of BEC to the diet restored the production performance of the laying hens. In addition, supplementation of 2,000 mg/kg BEC modulated gut health by reducing gut damage scores and modulating microbial composition, thereby promoting recovery of laying hens after coccidia and Clostridium perfringens challenge.

View all citing articles on Scopus

View full text

Trends in Microbiology

ReviewRethinking our understanding of the pathogenesis of necrotic enteritis in chickens

Section snippets

Removing antimicrobial growth promoters and the rise of necrotic enteritis

Necrotic enteritis in broiler chickens

Histopathology of NE lesions

Do Clostridium perfringens toxins play a part in NE?

NetB, the new toxin on the block

Vaccination with α-toxin

Identification of other vaccine antigens

Concluding remarks and future directions

Int. J. Antimicrob. Agents

Poult. Sci.

Poult. Sci.

Poult. Sci.

Poult. Sci.

Poult. Sci.

Poult. Sci.

Prev. Vet. Med.

Trends Microbiol.

Anaerobe

Poult. Sci.

Poult. Sci.

Anim. Feed Sci. Technol.

Res. Vet. Sci.

Vet. Microbiol.

Vet. Microbiol.

Vet. Microbiol.

Res. Vet. Sci.

Vaccine

Int. J. Food Microbiol.

Toxicon

Vet. Microbiol.

Subtherapeutic levels of antibiotics in poultry feeds and their effects on weight gain, feed efficiency, and bacterial cholyltaurine hydrolase activity

Appl. Environ. Microbiol.

Risk factors for the use of prescription antibiotics on UK broiler farms

J. Antimicrob. Chemother.

Clostridial enteritis is an often underestimated problem

World Poultry

Clostridial enteritis – a syndrome

World Poultry

Clostridial enteric diseases of domestic animals

Clin. Microbiol. Rev.

Virulence studies on chromosomal α-toxin and τ-toxin mutants constructed by allelic exchange provide genetic evidence for the essential role of α-toxin in Clostridium perfringens-mediated gas-gangrene

Mol. Microbiol.

Review
Rethinking our understanding of the pathogenesis of necrotic enteritis in chickens