Coccidia-induced mucogenesis promotes the onset of necrotic enteritis by supporting Clostridium perfringens growth

This study tested the hypothesis that a host mucogenic response to an intestinal coccidial infection promotes the onset of necrotic enteritis (NE). A chick NE model was used in which birds were inoculated with Eimeria acervulina and E. maxima and subsequently with Clostridium perfringens (EAM/CP). A second group of EAM/CP-infected birds was treated with the ionophore narasin (NAR/EAM/CP). These groups were compared to birds that were either non-infected (NIF), or infected only with E. acervulina and E. maxima (EAM), or C. perfringens (CP). The impact of intestinal coccidial infection and anti-coccidial treatment on host immune responses and microbial community structure were evaluated with histochemical-, cultivation- and molecular-based techniques. Barrier function was compromised in EAM/CP-infected birds as indicated by elevated CFUs for anaerobic bacteria and C. perfringens in the spleen when compared to NIF controls at day 20, with a subsequent increase in intestinal NE lesions and mortality at day 22. These results correlate positively with a host inflammatory response as evidenced by increased ileal interleukin (IL)-4, IL-10 and IFN-gamma RNA expression. Concurrent increases in chicken intestinal mucin RNA expression, and goblet cell number and theca size indicate that EAM/CP induced an intestinal mucogenic response. Correspondingly, the growth of mucolytic bacteria and C. perfringens as well as alpha toxin production was greatest in EAM/CP-infected birds. The ionophore narasin, which directly eliminates coccidia, reduced goblet cell theca size, IL-10 and IFN-gamma expression, the growth of mucolytic bacteria including C. perfringens, coccidial and NE lesions and mortality in birds that were co-infected with coccidia and C. perfringens. Collectively the data support the hypothesis that coccidial infection induces a host mucogenic response providing a growth advantage to C. perfringens, the causative agent of NE.     

Intestinal digesta viscosity decreases during coccidial infection in broilers

1. The effect of intestinal digesta viscosity on bird performance in chickens with coccidiosis was compared to those without coccidiosis. 2. Six hundred chicks were divided into five groups: one control group was fed a basal maize/soyabean-based diet and the other groups were fed the basal diet supplemented with 2, 4, 6 or 8 g carboxymethyl cellulose (CMC) per kg of feed. At 14 d of age half the birds were individually inoculated with sporulated oocysts of Eimeria acervulina and Eimeria praecox. 3. Intestinal digesta viscosity increased with increasing inclusion of CMC. This effect was considerably less pronounced in inoculated than in non-inoculated birds. 4. There was a significant negative effect on live weight gain and feed conversion ratio (FCR) with increasing CMC inclusion in non-inoculated birds, but in inoculated birds there was no clear relation between CMC inclusion and performance. Neither intestinal lesion scores, nor numbers of Clostridium pefringens in the caeca, were significantly affected by CMC inclusion. 5. Across all diets inoculation impaired growth rate by 9% and FCR by 8%, but did not affect the amount of C. perfringens in the caeca.     

Role of Coccidia in the occurrence of necrotic enteritis of chickens

Clostridium perfringens type A, Eimeria acervulina, and Eimeria necatrix were used to produce necrotic enteritis in chickens. The disease was produced in all groups of birds that received feed contaminated with C. perfringens. Mortality due to necrotic enteritis was highest (53%) in birds infected with E. acervulina before infection with clostridia. There was a significant difference in mortality rates between birds infected with E. acervulina and birds infected with E. necatrix before infection with C. perfringens. Mortality rates also differed significantly between the group infected with E. necatrix and the group that received only feed contaminated with C. perfringens. It was concluded that under field conditions, coccidia can play a significant role in the occurrence of necrotic enteritis when a sufficient number of toxigenic strain of C. perfringens type A is present. The pathological changes induced by clostridia and coccidia are described.     

Effect of a radiant energy-treated lysozyme antimicrobial blend on the control of clostridial necrotic enteritis in broiler chickens

A cage study was conducted to demonstrate the effect of Entegard REV, a lysozyme-based antimicrobial blend, on the performance of broiler chickens and necrotic enteritis (NE) disease reduction of birds that were challenged with Eimeria maxima and Clostridium perfringens. In the experiment, challenge by the infectious agents without medication resulted in impaired feed consumption, weight gain, and feed conversions and caused high incidence of gross NE lesions and NE mortality rate. Entegard REV included in feed at 200 g/metric ton (MT) was very effective in reducing negative health effects in the birds after NE challenge, and its ability to control the disease was not statistically different from a commonly used antibiotic growth promotant, bacitracin methylene disalicilate, at 55 g/MT.     

Clostridium perfringens alpha-toxin and NetB toxin antibodies and their possible role in protection against necrotic enteritis and gangrenous dermatitis in broiler chickens

Necrotic enteritis (NE) and gangrenous dermatitis (GD) are important infectious diseases of poultry. Although NE and GD share a common pathogen, Clostridium perfringens, they differ in other important aspects such as clinical signs, pathologic symptoms, and age of onset. The primary virulence factors of C perfringens are its four major toxins (alpha, beta, epsilon, iota) and the newly described NE B-like (NetB) toxin. While neutralizing antibodies against some C perfingens toxins are associated with protection against infection in mammals, the serologic responses of NE- and GD-afflicted birds to these toxins have not been evaluated. Therefore, we measured serum antibody levels to C perfringens alpha-toxin and NetB toxin in commercial birds from field outbreaks of NE and GD using recombinant toxin-based enzyme-linked immunosorbent assay (ELISA). Initially, we used this ELISA system to detect antibody titers against C perfringens alpha-toxin and NetB toxin that were increased in birds experimentally coinfected with Eimeria maxima and C perfringens compared with uninfected controls. Next, we applied this ELISA to field serum samples from flock-mated birds with or without clinical signs of NE or GD. The results showed that the levels of antibodies against both toxins were significantly higher in apparently healthy chickens compared to birds with clinical signs of NE or GD, suggesting that these antitoxin antibodies may play a role in protection against NE and GD.     

Effect of Eimeria tenella infection on the caecal population of lincomycin-resistant Clostridium perfringens introduced into chickens

The effect of Eimeria tenella infection on the caecal population of lincomycin-resistant Clostridium perfringens (KGW 851) newly introduced into chickens was studied. Four groups of chickens consisted of: (1) Uninoculated controls; (2) inoculated with C perfringens (KGW 851); (3) inoculated with E tenella; and (4) inoculated with C perfringens (KGW 851) followed by E tenella. Five chickens in each group were necropsied on each of days 0, 3, 5, 7, 10 and 14 after the E tenella inoculation. The mean total C perfringens counts in the caecal contents increased at five days after E tenella inoculation and reached maximum counts at seven days after the inoculation. Also lincomycin-resistant C perfringens readily established itself at approximately one 10th of the total C perfringens population in the presence or absence of E tenella infection.     

Effect of trypsin inhibitor activity in soya bean on growth performance, protein digestibility and incidence of sub-clinical necrotic enteritis in broiler chicken flocks

1. The effect of three different levels of dietary trypsin inhibitor activity (achieved by varying the amount of non-toasted full fat soya bean in replacement for toasted full fat soya bean) on the incidence of spontaneously-occurring sub-clinical necrotic enteritis (NE) in broiler chickens was compared. A fourth dietary treatment compared the effect of a diet that used potato protein concentrate as the major protein source. The determined trypsin inhibitor activity increased with the increasing content of non-toasted soya bean: 1·90, 6·21, 8·46 and 3·72 mg/g for the three soya bean diets (0, 100 and 200 g of non-toasted soya bean/kg) and the potato protein diet respectively. 2. Although increasing amounts of the non-toasted full-fat soya bean increased the feed intakes of the birds, there was a marked reduction in protein digestibility, weight gain and feed conversion efficiency. 3. There was a linear increase in sub-clinical NE lesions in the duodenum, jejunum, mid small intestine and ileum with increasing non-toasted soya bean. Caecal Clostridium perfringens counts increased with the increasing dietary content of non-toasted soya bean. Serum α-toxin antibodies were higher in the birds fed the 200 g non-toasted soya bean/kg diet compared with the other diets. 4. The results demonstrated that variation in the amount of non-toasted dietary soya bean not only affects growth performance of broilers but also affects the incidence of sub-clinical necrotic enteritis in the flock. Ensuring the lowest possible trypsin-inhibitor activity in soya bean samples is a valuable tool to improve the health and welfare of birds and in reducing the financial losses from this disease.     

Effect of avilamycin, tylosin and ionophore anticoccidials on Clostridium perfringens enterotoxaemia in chickens

In order to study the prophylactic and metaphylactic effect of antomicrobial growth promoters and ionophorous anticoccidials on the incidence of Cl. perfringens enterotoxaemia in chickens, experimental attempts were performed with 675 chickens in 27 trials. The birds were intraduodenally infected with Cl. perfringens type A (ATCC 3624). The following antimicrobial growth promoters and ionophore anticoccidials were used either on their own or in combination: avilamycin, narasin, monensin and tylosin. While infected and non-medicated trials showed an average incubation period of 1 week, clinical symptoms occurred 2-4 days later in infected and medicated birds. Avilamycin medicated birds had the longest incubation period. In the infected and non-medicated trials, a mortality rate of 16%-36% was noted within 3 weeks post infection. The avilamycin trials showed a mortality rate of 0-8% (0-2 birds died) and the narasin and monensin a mortality rate of 0-8%, respectively. In the combination groups (monensin + avilamycin or narasin + avilamycin), the mortality rate ranged from 0 to 4%. Tylosin showed a very good metaphylactic/therapeutic effect against Cl. perfringens enterotoxaemia. Following infection, medicated birds showed a significantly better bodyweight gain than the chickens, whose feeds had not been supplemented. From epidemiological point of view, the systematic prevention of coccidiosis is a key in the control of Cl. perfringens enterotoxaemia in chickens.     

Performance of Broilers Fed on Diets Containing Different Amounts of Chaya (Cnidoscolus aconitifolius) Leaf Meal

The performance and gut measurements of broilers fed on diets containing different amounts of chaya (Cnidoscolus aconitifolius) leaf meal (CLM) were examined in two experiments. In the first experiment, 60 Hubbard chickens (30 males and 30 females; 2 weeks old) were fed on five maize diets; these were formulated using 0, 150 (CLM150), 250 (CLM250) or 350 (CLM350) g CLM/kg, and the fifth diet contained soyabean. In the second experiment, 148 Ross male chicks, 1 day old, were fed on three isonitrogenous and isoenergetic maize–soyabean-based diets, which included 0 (control), 150 (C150) or 250 (C250) g CLM/kg. The diets were offered ad libitum for 2 or 3 weeks in the first and second experiments, respectively. Food intake, weight gain and the food:weight gain ratio were recorded. The weight of the gizzard and intestine and the weight and length of the caeca were also determined in the second experiment. In experiment 1, the birds fed on the maize–soyabean diet had a higher (p<0.05) weight gain and final weight than birds fed on maize only or on the CLM150 diets. There were no differences for any of the variables studied between the birds fed on the maize–soyabean diet and those fed on the CLM250, nor between males and females. In the second experiment, weight gain, food intake and the food:weight gain ratio for birds fed on C250 were lower (p<0.05) than those in birds fed on either the control or C150 diets. The weights of the gizzard and intestine were the lowest and the highest, respectively, in birds fed on C250 (p<0.05). The length and weight of the caecum from birds fed on the control diet were lower (p<0.05) than those of birds fed on either the C150 or C250 diets. The results from this study suggest that CLM may be included up to 150 g/kg in commercial diets without having an adverse effect on poultry performance, and may also be mixed with maize up to 250 g/kg to improve the performance of chickens fed on low-protein diets.     

The role of penicillin G potassium in managing Clostridium perfringens in broiler chickens

The efficacy of penicillin G potassium (Pot-Pen) administered via drinking water to manage necrotic enteritis (NE) was investigated in a Clostridium perfringens (CP) challenge study using 1600 broiler chickens assigned to one of four treatment groups: nonchallenged, nonmedicated; challenged, nonmedicated; challenged, Pot-Pen 0.2 g/L; challenged, Pot-Pen 0.4 g/L. Overall mortality due to NE was significantly reduced among Pot-Pen-treated pens; mortality due to other causes did not differ among the treatment groups. Among all birds, growth performance parameters were significantly improved among Pot-Pen-treated pens. When considering birds randomly sacrificed 4 days post-Pot-Pen initiation, mean NE lesion scores were greatest among the challenged, nonmedicated pens; only one of 80 randomly sacrificed birds treated with Pot-Pen had NE lesions. Among the nonmedicated control pens, body weight (BW) was significantly greater among birds that did not have NE-associated lesions. When sacrificed birds were stratified by NE lesion score, there were no significant differences in BW among the treatment groups. Results of this study suggest that CP-associated subclinical disease can significantly reduce broiler performance. Furthermore, the positive effects of treatment with Pot-Pen appeared to be associated with the prevention and/or treatment of NE-specific lesions.     

Etiology and pathogenesis of necrotic enteritis

Sporulated oocysts of Eimeria acervulina were administered orally to cage-housed broilers at a dose of 3.5 X 10(5) resulted in mild subclinical coccidiosis. Clostridium perfringens incorporated in feed at a level of 2.5 X 10(8) organisms/g. produced lesions characteristic of necrotic enteritis. Mortality of 8% (7/80) occurred in birds fed a ration inoculated with Cl. perfringens alone. Mortality of 35% (28/80) was observed in birds which received an oral dose of E. acervulina and which were fed simultaneously with a ration containing Cl. perfringens. Birds which were fed an inoculated ration two days after an oral dose of E. acervulina showed 41% (33/80) mortality. Birds which received an inoculated ration for two days before administration of an oral dose of E. acervulina demonstrated 18% mortality (15/80). Birds which were fed an inoculated ration four days after an oral dose of E. acervulina showed 10% mortality. Infection with E. acervulina reduced the pH of intestinal contents with a simultaneous depression in serum protein. A 39% increase in intestinal passage time from 178 to 248 minutes occurred on the fifth day after infection with E. acervulina. These experiments suggest that necrotic enteritis, attributed to proliferation of a toxigenic strain of Cl. perfringens, followed intestinal stasis and minimal lesions induced by mild intestinal coccidiosis.     

Effects of dietary tannic acid and vaccination on the course of coccidiosis in experimentally challenged broiler chicken

An experiment was carried out to assess the influence of tannic acid (TA) on integrity of the intestine in broiler chicks vaccinated against coccidiosis and challenged with the disease. In a 2 × 2 factorial design, the trial had five groups of 10 chickens each, including positive (group 2) and negative (group 1) controls. The chickens were kept on wood shavings and fed a commercial maize and soybean-based starter-grower diet. From day 1, groups 3 and 5 received TA (10 g kg(-1)) in their diet. On day 4, birds of groups 4 and 5 were vaccinated orally against coccidiosis with anti-coccidial vaccine, Livacox T?. Each dose of the vaccine contained 300-500 sporulated oocysts of each of Eimeria acervulina, Eimeria maxima and Eimeria tenella. On day 18, all experimental groups except for the negative (group 1) were challenged with 10-fold dose of Livacox T? to produce a mild coccidiosis infection. Faecal samples of individual birds were collected on day 23, and the number of faecal oocysts was determined. d-Xylose absorption test was also carried out on all birds on day 23. Immediately after d-xylose absorption test, all birds were killed humanely and the intestinal tract was removed, weighed and examined for gross lesions. Results showed that negative (group 1) and positive controls (group 2) had the highest and lowest levels of plasma d-xylose post-ingestion of the substrate, respectively. Vaccination and/or feeding TA raised the level of plasma d-xylose in infected birds, although this was not significant for TA-fed birds. Vaccination reduced but TA increased the total number of oocysts per gram of faeces. Birds of groups 2-5 had distinct intestinal lesions when compared with group 1. However, vaccination prevented intestinal lesions. Relative weights of intestinal parts were the lowest in group 1 and the highest in group 2. Vaccination but not TA reduced the relative weights of intestinal parts in infected birds. It was concluded that dietary tannins may reduce the efficacy of anticoccidial vaccines and alter the proper development of immunity against the disease.     

Factors affecting the incidence of necrotic enteritis, caecal carriage of Clostridium perfringens and bird performance in broiler chicks.

Two trials were conducted to study the effects of a competitive exclusion (CE) product BROILACT and the anticoccidial narasin on the incidence of necrotic enteritis (NE), the numbers of Clostridium perfringens (CP) in the caeca of broiler chicks and the performance of the birds. In trial 1 the effects of type of protein and partial replacement of a narasin containing diet with whole wheat were also studied. All groups of chicks were studied up to the point of slaughter at 43 days of age and after evisceration in a processing plant to determine slaughter yield. In trial 1, statistically significant results included the following: CE-treatment reduced total mortality, and incidence of NE, on diet containing animal but not vegetable protein. Caecal carriage of CP was also reduced, while slaughter yield increased. Narasin reduced caecal carriage of CP and increased both growth rate and slaughter yield in both trials. Whole wheat replacement improved feed conversion but reduced bird growth rate. In trial 2, both CE-treatment and narasin influenced feed intake, CE-treatment significantly only at days 22 and 44. Narasin improved feed conversion until 5 weeks of age and CE-treatment did so until 22 days of age. In both trials, there was also an interaction effect indicating that CE-treatment increased slaughter yield for birds that were not fed narasin.     

Influence of bacteria on Clostridium perfringens infections in young chickens

When monoflora chickens with Lactobacillus acidophilus or Streptococcus faecalis were inoculated with Clostridium perfringens either in broth culture or resuspended in Gifu anaerobic medium broth or supernatant fluid, few or no chickens died. Approximately 50% of germ-free chickens died after inoculation of C. perfringens culture, whereas no conventional birds died after inoculation of broth culture. C. perfringens in the contents of duodenum from germ-free chickens numbered about 10(4) colony-forming units (CFU)/g after inoculation 10(8) CFU broth culture per bird, but in gnotobiotic and conventional chickens this organism decreased or was not detected. When C. perfringens was cultured in intestinal contents collected from germ-free chickens, C. perfringens proliferated but alpha toxin was not detected. These findings indicate that the pathogenicity of C. perfringens was suppressed by L. acidophilus or S. faecalis administered previously or inhibited by normal intestinal flora.     

不同净能水平和赖氨酸净能比的低蛋白日粮对肥育猪生长性能和胴体品质的影响

选用61.08 kg杜×长×大三元杂交猪540头,随机分为9个处理,每处理6个重复,每重复10头猪,公、母各半。采用3×3两因素设计,日粮蛋白水平降低4个百分点,净能为9.83、10.04、10.25 MJ/kg,赖氨酸净能比为0.76、0.84、0.91 g/MJ,研究净能和赖氨酸净能比对肥育猪生长性能和胴体品质的影响。结果表明:在60~100 kg阶段,10.04 MJ/kg净能组的ADG显著高于9.83 MJ/kg组的(P<0.05);赖氨酸净能比为0.84 g/MJ组的ADG显著高于0.76 g/MJ组(P<0.05);二者互作不影响生长性能和胴体品质(P>0.05),综合评定在60~100 kg阶段,猪的低蛋白日粮净能水平和赖氨酸净能比分别为10.04 MJ/kg和0.84 g/MJ为宜。     

Studies on the stage of action of lasalocid against Eimeria tenella and Eimeria acervulina in the chicken

Broiler chickens in battery pens were either fed a diet containing 100 ppm lasalocid or no drug for 24 h prior to inoculation with sporulated oocysts of Eimeria tenella or Eimeria acervulina. Different groups of birds remained on medicated feed for 24, 48, 72, 96, 120 or 144 h after inoculation. Conversely, other groups started on an unmedicated diet, were given medicated feed at different times after oocyst inoculation. Starting lasalocid medication 24 h (E. tenella) or 48 h (E. acervulina) after inoculation reduced the lesions and improved the weight gain. There was no significant difference in performance of birds after withdrawal of the drug at 48 h (E. tenella) or 72 h (E. acervulina) and thereafter. Starting lasalocid medication at 96 or 120 h did not suppress but rather reduced oocyst production.     

Effects of Eimeria brunetti infection and dietary zinc on experimental induction of necrotic enteritis in broiler chickens.

Broilers infected with Eimeria brunetti and given dietary zinc were examined for experimental induction of necrotic enteritis. Inoculation with sporulated E. brunetti oocysts at 7 days of age was followed by 5 consecutive days of oral inoculation with cultured Clostridium perfringens. Feed was supplemented with zinc at 1000 ppm. Upon necropsy of broilers 6 days after coccidial inoculation, necrotic enteritis was found in 20% (2/10) of birds given both organisms and dietary zinc. Coccidial lesion scores were also highest in that group. Birds infected with E. brunetti and C. perfringens with no dietary zinc had significantly higher coccidiosis lesion scores (P less than 0.05) than groups inoculated with E. brunetti only, regardless of zinc supplementation. Alpha toxin levels in intestinal contents were low in groups infected with both organisms, regardless of zinc supplementation. Zinc was tested for effects of alpha toxin production in vitro. In the mid-log phase (6 hours incubation), a high level of alpha toxin was produced in zinc-supplemented media, but this was lost quickly in the presence of trypsin. Addition of zinc partly protected the toxin from the action of trypsin.     

Growth performance and immune response of two commercial broiler strains fed diets containing Lactobacillus cultures and oxytetracycline under heat stress conditions

1. Hubbard x Hubbard (HH) and Shaver x Shaver (SS) chicks given a dietary supplement of either 50 mg/kg oxytetracycline (OTC) or 1 g/kg Lactobacillus culture (LC) were exposed to 36 +/- 1 degrees C for 3 h daily from day (d) 21 to 42. 2. Prior to heat treatment, body weight (d 21) and weight gain (d 1 to d 21) of OTC and LC birds were greater than those fed the control diet. Chicks given LC had the best food efficiency followed by OTC and control birds during d 1 to d 21. Body weight (d 1 and d 21) and weight gain (d 1 to d 21) were greater for HH tlhan SS chicks. 3. After 3 weeks of heat exposure, birds receiving the LC diet had greater body weight and weight gain, higher food intake and lower food efficiency than OTC and control chicks. 4. Antibody production against Newcastle discase vaccine on d 21 was not affected by strain or diet. On d 42, while diet had negligible effect on this variable among the SS broilers, HH birds fed LC had higher antibody production than those on the control diet. 5. Neither strain nor diet had a significant effect on mortality.     

Oocyst output and transmission rates during successive infections with Eimeria acervulina in experimental broiler flocks

The infection dynamics of Eimeria species determine the clinical manifestation of the disease coccidiosis in poultry flocks, and a better understanding of the dynamics may contribute to improvement of control measures. Our aim was to study the course of infection and the transmission of Eimeria acervulina in groups of broilers by quantifying the transmission rate parameter and oocyst output. Three transmission experiments were carried out with groups of 20 male SPF broilers. At 2 days of age, one bird in each trial was orally inoculated with five sporulated E. acervulina oocysts (D0 post-inoculation, pi). One day after inoculation (D1 pi), the inoculated bird was housed with 19 non-inoculated contact birds. Individual faecal droppings were examined daily from D3-D32 pi to quantify the number of oocysts per gram faeces. The inoculated bird started shedding oocysts at D5 pi and contact birds between D10 and D17 pi. Contact birds that became infected due to oocyst excretion by the inoculated bird were characterized as first generation contact birds (C1). Contact birds excreting from D15 pi onwards (C2) became infected after the first C1 birds had started shedding and were considered to belong to a successive generation of the flock infection. Oocyst output was significantly lower for C1 compared to C2 birds, but the transmission rate parameter remained constant for both infection generations. These results suggest that although oocyst load increases, the transmission rate of E. acervulina remains constant between successive generations of infection in a flock.     

Responses of body composition, growth and food efficiency to dietary protein in genetically lean and fat broilers up to seven weeks of age

1. Growth, food intake and abdominal fatness were measured at 7 weeks of age in male and female broilers from genetically lean and fat lines fed on isoenergetic diets containing 160, 190, 210, 230 and 260 g crude protein/kg from hatching. 2. The mean proportions of abdominal fat decreased from 39 to 20 g/kg body weight in the fat line and from 26 to 11 g/kg in the lean line over this range of protein concentrations. 3. Protein requirements, for optimum growth as a proportion of the diet did not differ between the lines but the lean line was less tolerant of inadequate dietary protein. 4. In birds fed on the same diet, conversion efficiencies of food (FCE) and dietary protein (PCE) were always superior in the lean line but energetic efficiency (EE) was similar in good growing birds of both lines. 5. When dietary protein was adequate for growth and body compositions were equalised in the two lines (by feeding higher protein diets to the fat line), lean line birds had 10% better FCE, 6% better EE and 33% better PCE. 6. It is concluded that genetic selection is a more effective and economic means of producing leaner broilers than dietary manipulation.