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.     

Experimental reproduction of necrotic enteritis in the chicken. 1. Mono- and polyinfections with Clostridium perfringens and coccidia with reference to cage managemen]

Experimental mono-infection and poly-infection, using vegetative germs of a CL. perfringens Type A strain 1663 and its toxin as well as E. acervulina, necatrix or mitis oocysts, were applied to 100 SPF chicken aged seven days. While clostridial or coccidial mono-infection did not cause any loss and only unspecific pathologic-anatomic changes, polyinfection was accompanied by diarrhoea and slower weight increase, with 55.4 per cent of the chicken having been lost three to nine days from the outbreak of the infection. The pathologic-anatomic findings recorded from the chicken with poly-infections included haemorrhagic, ulcerative, and necrotic intestinal inflammations, primarily in the small intestin. The results of these experimental infections indicated a possible correlation between CL. perfringens and coccidia in the pathogenesis of necrotic enteritis.     

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.     

Efficacy of narasin in the prevention of necrotic enteritis in broiler chickens

The efficacy of narasin in the control of necrotic enteritis (NE) was investigated in a floor pen study of 2000 broiler chickens using a Clostridium perfringens feed inoculum challenge model. Treatments were 1) nonmedicated, nonchallenged; 2) nonmedicated, challenged; 3) narasin, nonchallenged; 4) narasin, challenged. Narasin was administered at 70 ppm in the feed from day 0 to trial termination on day 41. Challenge inoculum contained approximately 1 x 10(8) colony-forming units CP/ml and was administered from day 14 to day 16. In the unmedicated groups, challenged birds had significantly (P < 0.05) lower mean body weight and reduced feed efficiency at day 21 and significantly (P < 0.01) higher cumulative NE mortality at day 41 compared with unchallenged. Similarly, among unmedicated birds, those challenged had a significantly (P < 0.01) higher mean NE score on day 17 and significantly (P < 0.05) higher mean huddling scores on days 15-17 than unchallenged. Among challenged birds, those fed narasin had significantly (P < 0.05) higher mean body weight and improved feed efficiency at days 21 and 41 and significantly (P < 0.01) lower cumulative NE mortality at day 41 than unmedicated. Similarly, among challenged birds, those receiving narasin had a lower mean NE score on day 17 (P > 0.05) and significantly (P < 0.05) lower huddling scores on days 16 and 17 than unmedicated. Coccidiosis lesion scores were zero for birds euthanatized from all treatment groups on day 17, suggesting that the beneficial effects of narasin were not due to prevention of coccidiosis. This study thus provides evidence that narasin is effective in the prevention of necrotic enteritis in broiler chickens.     

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.     

影响鸡坏死性肠炎病例复制的关键因素

坏死性肠炎(necrotic enteritis,NE)是家禽中最重要的肠道疫病之一,呈世界性流行,严重危害养鸡业发展。人工复制鸡坏死性肠炎病例是研究该病发病机制及筛选有效药物的重要手段,然而在实际试验过程中,通常受实验动物、感染菌株、诱导病原等多方面因素影响而难以成功复制病例。鉴于此,现综述了影响鸡坏死性肠炎病例成功复制的关键因素,包括攻毒使用的实验动物、菌株毒素、菌株培养条件、诱导病因等,同时探讨了如何通过调整这些因素来改善人工复制鸡坏死性肠炎病例的严重程度,分析了鸡坏死性肠炎病例的病变评分系统,以期为鸡坏死性肠炎的实验室研究及综合防控提供理论基础。     

Reduced incidence of Clostridium perfringens-associated lesions and improved performance in broiler chickens treated with normal intestinal bacteria from adult fowl

The dosing of young chicks with cultures of normal gut flora has been termed "competitive exclusion" (CE). This study was undertaken to examine, under field conditions, the effect of CE treatment on counts of intestinal Clostridium perfringens (CP) and on the occurrence of CP-associated disease in broiler chickens. A farm having recurrent CP-associated health problems was selected as study site. The study comprised four broiler houses, with one treated and one untreated flock per house. Treated birds were sprayed with the CE product Broilact upon arrival at the farm. All flocks were offered feed containing the ionophorous anticoccidial agent narasin. The feed did not contain growth promoters. Treatment was associated with positive but statistically nonsignificant effects on gut health. Delayed intestinal proliferation of CP and delayed appearance of CP-associated gut lesions were found in CE-treated flocks. This delay was associated with improved production performance at slaughter.     

Effects of necrotic enteritis challenge on intestinal micro-architecture and mucin profile

1. This study investigated the effect of Eimeria spp./Clostridium perfringens induced necrotic enteritis and traditional antibiotic preventatives on intestinal micro-architecture and mucin profile. 2. A total of 600 Cobb 500 broiler chickens were randomly assigned to the following three groups: (i) unchallenged, (ii) challenged, and (iii) zinc bacitracin/monensin (ZnB/monensin) (n = 25 chickens/pen, 8 pens/group). The challenged and ZnB/monensin chickens were individually inoculated with Eimeria acervulina, E. maxima and E. tenella and C. perfringens type A (EHE-NE18) at 9 and 15 d post-hatch respectively, to induce necrotic enteritis. 3. The challenge procedure significantly decreased villus height, increased villus width and increased crypt depth in the challenged compared to the unchallenged chickens. Zinc bacitracin and monensin maintained villus-crypt structure similar to that of the unchallenged chickens. 4. Mucin profile was not affected by Eimeria spp./C. perfringens challenge as demonstrated by periodic acid-Schiff and high iron diamine-alcian blue pH 2 x 5 staining. Zinc bacitracin and monensin decreased the number of intestinal mucin-containing goblet cells. 5. Lectin histochemistry showed a trend towards greater Arachis hypogea (PNA) reactivity in unchallenged chickens. 6. In summary, Eimeria spp./C. perfringens challenge disrupted intestinal micro-architecture; however, challenge did not appear to affect intestinal mucin profile. Traditional antibiotics, zinc bacitracin and monensin maintained micro-architecture.     

A survey of sensitivity to anticoccidial drugs in 60 isolates of coccidia from broiler chickens in Brazil and Argentina

Coccidia were isolated from 90 broiler farms in 15 poultry-producing areas in Brazil and Argentina. Sixty isolates were tested for sensitivity to 7 anticoccidial drugs. The common species were: a) Eimeria tenella, 47 isolates; b) E. maxima, 49 isolates; c) E. acervulina, 44 isolates; d) E. mitis, 26 isolates; and e) E. brunetti, 12 isolates. Isolates were considered sensitive to drugs if intestinal lesion scores of medicated broilers were reduced by at least 50% compared with unmedicated infected broilers or if weight gain was at least 75% of that of uninfected birds in a 6-day laboratory test. According to lesion scores, there was evidence of resistance or seriously reduced sensitivity to monensin in 20 isolates, narasin in 29, salinomycin in 11, maduramicin in 1, clopidol in 36, amprolium in 40, and nicarbazin in 1. According to broiler weight gain, there was resistance to monensin in 36 isolates, narasin in 32, salinomycin in 28, maduramicin in 2, clopidol in 28, amprolium in 50, and nicarbazin in 4. These results suggested incomplete cross resistance of coccidia to polyether ionophorous drugs. The degree of resistance might be explained by previous patterns of use of these drugs.     

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

The present study examines the responses of broiler chickens to oral administration of Clostridium perfringens freshly isolated from field cases of necrotic enteritis (NE). The challenge studies included long-term exposure and short-term exposure, factored in with dietary and management variables including high levels of dietary components such as fish meal, meat meal, abrupt change of feed, and fasting. In the long-term exposure trials, the birds were orally inoculated daily, with 1 ml (1.0 or 2 x 10(8) CFU/ml) of an overnight culture of C. perfringens for 7 days. Short-term exposure trials involved challenge with 1 ml (3 x 10(10) CFU/ml) administered as a single dose. The responses of broilers to orally administered C. perfingens under laboratory controlled conditions are presented and discussed in the context of authentic field cases of necrotic enteritis. None of the challenge trials produced overt clinical signs of NE and there were no mortalities associated with oral exposure to high doses of C. perfringens. However, many of the challenged birds showed distinctly pronounced pathological changes in the intestinal tissue. On gross examination the responses in birds challenged orally with C. perfringens could be placed into two categories: (1) no apparent pathological changes in the intestinal tissue and (2) sub-clinical inflammatory responses with focal, multi-focal, locally extensive, or disseminated distribution throughout various sections of duodenum, jejunum, ileum, and ceca. In birds that responded with intestinal lesions, hyperemia and occasional hemorrhages were the main gross changes. In some birds, the mucosa was covered with a brownish material, but typically, the mucosa was lined by yellow or greenish, loosely adherent material. Mild gross changes were seen in some control birds, but both qualitatively and quantitatively, the lesions were distinctly more pronounced in the challenged birds. Upon histological examination, none of the experimentally exposed birds showed overt mucosal necrosis typical of field cases of NE, but typically the lamina propria was hyperemic and infiltrated with numerous inflammatory cells. Most significant changes were seen at the interface of the basal domain of enterocytes and lamina propria. Multifocally, these areas were extensively edematous, allowing for the substantial disturbance of the structural integrity between the lamina propria and the enterocytes. The lesions observed in the present study were consistently reproduced in all of our challenge trials, hence these responses may signify newly emerging patterns of sub-clinical enteric disorders in contemporary strains of poultry. The pathological changes observed in broilers challenged orally with C. perfringens in the present study, differ significantly from those reported previously, and must be clearly differentiated from those described in cases of NE or ulcerative enteritis. Although no overt necrosis of the intestinal mucosa typical of field cases of NE were observed in the present study, the birds challenged with C. perfringens showed strong inflammatory reaction to the introduced pathogens. The distinct features of the microscopic lesions were changes involving apparently normal enterocytes at the interface of the basal domain of villar epithelia and lamina propria. Although the pathological changes in the intestinal tissues observed in our trials appear to be rather subtle when compared to field cases of NE, the nature of these lesions suggest a significant negative effect on the digestive physiology of intestinal mucosa.     

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.     

柔嫩艾美耳球虫感染对鸡盲肠中一些正常厌氧微生物菌群动态的影响

通过对鸡感染柔嫩艾美耳球虫时盲肠中常见正常厌氧微生物的变化情况进行检测研究发现:鸡在感染球虫后第7、10、14天时盲肠中类杆菌(Bacteriodes)的数量显著减少(P<005),而梭状芽孢杆菌(Clostridium)则显著增多;在球虫感染后的第7、10天时,盲肠中的消化球菌(Peptostreptocuccus)和真杆菌(Eubacterium)呈显著下降(P<005)。上述结果从一个方面阐明了柔嫩艾美耳球虫感染导致对某些其它感染抵抗力下降的机制。     

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.     

Efficacy evaluation of lasalocid plus roxarsone combination medication with different geographic field strains of Eimeria acervulina

Performance of broiler chickens medicated with lasalocid alone (at 125 ppm) or in combination with roxarsone (at 50 ppm) was evaluated in battery and floorpen trials after challenge with geographically different field strains of coccidia containing predominately the upper intestinal species Eimeria acervulina. No significant difference in bird performance measured at 6 days postinfection (PI) was observed between lasalocid plus roxarsone-medicated (L+RM) or lasalocid-medicated (LM) birds challenged in separate battery trials with mixed-species inocula from Alabama or Georgia containing 92% or 88% E. acervulina, respectively. In contrast, L+RM birds challenged in another battery trial with a Louisiana mixed-species inoculum containing 92% E. acervulina showed significant reduction in average weight gain at 6 days PI compared with LM-challenged birds. A floorpen trial done with the same Louisiana inoculum showed significant reduction in average bird weight gain at 27 and 35 days of age (6 and 14 days PI) for L+RM-challenged birds compared with both unmedicated-nonchallenged (UMNC) control and LM-challenged birds. The LM+R groups were significantly lower in average bird weight at 27 days of age than the unmedicated-challenged controls. Feed conversions (FCs) for L+RM birds were significantly higher than those for the UMNC control birds during time of challenge (21-27 days of age) and for the 1-to-27-day-of-age time period. No significant difference in FC was seen between the UMNC and LM groups. Results of this study showed that performance of broiler birds medicated with lasalocid plus roxarsone could vary for geographically different mixed-species challenge inocula that contained predominately E. acervulina.     

Use of hybridoma antibodies and recombinant DNA technology in protozoan vaccine development

The use of hybridoma antibodies developed against the sporozoite stage of avian coccidia, coupled with genetic-engineering techniques, has made it possible to begin bird-immunization studies utilizing an Escherichia coli-elicited coccidial protein. The coccidia are currently controlled in the poultry industry by use of anticoccidial compounds, but it now may be possible to use the bird's own immune system for defense against the parasitic infection. Since the sporozoite stage, which initiates the infection in poultry, is quite complex and is made up of hundreds of proteins or antigens, hybridoma antibodies were produced to identify specific antigens. These antigens, once identified, were found in such minute amounts that it became necessary to utilize genetic engineering in order to produce enough protein for immunization studies. One such protein, designated 5401, has been shown to stimulate an antibody response in immunized birds and to impart partial protection against a coccidial challenge infection. The results of these studies indicate that development of a vaccine against coccidial parasites may someday be possible.     

Optimized necrotic enteritis model producing clinical and subclinical infection of Clostridium perfringens in broiler chickens

In this study we assessed the roles of Eimeria infection and dietary manipulation (feeding a diet with a high level of fishmeal) in an Australian necrotic enteritis (NE) challenge model in broiler chickens. An experiment was designed to test the hypothesis that Eimeria infection and dietary manipulation, i.e., inclusion of fishmeal in the diet, are necessary to induce NE experimentally. The results showed that the combination of Eimeria administration and fishmeal feeding had a significant effect on induction of clinical and subclinical Clostridium perfringens infection. The majority of the mortality that occurred during the second week of the trial was due to an NE outbreak following the C. perfringens challenge. The mortality rate of the birds was 12.00% for the high-fishmeal (HFM; 500 g/kg) group and 9.33% for the low-fishmeal (LFM; 250 g/kg) group when the birds were subjected to C. perfringens and Eimeria. Fishmeal alone did not induce significant mortality in birds challenged only with C. perfringens but showed a significantly higher C. perfringens count than the non-fishmeal (NFM) control group. Eimeria administration had a significant effect on NE-related mortality but did not have an effect on the C. perfringens count. In accordance with the time course of bird mortality, it can be determined that of the 3 successive days of oral gavage with C. perfringens, the first inoculation was essential for inducing NE, but the third had no additional effect on NE-related mortality. Also, reducing the fishmeal level from 500 to 250 g/kg had no negative impact on the reproducibility of the model. It may be concluded that NE can be consistently induced under experimental conditions by feeding broilers a diet containing 250 g/kg fishmeal, using a single inoculation with low numbers of Eimeria, administering one or two oral C. perfringens inoculations, and maintaining appropriate ambient temperatures and diets.     

Intestinal immunomodulation by vitamin A deficiency and lactobacillus-based probiotic in Eimeria acervulina-infected broiler chickens

In a 2 X 2 factorial study, a broiler starter ration was amended for vitamin A (control, C; deficient, A) and probiotic status (-, P) to investigate their modulatory effects onthe host immune system. Birds were inoculated orally with Eimeria acervulina (EA) oocysts, and disease susceptibility was evaluated by assessment of fecal oocyst shedding. Humoral and local cellular mediated immunity were assessed by evaluation of antibody and cytokine (interferon-gamma [IFN-gamma] and interleukin-2 [IL-2]) levels in sera and intestinal secretions on a 3-day interval after inoculation. Fecal oocyst shedding was highest (P < 0.05) in A- birds, followed by AP, C-, and CP birds. Feeding the probiotic reduced shed oocysts by 20% in A fed birds and by 26% in C fed birds. Intestinal IFN-gamma was relatively constant in all treatment groups except for A-, where it declined steadily and was lower (P < 0.05) from day 6 on. Serum IFN-gamma levels fluctuated within each treatment and over time were not revealing. Intestinal IL-2 was highest in CP birds at 3 and 9 days postinfection (DPI) and lowest in A- birds at 3, 9, and 12 DPI (P < 0.05); no difference between treatments was found at 6 DPI (P > 0.05). Eimeria-specific intestinal antibody (Ab) level was constant (P > 0.05) in C- birds but increased with time (P < 0.05) in A-, AP, and CP birds. Serum Ab levels were also constant in A- and CP birds but increased (P < 0.05) in C- and AP birds after 6 DPI. The data demonstrate for the first time a probiotic-enhanced immunity in vitamin A deficient birds. This study is also the first to demonstrate the probiotic effect on local cell-mediated immunity of chickens, best manifested by apparent lower intestinal invasion and development by EA, on the basis of higher IL-2 secretion and lower EA oocyst production.     

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.     

Antimicrobial susceptibility of Swedish, Norwegian and Danish isolates of Clostridium perfringens from poultry, and distribution of tetracycline resistance genes

This study was undertaken to determine the in vitro susceptibility of Clostridium perfringens, isolated from poultry to antimicrobials used in poultry production. The minimal inhibitory concentration (MIC) of eight antimicrobials, including the ionophoric coccidiostat narasin, was determined for 102 C. perfringens isolates, 58 from Sweden, 24 from Norway and 20 from Denmark. Susceptibility to each antimicrobial compound was determined by broth microdilution. The isolates were obtained from broilers (89), laying hens (9) and turkeys (4), affected by necrotic enteritis (NE) or by C. perfringens associated hepatitis (CPH), and from healthy broilers. All strains, regardless of origin, proved inherently susceptible to ampicillin, narasin, avilamycin, erythromycin and vancomycin. A low frequency of resistance to virginiamycin and bacitracin was also found. Resistance to tetracycline was found in strains isolated in all three countries; Sweden (76%), Denmark (10%) and Norway (29%). In 80% of the tetracycline-resistant isolates, the two resistance genes tetA(P) and tetB(P) were amplified by PCR whereas in 20% only the tetA(P) gene was detected. No tetM gene amplicon was obtained from any of the tetracycline-resistant isolates. The uniform susceptibility to narasin revealed in this study shows that the substance can still be used to control clostridiosis. In this study, C. perfringens also showed a low degree of resistance to most other antimicrobials tested. Despite the small amounts of tetracycline used in poultry, a considerable degree of resistance to tetracycline was found in C. perfringens isolates from Swedish broilers.