A field study of naturally occurring specific antibodies against Clostridium perfringens alpha toxin in Norwegian broiler flocks

Necrotic enteritis (NE), a disease associated with high numbers of the intestinal bacterium Clostridium perfringens, is common in intensive broiler production. Antimicrobial feed additives may control the disease, but their use is now being questioned in many countries. A field study was undertaken at the end of 1997 to study the level of naturally occurring specific humoral immunity against phospholipase C (PLC; C perfringens alpha toxin) in Norwegian broiler flocks. Blood samples were collected at hatch from 61 study flocks, and the sampling was repeated for 56 of the same flocks at processing. The level of specific antibodies against PLC was analyzed in an enzyme-linked immunosorbent assay (ELISA) test. Data on production performance and weekly mortality were recorded. The relationship between the age of the hens and the level of specific maternal antibodies in the progenies was studied. The association between the level of the maternal antibodies and the production performance, including mortality, was analyzed. The level of specific antibodies against PLC in day-old broiler flocks was relatively high and varied considerably compared with the levels in the broilers at processing. The progenies from the oldest hens had significantly higher levels of specific antibodies than the chicks from younger hens. No outbreak of NE occurred during the study period, making it impossible to analyze the association between naturally occurring specific immunity against PLC and the occurrence of the disease. However, the results showed that the flocks with high titers of specific maternal antibodies against PLC had lower mortality during the production period than flocks with low titers.     

The synergistic necrohemorrhagic action of Clostridium perfringens perfringolysin and alpha toxin in the bovine intestine and against bovine endothelial cells

Bovine necrohemorrhagic enteritis is a major cause of mortality in veal calves. Clostridium perfringens is considered as the causative agent, but there has been controversy on the toxins responsible for the disease. Recently, it has been demonstrated that a variety of C. perfringens type A strains can induce necrohemorrhagic lesions in a calf intestinal loop assay. These results put forward alpha toxin and perfringolysin as potential causative toxins, since both are produced by all C. perfringens type A strains. The importance of perfringolysin in the pathogenesis of bovine necrohemorrhagic enteritis has not been studied before. Therefore, the objective of the current study was to evaluate the role of perfringolysin in the development of necrohemorrhagic enteritis lesions in calves and its synergism with alpha toxin. A perfringolysin-deficient mutant, an alpha toxin-deficient mutant and a perfringolysin alpha toxin double mutant were less able to induce necrosis in a calf intestinal loop assay as compared to the wild-type strain. Only complementation with both toxins could restore the activity to that of the wild-type. In addition, perfringolysin and alpha toxin had a synergistic cytotoxic effect on bovine endothelial cells. This endothelial cell damage potentially explains why capillary hemorrhages are an initial step in the development of bovine necrohemorrhagic enteritis. Taken together, our results show that perfringolysin acts synergistically with alpha toxin in the development of necrohemorrhagic enteritis in a calf intestinal loop model and we hypothesize that both toxins act by targeting the endothelial cells.     

Purification of the alpha toxin of Clostridium perfringens type A by ultrafiltration and gel chromatography

Clostridium perfringens type A toxin produced in Jayko & Lichstein medium was subjected to various concentration and purification procedures. The results obtained with 3 different ultrafiltration membranes followed by gel filtration showed that by using Millipore PSED OHV10 and Amicon XM-100 filter membranes in combination, a three-hundred-and-fivefold purification could be achieved as against a twelvefold increase obtained with ammonium sulphate/acetone precipitation. The lecitovitelin test was more sensitive than the haemolytic activity in determining the alpha toxin activity. The optical density, measured at 280 nm, did not reveal any alpha toxin activity in the relevant toxic fractions.     

Morphologic and physiologic changes induced by Clostridium perfringens type A alpha toxin in the intestine of sheep

OBJECTIVE: To evaluate the morphologic and physiologic changes induced by Clostridium perfringens type A (alpha toxin in the ileum and colon of sheep. SAMPLE POPULATION: 16 ligated intestinal loops in 4 Merino lambs and 18 explants of ileum and colon from slaughtered lambs. PROCEDURE: alpha Toxin-induced fluid accumulation was evaluated in ligated ileal and colonic loops of sheep. Tissues were evaluated morphologically by use of gross and histologic examination. Effects of toxin on in vitro intestinal net water transport were tested in modified Ussing chambers. RESULTS: Ovine ileal and colonic loops incubated with C perfringens type A alpha toxin retained more fluid than control loops. Histologically, in the ileum of lambs inoculated with 300 LD50 of alpha toxin/mL, there was a mild to moderate multifocal infiltration of neutrophils in the lamina propria and submucosa. The colonic loops of lambs inoculated with 30 or 300 LD50 of alpha toxin/mL had excessive mucus in the lumen, a moderate amount of neutrophils mixed with mucus in the intestinal lumen, and moderate multifocal infiltration of the lamina propria and submucosa with neutrophils; the blood vessels of these layers were engorged with neutrophils. In vitro measurements of water transport also revealed inhibition of net epithelial water absorption in ileum and colon incubated with alpha toxin on the mucosal side. CONCLUSIONS AND CLINICAL RELEVANCE: These results indicate that alpha toxin induces alterations in sheep intestine. Clostridium perfringens type A organisms that produce alpha toxin could be responsible for diseases of intestinal origin in some ruminants.     

A comparative pathological study on canine necrotizing meningoencephalitis and granulomatous meningoencephalomyelitis

Canine necrotizing meningoencephalitis (NME) and granulomatous meningoencephalomyelitis (GME) were compared pathologically. Gross observation exhibited lateral ventricular dilation and discoloration, malacia and/or cavitation of the cerebrum in NME. On the contrary, gross changes were milder in GME, except for occasional visible granulomatous mass formation. Histopathologically, the lesions of NME were distributed predominantly in the cerebral cortex and various degrees of inflammatory and necrotic changes were observed according to clinical stages. Besides, microscopic lesions of GME were mainly distributed in the white matter of the cerebrum, cerebellum and brainstem, which are characterized by perivascular cuffing, multiple granulomas and leptomeningeal infiltrates. Although macrophages and lymphocytes were predominant in the inflammatory lesions of both disorders, macrophages in GME transformed into epithelioid cells and exhibited more massive infiltration. Although lectin RCA-1-reactive cells were numerous in both disorders, lysozyme immunoreactive cells in NME were fewer than that in GME. Parenchymal infiltration of MAC387-positive cells was common in GME and limited in NME. The number of CD3-positive lymphocytes in the GME lesions tended to be greater than in NME, though the difference was not statistically significant. Morphological and immunohistochemical differences of the lesions, in particular, the characteristics of infiltrative macrophages may reflect these different pathogeneses of the two disorders.     

霉菌毒素的毒害作用及其互作效应

<正>饲料霉变的典型特征是产生霉菌毒素,可造成高达10%的经济损失,是饲料工业和畜牧业生产中不可忽视的问题。霉菌毒素不但对动物产生毒害作用,而且可通过食物链危及人类健康。动物对霉菌毒素的临     

A necrotizing meningoencephalitis of pug dogs

Clinical and pathologic features of a sporadic, necrotizing meningoencephalitis affecting adolescent and mature pug dogs are described. Many of the affected animals were closely related. Acute and chronic forms occur, with clinical signs reflecting the pathologic affinity of the disease for the cerebral hemispheres. No etiologic agent has been identified. The extensive necrosis and affinity for the cerebral hemispheres are similar to alphatype herpesvirus encephalitides of other species.     

植物毒素免疫研究现状

植物毒素学和免疫学结合,以研究动物中毒性疾病的发生机理、诊断和防治,是一个全新的交叉性研究领域。就植物毒素免疫的免疫学基础、植物毒素人工抗原的研究和植物毒素免疫的临床研究及应用进行了综述。     

重组腐败梭菌α毒素的生物活性及其灭活疫苗的免疫效果

为了研究重组腐败梭菌α毒素的生物活性及其灭活疫苗的免疫效果,采用PCR法扩增腐败梭菌α毒素基因,构建重组表达质粒pET28α-csa,转化E.coli BL21(DE3);对表达产物的细胞毒性、小鼠致死毒力和抗毒素中和试验等生物活性进行鉴定,并将其制成灭活疫苗免疫家兔和靶动物绵羊,通过攻毒素试验和血清中和试验与类毒素疫苗进行比较。结果显示,重组α毒素能够与腐败梭菌抗毒素血清反应,具有细胞毒性,小鼠致死毒力超过300 MLD/mL,其毒性可以被腐败梭菌抗毒素中和。制成灭活疫苗免疫家兔,随着免疫剂量的增加血清中和效价随之提高,0.25 mL/只(含重组α毒素0.11 mg)免疫即可使家兔获得100%保护,免疫效果优于类毒素疫苗;0.50 mL/只(含重组α毒素0.22 mg)免疫绵羊攻毒保护率为75%。结果表明,重组腐败梭菌α毒素具有很好的生物活性,所制成的灭活疫苗可以作为预防羊快疫的候选疫苗。     

Pasteurella multocida toxin induces IL-6, but not IL-1 alpha or TNF alpha in fibroblasts.

Pasteurella multocida toxin (PMT) is the major virulence factor in Progressive Atrophic Rhinitis of swine. Other workers' previous findings that PMT was mitogenic for 3T3 fibroblasts, were confirmed in the present study. In addition, PMT stimulated 3T3 cells to release IL-6, but IL-1 alpha or TNF alpha were not detected in fibroblast supernatants sampled 24, 48, or 72 h after stimulation. In view of the role of IL-6 in osteoclastic bone resorption, these findings provide a new working hypothesis for investigations into the molecular pathogenesis of this important disease.     

Combined action of plant estrogens, F-2 toxin and natural estrogens

The investigation concerns the interaction of plant estrogens, F-2 toxin, and natural estrogen (estradiol-benzoate) as causative factors of eostrogenic changes. Plant estrogens and F-2 toxin were applicated at three and estradiol-benzoate at two dosage levels. The estrogenic effect was studied by bioassay on immature rats taking uterine weight, uterine liquid and vaginal opening as criteria. When uterine weight is taken as a criterion, plant estrogens and F-2 toxin, no matter if they are administered alone or together, regularly increase the effect of estradiol-benzoate, but when uterine fluid is taken as a criterion they always decrease the effect of estradiol-benzoate. According to the present results it seems obvious, that the mode of action of estrogenic substances occurring in plants considerably differ from that of natural estrogens. Thus the mechanisms which cause disturbances may be different from those of natural estrogens.     

A cooperative action of endothelin-1 with prostaglandin F(2alpha) on luteal function in the cow

Prostaglandin F(2alpha) (PGF(2alpha)) is the primary luteolysin in the cow, and luteal endothelin-1 (ET-1) interacts with PGF(2alpha) during the process of luteolysis. In contrast, a developing corpus luteum (CL) is refractory to exogenous administration of PGF(2alpha). Thus, the present study was aimed to investigate the functional relationship between ET-1 and PGF(2alpha) in the mid-CL (PGF(2alpha)-sensitive) and early-CL (PGF(2alpha)-refractory). In the mid-CL model, cows (n = 6/treatment) were assigned to receive one of five types of treatments on day 10 of the estrous cycle: (1) an injection of saline; control, (2) a 500 microg of PGF(2alpha) analogue (sufficient dose to induce luteolytis); full-PG, (3) an intraluteal injection of 0.25 mg ET-1; ET-1, (4) a 125 micro g of PGF(2alpha) (insufficient dose to induce luteolytis); 1/4PG or (5) an intraluteal injection of 0.25 mg ET-1 after administration of a insufficient dose of PGF(2alpha) analogue; 1/4PG/ET. In the early-CL model, cows were assigned to receive one of two types of treatments on day 5 of the estrous cycle: (1) a sufficient dose of PGF(2alpha) analogue; PG (n = 5) or (2) an intraluteal injection ET-1 after a sufficient dose of PGF(2alpha); PG/ET (n = 7). In the mid-CL model, 1/4PG/ET resulted in a rapid reduction of progesterone (P) concentrations similar to that in full-PG from the next day. However, the levels of P in 1/4PG/ET (1.5-2.5 ng/ml) kept significantly higher than that in full-PG (< 0.5 ng/ml). ET-1 or 1/4PG did not decrease plasma P concentrations (4-6 ng/ml). The plasma ET-1 levels increased with the full-PG administration. In the early-CL model, both treatments had no effect on plasma P increase and ET-1 levels. The overall results indicate that the intraluteal ET-1 injection after administration of insufficient dose of PGF(2alpha) induces the depression of P secretion in vivo during the mid luteal phase in the cow, supporting the concept that ET-1 is one of a local mediator of functional luteolysis in the cow. The result further indicates that the early-CL is not only PG-refractory but also ET-1-refractory.     

A study of experimental autoimmune encephalomyelitis in dogs as a disease model for canine necrotizing encephalitis

In the present study, the use of dogs with experimental autoimmune encephalomyelitis (EAE) as a disease model for necrotizing encephalitis (NE) was assessed. Twelve healthy dogs were included in this study. Canine forebrain tissues (8 g), including white and grey matter, were homogenized with 4 mL of phosphate-buffered saline for 5 min in an ice bath. The suspension was emulsified with the same volume of Freund''s complete adjuvant containing 1 mg/mL of killed Mycobacterium tuberculosis H37Ra. Under sedation, each dog was injected subcutaneously with canine brain homogenate at four sites: two in the inguinal and two in the axillary regions. A second injection (booster) was administered to all the dogs using the same procedure 7 days after the first injection. Clinical assessment, magnetic resonance imaging, cerebrospinal fluid analyses, necropsies, and histopathological and immunohistochemical examinations were performed for the dogs with EAE. Out of the 12 animals, seven (58%) developed clinically manifest EAE at various times after immunization. Characteristics of canine EAE models were very similar to canine NE, suggesting that canine EAE can be a disease model for NE in dogs.