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1.
Colony hybridizations with DNA probes for 3 heat-stable (STaP, STaH, and STb) enterotoxins and 1 heat-labile (LT) enterotoxin and for 4 adhesins (K99, F41, K88, 987P) were performed on 870 Escherichia coli isolates to determine pathotypes prevalent among enterotoxigenic E coli (ETEC) isolated from cattle in Belgium. One hundred thirty-two E coli isolates (15.2%) hybridized with probes STaP, K99, and/or F41. The 5 other probes were not hybridized by E coli isolates. Therefore, only STaP enterotoxin and K99 and F41 adhesins were virulence factors of ETEC isolated from cattle. Two major pathotypes accounted for 95% of the ETEC: STaP+K99+F41+ (67.4%) and STaP+K99+ (27.3%). The last 5% of probe-positive isolates had STaP+, STaP+F41+, or K99+F41+ minor pathotypes. Of 12 American ETEC isolates also assayed, 7 were positive with STb and/or 987P probes (pathotypes STaP+STb+, STaP+ 987P+, or STaP+STb+987P+) and may be porcine- rather than bovine-specific enteropathogens. The remaining 5 American ETEC isolates belonged to 3 minor pathotypes (STaP+, STaP+F41+, and K99+F41+) also found among Belgian E coli isolates. Such isolates may be derivatives of STaP+K99+F41+ or STaP+K99+ ETEC after in vivo or in vitro loss of virulence genes and/or non-ETEC isolates, which have acquired virulence genes by in vivo transfer.  相似文献   

2.
Pregnant gilts were vaccinated with two doses of alhydrogel adsorbed fimbrial antigens of Escherichia coli (K88ab, K88ac, K99 and 987P) supplemented with beta toxoid of Clostridium perfringens type C. Their piglets, and piglets of nonvaccinated gilts, were subsequently orogastrically challenged with one or other of the four fimbrial types of enteropathogenic E coli. Some of the vaccinated animals were reinjected with a single dose of the vaccine during second gestation and their piglets, and piglets of non-vaccinated sows, were challenged the same way as were litters of gilts. Blood serum and colostra were examined for antibodies to the four fimbrial antigens of E coli and for antitoxin to beta toxin of C perfringens type C. It was found that: (1) a highly significant reduction in mortality and morbidity was achieved in vaccinated litters against all four challenge strains of E coli; (2) excretion of K88ab and K88ac but not of K99 and 987P challenge strains was significantly reduced; (3) revaccination of sows by a single dose of the vaccine during second gestation conferred complete protection against mortality and highly significant protection against morbidity; (4) no correlation was noted between colostral or seroagglutinins to fimbrial antigens of E coli and mortality rates in litters challenged with homologous fimbrial types of E coli, but good correlation was found between colostral precipitins to K88 antigens and mortality rates in litters; (5) antitoxin value in 97 per cent of colostrum of vaccinated sows was 10 iu equivalent of C perfringens type C toxin or more per ml of colostrum.  相似文献   

3.
Enterotoxigenic Escherichia coli (ETEC)-associated post-weaning diarrhea (PWD) is economically one of the most important diseases for the swine industry. Porcine ETEC strains typically express K88 or F18 fimbria and heat-labile (LT) and/or heat-stable (STa, STb) enterotoxins. However, recent studies indicate that EAST1 toxin, adhesin involved in diffuse adherence (AIDA-I) and porcine attaching and effacing-associated factor (paa) may also be expressed by ETEC strains associated with diarrhea. To better understand the virulence factors of E. coli strains that cause PWD, we applied PCR to screen for K88, F18, F41, 987P and K99 fimbrial genes; LT, STa, STb, Stx2e and EAST1 toxic genes; and AIDA-I, paa and EAE adhesin genes in E. coli strains recently isolated from young pigs with PWD in the US. Of 304 E. coli isolates from diarrheic pigs submitted for testing, 175 (57.6%) strains possessed fimbrial genes: K88 (64.6%), F18 (34.3%), F41 (0.57%), K99 (0.57%), 987P (0); toxin genes: LT (57.7%), STb (72.6%), STa (27.4%), STx2e (17.4%), EAST1 (35%); and adhesin genes: AIDA-I (26.9%), paa (60%), EAE (1.1%). All toxin genes except the EAST1 toxin gene, were almost exclusively associated with K88+ or F18+ isolates, and most of these isolates carried multiple toxin genes. The non-fimbrial adhesin paa was found present in over half of the K88+ isolates. A total of 129 (42%) isolates carried no fimbrial genes, including 66 (21.7%) isolates that did not have any of the above virulence genes. These results suggest a broad array of virulence genes associated with PWD in pigs.  相似文献   

4.
Virulence of enterotoxigenic Escherichia coli (ETEC) is associated with fimbrial adhesins and enterotoxins such as heat-labile (LT) and/or heat-stable (ST) enterotoxins. Previous studies using a cell culture model suggest that exclusion of ETEC from attachment to epithelial cells requires expression of both an adhesin such as K88 (F4) fimbriae, and LT. To test the ability of non-pathogenic E. coli constructs to exclude virulent ETEC sufficiently to prevent clinical disease, we utilized a piglet ETEC challenge model. Thirty-nine 5-day-old piglets were inoculated with a placebo (control), or with either of the three K88(+)E. coli strains isogenic with regard to modified LT expression: 8017 (pBR322 plasmid vector control), non-toxigenic mutant 8221 (LT(R192G)) in pBR322, or 8488, with the LT gene fused to the STb gene in pBR322 (LT(R192G)-STb). Piglets were challenged with virulent ETEC Strain 3030-2 (K88(+)/LT/STb) 24h post-inoculation. K88ac receptor-positive piglets in the control group developed diarrhea and became dehydrated 12-24h post-challenge. Piglets inoculated with 8221 or 8488 did not exhibit clinical signs of ETEC disease; most piglets inoculated with 8017 showed diarrhea. Control pigs exhibited significant weight loss, increased blood total protein, and higher numbers of colony-forming units of 3030-2 E. coli in washed ileum and jejunum than treated pigs. This study shows for the first time that pre-inoculation with an avirulent strain expressing adhesive fimbriae and a non-toxic form of LT provides significant short term protection from challenge with a virulent ETEC strain that expresses the same fimbrial adhesion and enterotoxin.  相似文献   

5.
Cheng D  Sun H  Xu J  Gao S 《Veterinary microbiology》2005,110(1-2):35-39
F18ab and F18ac are important fimbrial colonization factors of verotoxigenic Escherichia coli (VTEC) and/or enterotoxigenic E. coli (ETEC) in weaned piglets with edema disease and/or diarrhea. To further investigate their prevalence and correlation to pathogenic E. coli, a duplex PCR, using three primers derived from the nucleotide sequence of the F18 major fimbrial subunit gene (fedA), and a direct agglutination test, using a monoclonal antibody specific for the antigenic factor 'a' of F18, were performed. Among 60VTEC, 24VTEC/ETEC and 24 ETEC isolates tested from weaned piglets with edema disease and/or diarrhea in different pig farms in the Jiangsu Province of China, 52 isolates (48.15%) were positive in the direct agglutination test and 63 isolates (58.33%) were positive in the duplex PCR. Among 63 PCR-positive isolates, 53 isolates (49.07%) were F18ab-positive and 10 isolates (9.26%) were F18ac-positive. In addition, the F18ab gene was more frequently detected in VTEC (61.67%) or VTEC/ETEC (62.50%) than in ETEC (4.17% only), while the F18ac gene was more frequently detected in VTEC/ETEC (33.33%) than in ETEC (8.33%) or VTEC (0%). Furthermore, F18ab was more frequently associated with Shiga toxin 2e (Stx2e), whereas F18ac was more frequently associated with enterotoxin ST I. These results suggest that the duplex PCR performed in this experiment is a more reliable method for identification of F18+E. coli, and that F18 is a more important virulence factor of VTEC and VTEC/ETEC.  相似文献   

6.
《Veterinary microbiology》1998,59(4):283-294
F41-positive and F41-negative derivatives of bovine enterotoxigenic Escherichia coli strain B41 carrying K88 or K88 and K99 plasmids were investigated for stability and expression of genes for their fimbrial antigens. Either K88 plasmid alone or both K88 and K99 plasmids could be maintained in these strains though stability could depend on culture medium. K99 antigen could be detected in each strain bearing K99 plasmid. Clones that produced K88 antigen or clones that did not produce this antigen could be isolated from each strain, except from the strain that possessed K99 plasmid in the strain that did not possess the ability to produce F41 antigen. Strains possessing K88 plasmid in the strain able to produce F41 antigen produced clones expressing either both K88 and F41 antigens, (also F41 appeared strongly expressed in some clones) or clones that produced only F41 antigen or no antigen at all. Clones that produced only K88 antigen or others that did not produce this antigen could be produced from a strain bearing only K88 plasmid and that did not possess the ability to produce F41 antigen. None of these strains bearing K88 plasmid alone or additionally K99 plasmid produced mannose-resistant hemagglutination of horse or sheep erythrocytes at 20°C as found for K99 and F41 ETEC natural strains, respectively. These results suggested that the structures of pili when several genetic determinants were present simultaneously may not be identical to those of original strains. In this study, clones expressing either one, two or three adhesin bearing antigens could be obtained from the strain B41.  相似文献   

7.
Attempts to develop live vaccines to protect against enterotoxigenic Escherichia coli (ETEC) infection by induction of both cell-mediated and mucosal immunity, and serum antibody responses have included use of recombinant Salmonella strains that produce K88 fimbrial antigens (Hone et al., 1988; Attridge et al., 1988; Morona et al., 1994). However, none of the recombinant Salmonella vectors has been licensed by the United States Department of Agriculture (USDA) for use as a live vaccine in pigs in the United States. A variant of Salmonella enterica ser. Choleraesuis strain 54 (SC54) is currently used as a safe and effective intranasal attenuated live vaccine in pigs. In order to expand the efficacy of this live vaccine strain, we sought to modify strain SC54 to express the K88 antigens of ETEC. To accomplish this, a plasmid-based system was used to integrate the K88 gene cluster into the chromosome of strain SC54 by site-specific recombination. The K88 antigens were expressed by strain SC54, and the gene cluster was stably maintained in the host.  相似文献   

8.
Enterotoxigenic Escherichia coli (ETEC) in farm animals.   总被引：11，自引：0，他引：11  
Animal diseases due to enterotoxigenic Escherichia coli (ETEC) typically appear as severe watery diarrhoea during the first few days of life (also a few days after weaning in pigs). ETEC adhere to the small intestinal microvilli without inducing morphological lesions and produce enterotoxins acting locally on enterocytes. This action results in the hypersecretion (of water and electrolytes) and reduced absorption. Adhesins and toxins are the two prominent virulence attributes of ETEC and the level of knowledge of these factors determines the chances for successful prevention and therapy of the disease. For animal ETEC the most common adhesins are the fimbriae (pili) on the surface: F4(K88), F5(K99), F6(987P), F41, F42, F165, F17 and F18. Enterotoxins (extracellular proteins or peptides) of animal ETEC are classified as heat-labile (LT) and heat-stable (ST) enterotoxins with further subdivisions (LTh-I, LTp-I, LTIIa, LTIIb, STaH, STaP, STb) according to antigenic and biological differences. Fimbriae and LT enterotoxins are made up of large molecular weight proteins which facilitate their utilisation in vaccines and their detection using immunodiagnostic systems. The adhesive fimbriae and enterotoxins of animal ETEC are plasmid determined (except F41 and F17). Virulence gene probes (DNA hybridisation, PCR) are specific and sensitive diagnostic and epidemiologic tools for the detection of ETEC. Based on genetic typing, the ETEC, in spite of limited serogroups, seem to represent a population of E. coli with a diverse genetic background.  相似文献   

9.
Faeces samples collected from diarrhoeic dairy calves in the first 8 weeks of life were examined for the presence of 5 enteropathogens. The majority of the 108 diarrhoea cases occurred in the first 5 weeks of life and a commercial ELISA kit detected bovine enteric coronavirus (BEC) in 38.9%, serogroup A rotavirus (RV) in 16.7% and K99 (F5) fimbrial adhesin-positive Escherichia coli (K99 ETEC) in 11.1 per cent. Concurrent infections of these enteropathogens were detected in 14.8% of samples (30.8% of samples positive for these agents). No evidence of cryptosporidial infection was found using a differential staining method on faecal smears nor was salmonella excretion detected. On 2 of the 8 farms only BEC was present; the other 6 farms were positive for all 3 agents. It is concluded that BEC is the major infectious cause of neonatal calf diarrhoea in the Ethiopian dairy herds studied with RV and K99 ETEC also contributing to morbidity, either alone or as mixed infections.  相似文献   

10.
The study comprises fifty 4 to 12 weeks old pigs that died from oedema disease or severe diarrhoea. Smears were prepared from the mucosa of duodenum, jejunum and ileum, and by immunofluorescence F107 fimbrial antigens were detected. E. coli. strains were isolated from the intestines and were characterised by slide agglutination (serogroup and F107 fimbriae production), by their cytotoxicity for Vero cells, and by gene amplification (genes coding for the major F107 subunit FedA, the toxin causing oedema disease SLT-IIv, and enterotoxins LTI, STIa and STII). F107 fimbriae were demonstrated in association with E. coli of serogroups O139:K12 and O141:K85a,b but not of serogroup O149:K91:F4a,c. Expression in culture of F107 fimbriae by some isolates gave additional evidence for production of these fimbriae by ETEC strains. The genetic determinant of SLT-IIv was found in association with F107, and could not be detected in serogroup O149:K91:F4a,c. Gene fedA was demonstrated in two isolates which were devoid of SLT-IIv. Most isolates from cases of oedema disease belonged to serogroup O139:K12 and did not contain enterotoxin genes. Isolates from pigs that suffered from diarrhoea were serotyped O141:K85a,b or O149:K91:F4a,c, and carried at least two enterotoxin genes in their genomes. In a small proportion of the cases F107 antigens were demonstrated in intestinal smears although gene fedA was not detected in the corresponding isolates. The results confirm the importance of F107 fimbriae as virulence factor in oedema disease E coli strains, but also demonstrate that F107 fimbriae can be found in association with postweaning diarrhoea isolates. In these latter strauns enterotoxins were always demonstrated, irrespective of the presence of toxin SLT-IIv.  相似文献   

11.
近年来，从华东地区患腹泻仔猪中分离到一些表达K88菌毛的大肠杆菌，这些菌株只与K88a因子单抗反应，而不与b、c、d因子单抗反应。通过K88常规血清交叉吸收试验、SDS-PAGE、Western印迹，表明这些菌株不仅与K88ac参考菌株C83907制备的c因子血清反应，而且与以分离株SEC586制备且经K88ab、K88ac、K88ad参考菌株吸收后的血清也反应。对分离株SEC586、SEC464的K88主要亚单位结构基因faeG的克隆、测序，发现该基因由846对核苷酸组成，编码菌毛主要亚单位的262个氨基酸及21个氨基酸的信号肽，比国外报道的K88ac FaeG亚单位（263个氨基酸）少了1个氨基酸，比K88ab、K88ad（265个氨基酸）少了3个氨基酸。SEC586、SEC464菌株的FaeG亚单位氨基酸序列的同源性为97．7％，它们与K88ac的同源性为94．7％和96．2％；与K88ab的同源性为90．1％和91．2％；与K88ad的同源性为87．0％和88，6％。结果表明，新分离的K88ac大肠杆菌黏附素主要亚单位已发生了部分变异。  相似文献   

12.
Faecal samples from 148 dogs with diarrhoea and from 15 healthy dogs were cultured for bacterial pathogens with enterotoxigenic properties. The aim of the study was to define the toxin profile (production of heat-labile [LT] and heat-stable [ST] toxins) and possible surface fimbrial antigens. Enterotoxigenic bacteria were isolated from 6 (4.1%) dogs with diarrhoea, four of these were Escherichia coli and two were Klebsiella pneumoniae. The E. coli strains and K. pneumoniae strains were producing both LT and ST toxins. The LT toxin from these strains was not neutralized by human anti-LT serum or anti-choleragen and did not cause coagglutination with Staphylococcus aureus coated with anti-human-LT. This suggests that the LT toxin produced by these canine isolates is non-identical to LT toxin from human strains.

Three of the ETEC strains were haemagglutinating and showed surface hydrophobic properties. Electron microscopy showed that canine ETEC isolates possessed fimbriae of two different types: thick (5–5.5 nm) and thin (2–3 nm).  相似文献   


13.
Enterotoxigenic Escherichia coli (ETEC) infections result in large economic losses in the swine industry worldwide. The organism causes diarrhea by adhering to and colonizing enterocytes in the small intestines. While much progress has been made in understanding the pathogenesis of ETEC, no homologous intestinal epithelial cultures suitable for studying porcine ETEC pathogenesis have been described prior to this report. In the current study, we investigated the adherence of various porcine ETEC strains to two porcine (IPEC-1 and IPEC-J2) and one human (INT-407) small intestinal epithelial cell lines. Each cell line was assessed for its ability to support the adherence of E. coli expressing fimbrial adhesins K88ab, K88ac, K88ad, K99, F41, 987P, and F18. Wild-type ETEC expressing K88ab, K88ac, and K88ad efficiently bound to both IPEC-1 and IPEC-J2 cells. An ETEC strain expressing both K99 and F41 bound heavily to both porcine cell lines but an E. coli strain expressing only K99 bound very poorly to these cells. E. coli expressing F18 adhesin strongly bound to IPEC-1 cells but did not adhere to IPEC-J2 cells. The E. coli strains G58-1 and 711 which express no fimbrial adhesins and those that express 987P fimbriae failed to bind to either porcine cell line. Only strains B41 and K12:K99 bound in abundance to INT-407 cells. The binding of porcine ETEC to IPEC-J2, IPEC-1 and INT-407 with varying affinities, together with lack of binding of 987P ETEC and non-fimbriated E. coli strains, suggests strain-specific E. coli binding to these cell lines. These findings suggest the potential usefulness of porcine intestinal cell lines for studying ETEC pathogenesis.  相似文献   

14.
DNA gene probes specific for genes encoding heat labile enterotoxin (LTI), heat stable enterotoxins (STIa, STII), vero cytotoxins (VT1, VT2), and adhesins K88 (F4), K99(F5), F41 and 987P(F6) were used to examine 873 isolates of E. coli from cases of diarrhoea (680 from pigs, 187 from cattle and six from sheep). A total of 188 were toxin gene positive and of these 84 belonged to the classical ETEC serogroups. Of the other 104 toxin gene positive strains, 80 hybridized with the VT2 probe of which 34 were from cases of porcine post-weaning diarrhoea belonging to serogroup 0138:K81 and 22 were untypable strains from cattle.  相似文献   

15.
A modified, double-antibody, enzyme-linked immunosorbent assay (ELISA) was developed to detect the K99 pilus antigen of enterotoxic Escherichia coli (ETEC) in feces of calves. Extremely high positive to negative ratios (greater than 200) were obtained by using monoclonal antisera as the primary antibody. Strong positive reactions were obtained with strains of E coli known to produce the K99 antigen; however, non-enteropathogenic E coli (strains not producing the K99 antigen), Salmonella, Proteus, Klebsiella, Pseudomonas, Staphylococcus, Streptococcus, and rotavirus produced negative results. Seventy-five fecal samples, 8 from healthy calves and 67 from calves with neonatal calf diarrhea were examined with the K99 ELISA for the presence of ETEC. Rotavirus test and fecal culture results were available on feces from calves with diarrhea and were used with the K99 ELISA results to determine the specific cause of the disease. Enterotoxic E coli was the predominant agent detected in the feces of 29 diarrheal calves less than 5 days of age. Mixed infections of rotavirus and ETEC were also common in these calves, but rotavirus infections alone were not detected. In 38 calves greater than or equal to 5 days, rotavirus was detected without ETEC. Of these calves, only 2 produced positive tests with the K99 ELISA. Salmonella sp and Proteus sp were detected from 5 of 67 calves with diarrhea.  相似文献   

16.
We tested hemolytic E. coli from 86 pigs with edema disease or colidiarrhea. They were tested serologically and with nonradioactive digoxigenin-dUTP labelled probes for the presence of enterotoxin or Shiga-like-toxin genes. By slide-agglutination we detected 38 cases with E. coli O149:K88, 28 with E. coli O139:82B and 20 with E. coli O141. E. coli of serogroup O149:K88 isolated from diarrheic pigs, reacted with the probes for LT and STb genes. Edema disease E. coli O139:82B reacted with the SLTII probe. E. coli O141, isolated from colidiarrhea or edema disease showed a diversity of toxin gene patterns. All the E. coli O141 from diarrheic pigs reacted with the probes for LT and STap in addition to SLTII. No strains isolated from pigs with edema disease possessed any of these enterotoxin genes. Gene probe technique confirmed the serological method as useful tool for diagnosing E. coli O149:K88 and O139:82B as ETEC or VTEC, respectively. On the other hand only the demonstration of toxin genes with probes could explain the pathological findings in the pigs shedding E. coli of serogroup O141.  相似文献   

17.
Coagglutination was used to detect K88 and K99 fimbrial antigens on Escherichia coli, and results were compared to an enzyme immuno assay (EIA). When pili suspensions were tested by both methods, 28 of 66 cultures were shown to have K88 and 11 of 31 cultures had K99 antigens. No pili suspensions were positive by coagglutination that were not positive by EIA. Testing of cell suspensions gave equivalent results to pili suspensions for K99 when tested by coagglutination. Two cell suspensions reacted with the K88 coagglutination which could not be confirmed by testing of pili suspensions, while a further 20 out of 43 cultures gave equivalent results with both cell and pili suspensions for K88 when tested by coagglutination.  相似文献   

18.
The presence of porcine toxigenic E. coli (ETEC, VTEC) in 28 piggeries (5% of total) of the central and northeast region of Argentina was studied for a better understanding of the epidemiology of porcine strains. Samples were taken by rectal swabs from healthy piglets and from those with diarrhoea, in addition to their dams. Between 5-10 colonies were isolated from each one of 223 animals sampled from 1992 to 1997. By using specific primers each strain was screened by PCR for VT1, VT2all, VT2e, STIa, and LTI toxin genes. Only strains positive for any of the toxins mentioned above were screened for STb. Their O serogroups were determined by agglutination. All of the above enterotoxins and verocytotoxins were found in E. coli isolated from the animals. The STIa gene was detected in E. coli isolated from 27/127 piglets with diarrhoea, in comparison with LTI (4/127 pigs). No toxin gene was amplified from E. coli isolated from either healthy piglets or their dams. When strains isolated from 48 piglets without diarrhoea but showing delayed growth were analysed by PCR, their toxin profile was determined to be VT1 (1/48 piglets), VT2all (5/48), STIa (1/48), LTI (3/48) and VT2e (3/48). Serogroup O64 prevailed among ETEC; O138 prevailed for ETEC/VTEC strains. This is the first extensive study regarding porcine toxigenic E. coli in Argentina and constitutes an important database for the implementation of prevention measures.  相似文献   

19.
A novel common colonization factor was detected in enterotoxigenic strains of Escherichia coli isolated from intestinal contents of piglets affected with postweaning diarrhoea. This factor was antigenically distinct from the previously described K88, K99, F41, 987P, CFAI, CFAII and Att25 fimbrial antigens. E. coli strains possessing this factor adhered to the pig intestinal brush borders and one strain, used in experimental infection in weanlings, colonized the intestinal epithelium and induced diarrhoea. Examination of 212 toxigenic strains of E. coli isolated from weanlings revealed the presence of the novel common colonization factor in 83 strains, belonging to serogroups O25, O108, O138, O141, O147 and O157. The antigen K88 was detected in 47 strains belonging to serogroups O8, O141, O147 and O149.  相似文献   

20.
The haemagglutinating properties (HA) of Escherichia coli possessing the fimbrial antigens K88 and K99 were significantly affected by tiamulin. HA for K88-positive strains was reduced at 1/10 to 1/40 minimum inhibitory concentration (MIC), while the corresponding values for the K99 strains were 1/2 to 1/10 MIC. Tiamulin concentrations of 3 to 12 micrograms ml-1 increased salt aggregation test values from between 1.4 and 1.5, to more than 2.  相似文献   

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