Prevalence of the enteroaggregative Escherichia coli heat-stable enterotoxin 1 gene and its relationship with fimbrial and enterotoxin genes in E. coli isolated from diarrheic piglets.

A total of 720 Escherichia coli strains isolated from diarrheic piglets on 756 swine farms were screened for the presence of the enteroaggregative E. coli heat-stable enterotoxin 1 (EAST1) gene by polymerase chain reaction (PCR). Escherichia coli strains that carried EAST1 genes were also tested by PCR for the presence of 4 fimbriae (F4, F5, F6, F41), 2 heat-stable enterotoxins (STa and STb), and 1 heat-labile enterotoxin (LT) gene. One hundred sixty-four (22.7%) of the 720 E. coli isolates carried genes for EAST1. Of these 164 isolates, 62 (37.8%) carried EAST1 genes only, 11 (6.7%) carried genes for at least 1 of the fimbrial adhesins, 51 (31.1%) carried genes for at least 1 of the enterotoxins, and 40 (23.8%) carried genes for at least 1 of the fimbrial adhesins and enterotoxins. Forty-six percent of strains that carried EAST1 genes carried STa genes, and 16% of strains that carried EAST1 genes carried F4. The isolation rate of enterotoxigenic E. coli strains carrying genes for EAST1 gene was 63%. The 6 major genotypes observed in this study (in decreasing order) were EAST1+, EAST1+STa+, EAST1+STa+STb+, EAST1+STa+F5+, EAST1+STa+F4+, and EAST1+STb+F4+. EAST1 is widely prevalent among diarrheagenic strains of E. coli and may represent an important virulence determinant in the pathogenesis of enteric colibacillosis of preweaned pigs.     

Prevalence of the enteroaggregative Escherichia coli heat-stable enterotoxin 1 (EAST1) gene in isolates in weaned pigs with diarrhea and/or edema disease

A total of 476 Escherichia coli isolated from weaned pigs with diarrhea and/or edema disease were screened for the presence of the enteroaggregative E. coli heat-stable enterotoxin 1 (EAST1) gene by polymerase chain reaction (PCR). E. coli strains that carried EAST1 genes were also tested by PCR for the presence of genes for five fimbriae (F4, F5, F6, F18 and F41), two heat-stable (STa and STb) and one heat-labile (LT) enterotoxin, and Shiga toxin 2e (Stx2e). One hundred and forty nine (31.3%) of the 476 E. coli isolates carried the gene for EAST1. Of these 149 isolates, 66 (44.3%) carried the east1 gene only and 83 (55.7%) carried genes for the fimbrial adhesins or enterotoxins. E. coli which carried east1 gene also possessed genes for STa or F4 frequently. EAST1 may represent an additional determinant in the pathogenesis of E. coli diarrhea in weaned pigs.     

猪源大肠杆菌热稳定肠毒素的检测

致病性大肠杆菌产生一种或多种肠毒素称为肠毒素型大肠杆菌(Enterotoxigenic E．COLI ETEC)．ETEC是引起幼畜腹泻的重要病原菌。ETEC能借助于所产生的菌毛抗原粘附于动物小肠黏膜。定居并产生作用于肠壁的外毒素，称为肠毒素。主要有两类肠毒素：一类是不耐热性肠毒素即热敏感肠     

Detection of the astA (EAST1) gene in attaching and effacing Escherichia coli from ruminants

A total of 206 attaching and effacing Escherichia coli strains from ruminants were analysed for the presence of the astA (EAST1) and bfpA genes. None of these strains was bfpA-positive. The percentage of enteropathogenic E. coli (EPEC) strains astA-positive found in healthy cattle (15.6%) suggests that this animal species may be a significant reservoir of atypical EPEC potentially pathogenic for humans.     

Analysis of the AIDA-I gene sequence and prevalence in Escherichia coli isolates from pigs with post-weaning diarrhoea and oedema disease

Three hundred and twenty-four strains of Escherichia coli isolated from weaned pigs with diarrhoea or oedema disease in Eastern China were screened by multiplex PCR for the presence of the gene encoding adhesin involved in diffuse adhesion I (AIDA-I). Two AIDA-I positive strains were subjected to analysis of the nucleotide sequence of the complete orfA and orfB of the AIDA gene. The AIDA-I positive E. coli isolates were also assessed for five fimbriae (F4, F5, F6, F18 and F41) by monoclonal antibodies and for toxin genes (STa, STb, LT, EAST1, Stx2e) by PCR. Twenty-one (6.5%) of the isolates possessed AIDA-I genes. Of these isolates, two carried AIDA-I genes as the only demonstrated virulence factors, and the remaining isolates carried other virulence factor genes. Comparing the AIDA-I sequence from porcine and human sources, a high homology of orfA both in porcine E. coli and human E. coli was observed. However, each orfB of the two porcine E. coli isolates was 3864 nucleotides long compared with 3861 for the E. coli 2787 orfB, and showed 96.5% homology to E. coli 2787. The data indicated (1) that AIDA-I may be an occasional virulence factor in post-weaning diarrhoea and oedema disease in pigs, (2) that it has the potential to transfer between porcine and human E. coli, and (3) that there is a genetic diversity in orfB between human and porcine E. coli.     

Investigation of putative CDT gene in Escherichia coli isolates from pigs with diarrhea

In this study, 98 Escherichia coli isolates from 42 diarrheic neonatal piglets were screened for the presence of cytolethal distending toxin coding gene by polymerase chain reaction (PCR). PCR yielded a single product which was specifically generated for E. coli cdt(+) control strain and not for other control strains. Twenty two (22.4%) of the isolates tested were cdtB positive, and 50% of the cdtB(+) isolates were also estII positive. The most prevalent pathotype was O32 cdtB(+) estII(+), which accounted for 59% of the cdtB positive strains. These results indicate an association between the presence of the cdtB gene and diarrhea, and support the need for further studies to determine the role of this toxin in diarrhea.     

Escherichia coli heat-stable enterotoxin in feces and intestines of calves with diarrhea

Two experiments were conducted to evaluate detection of Escherichia coli heat-stable enterotoxin (ST) in the feces of calves as a method for implicating E coli in neonatal calf diarrhea. The first experiment evaluated the use of the infant mouse test for detection of ST in the feces of calves with naturally occurring diarrhea. Simultaneous identification of bovine enteropathogenic strains of E coli (EEC) and of other infective agents implicated in neonatal calf diarrhea was attempted in these samples. The ST was detected with certainty in only 7 of 41 samples from calves less than or equal to 3 weeks old. Enteropathogenic E coli, however, was detected in 27 samples. In 23 of these 27 samples, EEC was the only recognizable diarrheagenic agent. In a small percentage of the samples, Salmonella, rotavirus, coronavirus, and cryptosporidium were recognized alone, in combination with each other, or with EEC. In the second experiment, 6 calves were fed colostrum from cows inoculated with the bovine EEC strain B44; 6 were given colostrum from cows vaccinated with non-EEC strain 28F, and 4 were given milk from nonvaccinated heifers. Two of the calves that were given colostrum from cows inoculated with strain B44 were challenge exposed with the non-EEC strain 28F. The remaining calves were challenge exposed with the EEc strain B44. Fecal samples were taken from these calves at intervals and were examined for the presence of ST and of the challenge-exposure organism. The ST was detected in approximately one half of the fecal samples obtained, and it was most often detected in the early stages of the induced diarrhea. Calves were observed to shed the challenge-exposure EEC strain for long periods in the absence of diarrhea or detectable amounts of ST in the feces. The ST was detectable in fecal samples when the diarrhea was severe and when the dry matter content of the fecal samples was low.     

Protection against neonatal Escherichia coli diarrhoea in pigs by vaccination of sows with a new vaccine that contains purified enterotoxic E. coli virulence factors F4ac, F4ab, F5 and F6 fimbrial antigens and heat-labile E. coli enterotoxin (LT) toxoid

The efficacy of a new vaccine against neonatal Escherichia coli diarrhoea in piglets containing purified F4ab, F4ac, F5 and F6 fimbriae and detoxified heat-labile toxin (LT) was tested in challenge experiments by the method described by the European Pharmacopoeia (3rd edn, EDQM, Council of Europe, Strasbourg, France). A group of 11 young sows from a herd without E. coli problems was vaccinated 6-8 and 2-4 weeks prior to expected farrowing and another group of nine young sows were non-vaccinated controls. Escherichia coli antibody titres were determined in serum samples taken from the sows before first vaccination and before farrowing and in colostrum samples. The newborn piglets were allowed to suckle colostrum from their mother immediately after birth. The piglets were marked with individually numbered ear tags. Approximately 12 h after birth, 118 piglets from vaccinated sows and 79 piglets from non-vaccinated control sows were challenged by oral instillation of 5 ml of a freshly prepared culture of one of the challenge strains [O8:K87:F4ab (LT+) or O149:K91:F4ac (LT+) or O9:K30:F5 or O9:K103:F6 respectively]. The challenge cultures contained as a mean 6.8x10(9) CFU/ml. After challenge the piglets were observed for 7 days and mortality and morbidity were recorded. Vaccinated sows developed significant levels of antibody titres in colostrum and serum. Control sows stayed at a low/seronegative level. The protective efficacy was excellent because 66.7-87.5% of the piglets from vaccinated sows remained without clinical signs after challenge. Only 0.0-28.0% of the piglets from non-vaccinated sows remained healthy and more than 47.1% of the piglets in this group died after challenge. It is concluded that the new vaccine is very effective in protection of piglets against neonatal E. coli diarrhoea.     

Preliminary characterization of Escherichia coli isolated from pigs with diarrhoea in Venezuela

The aetiology of neonatal porcine diarrhoea was studied in 15 different herds located in the north-western region of Venezuela. Of 56 strains of Escherichia coli analyzed, 16 (28.6%) were shown to produce heat-stable (STa) enterotoxin, as detected by infant mouse assay. Only four of these STa+ isolates also possessed the K88 pilus antigen, two were 987P+ and none possessed the K99 antigen, leaving 10 STa+ samples in which no pilus antigen was identified. Among the 40 STa negative samples were six K88+ specimens, one K99+, four 987P+, one which reacted as K88+ + K99+ and one K88+ + 987P+. Considering as pathogenic any strain showing at least one of the characters studied, pathogenic E. coli were detected with an overall frequency of 42.9%, being more prevalent during the second week of life. An electrophoretic analysis of the plasmid content of the field isolates of E. coli, revealed the presence of numerous species of extrachromosomal DNA, although no direction association could be made between a particular plasmid and any of the pathogenic characteristics identified. Results of Southern blot analysis indicate that the STa enterotoxin was preferentially encoded within an endemic plasmid of 4.9 Md. Other plasmids present in the E. coli isolates could be related to antibiotic resistance. With the exception of one strain, all E. coli isolates were resistant to more than one of the nine drugs tested; multiresistant E. coli were frequently isolated, including four strains which were resistant to seven antibiotics.     

Escherichia coli diarrhoea in pigs with or without the K88 receptor.

There was a high incidence of neonatal scours in 38 litters of pigs born at Compton in a four month period during 1978. The most important cause of the disease was an enteropathogenic Escherichia coli strain which possessed the K88 antigen. The Compton herd has been bred to produce pigs of three genotypes with respect to the presence or absence of the intestinal receptor for the K88 antigen. These are homozygous dominants (SS) and heterozygotes (Ss) susceptible to infection by virulent K88-positive E coli, and homozygous recessives (ss) resistant to the disease. The highest incidence of diarrhoea was in the susceptible progeny of resistant dams and susceptible sires. There was no K88 associated diarrhoea in resistant progeny or in susceptible progeny of susceptible dams.     

Escherichia coli heat-stable enterotoxin b (STb) in vivo internalization within rat intestinal epithelial cells

Heat-stable enterotoxin b (STb) is a low molecular weight toxin known to bind sulfatide, its receptor. The fate of STb bound to rat intestinal epithelium cells was followed using an anti-toxin gold labeled assay and transmission electron microscopy. The data suggest that STb toxin and the fusion protein maltose binding protein (MBP)-STb were internalized whereas its mutant I41 E-M42R with reduced hydrophobicity did not show internalization. There was a significant difference in the mean of gold particles per field between rat intestine incubated with STb or the fusion protein MBP-STb and the negative control consisting of intestine incubated with PBS alone. No subcellular compartment seems to be particularly aimed by the toxin as gold particles were randomly distributed within the cell.     

Effects of Escherichia coli heat-stable enterotoxin b on small intestinal villi in pigs, rabbits, and lambs

Culture supernates from two strains of E. coli were placed into different ligated intestinal sections (loops) of each animal. The two bacterial strains were identical except that one contained a plasmid carrying the heat-stable toxin b (STb) gene, while the other did not. Morphometric techniques were used to assess villous epithelial surface areas and mucosal volumes in both intestinal segments exposed to STb-positive (test) and to STb-negative (control) supernates. In pigs whose intestines were exposed to STb-positive supernatants for 2 hours, both villous epithelial surface area and mucosal volume were significantly smaller in test loops than in control loops (P less than 0.02). In test loops of pigs incubated for 1 hour, and in test loops of lambs incubated for 2 hours, there was a decrease in villous epithelial surface area which approached the test for significance but did not meet it (0.05 less than P less than 0.10). Rabbit test loops did not differ from rabbit control loops in either villous epithelial surface area or mucosal volume. Histological examination of the tissues from all three species revealed epithelial changes in porcine and ovine tissues only. In porcine and ovine tissues, epithelium at villous tips was seen to be cuboidal or squamous, or even to be absent. Villi with similarly altered epithelium were seen in control loops, but were seen much more frequently in test loops. These epithelial changes were seen as early as 30 minutes of incubation in pigs. Intestinal tissues from these pigs were examined by transmission electron microscopy, but no difference between test and control tissues was seen.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effect of salicylates on intestinal secretion in calves given (intestinal loops) Escherichia coli heat-stable enterotoxin

The inhibitory effect of salicylates on intestinal secretion in 1- to 5-day-old calves given Escherichia coli heat-stable enterotoxin (ST)-induced intestinal fluid response was investigated. Purified ST was diluted in isotonic saline solution to obtain 1:10, 1:25, 1:50, 1:75, and 1:100 dilutions. Each dilution (1 ml) was inoculated into ligated loops in the distal part of the jejunum of each calf. Acetylsalicylic acid (aspirin) given orally (100 mg/kg) at 4 hours before ST was inoculated did not substantially alter the intestinal fluid response to ST. Sodium salicylate (IV) infusion, begun simultaneously when, or at 1 hour after, ST was inoculated, significantly (P less than 0.05) decreased fluid accumulation in those loops inoculated with ST dilutions of 1:25 or greater. The sodium and potassium concentrations of the accumulated fluid did not differ significantly between or within treatment groups. These results indicate that sodium salicylate infusion may be beneficial in treating enterotoxic colibacillosis in calves. Aspirin given orally at the dose used in the present study, would not have any beneficial effect.     

Effect of jejunal loop location on the activity of Escherichia coli heat-stable enterotoxin in 4- to 5-week-old pigs

Heat-stable enterotoxin (STa) from Escherichia coli strain 431 was injected into the jejunum of 4 pigs from each of 3 litters, using a ligated intestinal loop assay (with loops beginning 1 m caudad to the pylorus). The jejunum was divided into 4 contiguous areas, with 4 loops in each area. Doses of 0, 10, 100, or 1,000 ng of purified toxin (10 ng/mouse unit) were injected into the loops within an area, using a 4 X 4 Latin square design. Fluid accumulation in the loops increased (P less than 0.05) with increasing concentrations of STa in pigs in all litters, but the magnitude of the response varied across litters. Fluid responses to the STa varied in the different areas of the jejunum, with pigs in 2 litters having a decrease (P less than 0.05) in the response to STa in the caudal areas. These data quantitate the variability within the different areas of the jejunum of the young pig.     

Genotypic prevalence of the adhesin involved in diffuse adherence in Escherichia coli isolates in pre-weaned pigs with diarrhoea in Korea

A total of 1002 Escherichia coli strains isolated from pre-weaned pigs with diarrhoea on 1114 swine farms were screened for the presence of the adhesin involved in diffuse adherence (AIDA) gene by polymerase chain reaction (PCR). Escherichia coli isolates that carried AIDA genes were also tested by PCR for the detection of five fimbriae (F4, F5, F6, F18 and F41), heat-stable (STa, STb) and heat-labile (LT) enterotoxin, enteroaggregative E. coli heat-stable enterotoxin 1 (EAST1), and Shiga toxin 2 oedema disease (Stx2e) genes. Twenty-three (2.3%) of the 1002 E. coli isolates carried the gene for AIDA. Among 23 isolates shown to carry genes for AIDA, three carried the AIDA gene as the only shown virulence factor. Other isolates carried other virulence factor genes in addition to AIDA. Four isolates carried genes for at least one of the fimbrial adhesins and enterotoxins. Sixteen isolates carried genes for enterotoxins only. The AIDA may represent an additional virulence determinant in pre-weaned pigs with diarrhoea.