Immunomagnetic separation and DNA hybridization for detection of enterotoxigenic Escherichia coli in a piglet model.

1991 ◽  
Vol 29 (10) ◽  
pp. 2259-2262 ◽  
Author(s):  
A Lund ◽  
Y Wasteson ◽  
O Olsvik
2010 ◽  
Vol 5 (04) ◽  
pp. 248-254 ◽  
Author(s):  
Nader Shahrokhi ◽  
Saeid Bouzari ◽  
Anis Jafari

Introduction: Enterotoxigenic Escherichia coli (ETEC) causes diarrhoea by producing heat-labile (LT) or heat-stable (ST) enterotoxins after colonizing the small intestine by means of colonization factors (CFs). Although detection of the toxins is sufficient for verification of ETEC isolates, toxin-positive strains may be further analyzed for the presence of CFs. Antibiotics may shorten the duration of diarrhoea caused by ETEC, but the rapid emergence of resistant strains limits their usefulness. Methodology: ETEC isolates collected 10 years apart were compared for the prevalence of toxin types, CFs and antibiotic resistance. DNA/DNA hybridization with digoxigenin (DIG)-labeled probes was used for the detection of toxin types, and CF-typing was performed by DNA hybridization using DIG-labeled probes for cfaD and CS6 with slide agglutination. Disk diffusion was used to determine antibiotic resistance. The presence of class 1 integrons was detected by PCR. Results: ST-positive isolates were the most prevalent among the isolates from 1988, but a significant shift towards LT-gene carriage was observed in the 1998 group. CFA/I and CFA/IV were the most common CF types within both groups. The most prevalent resistance patterns among these isolates were ACSTSXT followed by ASTSXT and ASSXT. Conclusion: Our study of the two groups of isolates showed that the rate of LT and ST gene carriage, as well as antibiotic resistance markers, has changed in the ten years separating the two bacterial populations. These variations show the importance of monitoring pathogenic bacteria to obtain a near realistic picture of the circulating bacterial pathogens.


1985 ◽  
Vol 151 (1) ◽  
pp. 124-130 ◽  
Author(s):  
P. Echeverria ◽  
J. Seriwatana ◽  
D. N. Taylor ◽  
C. Tirapat ◽  
W. Chaicumpa ◽  
...  

2004 ◽  
Vol 70 (3) ◽  
pp. 1708-1716 ◽  
Author(s):  
M. C. Pearce ◽  
C. Jenkins ◽  
L. Vali ◽  
A. W. Smith ◽  
H. I. Knight ◽  
...  

ABSTRACT This study investigated the shedding of Escherichia coli O26, O103, O111, O145, and O157 in a cohort of beef calves from birth over a 5-month period and assessed the relationship between shedding in calves and shedding in their dams, the relationship between shedding and scouring in calves, and the effect of housing on shedding in calves. Fecal samples were tested by immunomagnetic separation and by PCR and DNA hybridization assays. E. coli O26 was shed by 94% of calves. Over 90% of E. coli O26 isolates carried the vtx 1, eae, and ehl genes, 6.5% carried vtx 1 and vtx 2, and one isolate carried vtx 2 only. Serogroup O26 isolates comprised seven pulsed-field gel electrophoresis (PFGE) patterns but were dominated by one pattern which represented 85.7% of isolates. E. coli O103 was shed by 51% of calves. Forty-eight percent of E. coli O103 isolates carried eae and ehl, 2% carried vtx 2, and none carried vtx 1. Serogroup O103 isolates comprised 10 PFGE patterns and were dominated by two patterns representing 62.5% of isolates. Shedding of E. coli O145 and O157 was rare. All serogroup O145 isolates carried eae, but none carried vtx 1 or vtx 2. All but one serogroup O157 isolate carried vtx 2, eae, and ehl. E. coli O111 was not detected. In most calves, the temporal pattern of E. coli O26 and O103 shedding was random. E. coli O26 was detected in three times as many samples as E. coli O103, and the rate at which calves began shedding E. coli O26 for the first time was five times greater than that for E. coli O103. For E. coli O26, O103, and O157, there was no association between shedding by calves and shedding by dams within 1 week of birth. For E. coli O26 and O103, there was no association between shedding and scouring, and there was no significant change in shedding following housing.


1987 ◽  
Vol 50 (6) ◽  
pp. 487-489 ◽  
Author(s):  
VALERIE MITCHELL DAVIS

Previously a DNA hybridization assay was designed to detect the presence of and to enumerate enterotoxigenic foodborne Escherichia coli. The determinative step in the method involves autoradiographic analysis of the DNA from foodborne isolates after hybridization with a 32P-labeled probe specific for an enterotoxin gene. Dark spots appearing on the X-ray film after exposure indicate which colonies carry genes encoding the pathogenic determinant. A problem with this assay is the tendency of some colonies to detach from the nitrocellulose filters during hybridization or washing to remove the unbound probe DNA; this results in a false-negative interpretation in up to 60% of the samples processed at 80°C. By lowering the temperature to 70°C and increasing the incubation time to 3 h during in vacuo baking of filters, detachment (flotation) of colonies is reduced to about 37%. At 65°C only 2% of the colonies came off the filter after in vacuo baking of filters for 24 h. Another problem has been the inadequacy of exposure of X-ray film at −20°C when a −70°C freezer is not available. This problem can be alleviated by exposing the X-ray film in cassette holders “sandwiched” between slabs of dry ice (CO2 ice has a temperature of −78.5°C). These modifications improve the reliability and accuracy of this DNA colony hybridization method.


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