Verocytotoxin-producing Escherichia coli: a veterinary view

This overview places verocytotoxin-producing Escherichia coli (VTEC) in perspective with other E. coli types that cause disease in animals. VTEC O157 and other verocytotoxin-producing serotypes cause severe disease in man but to date, although other VTEC are found in animals, zoonosis appears to be associated with E. coli O157 only. The epidemiology of E. coli O157 in cattle has been studied in Scotland, and this work is described alongside current knowledge.     

Clinical significance of Verocytotoxin-producing Escherichia coli O157

In 1977, Konowalchuk and colleagues (Konowalchuk, J., Speirs, J.I. & Stavric, S. 1977 Infection and Immunity 18, 775–779) were the first to describe Verocytotoxin-producing strains of Escherichia coli or VTEC. The surveillance of infection caused by VTEC demonstrated strains of E. coli belonging to serogroup O157 as the main cause of human infection capable of causing haemorrhagic colitis (HC) and haemolytic uraemic syndrome (HUS). Infection with O157 VTEC results in a range of disease manifestations including abdominal cramps, vomiting and fever. This frequently leads to cases with bloody diarrhoea and HC, and approximately 10% of patients develop HUS. The symptoms of disease caused by VTEC O157 have been well documented and the pathogenic mechanisms expressed by VTEC have been the focus of considerable attention. However, the role of putative pathogenic mechanisms in the pathogenesis of disease is not fully understood. The aim of this review is to consider the clinical aspects of infection with strains of VT-producing E. coli O157 in terms of the putative pathogenic mechanisms expressed by these bacteria. This revised version was published online in November 2006 with corrections to the Cover Date.     

The serodiagnosis of parasitic infections

Recently, the term of clinical immunoparasitology has been coined to indicate the application of immunological methods to the laboratory diagnosis of parasitic infections. In particular, serological diagnosis (indirect diagnosis) is useful especially in the cases of toxocarosis, trichinellosis, echinococcosis, cysticercosis, toxoplasmosis, amoebic abscess, some filariasis, visceral leishmaniasis, schistosomiasis. When possible, for infections caused by protozoa or helminths, the "gold standard" is represented by direct diagnosis performed by microscopic and/or macroscopic observation of the parasite. In any case, immunological results must be interpreted in consideration of the clinical picture of the patient and confirmed possibly by finding the parasite or its genome, even using molecular methods. Furthermore, since the presence of specific antibodies can reveal an acquired infection, but not necessarily a disease, it is particularly helpful, in addition to a qualitative evaluation, a quantitative one, by determining the serum antibody titre. After recovery, the antibody levels decrease, however, they may persist for long periods, for this reason they do not help in evaluating the treatment outcome. Interpretation of serological results may be difficult when the patients originate from areas where the suspected infection is endemic, in that case, a serum positivity could reflect an old exposition to the parasite, therefore it is not related to the present clinical status. Furthermore, serology may frequently result falsely negative in not immunocompetent subjects (organ transplanted, HIV positive individuals, premature babies, diabetics). Clinicians can interpret correctly the serological results only if the Parasitology laboratory inform them about the significant diagnostic values, the sensitivity and the specificity of the test in use. At present time, many diagnostic kits for immunoparasitology are commercially available, and industries are developing newer and newer ones (which are not always validated). In relation to this aspect, it should be helpful, for each of parasitic infection, to establish reference centers, not only to control the quality of commercial kits, but also as a reference point to those laboratories which use "in house" kits. To this regard, the recent establishment of a European Centre for Control of Infectious Diseases will help. The antigen characteristics (crude, E/S, recombinant, synthetic) for assays searching for antibodies (IHA, IFA, EIA, WB) of different classes, the controls to choose for these assays, the specimen requirements will be discussed. The recent findings on the serological diagnosis of intestinal protozoa infections, malaria, leishmaniasis, echinococcosis, cysticercosis, trichinellosis, toxocariasis, schistosomiasis, strongyloidiasis will be presented.     

Verocytotoxin-producing Escherichia coli in chamois (Rupicapra rupicapra) and cattle in Austria

We assessed the prevalence of verotoxigenic Escherichia coli (VTEC) in chamois (Rupicapra rupicapra) and livestock grazing on a mountain pasture in Austria during June-August 2009. We detected VTEC throughout the sampling period in high numbers in cattle as well as in chamois, leading to the assumption that the degree of contamination of the environment is high. This is the first report of pathogenic E. coli identified in chamois, implicating chamois as a new potential reservoir of these zoonotic pathogens. Because the study area also serves recreational purposes, there is a risk of humans acquiring infection via direct or indirect contact.     

Verocytotoxin-producing Escherichia coli in wild birds and rodents in close proximity to farms

Wild animals living close to cattle and pig farms (four each) were examined for verocytotoxin-producing Escherichia coli (VTEC; also known as Shiga toxin-producing E. coli). The prevalence of VTEC among the 260 samples from wild animals was generally low. However, VTEC isolates from a starling (Sturnus vulgaris) and a Norway rat (Rattus norvegicus) were identical to cattle isolates from the corresponding farms with respect to serotype, virulence profile, and pulsed-field gel electrophoresis type. This study shows that wild birds and rodents may become infected from farm animals or vice versa, suggesting a possible role in VTEC transmission.     

Growth of pure cultures of Verocytotoxin-producing Escherichia coli in a range of enrichment media

Aims: This study compared the growth of different strains of Verocytotoxin-producing Escherichia coli (VTEC) in a range of selective enrichment media. Methods and Results: Turbidometric and impedance methods were used to determine the growth of VTEC in pure culture in different enrichment media. Ten strains failed to grow in buffered peptone water + vancomycin, cefsulodin, cefixime at 42°C and some failed to grow, or grew poorly in E. coli (EC) medium supplemented with 20 mg l⁻¹ novobiocin and modified EC supplemented with 20 mg l⁻¹ novobiocin at 37°C and 42°C. Individual VTEC strains were sensitive to the selective agents in some media. Statistical analysis of the conductance detection times of 10 strains showed no overall effect of temperature alone (P = 0·66) but there were significant (P < 0·001) effects as a result of the combination of medium and temperature and these two factors were influenced by strain. Conclusions: Growth of VTEC during enrichment is dependent on different factors alone or in combination. These include medium type, presence of certain selective agents or antibiotics, incubation temperature and the initial population of VTEC. Sensitivity to these conditions can be strain related. Significance and Impact of the Study: This study highlighted differences in the ability of some enrichment media to support the growth of VTEC, making them unsuitable for the isolation of VTEC, especially low numbers of non-O157 strains.     

Therapy of infections caused by mixed cultures ofEscherichia coli andStreptococcus faecalis

The antibacterial efficacy of ampicillin, mezlocillin, piperacillin, and cefotaxime against antagonistic mixed infections ofEscherichia coli andStreptococcus faecalis was studied by adopting an acute as well as a chronic animal model. Data obtained by both models revealed that ampicillin and, especially, mezlocillin were equally effective againstE. coli andS. faecalis whereas piperacillin and cefotaxime selected the enterococci from the mixed infections while eliminatingE. coli from the infected foci.     

Evaluation of serodiagnosis of toxoplasmosis by using the recombinant nucleoside triphosphate hydrolase isoforms expressed in Escherichia coli

The nucleoside triphosphate hydrolase (NTPase) isoforms termed, NTPase-I and NTPase-II of Toxoplasma gondii, were expressed in Escherichia coli as inclusion bodies and purified under denaturing condition. Furthermore, NTPase-I was refolded as an active form and purified under non-denaturing condition. The purified NTPase isoforms, both denatured and refolded, were tested for their usefulness as antigens for the serodiagnosis of acute toxoplasmosis in immunocompetent humans. The test was conducted by using the recombinant NTPase isoforms and comparing the enzyme linked immunosorbent assay (ELISA) absorbances with the Sabin-Feldman dye test titer. Seventy-three sera from dye test-positive patients, and 30 sera from subjects with no T. gondii infection were examined. The total positive rates in dye test positive sera were: 82% (60/73) for denatured NTPase-I; 78% (57/73) for denatured NTPase-II; and 63% (46/73) for refolded NTPase-I. For all three antigen types of recombinant NTPase, the positive rates of sera of acute toxoplasmosis suspected patients were 93% (13/14). A moderate correlation between the ELISA absorbance using these antigens and the dye test titer was observed with the correlation coefficients, 0.583 (r2) for denatured NTPase-I, 0.472 (r2) for denatured NTPase-II, and 0.604 (r2) for refolded NTPase-I in the linear regression analysis. There was no significant difference observed in the antigenicity between refolded and denatured NTPase-I, nor between the isoforms.     

Quantitative microbiological and clinical characteristics of diseases caused by enterotoxigenic Escherichia coli

For the first time the quantitative microbiological characteristics of diarrheas caused by enterotoxigenic E. coli (ETEC), serovars O15: H?, O34: H10, O78: H? O148: H? and O148: H28, in 5 patients are presented. The multiplication of ETEC in the patients' body has provided clinical material permitting the authors to reveal, for the first time, the capacity of ETEC for the colonization of the intestine and to judge of their etiological role in the diseases. A close relationship between the clinical manifestations of the disease and the number of ETEC per g of feces in its dynamics has been established. These data indicate that patients may be regarded as the possible sources of infection.     

Electrophysical analysis of Escherichia coli cell damage caused by silver ions]

The influence of Ag+ (0.5-10 microM) on Escherichia coli K-12 cells was studied by electrophoresis and electro-orientation spectroscopy methods. It was shown that the pH-dependency of the cell electrokinetic potential (phosphate-citrate buffer with ion strength 0.02) practically didn't changed after Ag+ treatment, but in low-conductive media electrophoretical mobility of intact and inactivated by heat (70 degrees, 15 min) cells gradually decreased as the Ag+ concentration increased. It was due to the Ag+ adsorption on the cell surface and could not be used for the definite characterization of the cell damage. The high-frequency decrease in the cell electro-orientation spectrum shifted to the region of lower frequencies, K+ was excreted by cells, slight raise of the medium pH occurred and significant changes of cell osmotic properties were observed as a result of Ag+ action. All these changes showed the disturbance of barrier properties of the cytoplasmic membrane. Besides the damaging action of Ag+ on cell membranes increased with the decrease of pH and decreased after the addition of Mg2+, Ca2+ and Sr2+ in low concentrations.     

Cell death caused by hyper-expression of a secretory exoglucanase in Escherichia coli

Induced expression of a gene fusion between the ompA leader sequence and the Cellulomonas fimi cex gene encoding a secretory exoglucanase, Exg, engineered in the Tac-cassette excretion vector was lethal to Escherichia coli. An exponentially growing culture harboring the recombinant construct suffered slow growth and 99.9% of its cells died within 60-100 min after induction. This abnormality was found to have a close correlation with the rapid increase in the relative amount of the OmpA/Exg fusion precursor (Pre-Exg) compared to its processed product (Mat-Exg). Analysis of subcellular fractions revealed the presence of Pre-Exg in the inner membrane of cultures expressing high levels but not low levels of Pre-Exg. As only Pre-Exg but not Mat-Exg was detectable in the cytoplasm, and Exg was shown by cross-linking experiments to be physically associated with the Sec proteins, it was concluded that secretion and processing of Pre-Exg took place in the SecYEG translocation machinery. The results were in line with the previous speculation that accumulation of unprocessed precursor proteins in the cytoplasmic membrane was detrimental, and supported the idea that cell death was caused by some unusual tie-up of Pre-Exg with the SecYEG translocation machinery, thus imposing an inhibitory effect on the secretion of endogenous secretory proteins. A new model, designated "Saturated Translocation," was proposed to explain the interchangeable lethal and non-lethal properties of Pre-Exg, and to address the possible scenarios that might occur in the course of cell death triggered by secretion of Pre-Exg.