E Van Driessche

Protection of just weaned pigs against infection with F18+Escherichia coli by non-immune plasma powder

The anti-colonization effect of porcine plasma powder against experimentally induced postweaning diarrhoea and oedema disease in just weaned piglets was examined. Piglets were infected with an Escherichia coli strain expressing F18ac fimbriae and producing SLTIIv- and LT-toxins. Reduced fecal excretion of the challenge strain and protection against clinical symptoms was obtained by daily supplementation of the feed with either 90 or 45 g of plasma powder. However, the piglets receiving 90 g of plasma powder a day showed diarrhoea and reduced weight gain compared to the piglets receiving 45 g of plasma powder a day. The diarrhoea was attributed to biogenic amines released from excessive protein in the diet.

Chicken egg yolk antibodies against F18ab fimbriae of Escherichia coli inhibit shedding of F18 positive E. coli by experimentally infected pigs

F18ab and F18ac are antigenic variants of a colonizing fimbria commonly found on E. coli associated with postweaning diarrhea and edema disease in pigs. Chicken F18ab antibodies were obtained by immunising hens with purified F18ab fimbriae. For their in vitro characterisation antibodies were isolated from diluted egg yolks by ammonium sulfate precipitation. In vitro adhesion tests demonstrated that the chicken F18ab antibodies inhibited attachment of F18ab positive E. coli bacteria to the intestinal mucosa. Just weaned piglets were experimentally infected with an F18ab positive edema disease strain of E. coli, or with an F18ac positive postweaning diarrhea E. coli strain. The animals were infected on the second day of a period during which chicken F18ab antibodies were added to their feed. During the same period, pigs of the control group received commercial eggs in which no F18 antibodies were detected. In both experimental infections the excretion of the F18 positive strain was reduced in pigs that received the F18ab antibodies as compared to the control animals. The F18ab antibodies diminished the cases of diarrhea and death in animals infected with F18ac positive E. coli.

N-terminal sequences of the α and β subunits of the lectin from the garden pea (Pisum sativum)

Lectins constitute a group of proteins mainly found in the seeds of a wide variety of plants and in certain invertebrates. These proteins have proven to be useful tools for the study of protein-carbohydrate interactions and the topography of cell surfaces. #en immobilised on Sepharose, lectins may be used for the fractionation of cells and for the isolation of glycoproteins and cell receptors. Certain lectins are able to induce mitogenesis in lymphocytes a phenomenon analogous to the stimulation of immune cells by a specific antigen [l-3] . Although lectins have been extensively studied for their biological properties, few structural data are available to relate their activity to their structure, except for Concanavalin A [4]. In this paper we describe the subunit structure and N-terminal sequences of the oand &subunits of the phytohemagglutinin of the garden pea (Pisum sutivum), a protein with sugar binding properties similar to those of concanavalin A.

The immuno-histochemical localization of lectin in pea seeds (Pisum sativum L.).

The lectin from the garden pea (Pisum sativum L.) has been localized at the ultrastructural level by the unlabeled peroxidase-antiperoxidase procedure of L.A. Sternberger et al. (1970, J. Histochem. Cytochem 18, 315–333) in 24 h imbibed seeds. Upon examination by light microscopy and transmission electron microscopy, the lectin was only found in the protein bodies of cotyledons and embryo axis. Cell walls as well as membraneous fractions were completely devoid of lectin. These results are discussed in relation to the possible physiological function of seed lectins.

Distribution of glucose/mannose-specific isolectins in pea (Pisum sativum L.) seedlings

We report on the distribution and initial characterization of glucose/mannose-specific isolectins of 4- and 7-d-old pea (Pisum sativum L.) seedlings grown with or without nitrate supply. Particular attention was payed to root lectin, which probably functions as a determinant of host-plant specificity during the infection of pea roots by Rhizobium leguminosarum bv. viciae. A pair of seedling cotyledons yielded 545±49 μg of affinity-purified lectin, approx. 25% more lectin than did dry seeds. Shoots and roots of 4-d-old seedlings contained 100-fold less lectin than cotyledons, whereas only traces of lectin could be found in shoots and roots from 7-d-old seedlings. Polypeptides with a subunit structure similar to the precursor of the pea seed lectin could be demonstrated in cotyledons, shoots and roots. Chromatofocusing and isoelectric focusing showed that seed and non-seed isolectin differ in composition. An isolectin with an isoelectric point at pH 7.2 appeared to be a typical pea seed isolectin, whereas an isolectin focusing at pH 6.1 was the major non-seed lectin. The latter isolectin was also found in root cell-wall extracts, detached root hairs and root-surface washings. All non-seed isolectins were cross-reactive with rabbit antiserum raised against the seed isolectin with an isolectric point at pH 6.1. A protein similar to this acidic glucose/mannose-specific seed isolectin possibly represents the major lectin to be encountered by Rhizobium leguminosarum bv. viciae in the pea rhizosphere and at the root surface. Growth of pea seedlings in a nitrate-rich medium neither affected the distribution of isolectins nor their hemagglutination activity; however, the yield of affinity-purified root lectin was significantly reduced whereas shoot lectin yield slightly increased. Agglutination-inhibition tests demonstrated an overall similar sugar-binding specificity for pea seed and non-seed lectin. However root lectin from seedlings grown with or without nitrate supplement, and shoot lectin from nitrate-supplied seedlings showed a slightly different spectrum of sugar binding. The absorption spectra obtained by circular dichroism of seed and root lectin in the presence of a hapten also differed. These data indicate that nutritional conditions may affect the sugar-binding activity of non-seed isolectin, and that despite their similarities, seed and non-seed isolectins have different properties that may reflect tissue-specialization.

THE USE OF NON-IMMUNE PLASMA POWDER IN THE PROPHYLAXIS OF NEONATAL ESCHERICHIA COLI DIARRHOEA IN CALVES

The protective use of plasma powder from cattle and swine against experimentally induced neonatal E. coli diarrhoea in colostrum‐deprived calves was examined. Diarrhoea was induced with a strain expressing F5+ fimbriae and a strain expressing F17+ fimbriae. In all groups supplemented with bovine plasma powder, diarrhoea and fever were less severe than in the control groups. For the groups infected with the F5+ E. coli strain, a reduction in excretion of the challenge strain by 2–4 orders of magnitude and by 1–2 orders of magnitude was seen when supplemented with bovine plasma powder at a dose of 25 g/l milk and 10 g/l milk, respectively. The bovine plasma powder showed also beneficial effects in the F17+ infected groups. No mortality, no septicaemia and no severe clinical signs were observed. Concerning the excretion of the E. coli F17+ strain in the faeces, no significant difference with the control group was found. Swine plasma powder showed little beneficial effect on E. coli diarrhoea in calves in this study.

Tissue treatment for whole mount internal lectin staining in the nematodes Caenorhabditis elegans, Panagrolaimus superbus and Acrobeloides maximus

Four different fixation schemes, using ten fluorescent-labelled lectins, were investigated for whole mount internal staining of three rhabditid nematodes: Caenorhabditis elegans, Panagrolaimus superbus and Acrobeloides maximus. Acetone-only fixation was found to give strong and reproducible staining, which could be prevented either by periodate treatment of the organisms or by specific inhibitory sugars of the lectins under investigation. Whereas the use of either phosphate or TRIS buffers had no effect on the staining pattern or the fluorescence intensity, the incubation time as well as the incubation temperature affected the staining reaction. The best results were obtained upon overnight incubation at 4° C: the lectin staining could be inhibited in all cases, except for the intestinal brush border of C. elegans by the lectin of Lens culinaris.

The subunit structure and N-terminal sequences of the α- and gb-subunits of the lentil lectin (Lens culinaris)

Lectins are proteins or glycoproteins present in many plants and are usually recognized by their ability to agglutinate erythrocytes. Agglutination of cells is in many cases inhibited by specific sugars, suggesting that the binding is to sugar residues on the cell surface. Very little is known of the mechanism by which the lectin leads to agglutination or cellsurface alteration. In order to understand the relationship between the activity and the structure of lectins, we have undertaken the determination of the amino acid sequence of several of these proteins. The lectin studied in this work is present in seeds of the lentil, Lens culinaris [ 1,2] . The protein has a molecular weight of approximately 49 000 and is composed of two types of subunits with molecular weights of 18 000 and 8000 [3] . The isolation and characterization of this hemagglutinin was described in our previous communication [4]. We report here the N-terminal sequences of the (Yand P-subunits of the lentil lectin. This protein has the same sugar-binding specificity as two other mitogenic lectins, Concanavalin A [5] and pea lectin [6], for which sequence studies were reported previously [7,8]. The comparison of these three proteins reveals surprising homologies.

Binding of biotinylated legume seed lectins with glycoproteins in blotted receptor-analogs: influence of incubation pH

Abstract SDS–PAGE and Western blotting of lectin inhibitors (bovine and porcine plasma powder, whole egg powder and fetuin) was performed and blots were incubated with several biotinylated lectins commercially available ( Phaseolus vulgaris E , Pisum sativum , Lens culinaris , Vicia faba ). In order to study the effect of pH on the binding of the lectins to glycoproteins present in the inhibitors, identical blots were incubated in buffers with different pH values, i.e. 3–7, respectively. Binding capacity of lectins to the glycoproteins in the inhibitors was very dependent on pH condition during incubation: for all the lectins involved in this experiment, pH values lower than 4 inhibited the binding process considerably. Results are discussed taking into account the low pH values prevailing in the stomach of pigs.

Chapter 20 Strategies for the prevention of E. coli infection in the young animal1

Publisher Summary This chapter discusses the strategies to prevent colonization of the gut by enterotoxigenic E. coli (ETEC). The chapter describes the attachment of E. coli , by virtue of fimbriae, to the intestinal wall, and the molecules involved both at the bacterial surface and on the intestinal epithelium. ETEC are pathogens that cause diarrhoea in humans and livestock. These bacteria colonize the small intestinal epithelium by of expressing adhesins (lectins) in the form of long proteinaceous appendages that protrude from their surface, and that are called fimbriae. Because of their size, architecture and extracellular expression, fimbriae can easily be obtained in a highly purified state and in appreciable quantities to be used as vaccine components. Purified fimbriae have proven to be strong immunogens, and humoral antibodies are readily generated in rodents, chickens and farm animals. Plant lectins are potential candidates to block intestinal mucosal E. coli lectin receptors, because most plant lectins are rather resistant to proteolytic degradation in the stomach and small intestine, and, can bind to glycoconjugates present in the intestinal mucosa. The beneficial effect of probiotics in protecting the host from intestinal disorders and/or colonization of the intestine is because of one or a combination of the following effects: (1) stimulation of the immune system; (2) competitive inhibition for bacterial adhesion sites on the intestinal surface; (3) degradation of intestinal toxin receptors; and (4) production of inhibitory substances such as bacteriocins, hydrogen peroxide or organic acids.