Worst-case scenarios for horizontal gene transfer from Lactococcus lactis carrying heterologous genes to Enterococcus faecalis in the digestive tract of gnotobiotic mice.

Since genetically modified (GM) lactic acid bacteria (LAB) might be released in open environments for future nutritional and medical applications, the purpose of this study was to determine an upper limit for the horizontal gene transfer (HGT) in the digestive tract (DT) from Lactococcus lactis carrying heterologous genes (lux genes encoding a bacterial luciferase) to Enterococcus faecalis. Two enterococcal wide host-range conjugative model systems were used: (i) a system composed of a mobilizable plasmid containing the heterologous lux genes and a native conjugative helper plasmid; and (ii) a Tn916-lux transposon. Both systems were tested under the most transfer-prone conditions, i.

Interrelationships between dairy product intake, microflora metabolism, faecal properties and plasmid dissemination in gnotobiotic mice.

We previously described the effects of intake of dairy products on plasmid dissemination in the digestive tract of gnotobiotic mice associated with human faecal flora (HFF) and found that yoghurt, heat-treated yoghurt (HTY) and milk reduced population levels of transconjugants compared with findings in mice fed a standard mouse diet. In the case of lactose intake, transconjugants were not detected. The aim of the present study was to assess the possible interrelationships between these observations and other variables (bacterial ecology, pH, moisture, enzyme activities, short-chain fatty acid (SCFA) contents, lactic acid contents).

Comparison of yoghurt, heat treated yoghurt, milk and lactose effects on plasmid dissemination in gnotobiotic mice.

The effect of yoghurt, heat-treated fermented milk, milk and lactose solution intake on plasmid transfer and establishment of the resulting transconjugants in the digestive tract of mice colonised with human faecal flora were examined. Yoghurt lowered the population level of transconjugants more efficiently than heat-treated fermented milk (-2 log and -1 log respectively) and indicated a beneficial effect of viable bacteria. On the other hand consumption of milk drastically inhibited the establishment of transconjugants, which were below the detection threshold of 10(2) UFC per g of faeces.

Influence of oral inoculation with plasmid-free human Escherichia coli on the frequency of diarrhea during the first year of life in human newborns.

Background: This study was carried out to determine whether early inoculation of the plasmid-free human Escherichia coli into human newborns would reduce the frequency of acute diarrhea during a 1-year period. The plasmid-free E. coli strain isolated from the fecal microbiota of a healthy adult was nontoxigenic in vivo and in vitro and sensitive to all usual antibiotics.

Experimental models of porcine post-weaning colibacillosis and their relationship to post-weaning diarrhoea and digestive disorders as encountered in the field.

The aim of this study was to develop a reliable model system of porcine post-weaning colibacillosis, and in doing so to assess the primary relationship of enterotoxigenic Escherichia coli to post-weaning diarrhoea and digestive disorders as encountered in the field. Six sequential experiments were carried out using 168 SPF piglets weaned into an optimal controlled environment at 28 days of age. The piglets were allocated to 23 treatment groups, 17 of which were inoculated either orally or intragastrically with enterotoxigenic strains of E.

Effects of yoghurt intake on plasmid transfer and colonisation with transconjugants in the digestive tract of mice associated with human faecal flora.

This study deals with the effects of yoghurt intake on wild-type and recombinant plasmid transfer from an exogenous Escherichia coli K12-derivative donor strain to an endogenous recipient strain in the digestive tract of mice associated with human faecal flora. We showed that the self-transmissible plasmid R388 was efficiently transferred to recipient strain PG1 in mice associated with human faecal flora (HFF-PG1) and that the resulting transconjugants (PG1-R388) became established at a high and maximal population level without any selective pressure. Using HFF-PG1 mice made it possible to determine whether yoghurt consumption decreases R388 transfer efficiency and the ability of transconjugant PG1-R388 to successfully colonise the digestive tract.

Effects of racecadotril and loperamide on bacterial proliferation and on the central nervous system of the newborn gnotobiotic piglet.

Methods: The effects of 4 days of oral administration of different doses of two drugs, an enkephalinase inhibitor (the antisecretory agent, racecadotril) and a mu-receptor agonist (loperamide), on intestinal growth of a bacterial nonpathogenic strain (Escherichia coli E 404) and on the central nervous system (CNS) were compared in newborn gnotobiotic piglets.

Results: The E. coli content of the proximal jejunum (segment S1) and the E.

Effect of fermented milk intake on plasmid transfer and on the persistence of transconjugants in the digestive tract of gnotobiotic mice.

Plasmid transfer occurs in the digestive tract and the transconjugants may become durably established. The aim of the present work is to investigate the effect of probiotics of plasmid transfer and on establishment of transconjugants in the gut. Plasmid transfers were carried out in the digestive tract of germ free mice associated with an E.

Recombinant plasmid mobilization between E. coli strains in seven sterile microcosms.

Transfer by mobilization of a pBR derivative recombinant plasmid lacking transfer functions (oriT+, tra-, mob-) from one E. coli K12 strain to another was investigated in seven sterile microcosms corresponding to different environments. These microcosms were chosen as representative of environments that genetically engineered microorganisms (GEMOs) encounter after accidental release, namely attached biomass in aquatic environments (biofilm), soil, seawater, freshwater, wastewater, mouse gut, and mussel gut, GEMOs survived in the same way as the host strains in all microcosms.

The porcine intestinal receptor for Escherichia coli K88ab, K88ac: regional localization on chromosome 13 and influence of IgG response to the K88 antigen.

The loci encoding the porcine intestinal receptors for Escherichia coli K88ab and K88ac (K88abR and K88acR) were firmly assigned to chromosome 13 by linkage analysis using a three-generation pedigree. The linear order of these loci and seven other markers on chromosome 13 was determined by multipoint analyses. The K88abR and K88acR loci were tightly linked (theta = 0.

Use of porcine fibrinogen as a model glycoprotein to study the binding specificity of the three variants of K88 lectin.

Known glycoproteins were used to determine the differences occurring in the binding specificities of the three variants of the K88 lectin in an approach essentially based on lectin blotting. During the screening, it was demonstrated that each variant of the K88 lectin biotinylated via its amino groups (NbioK88) exhibited a characteristic binding to the three chains of porcine fibrinogen. NbioK88ab weakly bound to A alpha chains, NbioK88ac bound to B beta and gamma chains, and NbioK88ad bound only to the gamma chain.

Characterization of O-glycan moieties of the 210 and 240 kDa pig intestinal receptors for Escherichia coli K88ac fimbriae.

The porcine brush border glycoproteins of 210 and 240 kDa, recognized by Escherichia coli K88ac fimbriae, contained O-linked oligosaccharides. The carbohydrate moieties were analysed by deglycosylation, lectin-binding and agglutination assays. Neuraminidase susceptibility of the 210 kDa receptor suggested that a sialoglycoprotein may act as receptor for the K88ac fimbriae.

Study of gene transfer in vitro and in the digestive tract of gnotobiotic mice from Lactococcus lactis strains to various strains belonging to human intestinal flora.

The use of genetically modified organisms (GMO) in dairy products requires evaluation of the DNA transfer capacity from such organisms among the human intestinal microflora. Thus, both in vitro and in vivo [in the digestive tract (DT) of mice] transfer from Lactococcus lactis donor strains of the conjugative plasmid pIL205 (CmR) and the non-conjugative plasmid pIL253 (EmR) to: (1) recipient strains isolated from human faecal flora Bacteroides sp., Bifidobacterium sp.

Gene transfer from engineered Lactococcus lactis strains to Enterococcus faecalis in the digestive tract of gnotobiotic mice.

The introduction of genetically modified organisms into food products requires an evaluation of the behaviour and the dissemination of foreign genes of such organisms among the human intestinal microflora. The conjugal transfer, both in vitro and in vivo (in mice digestive tract) of DNA from Lactococcus lactis donor strains to an Enterococcus faecalis strain isolated from human faecal flora was studied. We followed the transfer of (1) the self-transmissible plasmid pIL205; (2) two non-self-transmissible but mobilizable plasmids, pIL252 and pIL253; (3) one plasmid, pMS1.

Evidence for linkage between K88ab, K88ac intestinal receptors to Escherichia coli and transferrin loci in pigs.

Segregation at the loci coding for the K88ab and K88ac small intestinal receptors to E. coli adhesins (K88abR, K88acR) and at the transferrin (TF) locus was studied in 38 pig families including 273 piglets. The TF locus showed a segregation deviation towards the B variant while each of the K88 receptors behaved as a single autosomal dominant gene.

Colonization of the digestive tract of germ-free mice by genetically engineered strains of Lactococcus lactis: study of recombinant DNA stability.

The ability of genetically engineered Lactococcus lactis strains to become established in the digestive tract (DT) of germ-free mice was examined together with the stability of their genetic markers. Seven L. lactis strains were genetically modified by insertion of genetic markers on different replicons: chloramphenicol resistance gene cat was carried by self-transmissible plasmid pIL205, a derivative of plasmid pIP501; erythromycin resistance gene erm, originating from pAM beta 1, was inserted into non-transmissible plasmids pIL252 and pIL253 of low and high copy number respectively; erm gene from plasmid pMS1.

Adhesion of K99 fimbriated Escherichia coli to pig intestinal epithelium: correlation of adhesive and non-adhesive phenotypes with the sialoglycolipid content.

Evidence for the existence of two phenotypes of piglets born to experimental herds was obtained based on the susceptibility of intestinal brush borders to adhesion of K99-positive Escherichia coli. The enterocytes of the K99-receptive piglets displayed a characteristic sialoglycolipid pattern, with a higher content of the monosialoglycolipids II3NeuGc-LacCer (GM3Gc), IV3NeuGc-nLcOse4Cer (SPGGc) and IV3NeuAc-nLcOse4Cer (SPG) and the oligosialogangliosides IV3NeuAc,II3NeuAc-GgOse4Cer (GD1a), II3(NeuAc)2-GgOse3Cer (GD2), II3(NeuAc)2-GgOse4Cer (GD1b) and IV3NeuAc,II3(NeuAc)2-GgOse4Cer (GT1b) when compared to the gangliosides of non-receptive piglets. The gangliosides from enterocytes of the non-receptive piglets were mainly the monosialogangliosides II3NeuAc-GgOse3Cer (GM2) and II3NeuAc-LacCer (GM3), only traces of the other sialoglycolipids being detected.

Double-sandwich enzyme-linked immunosorbent assay for determination of Escherichia coli heat-labile porcine enterotoxins.

A "double-sandwich" ELISA for the detection and measurement of a heat-labile enterotoxin produced by porcine enterotoxigenic strains of Escherichia coli (LTp) is described. In contrast with other heat-labile toxins, LTp did not bind to agarose gels and exhibited a very low affinity for GM1 in the classical GM1-ELISA technique. The similarity of LTp with cholera toxin was confirmed by immunoblotting.

Proteins in the digesta of the pig: amino acid composition of endogenous, bacterial and fecal fractions.

When studying digestibility, the respective parts of the exogenous, endogenous and bacterial fractions in the digesta or feces must be measured. The proportions of proteins from different sources may be estimated by comparing their amino acid composition with those of reference sources. This study describes the composition of endogenous and microbial proteins, i.

Resistance of gnotobiotic large white and Chinese piglets to in vivo attachment of a K88ab enterotoxigenic Escherichia coli strain.

In vivo adhesion of a K88ab-positive enterotoxigenic Escherichia coli (ETEC) strain to the small intestinal wall of gnotobiotic, colostrum-deprived Chinese and Large White piglets was investigated. A non-enterotoxigenic, attachment factor-deprived E. coli strain, inoculated in association with the K88ab ETEC strain, was used as a marker to determine the content residues on the intestinal walls of gnotobiotic piglets.

[Effect of orally administered bovine lactoferrin and bovine IgG on the establishment of Escherichia coli in the digestive tract of gnotobiotic mice and human newborn infants].

The bacteriostatic effect of the association bovine lactoferrin (LF) and bovine IgG (IgG) was studied in vitro and in vivo against two Escherichia coli strains, S17 and EMO1, isolated from the faecal flora of mouse and man, respectively. These two strains were sensitive in vitro to the bacteriostatic effect of LF + IgG. A kinetic study of the in vivo establishment of E.

Intraspecific interactions between Escherichia coli strains in human newborns and in gnotobiotic mice and piglets.

A plasmid-free human Escherichia coli strain EMO, was inoculated to human newborns within two hours of life. It became established in the feces at a high population level. Ampicillin and tetracycline resistant E.

[Implantation of a strain of "Escherichia coli" in the digestive tract of human new-borns: barrier effect against antibioresistant "E. coli" (author's transl)].

Twenty-two healthy human new-borns were inoculated within two hours of birth with a strain of Escherichia coli. This strain was isolated from the faecal flora of an healthy adult and was plasmid-free, nontoxigenic in vitro or in vivo, and sensitive to all usual antibiotics. This strain became established at a high population level within two days in all new-borns and remained at a very high level during the following days in almost all cases (86%).

Antagonisms among isogenic strains of Escherichia coli in the digestive tracts of gnotobiotic mice.

We have observed that antagonisms occur between isogenic strains of Escherichia coli associated with gnotobiotic mice. The strains differed in the carriage of plasmids or in chromosomal mutations. The plasmid-free strains, in general, inhibited the establishment of plasmid-bearing strains in the gastrointestinal tract of mice.