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. We were not able to recover transconjugants from faecal samples with lactose supplementation, indicating a possible inhibition of plasmid transfer. Since the yoghurt, heat-treated fermented milk, milk and lactose solution contained approximately the same lactose concentration it is fair to speculate that lactose may contribute to the inhibiting effects of the various supplementations. The inhibitions described were not associated with other intestinal parameters like the intestinal transit time, the population levels of the recipient, or the donor and total anaerobic microflora. It is evident that other parameters need to be investigated such as the composition of the endogenous microflora and metabolic products.

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. Results indicated that yoghurt consumption had two effects: it reduced the efficacy of plasmid transfer 10-fold and decreased the transconjugant PG1-R388 population density 100-fold, compared to the control group. We also describe another HFF mouse model in which recipient PG1 was replaced by EM0 with which no plasmid transfer was observed. This model made it possible to demonstrate the potential promoting effect of yoghurt intake on transconjugant formation and establishment. Our results indicated no yoghurt effect; no transconjugants appeared in the digestive tract of HFF-EM0 mice fed on yoghurt or on standard food. In both mouse models, HFF-PG1 and HFF-EM0, yoghurt intake did not promote the mobilisation of either the non-self-transmissible plasmid pUB2380 or the recombinant plasmid pCE325.

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. coli K12 donor strain harboring three plasmids (R388, self-transmissible, pCE325 and pUB2380, mobilisable,) and an E. coli recipient strain, PG1, of human origin (Duval-Iflah et al., 1994). Milks fermented with either Lactobacillus bulgaricus or Streptococcus thermophilus or symbiosis, S85, of both strains were given daily as 1/3 of food diet. Fermented milks have no effect on the transfer of R388 and pUB2380 except a slight increase of TC(R388) with milk fermented with S85. Long term ingestion of milk fermented with S85 inhibited the formation and the establishment of transconjugants TC(pCE325). Milk fermented with L. bulgaricus lowered the population density of TC(pCE325) in animals where they were already established. This phenomenon was reversible, since the density of TC(pCE325) increased in the same animals after cessation of supplementation. Bacterial cultures obtained in MRS broth and given in state of drinking water were compared with fermented milks. Bacterial cultures with L. bulgaricus and with S85 favoured the establishment of TC(pCE325). There results indicate for the first time that probiotics have various effects on the formation and/or establishment of transconjugants in the gut of axenic mice. The effects depend on whether the probiotics were cultivated in milk or in MRS, indicating that bacterial metabolites and viable bacteria can be involved.

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., Peptostreptococcus sp. (strictly anaerobic bacterial strains) and Enterococcus faecalis, (2) a whole human faecal flora, was studied. In both cases, no gene transfer was observed to strictly anaerobic bacterial strains. DNA transfer was only observed to the E. faecalis strain: in vivo CmR E. faecalis transconjugants were isolated from sequentially multi-associated mice and when the recipient strains associated with the mice, they were a defined mixture of Bacteroides sp., Bifidobacterium sp., Peptostreptococcus sp. and E. faecalis strains. When mice were associated with the whole human faecal flora, the plasmid pIL205 was transferred into some facultative anaerobic streptococci. It was also shown that DNA transfer occurred even when the lactococcal donor strain was transient in the DT of the gnotobiotic host animals.

[Implantation of a strain of "Escherichia coli" in the digestive tract of human new-borns: barrier effect against antibioresistant "E. coli" (author's transl)]. / Implantation précoce d'une souche de Escherichia coli dans l'intestin de nouveau-nés humains: effet de barrière vis-à-vis de souches de E. coli antibiorésistantes.

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%). Strains of E. coli resistant to ampicillin or tetracycline were found in 6 of 22 inoculated infants as well as in 7 of the 24 control infants. These resistant strains remained at a very high level in the control infants but disappeared, or decreased to a subdominant level, in the inoculated infants. These results show that a totally innocuous strain of E. coli can exert in holoxenic conditions a barrier effect on antibiotic resistant E. coli strains. They also suggest the interest of inoculating such a strain at birth in order to prevent proliferation of potentially pathogenic bacteria.