Short courses of dual-strain probiotics appear to be effective in reducing necrotising enterocolitis.

AIM: Prophylactic probiotics to reduce necrotising enterocolitis (NEC) are mostly given for at least 28 days or until discharge. We describe the effects of a shorter duration dosing strategy. METHODS: Retrospective cohort study of neonates (birthweight 400-1500 g) in three neonatal intensive care units in Switzerland and Germany that embarked on probiotic prophylaxis given for 10 or 14 days, employing a fixed combination (Lactobacillus acidophilus plus Bifidobacterium infantis, each 10(9) CFU/day) licensed as a drug in Switzerland. Probiotics were initiated upon discontinuation of antibiotics, or on day 1-3 in infants without antibiotics. Repeat probiotic courses were given whenever antibiotics had been instituted and were discontinued. RESULTS: Birthweight and gestational age were similar in the two 24-month pre- and postimplementation cohorts. NEC rates fell from 33 of 633 (5.2%) to 8 of 591 infants alive at three days (1.4%; risk ratio (RR) 0.26, 95% confidence interval (CI) 0.12-0.55). The drop in NEC was significant both for infants of 400-999 g (6.4% to 2.5%) and 1000-1500 g birthweight (4.4% to 0.6%). Mortality was 5.1% (32/633) without, as opposed to 3.5% (21/591) with probiotics, respectively (RR 0.69, 95% CI 0.41-1.19). CONCLUSION: Short courses of a dual-strain probiotics appear to be effective in reducing NEC.

Agrobacterium-Mediated Transformation of Fusarium oxysporum: An Efficient Tool for Insertional Mutagenesis and Gene Transfer.

ABSTRACT Agrobacterium tumefaciens-mediated transformation (ATMT) has long been used to transfer genes to a wide variety of plants and has also served as an efficient tool for insertional mutagenesis. In this paper, we report the construction of four novel binary vectors for fungal transformation and the optimization of an ATMT protocol for insertional mutagenesis, which permits an efficient genetic manipulation of Fusarium oxysporum and other phytopathogenic fungi to be achieved. Employing the binary vectors, carrying the bacterial hygromycin B phosphotrans-ferase gene (hph) under the control of the Aspergillus nidulans trpC promoter as a selectable marker, led to the production of 300 to 500 hygromycin B resistant transformants per 1 x 10(6) conidia of F. oxysporum, which is at least an order of magnitude higher than that previously accomplished. Transformation efficiency correlated strongly with the duration of cocultivation of fungal spores with Agrobacterium tumefaciens cells and significantly with the number of Agrobacteruium tumefaciens cells present during the cocultivation period (r = 0.996; n = 3; P < 0.01). All transformants tested remained mitotically stable, maintaining their hygromycin B resistance. Growing Agrobacterium tumefaciens cells in the presence of acetosyringone (AS) prior to cocultivation shortened the time required for the formation of transformants but decreased to 53% the percentage of transformants containing a single T-DNA insert per genome. This increased to over 80% when Agrobacterium tumefaciens cells grown in the absence of AS were used. There was no correlation between the average copy number of T-DNA per genome and the colony diameter of the transformants, the period of cocultivation or the quantity of Agrobacterium tumefaciens cells present during cocultivation. To isolate the host sequences flanking the inserted T-DNA, we employed a modified thermal asymmetric interlaced PCR (TAIL-PCR) technique. Utilizing just one arbitrary primer resulted in the successful amplification of desired products in 90% of those transformants analyzed. The insertion event appeared to be a random process with truncation of the inserted T-DNA, ranging from 1 to 14 bp in size, occurring on both the right and left border sequences. Considering the size and design of the vectors described here, coupled with the efficiency and flexibility of this ATMT protocol, it is suggested that ATMT should be regarded as a highly efficient alternative to other DNA transfer procedures in characterizing those genes important for the pathogenicity of F. oxysporum and potentially those of other fungal pathogens.