Quinolone-resistant E. coli with various CTX-M beta-lactamase genes that are common in human infections worldwide were found in imported chicken breasts, indicating a possible source for gut colonization. Samples from Brazil were commonly positive for E. coli with CTX-M-2, the dominant bla(CTX-M) genotype from human infections in South America, which is currently rare in clinical infections in the UK. CTX-M-15, the dominant CTX-M type in human infections in the UK, was not found in chicken isolates, suggesting that the UK-reared chickens are not a reservoir of CTX-M-15.
Aims: To employ an in vitro screening regime to select a probiotic Bi®dobacterium strain to complement resistant starch (Hi-maizeä) in a synbiotic yoghurt. Methods and Results: Of 40 Bi®dobacterium isolates examined, only B. lactis Laftiä B94 possessed all of the required characteristics. This isolate hydrolysed Hi-maizeä, survived well in conditions simulating passage through the gastrointestinal tract and possessed technological properties suitable for yoghurt manufacture. It grew well at temperatures up to 45°C, and grew to a high cell yield in an industrial growth medium. In addition to resistant starch, the organism was able to utilize a range of prebiotics including inulin, and fructo-, galacto-, soybean-and xylo-oligosaccharides. Pulse ®eld gel electrophoresis of restriction enzyme cut chromosomal DNA revealed that B. lactis Laftiä B94 was very closely related to the B. lactis Type Strain (DSM 10140), and to the commercial strains B. lactis Bb-12 and B. lactis DS 920. However, B. lactis Laftiä B94 was the only one of these isolates that could hydrolyse Hi-maizeä. This phenotypic difference did not appear to be due to the presence of plasmid encoded amylase. Bi®dobacterium lactis Laftiä B94 survived without substantial loss of viability in synbiotic yoghurt containing Hi-maizeä during storage at 4°C for six weeks. Conclusions: Bi®dobacterium lactis Laftiä B94 is a promising new yoghurt culture that warrants further investigation to assess its probiotic potential. Signi®cance and Impact of the Study: In vitro screening procedures can be used to integrate complementary probiotic and prebiotic ingredients for new synbiotic functional food products.
In this study, two PCR-based methodologies were evaluated for potential use in the determination of DNA diversity between 20 commercial sugarcane hybrids and 6 'outgroup' varieties of S. spontaneum, S. officinarum and hybrids from early in the genealogy . The first method involved PCR amplification of sugarcane DNA in the presence of random, decamer primers (RAPDs), while the second protocol utilized specific microsatellite and telomere sequences as primers . A total of 41 RAPD primers (356 loci) were screened across the varieties of which 15 (160 loci) were used in the calculation of DNA diversity (expressed% similarity) . This varied from 61 to 95%, with most of the commercial varieties showing more than 80% similarity in their DNA . The RAPD data indicated that there had been a gradual decline in DNA diversity (84% reduction) from the early inter-specific crosses to the commercial hybrids, probably as a result of backcrossing and in-breeding strategies used in the previous 5 to 6 generations of sugarcane breeding . The microsatellite and telomere data produced a much greater range in DNA similarity values (25-91%), probably due to the fact that these primers detect highly variable regions of the genome . It is suggested that these specific primers would not be suitable for determination of DNA diversity, but could be used more effectively in the development of a methodology for routine, rapid identification of sugarcane varieties .
An 8.8-kb plasmid (pND302) was identified in Lactococcus lacti spp lactis M71 which encodes cadmium resistance (CdR). Most of the commercial lactococcal strains tested were sensitive to cadmium. Therefore, CdR should provide a useful selectable marker for constructing cloning vectors in lactococci. pND302 was mapped with a number of restriction enzymes and found to contain a unique EcoRI site suitable for cloning. Two E. coli/L lactis shuttle cloning vectors, pND304 and pND624, were constructed by subcloning of the E. coli plasmids pBR322 and pGEM-7Zf(+) containing a 1.6-kb gene encoding nisin resistance (NisR) of lactococcal origin into the EcoRI site of pND302, separately. The E. coli DNA component of pND624 was removed and the resulting plasmid, pND625, consisted of only lactococcal DNA, expressing NisR and CdR, with two synthetic polylinkers that contain multiple restriction sites for versatile cloning. Both pND302 and pND625 can be transformed by electroporation into L. lactis LMO230 at 10(3)/micrograms DNA and maintained stably in LMO230. The results indicated that pND302 and pND625 are potential food-grade cloning vectors for lactococci.
Plasmid pND852 (56 kb) encodes nisin resistance and was isolated from Lactococcus lactis ssp lactis (L. lactis) M138 by conjugation to L. lactis LM0230. It conferred strong resistance to the isometric-headed phage φ712 and partial resistance to the prolate-headed phage φc2. A 2.6 kb HpaII fragment encoding phage resistance was cloned into the streptococcal/Bacillus hybrid vector pGB301 to generate pND817. The mechanism of phage resistance encoded by pND817 involved abortive infection and this was illustrated by a reduction in burst size from 166 to 6 at 30°C and from 160 to 90 at 37°C. Partial resistance was therefore retained at 37°C. DNA sequencing revealed that the abortive infection was encoded by a single open reading frame (ORF), designated abiI, encoding a 332 amino acid protein. Neither abiI nor the predicted product showed significant homology to any existing sequence in the GenBank database. Frame shift mutation at the unique EcoRI site within the ORF resulted in loss of the Abi + phenotype, confirming that the ORF is responsible for the encoded phage resistance.
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