High-performance liquid chromatographic determination of 1-β- -arabinofuranosyl-E-5-(2-bromovinyl)uracil and its metabolite (E)-5-(2-bromovinyl)uracil in serum

A high-performance liquid chromatographic method was developed to assay 1-β- -arabinofuranosyl-E-5-(2-bromovinyl)uracil and its metabolite (E)-5-(2-bromovinyl)uracil in serum. The chloro analogue of the parent drug is used as internal standard. Human serum samples were assayed to establish the pharmacokinetic parameters. Acetonitrile, used as a protein precipitant, was evaporated to dryness and the residue, containing the analytes and internal reference, was dissolved in mobile phase prior to chromatographic analysis. The minimum quantifiable level was 0.02 μg of each analyte per ml of serum.     

Type V collagen as the target for type-3 fimbriae, enterobacterial adherence organelles

Tissue-binding specificity of the type-3 fimbriae of pathogenic enteric bacteria was determined using frozen sections of human kidney. A wild-type Klebsiella sp. strain and the recombinant strain Escherichia coli HB101(pFK12), both expressing type-3 fimbriae, as well as the purified type-3 fimbriae effectively bound to sites at or adjacent to tubular basement membranes, Bowman's capsule, arterial walls, and the interstitial connective tissue. Bacterial adherence to kidney was decreased after collagenase treatment of the tissue sections. Recombinant strains expressing type-3 fimbriae specifically adhered to type V collagen immobilized on glass slides, whereas other collagens, fibronectin or laminin did not support bacterial adherence. In accordance with these findings, specific binding of purified type-3 fimbriae to immobilized type V collagen was demonstrated. Specific adhesion to type V collagen was also seen with the recombinant strain HB101(pFK52/pDC17), which expresses the mrkD gene of the type-3 fimbrial gene cluster in association with the pap-encoded fimbrial filament of E. coli, showing that the observed binding was mediated by the minor lectin (MrkD) protein of the type-3 fimbrial filament. The interaction is highly dependent on the conformation of type V collagen molecules since type V collagen in solution did not react with the fimbriae. Specific binding to type V collagen was also exhibited by type-3 fimbriate strains of Yersinia and Salmonella, showing that the ability to use type V collagen as tissue target is widespread among enteric bacteria.     

The priB gene encoding the primosomal replication n protein of Escherichia coli.

The gene encoding protein n of the Escherichia coli primosome has been discovered in the rpsF-rpsR-rplI ribosomal protein operon and designated priB. The low copy number of PriB protein and the distinctive codon usage of its gene argue against its being a ribosomal protein. A strain which overproduces PriB was constructed and has been used to purify the protein to homogeneity. The overproduced protein behaves like that purified from wild-type cells.     

Induction of vancomycin resistance in Enterococcus faecium by non-glycopeptide antibiotics

Abstract Bacitracin and other antibiotics that inhibit late stages in peptidoglycan biosynthesis induce vancomycin resistance in a high-level, inducibly vancomycin-resistant strain of Enterococcus faecium . Exposure to bacitracin led to synthesis of the lactate-containing UDP-MurNAc-pentadepsipeptide precursor required for vancomycin resistance. These findings indicate that inhibition of peptidoglycan biosynthesis can lead to induction of vancomycin resistance and raise the possibility that multiple signals may serve to induce resistance.     

Binding to Ia protects an immunogenic peptide from proteolytic degradation

A 34 amino acid hen egg-white lysozyme (HEL) peptide was designed and synthesized to investigate if an immunogenic peptide once bound to an Ia molecule becomes proteolytically inaccessible. The determinant recognized by T cells, HEL(52-61) was composed of L-amino acids whereas the 12 amino acid extension on each side of this core were composed of D-epimers. This peptide, HEL(40-73) was resistant to proteolysis, except in the core region, where any cleavage would destroy the determinant. Initially HEL(40-73) was shown to be able to stimulate the HEL specific T cell, 3A9, indicating that an I-Ak molecule can bind and present large peptides that extend beyond the theoretical binding groove. HEL(40-73) was then used to examine the proteolytic sensitivity of determinants recognized by T cells. If HEL(40-73) was treated with chymotrypsin before binding to I-Ak, the determinant was totally destroyed; however, if HEL(40-73) was allowed to first bind to I-Ak, then the determinant became resistant to chymotrypsin cleavage. Thus an Ia molecule can protect a determinant from proteolytic degradation, a finding that has important implications for proposed pathways of Ag processing.     

Efficacy of beta-glucuronidase assay for identification of Escherichia coli by the defined-substrate technology

In 1976, Kilian and Bulow described the association of beta-glucuronidase with the genus Escherichia (97% positive) and suggested that a beta-glucuronidase assay would be a useful identification test. Since that report, papers about the sensitivity and specificity of this enzyme for the identification of Escherichia coli from clinical sources, food, seawater, potable-water supplies, and various environmental sources have appeared. A study was undertaken to determine the efficacy and specificity of the defined-substrate technology beta-glucuronidase (Colilert) assay for the identification of this species from fecal samples. A total of 460 human, 105 cow, and 55 horse E. coli isolates were tested. Results showed 95.5% beta-glucuronidase-positive isolates in 24 h and 99.5% positive after 28 h of incubation. Only one E. coli isolate was negative. There were no significant differences in the percentage of beta-glucuronidase-positive isolates among the human or animal isolates. There were no non-E. coli isolates that were positive. All subjects carried beta-glucuronidase-positive E. coli.     

When Landscaping Goes Bad: The Incipient Invasion of Mahonia bealei in the Southeastern United States

Woodlots are forest islands embedded within an urban matrix, and often represent the only natural areas remaining in suburban areas. Woodlots represent critical conservation areas for native plants, and are important habitat for wildlife in urban areas. Invasion by non-indigenous (NIS) plants can alter ecological structure and function, and may be especially severe in remnant forests where NIS propagule pressure is high. Woody shrubs in the Family Berberidaceae have been well documented as invaders of the forest–urban matrix in North America. Mahonia bealei (Berberidaceae) is a clonal shrub native to China, and is a popular ornamental in the Southeastern United States. Mahoni bealei is listed as “present” on some local and state floras, but almost nothing is known regarding its invasion potential in the United States. We sampled 15 woodlots in Clemson, South Carolina, to assess the invasion of M. bealei and other woody non-indigenous species (NIS). M. bealei invaded 87% of the woodlots surveyed and species richness of NIS on these woodlots varied from 5 to 14. Stepwise-multiple regression indicated that less canopy cover and older M. bealei predicted greater abundance of M. bealei , and that not all subdivisions were equally invaded (P < 0.0001; r² = 0.88). The impact of M. bealei on native flora and fauna may be considerable, and it is likely to continue to spread in the Southeastern United States. M. bealei should be recognized as an aggressive invader in the Southeastern United States, with the potential for negative impacts on native flora and fauna.