The capsular polysaccharide was isolated from Escherichia coli 010 : K5 : H4; it could not be obtained from a uncapsulated ( K Y ) mutant. It contains N-acetylglucosamine and glucuronic acid in a molar ratio of 1 : 1. Acid hydrolysis of the acidic polysaccharide as well as Smith degradation and degradation by deamination of the carboxyl-reduced polysaccharide suggested that the polysaccharide is composed of a disaccharide repeating unit. The data obtained by methylation analysis and nuclear magnetic resonance spectroscopy indicated that the repeating sequence of the capsular polysaccharide is the 4-fi-glucuronyl-I ,4-a-N-acetylglucosaminyl unit. This structure is similar to that of desulfo-heparin.It has been shown previously that pathogenicity of Escherichia coli bacteria correlated with the chemical and physical properties of their surface polysaccharide antigens [I, 21. E. coli strains with certain capsular polysaccharide antigens (K antigens) are frequently associated with urinary tract infections, especially in children [2 -41. The K antigens which are most prominent in these extraintestinal infections are KI, K2, K3, K5, K12 and K13. Encapsulated bacteria give rise to antibodies against K antigens in man and experimental animals. However, antibodies against the K5 polysaccharide antigen are only very rarely found after immunization with E. coli exhibiting the K5 antigen. It was therefore desirable to elucidate the structure of the K5 antigen in the hope that structural considerations may help to explain the poor immunogenicity of this important bacterial surface antigen.The following structural information of the above-mentioned K antigenic capsular polysaccharides of E. coli is available : the K1 antigen is a homopolysaccharide consisting of cc-2,S-linked neuraminic acid [5], the K2 antigen is a teichoicacid-like polymer consisting of galactopyranosyl-glycerol phosphate and galactofuranosyl-glycerol phosphate units in a molar ratio of 2: 1 [6] and the K13 antigen is a heteropolysaccharide with a repeating sequence of 3-linked ribose and 7-linked 2-keto-3-deoxy-manno-octulosonic acid (dOclA) [7]. In preliminary studies we have found that the K12 antigen consists of L-rhamnose and dOclA in a molar ratio of 2: 1 (unpublished results). In this publication we report the structure of the KS antigen from E. coli 010 : K5 : H4, compare it with similar bacterial polysaccharides and discuss the low immunogenicity of this capsular polysaccharide. MATERIALS AND METHODS Bacteria and CultivationEscherichia coli strain Bi 8337-41 (010 : K5 : H4) was used. It was obtained by Drs I. and F. Qrskov of the International Abbreviation. dOclA, 2-kelo-3-deoxy-~-manno-octulosonic acid Enzyme. b-Glucuronidase (EC 3.2.1.31).Escherichia Centre, Copenhagen. The growth conditions for the isolation of the capsular polysaccharide have been described previously [6,7]. Isolation and Purijication qf the Capsular PolysaccharideThe acidic capsular polysaccharide and the bacterial cells were precipitated from the liquid cultures by the additi...

SummaryStaphylococcus aureus and Staphylococcus epidermidis can cause serious chronic and recurrent infections that are difficult to eradicate. An important pathogenicity factor in these infections caused by S. aureus is its ability to be internalized by non-professional phagocytes thereby evading the host immune system and antibiotic treatment. Here, we report a novel mechanism involved in staphylococcal internalization by host cells, which is mediated by the major autolysin/adhesins Atl and AtlE from S. aureus and S. epidermidis respectively. In a flow cytometric internalization assay, atl and atlE mutants are significantly reduced in their capacities to be internalized by endothelial cells. Moreover, pre-incubation of endothelial cells with recombinant Atl dose-dependently inhibited internalization. As putative Atl-host cell receptor, the heat shock cognate protein Hsc70 was identified by mass spectrometry. The importance of Hsc70 in internalization was demonstrated by the inhibition of S. aureus internalization with antiHsc70 antibodies. In conclusion, this novel Atl-or AtlE-mediated internalization mechanism may represent a 'back-up' mechanism in S. aureus internalization, while it may represent the major or even sole mechanism involved in the internalization of coagulase-negative staphylococci and thus may play an important role in the pathogenesis of chronic and relapsing infections with these serious pathogens.

e Uropathogenic Escherichia coli (UPEC) is the most common cause of community-and hospital-acquired urinary tract infections (UTIs). Isolates from uncomplicated community-acquired UTIs express a variety of virulence traits that promote the efficient colonization of the urinary tract. In contrast, nosocomial UTIs can be caused by E. coli strains that differ in their virulence traits from the community-acquired UTI isolates. UPEC virulence markers are used to distinguish these facultative extraintestinal pathogens, which belong to the intestinal flora of many healthy individuals, from intestinal pathogenic E. coli (IPEC). IPEC is a diarrheagenic pathogen with a characteristic virulence gene set that is absent in UPEC. Here, we characterized 265 isolates from patients with UTIs during inpatient or outpatient treatment at a hospital regarding their phylogenies and IPEC or UPEC virulence traits. Interestingly, 28 of these isolates (10.6%) carried typical IPEC virulence genes that are characteristic of enteroaggregative E. coli (EAEC), Shiga toxin-producing E. coli (STEC), and atypical enteropathogenic E. coli (aEPEC), although IPEC is not considered a uropathogen. Twenty-three isolates harbored the astA gene coding for the EAEC heat-stable enterotoxin 1 (EAST1), and most of them carried virulence genes that are characteristic of UPEC and/or EAEC. Our results indicate that UPEC isolates from hospital patients differ from archetypal community-acquired isolates from uncomplicated UTIs by their spectrum of virulence traits. They represent a diverse group, including EAEC, as well as other IPEC pathotypes, which in addition contain typical UPEC virulence genes. The combination of typical extraintestinal pathogenic E. coli (ExPEC) and IPEC virulence determinants in some isolates demonstrates the marked genome plasticity of E. coli and calls for a reevaluation of the strict pathotype classification of EAEC.

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Strong focusing on diffraction-limited spots is essential for many photonic applications and is particularly relevant for optical trapping; however, all currently used approaches fail to simultaneously provide flexible transportation of light, straightforward implementation, compatibility with waveguide circuitry, and strong focusing. Here, we demonstrate the design and 3D nanoprinting of an ultrahigh numerical aperture meta-fibre for highly flexible optical trapping. Taking into account the peculiarities of the fibre environment, we implemented an ultrathin meta-lens on the facet of a modified single-mode optical fibre via direct laser writing, leading to a diffraction-limited focal spot with a record-high numerical aperture of up to NA ≈ 0.9. The unique capabilities of this flexible, cost-effective, bio- and fibre-circuitry-compatible meta-fibre device were demonstrated by optically trapping microbeads and bacteria for the first time with only one single-mode fibre in combination with diffractive optics. Our study highlights the relevance of the unexplored but exciting field of meta-fibre optics to a multitude of fields, such as bioanalytics, quantum technology and life sciences.

Commonly used antimicrobials show poor cellular uptake and often have limited access to intracellular targets, resulting in low antimicrobial activity against intracellular pathogens. An efficient delivery system to transport these drugs to the intracellular site of action is needed. Cell-penetrating peptides (CPPs) mediate the internalization of biologically active molecules into the cytoplasm. Here, we characterized two CPPs, ␣1H and ␣2H, derived from the Yersinia enterocolitica YopM effector protein. These CPPs, as well as Tat (trans-activator of transcription) from HIV-1, were used to deliver the antibiotic gentamicin to target intracellular bacteria. The YopM-derived CPPs penetrated different endothelial and epithelial cells to the same extent as Tat. CPPs were covalently conjugated to gentamicin, and CPP-gentamicin conjugates were used to target infected cells to kill multiple intracellular Gram-negative pathogenic bacteria, including Escherichia coli K1, Salmonella enterica serovar Typhimurium, and Shigella flexneri. Taken together, CPPs show great potential as delivery vehicles for antimicrobial agents and may contribute to the generation of new therapeutic tools to treat infectious diseases caused by intracellular pathogens.

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