Eight hundred and eighty-three strains of Campylobacter spp. isolated between 1982 and 1989 from human stools and poultry products were screened for quinolone resistance. In this period the prevalence of resistant strains isolated from poultry products increased from 0% to 14%. During the same period the prevalence in man increased from 0% to 11%. The emergence of quinolone resistance has implications for the identification of campylobacter up to species level: the susceptibility for nalidixic acid can no longer be used as a criterion for identification in the laboratory. The rapid emergence of resistant campylobacter may also have important implications for the treatment and prophylaxis of diarrhoeal disease. The increase of quinolone resistance coincides with the increasing use of fluoroquinolones in human and veterinary medicine. Extensive use of enrofloxacin in poultry and the almost exclusive transmission route of campylobacter from chicken to man, in The Netherlands, suggests that the resistance observed is mainly due to the use of enrofloxacin in the poultry industry.

Caenorhabditis elegans is currently introduced as a new, facile, and cheap model organism to study the pathogenesis of gram-negative bacteria such as Pseudomonas aeruginosa and Salmonella enterica serovar Typhimurium. The mechanisms of killing involve either diffusible exotoxins or infection-like processes. Recently, it was shown that also some gram-positive bacteria kill C. elegans, although the precise mechanisms of killing remained open. We examined C. elegans as a pathogenesis model for the gram-positive bacterium Streptococcus pyogenes, a major human pathogen capable of causing a wide spectrum of diseases. We demonstrate that S. pyogenes kills C. elegans, both on solid and in liquid medium. Unlike P. aeruginosa and S. enterica serovar Typhimurium, the killing by S. pyogenes is solely mediated by hydrogen peroxide. Killing required live streptococci; the killing capacity depends on the amount of hydrogen peroxide produced, and killing can be inhibited by catalase. Major exotoxins of S. pyogenes are not involved in the killing process as confirmed by using specific toxin inhibitors and knockout mutants. Moreover, no accumulation of S. pyogenes in C. elegans is observed, which excludes the involvement of infection-like processes. Preliminary results show that S. pneumoniae can also kill C. elegans by hydrogen peroxide production. Hydrogen peroxide-mediated killing might represent a common mechanism by which gram-positive, catalase-negative pathogens kill C. elegans.Streptococcus pyogenes is a major human pathogen, causing a wide spectrum of pyogenic infections such as tonsillitis, pharyngitis, scarlet fever, and skin inflammation. The organism can also invade tissues and cells and cause life-threatening diseases such as necrotizing fasciitis and toxic shock syndrome. Untreated infections can lead to serious postinfection complications in the form of rheumatic heart disease and glomerulonephritis in predisposed individuals (22). The pathogenesis of streptococci is so complex that the infections caused by these organisms and their sequelae have not been completely understood. Bacterial factors, host factors, and an abnormal immune response determine the outcome of the infection. One of the difficulties in understanding the streptococcus-host interaction is the lack of a suitable animal model. Recently a mouse model of invasive streptococcal infections has been developed (16), but there remains a great need for new animal models to understand streptococcal pathogenesis.Ausubel and coworkers have introduced Caenorhabditis elegans as a new, facile, and cheap model organism to study the pathogenesis of the gram-negative bacteria Pseudomonas aeruginosa (5,15,(23)(24)(25)27) and Salmonella enterica serovar Typhimurium (1, 2, 13), and Garsin et al. showed that the gram-positive bacteria Enterococcus faecalis and Streptococcus pneumoniae kill C. elegans (8). Hodgkin et al. demonstrated that the genetically amenable nematode C. elegans is ideally suited to identify host factors (12).We examined C. elegans as a pathogenes...

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Oligosaccharide (OS)-protein conjugates are promising candidate vaccines against encapsulated bacteria, such as Haemophilus influenzae, Neisseria meningitidis, and Streptococcus pneumoniae. Although the effects of several variables such as OS chain length and protein carrier have been studied, little is known about the influence of adjuvants on the immunogenicity of OS-protein conjugates. In this study, a minimal protective trisaccharide epitope of Streptococcus pneumoniae type 3 conjugated to the cross-reacting material of diphtheria toxin was used for immunization of BALB/c mice in the presence of different adjuvants. Subsequently, half of the mice received a booster immunization with conjugate alone. Independent of the use and type of adjuvant, all mice produced long-lasting anti-polysaccharide type 3 (PS3) antibody levels, which provided full protection against challenge with pneumococcal type 3 bacteria. All adjuvants tested increased the anti-PS3 antibody levels and opsonic capacities as measured by an enzyme-linked immunosorbent assay and an in vitro phagocytosis assay. The use of QuilA or a combination of the adjuvants CpG and dimethyl dioctadecyl ammonium bromide resulted in the highest phagocytic capacities and the highest levels of Th1-related immunoglobulin G (IgG) subclasses. Phagocytic capacity correlated strongly with Th1-associated IgG2a and IgG2b levels, to a lesser extent with Th2-associated IgG1 levels, and weakly with thiocyanate elution as a measure of avidity. Thus, the improved immunogenicity of OS-protein conjugates was most pronounced for Th1-directing adjuvants.

Lipid II is a crucial component in bacterial cell wall synthesis [Breukink, E., et al. (1999) Science 286, 2361-2364. It is the target of a number of important antibiotics, which include vancomycin and nisin [Breukink, E., and de Kruijff, B. (2006) Nat. ReV. Drug DiscoVery 5,[321][322][323][324][325][326][327][328][329][330][331][332]. Here we show that a hybrid antibiotic that consists of vancomycin and nisin fragments is significantly more active than the separate fragments against vancomycin resistant entercocci (VRE). Three different hybrids were synthesized using click chemistry and compared. Optimal spacer lengths and connection points were predicted using computer modeling.Vancomycin and nisin are members of two very different classes of antibiotics that both target the essential cell wall precursor lipid II. The glycopeptide vancomycin binds with high affinity to the tripeptide part (Lys-D-Ala-D-Ala) of lipid II (3), which is also present in the immature cell wall prior to cross-linking. The N-terminal fragment (residues 1-12) of the antimicrobial peptide nisin binds the unique pyrophosphate part present on lipid II (4). The molecular mechanism of action of both antibiotics is very different but starts in both cases with the aforementioned noncovalent binding to lipid II.Upon binding, vancomycin mainly prevents the growing oligosaccharide cell wall structure from strengthening through peptide cross-linking. This results in bacterial cell death due to osmotic pressure. Resistance to vancomycin (vanA or vanB type resistance) due to a substitution of D-Lac for the terminal D-Ala residue results in a decreased level of vancomycin binding and therefore decreased efficacy (5). One approach for improving or modifying the activity of vancomycin has focused on peripheral synthetic modifications of vancomycin. Hydrophobic appendages have been the most successful modifications and have altered the mode of action of vancomycin (6, 7). A second approach has been the synthesis of vancomycin mimics (8,9). This approach has mainly focused on repairing the disrupted ligand binding by modification of the carboxylate binding pocket in small constrained peptides that mimic vancomycin, therefore attempting to restore the original mode of action of vancomycin, although by binding strongly to D-Lac. The mode of action of nisin is a so-called targeted pore forming mechanism that involves the N-terminal part of nisin [nisin(1-12)]. This part binds to the pyrophosphate portion of the lipid II molecule through five hydrogen bonds. Subsequently, when the C-terminal fragment of nisin inserts into the membrane and associates with other lipid II-nisin complexes, lethal pore formation occurs (10). The lantibiotic mutacin 1140, which is structurally very similar, especially the N-terminal part, also binds to lipid II. However, for this antibiotic, the subsequent pore formation is not observed (11). High-affinity binding to lipid II, whether or not it leads to pore formation, is thus effective in disrupting the cell wall sy...