The low concentration of free iron in body fluids creates bacteriostatic conditions for many microorganisms and is therefore an important defense factor of the body against invading bacteria. Pathogenic bacteria have developed several mechanisms for acquiring iron from the host. Siderophore-mediated iron uptake involves the synthesis of low molecular weight iron chelators called siderophores which compete with the host iron-binding glycoproteins lactoferrin (LF) and transferrin (TF) for iron. Other ways to induce iron uptake, without the mediation of siderophores, are the possession of outer membrane protein receptors that actually recognize the complex of TF or LF with iron, resulting in the internalization of this metal, and the use of heme-compounds released into the circulation after lysis of erythrocytes. In this review, the nonsiderophore-mediated iron-uptake systems used by certain pathogenic bacteria are emphasized. The possible contribution of these iron-uptake systems to the virulence of pathogens is also discussed.

Lactoferrin (LF), a cationic 80-kDa protein present in polymorphonuclear leukocytes and in mucosal secretions, is known to have antibacterial effects on gram-negative bacteria, with a concomitant release of lipopolysaccharides (LPS, endotoxin). In addition, LF is known to decrease LPS-induced cytokine release by monocytes and LPS priming of polymorphonuclear leukocytes. Its mechanism of action is incompletely understood. We have now demonstrated by in vitro-binding studies that LF binds directly to isolated lipid A and intact LPS of clinically relevant serotypes of the species which most frequently cause bacteremia (Escherichia coli, Klebsiella pneumoniae, and Pseudomonas aeruginosa), as well as to lipid A and LPS of mucosal pathogens (among others, Neisseria meningitides and Haemophilus influenzae). Binding to LPS was inhibitable by lipid A and polymyxin B but not by KDO (3-deoxy-D-manno-octulosonate), a glycoside residue present in the inner core of LPS. Binding of LF to lipid A was saturable, and an affinity constant of 2 x 109 M-1 was calculated for the LF-lipid A interaction. Our data may explain, in part, the mechanism whereby LF exerts its antibacterial and anti-endotoxic effects. Further studies on the ability of LF to block the detrimental effects of LPS, both in vitro and in vivo, are warranted.

Summary. The importance of K antigen of Klebsiella as a virulence factor was studied in nine pairs of K + and K-strains, each pair isogenic apart from the.presence of K antigen. Loss of K antigen by nine K + strains resulted in the reduced virulence of their K-variants in a mouse-skin model. This reduced virulence of K-strains for mice may be explained in all strains by a higher degree of phagocytosis as measured by chemiluminescence response of human polymorphonuclear leukocytes (PMNL) and in most strains by enhanced killing by either human PMNL or human serum or both. Although the protective role of the K antigen in serum-induced killing and killing by PMNL was generally evident, our results also suggested that other virulence factors were sometimes involved.

We have investigated the distribution of the various core types (Rl, R2, R3, in 138 Escherichia coli isolates obtained from positive blood cultures. Rabbit antisera, raised against five rough strains expressing the respective core types, were made monospecific by extensive absorption. The reactivity of the antisera was tested in ELISA with bacterial cells that had been autoclaved for full exposure of core epitopes. One hundred and thirty strains could be typed directly, while eight strains required prior digestion with proteinase K for removal of cross-reactions. Ninety-four of the strains (68 %) expressed the R1 type, and 9 (6*5%), 12 (8*7%), 7 (5.1 %) and 3 (202%) strains expressed the R2, R3, R4 and K-12 core types, respectively. An R1R4 mixed core type, hitherto not yet described, was found in 13 (9.4%) strains. Results obtained with polyclonal antisera were in agreement with those obtained with monoclonal antibodies to the R1, R2 and R3 core types. Core typing may serve as an additional serological marker next to conventional typing of 0-, H-and K-antigens. Microbiology andUnit, Vrije Universiteit, van der Boechorststraat 7, 1081 BT Amsterdam, The Netherlands

Detection and genotyping of Chlamydia trachomatis were optimized by using a polymerase chain reaction (PCR) and restriction fragment length polymorphism (RFLP) analysis performed directly with crude cells of cervical scrapes. Different PCR pretreatment methods were evaluated on samples which were positive for C. trachomatis by cell culture. In comparison with DNA extraction and different proteolytic digestion methods, a simple pretreatment of 10 min of boiling appeared to be optimal for PCR amplification. Crude samples (n = 209) were first screened for C. trachomatis by both cell culture and plasmid PCR. Subsequently, positive samples found by plasmid PCR were subjected to a direct ompi PCR-based RFLP analysis to differentiate C. trachomatis serovars A to K, Ba, Da, and Li to L3 and serovariant D-. All cervical scrapes that were found positive for C. trachomatis by cell culture (n = 30) were also positive by plasmid PCR and ompl PCR and could be easily genotyped. In addition, of the culture-negative group, eight samples were found positive by plasmid PCR. Five of these eight samples were also positive by ompi PCR; of these five, two were positive by a nested ompi PCR. Genotyping by RFLP analysis of the 35 ompi PCR-positive samples showed that serovars D, E, and F are the most prevalent types found in cervical scrapes, while serovariant Dwas also detected. This study shows that direct PCR and PCR-based RFLP analysis are feasible for detection and genotyping of C. trachomatis in cervical scrapes and are more sensitive than culture-based serotyping.