Pathogenesis and prevention of risk of cardiovascular events in patients with pneumococcal community-acquired pneumonia.

It is now well recognized that cardiovascular events (CVE) occur quite commonly, both in the acute phase and in the long-term, in patients with community-acquired pneumonia (CAP). CVE have been noted in up to 30% of patients hospitalized with all-cause CAP. One systematic review and meta-analysis of hospitalized patients with all-cause CAP noted that the incidence rates for overall cardiac events were 17.7%, for incident heart failure were 14.1%, for acute coronary syndromes were 5.3% and for incident cardiac arrhythmias were 4.7%. In the case of pneumococcal CAP, almost 20% of patients studied had one or more of these cardiac events. Recent research has provided insights into the pathogenesis of the acute cardiac events occurring in pneumococcal infections. With respect to the former, key involvements of the major pneumococcal protein virulence factor, pneumolysin, are now well documented, whilst systemic platelet-driven neutrophil activation may also contribute. However, events involved in the pathogenesis of the long-term cardiovascular sequelae remain largely unexplored. Emerging evidence suggests that persistent antigenaemia may predispose to the development of a systemic pro-inflammatory/prothrombotic phenotype underpinning the risk of future cardiovascular events. The current manuscript briefly reviews the occurrence of cardiovascular events in patients with all-cause CAP, as well as in pneumococcal and influenza infections. It highlights the close interaction between influenza and pneumococcal pneumonia. It also includes a brief discussion of mechanisms of the acute cardiac events in CAP. However, the primary focus is on the prevalence, pathogenesis and prevention of the longer-term cardiac sequelae of severe pneumococcal disease, particularly in the context of persistent antigenaemia and associated inflammation.

Mannose-sensitive and Gal-Gal binding Escherichia coli pili from recombinant strains. Chemical, functional, and serological properties.

Chromosomal genes encoding the MS and Gal-Gal binding properties have been cloned into separate recombinants and their respective pili characterized. Hapten inhibition of hemagglutination with synthetic carbohydrate receptor analogues and carbohydrate-adsorbed latex agglutination studies indicate that Gal-Gal and MS pili collectively exhibit the binding properties of the parent strain. MS pili migrated in SDS-PAGE with an Mr of 19 kdaltons and 17 kdaltons; the Mr of Gal-Gal pili was 17.5 kdaltons. The pili are chemically similar by amino acid composition and when the N-terminal cysteines are aligned, 8 of the 13 residues between positions 9 and 22 are homologous. Further, carboxy-terminal sequence homology was inferred from the carboxypeptidase digestion of a MS pili and the sequence of a carboxy-terminal tryptic peptide from Gal-Gal pili.

The PapG-adhesin at the tip of P-fimbriae provides Escherichia coli with a competitive edge in experimental bladder infections of cynomolgus monkeys.

Human urinary tract infection is an infectious disease that depends on a series of host-microbial interactions. The bacteria first colonize the colon and then the periurethral/vaginal areas; they ascend to and infect first the bladder and then the kidneys. Expression of Escherichia coli P-fimbriae constitutes the strongest correlation to renal pathogenicity, but is also related to first-time cystitis in children. The role of P-fimbriae in the preceding steps in the infectious process is unknown. To examine this, we constructed, from a P-fimbriated E. coli strain with a class II G-adhesin preferentially binding to globoside, one isogenic mutant lacking the G-adhesin and another isogenic mutant in which we replaced the papG class II allele with a class III adhesin preferentially binding to the Forssman antigen. We report here the comparison of the adhesin knockout mutant (DS17-8) and the class-switch mutant (DS17-1) with the wild-type (DS17) for in vivo colonization of the gut, vagina, and bladder of cynomolgus monkeys. It was recently shown that the class II tip G-adhesin is a prerequisite for acute pyelonephritis to occur in the monkey model in the absence of other kidney-specific adhesins or obstruction of the urinary flow. Here we show that it is not required for bladder infection but gives a competitive advantage in mixed infections. In the vagina and colon, the G-adhesin gives no competitive advantage.

Induction of gene expression in Escherichia coli after pilus-mediated adherence.

The induction of cascades of virulence factors after contact between bacteria and host cells was investigated. P-pili mediate the binding of uropathogenic Escherichia coli to its host cell receptor. After P-pili binding there was transcriptional activation of a sensor-regulator protein that is essential for the bacterial iron-starvation response. An insertion mutation of the sensor-regulator gene eliminated the ability of uropathogenic E. coli to produce siderophores and their iron-regulated membrane receptors, thereby abolishing their ability to grow in urine. These results suggest that P-pilus-mediated attachment may be an important part of the sensor-regulatory process involved in uropathogenic E. coli urinary tract infection.

Attachment of Helicobacter pylori to human gastric epithelium mediated by blood group antigens.

Helicobacter pylori is associated with development of gastritis, gastric ulcers, and adenocarcinomas in humans. The Lewis(b) (Le(b)) blood group antigen mediates H. pylori attachment to human gastric mucosa. Soluble glycoproteins presenting the Leb antigen or antibodies to the Leb antigen inhibited bacterial binding. Gastric tissue lacking Leb expression did not bind H. pylori. Bacteria did not bind to Leb antigen substituted with a terminal GalNAc alpha 1-3 residue (blood group A determinant), suggesting that the availability of H. pylori receptors might be reduced in individuals of blood group A and B phenotypes, as compared with blood group O individuals.

Coordinate regulation of beta-lactamase induction and peptidoglycan composition by the amp operon.

The amp operon, which is located on the Escherichia coli chromosome, modulates the induction of plasmid-borne beta-lactamase genes by extracellular beta-lactam antibiotics. This suggests that the gene products AmpD and AmpE may function in the transduction of external signals. beta-Lactam antibiotics are analogs of cell wall components that can be released during cell wall morphogenesis of enterobacteria. The amp operon was studied to determine its importance in signal transduction during cell wall morphogenesis. The peptidoglycan compositions of amp mutants were determined by high-performance liquid chromatography and fast atom bombardment mass spectrometry. When a chromosomal or plasmid-borne copy of ampD was present, the amount of pentapeptide-containing muropeptides in the cell wall increased upon addition of the cell wall constituent diaminopimelic acid to the growth medium. These results suggest that beta-lactamase induction and modulation of the composition of the cell wall share elements of a regulatory circuit that involves AmpD. Escherichia coli requires AmpD to respond to extracellular signaling amino acids, such as diaminopimelic acid, and this signal transduction system may regulate peptidoglycan composition in response to cell wall turnover products.

Cloning, expression, purification, crystallization and preliminary X-ray analysis of the pilus-associated sortase C from Streptococcus pneumoniae.

The pilus-associated sortase C from Streptococcus pneumoniae (SrtC or Srt-2) acts as a polymerase for the pilus subunit proteins RrgA and RrgB. Here, the crystallization and preliminary X-ray diffraction analysis of three crystal forms of SrtC are reported. One crystal form belongs to space group P2(1)2(1)2(1), with unit-cell parameters a = 48.9, b = 96.9, c = 98.9 A, alpha = beta = gamma = 90 degrees . The other two crystal forms belong to space group P222, with unit-cell parameters a = 48.8, b = 97.2, c = 99.2 A, alpha = beta = gamma = 90 degrees and a = 48.6, b = 96.5, c = 98.8 A, alpha = beta = gamma = 90 degrees , respectively. Preliminary analysis indicates the presence of two molecules in the asymmetric unit of the crystal for all three forms.

Biogenesis of the bacterial pilus.

The assembly of surface structures in gram-negative bacteria requires specialized secretion and chaperone systems localized on both sides of the cytoplasmic membrane. Major advances have been made over the last year in understanding how these systems form part of a general strategy used by bacteria to cap and target interactive subunits imported into the periplasmic space to outer membrane uncapping and assembly sites.

Induction of protective immunity after escherichia coli bladder infection in primates. Dependence of the globoside-specific P-fimbrial tip adhesin and its cognate receptor.

Clinical observations suggest that immune mechanisms affect etiology and course of recurrent cystitis. A primate infection model was used to show that primary bladder infection with a uropathogenic P-fimbriated strain (binding to globoside present in the bladder wall) protects against rechallenge with homologous as well as heterologous Escherichia coli strains for up to 5-6 mo. In contrast, mutant derivatives producing P-fimbriae either lacking the tip adhesin protein or carrying an adhesin for which no bladder receptor was present, were unable to induce protection, even though they generated bladder infections of similar duration as the wild type. Therefore, the protective effect mediated by the adhesin seemed to depend upon the presence of its cognate receptor. Since the wild strain also mediated protection against mutants that lacked the adhesin, our data suggest that the globoside-binding PapG adhesin acts as an adjuvant during infection to enhance a specific response against other bacterial antigens. In fact, the globoside-binding strain DS17, but not the mutant DS17-1, unable to bind to membrane-bound globoside, elicited a secretory IgA response to LPS in urine. These in vivo findings suggest that bacterial adhesin-ligand interactions may have signaling functions of importance for the immune response.

Fimbriae-mediated host-pathogen cross-talk.

Recent studies show that the coupling of fimbrial adhesins of uropathogenic Escherichia coli and pathogenic Neisseria species to their complementary receptors on host cells is a dynamic event, involving specific signaling to the bacteria as well as to the host cell. These studies have unveiled intriguing and novel mechanisms by which bacteria utilize their fimbriae to promote virulence at the mucosal surface and in deeper tissue.

Helicobacter pylori: molecular basis for host recognition and bacterial adherence.

The bacterium Helicobacter pylori is tropic for epithelial cells and the mucus layer in the stomach lining, and is associated with the development of gastritis, ulcers and possibly also gastric malignancies. Adherence to the gastric epithelial cells is mediated by fucosylated blood-group antigens associated with blood-group O phenotype, which could explain the higher prevalence of ulcerative disease in individuals with this blood group.

The origin of cecropins; implications from synthetic peptides derived from ribosomal protein L1.

We recently showed that Helicobacter pylori grown on plates produce cecropin-like antibacterial peptides to which H. pylori is resistant. This antibacterial activity was traced to fragments from the N-terminus of ribosomal protein L1 (Pütsep et al., Nature, April 22, 1999). The evolutionary suggestion from this finding has now been extended by the synthesis of eight peptides with sequences taken from the N-terminus of ribosomal protein L1 (RpL1) of five different species. Two peptides of different length derived from H. pylori RpL1 showed a potent antibacterial activity, while a peptide with the sequence from Escherichia coli was 20 times less active. Like cecropins the H. pylori peptides were not cytolytic. We suggest that the cecropins have evolved from ribosomal protein L1 of an ancestral intracellular pathogen that developed to a symbiont ending as an organelle. When the R1 gene moved into the host nucleus, a duplication provided a copy from which today cecropins could have evolved.

beta-Lactamase induction in gram-negative bacteria is intimately linked to peptidoglycan recycling.

A number of Gram-negative organisms normally express a chromosomally mediated class C beta-lactamase that is inducible by beta-lactam antibiotics. Data have recently emerged suggesting a close link between beta-lactamase induction and the recycling of released muramyl peptides from the bacterial peptidoglycan. Thus the AmpG transporter is responsible for the uptake into the cell of GlcNAc-anhMurNAc-tripeptide. A mutant unable to express AmpG is therefore unable to recycle the cell wall and is at the same time not possible to induce by a beta-lactam. Once inside the cytosol the above muramyl peptide and its derivative anhMurNAc-tripeptide is degraded by the cytosolic AmpD amidase that specifically releases the tripeptide from cytosolic muramyl peptides brought into the cell via AmpG. Mutants unable to produce AmpD are blocked in a cytosolic step for cell wall recycling and accumulate large amounts of cytosolic anhMurNAc-tripeptide. It is believed that cytosolic muramyl peptides can act as ligands for the beta-lactamase regulator AmpR to activate expression of beta-lactamase. AmpD mutants, therefore, constitutively overproduce the chromosomal beta-lactamase and are beta-lactam resistant. In wild-type strains beta-lactams that result in an increased cell wall breakdown will cause an increase in the cytosol of muramyl peptides leading to beta-lactamase induction. Mutants affected in the ampD gene arise readily during treatment with third-generation cephalosporins. Since these mutants lack a functional cell wall recycling system they may be at a disadvantage in the absence of selection. However, since muramyl peptides may act as cytotoxins, especially for respiratory epithelial cells, ampD mutants due to their large accumulation of anhMurNAc-tripeptide may be altered in their pathogenic properties as compared to wild-type cells possessing a normal cell wall recycling system.

Mutation of serine residue 318 in the class C beta-lactamase of Enterobacter cloacae 908R.

The involvement of the serine residue 318 in the specificity of a class C beta-lactamase was investigated. Multiple site-directed mutants at this position were generated using a polymerase chain reaction technique. These mutants were then probed for their activity towards various beta-lactam compounds. One mutant, S318G was further purified and its physico-chemical and catalytic properties determined. It was shown that the observed minimal inhibitory concentration values of this mutant could be correlated to its kinetic properties using a 'diffusion-hydrolysis' model. However, the data showed that residue 318 has little influence on the specificity of class C beta-lactamases.

Antibiotic tolerance in pneumococci.

When bacteria such as Staphylococcus aureus and Streptococcus pneumoniae are exposed to lytic antibiotics such as penicillin and vancomycin, a self-induced killing process is initiated in the organism. This killing occurs via both non-lytic and lytic processes. Recent data suggest that the non-lytic killing system, which might affect the cytoplasmic membrane, secondarily activates murein hydrolases that eventually lyse the cell. Disturbances in this suicide pathway can lead to antibiotic tolerance, a process whereby the antibiotic still exerts its bacteriostatic effects but the self-induced killing system is impaired. In mutants obtained in vitro, signaling pathways have been affected that show either increased or decreased antibiotic-induced killing. Among clinical isolates of S. pneumoniae that are tolerant to penicillin and/or vancomycin, we do not yet know whether these signaling pathways are affected. We could, however, demonstrate that the activity of murein hydrolases is negatively controlled by the production of capsular polysaccharides in one vancomycin-tolerant isolate. Hence, type and level of capsular expression might constitute one factor that determines the degree of lysis, once the killing signal has been elicited by the antibiotic.