In vitro activity of tigecycline and molecular characterization of extended-spectrum beta-lactamase-producing Escherichia coli and Klebsiella pneumoniae isolates from a university hospital in south-western Germany.

BACKGROUND: Extended-spectrum ß-lactamase (ESBL)-producing organisms are spreading worldwide in hospital and community settings. METHODS: A total of 328 unduplicated ESBL-producing Enterobacteriaceae isolated in 2008 and 2009 at the University Hospital of Tübingen were analysed retrospectively. RESULTS: Escherichia coli (n = 253) and Klebsiella spp. (n = 46) were the most frequent ESBL-producing species. The ESBL rates among E. coli and Klebsiella spp. increased from 3.8 and 2.1%, respectively, in 2008, to 5.2 and 2.4%, respectively, in 2009. Two E. coli and 3 Klebsiella pneumoniae ESBL producers were non-susceptible to ertapenem, most likely due to loss of porins. Antimicrobial susceptibility testing of selected, molecularly characterized ESBL producers revealed susceptibility to tigecycline among 97.9% (191/195) of the E. coli and 78.8% (26/33) of the K. pneumoniae isolates. PCR analysis and sequencing showed the presence of CTX-M-type enzymes in 91.3% of the E. coli and 87.9% of the K. pneumoniae isolates, whereby bla(CTX-M-15) was the most frequent ESBL gene both in E. coli (50.0%) and K. pneumoniae (51.5%). Only 7 single cases of potential patient-to-patient transmissions of E. coli strains were observed. CONCLUSIONS: Our data suggest that the increase in ESBL-producing E. coli and K. pneumoniae isolates at our hospital is mainly caused by growing import of Enterobacteriaceae harbouring CTX-M-type ESBLs.

BK channels in innate immune functions of neutrophils and macrophages.

Oxygen-dependent antimicrobial activity of human polymorphonuclear leukocytes (PMNs) relies on the phagocyte nicotinamide adenine dinucleotide phosphate (NADPH) oxidase to generate oxidants. As the oxidase transfers electrons from NADPH the membrane will depolarize and concomitantly terminate oxidase activity, unless there is charge translocation to compensate. Most experimental data implicate proton channels as the effectors of this charge compensation, although large-conductance Ca2+-activated K+ (BK) channels have been suggested to be essential for normal PMN antimicrobial activity. To test this latter notion, we directly assessed the role of BK channels in phagocyte function, including the NADPH oxidase. PMNs genetically lacking BK channels (BK(-/-)) had normal intracellular and extracellular NADPH oxidase activity in response to both receptor-independent and phagocytic challenges. Furthermore, NADPH oxidase activity of human PMNs and macrophages was normal after treatment with BK channel inhibitors. Although BK channel inhibitors suppressed endotoxin-mediated tumor necrosis factor-alpha secretion by bone marrow-derived macrophages (BMDMs), BMDMs of BK(-/-) and wild-type mice responded identically and exhibited the same ERK, PI3K/Akt, and nuclear factor-kappaB activation. Based on these data, we conclude that the BK channel is not required for NADPH oxidase activity in PMNs or macrophages or for endotoxin-triggered tumor necrosis factor-alpha release and signal transduction BMDMs.

Investigation of mechanisms involved in phagocytosis of Legionella pneumophila by human cells.

Legionella pneumophila, the causative agent of Legionnaires' disease, is able to survive and multiply efficiently in a variety of mammalian cells. By using in vitro assays, the uptake of L. pneumophila into monocytes has shown to be mediated, at least in part, through attachment of complement-coated bacteria to complement receptors, but complement-independent phagocytosis could also be demonstrated. Since complement levels in the human lung are normally low, the role of complement-dependent phagocytosis in the pathogenesis of Legionnaires' disease is doubtful. However, the contribution of other potential phagocytosis-related host cell surface molecules to the phagocytosis of L. pneumophila has never been investigated. We therefore analyzed the role of complement receptors 1 (CD35) and 3 (CD11b/18), the lipopolysaccharide (LPS) receptor (CD14), the beta(1)-integrin chain of the fibronectin receptor (CD29), the intercellular adhesion molecule 1 (ICAM-1, CD54) and the transferrin receptor (CD71) in the complement-independent uptake of L. pneumophila. To exclude any influence of culture conditions onto phagocytosis rates, we compared a fresh clinical isolate with an agar-adapted isolate of L. pneumophila. In addition, we used three different host cell types (MM6, HeLa and Jurkat cells) expressing different rates of complement receptors. We could show that both strains of L. pneumophila were phagocytized by the three host cell lines to the same extent, but intracellular multiplication was only found in MM6 and, although to a much lesser degree, in Jurkat cells. Preincubation of MM6 cells with monoclonal antibodies directed against the above cited phagocytosis-related receptors did not result in inhibition of L. pneumophila uptake. We therefore conclude that typical phagocytosis-related cell surface receptors are not involved in the complement-independent phagocytosis of L. pneumophila.

Accuracy of serum LDH elevation for the diagnosis of Pneumocystis jiroveci pneumonia.

In 328 immunocompromised patients, 105 with and 193 without Pneumocystis jiroveci pneumonia (PCP), serum lactate dehydrogenase (LDH) was analysed retrospectively, taking into consideration the time interval from the onset of symptoms to the start of specific therapy. 97 of the 105 PCP patients were negative for human immunodeficiency virus (HIV). Eight were positive. Of the 193 patients without PCP 134 were HIV-negative and 59 were HIV-positive. In HIV-negative patients the sensitivity of LDH elevation was 63% and specificity 43%. In HIV-positive patients sensitivity was 100% and specificity 47%. The overall accuracy of LDH for the diagnosis of PCP was 52%, 51% in HIV-negative and 58% in HIV-positive patients. Except for its sensitivity in HIV-positive patients, the value of LDH for the diagnosis of PCP should not be overestimated.