In vivo proteomics identifies the competence regulon and AliB oligopeptide transporter as pathogenic factors in pneumococcal meningitis

Streptococcus pneumoniae (pneumococci) is a leading cause of severe bacterial meningitis in many countries worldwide. To characterize the repertoire of fitness and virulence factors predominantly expressed during meningitis we performed niche-specific analysis of the in vivo proteome in a mouse meningitis model, in which bacteria are directly inoculated into the cerebrospinal fluid (CSF) cisterna magna. We generated a comprehensive mass spectrometry (MS) spectra library enabling bacterial proteome analysis even in the presence of eukaryotic proteins. We recovered 200,000 pneumococci from CSF obtained from meningitis mice and by MS we identified 685 pneumococci proteins in samples from in vitro filter controls and 249 in CSF isolates. Strikingly, the regulatory two-component system ComDE and substrate-binding protein AliB of the oligopeptide transporter system were exclusively detected in pneumococci recovered from the CSF. In the mouse meningitis model, AliB-, ComDE-, or AliB-ComDE-deficiency resulted in attenuated meningeal inflammation and disease severity when compared to wild-type pneumococci indicating the crucial role of ComDE and AliB in pneumococcal meningitis. In conclusion, we show here mechanisms of pneumococcal adaptation to a defined host compartment by a proteome-based approach. Further, this study provides the basis of a promising strategy for the identification of protein antigens critical for invasive disease caused by pneumococci and other meningeal pathogens

Replacement Of The Boiler Feedpumps During The Retrofit Of The 500 Mw Units At A German Power Station

Lecturepg. 15The Janschwalde Power Station consists of six 500 MW units that were commissioned between 1981 and 1989. These units, which were equipped with turbo feedpumps manufactured in Russia, had an average efficiency of only 78 percent and a mean time between overhauls of 25,000 hr. The test was to adapt these boiler feedpumps at the lowest possible cost (only minor modifications to the base and the pipework were envisaged), and to achieve a payback time of four to five years by improving the pump efficiency and reducing operating costs. This was accomplished by fitting new cartridges into the existing barrel casings. These cartridges consisted of six-stages of a well-proved hydraulic with a specific speed of 1700 operating at the nominated turbine speed of 5600 to 5700 rpm. The processing of the contract within the pump manufacturer’s organization is described, along with the extensive coordination and quality assurance within a tight production schedule (delivery time of only seven months before the trail operation of the first pump unit). The efficiency of the new pumps was determined onsite by means of thermodynamic measurement. The specific features of the pumps and the proof efficiency in the power station by means of thermodynamic measurement are defined in detail

Adenosine Triphosphate Neutralizes Pneumolysin-induced Neutrophil Activation.

Background: In tissue infections, adenosine triphosphate (ATP) is released into extracellular space and contributes to purinergic chemotaxis. Neutrophils are important players in bacterial clearance and recruited to the site of tissue infections. Pneumococcal infections can lead to uncontrolled hyper-inflammation of the tissue along with substantial tissue damage through excessive neutrophil activation and uncontrolled granule release. We aimed to investigate the role of ATP in neutrophil response to pneumococcal infections. Methods: Primary human neutrophils were exposed to the pneumococcal strain TIGR4 and its pneumolysin deficient mutant or directly to different concentrations of recombinant pneumolysin. Neutrophil activation was assessed by measurement of secreted azurophilic granule protein resistin and profiling of the secretome, using mass spectrometry. Results: Pneumococci are potent inducers of neutrophil degranulation. Pneumolysin was identified as a major trigger of neutrophil activation. This process is partially lysis independent and inhibited by ATP. Pneumolysin and ATP interact with each other in the extracellular space leading to reduced neutrophil activation. Proteome analyses of the neutrophil secretome confirmed that ATP inhibits pneumolysin-dependent neutrophil activation. Conclusions: Our findings suggest that despite its cytolytic activity, pneumolysin serves as a potent neutrophil activating factor. Extracellular ATP mitigates pneumolysin induced neutrophil activation