Listeria monocytogenes tyrosine phosphatases affect wall teichoic acid composition and phage resistance

Tyrosine phosphatase (PTP)-like proteins exist in many bacteria and are segregated into two major groups: low molecular weight and conventional. The latter group also has activity as phosphoinositide phosphatases. These two kinds of PTP are suggested to be involved in many aspects of bacterial physiology including stress response, DNA binding proteins, virulence, and capsule/cell wall production. By annotation, Listeria monocytogenes possesses two potential low molecular weight and two conventional PTPs. Using L. monocytogenes wild-type (WT) strain 10403S, we have created an in-frame deletion mutant lacking all four PTPs, as well as four additional complemented strains harboring each of the PTPs. No major physiological differences were observed between the WT and the mutant lacking all four PTPs. However, the deletion mutant strain was resistant to Listeria phages A511 and P35 and sensitive to other Listeria phages. This was attributed to reduced attachment to the cell wall. The mutant lacking all PTPs was found to lack N-acetylglucosamine in its wall teichoic acid. Phage sensitivity and attachment was rescued in a complemented strain harboring a low molecular weight PTP (LMRG1707

IL-1 enhances expansion, effector function, tissue localization, and memory response of antigen-specific CD8 T cells

Here, we show that interleukin-1 (IL-1) enhances antigen-driven CD8 T cell responses. When administered to recipients of OT-I T cell receptor transgenic CD8 T cells specific for an ovalbumin (OVA) peptide, IL-1 results in an increase in the numbers of wild-type but not IL1R1−/− OT-I cells, particularly in spleen, liver, and lung, upon immunization with OVA and lipopolysaccharide. IL-1 administration also results in an enhancement in the frequency of antigen-specific cells that are granzyme B+, have cytotoxic activity, and/ or produce interferon γ (IFN-γ). Cells primed in the presence of IL-1 display enhanced expression of granzyme B and increased capacity to produce IFN-γ when rechallenged 2 mo after priming. In three in vivo models, IL-1 enhances the protective value of weak immunogens. Thus, IL-1 has a marked enhancing effect on antigen-specific CD8 T cell expansion, differentiation, migration to the periphery, and memory

Identification of an N-terminal 27 kDa fragment of Mycoplasma pneumoniae P116 protein as specific immunogen in M. pneumoniae infections

<p>Abstract</p> <p>Background</p> <p><it>Mycoplasma pneumoniae </it>is an important cause of respiratory tract infection and is increasingly being associated with other diseases such as asthma and extra-pulmonary complications. Considerable cross-reactivity is known to exist between the whole cell antigens used in the commercial serological testing assays. Identification of specific antigens is important to eliminate the risk of cross-reactions among different related organisms. Adherence of <it>M. pneumoniae </it>to human epithelial cells is mediated through a well defined apical organelle to which a number of proteins such as P1, P30, P116 and HMW1-3 have been localized, and are being investigated for adhesion, gliding and immunodiagnostic purposes.</p> <p>Methods</p> <p>A 609 bp fragment P116_(N-27),corresponding to the N-terminal region of <it>M. pneumoniae </it>P116 gene was cloned and expressed. A C-terminal fragment P1_(C-40),of P1 protein of <it>M. pneumoniae </it>was also expressed. Three IgM ELISA assays based on P116_(N-27),P1_(C-40)and (P116 _(N-27)+ P1_(C-40)) proteins were optimized and a detailed analysis comparing the reactivity of these proteins with a commercial kit was carried out. Comparative statistical analysis of these assays was performed with the SPSS version 15.0.</p> <p>Results</p> <p>The expressed P116_(N-27)protein was well recognized by the patient sera and was immunogenic in rabbit. P1_(C-40)of <it>M. pneumoniae </it>was also immunogenic in rabbit. In comparison to the reference kit, which is reported to be 100% sensitive and 75% specific, ELISA assay based on purified P116_(N-27),P1_(C-40)and (P116_(N-27)+ P1_(C-40)) proteins showed 90.3%, 87.1% and 96.8% sensitivity and 87.0%, 87.1% and 90.3% specificity respectively. The p value for all the three assays was found to be < 0.001, and there was a good correlation and association between them.</p> <p>Conclusion</p> <p>This study shows that an N-terminal fragment of P116 protein holds a promise for serodiagnosis of <it>M. pneumoniae </it>infection. The IgM ELISA assays based on the recombinant proteins seem to be suitable for the use in serodiagnosis of acute <it>M. pneumoniae </it>infections. The use of short recombinant fragments of P116 and P1 proteins as specific antigens may eliminate the risk of cross-reactions and help to develop a specific and sensitive immunodiagnostic assay for <it>M. pneumoniae </it>detection.</p

Quantum mechanical studies of lincosamides

Lincosamides are a class of antibiotics used both in clinical and veterinary practice for a wide range of pathogens. This group of drugs inhibits the activity of the bacterial ribosome by binding to the 23S RNA of the large ribosomal subunit and blocking protein synthesis. Currently, three X-ray structures of the ribosome in complex with clindamycin are available in the Protein Data Bank, which reveal that there are two distinct conformations of the pyrrolidinyl propyl group of the bound clindamycin. In this work, we used quantum mechanical methods to investigate the probable conformations of clindamycin in order to explain the two binding modes in the ribosomal 23S RNA. We studied three lincosamide antibiotics: clindamycin, lincomycin, and pirlimycin at the B3LYP level with the 6-31G** basis set. The focus of our work was to connect the conformational landscape and electron densities of the two clindamycin conformers found experimentally with their physicochemical properties. For both functional conformers, we applied natural bond orbital (NBO) analysis and the atoms in molecules (AIM) theory, and calculated the NMR parameters. Based on the results obtained, we were able to show that the structure with the intramolecular hydrogen bond C=O…H–O is the most stable conformer of clindamycin. The charge transfer between the pyrrolidine-derivative ring and the six-atom sugar (methylthiolincosamide), which are linked via an amide bond, was found to be the dominant factor influencing the high stability of this conformer

Elimination, reversal, and directional bias of optical diffraction

We experimentally demonstrate the manipulation of optical diffraction, utilizing the atomic thermal motion in a hot vapor medium of electromagnetically-induced transparency (EIT). By properly tuning the EIT parameters, the refraction induced by the atomic motion may completely counterbalance the paraxial free-space diffraction and by that eliminates the effect of diffraction for arbitrary images. By further manipulation, the diffraction can be doubled, biased asymmetrically to induced deflection, or even reversed. The latter allows an experimental implementation of an analogy to a negative-index lens

Bacteria Modulate the CD8+ T Cell Epitope Repertoire of Host Cytosol-Exposed Proteins to Manipulate the Host Immune Response

The main adaptive immune response to bacteria is mediated by B cells and CD4+ T-cells. However, some bacterial proteins reach the cytosol of host cells and are exposed to the host CD8+ T-cells response. Both gram-negative and gram-positive bacteria can translocate proteins to the cytosol through type III and IV secretion and ESX-1 systems, respectively. The translocated proteins are often essential for the bacterium survival. Once injected, these proteins can be degraded and presented on MHC-I molecules to CD8+ T-cells. The CD8+ T-cells, in turn, can induce cell death and destroy the bacteria's habitat. In viruses, escape mutations arise to avoid this detection. The accumulation of escape mutations in bacteria has never been systematically studied. We show for the first time that such mutations are systematically present in most bacteria tested. We combine multiple bioinformatic algorithms to compute CD8+ T-cell epitope libraries of bacteria with secretion systems that translocate proteins to the host cytosol. In all bacteria tested, proteins not translocated to the cytosol show no escape mutations in their CD8+ T-cell epitopes. However, proteins translocated to the cytosol show clear escape mutations and have low epitope densities for most tested HLA alleles. The low epitope densities suggest that bacteria, like viruses, are evolutionarily selected to ensure their survival in the presence of CD8+ T-cells. In contrast with most other translocated proteins examined, Pseudomonas aeruginosa's ExoU, which ultimately induces host cell death, was found to have high epitope density. This finding suggests a novel mechanism for the manipulation of CD8+ T-cells by pathogens. The ExoU effector may have evolved to maintain high epitope density enabling it to efficiently induce CD8+ T-cell mediated cell death. These results were tested using multiple epitope prediction algorithms, and were found to be consistent for most proteins tested

Cooperative Transition between Open and Closed Conformations in Potassium Channels

Potassium (K+) ion channels switch between open and closed conformations. The nature of this important transition was revealed by comparing the X-ray crystal structures of the MthK channel from Methanobacterium thermoautotrophicum, obtained in its open conformation, and the KcsA channel from Streptomyces lividans, obtained in its closed conformation. We analyzed the dynamic characteristics and energetics of these homotetrameric structures in order to study the role of the intersubunit cooperativity in this transition. For this, elastic models and in silico alanine-scanning mutagenesis were used, respectively. Reassuringly, the calculations manifested motion from the open (closed) towards the closed (open) conformation. The calculations also revealed a network of dynamically and energetically coupled residues. Interestingly, the network suggests coupling between the selectivity filter and the gate, which are located at the two ends of the channel pore. Coupling between these two regions was not observed in calculations that were conducted with the monomer, which emphasizes the importance of the intersubunit interactions within the tetrameric structure for the cooperative gating behavior of the channel

High Prevalence of Mycoplasma pneumoniae and Chlamydia pneumoniae in Children with Acute Respiratory Infections from Lima, Peru

Background Mycoplasma pneumoniae and Chlamydia pneumoniae are atypical pathogens responsible for pneumonia and a leading cause of morbidity and mortality in low income countries. The study objective is to determine the prevalence of this pathogens in Peruvian children with acute respiratory infections. Methods A consecutive cross-sectional study was conducted in Lima, Peru from May 2009 to September 2010. A total of 675 children admitted with clinical diagnoses of acute respiratory infections were tested for Mycoplasma pneumoniae and Chlamydia pneumoniae detection by polymerase chain reaction (PCR), and clinical symptoms were registered by the attending physician. Results Mycoplasma pneumonia was detected in 25.19% (170/675) of nasopharyngeal samples and Chlamydia pneumonia in 10.52% (71/675). The most common symptoms in patients with these atypical pathogens were rhinorrhea, cough and fever. A higher prevalence of Mycoplasma pneumoniae cases were registered in summer, between December 2009 and March 2010. Conclusions Mycoplasma pneumoniae and Chlamydia pneumonia are a significant cause of morbidity in Peruvian children with acute respiratory infections (ARI). Further studies should evaluate the use of reliable techniques such as PCR in Peru in order to avoid underdiagnoses of these atypical pathogens