The discovery of novel pathogenic mechanisms engaged during bacterial infections requires the evolution of advanced techniques. Here, we evaluate the dual polarity matrix norharmane (NRM) to improve detection of bacterial lipid A (endotoxin), from host and vector tissues infected withFrancisella novicida (Fn). We evaluated NRM for improved detection and characterization of a wide range of lipids in both positive and negative polarities, including lipid A and phospholipids across a range of matrix-assisted laser desorption-ionization-coupled applications. NRM matrix improved the limit of detection (LOD) for monophosphoryl lipid A (MPLA) down to picogram level representing a 10-fold improvement of LOD versus 2,5-dihydroxybenzoic acid and 100-fold improvement of LOD versus 9-aminoacridine (9-AA). Improved LOD for lipid A subsequently facilitated detection of theFn lipid A major ion (m/z 1665) from extracts of infected mouse spleen and the temperature-modifiedFn lipid A atm/z 1637 from infectedDermacentor variabilis ticks. Finally, we simultaneously mapped bacterial phospholipid signatures within anFn-infected spleen along with an exclusively host-derived inositol-based phospholipid (m/z 933) demonstrating coprofiling of the host–pathogen interaction. Expanded use of NRM matrix in other infection models and endotoxin-targeting imaging experiments will improve our understanding of the lipid interactions at the host–pathogen interface.

Infectious diseases have a substantial global health impact. Clinicians need rapid and accurate diagnoses of infections to direct patient treatment and improve antibiotic stewardship. Current technologies employed in routine diagnostics are based on bacterial culture followed by morphological trait differentiation and biochemical testing, which can be time-consuming and labor-intensive. With advances in mass spectrometry (MS) for clinical diagnostics, the U.S. Food and Drug Administration has approved two microbial identification platforms based on matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) MS analysis of microbial proteins. We recently reported a novel and complementary approach by comparing MALDI-TOF mass spectra of microbial membrane lipid fingerprints to identify ESKAPE pathogens. However, this lipid-based approach used a sample preparation method that required more than a working day from sample collection to identification. Here, we report a new method that extracts lipids efficiently and rapidly from microbial membranes using an aqueous sodium acetate (SA) buffer that can be used to identify clinically relevant Gram-positive and -negative pathogens and fungal species in less than an hour. The SA method also has the ability to differentiate antibiotic-susceptible and antibiotic-resistant strains, directly identify microbes from biological specimens, and detect multiple pathogens in a mixed sample. These results should have positive implications for the manner in which bacteria and fungi are identified in general hospital settings and intensive care units.

Background Methadone client retention levels and treatment doses of patients vary widely in methadone clinics across China. Because methadone clinics have been available in China only recently, this study explored the relationship between methadone dosage and client retention in methadone maintenance programmes in Guizhou province. Methods The study used a prospective cohort study design. Injecting and non-injecting heroin-using clients who had been treated for no more than two and half months in one of eight methadone maintenance treatment clinics in Guizhou province were recruited into the cohort, beginning on 3 June 2006 and followed up until 1 June 2007. A total of 1003 participants were enrolled. Face-to-face interviews were conducted to collect baseline information, and clients’ daily doses were recorded. Results The 14-month retention rate was 56.2%. Controlling for other covariates in the multivariate Cox model, a higher methadone dose was found to predict higher client retention. Retention was also associated with intention to remain in treatment for life and the clinic attended. Conclusion Clients need to receive an adequate methadone dose to assure continued retention. Patients who expect to be treated for life have higher retention rates than patients who anticipate only short-term treatment. Key factors associated with successful clinics in China need to be elucidated.

We employed top-and middle-down analyses with multiple fragmentation techniques including electron transfer dissociation (ETD), electron capture dissociation (ECD), and matrix-assisted laser desorption ionization in-source decay (MALDI-ISD) for characterization of a reference monoclonal antibody (mAb) IgG1 and a fusion IgG protein. Fourier transform ion cyclotron resonance (FT-ICR) or high performance liquid chromatography electrospray ionization (HPLC-ESI) on an Orbitrap was employed. These experiments provided a comprehensive view on the protein species; especially for different glycosylation level in these two proteins, which showed good agreement with oligosaccharide profiling. Top-and middle-down MS provided additional information regarding glycosylation sites and different combinational protein species that were not available from oligosaccharide mapping or conventional bottom-up analysis. Finally, incorporating a limited enzymatic digestion by immunoglobulin G-degrading enzyme of Streptococcus pyogene (IdeS) with MALDI-ISD analysis enabled extended sequence coverage of the internal region of protein without pre-fractionation. Biological significance: Oligosaccharide profiling together with top-and middle-down methods enabled: 1) detection of heterogeneous glycosylated protein species and sites in intact IgG1 and fusion proteins with high mass accuracy, 2) estimation of relative abundance levels of protein species in the sample, 3) confirmation of the protein termini structural information, and 4) improved sequence coverage by MALDI-ISD analysis for the internal regions of the proteins without sample pre-fractionation.

Background The aim of this study was to clarify the potential function of LINC00922 in regulating the progression of lung cancer and its underling mechanism. Material/Methods Relative levels of LINC00922 in lung cancer tissues and cell lines was determined by quantitative polymerase chain reaction. Correlation between LINC00922 levels and pathological indexes of lung cancer patients was analyzed through the chi-square test. Subsequently, regulatory effects of LINC00922 on the proliferative, migratory, and invasive capacities of PC9 and A549 cells were evaluated. Western blot was conducted to determine the role of LINC00922 in mediating protein levels of CXCR4, E-cadherin, and vimentin. Through dual-luciferase reporter gene assay and functional experiments, the potential function of LINC00922/miRNA-204/CXCR4 regulatory loop in mediating the progression of lung cancer was explored. Results LINC00922 was highly expressed in lung cancer and correlated to the poor prognosis of lung cancer patients. Overexpression of LINC00922 accelerated PC9 and A549 cells to proliferate, migrate, and invade. CXCR4 was upregulated in lung cancer tissues and cells, which promoted lung cancer cells to migrate and invade. LINC00922 regulated the level of CXCR4 and directly bound to miRNA-204/CXCR4. LINC00922 mediated the cellular behaviors of lung cancer cells via targeting the miRNA-204/CXCR4 axis. Conclusions LINC00922 was upregulated in lung cancer, and accelerated lung cancer cells to proliferate, migrate, and invade via targeting the miRNA-204/CXCR4 axis.