Hassingratliff2920

In March 2020, the Rare and Imported Pathogens Laboratory at the UK Health Security Agency (UKHSA) (formerly Public Health England [PHE]) Porton Down, was tasked by the Department of Health and Social Care with setting up a national surveillance laboratory facility to study SARS-CoV-2 antibody responses and population-level sero-surveillance in response to the growing SARS-CoV-2 outbreak. In the following 12 months, the laboratory tested more than 160,000 samples, facilitating a wide range of research and informing UKHSA, DHSC, and UK government policy. Here we describe the implementation and use of the Euroimmun anti-SARS-CoV-2 IgG assay and provide an extended evaluation of its performance. We present a markedly improved overall sensitivity of 91.39% (≥14 days 92.74%, ≥21 days 93.59%) compared to our small-scale early study, and a specificity of 98.56%. In addition, we detail extended characteristics of the Euroimmun assay intra- and interassay precision, correlation to neutralization, and assay linearity. IMPORTANCE Serology assays have been useful in determining those with previous SARS-CoV-2 infection in a wide range of research and serosurveillance projects. However, assays vary in their sensitivity at detecting SARS-CoV-2 antibodies. Here, we detail an extended evaluation and characterization of the Euroimmun anti-SARS-CoV-2 IgG assay, one that has been widely used within the United Kingdom on over 160,000 samples to date.Members of the gut-dwelling Bacteroides genus have remarkable abilities in degrading a diverse set of fiber polysaccharide structures, most of which are found in the mammalian diet. As part of their metabolism, they convert these fibers to organic acids that can in turn provide energy to their host. While many studies have identified and characterized the genes and corresponding proteins involved in polysaccharide degradation, relatively little is known about Bacteroides genes involved in downstream metabolic pathways. Bacteroides thetaiotaomicron is one of the most studied species from the genus and is representative of this group in producing multiple organic acids as part of its metabolism. We focused here on several organic acid synthesis pathways in B. thetaiotaomicron, including those involved in formate, lactate, propionate, and acetate production. We identified potential genes involved in each pathway and characterized these through gene deletions coupled to growth assays and organic acid quantificati also be directly applied to the Bacteroides. click here By investigating multiple genetic pathways for organic acid production in Bacteroides thetaiotaomicron, we provide a basis for deeper understanding of these pathways. The work further enables greater understanding of Bacteroides-host relationships, as well as inter-species relationships in the microbiota, which are of importance for both human and animal gut health.WCK 5222 (cefepime/zidebactam) is a β-lactam/β-lactamase inhibitor combination that is effective against a broad range of highly drug-resistant bacterial pathogens, including those producing metallo-β-lactamase. In this study, we isolated a multidrug-resistant Pseudomonas aeruginosa clinical strain that is resistant to a variety of β-lactam antibiotics and the ceftazidime-avibactam combination. A metallo-β-lactamase gene blaDIM-2 was identified on a self-transmissible megaplasmid in the strain, which confers the resistance to β-lactam antibiotics, leaving WCK 5222 potentially one of the last treatment resorts. In vitro passaging assay combined with whole-genome sequencing revealed mutations in the pbpA gene (encoding the zidebactam target protein PBP2) in the evolved resistant mutants. Among the mutations, a V516M mutation increased the bacterial virulence in a murine acute pneumonia model. Reconstitution of the mutations in the reference strain PAO1 verified their roles in the resistance to zidebactam and ret mutations in the zidebactam target protein PBP2 play a major role in the bacterial resistance to WCK 5222. We further demonstrated that the mutations reduced the affinities between PBP2 and zidebactam and resulted in functional resistance of PBP2 to zidebactam.Pore-forming toxins (PFTs) are commonly produced by pathogenic bacteria, and understanding them is key to the development of virulence-targeted therapies. Streptococcus agalactiae, or group B Streptococcus (GBS), produces several factors that enhance its pathogenicity, including the PFT β-hemolysin/cytolysin (βhc). Little is understood about the cellular factors involved in βhc pore formation. We conducted a whole-genome CRISPR-Cas9 forward genetic screen to identify host genes that might contribute to βhc pore formation and cell death. While the screen identified the established receptor, CD59, in control experiments using the toxin intermedilysin (ILY), no clear candidate genes were identified that were required for βhc-mediated lethality. Of the top targets from the screen, two genes involved in membrane remodeling and repair represented candidates that might modulate the kinetics of βhc-induced cell death. Upon attempted validation of the results using monoclonal cell lines with targeted disruption of these genes, no effect on βhc-mediated cell lysis was observed. The CRISPR-Cas9 screen results are consistent with the hypothesis that βhc does not require a single nonessential host factor to mediate target cell death. IMPORTANCE CRISPR-Cas9 forward genetic screens have been used to identify host cell targets required by bacterial toxins. They have been used successfully to both verify known targets and elucidate novel host factors required by toxins. Here, we show that this approach fails to identify host factors required for cell death due to βhc, a toxin required for GBS virulence. These data suggest that βhc may not require a host cell receptor for toxin function or may require a host receptor that is an essential gene and would not be identified using this screening strategy.Cholesterol is the most abundant lipid in the erythrocyte. During its blood-stage development, the malaria parasite establishes an active cholesterol gradient across the various membrane systems within the infected erythrocyte. Interestingly, some antimalarial compounds have recently been shown to disrupt cholesterol homeostasis in the intraerythrocytic stages of Plasmodium falciparum. These studies point to the importance of cholesterol for parasite growth. Previously, reduction of cholesterol from the erythrocyte membrane by treatment with methyl-β-cyclodextrin (MβCD) was shown to inhibit parasite invasion and growth. In addition, MβCD treatment of trophozoite-stage P. falciparum was shown to result in parasite expulsion from the host cell. We have revisited these phenomena by using live video microscopy, ultrastructural analysis, and response to antimalarial compounds. By using time-lapse video microscopy of fluorescently tagged parasites, we show that MβCD treatment for just 30 min causes dramatic expulsior challenge in much of the world. Symptoms of malaria are caused by the growth of parasites belonging to Plasmodium spp. inside the red blood cells (RBCs), leading to their destruction. The parasite depends upon its host for much of its nutritional needs. Cholesterol is a major lipid in the RBC plasma membrane, which is the only source of this lipid for malaria parasites. We have previously shown that certain new antimalarial compounds disrupt cholesterol homeostasis in P. falciparum. Here, we use live time-lapse video microscopy to show dramatic expulsion of the parasite from the host RBC when the cholesterol content of the RBC is reduced. Remarkably, this expulsion is inhibited by the antimalarials that disrupt lipid homeostasis. We also show stereospecificity of cholesterol in supporting parasite growth inside RBC. Overall, these results point to a critical role of cholesterol in the physiology of malaria parasites.The COVID-19 pandemic caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is an unprecedented event requiring frequent adaptation to changing clinical circumstances. Convalescent immune plasma (CIP) is a promising treatment that can be mobilized rapidly in a pandemic setting. We tested whether administration of SARS-CoV-2 CIP at hospital admission could reduce the rate of ICU transfer or 28-day mortality or alter levels of specific antibody responses before and after CIP infusion. In a single-arm phase II study, patients >18 years-old with respiratory symptoms with confirmed COVID-19 infection who were admitted to a non-ICU bed were administered two units of CIP within 72 h of admission. Levels of SARS-CoV-2 detected by PCR in the respiratory tract and circulating anti-SARS-CoV-2 antibody titers were sequentially measured before and after CIP transfusion. Twenty-nine patients were transfused high titer CIP and 48 contemporaneous comparable controls were identified. All classes of antibo II trial provides a suggestion that the antiviral effects of CIP from early in the COVID-19 pandemic may delay progression to critical illness and death in specific patient populations. This study informs the optimal timing and potential population of use for CIP in COVID-19, particularly in settings without access to other interventions, or in planning for future coronavirus pandemics.The analysis of biological fluids is crucial for the diagnosis and monitoring of diseases causing effusions and helps in the diagnosis of infectious diseases. The gold standard method for cell count in biological fluids is the manual method using counting chambers. The microbiological routine procedures consist of Direct Gram staining and culture on solid or liquid media. We evaluate the analytical performance of SYSMEX UF4000 (Sysmex, Kobe, Japan) and Sysmex XN10 (Sysmex, Kobe, Japan) in comparison with cytological and microbiological routine procedures. A total of 526 biological fluid samples were included in this study (42 ascitic, 31 pleural, 31 peritoneal, 125 cerebrospinal, 281 synovial, and 16 peritoneal dialysis fluids). All samples were analyzed by flow cytometry and subsequently processed following cytological and/or microbiological routine procedures. With regard to cell counts, UF4000 (Sysmex, Kobe, Japan) showed a performance that was at least equivalent to those of the reference methods and supe a single laboratory, 24/7). It would also minimize the quantity of sample required. The performance of cytological and microbiological analysis by the flow cytometer UF 4000 (Sysmex, Kobe, Japan) has never been evaluated yet. This study has shown that bacterial count by flow cytometry using UF4000 (Sysmex, Kobe, Japan) could be used upstream of microbiological routine procedures to improve and to accelerate the diagnosis of infection of biological fluid samples.The microbiota plays an important role in human health and disease development. The lung microbiota profile in pulmonary tuberculosis (TB) patients and the effects of anti-TB treatment on the profile need to be determined thoroughly and comprehensively. This study primarily aimed to determine the lung microbiota profile associated with pulmonary TB and characterize the longitudinal changes during anti-TB treatment. A total of 53 participants, comprising 8 healthy individuals, 12 untreated pulmonary TB patients, 15 treated pulmonary TB patients, 11 cured pulmonary TB patients, and 7 lung cancer patients, were recruited in the present study. Bronchioalveolar lavage fluid (BALF) samples were collected from the above participants, and throat swabs were taken from healthy individuals. Microbiomes in the samples were examined using metagenomic next-generation sequencing (mNGS). Differences in microbiota profiles were determined through a comparison of the indicated groups. Our findings indicated that the BALF samples displayed decreased richness and diversity of the microbiota compared to those of the throat swab samples, and these two kinds of samples exhibited obvious separation on principal-coordinate analysis (PCoA) plots.