Potential of Molecular Culture in Early Onset Neonatal Sepsis Diagnosis: A Proof of Principle Study.

Delay in the time-to-positivity of a peripheral blood culture (PBC), the gold standard for early onset neonatal sepsis (EOS) diagnosis, has resulted in excessive use of antibiotics. In this study, we evaluate the potential of the rapid Molecular Culture (MC) assay for quick EOS diagnosis. In the first part of this study, known positive and spiked blood samples were used to assess the performance of MC.

Predicting treatment response to vancomycin using bacterial DNA load as a pharmacodynamic marker in premature and very low birth weight neonates: A population PKPD study.

While positive blood cultures are the gold standard for late-onset sepsis (LOS) diagnosis in premature and very low birth weight (VLBW) newborns, these results can take days, and early markers of possible treatment efficacy are lacking. The objective of the present study was to investigate whether the response to vancomycin could be quantified using bacterial DNA loads (BDLs) determined by real-time quantitative polymerase chain reaction (RT-qPCR). VLBW and premature neonates with suspected LOS were included in a prospective observational study.

The SARS-CoV-2 viral load in COVID-19 patients is lower on face mask filters than on nasopharyngeal swabs.

Face masks and personal respirators are used to curb the transmission of SARS-CoV-2 in respiratory droplets; filters embedded in some personal protective equipment could be used as a non-invasive sample source for applications, including at-home testing, but information is needed about whether filters are suited to capture viral particles for SARS-CoV-2 detection. In this study, we generated inactivated virus-laden aerosols of 0.3-2 microns in diameter (0.

Moderate positive predictive value of a multiplex real-time PCR on whole blood for pathogen detection in critically ill patients with sepsis.

A novel multiplex real-time PCR for bloodstream infections (BSI-PCR) detects pathogens directly in blood. This study aimed at determining the positive predictive value (PPV) of BSI-PCR in critically ill patients with sepsis. We included consecutive patients with presumed sepsis upon admission to the intensive care unit (ICU).

Development and first evaluation of a novel multiplex real-time PCR on whole blood samples for rapid pathogen identification in critically ill patients with sepsis.

Molecular tests may enable early adjustment of antimicrobial therapy and be complementary to blood culture (BC) which has imperfect sensitivity in critically ill patients. We evaluated a novel multiplex real-time PCR assay to diagnose bloodstream pathogens directly in whole blood samples (BSI-PCR). BSI-PCR included 11 species- and four genus-specific PCRs, a molecular Gram-stain PCR, and two antibiotic resistance markers.

Evaluation of a real-time PCR assay for detection and quantification of bacterial DNA directly in blood of preterm neonates with suspected late-onset sepsis.

Background: Rapid and accurate diagnosis of neonatal sepsis is highly warranted because of high associated morbidity and mortality. The aim of this study was to evaluate the performance of a novel multiplex PCR assay for diagnosis of late-onset sepsis and to investigate the value of bacterial DNA load (BDL) determination as a measure of infection severity.

Methods: This cross-sectional study was conducted in a neonatal intensive care unit.

Presence of mcr-1-positive Enterobacteriaceae in retail chicken meat but not in humans in the Netherlands since 2009.

Recently, the plasmid-mediated colistin resistance gene mcr-1 was found in Enterobacteriaceae from humans, pigs and retail meat in China. Several reports have documented global presence of the gene in Enterobacteriaceae from humans, food animals and food since. We screened several well-characterised strain collections of Enterobacteriaceae, obtained from retail chicken meat and hospitalised patients in the Netherlands between 2009 and 2015, for presence of colistin resistance and the mcr-1 gene.

A novel quantitative PCR assay for the detection of Streptococcus pneumoniae using the competence regulator gene target comX.

Streptococcus pneumoniae is responsible for an estimated 1.6 million deaths worldwide every year. While rapid detection and timely treatment with appropriate antibiotics is preferred, this is often difficult due to the amount of time that detection with blood cultures takes.

Experimental Methods for Studying the BAM Complex in Neisseria meningitidis.

Neisseria meningitidis is a human pathogen. It is intensively studied for host-pathogen interactions and vaccine development. However, its favorable growth properties, genetic accessibility, and small genome size also make it an excellent model organism for studying fundamental biological processes, such as outer membrane biogenesis.

Transport of lipopolysaccharide to the Gram-negative bacterial cell surface.

Lipopolysaccharides (LPS) are major lipidic components of the outer membrane of most Gram-negative bacteria. They form a permeability barrier that protects these bacteria from harmful compounds in the environment. In addition, they are important signaling molecules for the innate immune system.

Development of a multiplex real-time PCR assay for the rapid diagnosis of neonatal late onset sepsis.

The diagnosis of late onset sepsis (LOS), a severe condition with high prevalence in preterm infants, is hampered by the suboptimal sensitivity and long turnaround time of blood culture. Detection of the infecting pathogen directly in blood by PCR would provide a much more timely result. Unfortunately, PCR-based assays reported so far are labor intensive and often lack direct species identification.

Involvement of Neisseria meningitidis lipoprotein GNA2091 in the assembly of a subset of outer membrane proteins.

GNA2091 of Neisseria meningitidis is a lipoprotein of unknown function that is included in the novel 4CMenB vaccine. Here, we investigated the biological function and the subcellular localization of the protein. We demonstrate that GNA2091 functions in the assembly of outer membrane proteins (OMPs) because its absence resulted in the accumulation of misassembled OMPs.

Species-specificity of the BamA component of the bacterial outer membrane protein-assembly machinery.

The BamA protein is the key component of the Bam complex, the assembly machinery for outer membrane proteins (OMP) in gram-negative bacteria. We previously demonstrated that BamA recognizes its OMP substrates in a species-specific manner in vitro. In this work, we further studied species specificity in vivo by testing the functioning of BamA homologs of the proteobacteria Neisseria meningitidis, Neisseria gonorrhoeae, Bordetella pertussis, Burkholderia mallei, and Escherichia coli in E.

Ght protein of Neisseria meningitidis is involved in the regulation of lipopolysaccharide biosynthesis.

Lipopolysaccharide (LPS) is a major component of the outer membrane of Gram-negative bacteria and is responsible for the barrier function of this membrane. A ght mutant of Neisseria meningitidis that showed increased sensitivity to hydrophobic toxic compounds, suggesting a breach in this permeability barrier, was previously described. Here, we assessed whether this phenotype was possibly caused by a defect in LPS transport or synthesis.

Zinc piracy as a mechanism of Neisseria meningitidis for evasion of nutritional immunity.

The outer membrane of Gram-negative bacteria functions as a permeability barrier that protects these bacteria against harmful compounds in the environment. Most nutrients pass the outer membrane by passive diffusion via pore-forming proteins known as porins. However, diffusion can only satisfy the growth requirements if the extracellular concentration of the nutrients is high.

Comparison of pathogen DNA isolation methods from large volumes of whole blood to improve molecular diagnosis of bloodstream infections.

For patients suffering from bloodstream infections (BSI) molecular diagnostics from whole blood holds promise to provide fast and adequate treatment. However, this approach is hampered by the need of large blood volumes. Three methods for pathogen DNA isolation from whole blood were compared, i.

Assembly of bacterial outer membrane proteins.

Various methods that are routinely used to study the subcellular localization of membrane proteins in wild-type Gram-negative bacteria fall short in genetic studies addressing the biogenesis of outer membrane proteins (OMPs). Here, we describe three biochemical methods that can be used in such studies to evaluate the proper assembly of OMPs into the outer membrane. The methods are based on (1) the differential electrophoretic mobility of folded and nonnative OMPs, (2) the intrinsically high protease resistance of folded OMPs, and (3) the observation that integral membrane proteins are not extracted from the membrane in solutions containing high concentrations of urea.

Effect of exercise and weight loss in people who have hip osteoarthritis and are overweight or obese: a prospective cohort study.

Background: Osteoarthritis (OA) is the most common joint disorder in the world and is recognized as a substantial source of disability. For people with OA of the knee, exercise in combination with weight loss is a proven, effective, conservative treatment option, yet evidence is lacking for people with hip OA.

Objective: The aim of this study was to obtain preliminary evidence of the effect of a program of exercise in combination with weight loss on physical function in people who have hip OA and are overweight or obese.

Cellular solid-state nuclear magnetic resonance spectroscopy.

Decrypting the structure, function, and molecular interactions of complex molecular machines in their cellular context and at atomic resolution is of prime importance for understanding fundamental physiological processes. Nuclear magnetic resonance is a well-established imaging method that can visualize cellular entities at the micrometer scale and can be used to obtain 3D atomic structures under in vitro conditions. Here, we introduce a solid-state NMR approach that provides atomic level insights into cell-associated molecular components.

Generating knock-out and complementation strains of Neisseria meningitidis.

The human-restricted pathogens Neisseria meningitidis and Neisseria gonorrhoeae are naturally competent for DNA uptake. This trait has been exploited extensively for genetic manipulation of these bacteria in the laboratory. Most transformation protocols were developed for N.