Publications by authors named "Victor Nizet"

468 Publications

Potent Activity of Ertapenem Plus Cefazolin Within Staphylococcal Biofilms: A Contributing Factor in the Treatment of Methicillin-Susceptible Endocarditis.

Open Forum Infect Dis 2022 May 23;9(5):ofac159. Epub 2022 Mar 23.

Sharp Memorial Hospital, San Diego, California, USA.

Background: Besides antistaphylococcal beta-lactams and source control, there are limited validated antimicrobial salvage options in patients with prolonged methicillin-susceptible (MSSA) bacteremia, including infective endocarditis (IE).

Methods: MSSA IE cases treated with ertapenem (ETP) plus cefazolin (CZ) were compared with matched IE cases treated with standard beta-lactam monotherapy. The bactericidal activity of ETP plus CZ was also compared with nafcillin (NAF), CZ, and ETP alone using an in vitro MSSA biofilm model.

Results: The median duration of bacteremia experienced by patients (n = 12) while on CZ or NAF was 4 days (range 1-16 days) compared with 1 day (range 1-3 days) for patients (n = 5) treated with ETP + CZ ( = .01, Mann-Whitney test). Cefazolin and NAF alone or in combination did not achieve biofilm eradication at clinically relevant concentrations. However, the addition of ETP to CZ led to bactericidal eradication within biofilms at standard dosing.

Conclusions: Ertapenem reduces CZ concentrations required to eradicate MSSA biofilms to those achievable in vivo by standard dosing, translating into shorter bacteremia duration in patients with MSSA endocarditis. Larger studies are needed to investigate ETP plus CZ therapy in the treatment of biofilm-related MSSA infections such as endocarditis.
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http://dx.doi.org/10.1093/ofid/ofac159DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC9045957PMC
May 2022

Impact of Clopidogrel on Clinical Outcomes in Patients with Staphylococcus aureus Bacteremia: a National Retrospective Cohort Study.

Antimicrob Agents Chemother 2022 Apr 13:e0211721. Epub 2022 Apr 13.

University of California, San Diego School of Medicine, La Jolla, California, USA.

Activated platelets have known antimicrobial activity against Staphylococcus aureus. Accelerated clearance of platelets induced by S. aureus can result in thrombocytopenia and increased mortality in patients. Recent studies suggest that P2Y12 inhibition protects platelets from accelerated clearance. We therefore evaluated the effect of P2Y12 inhibition on clinical outcomes in patients with S. aureus bacteremia across a large national cohort. Our retrospective cohort (2010 to 2018) included patients admitted to Veterans Affairs (VA) hospitals with blood cultures positive for S. aureus and treated with standard-of-care antibiotics. Employing propensity score-matched Cox proportional hazards regression models, we compared clinical outcomes in patients treated with clopidogrel for at least the 30 days prior to admission and continuing for at least 5 days after admission to patients without any P2Y12 inhibitor use in the year preceding admission. Mortality was significantly lower among clopidogrel users than P2Y12 inhibitor nonusers ( = 147 propensity score-matched pairs): the inpatient mortality hazard ratio (HR) was 0.11 (95% confidence interval [CI], 0.01 to 0.86), and 30-day mortality HR was 0.43 (95% CI, 0.19 to 0.98). There were no differences in 30-day readmission, 30-day S. aureus reinfection, microbiological clearance, or thrombocytopenia. Clopidogrel use at the time of infection reduced in-hospital mortality by 89% and 30-day mortality by 57% among a cohort of patients with S. aureus bacteremia. These results support the need to further study the use of P2Y12 inhibitors as adjunctive therapy in S. aureus bloodstream infections.
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http://dx.doi.org/10.1128/aac.02117-21DOI Listing
April 2022

Inhibition of histamine accumulation by novel histamine-degrading species of sp. isolated from goats and sheep milk.

Food Sci Nutr 2022 Feb 9;10(2):354-362. Epub 2022 Jan 9.

Department of Food Hygiene and Public Health School of Veterinary Medicine Shiraz University Shiraz Iran.

Histamine is an active amine compound that occurs in various fermented foods that may cause adverse effects on the human health. Certain microorganisms are able to degrade histamine by an oxidative deamination reaction. Therefore, the present study aimed to quantify histamine-forming and/or -degrading activity of the isolates derived from milk of goat and sheep herds, in Iran, by the capillary zone electrophoresis (CZE) method; and we evaluated the molecular characteristics of staphylococcal isolates. Among 243 staphylococcal isolates, 29 histamine-degrading bacteria were identified. One of these isolates, identified as , No. 605, exhibited the highest activity compared to others, degrading available histamine to 58.33% within 24 h. By polymerase chain reaction (PCR) analysis, the isolate, No. 605 that exhibited remarkable histamine-degrading activity lacked the genes encoding coagulase and DNase, nor did it harbor any of the five classical enterotoxin genes. This is the first report to show that seven species, including , , , , . , . , and , were able to degrade histamine, which were hitherto not known to have this capacity. Therefore, histamine-degrading activity is a definite criterion to introduce fermenting organisms able to decrease histamine content in different food products.
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http://dx.doi.org/10.1002/fsn3.2723DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8825735PMC
February 2022

Human Milk Oligosaccharides Reduce Murine Group B Vaginal Colonization with Minimal Impact on the Vaginal Microbiota.

mSphere 2022 Feb 5;7(1):e0088521. Epub 2022 Jan 5.

Department of Molecular Virology and Microbiology, Baylor College of Medicinegrid.39382.33, Houston, Texas, USA.

Group B Streptococcus (GBS) colonizes the vaginal mucosa of a significant percentage of healthy women and is a leading cause of neonatal bacterial infections. Currently, pregnant women are screened in the last month of pregnancy, and GBS-positive women are given antibiotics during parturition to prevent bacterial transmission to the neonate. Recently, human milk oligosaccharides (HMOs) isolated from breastmilk were found to inhibit GBS growth and biofilm formation , and women that make certain HMOs are less likely to be vaginally colonized with GBS. Using human vaginal epithelial cells and a murine vaginal colonization model, we tested the impact of HMO treatment on GBS burdens and the composition of the endogenous microbiota by 16S rRNA amplicon sequencing. HMO treatment reduced GBS vaginal burdens with minimal alterations to the vaginal microbiota. HMOs displayed potent inhibitory activity against GBS , but HMO pretreatment did not alter adherence of GBS or the probiotic Lactobacillus rhamnosus to human vaginal epithelial cells. In addition, disruption of a putative GBS glycosyltransferase (Δ_0913) rendered the bacterium largely resistant to HMO inhibition and but did not compromise its adherence, colonization, or biofilm formation in the absence of HMOs. We conclude that HMOs are a promising therapeutic bioactive to limit GBS vaginal colonization with minimal impacts on the vaginal microenvironment. During pregnancy, GBS ascension into the uterus can cause fetal infection or preterm birth. In addition, GBS exposure during labor creates a risk of serious disease in the vulnerable newborn and mother postpartum. Current recommended prophylaxis consists of administering broad-spectrum antibiotics to GBS-positive mothers during labor. Although antibiotics have significantly reduced GBS neonatal disease, there are several unintended consequences, including altered neonatal gut bacteria and increased risk for other types of infection. Innovative preventions displaying more targeted antimicrobial activity, while leaving the maternal microbiota intact, are thus appealing. Using a mouse model, we found that human milk oligosaccharides (HMOs) reduce GBS burdens without perturbing the vaginal microbiota. We conclude that HMOs are a promising alternative to antibiotics to reduce GBS neonatal disease.
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http://dx.doi.org/10.1128/msphere.00885-21DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8730812PMC
February 2022

Dexmedetomidine does not directly inhibit neutrophil extracellular trap production.

Br J Anaesth 2022 02 19;128(2):e51-e54. Epub 2021 Nov 19.

Department of Anesthesiology, Division of Critical Care Medicine, University of California San Diego, La Jolla, CA, USA. Electronic address:

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http://dx.doi.org/10.1016/j.bja.2021.11.015DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8603039PMC
February 2022

Bicarbonate modulates delafloxacin activity against MDR Staphylococcus aureus and Pseudomonas aeruginosa.

J Antimicrob Chemother 2022 02;77(2):433-442

Collaborative to Halt Antibiotic-Resistant Microbes (CHARM), UC San Diego, La Jolla, CA 92093, USA.

Objectives: To investigate the utility of recently approved delafloxacin and other fluoroquinolones against leading MDR bacterial pathogens under physiologically relevant conditions.

Methods: MIC and MBC assays were conducted for MDR strains of Staphylococcus aureus, Pseudomonas aeruginosa, Acinetobacter baumannii and Klebsiella pneumoniae in the standard antibiotic susceptibility testing medium CAMHB, amended Roswell-Park Memorial Institute tissue culture medium (RPMI+) or 20% fresh human whole blood. In vivo correlation of in vitro findings was performed in a murine P. aeruginosa pneumonia model. Mechanistic bases for the findings were explored by altering media conditions and with established fluoroquinolone accumulation assays.

Results: Fluoroquinolone MICs were increased in RPMI+ compared with CAMHB for all four MDR pathogens. Specifically, delafloxacin MICs were increased 32-fold versus MDR S. aureus and 8-fold versus MDR P. aeruginosa. MBC assays in 20% human whole blood and a murine MDR P. aeruginosa pneumonia model both confirmed that delafloxacin activity was reduced under physiological conditions. Bicarbonate (HCO3-), a key component of host physiology found in RPMI+ but absent from CAMHB, dictated delafloxacin susceptibility in CAMHB and RPMI+ by impairing its intracellular accumulation.

Conclusions: Standard in vitro antibiotic susceptibility testing conditions overpredicted the effectiveness of delafloxacin against MDR pathogens by failing to capture the role of the biological buffer HCO3- to impair delafloxacin accumulation. This work showcases limitations of our current antibiotic susceptibility testing paradigm and highlights the importance of understanding host microenvironmental conditions that impact true clinical efficacy.
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http://dx.doi.org/10.1093/jac/dkab421DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8809187PMC
February 2022

Editorial: Host-Pathogen Interactions During Pneumococcal Infection.

Front Cell Infect Microbiol 2021 25;11:752959. Epub 2021 Oct 25.

Department of Pediatrics, University of California (UC), San Diego, La Jolla, CA, United States.

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http://dx.doi.org/10.3389/fcimb.2021.752959DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8573109PMC
November 2021

Uremic serum damages endothelium by provoking excessive neutrophil extracellular trap formation.

Sci Rep 2021 11 2;11(1):21439. Epub 2021 Nov 2.

Department of Clinical Immunology, Hallym University Sacred Heart Hospital, Anyang, South Korea.

Cardiovascular disease (CVD) is the leading cause of death in patients with chronic kidney disease (CKD). Endothelial cell (EC) dysfunction is a key CKD-specific risk factor; however, the mechanisms by which uremia harms the endothelium are still unclear. We report a role for excessive neutrophil extracellular trap (NET) formation induced by uremic serum on EC injury. Level of plasma nucleosome and myeloperoxidase-DNA, established in vivo markers of NETs, as well as intracellular adhesion molecule (ICAM)-1 were measured in hemodialysis (HD) patients and healthy volunteers (HV) and their prognostic role evaluated. For in vitro studies, HV-derived neutrophils and differentiated HL-60 cells by retinoic acid were used to determine the effect of uremic serum-induced NETs on human umbilical vein EC (HUVEC). The level of in vivo NETs was significantly higher in incident HD patients compared to HV, and these markers were strongly associated with ICAM-1. Specifically, nucleosome and ICAM-1 levels were independent predictors of a composite endpoint, all-cause mortality, or vascular access failure. In vitro, HD-derived uremic serum significantly increased NET formation both in dHL-60 and isolated neutrophils compared to control serum, and these NETs decreased EC viability and induced their apoptosis. In addition, the level of ICAM-1, E-selectin and von Willebrand factor in HUVEC supernatant was significantly increased by uremic serum-induced NETs compared to control serum-induced NETs. Dysregulated neutrophil activities in the uremic milieu may play a key role in vascular inflammatory responses. The high mortality and CVD rates in ESRD may be explained in part by excessive NET formation leading to EC damage and dysfunction.
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http://dx.doi.org/10.1038/s41598-021-00863-wDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8563801PMC
November 2021

The S Protein of Group B Is a Critical Virulence Determinant That Impacts the Cell Surface Virulome.

Front Microbiol 2021 14;12:729308. Epub 2021 Oct 14.

Department of Pharmacology, University of California San Diego, La Jolla, CA, United States.

Group B (GBS, ) is a human commensal and occasional pathogen that remains a leading cause of neonatal sepsis and meningitis with increasing disease burden in adult populations. Although programs for universal screening in pregnancy to guide intrapartum prophylaxis have reduced GBS invasive disease burden resulting from mother-to-newborn transfer during birth, better knowledge of disease mechanisms may elucidate new strategies to reduce antibiotic exposure. In our efforts to expand the knowledge base required for targeted anti-virulence therapies, we identified a GBS homolog for a recently identified virulence determinant of group A , S protein, and evaluated its role in GBS pathogenesis. A GBS S protein deletion mutant, Δ, showed altered cell-surface properties compared to the WT parent strain, including defective retention of its surface polysaccharide. Quantitative proteome analysis of enzymatically shaved surface epitopes of the GBS Δ mutant revealed a dysregulated cell surface virulome, with reduced abundance of several protein and glycoprotein components. The Δ mutant showed markedly attenuated virulence in a murine model of GBS systemic infection, with increased proteasome activity detected in the spleens of animals infected with the Δ mutant. These results expand the key roles S protein plays in streptococcal pathogenesis and introduces a new GBS virulence determinant and potential target for therapy development.
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http://dx.doi.org/10.3389/fmicb.2021.729308DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8551713PMC
October 2021

Bacterial Membrane-Derived Vesicles Attenuate Vancomycin Activity against Methicillin-Resistant .

Microorganisms 2021 Sep 29;9(10). Epub 2021 Sep 29.

Research Group for Host-Microbe Interactions, UiT-The Arctic University of Norway, 9037 Tromsø, Norway.

Methicillin-resistant (MRSA) has evolved numerous antimicrobial resistance mechanisms and is identified as a serious public health threat by the World Health Organization and U.S. Centers for Disease Control and Prevention. The glycopeptide vancomycin (VAN) remains a cornerstone of therapy for severe MRSA infections despite increasing reports of therapeutic failure in hospitalized patients with bacteremia or pneumonia. Recently, the role of released bacterial-derived membrane vesicles (MVs) in antibiotic resistance has garnered attention. Here we examined the effect of exogenous MRSA-derived MVs on VAN activity against MRSA in vitro, using minimum inhibitory concentration and checkerboard assays, and ex vivo, incorporating components of host innate immunity such as neutrophils and serum complement present in blood. Additionally, the proteome of MVs from VAN-exposed MRSA was characterized to determine if protein expression was altered. The presence of MVs increased the VAN MIC against MRSA to values where clinical failure is commonly observed. Furthermore, the presence of MVs increased survival of MRSA pre-treated with sub-MIC concentrations of VAN in whole blood and upon exposure to human neutrophils but not human serum. Unbiased proteomic analysis also showed an elevated expression of MV proteins associated with antibiotic resistance (e.g., ) or proteins that are functionally linked to cell membrane/wall metabolism. Together, our findings indicate MRSA-derived MVs are capable of lowering susceptibility of the pathogen to VAN, whole-blood- and neutrophil-mediated killing, a new pharmacodynamic consideration for a drug increasingly linked to clinical treatment failures.
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http://dx.doi.org/10.3390/microorganisms9102055DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8539228PMC
September 2021

Endothelial Heparan Sulfate Mediates Hepatic Neutrophil Trafficking and Injury during Staphylococcus aureus Sepsis.

mBio 2021 10 21;12(5):e0118121. Epub 2021 Sep 21.

Department of Cellular and Molecular Medicine, University of California, San Diegogrid.266100.3, La Jolla, California, USA.

Hepatic failure is an important risk factor for poor outcome in septic patients. Using a chemical tagging workflow and high-resolution mass spectrometry, we demonstrate that rapid proteome remodeling of the vascular surfaces precedes hepatic damage in a murine model of Staphylococcus aureus sepsis. These early changes include vascular deposition of neutrophil-derived proteins, shedding of vascular receptors, and altered levels of heparin/heparan sulfate-binding factors. Modification of endothelial heparan sulfate, a major component of the vascular glycocalyx, diminishes neutrophil trafficking to the liver and reduces hepatic coagulopathy and organ damage during the systemic inflammatory response to infection. Modifying endothelial heparan sulfate likewise reduces neutrophil trafficking in sterile hepatic injury, reflecting a more general role of heparan sulfate contribution to the modulation of leukocyte behavior during inflammation. Vascular glycocalyx remodeling is critical to sepsis pathology, but the glycocalyx components that contribute to this process remain poorly characterized. This article shows that during Staphylococcus aureus sepsis, the liver vascular glycocalyx undergoes dramatic changes in protein composition associated with neutrophilic activity and heparin/heparan sulfate binding, all before organ damage is detectable by standard circulating liver damage markers or histology. Targeted manipulation of endothelial heparan sulfate modulates S. aureus sepsis-induced hepatotoxicity by controlling the magnitude of neutrophilic infiltration into the liver in both nonsterile and sterile injury. These data identify an important vascular glycocalyx component that impacts hepatic failure during nonsterile and sterile injury.
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http://dx.doi.org/10.1128/mBio.01181-21DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8546592PMC
October 2021

Immunobiology of the Classical Lancefield Group A Streptococcal Carbohydrate Antigen.

Infect Immun 2021 11 20;89(12):e0029221. Epub 2021 Sep 20.

Biomedical Sciences Graduate Program, UC San Diego, La Jolla, California, USA.

Group A Streptococcus (GAS) is a preeminent human bacterial pathogen causing hundreds of millions of infections each year worldwide. In the clinical setting, the bacterium is easily identified by a rapid antigen test against the group A carbohydrate (GAC), a polysaccharide that comprises 30 to 50% of the GAS cell wall by weight. Originally described by Rebecca Lancefield in the 1930s, GAC consists of a polyrhamnose backbone and a -acetylglucosamine (GlcNAc) side chain. This side chain, the species-defining immunodominant antigen, is potentially implicated in autoreactive immune responses against human heart or brain tissue in poststreptococcal rheumatic fever or rheumatic heart disease. The recent discovery of the genetic locus encoding GAC biosynthesis and new insights into its chemical structure have provided novel insights into the assembly of the polysaccharide, its contribution to immune evasion and virulence, and ideas for safely harnessing its natural immunogenicity in vaccine design. This minireview serves to summarize the emerging new literature on GAC, the eponymous cell well antigen that provides structural integrity to GAS and directly interfaces with host innate and adaptive immune responses.
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http://dx.doi.org/10.1128/IAI.00292-21DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8594609PMC
November 2021

Heat shock protein 27 activity is linked to endothelial barrier recovery after proinflammatory GPCR-induced disruption.

Sci Signal 2021 Aug 31;14(698):eabc1044. Epub 2021 Aug 31.

Department of Pharmacology, University of California, San Diego, La Jolla, CA 92093, USA.

Vascular inflammation causes endothelial barrier disruption and tissue edema. Several inflammatory mediators act through G protein–coupled receptors (GPCRs), including protease-activated receptor-1 (PAR1), to elicit inflammatory responses. The activation of PAR1 by its ligand thrombin stimulates proinflammatory, p38 mitogen-activated protein kinase (MAPK) signaling that promotes endothelial barrier disruption. Through mass spectrometry phosphoproteomics, we identified heat shock protein 27 (HSP27), which exists as a large oligomer that binds to actin, as a promising candidate for the p38-mediated regulation of barrier integrity. Depletion of HSP27 by siRNA enhanced endothelial cell barrier permeability and slowed recovery after thrombin stimulation. We further showed that two effector kinases of p38 MAPK, MAPKAPK2 (MK2) and MAPKAPK3 (MK3), differentially phosphorylated HSP27 at Ser, Ser, and Ser. Whereas inhibition of thrombin-stimulated p38 activation blocked HSP27 phosphorylation at all three sites, inhibition of MK2 reduced the phosphorylation of only Ser and Ser. Inhibition of both MK2 and MK3 was necessary to attenuate Ser phosphorylation. Thrombin-stimulated p38-MK2-MK3 signaling induced HSP27 oligomer disassembly. However, a phosphorylation-deficient mutant of HSP27 exhibited defective oligomer disassembly and altered the dynamics of barrier recovery after thrombin stimulation. Moreover, blocking HSP27 oligomer reassembly with the small-molecule inhibitor J2 enhanced endothelial barrier permeability in vitro and vascular leakage in vivo in response to PAR1 activation. These studies reveal the distinct regulation of HSP27 phosphorylation and function induced by the GPCR-stimulated p38-MK2-MK3 signaling axis that controls the dynamics of endothelial barrier recovery in vitro and vascular leakage in vivo.
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http://dx.doi.org/10.1126/scisignal.abc1044DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8538426PMC
August 2021

Streptolysins are the primary inflammasome activators in macrophages during Streptococcus pyogenes infection.

Immunol Cell Biol 2021 11 18;99(10):1040-1052. Epub 2021 Sep 18.

Australian Infectious Diseases Research Centre, School of Chemistry and Molecular Biosciences, The University of Queensland, St Lucia, QLD, Australia.

Group A Streptococcus (GAS) is a Gram-positive bacterial pathogen that causes an array of infectious diseases in humans. Accumulating clinical evidence suggests that proinflammatory interleukin (IL)-1β signaling plays an important role in GAS disease progression. The host regulates the production and secretion of IL-1β via the cytosolic inflammasome pathway. Activation of the NLR family pyrin domain-containing 3 (NLRP3) inflammasome complex requires two signals: a priming signal that stimulates increased transcription of genes encoding the components of the inflammasome pathway, and an activating signal that induces assembly of the inflammasome complex. Here we show that GAS-derived lipoteichoic acid can provide a priming signal for NLRP3 inflammasome activation. As only few GAS-derived proteins have been associated with inflammasome-dependent IL-1β signaling, we investigated novel candidates that might play a role in activating the inflammasome pathway by infecting mouse bone marrow-derived macrophages and human THP-1 macrophage-like cells with a panel of isogenic GAS mutant strains. We found that the cytolysins streptolysin O (SLO) and streptolysin S are the main drivers of IL-1β release in proliferating logarithmic phase GAS. Using a mutant form of recombinant SLO, we confirmed that bacterial pore formation on host cell membranes is a key mechanism required for inflammasome activation. Our results suggest that streptolysins are major determinants of GAS-induced inflammation and present an attractive target for therapeutic intervention.
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http://dx.doi.org/10.1111/imcb.12499DOI Listing
November 2021

Machine Learning of Bacterial Transcriptomes Reveals Responses Underlying Differential Antibiotic Susceptibility.

mSphere 2021 08 25;6(4):e0044321. Epub 2021 Aug 25.

Department of Bioengineering, University of California-San Diego, La Jolla, California, USA.

antibiotic susceptibility testing often fails to accurately predict drug efficacies, in part due to differences in the molecular composition between standardized bacteriologic media and physiological environments within the body. Here, we investigate the interrelationship between antibiotic susceptibility and medium composition in Escherichia coli K-12 MG1655 as contextualized through machine learning of transcriptomics data. Application of independent component analysis, a signal separation algorithm, shows that complex phenotypic changes induced by environmental conditions or antibiotic treatment are directly traced to the action of a few key transcriptional regulators, including RpoS, Fur, and Fnr. Integrating machine learning results with biochemical knowledge of transcription factor activation reveals medium-dependent shifts in respiration and iron availability that drive differential antibiotic susceptibility. By extension, the data generation and data analytics workflow used here can interrogate the regulatory state of a pathogen under any measured condition and can be applied to any strain or organism for which sufficient transcriptomics data are available. Antibiotic resistance is an imminent threat to global health. Patient treatment regimens are often selected based on results from standardized antibiotic susceptibility testing (AST) in the clinical microbiology lab, but these tests frequently misclassify drug effectiveness due to their poor resemblance to actual host conditions. Prior attempts to understand the combined effects of drugs and media on antibiotic efficacy have focused on physiological measurements but have not linked treatment outcomes to transcriptional responses on a systems level. Here, application of machine learning to transcriptomics data identified medium-dependent responses in key regulators of bacterial iron uptake and respiratory activity. The analytical workflow presented here is scalable to additional organisms and conditions and could be used to improve clinical AST by identifying the key regulatory factors dictating antibiotic susceptibility.
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http://dx.doi.org/10.1128/mSphere.00443-21DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8386450PMC
August 2021

Environmental conditions dictate differential evolution of vancomycin resistance in Staphylococcus aureus.

Commun Biol 2021 06 25;4(1):793. Epub 2021 Jun 25.

Department of Bioengineering, University of California San Diego, La Jolla, CA, USA.

While microbiological resistance to vancomycin in Staphylococcus aureus is rare, clinical vancomycin treatment failures are common, and methicillin-resistant S. aureus (MRSA) strains isolated from patients after prolonged vancomycin treatment failure remain susceptible. Adaptive laboratory evolution was utilized to uncover mutational mechanisms associated with MRSA vancomycin resistance in a physiological medium as well as a bacteriological medium used in clinical susceptibility testing. Sequencing of resistant clones revealed shared and media-specific mutational outcomes, with an overlap in cell wall regulons (walKRyycHI, vraSRT). Evolved strains displayed similar properties to resistant clinical isolates in their genetic and phenotypic traits. Importantly, resistant phenotypes that developed in physiological media did not translate into resistance in bacteriological media. Further, a bacteriological media-specific mechanism for vancomycin resistance associated with a mutated mprF was confirmed. This study bridges the gap between the understanding of clinical and microbiological vancomycin resistance in S. aureus and expands the number of allelic variants (18 ± 4 mutations for the top 5 mutated genes) that result in vancomycin resistance phenotypes.
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http://dx.doi.org/10.1038/s42003-021-02339-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8233327PMC
June 2021

Hypoxia-Inducible Factor 1 Alpha Is Dispensable for Host Defense of Group B Streptococcus Colonization and Infection.

J Innate Immun 2021 21;13(6):391-403. Epub 2021 May 21.

Division of Host-Microbe Systems & Therapeutics, Department of Pediatrics, UC San Diego, La Jolla, California, USA.

Group B Streptococcus (GBS) is a leading cause of neonatal morbidity and mortality, and the primary source of exposure is the maternal vagina. Intrapartum antibiotic prophylaxis for GBS-positive mothers has reduced the incidence of GBS early-onset disease, however, potential long-lasting influence of an antibiotic-altered neonatal microbiota, and the frequent clinical sequelae in survivors of invasive GBS infection, compels alternative treatment options for GBS. Here, we examined the role of transcription factor hypoxia-inducible factor 1 alpha (HIF-1α), widely recognized as a regulator of immune activation during infection, in the host response to GBS. Given the importance of endogenous HIF-1α for innate immune defense, and the potential utility of HIF-1α stabilization in promoting bacterial clearance, we hypothesized that HIF-1α could play an important role in coordinating host responses to GBS in colonization and systemic disease. Counter to our hypothesis, we found that GBS infection did not induce HIF-1α expression in vaginal epithelial cells or murine macrophages, nor did HIF-1α deficiency alter GBS colonization or pathogenesis in vivo. Furthermore, pharmacological enhancement of HIF-1α did not improve control of GBS in pathogenesis and colonization models, while displaying inhibitory effects in vaginal epithelial cytokines and immune cell killing in vitro. Taken together, we conclude that HIF-1α is not a prominent aspect of the host response to GBS colonization or invasive disease, and its pharmacological modulation is unlikely to provide significant benefit against this important neonatal pathogen.
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http://dx.doi.org/10.1159/000515739DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8613573PMC
January 2022

Current Paradigms of Combination Therapy in Methicillin-Resistant Staphylococcus aureus (MRSA) Bacteremia: Does it Work, Which Combination, and For Which Patients?

Clin Infect Dis 2021 12;73(12):2353-2360

Division of Host-Microbe Systems and Therapeutics, Center for Immunity, Infection and Inflammation, University of California-San Diego School of Medicine, La Jolla, California, USA.

The last several years have seen an emergence of literature documenting the utility of combination antimicrobial therapy, particularly in the salvage of refractory methicillin-resistant Staphylococcus aureus (MRSA) bacteremia. Recent clinical data are shaping conundrums of which regimens may be more beneficial, which can be potentially harmful, and which subset of patients stand to benefit from more aggressive treatment regimens than called for by current standards. In addition, the incorporation of combination therapy for MRSA bacteremia should be accompanied by the reminder that antimicrobial therapy does not need to be uniform for the entire duration, with an early intensive phase in high inoculum infections (eg, with combination therapy), followed by a consolidation phase (ie, monotherapy). This review and perspective consolidates the recent data on this subject and directs future goals in filling the knowledge gaps to methodically move forward towards improving patient outcomes.
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http://dx.doi.org/10.1093/cid/ciab452DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8826002PMC
December 2021

Ticagrelor Increases Platelet-Mediated Staphylococcus aureus Killing, Resulting in Clearance of Bacteremia.

J Infect Dis 2021 11;224(9):1566-1569

Department of Pediatrics, University of California, San Diego School of Medicine, La Jolla, California, USA.

Platelets are a critical immune defense against Staphylococcus aureus bloodstream infections. Staphylococcus aureus α-toxin is a virulence factor that decreases platelet viability and accelerates platelet clearance. It has been shown that ticagrelor blocks α-toxin-mediated platelet injury and resulting thrombocytopenia, protecting mice in a lethal S. aureus sepsis model. We now present the use of ticagrelor as adjunctive therapy in a patient with a S. aureus endovascular infection and thrombocytopenia, associated with restoration of platelet count and bacteremia clearance. Ticagrelor enhanced platelet killing of the S. aureus bloodstream isolate from the treated patient in vitro.
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http://dx.doi.org/10.1093/infdis/jiab146DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8599881PMC
November 2021

Streptococcus pyogenes upregulates arginine catabolism to exert its pathogenesis on the skin surface.

Cell Rep 2021 03;34(13):108924

Department of Oral and Molecular Microbiology, Osaka University Graduate School of Dentistry, Suita, Osaka 565-0871, Japan. Electronic address:

The arginine deiminase (ADI) pathway has been found in many kinds of bacteria and functions to supplement energy production and provide protection against acid stress. The Streptococcus pyogenes ADI pathway is upregulated upon exposure to various environmental stresses, including glucose starvation. However, there are several unclear points about the advantages to the organism for upregulating arginine catabolism. We show that the ADI pathway contributes to bacterial viability and pathogenesis under low-glucose conditions. S. pyogenes changes global gene expression, including upregulation of virulence genes, by catabolizing arginine. In a murine model of epicutaneous infection, S. pyogenes uses the ADI pathway to augment its pathogenicity by increasing the expression of virulence genes, including those encoding the exotoxins. We also find that arginine from stratum-corneum-derived filaggrin is a key substrate for the ADI pathway. In summary, arginine is a nutrient source that promotes the pathogenicity of S. pyogenes on the skin.
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http://dx.doi.org/10.1016/j.celrep.2021.108924DOI Listing
March 2021

Repurposed drugs block toxin-driven platelet clearance by the hepatic Ashwell-Morell receptor to clear bacteremia.

Sci Transl Med 2021 03;13(586)

Biomedical Sciences Graduate Program, UC San Diego, La Jolla, CA 92093, USA.

(SA) bloodstream infections cause high morbidity and mortality (20 to 30%) despite modern supportive care. In a human bacteremia cohort, we found that development of thrombocytopenia was correlated to increased mortality and increased α-toxin expression by the pathogen. Platelet-derived antibacterial peptides are important in bloodstream defense against SA, but α-toxin decreased platelet viability, induced platelet sialidase to cause desialylation of platelet glycoproteins, and accelerated platelet clearance by the hepatic Ashwell-Morell receptor (AMR). Ticagrelor (Brilinta), a commonly prescribed P2Y12 receptor inhibitor used after myocardial infarction, blocked α-toxin-mediated platelet injury and resulting thrombocytopenia, thereby providing protection from lethal SA infection in a murine intravenous challenge model. Genetic deletion or pharmacological inhibition of AMR stabilized platelet counts and enhanced resistance to SA infection, and the anti-influenza sialidase inhibitor oseltamivir (Tamiflu) provided similar therapeutic benefit. Thus, a "toxin-platelet-AMR" regulatory pathway plays a critical role in the pathogenesis of SA bloodstream infection, and its elucidation provides proof of concept for repurposing two commonly prescribed drugs as adjunctive therapies to improve patient outcomes.
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http://dx.doi.org/10.1126/scitranslmed.abd6737DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC9121309PMC
March 2021

Driving to Safety: CRISPR-Based Genetic Approaches to Reducing Antibiotic Resistance.

Trends Genet 2021 08 19;37(8):745-757. Epub 2021 Mar 19.

Tata Institute for Genetics and Society, University of California, San Diego, 9500 Gilman Drive, La Jolla, CA 92093-0349, USA; Collaborative to Halt Antibiotic-Resistant Microbes, Department of Pediatrics, University of California, San Diego, 9500 Gilman Drive, La Jolla, CA 92093-0687, USA; Skaggs School of Pharmacy & Pharmaceutical Sciences, University of California, San Diego, 9500 Gilman Drive, La Jolla, CA 92093-0687, USA.

Bacterial resistance to antibiotics has reached critical levels, skyrocketing in hospitals and the environment and posing a major threat to global public health. The complex and challenging problem of reducing antibiotic resistance (AR) requires a network of both societal and science-based solutions to preserve the most lifesaving pharmaceutical intervention known to medicine. In addition to developing new classes of antibiotics, it is essential to safeguard the clinical efficacy of existing drugs. In this review, we examine the potential application of novel CRISPR-based genetic approaches to reducing AR in both environmental and clinical settings and prolonging the utility of vital antibiotics.
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http://dx.doi.org/10.1016/j.tig.2021.02.007DOI Listing
August 2021

The lytic polysaccharide monooxygenase CbpD promotes Pseudomonas aeruginosa virulence in systemic infection.

Nat Commun 2021 02 23;12(1):1230. Epub 2021 Feb 23.

Faculty of Chemistry, Biotechnology and Food Science, Norwegian University of Life Sciences (NMBU), Ås, Norway.

The recently discovered lytic polysaccharide monooxygenases (LPMOs), which cleave polysaccharides by oxidation, have been associated with bacterial virulence, but supporting functional data is scarce. Here we show that CbpD, the LPMO of Pseudomonas aeruginosa, is a chitin-oxidizing virulence factor that promotes survival of the bacterium in human blood. The catalytic activity of CbpD was promoted by azurin and pyocyanin, two redox-active virulence factors also secreted by P. aeruginosa. Homology modeling, molecular dynamics simulations, and small angle X-ray scattering indicated that CbpD is a monomeric tri-modular enzyme with flexible linkers. Deletion of cbpD rendered P. aeruginosa unable to establish a lethal systemic infection, associated with enhanced bacterial clearance in vivo. CbpD-dependent survival of the wild-type bacterium was not attributable to dampening of pro-inflammatory responses by CbpD ex vivo or in vivo. Rather, we found that CbpD attenuates the terminal complement cascade in human serum. Studies with an active site mutant of CbpD indicated that catalytic activity is crucial for virulence function. Finally, profiling of the bacterial and splenic proteomes showed that the lack of this single enzyme resulted in substantial re-organization of the bacterial and host proteomes. LPMOs similar to CbpD occur in other pathogens and may have similar immune evasive functions.
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http://dx.doi.org/10.1038/s41467-021-21473-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7902821PMC
February 2021

More than a Pore: Nonlytic Antimicrobial Functions of Complement and Bacterial Strategies for Evasion.

Microbiol Mol Biol Rev 2021 02 27;85(1). Epub 2021 Jan 27.

Division of Host-Microbe Systems and Therapeutics, Department of Pediatrics, UC San Diego, La Jolla, California, USA

The complement system is an evolutionarily ancient defense mechanism against foreign substances. Consisting of three proteolytic activation pathways, complement converges on a common effector cascade terminating in the formation of a lytic pore on the target surface. The classical and lectin pathways are initiated by pattern recognition molecules binding to specific ligands, while the alternative pathway is constitutively active at low levels in circulation. Complement-mediated killing is essential for defense against many Gram-negative bacterial pathogens, and genetic deficiencies in complement can render individuals highly susceptible to infection, for example, invasive meningococcal disease. In contrast, Gram-positive bacteria are inherently resistant to the direct bactericidal activity of complement due to their thick layer of cell wall peptidoglycan. However, complement also serves diverse roles in immune defense against all bacteria by flagging them for opsonization and killing by professional phagocytes, synergizing with neutrophils, modulating inflammatory responses, regulating T cell development, and cross talk with coagulation cascades. In this review, we discuss newly appreciated roles for complement beyond direct membrane lysis, incorporate nonlytic roles of complement into immunological paradigms of host-pathogen interactions, and identify bacterial strategies for complement evasion.
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http://dx.doi.org/10.1128/MMBR.00177-20DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8549852PMC
February 2021

Exploring the Impact of Ketodeoxynonulosonic Acid in Host-Pathogen Interactions Using Uptake and Surface Display by Nontypeable Haemophilus influenzae.

mBio 2021 01 19;12(1). Epub 2021 Jan 19.

Glycobiology Research and Training Center, University of California, San Diego, La Jolla, California, USA

Surface expression of the common vertebrate sialic acid (Sia) -acetylneuraminic acid (Neu5Ac) by commensal and pathogenic microbes appears structurally to represent "molecular mimicry" of host sialoglycans, facilitating multiple mechanisms of host immune evasion. In contrast, ketodeoxynonulosonic acid (Kdn) is a more ancestral Sia also present in prokaryotic glycoconjugates that are structurally quite distinct from vertebrate sialoglycans. We detected human antibodies against Kdn-terminated glycans, and sialoglycan microarray studies found these anti-Kdn antibodies to be directed against Kdn-sialoglycans structurally similar to those on human cell surface Neu5Ac-sialoglycans. Anti-Kdn-glycan antibodies appear during infancy in a pattern similar to those generated following incorporation of the nonhuman Sia -glycolylneuraminic acid (Neu5Gc) onto the surface of nontypeable (NTHi), a human commensal and opportunistic pathogen. NTHi grown in the presence of free Kdn took up and incorporated the Sia into its lipooligosaccharide (LOS). Surface display of the Kdn within NTHi LOS blunted several virulence attributes of the pathogen, including Neu5Ac-mediated resistance to complement and whole blood killing, complement C3 deposition, IgM binding, and engagement of Siglec-9. Upper airway administration of Kdn reduced NTHi infection in human-like null (Neu5Gc-deficient) mice that express a Neu5Ac-rich sialome. We propose a mechanism for the induction of anti-Kdn antibodies in humans, suggesting that Kdn could be a natural and/or therapeutic "Trojan horse" that impairs colonization and virulence phenotypes of free Neu5Ac-assimilating human pathogens. All cells in vertebrates are coated with a dense array of glycans often capped with sugars called sialic acids. Sialic acids have many functions, including serving as a signal for recognition of "self" cells by the immune system, thereby guiding an appropriate immune response against foreign "nonself" and/or damaged cells. Several pathogenic bacteria have evolved mechanisms to cloak themselves with sialic acids and evade immune responses. Here we explore a type of sialic acid called "Kdn" (ketodeoxynonulosonic acid) that has not received much attention in the past and compare and contrast how it interacts with the immune system. Our results show potential for the use of Kdn as a natural intervention against pathogenic bacteria that take up and coat themselves with external sialic acid from the environment.
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http://dx.doi.org/10.1128/mBio.03226-20DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7845648PMC
January 2021

Novel Models of Colonization and Disease Reveal Modest Contributions of M-Like (SCM) Protein.

Microorganisms 2021 Jan 16;9(1). Epub 2021 Jan 16.

Department of Molecular Virology and Microbiology, Baylor College of Medicine, 1 Baylor Plaza, Houston, TX 77030, USA.

is a common colonizing bacterium of the urogenital tract of cats and dogs that can also cause invasive disease in these animal populations and in humans. Although the virulence mechanisms of are not well-characterized, an M-like protein, SCM, has recently identified been as a potential virulence factor. SCM is a surface-associated protein that binds to host plasminogen and IgGs suggesting its possible importance in host-pathogen interactions. In this study, we developed in vitro and ex vivo blood component models and murine models of vaginal colonization, systemic infection, and dermal infection to compare the virulence potential of the zoonotic vaginal isolate G361 and its isogenic SCM-deficient mutant (G361∆). We found that while establishes vaginal colonization and causes invasive disease in vivo, the contribution of the SCM protein to virulence phenotypes in these models is modest. We conclude that SCM is dispensable for invasive disease in murine models and for resistance to human blood components ex vivo, but may contribute to mucosal persistence, highlighting a potential contribution to the recently appreciated genetic diversity of SCM across strains and hosts.
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http://dx.doi.org/10.3390/microorganisms9010183DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7829700PMC
January 2021

A Novel N4-Like Bacteriophage Isolated from a Wastewater Source in South India with Activity against Several Multidrug-Resistant Clinical Pseudomonas aeruginosa Isolates.

mSphere 2021 01 13;6(1). Epub 2021 Jan 13.

School of Biotechnology, Amrita Vishwa Vidyapeetham, Amritapuri, Kerala, India

Multidrug-resistant community-acquired infections caused by the opportunistic human pathogen are increasingly reported in India and other locations globally. Since this organism is ubiquitous in the environment, samples such as sewage and wastewater are rich reservoirs of bacteriophages. In this study, we report the isolation and characterization of a novel N4-like lytic bacteriophage, vB_Pae_AM.P2 (AM.P2), from wastewater in Kerala, India. AM.P2 is a double-stranded DNA podovirus that efficiently lyses the model strain, PAO1, at a multiplicity of infection as low as 0.1 phage per bacterium and resistance frequency of 6.59 × 10 Synergy in bactericidal activity was observed between AM.P2 and subinhibitory concentrations of the antibiotic ciprofloxacin. Genome sequencing of AM.P2 revealed features similar to those of the N4-like phages LUZ7 and KPP21. As judged by two independent assay methods, spot tests and growth inhibition, AM.P2 successfully inhibited the growth of almost 30% of strains from a contemporary collection of multidrug-resistant clinical isolates from South India. Thus, AM.P2 may represent an intriguing candidate for inclusion in bacteriophage cocktails developed for various applications, including water decontamination and clinical bacteriophage therapy. In India, multidrug resistance determinants are much more abundant in community-associated bacterial pathogens due to the improper treatment of domestic and industrial effluents. In particular, a high bacterial load of the opportunistic pathogen in sewage and water bodies in India is well documented. The isolation and characterization of bacteriophages that could target emerging strains, representing possible epicenters for community-acquired infections, could serve as a useful alternative tool for various applications, such as phage therapy and environmental treatment. Continuing to supplement the repertoire of broad-spectrum bacteriophages is an essential tool in confronting this problem.
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http://dx.doi.org/10.1128/mSphere.01215-20DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7845610PMC
January 2021

Identifying the effect of vancomycin on health care-associated methicillin-resistant Staphylococcus aureus strains using bacteriological and physiological media.

Gigascience 2021 01;10(1)

Department of Bioengineering, University of California, 9500 Gilman Dr, La Jolla, CA 92093, USA.

Background: The evolving antibiotic-resistant behavior of health care-associated methicillin-resistant Staphylococcus aureus (HA-MRSA) USA100 strains are of major concern. They are resistant to a broad class of antibiotics such as macrolides, aminoglycosides, fluoroquinolones, and many more.

Findings: The selection of appropriate antibiotic susceptibility examination media is very important. Thus, we use bacteriological (cation-adjusted Mueller-Hinton broth) as well as physiological (R10LB) media to determine the effect of vancomycin on USA100 strains. The study includes the profiling behavior of HA-MRSA USA100 D592 and D712 strains in the presence of vancomycin through various high-throughput assays. The US100 D592 and D712 strains were characterized at sub-inhibitory concentrations through growth curves, RNA sequencing, bacterial cytological profiling, and exo-metabolomics high throughput experiments.

Conclusions: The study reveals the vancomycin resistance behavior of HA-MRSA USA100 strains in dual media conditions using wide-ranging experiments.
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http://dx.doi.org/10.1093/gigascience/giaa156DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7794652PMC
January 2021

Elongated neutrophil-derived structures are blood-borne microparticles formed by rolling neutrophils during sepsis.

J Exp Med 2021 03;218(3)

La Jolla Institute for Immunology, La Jolla, CA.

Rolling neutrophils form tethers with submicron diameters. Here, we report that these tethers detach, forming elongated neutrophil-derived structures (ENDS) in the vessel lumen. We studied ENDS formation in mice and humans in vitro and in vivo. ENDS do not contain mitochondria, endoplasmic reticulum, or DNA, but are enriched for S100A8, S100A9, and 57 other proteins. Within hours of formation, ENDS round up, and some of them begin to present phosphatidylserine on their surface (detected by annexin-5 binding) and release S100A8-S100A9 complex, a damage-associated molecular pattern protein that is a known biomarker of neutrophilic inflammation. ENDS appear in blood plasma of mice upon induction of septic shock. Compared with healthy donors, ENDS are 10-100-fold elevated in blood plasma of septic patients. Unlike neutrophil-derived extracellular vesicles, most ENDS are negative for the tetraspanins CD9, CD63, and CD81. We conclude that ENDS are a new class of bloodborne submicron particles with a formation mechanism linked to neutrophil rolling on the vessel wall.
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http://dx.doi.org/10.1084/jem.20200551DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7721910PMC
March 2021
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