G protein-coupled estrogen receptor agonist G-1 decreases ADAM10 levels and NLRP3-inflammasome component activation in response to Staphylococcus aureus alpha-hemolysin.

The G protein-coupled estrogen receptor, also known as GPER1 or originally GPR30, is found in various tissues, indicating its diverse functions. It is typically present in immune cells, suggesting its role in regulating immune responses to infectious diseases. Our previous studies have shown that G-1, a selective GPER agonist, can limit the pathogenesis mediated by Staphylococcus aureus alpha-hemolysin (Hla).

Nonconformity of biofilm formation in vivo and in vitro based on Staphylococcus aureus accessory gene regulator status.

Staphylococcus aureus is an opportunistic, pathogenic bacteria that causes significant morbidity and mortality. As antibiotic resistance by S. aureus continues to be a serious concern, developing novel drug therapies to combat these infections is vital.

Quantitative Hemolysis Assays.

Many strains of Staphylococcus aureus produce a variety of cytolysins that target many different cell types to both fight the immune system and acquire nutrients. This includes hemolysins which destroy erythrocytes and are well studied virulence factors. Traditionally, hemolysin activity is measured on blood agar plates due to the simplicity of the assay.

Peptide-Conjugated Phosphorodiamidate Morpholino Oligomers Retain Activity against Multidrug-Resistant Pseudomonas aeruginosa and .

Most antimicrobials currently in the clinical pipeline are modifications of existing classes of antibiotics and are considered short-term solutions due to the emergence of resistance. represents a major challenge for new antimicrobial drug discovery due to its versatile lifestyle, ability to develop resistance to most antibiotic classes, and capacity to form robust biofilms on surfaces and in certain hosts such as those living with cystic fibrosis (CF). A precision antibiotic approach to treating could be achieved with an antisense method, specifically by using peptide-conjugated phosphorodiamidate morpholino oligomers (PPMOs).

Complement Receptor 3 Contributes to the Sexual Dimorphism in Neutrophil Killing of .

We previously reported sex differences in innate susceptibility to skin infection and that bone marrow neutrophils (BMN) from female mice have an enhanced ability to kill ex vivo compared with those of male mice. However, the mechanism(s) driving this sex bias in neutrophil killing have not been reported. Given the role of opsonins such as complement, as well as their receptors, in recognition and clearance, we investigated their contribution to the enhanced bactericidal capacity of female BMN.

Vaccination with VLPs Presenting a Linear Neutralizing Domain of Hla Elicits Protective Immunity.

The pore-forming cytotoxin α-hemolysin, or Hla, is a critical virulence factor that promotes infection by causing tissue damage, excessive inflammation, and lysis of both innate and adaptive immune cells, among other cellular targets. In this study, we asked whether a virus-like particle (VLP)-based vaccine targeting Hla could attenuate Hla-mediated pathogenesis. VLPs are versatile vaccine platforms that can be used to display target antigens in a multivalent array, typically resulting in the induction of high titer, long-lasting antibody responses.

Staphylococcus aureus Fatty Acid Kinase FakA Modulates Pathogenesis during Skin Infection via Proteases.

fatty acid kinase FakA is necessary for the incorporation of exogenous fatty acids into the lipid membrane. We previously demonstrated that the inactivation of leads to decreased α-hemolysin (Hla) production but increased expression of the proteases SspAB and aureolysin , and that the Δ mutant causes larger lesions than the wild type (WT) during murine skin infection. As expected, necrosis is Hla dependent in the presence or absence of FakA, as both and Δ mutants are unable to cause necrosis of the skin.

GPER activation protects against epithelial barrier disruption by Staphylococcus aureus α-toxin.

Sex bias in innate defense against Staphylococcus aureus skin and soft tissue infection (SSTI) is dependent on both estrogen production by the host and S. aureus secretion of the virulence factor, α-hemolysin (Hla). The impact of estrogen signaling on the immune system is most often studied in terms of the nuclear estrogen receptors ERα and ERβ.

Morpholino oligomers tested in vitro, in biofilm and in vivo against multidrug-resistant Klebsiella pneumoniae.

Background: Klebsiella pneumoniae is an opportunistic pathogen and many strains are multidrug resistant. KPC is one of the most problematic resistance mechanisms, as it confers resistance to most β-lactams, including carbapenems. A promising platform technology for treating infections caused by MDR pathogens is the nucleic acid-like synthetic oligomers that silence bacterial gene expression by an antisense mechanism.

Antisense Inhibitors Retain Activity in Pulmonary Models of Burkholderia Infection.

The Burkholderia cepacia complex is a group of Gram-negative bacteria that are opportunistic pathogens in immunocompromised individuals, such as those with cystic fibrosis (CF) or chronic granulomatous disease (CGD). Burkholderia are intrinsically resistant to many antibiotics and the lack of antibiotic development necessitates novel therapeutics. Peptide-conjugated phosphorodiamidate morpholino oligomers are antisense molecules that inhibit bacterial mRNA translation.

MCR-1 Inhibition with Peptide-Conjugated Phosphorodiamidate Morpholino Oligomers Restores Sensitivity to Polymyxin in .

In late 2015, the first example of a transferrable polymyxin resistance mechanism in Gram-negative pathogens, MCR-1, was reported. Since that report, MCR-1 has been described to occur in many Gram-negative pathogens, and the mechanism of MCR-1-mediated resistance was rapidly determined: an ethanolamine is attached to lipid A phosphate groups, rendering the membrane more electropositive and repelling positively charged polymyxins. Acquisition of MCR-1 is clinically significant because polymyxins are frequently last-line antibiotics used to treat extensively resistant organisms, so acquisition of this mechanism might lead to pan-resistant strains.

Synthetic Antibacterial Peptide Exhibits Synergy with Oxacillin against MRSA.

One proposed solution to the crisis of antimicrobial resistant (AMR) infections is the development of molecules that potentiate the activity of antibiotics for AMR bacteria, such as methicillin-resistant (MRSA). Rather than develop broad spectrum compounds, we developed a peptide that could potentiate the activity of a narrow spectrum antibiotic, oxacillin. In this way, the combination treatment could narrowly target the resistant pathogen and limit impact on host flora.

Employing high-frequency alternating magnetic fields for the non-invasive treatment of prosthetic joint infections.

Treatment of prosthetic joint infection (PJI) usually requires surgical replacement of the infected joint and weeks of antibiotic therapy, due to the formation of biofilm. We introduce a non-invasive method for thermal destruction of biofilm on metallic implants using high-frequency (>100 kHz) alternating magnetic fields (AMF). In vitro investigations demonstrate a >5-log reduction in bacterial counts after 5 minutes of AMF exposure.

VLP-based vaccine induces immune control of Staphylococcus aureus virulence regulation.

Staphylococcus aureus is the leading cause of skin and soft tissue infections (SSTIs) and mounting antibiotic resistance requires innovative treatment strategies. S. aureus uses secreted cyclic autoinducing peptides (AIPs) and the accessory gene regulator (agr) operon to coordinate expression of virulence factors required for invasive infection.

Inhibition of Bacterial Growth by Peptide-Conjugated Morpholino Oligomers.

Morpholino oligomers (MOs) are antisense molecules designed for sequence-specific binding of target mRNA. In bacteria, inhibition is hypothesized to occur by preventing translation initiation. Cell-penetrating peptides may be conjugated to the 5'- or 3'-termini of an MO to enhance cellular entry and therefore inhibition.

Peptide-conjugated phosphorodiamidate morpholino oligomer (PPMO) restores carbapenem susceptibility to NDM-1-positive pathogens in vitro and in vivo.

Objectives: The objective of this study was to test the efficacy of an inhibitor of the New Delhi metallo-β- lactamase (NDM-1). Inhibiting expression of this type of antibiotic-resistance gene has the potential to restore antibiotic susceptibility in all bacteria carrying the gene.

Methods: We have constructed a peptide-conjugated phosphorodiamidate morpholino oligomer (PPMO) that selectively inhibits the expression of NDM-1 and examined its ability to restore susceptibility to meropenem in vitro and in vivo .

ω-Hydroxyemodin limits staphylococcus aureus quorum sensing-mediated pathogenesis and inflammation.

Antibiotic-resistant pathogens are a global health threat. Small molecules that inhibit bacterial virulence have been suggested as alternatives or adjuncts to conventional antibiotics, as they may limit pathogenesis and increase bacterial susceptibility to host killing. Staphylococcus aureus is a major cause of invasive skin and soft tissue infections (SSTIs) in both the hospital and community settings, and it is also becoming increasingly antibiotic resistant.

Development of a mimotope vaccine targeting the Staphylococcus aureus quorum sensing pathway.

A major hurdle in vaccine development is the difficulty in identifying relevant target epitopes and then presenting them to the immune system in a context that mimics their native conformation. We have engineered novel virus-like-particle (VLP) technology that is able to display complex libraries of random peptide sequences on a surface-exposed loop in the coat protein without disruption of protein folding or VLP assembly. This technology allows us to use the same VLP particle for both affinity selection and immunization, integrating the power of epitope discovery and epitope mimicry of traditional phage display with the high immunogenicity of VLPs.

Identification of the Staphylococcus aureus vfrAB operon, a novel virulence factor regulatory locus.

During a screen of the Nebraska Transposon Mutant Library, we identified 71 mutations in the Staphylococcus aureus genome that altered hemolysis on blood agar medium. Although many of these mutations disrupted genes known to affect the production of alpha-hemolysin, two of them were associated with an apparent operon, designated vfrAB, that had not been characterized previously. Interestingly, a ΔvfrB mutant exhibited only minor effects on the transcription of the hla gene, encoding alpha-hemolysin, when grown in broth, as well as on RNAIII, a posttranscriptional regulatory RNA important for alpha-hemolysin translation, suggesting that VfrB may function at the posttranscriptional level.

Nox2 modification of LDL is essential for optimal apolipoprotein B-mediated control of agr type III Staphylococcus aureus quorum-sensing.

Staphylococcus aureus contains an autoinducing quorum-sensing system encoded within the agr operon that coordinates expression of virulence genes required for invasive infection. Allelic variation within agr has generated four agr specific groups, agr I-IV, each of which secretes a distinct autoinducing peptide pheromone (AIP1-4) that drives agr signaling. Because agr signaling mediates a phenotypic change in this pathogen from an adherent colonizing phenotype to one associated with considerable tissue injury and invasiveness, we postulated that a significant contribution to host defense against tissue damaging and invasive infections could be provided by innate immune mechanisms that antagonize agr signaling.

Unprecedented C-2 arylation of indole with diazonium salts: Syntheses of 2,3-disubstituted indoles and their antimicrobial activity.

A novel reaction of indole with aryldiazonium salts leading to the formation of 2-aryl-3-(arylazo)indoles was discovered. The products were found to possess potent anti-MRSA and anti-LLVRE activities. The SAR studies indicate that the potentially metabolically labile azo functionality can be replaced with ether oxygen and thioether sulfur atoms without any loss of activity.