Biology subjects

Urish, K. L.

Publications and source records attributed to Urish, K. L..

3 recordsLinked to original sources

bioRxiv2025

Methicillin-Resistant Staphylococcus aureus has Phenotypic Variation in mecA Expression that Alters Antibiotic Sensitivity

Methicillin resistant Staphylococcus aureus (MRSA) sepsis has a high rate of morbidity and mortality. Multiple clinical studies have demonstrated improved outcomes when MRSA sepsis is treated with dual antibiotic therapy that includes a {beta}-lactam antibiotic such as cefazolin. This is a paradox as MRSA should be inherently resistant to this class of antibiotics. We report a serendipitous observation revealed a phenotype where MRSA became sensitive to cefazolin when cultured in a physiologic relevant media of fetal bovine serum as well as in synovial fluid. This could be observed across multiple clinical isolates. Expected resistance was maintained when cultured in Muller Hinton Broth (MHB). MRSA {beta}-lactam antibiotic resistance is mediated by PBP2a, a penicillin-binding protein encoded by mecA. We hypothesized that this phenotype of antibiotic sensitivity in physiologic medium was based, in part, on levels of PBP2a expression and post-translational modifications of peptidoglycan wall teichoic acid (WTA). We therefore conducted quantitative RT-PCR analysis and Western blotting which demonstrated limited mecA expression in the mRNA level and limited PBP2a protein level when cultured in FBS or synovial fluid as compared to the clinical microbiology standard MHB, respectively. Whole genome sequencing of loss of function mutants generated through serial passaging in FBS revealed that the clp family of proteins and rpo genes were involved in {beta}-lactam resistance. Cell wall peptidoglycan analysis suggested that WTA glycosylation was altered between {beta}-lactam resistant and sensitive MRSA phenotypes. Together, this suggests pathways for clpP, rpoB, and WTA glycosylation can be new potential targets for MRSA treatment. IMPORTANCEThis study demonstrates that MRSA has phenotypic variation in mecA expression, PBP2a protein levels, and patterns of wall teichoic acid glycosylation based on environmental conditions which can result in a phenotype of {beta}-lactam antibiotic sensitivity. The presence of an antibiotic resistance gene does not necessarily result in antibiotic resistance. This provides a possible explanation for the clinical observations of potential therapeutic benefit of {beta}-lactam antibiotics in dual therapy treatment of MRSA sepsis and suggests new therapeutic strategies to combat multidrug resistant bacteria. More importantly, identifying genes potentially responsible for this phenotype offers new potential therapeutic targets for the treatment of multidrug resistant infection.

microbiology↗

bioRxiv2020

Genotypic diversity between surgical and nasal Staphylococcus aureus isolates

Staphylococcus aureus is a common organism in periprosthetic joint infection (PJI). Little is known about S. aureus genetic diversity in PJI as compared to nasal carriage. We hypothesized PJI S. aureus strains would be associated with increased virulence as compared to those from nasal carriage. Whole genome sequencing and multilocus sequence typing (MLST) was performed to genotype these two populations at high resolution. MLST revealed a variety of genotypes in both populations but many belonged to the most common clonal complexes. In nasal cultures, 69% of strains were of clonal complexes CC5, CC8, and CC30. In PJI cultures, only 51% could be classified in these common clonal complexes. Remaining strains were atypical, and these atypical strains in PJI were associated with poor host status and compromised immune conditions. Mutations in genes involved in fibronectin binding (ebh, fnbA, clfA, clfB) systematically distinguished later PJI isolates from the first PJI isolate from each patient. S. aureus isolated from nasal carriage and PJI specimens differ significantly, with the latter being more diverse. Strains associated with lower pathogenicity tended to be found in immunocompromised patients, suggesting the host immune system plays an important role in preventing PJI. Repeated mutations in S. aureus genes associated with extracellular matrix binding were identified suggesting an adaptive, parallel evolution in S. aureus during the development of PJI.

microbiology↗

bioRxiv2019

The Toxin Antitoxin MazEF Drives Staphylococcus aureus Chronic Infection

Staphylococcus aureus is the major organism responsible for surgical implant infections. Antimicrobial treatment of these infections often fails leading to expensive surgical intervention and increased risk of mortality to the patient. The challenge in treating these infections is associated with the high tolerance of S. aureus biofilm to antibiotics. MazEF, a toxin-antitoxin system, is thought to be an important regulator of this phenotype, but its physiological function in S. aureus is controversial. Here, we examined the role of MazEF in developing chronic infections by comparing growth and antibiotic tolerance phenotypes in three S. aureus strains to their corresponding strains with disruption of mazF expression. Strains lacking mazF production showed increased biofilm growth, and decreased biofilm antibiotic tolerance. Deletion of icaADBC in the mazF::tn background suppressed the growth phenotype observed with mazF-disrupted strains, suggesting the phenotype was ica-dependent. We confirmed these phenotypes in our murine animal model. Loss of mazF resulted in increased bacterial burden and decreased survival rate compared to its wild-type strain demonstrating that loss of the mazF gene caused an increase in S. aureus virulence. Although lack of mazF gene expression increased S. aureus virulence, it was more susceptible to antibiotics in vivo. Combined, the ability of mazF to inhibit biofilm formation and promote biofilm antibiotic tolerance plays a critical role in transitioning from an acute to chronic infection that is difficult to eradicate with antibiotics alone.\n\nImportanceSurgical infections are one of the most common types of infections obtained in a hospital. Staphylococcus aureus is the most common pathogen associated with this infection. These infections are resilient and difficult to eradicate as the bacteria form a biofilm, a community of bacteria held together by an extracellular matrix. Compared to bacteria floating in liquid, bacteria in a biofilm are more resistant to antibiotics. The mechanism behind how bacteria develop this resistance and establish a chronic infection is unknown. We demonstrate that mazEF, a toxin-antitoxin gene, inhibits biofilm formation and promotes biofilm antibiotic tolerance which allows S. aureus to transition from an acute to chronic infection that cannot be eradicated with antibiotics but is less virulent. This gene not only makes the bacteria more tolerant to antibiotics but makes the bacteria more tolerant to the host.

microbiology↗