Browsing by Author "Ramos-Hegazy, Layla"

Now showing 1 - 3 of 3
  • Thumbnail Image
    Item
    Biofilm and Virulence Regulation of the Cystic Fibrosis Associated Pathogens, Stenotrophomonas maltophilia and Pseudomonas aeruginosa
    (2020-05) Ramos-Hegazy, Layla; Anderson, Gregory; Perrin, Benjamin; Slayback-Barry, Denise
    Cystic fibrosis (CF) is a fatal, incurable genetic disease that affects over 30,000 people in the United States alone. People with this disease have a homozygous mutation in the cystic fibrosis transmembrane conductance regulator (CFTR) which causes defects in chloride transport and leads to build up of mucus in the lungs and disruption of function in various organs. CF patients often suffer from chronic bacterial infections within the lungs, wherein the bacteria persist as a biofilm, leading to poor prognosis. Two of these pathogens, Stenotrophomonas maltophilia and Pseudomonas aeruginosa, are often found in the lungs of patients with CF and are an increasing medical concerns due to their intrinsic antimicrobial resistance. Both species can readily form biofilms on biotic and abiotic surfaces such as intravascular devices, glass, plastic, and host tissue. Biofilm formation starts with bacterial attachment to a surface and/or adjacent cells, initiating the acute infection stage. Chronic, long-term infection involves subsequent or concurrent altered genetic regulation, including a downregulation of virulence factors, resulting in the bacteria committing to a sessile lifestyle, markedly different from the planktonic one. Many of these genetic switches from an acute to chronic lifestyle are due to pressures from the host immune system and lead to permanently mutated strains, most likely an adaptive strategy to evade host immune responses. Biofilms are extremely problematic in a clinical setting because they lead to nosocomial infections and persist inside the host causing long-term chronic infections due to their heightened tolerance to almost all antibiotics. Understanding the genetic networks governing biofilm initiation and maintenance would greatly reduce consequences for CF and other biofilm-related infections and could lead to the development of treatments and cures for affected patients. This study showed that in S. maltophilia, isogenic deletion of phosphoglycerate mutase (gpmA) and two chaperone-usher pilin subunits, S. maltophilia fimbrae-1 (smf-1) and cblA, lead to defects in attachment on abiotic surfaces and cystic fibrosis derived bronchial epithelial cells (CFBE). Furthermore, Δsmf-1 and ΔcblA showed defects in long-term biofilm formation, mimicking that of a chronic infection lifestyle, on abiotic surfaces and CFBE as well as stimulating less of an immune response through TNF-α production. This study also showed that in P. aeruginosa, the Type III secretion system (T3SS), an important virulence factor activated during the acute stage of infection, is downregulated when polB, a stress-induced alternate DNA polymerase, is overexpressed. This downregulation is due to post-transcriptional inhibition of the master regulatory protein, ExsA. Taken together, this project highlights important genes involved in the acute and chronic infection lifestyle and biofilm formation in S. maltophilia and genetic switches during the acute infection lifestyle in P. aeruginosa.
  • Thumbnail Image
    Item
    DNA alternate polymerase PolB mediates inhibition of type III secretion in Pseudomonas aeruginosa
    (Elsevier, 2021) Chakravarty, Shubham; Ramos-Hegazy, Layla; Gasparovic, Abigail; Anderson, Gregory G.; Biology, School of Science
    Opportunistic pathogen Pseudomonas aeruginosa uses a variety of virulence factors to cause acute and chronic infections. We previously found that alternate DNA polymerase gene polB inhibits P. aeruginosa pyocyanin production. We investigated whether polB also affects T3SS expression. polB overexpression significantly reduced T3SS transcription and repressed translation of the master T3SS regulator ExsA, while not affecting exsA mRNA transcript abundance. Further, polB does not act through previously described genetic pathways that post-transcriptionally regulate ExsA. Our results show a novel T3SS regulatory component which may lead to development of future drugs to target this mechanism.
  • Thumbnail Image
    Item
    Phosphoglycerate mutase affects Stenotrophomonas maltophilia attachment to biotic and abiotic surfaces
    (Elsevier, 2020) Ramos-Hegazy, Layla; Chakravarty, Shubham; Anderson, Gregory G.; Biology, School of Science
    Stenotrophomonas maltophilia biofilm formation is of increasing medical concern, particularly for lung infections. However, the molecular mechanisms facilitating the biofilm lifestyle in S. maltophilia are poorly understood. We generated and screened a transposon mutant library for mutations that lead to altered biofilm formation compared to wild type. One of these mutations, in the gene for glycolytic enzyme phosphoglycerate mutase (gpmA), resulted in impaired attachment on abiotic and biotic surfaces. As adherence to a surface is the initial step in biofilm developmental processes, our results reveal a unique factor that could affect S. maltophilia biofilm initiation and, possibly, subsequent development.