Browsing by Subject "Antimicrobial Peptides"

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    The Haemophilus ducreyi SAP Transporter Contributes to Antimicrobial Peptide Resistance
    (2009-09-30T19:56:19Z) Mount, Kristy Lee Beavers; Margaret E. Bauer, PhD
    Haemophilus ducreyi is the causative agent of the genital ulcer disease chancroid, which has been shown to facilitate the transmission of HIV. H. ducreyi is likely exposed to multiple sources of antimicrobial peptides in vivo. APs are small, cationic molecules with both bactericidal and immunomodulatory functions. Because H. ducreyi is able to establish and maintain an infection in an environment rich with antimicrobial peptides, we hypothesized that the bacterium was resistant to the bactericidal effects of these peptides. Using a 96-well AP bactericidal assay, we examined H. ducreyi susceptibility to eight human APs likely to be encountered at the site of infection, including the α-defensins human neutrophil peptide-1, human neutrophil peptide-2, human neutrophil peptide-3, and human defensin 5, the β-defensins human β defensin-2, human beta defensin-3, and human beta defensin-4, and the human cathelicidin, LL-37. H. ducreyi survival was compared to the survival of Escherichia coli ML35, a strain known to be susceptible to several antimicrobial peptides. H. ducreyi was significantly more resistant than E. coli ML35 to the bactericidal effects of all peptides tested. Furthermore, we found that representative class I and class II strains of H. ducreyi were each resistant to APs of each functional category, indicating that resistance to antimicrobial peptides could represent a conserved method of pathogenesis for H. ducreyi as a species. The H. ducreyi genome contains a homolog for the Sap influx transporter. To study the role of the H. ducreyi Sap transporter in AP resistance, we generated an isogenic sapA mutant and used the 96-well AP bactericidal assay to compare the AP susceptibility profiles of wild-type H. ducreyi, the sapA mutant and the sapA trans-complement to α-defensins, β-defensins, and LL-37. We observed a 25% decrease in the survival of the sapA mutant when it was exposed to LL-37. These findings suggest that the H. ducreyi Sap transporter plays a role in H. ducreyi resistance to LL-37, but it is likely that other AP resistance mechanisms co-exist within the bacterium.
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    Innate Immune Roles of Alpha-Defensin 1-3 in Neutralizing Uropathogenic Escherichia Coli
    (2024-05) Canas Kouaifati, Jorge Jose; Hains, David S.; Kaplan, Mark H.; Nelson, David E.; Dent, Alexander L.; Eadon, Michael T.
    Urinary tract infections (UTIs) are characterized by microbial colonization of the bladder, ureters or/and kidneys. Host-pathogen interactions compound to drive UTI susceptibility. The host innate immune system is composed of physio-chemical barriers and broad-spectrum potent effector responses that prevent pathogenic host colonization and bystander damage. Uropathogenic Escherichia coli (UPEC) isolates account for the majority of reported UTI cases. Virulence factors that are expressed by CFT073/UPEC heighten pathogenesis by permitting preferential invasion towards kidneys (pyelonephritis). During the acute invasion of the urinary tract tissues, pathogen molecular patterns and host-damage signals represent triggers that lead to host-defense responses. The release of antimicrobial peptides (AMPs) prevents microbial attachment to the epithelium. α-Defensin 1-3 are host-defense AMPs that work by eliciting microbial membrane interactions and inducing immunomodulatory effects. In humans, the DEFA1A3 locus encodes for three α-Defensin peptides (1-3). Chromosome 8 can harbor copy number polymorphism of the DEFA1A3 locus that range from 3-16 copies per diploid genome. Low copy numbers of DEFA1A3 have been associated with increased UTI risk in children (< 5 copies). On the other hand, patients with > 8 per copies per diploid genome have improved antibiotic therapy outcomes. In this thesis, we establish a pipeline to characterize α-Defensin 1-3 mechanisms of action, dissect contributors of expression at the cellular level, and protective DEFA1A3 gene-dose-dependent effects in the pyelonephritis setting. Using a manipulable mouse model expressing transgenic human DEFA1A3 gene copies, we explored dynamics of inducible α-Defensin 1-3 expression in the UPEC-infected kidney. Additionally, in vitro combinations of α-Defensin 1-3 and other host-defense AMPs work in concert to drive diverse cooperative action effects against UPEC. Methods and findings from my research in this thesis improve the current biomedical approaches to study AMP functions. Collectively, my results expand the understanding of DEFA1A3 polymorphic locus as a stratification UTI biomarker and exploration for the pre-clinical evaluation of kidney α-Defensin 1-3 expression as a potential therapeutic target for UTIs and other infectious diseases.