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- All Subjects: Staphylococcus aureus
- Creators: Reyes del Valle, Jorge
Description
The viscous lung mucus of cystic fibrosis (CF) patients is characterized by oxygen gradients, which creates a unique niche for bacterial growth. Pseudomonas aeruginosa and Staphylococcus aureus, two predominant microorganisms chronically infecting the airways of CF patients, typically localize in hypoxic regions of the mucus. While interspecies interactions between P. aeruginosa and S. aureus have been reported, little is known about the role of low oxygen in regulating these interactions. Studying interspecies interactions in CF lung disease is important as evidence suggests that microbial community composition governs disease progression. In this study, P. aeruginosa lab strain PAO1 and two primary clinical isolates from hypoxic tissues were cultured alone, or in combination, with methicillin resistant S. aureus (MRSA) strain N315 under hypoxic or normoxic conditions. Herein, it is shown for the first time that low oxygen conditions relevant to the CF lung affect the competitive behavior between P. aeruginosa and S. aureus. Specifically, S. aureus was able to better survive competition in hypoxic versus normoxic conditions. Competition data from different oxygen concentrations were consistent using PAO1 and clinical isolates even though differences in the level of competition were observed. PAO1 strains carrying mutations in virulence factors known to contribute to S. aureus competition (pyocyanin/phzS, elastase/lasA and lasI quorum sensing/lasI) were used to determine which genes play a role in the differential growth inhibition. The lasA and lasI mutants competed less effectively with S. aureus regardless of the oxygen level present in the culture compared to the isogenic wild type strain. These results are consistent with previous findings that elastase and lasI quorum sensing play a role in competitive behavior of P. aeruginosa and S. aureus. Interestingly, the phzS mutant competed less effectively in hypoxic conditions suggesting that pyocyanin may be important in microaerophilic conditions. This study demonstrates that oxygen plays a role in competition between P. aeruginosa and S. aureus and contributes to understanding CF environmental factors that may regulate microbial community dynamics important for disease progression with potential for development of therapeutic avenues.
ContributorsLedesma Barrera, Maria Alexandra (Author) / Nickerson, Cheryl A. (Thesis advisor) / Reyes del Valle, Jorge (Committee member) / Clark-Curtiss, Josephine (Committee member) / Stout, Valerie (Committee member) / Ott, C M (Committee member) / Arizona State University (Publisher)
Created2014
Description
Dengue virus infects millions of people every year. Yet there is still no vaccine available to prevent it. Here we use a neutralizing epitope determinant on the dengue envelope (E) protein as an immunogen to be vectored by a measles virus (MV) vaccine. However the domain III (DIII) of the dengue 2 E protein is too small to be immunogenic by itself. In order for it to be displayed on a larger particle, it was inserted into the amino terminus of small hepatitis B surface antigen (HBsAg, S) coding sequence. To generate the recombinant MV vector and verify the efficiency of this concept, a reverse genetics system was used where the MV vectors express one or two additional transcription units to direct the assembly of hybrid HBsAg particles. Two types of recombinant measles virus were produced: pB(+)MVvac2(DIII-S,S)P and pB(+)MVvac2(DIII-S)N. Virus recovered from pB(+)MVvac2(DIII-S,S)P was viable. An ELISA assay was performed to demonstrate the expression and secretion of HBsAg. Supernatant from MVvac2(DIII-S,S)P infected cells confirmed that hybrid HBsAg-domain III particles with a density similar to traditional HBsAg particles were released. Characteristics of the subviral particle have been analyzed for the successful incorporation of domain III. The replication fitness of the recombinant MV was evaluated using multi-step growth kinetics and showed reduced replication fitness when compared to the parental strain MVvac2. This demonstrates that viral replication is hindered by the addition of the two inserts into MV genome. Further analysis of MVvac2(DIII-S)N is needed to justify immune response studies in a small animal model using both of the generated recombinant vectors.
ContributorsHarahap, Indira Saridewi (Author) / Reyes del Valle, Jorge (Thesis director) / Hogue, Brenda (Committee member) / Misra, Rajeev (Committee member) / Barrett, The Honors College (Contributor) / T. Denny Sanford School of Social and Family Dynamics (Contributor) / School of Human Evolution and Social Change (Contributor) / School of Life Sciences (Contributor)
Created2014-05