Microbiology / Infectious Disease

Permanent URI for this collectionhttps://hdl.handle.net/20.500.12503/21740

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    Next-generation sequencing and cytokine analysis of bronchoalveolar lavage samples from mechanically ventilated trauma patients
    (2015-03) Smith, Ashley D.; Zhang, Yan; Huebinger, Ryan M.; Ireland, Sara J.; Monson, Nancy L.; Allen, Michael
    Purpose: Mechanically ventilated trauma patients are at high risk for ventilator-associated pneumonia (VAP). VAP diagnosis relies on several clinical factors including the identification of a specific pathogen by culture-dependent techniques. Often, patients exhibit signs of VAP, yet the hospital lab is unable to culture a pathogen from a bronchoalveolar lavage (BAL) sample. We hypothesize that these culture-negative, presumptive positive patients are infected with potentially pathogenic bacteria that are not detected by traditional culture techniques. Methods: Culture-positive and -negative hospital results were tested again by Sanger and next-generation sequencing (NGS) on the Ion Torrent PGM. Sample cytokine levels were determined using a Bio-Plex Pro Human Th17 cytokine panel. Results: No significant difference was seen between the identification methods in culture-positive BAL. However, NGS analysis of culture-negative BAL identified hundreds of bacterial genera, including a group of patients that exhibited similar bacterial composition, diversity, and abundance. This group was significantly different from other culture-negative BAL that were dominated by one or two suspected pathogens. Culture-negative BAL contained significantly less IL-1β, IL-6 and TNF-α than culture-positive BAL. Conclusions: NGS is a valuable method for pathogen identification, particularly for difficult to culture BAL. Culture-positive BAL exhibit less bacterial diversity and increased cytokine production than culture-negative BAL. The grouping of culture-negative BAL with similar characteristics may denote a core lung microbiome.
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    Clostridium difficile Ribotype 027 Virulence Phenotype Influences Disease Severity when Compared to Ribotype Non-027 Strains in the Hamster CDAD Model
    (2015-03) Vitucci, John C.; Pulse, Mark; Orlowski, Ashley; Simecka, Jerry
    Introduction: C. difficile ribotype 027 (RT027) is the epidemic strain found primarily in North America, and, recently, a strain of C. difficile has been named a superbug by the CDC. Studies have suggested an enhanced virulence phenotype for RT027 such as increased toxin production, but the impact on disease severity on in vivo models is not well understood. This study describes the in vitro characterization of important virulence factors for several RT027 and non-RT027 C. difficile clinical isolates, and how these characteristics may impact disease severity in the hamster C. difficile associated disease (HCDAD) model. Materials and Methods: Six RT027 and six non-RT027 clinical isolates were evaluated in vitro for total spore counts and Toxin A/B titers in 72H broth cultures. Spore counts were generated from heat/ethanol shock culture samples and plated onto CCFA containing 0.1% taurocholate, and toxin A/B titers were determined from spent broth with the tgcBIOMICS ELISA assay. The HCDAD studies involved infecting male Golden Syrian hamsters with 72H broth cultures of two RT027 and two non-RT027 isolates, followed by subcutaneous administration of 10 mg/kg clindamycin 24H post-infection. One group of infected hamsters was orally treated with 20 mg/kg vancomycin once a day for 3 days following clindamycin administration, while the other group remained untreated. Survival was monitored for 11 days after infection and 3 hamsters were euthanized at set time points to determine cecal fluid associated the CFU/spore counts and Toxin A/B titers. Results: The RT027 and the non-RT027 strains generated similar mean CFU/mL in 72H broth cultures, while the mean spore counts were 548 spores/mL for the RT027 strains and 273 spores/mL for the non-RT027 strains. In addition, the 72H broth-associated mean toxin A/B titers were 2.8-fold higher for RT027 strains when compared to the 72H titers of non-RT027 strains. In the HCDAD studies 14% of the non-027 infected hamsters became moribund, while 71% of the hamsters infected with the RT027 isolates became moribund. The mean cecal fluid Toxin A/B titers for RT027 infected hamsters were 2.3 to 9-fold higher than the titers for non-RT027 infected hamsters. Conclusion: The results highlight that C. difficile RT027 isolates, when compared to non-RT027 clinical isolates, have enhanced virulence that corresponds to a more severe disease in the HCDAD model. Understanding why these strains have this phenotype, in vivo as compared to in vitro, is important for treatment as new ribotypes, with varying virulence, are continuously emerging.
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    Methods to determine the true rate of side effects caused by medications: A case study with antiretroviral medications and diarrhea.
    (2015-03) Bavafa, Azadeh; Killam-Worrall, Lisa; Clay, Patrick G.
    Introduction: There have been discrepancies between the rates of adverse events reported in the medication’s package insert and what have been seen in clinical settings. This difference causes confusion for clinicians and patients and interferes with pharmacists’ attempts to establish the necessary rapport with patient to improve compliance. Hypothesis: The incidence of selected adverse event for a class of medication would not be same as those found the package insert. Goals: The goal of this project is to conduct a systematic review of current literature to identify the true rate of adverse events associated with common Antiretroviral (ARV) medication regimens. Methods: A systematic search of primary literatures was conducted to evaluate the incidence rates of selected adverse events associated with most customary medication regimens recommended by HIV(Human Immunodeficiency Virus)/AIDS (Acquired Immune Deficiency Syndrome) medical practice guidelines. Initially, Inclusion and exclusion criteria were established. The keywords collected were used to search the primary literatures and documents produced during the drug approval process. Finally, the gathered data was tabulated and analyzed to determine “true” rate of adverse events. Results: The incidence rates of diarrhea among HIV infected—treatment Naïve—males and non-pregnant females ages: 18-65, were extracted from various sources including: the medication’s package insert, pertaining briefing document, clinical trials submitted to FDA, and published—unpublished phase III clinical trials. The medication regimens under investigation included: efavirenz (EFV) / emtricitabine (FTC)/ tenofovir( TDF) (Atripla), raltegravir(RAL)/FTC/ TDF, and rilpivirine(RPV)/ FTC/ TDF(Complera). The values reported, varied from one source to another. For example, the rate reported in Atripla’s package insert was 9%, and yet nothing was documented in its briefing document. In addition, the average rates (all causes and grades) extracted from submitted clinical trials—published and unpublished phase III clinical trials, were respectively: 22.9% (10.14%-35.6%) and 13.43% (0%-33.9%). The same method was used for the other two medication regimens, and similar trends were observed. Conclusion: Preliminary results of case study show the rate of diarrhea reported in package insert was different from what was extrapolated from published and unpublished phase III trials. Further analysis will be conducted to confirm this finding.