Browsing by Subject "Bacteria"
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Item A Novel sRNA Member of the Carbon Storage Regulatory System of Escherichia Coli(2002-12-01) Weilbacher, Thomas; Jerry SimeckaWeilbacher, Thomas S., A Novel sRNA Member of the Carbon Storage Regulatory System of Escherichi coli. Master of Science (Microbiology & Immunology), December, 2002, 57 pp., 2 tables, 12 illustrations, bibliography, 44 titles. Small untranslated RNAs (sRNAs) perform a variety of important functions in bacterial systems. The 245 nt sRNA of Escherichia coli K-12, CsrC, was uncovered using a genetic screen for genes that regulate glycogen biosynthesis. CsrC RNA binds multiple copies of CsrA, a protein that post-transcriptionally regulates central carbon flux, biofilm formation, and motility in E. coli. CsrC antagonizes the regulatory effects of CsrA, presumably by sequestering this protein. The discovery of CsrC is intriguing, in that a similar sRNA, CsrB, performs essentially the same function. Both of these sRNAs possess similar imperfect repeat sequences (18 in CsrB, 9 in CsrC), primarily localized in the loops of predicted hairpins, which may serve as CsrA binding elements. Transcription of csrC increases as the culture approaches the stationary phase of growth and is activated by CsrA and the response regulator UvrY. Complementation and in vitro transcription-translation experiments reveal that CsrA effects on csrC are mediated indirectly, through UvrY. Because CsrB and CsrC antagonize the activity of CsrA and are dependent on CsrA for their synthesis, a csrB null mutation causes a modest compensatory increase in CsrC levels and vice versa. An updated model for the signaling circuitry of the Csr system is discussed.Item A Study of the Effectiveness and Tolerability of Weekly Rifapentine/Isoniazid for Three Months Versus Daily Isoniazid for Nine Months for the Treatment of Latent Tuberculosis Infection(2004-11-01) Lemp, Jessie; Patricia Gwirtz; Walter McConathy; Richard EasomLemp, Jessie M. A Study of the Effectiveness and Tolerability of Weekly Rifapentine/Isoniazid for Three Months Versus Daily Isoniazid for Nine Months for the Treatment of Latent Tuberculosis Infection. Master of Science, November, 2004, 107 pp., 4 tables, 4 figures, references, 29 titles. The standard treatment for latent tuberculosis infection, nine months of daily isoniazid, is effective at preventing active tuberculosis; however, its full benefits are limited by non-adherence. A shorter intermittent regimen of rifapentine plus isoniazid once weekly for three months is equally effective as the standard regimen in animal models. This regimen facilitates the use of directly observed therapy, a method that significantly improves adherence. The Center for Disease Control is sponsoring Study 26 to test the effectiveness and tolerability the three-month rifapentine based regimen in latently infected persons with risk factors for progression to active tuberculosis. This thesis will describe the background rationale and methods for the clinical trial, and the internship experience.Item Assessment and Identification of Areas for Improvement of a Local Health Department Food Safety Program(2008-05-01) Harris, Ann MarieHarris, Ann Marie. Assessment and Identification of Areas for Improvement of a Local Health Department Food Safety Program. Master of Public Health (Environmental Health), May 2008, 14 pp. 1 table, 1 figure, references, 14 titles. The Fort Worth Public Health Department (FWPHD) established a standardized assessment to compare compliance rates for risk factors contributing to foodborne illness. The FWPHD identified significantly higher compliance rates in four out of six risk factors. Risk factors posing the greatest risk for out of compliance observations included threats from contaminated equipment and chemical/other hazards. Fast food establishments had a significantly greater risk for contaminated equipment (OR=1.81; CI=1.27, 2.58). Chemical/other hazards was the only risk factor with a higher overall out of compliance rate than the FDA. The FWPHD can now accurately track the effectiveness of training and education programs for food handlers, consumer health specialists, and the overall inspection process.Item Creating novel purification and biochemical characterization protocols for C. collagenase from Clostridium histolyticum, developing a new emergency medicine product, and formulating several novel therapies for chronic and acute wound treatment(2017-05-01) Mars, Jason P.; Jerry W. Simecka; Patricia A. Gwirtz; Aleksa JovanovicThe pharmaceutical industry not only includes infinite areas of specialization, but also consists of distinct areas that do not typically overlap. Biotechnology is the branch of medicinal research that bridges the gap between the fields within the pharmaceutical industry by being able to take on the challenges that require knowledge of a vast range of information. This practicum was organized to put the scientific knowledge and the interdisciplinary practices of biotechnology to use in a modern day, pharmaceutical company specializing in wound therapy and skincare: Smith & Nephew Biotherapeutics. Wound therapy has the widest range of application due to being one of the few fields that affects everyone, regardless of medical disposition. The specific goals of this practicum were: to develop novel purification and biochemical characterization protocols for C. collagenase from Clostridium histolyticum to replace current production methods of Santyl®, to develop a working prototype of a venom-based, hemostatic film, and perform reformulation, quality control, troubleshooting, and verification testing on samples of Regranex®, Iodosorb “Max”, and EU-Collagenase. Every goal presented was approached with the end results of saving Smith & Nephew costs, reducing bioburden of production, and creating more efficient protocols to bring Smith & Nephew into the modern age.Item Effects of Serine Protease-Like F (SPLF) on Alpha-Toxin Expression in Staphylococcus aureus(2003-05-01) Pulse, Mark E.; Mart Hart; Jerry Simecka; Ming-Chi WuPulse, Mark E., Effects of Serine protease-like F (Sp1F) on Alpha-Toxin Expression in Staphylococcus aureus. Master’s of Science (Microbiology). May 2003. Pages-67. Table-5. Figures-9. Transposon mutagenesis (Tn551) was used to generate agr-suppressor mutations in the agr-null Staphylococcus aureus strain RN6911 (Δagr;;tmn). Firty-four suppressor mutants displaying changes in hemolysin, protease, and lipase activities were isolated, and only twenty-six mutants contained Tn551 within their chromosomes. Transposon insertion sites for seven mutants were determined by sequencing amplicons generated by arbitrary-PCR. One of the insertion sites was within the serine protease-like F (spʅF) gene. Alpha-toxin message levels for the spʅF mutant were similar to RN6911. However, alpha-toxin activity in spent media isolated from the spʅF mutant were similar to RN6911. However, alpha-toxin activity in spent media isolated from the spʅF mutant was increased ten-fold as compared to RN6911. Transduction of the spʅf:Tn551 mutation back into the parental strain verified the link between the phenotype and the mutation. Whole cell lysates from Escherichia coli cells containing a plasmid copy of spʅF displayed protease activity on casein. These data suggest that SpʅF may be post-translationally modifying alpha-toxin through proteolysis.Item Effects of the Pleiotropic Gene csrA on Glycogen Metabolism in Escherichia Coli(1995-06-01) Yang, Honghui; Ming-Chi Wu; Wayne NicholsonYang, Honghui, Effects of the pleiotropic gene csrA on glycogen metabolism in Escherichia coli. Master of Science (Biomedical Sciences), June, 1995, 78 pp., 3 tables, 17 illustrations, bibliography, 59 titles. The csrA gene negatively regulates the expression of four genes glgB, glgC, glgA and glgS involved in glycogen synthesis. It also negatively regulates glgY, which encodes the enzyme glycogen phoshorylase involved in glycogen degradation, but no effect was observed on the glycogen debranching enzyme in this pathway. In addition, csrA exhibits a positive effect on the glycolytic enzyme triosephosphate isomerase. No significant effects were observed on the expression of two genes (zwf & gnd) participating in the pentose phosphate pathway. In vitro expression of glgB, glgC and glgA was specifically inhibited by cell extracts containing the csrA gene product (CsrA). This study provides evidence that csrA encodes an important regulator of intermediary carbon metabolism in Escherichia coli.Item Epidemiologic Assessment of a Targeted Tuberulosis Screening and Treatment Program Based on Geographic and Molecular Clustering(2005-08-01) Moonan, Patrick KevinMoonan, Patrick K., Epidemiologic Assessment of a Targeted Tuberculosis Screening and Treatment Program Based on Geographic and Molecular Clustering. Doctor of Public Health (Disease Prevention and Control), August 2005, 93 pp., 12 tables, 7 illustrations, bibliography, 148 tables. One of the primary goals of the tuberculosis elimination strategy is to interrupt the transmission of mycobacterium tuberculosis (TB). The most effective way to accomplish this goal is to identify and treat individuals who have active tuberculosis. However, even in highly effected tuberculosis control programs, M. tuberculosis continues to be transmitted to others, largely because most transmission occurs before diagnosis and initiation of therapy. Under the current recommendations, testing should be targeted at specific high-risk populations. While a strategy of targeted testing and treatment of persons most likely to develop tuberculosis is attractive, it is uncertain how best to accomplish this goal. This is the first study to assess the use of geographic and molecular surveillance in guiding a targeted tuberculosis screening and treatment of active tuberculosis and latent tuberculosis infection that monitors potential transmission in a defined high risk geographic area. The results of this geographically targeted program demonstrate significant yield for discovering active cases, latent tuberculosis infection, and recent transmission (TST converters). In this setting, geographically targeted screening identified as many as 19.8 tuberculosis cases per 1,000 persons screened and as many as 292.4 latent tuberculosis infections per 1,000 persons screened. Additionally, successful treatment of these individuals reduced the number of both cases and latent infection identified. Over a three-year period the case detection rate, latent infection detection rate, and TST conversion rate was reduced by 335%, 171% and 285% respectively.Item Factors Associated with Multi-Drug Resistance among Patients with Streptoccus pneumoniae Ear Infections(2004-05-01) Mendoza, Belinda A.; Francisco Soto Mas; Chiehwen Ed Hsu; Antonio ReneMendoza, Belinda A., Factors Associated with Multi-Drug Resistance among Patients with Streptoccus pneumoniae Ear Infections. Master of Public Health (Social and Behavioral Sciences), May 2004, 27 pp., 6 tables, 1 figure, references, 9 titles. Clinical trials play an important role in the development of new medical treatments. The purpose of this study is to describe patients participating in a clinical trial and at analyze the socio-demographic characteristics of patients with susceptible and multi-drug resistant Streptococcus pneumoniae ear infections. At the conclusion of this study, a socio-demographic description of clinical trial participants was obtained and the results were slightly younger than patients with susceptible S. pneumoniae ear infections and were more likely to attend day care.Item Fecal Coliforms in the Rio Grande: A Risk to Human Health(2004-12-01) Tompkins, Erin L.; Thomas Vaughan; Claudia S. CogginTompkins, Erin L., Fecal Coliforms in the Rio Grande: A Risk to Human Health. Master of Public Health (Environmental Health), December 2004, 45 pages, bibliography, 33 titles. The Rio Grande around Laredo, Texas/Nuevo Laredo, Mexico was designated for primary contact reaction by the EPA. However, monthly sampling over a ten-year period in this section of the river may show otherwise. Fecal contamination of the Rio Grande in this area may be a source of illness to the population. Four sites in Laredo area were tested for fecal coliform density and rate of flow. Rainfall data from the USGS was used for comparisons. The rate of flow of the Rio Grande had an impact on fecal coliform density at one site measured. Rainfall in Laredo had an impact on fecal coliform density at two measured sites, and was a significant predictor of density at these sites as well. A review of the designation for this river segment is recommended. More research is needed to determine the exposed population, and effects of high coliform densities on downstream communities.Item Positive Regulation of Acetate Metabolism and Motility by the RNA-Binding Protein CsrA in Escherichia coli(2000-08-01) Wei, Bangdong L.; Jerry Simecka; Ming-Chi Wu; Stephen R. GrantWei, Bangdong L., Positive Regulation of Acetate Metabolism and Motility by the RNA-binding Protein CsrA in Escherichia coli. Doctor of Philosophy (Biomedical Sciences), August, 2000, 118 pp., 5 tables, 19 illustrations, bibliography, 175 titles. The carbon storage regulatory (Csr) system consists of a small RNA-binding effector protein, CsrA, and non-coding RNA, CsrB. CsrA acts as a global regulator and modulates specific mRNA stability in Escherichia coli. It regulates central carbon metabolism, physiology, and cell surface properties on a broad scale. In this study, the regulatory roles of csrA in acetate metabolism and motility were examined. The csrA gene was demonstrated to positively regulate acetyl-CoA synthetase and isocitrate lyase, while it did not affect phosphotransacetylase, isocitrate dehydrogenase, or citrate synthase. As a result, growth of csrA rpoS mutant strains was very poor on acetate as a sole carbon source. Surprisingly, growth also was inhibited specifically by the addition of modest amounts of acetate to rich media. Cultures grown in the presence of ≥25 mM acetate consisted substantially of glycogen biosynthesis (glg) mutants, which were no longer inhibited by acetate. Several classes of glg mutations were mapped to known and novel loci. The TCA cycle intermediates or pyruvate, but not glucose, galactose or glycerol, restored growth and prevented the glg mutations in the presence of acetate. Furthermore, amino acid uptake was inhibited by acetate specifically in the csrA rpoS strain. Apparently, central carbon flux imbalance, inhibition of amino acid uptake, and a deficiency in acetate metabolism are combined to cause metabolic stress by depleting the TCA cycle. The csrA gene was essential for motility and flagellum biosynthesis. Further studies elucidated the molecular mechanism by which CsrA positively regulates flagellum synthesis. Purified recombinant CsrA protein, which was isolated as a ribonucleoprotein complex consisting of one single CsrB molecule and ~18 CsrA subunits, directly stimulated the coupled transcription-translation of flhDC::lacZ in S-30 extracts and bound specifically to the 5’ non-coding segment of flhDC mRNA in mobility shift assay. The steady state level of flhDC mRNA was higher and its half-life was ~3-fold greater in a csrA wild type versus a csrA::kanR mutant strain, as shown by RT-PCR. Thus, CsrA is able to stimulate flhDC gene expression by a post-transcriptional mechanism that resembles its function in repression.Item Predictors of Complicated Staphylococcus Aureus Bacteremia: A Retrospective Validation Study(2008-04-01) Krishnamurthy, Pramod; Fischbach, Lori; Cardarelli, Roberto; Coggin, Claudia S.Krishnamurthy, P., Predictors of Complicated Staphylococcus aureus Bacteremia (SAB): A Retrospective Validation Study. Master of Public Health (Epidemiology), April 2008, 57 pp, 9 tables, 1 illustration, bibliography, 39 titles. SAB often has a complicated clinical course and it is important to identify those at risk for complications to guide management. We conducted a validation study of a clinical prediction tool that uses a scoring system to predict the likelihood of developing complicated SAB. Chapter I is a review of background literature and rationale for our study. Chapter II has sections describing the study design, methods, eligibility criteria, statistical analysis and a summary of the results. We observed significantly higher complications among patients with SAB in our validation study. The prediction tool is not a valid predictor of complicated SAB and we recommend better prediction models to accurately predict complications of SAB.Item Quorum Sensing in Sinorhizobium meliloti(2008-12-01) Patankar, Arati V.; Juan E. Gonzales; Jerry W. Simecka; Stephen O. MathewPatankar, Arati V., Quorum Sensing in Sinorhizobium meliloti. Doctor of Philosophy (Microbiology and Immunology), December 2008, 170 pp., 14 tables, 23 illustrations, bibliography, 212 titles. The overall goal of this study was to elucidate the role of a series of transcriptional regulators and potential signal molecules in the coordination of gene regulation in Sinorhizobium meliloti. The agriculturally important gram-negative soil bacterium S. meliloti, forms a symbiotic association with its host legume, Medicago sativa (alfalfa); thereby serving as a good model for studying host-bacterial interactions. Often, bacteria associated with eukaryotic hosts utilize global gene regulatory systems to coordinate their behavior in order to establish pathogenic or symbiotic associations. Quorum sensing is one such form of bacterial gene regulation which is mediated by signaling molecules and regulatory proteins in a population density dependent manner. In S. meliloti, the process of quorum sensing has been shown to play an important role in the relationship with its host plant. Control of essential processes such as plant nodulation and exopolysaccharide production has been attributed to the Sin/ExpR quorum-sensing system of S. meliloti. Interestingly, S. meliloti contains four additional (SMc04032, SMc00658, Smc00878 and SMc00877) putative quorum-sensing response regulators whose regulatory network was not known. The predicted protein sequences of these genes contain features typical of the LuxR family of proteins i.e., an N-terminal signal binding domain and C-terminal helix-turn-helix DNA biding domain. In order to identify their regulatory role, mutants of the response regulators were constructed and their expression profile was determined by employing genome-wide microarray and real-time PCR expression analysis. Through these analyses, it was determined that the SMc004032 locus controls expression of genes involved in the active methyl cycle, while the SMc00658, SMc00878 and SMc00877 loci control expression of genes from the denitrification of pathway of S. meliloti. Further, through phenotypic studies it was established that SMc04032 impacts stress response adaptation, and effective competition for plant nodulation. This suggests that SMc04032 could play a role in bacterial survival in the soil as well as within the host. The ability to denitrify is highly variable in different strains of S. meliloti. Through growth and enzymatic assays, it was established that the wild-type strain of this study, S. meliloti Rm8530, is a partial dentrifier in which, the capacity to metabolize nitrate is impaired. It was further determined that SMc00658, SMc00878 and SMc0877 modulated nitrite reductase activity under aerobic conditions, implying that these genes are involved in aerobic denitrification and therefor probably play a role in detoxification in S. meliloti. Based on the sequenced-genome analysis, S. meliloti possess homologs of other mediators of quorum sensing, that might be responsible for the synthesis of novel signal molecules. Bioreporter strains and mass spectrometry analysis were employed to identify production of cyclic dipeptides in S. meliloti. These compounds have been previously reported as quorum-sensing signal molecules in several bacteria. The results presented in this study provide a better understanding of S. meliloti’s metabolic and physiological properties and will be fundamental in future studies of bacterial interaction with its host and survival within its ecological niche.Item Resistance of Bacillus Subtilis Spores Lacking Either Nucleotide Excision Repair or Spore Photoproduct Lyase to Ultraviolet (UV) Radiation from Artificial or Natural Sources(1996-06-01) Xue, Yaming; Tony Romeo; Ming-Chi Wu; Wayne L. NicholsonXue, Yaming, Resistance of Bacillus subtilis spores lacking either nucleotide excision repair or spore photoproduct lyase to ultraviolet (UV) radiation from artificial or natural sources. Master of Science (Biomedical Sciences), June 1996, pp., 4 tables, 12 illustrations, references, 38 titles. Exposure of bacterial spores to UV radiation causes the accumulation of a unique pyrimidine dimer in the DNA, “spore photoproduct” (SP). In Bacillus subtilis, two distinct DNA repair pathways are used for removal of SP: general nucleotide excision repair (the uvr parthway), or the SP-specific enzyme SP lyase (the spl pathway). Spores of four strains of Bacillus subtilis differing in their repair capabilities were irradiated under either artificial or solar UV. To determine the biologically-relevant cumulative UV dose under each irradiation condition, a sporocidal dosimeter was constructed. The results showed: (i) Both uvr and spl pathways contributed to the survival of spores under all tested conditions. The spl pathway was more efficient than uvr pathway in repairing the DNA damage caused by UV-C and solar UV-A, but no significant difference was noted in repairing DNA damage caused by UV-B or full-spectrum solar UV. (ii) Exposure of spores to solar UV can cause cellular lethal damage which is reparable by neither repair pathway.Item The Effect of CsrA on Biofilm Development in Escherichia coli(2001-05-01) Jackson, Debra White; Julian Borejdo; Richard Easom; Jerry SimeckaJackson, Debra W., The Effect of CsrA on Biofilm Development in Escherichia coli. Doctor of Philosophy (Biomedical Sciences), May 2001, 127 pp., 2 tables, 15 illustrations, bibliography, 138 titles. CsrA, carbon storage regulator, is a small RNA-binding protein that acts as a global regulator and modulates specific mRNA stability in Escherichia coli. CsrA regulates central carbon metabolism in addition to flagella biogenesis. In this study, the phylogenetic distribution of csrA and its role in Escherichia coli biofilm development were examined. CsrA homologs were examined using Southern hybridization experiment and by analyzing existing sequencing data and was found to be widespread among eubacteria. CsrA was shown to be capable of acting as a genetic switch for biofilm formation and dispersal. A csrA mutant of E. coli was shown to increase biofilm formation and exhibit apparent pillars and channels characteristic of a mature biofilm. Over-expression of csrA completely inhibited biofilm formation in E. coli K-12 and decreased biofilm formation in related enteric pathogens. Induction of csrA expression from a multicopy plasmid caused dispersal of a pre-formed biofilm. Gene expression studies revealed that csrA expression is dynamically regulated during biofilm formation. Several outer-membrane factors and global regulators that have been implicated in biofilm formation were examined for effects on biofilm formation in a csrA mutant. Crystal violet adherence assays revealed that flagella and type I pili affect biofilm formation in a scrA mutant strain; however, colonic acid and curli fimbriae did not exhibit quantitative effects on biofilm formation in the csrA mutant, but the stationary phase sigma factor, RpoS, had no quantitative effect on csrA mutant biofilm formation. Therefore, a csrA mutant will form a biofilm in the absence of each of these outer-membrane factors and global regulatory factors of biofilm formation. The effects of csrA on biofilm formation were found to be mediated in part through its effects on intracellular glycogen metabolism. Thus the redirection of carbon flux, in response to environmental and/or physiological cues, is important for biofilm development.Item The Effects of Two Staphylococcal Global Regulators (agr and sar) on Acid Phosphatase production in Staphylococcus aureus(2003-05-01) Agouna-Deciat, Bahrka Olivier; Jerry Simecka; Michael SmithAgouna-Deciat, B. Olivier, The Effect of Two Staphylococcal Global Regulators (agr and sar) on Acid Phosphatase Production in Staphylococcus aureus. Master of Science (Molecular Biology and Immunology), May 2003, 75 pp., 3 tables, 14 illustrations, 16 titles. Staphylococcus aureus produces an extensive number of cell-surface associated proteins, extracellular proteins and enzymes that contribute to its virulence. The key to better preventative or curative approaches resides in identifying and targeting the very genes and their products that play major roles in the survival of the bacteria within the host and the establishment of diseases. Two well known regulatory loci, the accessory gene regulatory (agr) and the staphylococcal accessory regulator (sar), control the expression of most S. aureus genes that encode for its virulence factors. Other virulence gene regulators have recently been isolated. Over 40 proteins and enzymes produced by S. aureus have been identified and several of them have been linked to staphylococcal pathogenesis. In this study, we attempt to determine the role of agr and sar in the regulation of the production of a secreted staphylococcal acid phosphatase (Sap) suspected to contribute to virulence.Item The Impact of the Mycoplasma pulmonis MALP-2 Homologue on Disease Progression(2008-04-01) Spear, Marcia G.; Simecka, Jerry W.; Hodge, Lisa M.; Mathew, Porunelloor A.Spear, Marcia. The Impact of Mycoplasma pulmonis MALP-2 Homologue on Disease Progression. Master of Science (Biomedical Sciences), April 2008. 64 pp., 3 tables, 8 illustrations. Using Mycoplasma pulmonis, this project looked at a possible critical component in mycoplasma disease, the MALP-2 homologue lipoprotein. Studies demonstrated other lipoproteins besides the MALP-2 homologue were critical for in vivo disease progression and in vitro macrophage IL-6, IL-12, and TNF-α cytokine production. This trend was also seen human endothelial kidney (HEK) cells transfected with toll-like receptor 1 (TLR2) and the heterodimer TLR2/6. An increase in IL-8 cytokine production seen in all stimulated HEK cell lines, indicating the lipoproteins involved in cell interactions are TLR2 mediated. This project suggests the M. pulmonis MALP-2 homologue is not the main lipoprotein involved in disease progression and cell interactions, indicating the MALP-2 homologue may not be an ideal target for vaccines or antibiotics.Item Variations of the lung microbiome and immune response in mechanically ventilated surgical patients(PLOS, 2018-10-24) Huebinger, Ryan M.; Smith, Ashley D.; Zhang, Yan; Monson, Nancy L.; Ireland, Sara J.; Barber, Robert C.; Kubasiak, John C.; Minshall, Christian T.; Minei, Joseph P.; Wolf, Steven E.; Allen, Michael S.Mechanically ventilated surgical patients have a variety of bacterial flora that are often undetectable by traditional culture methods. The source of infection in many of these patients remains unclear. To address this clinical problem, the microbiome profile and host inflammatory response in bronchoalveolar lavage samples from the surgical intensive care unit were examined relative to clinical pathology diagnoses. The hypothesis was tested that clinical diagnosis of respiratory tract flora were similar to culture positive lavage samples in both microbiome and inflammatory profile. Bronchoalveolar lavage samples were collected in the surgical intensive care unit as standard of care for intubated individuals with a clinical pulmonary infection score of >6 or who were expected to be intubated for >48 hours. Cytokine analysis was conducted with the Bioplex Pro Human Th17 cytokine panel. The microbiome of the samples was sequenced for the 16S rRNA region using the Ion Torrent. Microbiome diversity analysis showed the culture-positive samples had the lowest levels of diversity and culture negative with the highest based upon the Shannon-Wiener index (culture positive: 0.77 ± 0.36, respiratory tract flora: 2.06 ± 0.73, culture negative: 3.97 ± 0.65). Culture-negative samples were not dominated by a single bacterial genera. Lavages classified as respiratory tract flora were more similar to the culture-positive in the microbiome profile. A comparison of cytokine expression between groups showed increased levels of cytokines (IFN-g, IL-17F, IL-1B, IL-31, TNF-a) in culture-positive and respiratory tract flora groups. Culture-positive samples exhibited a more robust immune response and reduced diversity of bacterial genera. Lower cytokine levels in culture-negative samples, despite a greater number of bacterial species, suggest a resident nonpathogenic bacterial community may be indicative of a normal pulmonary environment. Respiratory tract flora samples were most similar to the culture-positive samples and may warrant classification as culture-positive when considering clinical treatment.