DOCSLIB.ORG

Conjugative Transfer Pathways of High-Level Mupirocin Resistance and Conjugative Transfer Genes in Staphylococcus

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
Conjugative Transfer Pathways of High-Level Mupirocin Resistance and Conjugative Transfer Genes in Staphylococcus East Tennessee State University Digital Commons @ East Tennessee State University Electronic Theses and Dissertations Student Works 5-2006 Conjugative Transfer Pathways of High-Level Mupirocin Resistance and Conjugative Transfer Genes in Staphylococcus. Danielle Barnard East Tennessee State University Follow this and additional works at: https://dc.etsu.edu/etd Part of the Bacteriology Commons Recommended Citation Barnard, Danielle, "Conjugative Transfer Pathways of High-Level Mupirocin Resistance and Conjugative Transfer Genes in Staphylococcus." (2006). Electronic Theses and Dissertations. Paper 2188. https://dc.etsu.edu/etd/2188 This Thesis - Open Access is brought to you for free and open access by the Student Works at Digital Commons @ East Tennessee State University. It has been accepted for inclusion in Electronic Theses and Dissertations by an authorized administrator of Digital Commons @ East Tennessee State University. For more information, please contact [email protected]. Conjugative Transfer Pathways of High-level Mupirocin Resistance and Conjugative Transfer Genes in Staphylococcus A thesis presented to the faculty of the Department of Biological Sciences East Tennessee State University In partial fulfillment of the requirements for the degree Masters of Science in Biological Sciences by Danielle Barnard May 2006 Foster Levy Ph.D., Co-Chair Elaine Walker Ph.D., Co-Chair Scott Champney Ph.D. Keywords: Antibiotic Resistance, Conjugative Transfer, Staphylococcus, Mupirocin, tra Genes ABSTRACT Conjugative Transfer Pathways of High-level Mupirocin Resistance and Conjugative Transfer Genes in Staphylococcus by Danielle Barnard To combat widespread infections caused by Staphylococcus aureus, mupirocin was introduced at the Veterans Affairs Medical Center, Mountain Home, Tennessee. Soon after introduction, high- level mupirocin-resistance emerged. The rapid emergence was hypothesized to be due to conjugative transfer of the mupA resistance gene from S. epidermidis to S. aureus. Results have shown that transfer of high-level mupirocin-resistance from S. aureus donors commonly occurs. However, transfer from naturally-occurring S. epidermidis donors was not attainable. Staphylococcus epidermidis transconjugants, however, were capable of serving as donors. Further examination of non-transmissibility included PCR analysis of conjugative transfer genes (tra genes) in capable and non-capable donors. Results confirmed that capable donors possess full-length copies of selected transfer genes. Non-capable donors varied in the presence/absence of full-length copies of transfer genes, but none had all three genes. The genetic differences among non-capable donors suggest that non-transmissibility has arisen independently in different strains via gene deletions and recombinations. 2 DEDICATION In Loving Memory of Agnes Bailey, my “Granny”, 1925-2004. “One In A Million” 3 ACKNOWLEDGMENTS I would like to acknowledge the two wonderful co-chairs of my thesis committee, Dr. Foster Levy and Dr. Elaine Walker. I would like to thank them for introducing me to the world of basis science research. Dr. Levy and Dr. Walker have provided their expertise and insight concerning the field of antibiotic resistance as they have so capably served as my mentors during this thesis research. I am forever indebted to them for their countless hours of supervision, guidance, and support throughout my research. I would also like to thank Dr. Scott Champney, committee member, for all of his advice and comments on my research project. Dr. Champney’s suggestions concerning the molecular component of my research were most helpful. I would like to thank the ETSU Department of Biological Sciences for the opportunity to conduct research for my Masters of Science degree and for affording me the opportunity to serve as a graduate assistant during my master’s degree program. I would like to acknowledge the following entities for their financial support of my research: GlaxoSmithKline and ETSU’s School of Graduate Studies for the Research Award. Many thanks to others who have provided me with support and encouragement during my research including: Scott Reynolds, Susan Reynolds, Brandi Earp, Samara Miller, and Kristi Tipton. Lastly, I would like to acknowledge my parents, Lynn and Janet Barnard, for their continuous love, support, and patience during my educational endeavors. 4 CONTENTS Page ABSTRACT ........................................................................................................................ 2 DEDICATION....................................................................................................................... 3 ACKNOWLEDGMENTS ..................................................................................................... 4 LIST OF TABLES................................................................................................................. 7 LIST OF FIGURES ............................................................................................................... 8 Chapter 1. INTRODUCTION ........................................................................................................ 9 Pathogenic Bacteria and Antibiotic Resistance ......................................................... 9 Staphylococcus........................................................................................................... 10 MRSA and Nosocomial Infections………………………………………………… 11 Mupirocin................................................................................................................... 12 Horizontal Gene Exchange........................................................................................ 14 Conjugative Transfer Genes...................................................................................... 15 Resistance Reservoirs................................................................................................ 16 Research Approach.................................................................................................... 18 2. MATERIAL AND METHODS .................................................................................... 19 Bacterial Samples ...................................................................................................... 19 Conjugative Transfer Assays .................................................................................... 21 Restriction-Deficient Recipient................................................................................. 24 Pre-exposure to a β-lactam Agent………………………………………………….. 25 PCR DNA Preparations……………………………………………………………. 25 5 Chapter Page PCR Primers............................................................................................................... 26 PCR Reactions and Conditions.................................................................................. 27 3. RESULTS...................................................................................................................... 29 Conjugations Involving S. aureus and S. epidermidis .............................................. 29 Conjugations Involving non-S. epidermidis CNS...................................................... 34 Restriction-Deficient Recipient................................................................................. 36 Pre-exposure of Recipients to a β-lactam agent......................................................... 37 PCR Analyses of mupA and Conjugative Transfer Genes......................................... 38 4. DISCUSSION................................................................................................................ 43 Interspecific Transfer from S. epidermidis Donors .................................................. 43 Transfer of Mupirocin Resistance Interspecifically................................................... 44 Explanations for Non-transmissibility....................................................................... 45 Conjugative Transfer Genes...................................................................................... 46 Conclusions................................................................................................................ 47 REFERENCES ..................................................................................................................... 49 APPENDIX: Conjugative Transfer Rates............................................................................. 52 VITA ………........................................................................................................................ 53 6 LIST OF TABLES Table Page 1. Characteristics of Staphylococcal Strains Used in Conjugative Transfer Assays and PCR Analyses.......................................................................................................... 20 2. Number of Staphylococcal Tested as Donors and Recipients in Filter Mating Experiments ............................................................................................................ 24 3. Primers Used in PCR Analysis ................................................................................... 28 4. Intra- and Interspecific Conjugative Transfer of Mupirocin Resistance .................... 30 5. Summary of Conjugative Transfers for Donor and Recipient Combinations Involving S. aureus and S. epidermidis .................................................................. 31 6. 2x2 Contingency Table Showing Similar Frequency of Successful Conjugative Transfer of High-level Mupirocin Resistance
Load more

Recommended publications
  • 85224601.Pdf
    View metadata, citation and similar papers at core.ac.uk brought to you by CORE provided by RERO DOC Digital Library RESEARCH LETTER Construction and characterization of Enterococcus faecalis CG110/gfp/pRE25Ã, a tool for monitoring horizontal gene transfer in complex microbial ecosystems Martina C. Haug, Sabine A. Tanner, Christophe Lacroix, Leo Meile & Marc J.A. Stevens Laboratory of Food Biotechnology, Institute of Food, Nutrition and Health, ETH Zurich, Zurich, Switzerland Correspondence: Leo Meile, Laboratory of Abstract Food Biotechnology, Institute of Food, Nutrition and Health, ETH Zurich, Enterococci are among the most notorious bacteria involved in the spread of Schmelzbergstrasse 7, CH-8092 Zurich, antibiotic resistance (ABR) determinants via horizontal gene transfer, a process that Switzerland. Tel.: 141 44 632 33 62; fax: leads to increased prevalence of antibiotic-resistant bacteria. In complex microbial 141 44 632 14 03; e-mail: communities with a high background of ABR genes, detection of gene transfer is [email protected] possible only when the ABR determinant is marked. Therefore, the conjugative multiresistance plasmid pRE25, originating from a sausage-associated Enterococcus Received 6 August 2010; revised 8 September faecalis, was tagged with a 34-bp random sequence marker spliced by tet(M). The 2010; accepted 29 September 2010. plasmid constructed, designated pRE25Ã, was introduced into E. faecalis CG110/ Final version published online 28 October 2010. gfp, a strain containing a gfp gene as chromosomal marker. The plasmid pRE25Ã is DOI:10.1111/j.1574-6968.2010.02131.x fully functional compared with its parental pRE25, occurs at one to two copies per chromosome, and can be transferred to Listeria monocytogenes and Listeria innocua À6 À8 Editor: Andre´ Klier at frequencies of 6 Â 10 to 8 Â 10 transconjugants per donor.
    [Show full text]
  • Klinicky Významné Rezistentní Plazmidy – Studium Biologické Zátěže a Stability Úspěšných Mobilních Genetických Elementů
    MASARYKOVA UNIVERZITA PŘÍRODOVĚDECKÁ FAKULTA ÚSTAV EXPERIMENTÁLNÍ BIOLOGIE Klinicky významné rezistentní plazmidy – studium biologické zátěže a stability úspěšných mobilních genetických elementů Diplomová práce Tomáš Nohejl Vedoucí práce: doc. RNDr. Monika Dolejská, Ph.D. Brno 2018 Bibliografický záznam Autor: Bc. Tomáš Nohejl Přírodovědecká fakulta, Masarykova univerzita Ústav experimentální biologie Název práce: Klinicky významné rezistentní plazmidy – studium biologické zátěže a stability úspěšných mobilních genetických elementů Studijní program: Experimentální biologie Studijní obor: Molekulární biologie a genetika Vedoucí práce: doc. RNDr. Monika Dolejská, Ph.D. Akademický rok: 2017/2018 Počet stran 78 Klíčová slova: Antibiotická rezistence; plazmid; horizontální přenos genetické informace; HGT; konjugace; PMQR; chinolony; fitness; perzistence Bibliographic entry Author: Bc. Tomáš Nohejl Faculty of Science, Masaryk University Department of Experimental Biology Title of Thesis: Clinically important resistance plasmids - the study of biological burden and stability of successful mobile genetic elements Degree programme: Experimental biology Field of Study: Molecular Biology and Genetics Supervisor: doc. RNDr. Monika Dolejská, Ph.D. Academic Year: 2017/2018 Number of Pages: 78 Keywords: Antibiotic resistance; plasmid; horizontal gene transfer; HGT; conjugation; PMQR; quinolones; fitness; persistence Abstrakt Antibiotická rezistence představuje jeden z nejzávažnějších globálních problémů současnosti. Šíření genů kódující antibiotickou
    [Show full text]
  • ESBL-Producing Escherichia Coli in Swedish Gulls—A Case of Environmental Pollution from Humans?
    RESEARCH ARTICLE ESBL-producing Escherichia coli in Swedish gullsÐA case of environmental pollution from humans? Clara Atterby1, Stefan BoÈrjesson2*, Sofia Ny3,4, Josef D. JaÈrhult1,5, Sara Byfors3, Jonas Bonnedahl6,7,8 1 Zoonosis Science Center, Department of Medical Biochemistry and Microbiology, Uppsala University, Uppsala, Sweden, 2 Department of Animal Health and Antimicrobial strategies, National Veterinary Institute (SVA), Uppsala, Sweden, 3 Public Health Agency of Sweden, Stockholm, Sweden, 4 Department of a1111111111 Laboratory Medicine, Karolinska Institute, Stockholm, Sweden, 5 Section of Infectious Diseases, Department a1111111111 of Medical Sciences, Uppsala University, Uppsala, Sweden, 6 Center for Ecology and Evolution in Microbial a1111111111 Model Systems, Linnaeus University, Kalmar,Sweden, 7 Department of Infectious Diseases, Kalmar County a1111111111 Council, Kalmar, 8 Department of Clinical and Experimental Medicine, LinkoÈping University, LinkoÈping, a1111111111 Sweden * [email protected] OPEN ACCESS Abstract Citation: Atterby C, BoÈrjesson S, Ny S, JaÈrhult JD, ESBL-producing bacteria are present in wildlife and the environment might serve as a resis- Byfors S, Bonnedahl J (2017) ESBL-producing Escherichia coli in Swedish gullsÐA case of tance reservoir. Wild gulls have been described as frequent carriers of ESBL-producing environmental pollution from humans? PLoS ONE E. coli strains with genotypic characteristics similar to strains found in humans. Therefore, 12(12): e0190380. https://doi.org/10.1371/journal. potential dissemination of antibiotic resistance genes and bacteria between the human pop- pone.0190380 ulation and wildlife need to be further investigated. Occurrence and characterization of Editor: Michael Lierz, Justus-Liebeig University ESBL-producing E. coli in Swedish wild gulls were assessed and compared to isolates from Giessen, GERMANY humans, livestock and surface water collected in the same country and similar time-period.
    [Show full text]
  • FINAL REPORT the Molecular Biology of Nitroamine Degradation in Soils
    FINAL REPORT The Molecular Biology of Nitroamine Degradation in Soils SERDP Project ER-1608 JULY 2015 Stuart E. Strand University of Washington Neil C. Bruce University of York David A. Stahl University of Washington Jalal Hawari National Research Council of Canada Distribution Statement A This report was prepared under contract to the Department of Defense Strategic Environmental Research and Development Program (SERDP). The publication of this report does not indicate endorsement by the Department of Defense, nor should the contents be construed as reflecting the official policy or position of the Department of Defense. Reference herein to any specific commercial product, process, or service by trade name, trademark, manufacturer, or otherwise, does not necessarily constitute or imply its endorsement, recommendation, or favoring by the Department of Defense. Form Approved REPORT DOCUMENTATION PAGE OMB No. 0704-0188 The public reporting burden for this collection of information is estimated to average 1 hour per response, including the time for reviewing instructions, searching existing data sources, gathering and maintaining the data needed, and completing and reviewing the collection of information. Send comments regarding this burden estimate or any other aspect of this collection of information, including suggestions for reducing the burden, to Department of Defense, Washington Headquarters Services, Directorate for Information Operations and Reports (0704-0188), 1215 Jefferson Davis Highway, Suite 1204, Arlington, VA 22202-4302. Respondents should be aware that notwithstanding any other provision of law, no person shall be subject to any penalty for failing to comply with a collection of information if it does not display a currently valid OMB control number.
    [Show full text]
  • Bamidele Tolulope, Odumosu
    MOLECULAR CHARACTERISATION OF MULTIDRUG-RESISTANT Pseudomonas aeruginosa IN SOUTHWESTERN NIGERIA. BY BAMIDELE TOLULOPE, ODUMOSU B.Sc Microbiology (OOU), M.Sc. Pharmaceutical Microbiology (Ibadan) A thesis in the Department of Pharmaceutical Microbiology Submitted to the Faculty of Pharmacy in partial fulfillment of the requirements for the degree of DOCTOR OF PHILOSOPHY of the UNIVERSITY OF IBADAN, IBADAN, NIGERIA. APRIL, 2013 i CERTIFICATION I certify that this project was carried out by Mr B.T. Odumosu under my supervision in the Department of Pharmaceutical Microbiology, University of Ibadan, Nigeria. ----------------------------------------------------------- Supervisor Bola A. Adeniyi, B.Sc, M.Sc (Ilorin), Ph.D. (Ibadan) Professor, Department of Pharmaceutical Microbiology University of Ibadan, Ibadan, Nigeria. ii DEDICATION This research work is dedicated to Almighty God, the God of Jacob who answers me in times of my distress and has been with me wherever I have gone. To you alone is glory. And to the loving memory of my beloved mother, Florence Titilayo Adunni Odumosu, continue to rest in the bosom of our dear Lord (amen). Mummy how I wish you were here! iii ACKNOWLEDGMENTS I acknowledge the love and support of Professor Bolanle Adeniyi who has been my supervisor since the days of my Master of Science (M.Sc) till present time. I enjoyed her commitment and dedication towards the successful completion of my postgraduate study, “Thank you ma”. I also want to acknowledge Professor Ram Chandra who was my host and supervisor in India and ensured I never lacked support as regarding my training and research in Lucknow India. My appreciation goes to the head of Department of Pharmaceutical Microbiology, Dr O.
    [Show full text]
  • Aus Dem Institut Für Pflanzenbau Und Pflanzenzüchtung Der Christian-Albrechts-Universität Zu Kiel
    Aus dem Institut für Pflanzenbau und Pflanzenzüchtung der Christian-Albrechts-Universität zu Kiel Hybrid Assembly of Whole Genome Shotgun Sequences of Two Sugar Beet (Beta vulgaris L.) Translocation Lines Carrying the Beet Cyst Nematode Resistance Gene Hs1-2 and Functional Analysis of Candidate Genes der Agrar- und Ernährungswissenschaftlichen Fakultät der Christian-Albrechts-Universität zu Kiel vorgelegt von M.Sc. Sarah Christina Jäger aus Gießen Kiel, 06. Dezember 2012 Dekan: Prof. Dr. Rainer Horn 1. Berichterstatter: Prof. Dr. Christian Jung 2. Berichterstatter: Prof. Dr. Andre Franke Tag der mündlichen Prüfung: 31.01.2013 Table of Contents I Table of Contents Table of Contents ...................................................................................................................................... I Abbreviations .......................................................................................................................................... V List of Figures ........................................................................................................................................ XI List of Tables ......................................................................................................................................... XV 1 General Introduction ........................................................................................................................ 1 1.1 Recent Advances in Plant Genomics ........................................................................................
    [Show full text]
  • Expansion of Highly Stable Blaoxa-10 Β-Lactamase Family Within Diverse
    Maurya et al. BMC Res Notes (2017) 10:145 DOI 10.1186/s13104-017-2467-2 BMC Research Notes RESEARCH ARTICLE Open Access Expansion of highly stable blaOXA-10 β-lactamase family within diverse host range among nosocomial isolates of Gram-negative bacilli within a tertiary referral hospital of Northeast India Anand Prakash Maurya1, Debadatta Dhar2, Mridul Kumar Basumatary1, Deepjyoti Paul1, Birson Ingti1, Debarati Choudhury3, Anupam Das Talukdar3, Atanu Chakravarty2, Shweta Mishra4 and Amitabha Bhattacharjee1* Abstract Background: The current study reports dissemination of highly stable blaOXA-10 family of beta lactamases among diverse group of nosocomial isolates of Gram-negative bacilli within a tertiary referral hospital of the northern part of India. Methods: In the current study, a total number of 590 Gram negative isolates were selected for a period of 1 year (i.e. 1st November 2011–31st October 2012). Members of Enterobacteriaceae and non fermenting Gram negative rods were obtained from Silchar Medical College and Hospital, Silchar, India. Screening and molecular characterization of β-lactamase genes was done. Integrase gene PCR was performed for detection and characterization of integrons and cassette PCR was performed for study of the variable regions of integron gene cassettes carrying blaOXA-10. Gene transferability, stability and replicon typing was also carried out. Isolates were typed by ERIC as well as REP PCR. Results: Twenty-four isolates of Gram-negative bacilli that were harboring blaOXA-10 family (OXA-14, and OXA16) with fact that resistance was to the extended cephalosporins. The resistance determinant was located within class I inte- gron in five diverse genetic contexts and horizontally transferable in Enterobacteriaceae, was carried through IncY type plasmid.
    [Show full text]
  • Analysis of the Novel Lipid Transfer Protein Anchored at Membrane Contact Sites (LAM) Family
    LONDON’S GLOBAL UNIVERSITY Analysis of the novel Lipid transfer protein Anchored at Membrane contact sites (LAM) family A thesis submitted to the University College London in the fulfilment of the requirements for the degree of Doctor of Philosophy Supervisor: Dr Tim Levine Department of Cell Biology, Faculty of Brain Science, Institute of Ophthalmology 2017 Declaration I, Louise Wong, confirm that the work presented in this thesis is my own. Where information has been derived from other sources, I confirm that this has been indicated. June 2017 1 2 Abstract Membrane contact sites are dynamic structures where two organelles come into close proximity to regulate and facilitate the flow of material and information between them. One type of inter-organelle communication is lipid exchange, which is essential for membrane maintenance and in response to environmental and cellular stimuli. We recently discovered a new family of Lipid transfer proteins Anchored at Membrane contact sites (LAMs) that is present in all eukaryotes. LAM proteins are integral Endoplasmic Reticulum (ER) proteins containing at least one domain that is structurally similar to the StARkin domain superfamily, a specialised fold that can bind amphipathic ligands such as lipids. The budding yeast, Saccharomyces cerevisiae, has six such proteins: Lam1p-6p. Lam1p-4p are located at contacts between the ER and the plasma membrane (PM), and Lam1p-3p are implicated in retrograde sterol traffic between the ER and PM. The PM contains a high concentration of sterol where it increases rigidity by altering the packing characteristics of the phospholipids in the bilayer. Sterol is also important in the ER, where its levels are low but it is both synthesised and sensed.
    [Show full text]