The role of the non-ribosomal peptide synthetase AusAB and its product phevalin in intracellular virulence of Staphylococcus aureus: = Die Rolle der nicht-ribosomalen Peptidsynthetase AusAB und ihres Produktes Phevalin in der intrazellulären Virulenz von Staphylococcus aureus
Gespeichert in:
| 1. Verfasser: | |
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| Format: | Abschlussarbeit Buch |
| Sprache: | English |
| Veröffentlicht: |
Würzburg
2016
|
| Schlagworte: | |
| Online-Zugang: | kostenfrei Inhaltsverzeichnis Inhaltsverzeichnis |
| Beschreibung: | 186 Seiten Illustrationen, Diagramme 30 cm |
Internformat
MARC
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| 100 | 1 | |a Blättner, Sebastian |d ca. 21. Jh. |e Verfasser |0 (DE-588)1132839521 |4 aut | |
| 245 | 1 | 0 | |a The role of the non-ribosomal peptide synthetase AusAB and its product phevalin in intracellular virulence of Staphylococcus aureus |b = Die Rolle der nicht-ribosomalen Peptidsynthetase AusAB und ihres Produktes Phevalin in der intrazellulären Virulenz von Staphylococcus aureus |c submitted by Sebastian Blättner fraom München |
| 246 | 1 | 1 | |a Die Rolle der nicht-ribosomalen Peptidsynthetase AusAB und ihres Produktes Phevalin in der intrazellulären Virulenz von Staphylococcus aureus |
| 264 | 1 | |a Würzburg |c 2016 | |
| 300 | |a 186 Seiten |b Illustrationen, Diagramme |c 30 cm | ||
| 336 | |b txt |2 rdacontent | ||
| 337 | |b n |2 rdamedia | ||
| 338 | |b nc |2 rdacarrier | ||
| 502 | |b Dissertation |c Julius-Maximilians-Universität Würzburg |d 2017 | ||
| 546 | |a Zusammenfassung in deutscher und englischer Sprache | ||
| 650 | 0 | 7 | |a Staphylococcus aureus |0 (DE-588)4182912-8 |2 gnd |9 rswk-swf |
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| 655 | 7 | |0 (DE-588)4113937-9 |a Hochschulschrift |2 gnd-content | |
| 689 | 0 | 0 | |a Staphylococcus aureus |0 (DE-588)4182912-8 |D s |
| 689 | 0 | 1 | |a Virulenz |0 (DE-588)4129834-2 |D s |
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| 912 | |a ebook | ||
| 999 | |a oai:aleph.bib-bvb.de:BVB01-029741164 | ||
Datensatz im Suchindex
| _version_ | 1804177569307688960 |
|---|---|
| adam_text | THE ROLE OF THE NON-RIBOSOMAL PEPTIDE SYNTHETASE AUSAB AND ITS
PRODUCT PHEVALIN IN INTRACELLULAR VIRULENCE OF STAPHYLOCOCCUS AUREUS
DIE ROLLE DER NICHT-RIBOSOMALEN PEPTIDSYNTHETASE AUSAB UND IHRES
PRODUKTES PHEVALIN IN DER INTRAZELLULAEREN VIRULENZ VON
STAPHYLOCOCCUS
AUREUS
DOCTORAL THESIS FOR A DOCTORAL DEGREE
AT THE GRADUATE SCHOOL OF LIFE SCIENCES,
JULIUS-MAXIMILIANS-UNIVERSITAET WUERZBURG,
SECTION INFECTION AND IMMUNITY
SUBMITTED BY
SEBASTIAN BLAETTNER
FROM
MUENCHEN
WUERZBURG 2016
TABLE OF CONTENT
TABLE OF
CONTENT............................................................................................................................................1
1.
SUMMARY...................................................................................................................................................
5
1.1
ABSTRACT...............................................................................................................................................
5
1.2 ZUSAMMENFASSUNG
......................
7
2.
INTRODUCTION..............................................................................................................................................
9
2.1 STAPHYLOCOCCUS
AUREUS.......................................................................................................................
9
2.1.1 GENERAL
INFORMATION....................................................................................................................
9
2.1.2 PREVALENCE O F S.
AUREUS.............................................................................................................10
2.1.2.1
COLONIZATION.......................................................................................................................
10
2.1.2.2
EPIDEMIOLOGY.....................................................................................................................
10
2.1.2.3 S. AUREUS STRAINS OF SEQUENCE TYPE 8: THE SUCCESS OF
USA300......................................11
2.2 VIRULENCE FACTORS OF S.
AUREUS...........................................................................................................12
2.2.1
TOXINS.........................................................................................................................................12
2.2.2.1 ALPHA-HEMOLYSIN
(A-TOXIN)................................................................................................
12
2.2.2.2 BETA-HEMOLYSIN
(SS-TOXIN)..................................................................................................
13
2.2.2.3
LEUKOCIDINS........................................................................................................................
14
2.2.2.4
ENTEROTOXINS..................................................................................................................
16
2.2.2 SECRETED PROTEASES AND
NUCLEASES...........................................................................................
17
2.2.3 COMPLEMENT INHIBITION
FACTORS......................................................................................
17
2.2.4 PHENOL SOLUBLE
MODULINS...........................................................................................................18
2.2.5 ADHESION TO NON-ORGANIC AND ORGANIC
SURFACES.......................................................................
21
2.2.6 SMALL MOLECULE PRODUCTS OF S. AUREUS AND THEIR ASSOCIATION WITH
VIRULENCE
.........................
22
2.2.7 REGULATION OF VIRULENCE IN S.
AUREUS.........................................................................................23
2.2.7.1 THE AGR QUORUM SENSING
SYSTEM......................................................................................23
2.2.7.2 SAERS TWO-COMPONENT
SYSTEM.........................................................................................26
2.2.73 MARR-TYPE TRANSCRIPTIONAL REGULATORS: THE SARA PROTEIN FAM
ILY.....................................27
2.2.7.4
CODY...................................................................................................................................
27
2.2.7.5 ALTERNATIVE SIGMA FACTOR B
(OB).........................................................................................28
2.3 INTRACELLULAR PERSISTENCE AND
VIRULENCE...........................................................................................29
2.3.1 INVASION IN NON-PROFESSIONAL PHAGOCYTES
................................................................................
29
2.3.2 SMALL COLONY VARIANTS
(SCV)......................................................................................................
30
2.3.3 INTRACELLULAR CYCLING OF 5.
AUREUS...........................................................................................
32
2.3.45. AUREUS INDUCED HOST CELL
DEATH............................................................................................33
2.4
S. AUREUS MODULATION OF PHAGOSOME MATURATION AND
INTEGRITY.....................................................34
2.5 AIM OF THIS
WORK.................................................................................................................................37
3. MATERIAL AND
METHODS...........................................................................................................................
38
3.1 M
ATERIAL.............................................................................................................................................
38
3.1.1 BACTERIAL
STRAINS.........................................................................................................................
38
3.1.2 CELL LINES
.......................................
39
3.1.3
PLASMIDS......................................................................................................................................39
3.1.4
OLIGONUCLEOTIDES........................................................................................................................
40
3.1.5
ENZYMES......................................................................................................................................42
3.1.6 BUFFER AND
MEDIA.......................................................................................................................
42
3.1.7
KITS..............................................................................................................................................
44
3.1.8
CHEMICALS...................................................................................................................................44
3.1.9 TECHNICAL
EQUIPMENT.................................................................................................................47
3.1.10
SOFTWARE...................................................................................................................................47
3.2 M
ETHODS.....................................................................................................................................
48
3.2.1 BACTERIAL CULTURE
METHODS.........................................................................................................48
3.2.2 GENETICAL MANIPULATION OF
BACTERIA..........................................................................................
49
3.2.3 S. AUREUS KNOCK-OUT
STRATEGIES.................................................................................................
51
3.2.4 CELL CULTURE
TECHNIQUES..............................................................................................................53
3.2.5 CELL INFECTION
PROTOCOLS.............................................................................................................54
3.2.6 PMN
TESTS...................................................................................................................................58
3.2.7 DETECTION OF AUSAB GENE EXPRESSION AND AUREUSIMINES PRODUCTION
....................................
59
3.2.8 DETECTION OF THE MITOCHONDRIA PHENOTYPE OF
PHEVALIN..........................................................61
3.2.9 DIFFERENCES IN THE PHOSPHOPROTEOME OF PHEVALIN-TREATED CELLS
............................................
62
3.2.10 MOUSE INFECTION
MODELS....................................................... 62
3.2.11.1 PULMONARY LUNG
INFECTION...............................................................................................62
3.2.11.2 MUSCLE ABSCESS MODEL
..................................................................
64
4.
RESULTS.....................................................................................................................................................
65
4.1 GENERATION OF A TRANSPOSON INSERTION MUTANT LIBRARY IN S. AUREUS
LAC........................................65
4.2 SCREENING FOR SINGLE GENE MUTANTS STRAINS DEFICIENT IN PHAGOSOMAL
ESCAPE USING AUTOMATED
MICROSCOPY.........................................................................................................................................
67
4.3 THE NON-RIBOSOMAL PEPTIDE SYNTHETASE CRUSAB AND ITS INFLUENCE ON
PHAGOSOMAL ESCAPE
.........
74
4.3.1 KINETICS OF ESCAPE IN AUSAB M UTANTS
.....................................................................................
77
4.3.2 UPLC ANALYSIS OF MUTANT STRAIN
SUPERNATANTS........................................................................
80
4.3.3 GROWTH PHASE DEPENDENCY OF AUREUSIMINE PRODUCTION
.......................................................
81
4.3.4 AUREUSIMINES ARE PRODUCED BY INTRACELLULAR 5. AUREUS
.........................................................
82
4.3.5 GENETIC COMPLEMENTATION OF PHEVALIN SYNTHESIS RESTORES PHAGOSOMAL
ESCAPE OF AN CRIVSB
MUTANT........................................................................................................................................83
4.4 MUTANTS IN AUSA AND OUSB DO NOT AFFECT GROWTH IN BROTH, HEMOLYSIS
BUT DO AFFECT INTRACELLULAR
CYTOTOXICITY........................................................................................................................................84
4.4.1 MUTANTS IN AUSA AND OUSB DO NOT SHOW A GROWTH DEFECT
.....................................................
84
4.4.2 NO DIFFERENCES IN HEMOLYSIS BETWEEN AUSAB MUTANTS AND WILD TY P E
.................................
85
4.4.3 INVASION INTO HOST CELLS NOT AFFECTED BY CRUSAB MUTATION
......................................................
86
4.4.4 AUSAB MUTANTS ARE ATTENUATED IN INTRACELLULAR CYTOTOXICITY
..................................................
87
4.4.5 CELL DEATH OF HOST CELLS BY S. AUREUS SUPERNATANT NOT INFLUENCED
BY PHEVALIN
....................
89
4.5 ADDITION OF SYNTHETIC PHEVALIN CAN RESTORE AND ENHANCE PHAGOSOMAL
ESCAPE OF S. AUREUS
......
91
4.5.1 PHEVALIN ASSOCIATES WITH HOST CELLS WHEN ADDED EXTRACELLULARLY
............................................
92
4.5.2 PHEVALIN DOES NOT PROMOTE THE PHAGOSOMAL RELEASE OF
FIXATION-KILLED BACTERIA
...................
93
4.5.3 PHEVALIN DOES NOT ACT AS A CLASSICAL CALPAIN
INHIBITOR..............................................................94
4.5.4 PHEVALIN DOES NOT EXHIBIT ANTIMICROBIAL
PROPERTIES................................................................95
4.6 AUREUSIMINE EXPRESSION PROMOTES SURVIVAL AND CYTOTOXICITY IN
LEUKOCYTES
...............................
97
4.6.1 CPU IN PMN ARE NOT INFLUENCED BY AUREUSIMINE PRODUCTION
............................................
97
4.6.2 INTRACELLULAR CYTOTOXICITY IN PMN IS DECREASED IN AUSAB M UTANTS
.........................................
98
4.6.3 PHEVALIN REDUCES CALCIUM FLUXES IN RESPONSE TO FPR-RECEPTOR
STIMULI IN PMN
......................
99
4.6.4 SURVIVAL AND CYTOTOXICITY OF AUREUSIMINE MUTANTS ARE DIMINISHED IN
MACROPHAGES
..........
102
4.7 THE ROLE OF OUSAB IN IN VIVO MOUSE INFECTION
.........................................
104
4.7.1 MOUSE PULMONARY LUNG INFECTION
MODEL................................................................................104
4.7.2 MOUSE MUSCLE ABSCESS INFECTION
MODEL.................................................................................106
4.8 TOWARDS IDENTIFICATION OF THE HOST TARGET OF S. OUREUS-PRODUCED
PHEVALIN
..............................
107
4.9 MITOCHONDRIA ELONGATION CAUSED BY PHEVALIN TRE ATM E N
T............................................................I L L
4.9.1 MITOCHONDRIAL ATP PRODUCTION IS INCREASED IN PHEVALIN TREATED
CELLS.............................114
5. DISCUSSION............................................... 116
5.1 S. AUREUS TRANSPOSON MUTANT SCREEN REVEALS SEVERAL GENE PRODUCTS TO
BE INVOLVED IN
PHAGOSOMAL ESCAPE
...........................................
116
5.2 THE NON-RIBOSOMAL PEPTIDE SYNTHETASE AUSAB INVOLVED IN PHAGOSOMAL
ESCAPE OF S. AUREUS . 120
5.3 AUREUSIMINE DEFICIENCY DOES NOT AFFECT STAPHYLOCOCCAL GROWTH OR
INVASION INTO EPITHELIAL CELLS
BUT DOES REDUCE HOST CELL D EA TH
.....................................................................................................122
5.4 PHEVALIN BUT NOT TYRVALIN ACTS IN PHAGOSOMAL ESCAPE OF 5. AUREUS
...
.........................................123
5.5 PHEVALIN PRODUCTION PROMOTES
S. AUREUS INTRACELLULAR SURVIVAL AND CYTOTOXICITY IN LEUKOCYTES 125
5.6 THE 5. AUREUS NRPS INFLUENCES INFECTION OUTCOME IN A MURINE LUNG
INFECTION MODEL ................129
5.7 HOST CELL PROTEOME CHANGES AFTER PHEVALIN TREATMENT INDICATE DIRECT
EFFECT ON HOST CELL GENE
EXPRESSION.......................................................................................................................................
131
5.8 MITOCHONDRIAL ELONGATION BY PHEVALIN TRE ATM E N
T........................................................................133
5.9 CONCLUSION AND
OUTLOOK..................................................................................................................
136
6.
REFERENCES............................................................................................................................................138
7.
APPENDIX..............................................................................................................................................
169
7.1
ABBREVATIONS...................................................................................................................................169
7.2 LIST OF
FIGURES...................................................................................................................................171
7.3 LIST OF
TABLES....................................................................................................................................
172
7.4 SUPPLEMENTARY
INFORMATIONS..........................................................................................................173
7.5 PUBLICATIONS AND POSTER
PRESENTATIONS..........................................................................................
182
7.6
DANKSAGUNG.....................................................................................................................................
183
7.7 A
FFIDAVIT...........................................................................................................................................
184
7.8 CURRICULUM
VITAE..............................................................................................................................185
|
| any_adam_object | 1 |
| author | Blättner, Sebastian ca. 21. Jh |
| author_GND | (DE-588)1132839521 |
| author_facet | Blättner, Sebastian ca. 21. Jh |
| author_role | aut |
| author_sort | Blättner, Sebastian ca. 21. Jh |
| author_variant | s b sb |
| building | Verbundindex |
| bvnumber | BV044338062 |
| collection | ebook |
| ctrlnum | (OCoLC)1004520081 (DE-599)DNB1132730716 |
| dewey-full | 570 |
| dewey-hundreds | 500 - Natural sciences and mathematics |
| dewey-ones | 570 - Biology |
| dewey-raw | 570 |
| dewey-search | 570 |
| dewey-sort | 3570 |
| dewey-tens | 570 - Biology |
| discipline | Biologie |
| format | Thesis Book |
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| physical | 186 Seiten Illustrationen, Diagramme 30 cm |
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| publishDateSort | 2016 |
| record_format | marc |
| spelling | Blättner, Sebastian ca. 21. Jh. Verfasser (DE-588)1132839521 aut The role of the non-ribosomal peptide synthetase AusAB and its product phevalin in intracellular virulence of Staphylococcus aureus = Die Rolle der nicht-ribosomalen Peptidsynthetase AusAB und ihres Produktes Phevalin in der intrazellulären Virulenz von Staphylococcus aureus submitted by Sebastian Blättner fraom München Die Rolle der nicht-ribosomalen Peptidsynthetase AusAB und ihres Produktes Phevalin in der intrazellulären Virulenz von Staphylococcus aureus Würzburg 2016 186 Seiten Illustrationen, Diagramme 30 cm txt rdacontent n rdamedia nc rdacarrier Dissertation Julius-Maximilians-Universität Würzburg 2017 Zusammenfassung in deutscher und englischer Sprache Staphylococcus aureus (DE-588)4182912-8 gnd rswk-swf MRSA (DE-588)7501248-0 gnd rswk-swf Virulenz (DE-588)4129834-2 gnd rswk-swf (DE-588)4113937-9 Hochschulschrift gnd-content Staphylococcus aureus (DE-588)4182912-8 s Virulenz (DE-588)4129834-2 s MRSA (DE-588)7501248-0 s DE-604 Erscheint auch als Online-Ausgabe, PDF urn:nbn:de:bvb:20-opus-146662 https://nbn-resolving.org/urn:nbn:de:bvb:20-opus-146662 Resolving-System kostenfrei Volltext B:DE-101 application/pdf http://d-nb.info/1132730716/04 Resolving-System Inhaltsverzeichnis DNB Datenaustausch application/pdf http://bvbr.bib-bvb.de:8991/F?func=service&doc_library=BVB01&local_base=BVB01&doc_number=029741164&sequence=000001&line_number=0001&func_code=DB_RECORDS&service_type=MEDIA Inhaltsverzeichnis |
| spellingShingle | Blättner, Sebastian ca. 21. Jh The role of the non-ribosomal peptide synthetase AusAB and its product phevalin in intracellular virulence of Staphylococcus aureus = Die Rolle der nicht-ribosomalen Peptidsynthetase AusAB und ihres Produktes Phevalin in der intrazellulären Virulenz von Staphylococcus aureus Staphylococcus aureus (DE-588)4182912-8 gnd MRSA (DE-588)7501248-0 gnd Virulenz (DE-588)4129834-2 gnd |
| subject_GND | (DE-588)4182912-8 (DE-588)7501248-0 (DE-588)4129834-2 (DE-588)4113937-9 |
| title | The role of the non-ribosomal peptide synthetase AusAB and its product phevalin in intracellular virulence of Staphylococcus aureus = Die Rolle der nicht-ribosomalen Peptidsynthetase AusAB und ihres Produktes Phevalin in der intrazellulären Virulenz von Staphylococcus aureus |
| title_alt | Die Rolle der nicht-ribosomalen Peptidsynthetase AusAB und ihres Produktes Phevalin in der intrazellulären Virulenz von Staphylococcus aureus |
| title_auth | The role of the non-ribosomal peptide synthetase AusAB and its product phevalin in intracellular virulence of Staphylococcus aureus = Die Rolle der nicht-ribosomalen Peptidsynthetase AusAB und ihres Produktes Phevalin in der intrazellulären Virulenz von Staphylococcus aureus |
| title_exact_search | The role of the non-ribosomal peptide synthetase AusAB and its product phevalin in intracellular virulence of Staphylococcus aureus = Die Rolle der nicht-ribosomalen Peptidsynthetase AusAB und ihres Produktes Phevalin in der intrazellulären Virulenz von Staphylococcus aureus |
| title_full | The role of the non-ribosomal peptide synthetase AusAB and its product phevalin in intracellular virulence of Staphylococcus aureus = Die Rolle der nicht-ribosomalen Peptidsynthetase AusAB und ihres Produktes Phevalin in der intrazellulären Virulenz von Staphylococcus aureus submitted by Sebastian Blättner fraom München |
| title_fullStr | The role of the non-ribosomal peptide synthetase AusAB and its product phevalin in intracellular virulence of Staphylococcus aureus = Die Rolle der nicht-ribosomalen Peptidsynthetase AusAB und ihres Produktes Phevalin in der intrazellulären Virulenz von Staphylococcus aureus submitted by Sebastian Blättner fraom München |
| title_full_unstemmed | The role of the non-ribosomal peptide synthetase AusAB and its product phevalin in intracellular virulence of Staphylococcus aureus = Die Rolle der nicht-ribosomalen Peptidsynthetase AusAB und ihres Produktes Phevalin in der intrazellulären Virulenz von Staphylococcus aureus submitted by Sebastian Blättner fraom München |
| title_short | The role of the non-ribosomal peptide synthetase AusAB and its product phevalin in intracellular virulence of Staphylococcus aureus |
| title_sort | the role of the non ribosomal peptide synthetase ausab and its product phevalin in intracellular virulence of staphylococcus aureus die rolle der nicht ribosomalen peptidsynthetase ausab und ihres produktes phevalin in der intrazellularen virulenz von staphylococcus aureus |
| title_sub | = Die Rolle der nicht-ribosomalen Peptidsynthetase AusAB und ihres Produktes Phevalin in der intrazellulären Virulenz von Staphylococcus aureus |
| topic | Staphylococcus aureus (DE-588)4182912-8 gnd MRSA (DE-588)7501248-0 gnd Virulenz (DE-588)4129834-2 gnd |
| topic_facet | Staphylococcus aureus MRSA Virulenz Hochschulschrift |
| url | https://nbn-resolving.org/urn:nbn:de:bvb:20-opus-146662 http://d-nb.info/1132730716/04 http://bvbr.bib-bvb.de:8991/F?func=service&doc_library=BVB01&local_base=BVB01&doc_number=029741164&sequence=000001&line_number=0001&func_code=DB_RECORDS&service_type=MEDIA |
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