View/Open - ARAN - National University of Ireland, Galway

More documents

Recommendations

Info

Chapter 2 2.3. Methods for examining biofilm density and substrata 2.3.1. Atomic Force Microscopy (AFM) Atomic force microscopy uses a scanning cantilever or probe that taps the surface of a specimen on contact. The probe then gently scans the surface and based on tapping of the surface encountered. A laser beam is aimed at the outer area of the probe. The electrons emitted from the laser are deflected back to a photo-detector which deciphers the distance of the probe from the surface at each scanned point to measure surface roughness of the overall sample [172] . 2.3.2. Scanning Electron Microscopy (SEM) SEM is a widely used method for examining biofilm on surfaces [114, 122, 173-174]. SEM has previously been used and standardised by Williams and colleagues to confirm biofilm growth of Staphylococcus epidermidis established using the CBR [174]. Gilmore et al. also used SEM analysis to confirm dense biofilm formation by Proteus mirabilis using the CBR [122]. The basic principle of SEM is that a beam of electrons are emitted from an electrode gun and accelerated at high voltage (e.g. 20 kiloVolts - kV) onto a specimen. Once the beam of electrons hits the specimen, the electrons rebound off the surface and are collected through the secondary electron imaging (SEI) collector [175]. The SEI collector deciphers the electrons collected and coverts them to digital analogs or signals. The signals are then converted to grey-scale pixilation using computer programming [175]. In order to optimize conditions for SEM visualization, the specimens are fixed (primary and secondary fixatives) which preserve the cellular details of the biofilm [174]. After fixation the specimens are dehydrated in a series of ethanol to remove excess water from the sample [175]. Any water remaining may destroy the sample if overheated in the SEM chamber. The ethanol series is followed by the addition of a critical point dryer to eliminate surface tension from liquids on the surface of the specimen. Hexamethyldisilizane (HMDS) is now recognised as a safer option for Page 49
Chapter 2 critical point drying due to safety concerns of critical point drying chambers (due to over heating or excessive pressure). After drying the specimens are coated in a thin layer of gold (or other conductive materials). This allows the specimen to emit the electrons when they hit off the surface and therefore be collected by the detector. The use of gold coating has been demonstrated to produce sharper images of biofilm cells compared with specimens that are not coated. The specimens are also mounted onto carbon double sided stickers and metal stubs to prevent movement during the process (movement could be caused by the vacuum created) the carbon and metal also provide conductivity. 2.3.3. Other methods for examining and enumerating biofilm cells A number of authors have reported on the use staining methods such as LIVE⁄DEAD® biofilm viability kits with confocal laser scanning microscopy (CLSM) and computer programmes to measure biofilm density relative to the surface area of the biofilm substrata [123, 176-178]. The LIVE/DEAD® stain contains the nucleic acid binding dyes SYTO® 9 green and propidium iodide. When used in combination, the SYTO® 9 green attaches to all cells while the propidium iodide penetrates disrupted cell membranes (nonviable cells). Hadi et al. described the use of LIVE/DEAD® stain to examine the percentage coverage P. aeruginosa biofilm formed on polytetrafluoroethylene (plastic) coupons developed using the CBR [123]. The use of LIVE⁄DEAD® stain [176-177] and SYTO® 9 green [178] to investigate biofilm formation using hydroxyapatite (dental ceramic) disks, glass flow-cells and microtitre plate models has also been reported elsewhere. However due to the surface properties of the coupons used in this thesis, in particular polycarbonate, concrete and tile, there was a high level of autofluorescence, when examined using confocal laser scanning microscopy. Autofluorescence of the surface may result in inaccurate measurement of the biofilm or lack of discrimination between the surface Page 50
Page 1:
Salmonella enterica - biofilm forma
Page 4 and 5:
Table of Contents Page Number 1.16.
Page 6 and 7:
Table of Contents Page Number 4.1.2
Page 8 and 9:
Table of Contents Page Number 6.8.4
Page 10 and 11:
List of Abbreviations Table Page Nu
Page 12 and 13:
List of Abbreviations List of Abbre
Page 14 and 15:
Summary of Content Summary of Conte
Page 16 and 17:
“To show your true ability is alw
Page 18 and 19:
Acknowledgements ever met, Fiona th
Page 20 and 21: Acknowledgements This Ph.D. thesis
Page 22 and 23: Chapter 1 1.1. Salmonella Salmonell
Page 24 and 25: Chapter 1 may impose enormous costs
Page 26 and 27: Chapter 1 cracks or penetration of
Page 28 and 29: Chapter 1 detected by agglutination
Page 30 and 31: Chapter 1 become colonized with >10
Page 32 and 33: Chapter 1 antigens [z27],[z45] [1].
Page 34 and 35: Chapter 1 S. Agona has also been im
Page 36 and 37: Chapter 1 isolated from undercooked
Page 38 and 39: Chapter 1 The S. Typhimurium strain
Page 40 and 41: Chapter 1 ecosystem with the use of
Page 42 and 43: Chapter 1 [82]. Previous research h
Page 44 and 45: Chapter 1 to genes involved in flag
Page 46 and 47: Chapter 1 observation has also been
Page 48 and 49: Chapter 1 [119] and flow cell react
Page 50 and 51: Chapter 1 based disinfectant) the s
Page 52 and 53: Chapter 1 formation of two strains
Page 54 and 55: Chapter 1 shield the S. Agona cells
Page 56 and 57: Chapter 1 Key Objectives of this st
Page 58 and 59: Chapter 2 2. Abstract Food-borne pa
Page 60 and 61: Chapter 2 indicated that there may
Page 62 and 63: Chapter 2 taken from industrial set
Page 64 and 65: Chapter 2 2.1.4. CDC Biofilm Reacto
Page 66 and 67: Chapter 2 can be performed without
Page 68 and 69: Chapter 2 To summarise, previous au
Page 72 and 73: Chapter 2 and the biofilm formed th
Page 74 and 75: Chapter 2 2.4. Statistical Analysis
Page 76 and 77: Chapter 2 Table 2.1: Strain charact
Page 78 and 79: Chapter 2 XII. The gas port and med
Page 80 and 81: Chapter 2 II. Primary fixative cons
Page 82 and 83: Chapter 2 IX. Once the conditions w
Page 84 and 85: Chapter 2 Figure 2.1: Image of CBR
Page 86 and 87: Chapter 2 2.6. Results 2.6.1. Asses
Page 88 and 89: Chapter 2 2.6.3. SEM Analysis of su
Page 90 and 91: Chapter 2 Figure 2.6: Complete remo
Page 92 and 93: Chapter 2 2.6.5. Mean log 10 densit
Page 94 and 95: Chapter 2 2.6.6. Mean log 10 densit
Page 96 and 97: Chapter 2 Table 2.5: Mean log 10 de
Page 98 and 99: Chapter 2 The results displayed in
Page 100 and 101: Chapter 2 Table 2.7: Difference bet
Page 102 and 103: Chapter 2 Table 2.8: The mean log 1
Page 104 and 105: Chapter 2 Figure 2.7: Graph of the
Page 106 and 107: Chapter 2 also used “high” soni
Page 108 and 109: Chapter 2 2.8. Summary All 13 strai
Page 110 and 111: Chapter 3 3. Abstract It has been e
Page 112 and 113: Chapter 3 classified as persistent
Page 114 and 115: Chapter 3 was plated onto TSA for e
Page 116 and 117: Chapter 3 Figure 3.1: The mean log
Page 118 and 119: Chapter 3 Figure 3.3: SEM image of
Page 120 and 121:
Chapter 3 Table 3.2: Difference bet
Page 122 and 123:
Chapter 3 3.4.3. Intra-serovar vari
Page 124 and 125:
Chapter 3 3.4.4. Strain variation i
Page 126 and 127:
Chapter 3 3.4.5. The density of bio
Page 128 and 129:
Chapter 3 3.5. Discussion As illust
Page 130 and 131:
Chapter 3 As discussed in section 3
Page 132 and 133:
Chapter 3 increased biofilm appeare
Page 134 and 135:
Chapter 4 Examining the efficacy of
Page 136 and 137:
Chapter 4 through laboratory based
Page 138 and 139:
Chapter 4 acid and quaternary ammon
Page 140 and 141:
Chapter 4 the surface the temperatu
Page 142 and 143:
Chapter 4 availability of multiple
Page 144 and 145:
Chapter 4 peracetic acid (100, 200,
Page 146 and 147:
Chapter 4 As discussed previously,
Page 148 and 149:
Chapter 4 Dey/Engley (Difco) neutra
Page 150 and 151:
Chapter 4 sterilisation was achieve
Page 152 and 153:
Chapter 4 IX. Aliquots of 100µl of
Page 154 and 155:
Chapter 4 XV. XVI. XVII. XVIII. XIX
Page 156 and 157:
Chapter 4 4.3. Results 4.3.1. Suspe
Page 158 and 159:
Chapter 4 Table 4.3: Mean log 10 de
Page 160 and 161:
Chapter 4 Benzalkonium chloride was
Page 162 and 163:
Chapter 4 4.4. Discussion Suspensio
Page 164 and 165:
Chapter 4 effective at eliminating
Page 166 and 167:
Chapter 4 Surprisingly, Nguyen et a
Page 168 and 169:
Chapter 4 though a contact time of
Page 170 and 171:
Chapter 4 provide a more standardis
Page 172 and 173:
Chapter 4 hours instead of the stan
Page 174 and 175:
Chapter 5 5. Abstract Examining bio
Page 176 and 177:
Chapter 5 dry and rough (bdar) morp
Page 178 and 179:
Chapter 5 absence of curli and cell
Page 180 and 181:
Chapter 5 studies such as the micro
Page 182 and 183:
Chapter 5 discussed in chapter 4. T
Page 184 and 185:
Chapter 5 5.2. Methods 5.2.1. Colon
Page 186 and 187:
Chapter 5 XIII. XIV. XV. XVI. XVII.
Page 188 and 189:
Chapter 5 XVI. XVII. XVIII. XIX. XX
Page 190 and 191:
Chapter 5 5.3.2. Repeatability of m
Page 192 and 193:
Chapter 5 Figure 5.2: Stained biofi
Page 194 and 195:
Chapter 5 was formed at room temper
Page 196 and 197:
Chapter 5 Table 5.3: The density of
Page 198 and 199:
Chapter 5 5.3.4. The density of bio
Page 200 and 201:
Chapter 5 Table 5.6: The difference
Page 202 and 203:
Chapter 5 Table 5.7: The difference
Page 204 and 205:
Chapter 5 Table 5.8: Summary of ass
Page 206 and 207:
Chapter 5 interpretation therefore
Page 208 and 209:
Chapter 5 measures of S. enterica b
Page 210 and 211:
Chapter 5 cells peaks [165, 191]. D
Page 212 and 213:
Chapter 5 5.5. Limitations of the s
Page 214 and 215:
Chapter 6 Discussion of S. enterica
Page 216 and 217:
Chapter 6 However, a number of auth
Page 218 and 219:
Chapter 6 It is of interest to exam
Page 220 and 221:
Chapter 6 were similar at 48 hours.
Page 222 and 223:
Chapter 6 attributable to a small n
Page 224 and 225:
Chapter 6 chosen as an appropriate
Page 226 and 227:
Chapter 6 undetermined. However, th
Page 228 and 229:
Chapter 6 reason for this may be du
Page 230 and 231:
Chapter 6 Moreover, based on the re
Page 232 and 233:
Chapter 6 responsible for increased
Page 234 and 235:
Chapter 6 not fully capture the ran
Page 236 and 237:
Chapter 6 However, the extent of bi
Page 238 and 239:
Chapter 6 6.10. Conclusions and rec
Page 240 and 241:
Chapter 6 studies may solve some of
Page 242 and 243:
Bibliography Bibliography Page 221
Page 244 and 245:
Bibliography 12. T. Takata, J.L., H
Page 246 and 247:
Bibliography 29. Barker, J., M. Nae
Page 248 and 249:
Bibliography 47. Threlfall, E.J., H
Page 250 and 251:
Bibliography 65. CDC. Investigation
Page 252 and 253:
Bibliography 83. Ledeboer, N., Frye
Page 254 and 255:
Bibliography 100. Chia, T., Goulter
Page 256 and 257:
Bibliography 117. Buckingham-Meyer,
Page 258 and 259:
Bibliography 133. European Committe
Page 260 and 261:
Bibliography Supernatant of a Hafni
Page 262 and 263:
Bibliography 165. Díez-García, M.
Page 264 and 265:
Bibliography cultivation of Staphyl
Page 266 and 267:
Bibliography Compounds. 2012. Appli
Page 268 and 269:
Bibliography 215. Buck, et al., A q
Page 270 and 271:
Bibliography Canadian Journal of Ps
Page 272 and 273:
Appendix 1 Appendix 1— List of Re
Page 274 and 275:
Appendix 1 Appendix 1 - List of Equ
Page 276 and 277:
Appendix 2 Colour index High outlie
Page 278 and 279:
Page 280 and 281:
Page 282 and 283:
Page 284 and 285:
Page 286 and 287:
Appendix 3 Appendix 3-Figure 2: PFG
Page 288 and 289:
Appendix 4 Poster Presentation Envi
show all

View/Open - ARAN - National University of Ireland, Galway

You also want an ePaper? Increase the reach of your titles

Delete template?

Save as template?