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Chapter 5 discussed in chapter 4. The reasons for the switch most likely relates to the fact that crystal violet staining stains all cellular biomass contained in the wells of the plates and therefore does not discriminate between viable and non-viable cells [231]. Alternative methods of using fluorescent stains to detect live cells [231] or both viable and non viable cells have been reported in recent years. Pitts et al. calculated percentage reduction of viable cells in a biofilm through measuring the absorbance of a cell respiratory stain (5-cyano-2,3-ditolyl tetrazolium chloride or CTC) post treatment with chlorine [231]. The percentage reduction in active metabolism was then used to evaluate the efficacy of the disinfectant treatments (reduction calculated from untreated wells – treated wells). This method was compared to the microtitre plate method (percentage removal of biomass) and enumeration of viable cells using the plate count method. The results suggest that there was an overall trend for all methods to display less viable biofilm cells (P. aeruginosa and S. epidermidis) after exposure to increased concentrations of chlorine. The percentage reduction using CTC corresponded with the mean log 10 reduction results achieved through use of the plate count method, while the crystal violet methods did not show a similar level of reduction [231]. As discussed in chapter 4, according to the CEN standards EN13697:2001, a 4log 10 reduction in viable count is necessary to demonstrate the efficacy of a disinfectant against a bacterial suspension attached to a surface [133]. Although the criteria that can be applied to assess disinfectant efficacy by percentage using LIVE/DEAD staining is not defined is not defined in the CEN standard EN13697:2001. Vestby et al. also investigated the use of synthetic furanone to inhibit biofilm formation using a microtitre plate based system [113]. They examined the effect of pre-exposure to furanone followed by treatment Page 161
Chapter 5 with a 0.05% concentration of sodium hypochlorite and 0.02% concentration of benzalkonium chloride after biofilm development. The effect of furanone was assessed by measuring the optical density of biofilm formed on a microtitre plate, while the effect of the disinfectants on the established biofilm was measured through the use of plate count method [113]. The results indicated that pre-exposure to furanone increased the efficacy of the disinfectant agents. There was a ~2 log 10 difference in the number of viable cells recovered following treatment with furanone and sodium hypochlorite in comparison to the disinfectant alone. There was also a ~1 log 10 difference in the number of cells recovered following treatment with furanone and benzalkonium chloride compared with the benzalkonium chloride alone [113]. However, even the combined treatment with furanone and sodium hypochlorite only reached a 4 log 10 reduction (log 10 density of biofilm recovered from the untreated surface– log 10 density of biofilm recovered from the treated surface) for 1 of the two strains of S. Agona examined [113]. Moreover, it is important to note that none of the agents used by Vestby et al. completely eliminated all viable cells from the biofilm [113]. Against this background the objectives in this chapter are to firstly examine if dense biofilm forming potential can be predicted using congo red agar and if this method can be used as an appropriate screening method to identify dense biofilm forming strains. Secondly, to investigate the use of the microtitre plate method to examine biofilm density of S. enterica strains and the ability of the strains to withstand disinfectant treatment using a microtitre plate based method. Page 162
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Salmonella enterica - biofilm forma
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Table of Contents Page Number 1.16.
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Table of Contents Page Number 4.1.2
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Table of Contents Page Number 6.8.4
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List of Abbreviations Table Page Nu
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List of Abbreviations List of Abbre
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Summary of Content Summary of Conte
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“To show your true ability is alw
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Acknowledgements ever met, Fiona th
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Acknowledgements This Ph.D. thesis
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Chapter 1 1.1. Salmonella Salmonell
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Chapter 1 may impose enormous costs
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Chapter 1 cracks or penetration of
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Chapter 1 detected by agglutination
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Chapter 1 become colonized with >10
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Chapter 1 antigens [z27],[z45] [1].
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Chapter 1 S. Agona has also been im
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Chapter 1 isolated from undercooked
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Chapter 1 The S. Typhimurium strain
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Chapter 1 ecosystem with the use of
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Chapter 1 [82]. Previous research h
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Chapter 1 to genes involved in flag
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Chapter 1 observation has also been
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Chapter 1 [119] and flow cell react
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Chapter 1 based disinfectant) the s
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Chapter 1 formation of two strains
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Chapter 1 shield the S. Agona cells
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Chapter 1 Key Objectives of this st
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Chapter 2 2. Abstract Food-borne pa
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Chapter 2 indicated that there may
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Chapter 2 taken from industrial set
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Chapter 2 2.1.4. CDC Biofilm Reacto
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Chapter 2 can be performed without
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Chapter 2 To summarise, previous au
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Chapter 2 2.3. Methods for examinin
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Chapter 2 and the biofilm formed th
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Chapter 2 2.4. Statistical Analysis
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Chapter 2 Table 2.1: Strain charact
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Chapter 2 XII. The gas port and med
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Chapter 2 II. Primary fixative cons
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Chapter 2 IX. Once the conditions w
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Chapter 2 Figure 2.1: Image of CBR
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Chapter 2 2.6. Results 2.6.1. Asses
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Chapter 2 2.6.3. SEM Analysis of su
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Chapter 2 Figure 2.6: Complete remo
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Chapter 2 2.6.5. Mean log 10 densit
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Chapter 2 2.6.6. Mean log 10 densit
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Chapter 2 Table 2.5: Mean log 10 de
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Chapter 2 The results displayed in
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Chapter 2 Table 2.7: Difference bet
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Chapter 2 Table 2.8: The mean log 1
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Chapter 2 Figure 2.7: Graph of the
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Chapter 2 also used “high” soni
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Chapter 2 2.8. Summary All 13 strai
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Chapter 3 3. Abstract It has been e
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Chapter 3 classified as persistent
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Chapter 3 was plated onto TSA for e
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Chapter 3 Figure 3.1: The mean log
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Chapter 3 Figure 3.3: SEM image of
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Chapter 3 Table 3.2: Difference bet
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Chapter 3 3.4.3. Intra-serovar vari
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Chapter 3 3.4.4. Strain variation i
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Chapter 3 3.4.5. The density of bio
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Chapter 3 3.5. Discussion As illust
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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 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
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Page 226 and 227: Chapter 6 undetermined. However, th
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Chapter 6 responsible for increased
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Chapter 6 not fully capture the ran
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Chapter 6 However, the extent of bi
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Chapter 6 6.10. Conclusions and rec
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Chapter 6 studies may solve some of
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Bibliography Bibliography Page 221
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Bibliography 12. T. Takata, J.L., H
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Bibliography 29. Barker, J., M. Nae
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Bibliography 47. Threlfall, E.J., H
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Bibliography 65. CDC. Investigation
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Bibliography 83. Ledeboer, N., Frye
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Bibliography 100. Chia, T., Goulter
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Bibliography 117. Buckingham-Meyer,
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Bibliography 133. European Committe
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Bibliography Supernatant of a Hafni
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Bibliography 165. Díez-García, M.
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Bibliography cultivation of Staphyl
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Bibliography Compounds. 2012. Appli
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Bibliography 215. Buck, et al., A q
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Bibliography Canadian Journal of Ps
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Appendix 1 Appendix 1— List of Re
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Appendix 1 Appendix 1 - List of Equ
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Appendix 2 Colour index High outlie
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Appendix 3 Appendix 3-Figure 2: PFG
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Appendix 4 Poster Presentation Envi
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