Development of a simple in vitro test system for the disinfection of bacterial biofilms

Päivi Härkönen, Satu Salo, Tiina Mattila-Sandholm, Gun Wirtanen, D. Allison, P. Gilbert

Research output: Contribution to journalArticleScientificpeer-review

18 Citations (Scopus)

Abstract

This paper evaluates the use of poloxamer-hydrogel biofilm-constructs for the routine evaluation of biocide formulations at use strength. Aqueous solutions of Poloxamer P127 show thermo-reversible gelation, being liquid at temperatures <15°C yet robust gels at temperatures >15°C. Chilled poloxamer solutions (30%w/v) were made up m tryptone soya broth and inoculated with c104–5 cfu/ml of stationary phase cultures of eleven foodbome spoilage and pathogen test-strains, including Listeria and Salmonella spp. Drops (200µl) were placed, in triplicate, onto pre-warmed, sterile stainless steel discs (8mm diam.) held in sealed-Petn dishes. These were incubated for 5h at 30 °C during which time all strains grew well giving between 106–7 cfu/ml. Previous work had demonstrated that cells grown within poloxamer hydrogels m this manner adopted biofilm-phenotypes which were distinct from those of planktontcally grown cells and which were characteristically resistant to a range of bioctdes. Incubated poloxamer gels, together with their supports were transferred to solutions (IOrnl) of four commercial biocide formulations: (t) a buffered (pH 10) tertiary alkyl amine formulation in an amphoteric surfactant (TARS), (ii) a hydrogen peroxide, peracefc acid, acetic acrd (pH 1) formulation (HPPA), (iii) sodium hypochlorite and sodium hydroxide formulation (pil 13) (HYPO), and (iv) isopropyl alcohol in lactic acid (pH 2.3) (IPA). After 5min the test pieces were removed and transferred to neutralizer at 15°C. The gels dispersed rapidly releasing the cells and enabling viable counts to be made. All formulations effected a >5-log kill of planktonic challenges within smin whilst the levels of killing effected within the biofilm-constructs varied between 0.5 and 3-log reductions. The results were highly reproducible with patterns of susceptibility varying both as a function of organism, biocide-type and concentration. The experiments strongly support the view that poloxamer constructs might find application m trials and testing of new disinfectant formulations.
Original languageEnglish
Pages (from-to)219-225
Number of pages7
JournalWater Science and Technology
Volume39
Issue number7
DOIs
Publication statusPublished - 1999
MoE publication typeA1 Journal article-refereed

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Biocides
Disinfection
Biofilms
disinfection
biofilm
pesticide
Hydrogels
Amphoteric surfactants
Gels
gel
Sodium
sodium
Spoilage
Listeria
Disinfectants
Salmonella
Pathogens
Gelation
Lactic acid
Acetic acid

Cite this

Härkönen, Päivi ; Salo, Satu ; Mattila-Sandholm, Tiina ; Wirtanen, Gun ; Allison, D. ; Gilbert, P. / Development of a simple in vitro test system for the disinfection of bacterial biofilms. In: Water Science and Technology. 1999 ; Vol. 39, No. 7. pp. 219-225.
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title = "Development of a simple in vitro test system for the disinfection of bacterial biofilms",
abstract = "This paper evaluates the use of poloxamer-hydrogel biofilm-constructs for the routine evaluation of biocide formulations at use strength. Aqueous solutions of Poloxamer P127 show thermo-reversible gelation, being liquid at temperatures <15°C yet robust gels at temperatures >15°C. Chilled poloxamer solutions (30{\%}w/v) were made up m tryptone soya broth and inoculated with c104–5 cfu/ml of stationary phase cultures of eleven foodbome spoilage and pathogen test-strains, including Listeria and Salmonella spp. Drops (200µl) were placed, in triplicate, onto pre-warmed, sterile stainless steel discs (8mm diam.) held in sealed-Petn dishes. These were incubated for 5h at 30 °C during which time all strains grew well giving between 106–7 cfu/ml. Previous work had demonstrated that cells grown within poloxamer hydrogels m this manner adopted biofilm-phenotypes which were distinct from those of planktontcally grown cells and which were characteristically resistant to a range of bioctdes. Incubated poloxamer gels, together with their supports were transferred to solutions (IOrnl) of four commercial biocide formulations: (t) a buffered (pH 10) tertiary alkyl amine formulation in an amphoteric surfactant (TARS), (ii) a hydrogen peroxide, peracefc acid, acetic acrd (pH 1) formulation (HPPA), (iii) sodium hypochlorite and sodium hydroxide formulation (pil 13) (HYPO), and (iv) isopropyl alcohol in lactic acid (pH 2.3) (IPA). After 5min the test pieces were removed and transferred to neutralizer at 15°C. The gels dispersed rapidly releasing the cells and enabling viable counts to be made. All formulations effected a >5-log kill of planktonic challenges within smin whilst the levels of killing effected within the biofilm-constructs varied between 0.5 and 3-log reductions. The results were highly reproducible with patterns of susceptibility varying both as a function of organism, biocide-type and concentration. The experiments strongly support the view that poloxamer constructs might find application m trials and testing of new disinfectant formulations.",
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Härkönen, P, Salo, S, Mattila-Sandholm, T, Wirtanen, G, Allison, D & Gilbert, P 1999, 'Development of a simple in vitro test system for the disinfection of bacterial biofilms', Water Science and Technology, vol. 39, no. 7, pp. 219-225. https://doi.org/10.1016/S0273-1223(99)00171-7

Development of a simple in vitro test system for the disinfection of bacterial biofilms. / Härkönen, Päivi; Salo, Satu; Mattila-Sandholm, Tiina; Wirtanen, Gun; Allison, D.; Gilbert, P.

In: Water Science and Technology, Vol. 39, No. 7, 1999, p. 219-225.

Research output: Contribution to journalArticleScientificpeer-review

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T1 - Development of a simple in vitro test system for the disinfection of bacterial biofilms

AU - Härkönen, Päivi

AU - Salo, Satu

AU - Mattila-Sandholm, Tiina

AU - Wirtanen, Gun

AU - Allison, D.

AU - Gilbert, P.

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N2 - This paper evaluates the use of poloxamer-hydrogel biofilm-constructs for the routine evaluation of biocide formulations at use strength. Aqueous solutions of Poloxamer P127 show thermo-reversible gelation, being liquid at temperatures <15°C yet robust gels at temperatures >15°C. Chilled poloxamer solutions (30%w/v) were made up m tryptone soya broth and inoculated with c104–5 cfu/ml of stationary phase cultures of eleven foodbome spoilage and pathogen test-strains, including Listeria and Salmonella spp. Drops (200µl) were placed, in triplicate, onto pre-warmed, sterile stainless steel discs (8mm diam.) held in sealed-Petn dishes. These were incubated for 5h at 30 °C during which time all strains grew well giving between 106–7 cfu/ml. Previous work had demonstrated that cells grown within poloxamer hydrogels m this manner adopted biofilm-phenotypes which were distinct from those of planktontcally grown cells and which were characteristically resistant to a range of bioctdes. Incubated poloxamer gels, together with their supports were transferred to solutions (IOrnl) of four commercial biocide formulations: (t) a buffered (pH 10) tertiary alkyl amine formulation in an amphoteric surfactant (TARS), (ii) a hydrogen peroxide, peracefc acid, acetic acrd (pH 1) formulation (HPPA), (iii) sodium hypochlorite and sodium hydroxide formulation (pil 13) (HYPO), and (iv) isopropyl alcohol in lactic acid (pH 2.3) (IPA). After 5min the test pieces were removed and transferred to neutralizer at 15°C. The gels dispersed rapidly releasing the cells and enabling viable counts to be made. All formulations effected a >5-log kill of planktonic challenges within smin whilst the levels of killing effected within the biofilm-constructs varied between 0.5 and 3-log reductions. The results were highly reproducible with patterns of susceptibility varying both as a function of organism, biocide-type and concentration. The experiments strongly support the view that poloxamer constructs might find application m trials and testing of new disinfectant formulations.

AB - This paper evaluates the use of poloxamer-hydrogel biofilm-constructs for the routine evaluation of biocide formulations at use strength. Aqueous solutions of Poloxamer P127 show thermo-reversible gelation, being liquid at temperatures <15°C yet robust gels at temperatures >15°C. Chilled poloxamer solutions (30%w/v) were made up m tryptone soya broth and inoculated with c104–5 cfu/ml of stationary phase cultures of eleven foodbome spoilage and pathogen test-strains, including Listeria and Salmonella spp. Drops (200µl) were placed, in triplicate, onto pre-warmed, sterile stainless steel discs (8mm diam.) held in sealed-Petn dishes. These were incubated for 5h at 30 °C during which time all strains grew well giving between 106–7 cfu/ml. Previous work had demonstrated that cells grown within poloxamer hydrogels m this manner adopted biofilm-phenotypes which were distinct from those of planktontcally grown cells and which were characteristically resistant to a range of bioctdes. Incubated poloxamer gels, together with their supports were transferred to solutions (IOrnl) of four commercial biocide formulations: (t) a buffered (pH 10) tertiary alkyl amine formulation in an amphoteric surfactant (TARS), (ii) a hydrogen peroxide, peracefc acid, acetic acrd (pH 1) formulation (HPPA), (iii) sodium hypochlorite and sodium hydroxide formulation (pil 13) (HYPO), and (iv) isopropyl alcohol in lactic acid (pH 2.3) (IPA). After 5min the test pieces were removed and transferred to neutralizer at 15°C. The gels dispersed rapidly releasing the cells and enabling viable counts to be made. All formulations effected a >5-log kill of planktonic challenges within smin whilst the levels of killing effected within the biofilm-constructs varied between 0.5 and 3-log reductions. The results were highly reproducible with patterns of susceptibility varying both as a function of organism, biocide-type and concentration. The experiments strongly support the view that poloxamer constructs might find application m trials and testing of new disinfectant formulations.

U2 - 10.1016/S0273-1223(99)00171-7

DO - 10.1016/S0273-1223(99)00171-7

M3 - Article

VL - 39

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EP - 225

JO - Water Science and Technology

JF - Water Science and Technology

SN - 0273-1223

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