Bacteriophage Isolated from Sewage Eliminates and Prevents the Establishment of Escherichia Coli Biofilm.

Roberto Sousa Dias,Leandro Licursi De Oliveira,Sergio Oliveira De Paula,Karla Veloso Gonçalves Ribeiro,Cleberson Ribeiro,Silvia Almeida Cardoso,Jose Cola Zanuncio

doi:10.15171/apb.2018.011

Abstract

Purpose: Biofilm growth exerts a negative impact on industry and health, necessitating the development of strategies to control. The objective of this work was study the lytic activity of the phage isolated from the sewage network in the formation and degradation of Escherichia coli biofilms.Methods: E. coli cultures were incubated in 96-well polystyrene microplates under controlled conditions to evaluate the biofilm formation. The E. coli cultures and established biofilms were treated with the suspensions of the vB_EcoM-UFV017 (EcoM017) bacteriophage obtained from sewage for 24 hours. The E. coli bacterial density was measured using absorbance at 600 nm and the biofilms were measured by crystal violet staining. Polystyrene coupons were used as support for Scanning Electron Microscopy and Confocal Microscopy to evaluate biofilm formation.Results: The E. coli strains formed biofilms in polystyrene microplates after 48 hours’ incubation. The highest EcoM017 phage titer, in the prevention and degradation experiments, reduced the bacterial growth and the quantity of biofilm formed by E. coli in 90.0% and 87.5%, respectively. The minimum dose capable of reducing the biofilms of this bacterium was 101 PFU/mL after 24 hours. The preformed E. coli biofilm mass was reduced 79% post exposure to the phage in the degradation assay. Microscopic analysis confirmed the results obtained in the plates assays.Conclusion: The EcoM017 phage prevented biofilm formation and degraded the E. coli-established ones. The EcoM017 phage isolated from sewage can reduce bacterial attachment and lyse the E. coli associated biofilm cells, offering biotechnological potential applicability for this phage.

Highlights

Escherichia coli, one of the main components of the normal flora of the digestive tract of humans and animals,[1] are usually harmless, some strains are pathogenic and can cause intra- and extra-intestinal diseases.[2]
It has been shown that E. coli 30 can form more biofilm when grown in media with enrofloxacin, which is commonly used in the treatment of E. coli bovine mastitis, showing the importance to develop alternatives to the treatment of mastitis caused by this isolate.[23]
This was reported for a phage mixture where Escherichia coli O157:H7 biofilms formed on different surfaces, times and temperatures showed pronounced decrease in its biofilm after addition of phage mixture.[34]

Summary

Introduction

Escherichia coli, one of the main components of the normal flora of the digestive tract of humans and animals,[1] are usually harmless, some strains are pathogenic and can cause intra- and extra-intestinal diseases.[2]. E. coli is the most important and frequently agent isolated from the clinical cases of environmental mastitis causing milk production losses and the death of 10% of the animals.[6,7] This organism is capable of causing clinical mastitis depending on the immunological status of animal.[8]

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