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Open Access Methodology article

Method to grow Actinobacillus pleuropneumoniae biofilm on a biotic surface

Yannick DN Tremblay1, Cynthia Lévesque1, Ruud PAM Segers2 and Mario Jacques1*

Author Affiliations

1 Groupe de recherche sur les maladies infectieuses du porc, Faculté de médecine vétérinaire, Université de Montréal, 3200 rue Sicotte, St-Hyacinthe, Québec J2S 7C6, Canada

2 Microbiological R&D, MSD Animal Health, 5831 AN, Boxmeer, The Netherlands

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BMC Veterinary Research 2013, 9:213  doi:10.1186/1746-6148-9-213

Published: 20 October 2013

Abstract

Background

Actinobacillus pleuropneumoniae is a Gram-negative bacterium and a member of the Pasteurellaceae family. This bacterium is the causative agent of porcine pleuropneumonia, which is a highly contagious respiratory disease causing important economical losses to the worldwide pig industry. It has been shown that A. pleuropneumoniae can form biofilms on abiotic surfaces (plastic and glass). Although in vitro models are extremely useful to gain information on biofilm formation, these models may not be representative of the conditions found at the mucosal surface of the host, which is the natural niche of A. pleuropneumoniae.

Results

In this paper, we describe a method to grow A. pleuropneumoniae biofilms on the SJPL cell line, which represents a biotic surface. A non-hemolytic, non-cytotoxic mutant of A. pleuropneumoniae was used in our assays and this allowed the SJPL cell monolayers to be exposed to A. pleuropneumoniae for longer periods. This resulted in the formation of biofilms on the cell monolayer after incubations of 24 and 48 h. The biofilms can be stained with fluorescent probes, such as a lectin against the polymer of N-acetyl-D-glucosamine present in the biofilm matrix, and easily observed by confocal laser scanning microscopy.

Conclusions

This is the first protocol that describes the formation of an A. pleuropneumoniae biofilm on a biotic surface. The advantage of this protocol is that it can be used to study biofilm formation in a context of host-pathogen interactions. The protocol could also be adapted to evaluate biofilm inhibitors or the efficacy of antibiotics in the presence of biofilms.

Keywords:
Biofilm; Biotic surface; SJPL cell line; Actinobacillus pleuropneumoniae; Confocal laser scanning microscopy; Host-pathogen interaction