The transcriptional programme of Salmonella enterica serovar Typhimurium reveals a key role for tryptophan metabolism in biofilms

<p>Abstract</p> <p>Background</p> <p>Biofilm formation enhances the capacity of pathogenic <it>Salmonella </it>bacteria to survive stresses that are commonly encountered within food processing and during host infection. The persistence of <it>Salmonella </it>within the food chain has become a major health concern, as biofilms can serve as a reservoir for the contamination of food products. While the molecular mechanisms required for the survival of bacteria on surfaces are not fully understood, transcriptional studies of other bacteria have demonstrated that biofilm growth triggers the expression of specific sets of genes, compared with planktonic cells. Until now, most gene expression studies of <it>Salmonella </it>have focused on the effect of infection-relevant stressors on virulence or the comparison of mutant and wild-type bacteria. However little is known about the physiological responses taking place inside a <it>Salmonella </it>biofilm.</p> <p>Results</p> <p>We have determined the transcriptomic and proteomic profiles of biofilms of <it>Salmonella enterica </it>serovar Typhimurium. We discovered that 124 detectable proteins were differentially expressed in the biofilm compared with planktonic cells, and that 10% of the <it>S</it>. Typhimurium genome (433 genes) showed a 2-fold or more change in the biofilm compared with planktonic cells. The genes that were significantly up-regulated implicated certain cellular processes in biofilm development including amino acid metabolism, cell motility, global regulation and tolerance to stress. We found that the most highly down-regulated genes in the biofilm were located on <it>Salmonella </it>Pathogenicity Island 2 (SPI2), and that a functional SPI2 secretion system regulator (<it>ssrA</it>) was required for <it>S</it>. Typhimurium biofilm formation. We identified STM0341 as a gene of unknown function that was needed for biofilm growth. Genes involved in tryptophan (<it>trp</it>) biosynthesis and transport were up-regulated in the biofilm. Deletion of <it>trpE </it>led to decreased bacterial attachment and this biofilm defect was restored by exogenous tryptophan or indole.</p> <p>Conclusions</p> <p>Biofilm growth of <it>S</it>. Typhimurium causes distinct changes in gene and protein expression. Our results show that aromatic amino acids make an important contribution to biofilm formation and reveal a link between SPI2 expression and surface-associated growth in <it>S</it>. Typhimurium.</p

Influence of flow on the structure of bacterial biofilms.

Bacteria attached to surfaces in biofilms are responsible for the contamination of industrial processes and many types of microbial infections and disease. Once established, biofilms are notoriously difficult to eradicate. A more complete understanding of how biofilms form and behave is crucial if we are to predict, and ultimately control, biofilm processes. A major breakthrough in biofilm research came in the early 1990’s when confocal scanning laser microscopy (CSLM) showed that biofilms formed complex structures which could facilitate nutrient exchange. We have recently found that biofilms growing in turbulent flow can also be temporally complex. Structures such as cell clusters and ripples can migrate downstream along solid surfaces. Further, biofilm viscoelasticity allows the biofilm to structurally deform when exposed to varying shear stresses

Limits to growth and what keeps a biofilm finite

Two of the factors, shear erosion and diffusive mass transfer, which limit the growth of heterogeneous biofilms are considered. For permeable beds of particulates, with a regulated throughflow, equating shear induced erosion and biofilm growth, leads to estimates of biofilm thickness and activity which conform with experimental measurements. In the more open environments of pipes and channels, increased thickness of biofilm is not directly balanced by increased cell erosion from the biofilm surface. However increasing thickness leads to growth limitations as diffusion limits the rate of mass transfer to cells deep in the film. For heterogeneous biofilms , consisting of complex clusters intersected by channels, mass transfer into the biofilm is by a combination of advective flow in the channels and diffusive transfer in clusters. In this paper we have considered mass transfer into simplified cluster forms, that is cylinders and hemispheres. Using the concept of critical dimension we have explored some of the implications of these simplified structures. We discuss the limitation to this approach as fluid shear alters the form of these simplified clusters. The viscoelastic properties of the biofilm clusters are being investigated and should allow better prediction of the effect of lateral shear on simple forms. The advection in biofilm channels and the related mass transfer processes needs further investigation

Consensus model of biofilm structure

Biofilms have been defined in various ways by various researchers. The definition is usually structured to be all inclusive of the many environments that biofilms are found and disciplines that the subject covers. Characklis and Marshall (1990) define a biofilm as consisting of “cells immobilized at a substratum and frequently embedded in an organic polymer matrix of microbial origin”. A broader definition is supplied by Costerton et al. (1995) who defined biofilms as “matrix-enclosed bacterial populations adherent to each other and/or to surfaces or interfaces”. It might be easiest to define biofilms in terms of what they are not - single cells homogeneously dispersed in fluid, the well mixed batch culture of which much of contemporary microbiology is based. Structural organisation is a characteristic feature of biofilms which distinguishes biofilm cultures from conventional suspended cultures, with or without an association with an interface. Biofilm structure is a recurrent topic of discussion among biofilm researchers generally and has been featured in a number of presentations at the first two British Biofilm Club Gregynog meetings. Much discussion time has been spent in search of a “universal” conceptual biofilm model describing biofilm structure (Handley 1995). The existence of such a model is appealing but given the enormous diversity of biofilms is it possible to characterise all biofilms with a single conceptual model? And if we do agree on a working model how useful will such a model be? Possibly we should not restrict a biofilm model to certain structural constraints but instead look for common features or basic building blocks of biofilms which could be readily incorporated into different structural models in a modular fashion

Influence of surfaces on sulphidogenic bacteria

Sulphidogenic bacteria in oil reservoirs are of great economic importance in terms of souring, fouling and corrosion. Mixed cultures containing these bacteria were isolated from chalk formations in North Sea oil reservoirs. These were thermophilic cultures, growing optimally at 60°C. Oil formations are porous matrices, providing a very large surface area and ideal conditions for bacterial attachment, survival and growth. This study included assessments of sulphide production rates of thermophilic (t-)sulphidogen consortia with and without additional surfaces. The availability of a surface contributed significantly to the rate and extent of sulphide generation. Surfaces were offered in varying amounts to growing planktonic cultures: significantly more sulphide was produced from cultures in contact with a surface than from identical cultures in the absence of a surface. In another series of experiments, t-sulphidogens were added to chalk rock chips in the presence of nutrients and incubated for several months. This resulted in rapid sulphide generation, the final concentration being related to the initial nutrient concentration. Subsequent nutrient addition resulted in renewed sulphide generation. It is suggested that bacteria in reservoirs can withstand long periods of nutrient deprivation while attached within the porous rock matrix and opportunistically utilise nutrients when they become available