In traditional models of in vitro biofilm development, individual bacterial cells seed a surface, multiply, and mature into multicellular, three-dimensional structures. Much research has been devoted to elucidating the mechanisms governing the initial attachment of single cells to surfaces. However, in natural environments and during infection, bacterial cells tend to clump as multicellular aggregates, and biofilms can also slough off aggregates as a part of the dispersal process. This makes it likely that biofilms are often seeded by aggregates and single cells, yet how these aggregates impact biofilm initiation and development is not known. Here we use a combination of experimental and computational approaches to determine the relative fitness of single cells and preformed aggregates during early development of Pseudomonas aeruginosa biofilms. We find that the relative fitness of aggregates depends markedly on the density of surrounding single cells, i.e., the level of competition for growth resources. When competition between aggregates and single cells is low, an aggregate has a growth disadvantage because the aggregate interior has poor access to growth resources. However, if competition is high, aggregates exhibit higher fitness, because extending vertically above the surface gives cells at the top of aggregates better access to growth resources. Other advantages of seeding by aggregates, such as earlier switching to a biofilm-like phenotype and enhanced resilience toward antibiotics and immune response, may add to this ecological benefit. Our findings suggest that current models of biofilm formation should be reconsidered to incorporate the role of aggregates in biofilm initiation.

We have become increasingly aware that during infection, pathogenic bacteria often grow in multicellular biofilms which are often highly resistant to antibacterial strategies. In order to understand how biofilms form and contribute to infection, in vitro biofilm systems such as microtitre plate assays and flow cells, have been heavily used by many research groups around the world. Whilst these methods have greatly increased our understanding of the biology of biofilms, it is becoming increasingly apparent that many of our in vitro methods do not accurately represent in vivo conditions. Here we present a systematic review of the most widely used in vitro biofilm systems, and we discuss why they are not always representative of the in vivo biofilms found in chronic infections. We present examples of methods that will help us to bridge the gap between in vitro and in vivo biofilm work, so that our bench-side data can ultimately be used to improve bedside treatment.

Plant-derived compounds and other natural substances are a rich potential source of compounds that kill or attenuate pathogens that are resistant to current antibiotics. Medieval societies used a range of these natural substances to treat conditions clearly recognizable to the modern eye as microbial infections, and there has been much debate over the likely efficacy of these treatments. Our interdisciplinary team, comprising researchers from both sciences and humanities, identified and reconstructed a potential remedy for Staphylococcus aureus infection from a 10th century Anglo-Saxon leechbook. The remedy repeatedly killed established S. aureus biofilms in an in vitro model of soft tissue infection and killed methicillin-resistant S. aureus (MRSA) in a mouse chronic wound model. While the remedy contained several ingredients that are individually known to have some antibacterial activity, full efficacy required the combined action of several ingredients, highlighting the scholarship of premodern doctors and the potential of ancient texts as a source of new antimicrobial agents.

Bacterial biofilms are usually assumed to originate from individual cells deposited on a surface. However, many biofilm-forming bacteria tend to aggregate in the planktonic phase so that it is possible that many natural and infectious biofilms originate wholly or partially from pre-formed cell aggregates. Here, we use agent-based computer simulations to investigate the role of pre-formed aggregates in biofilm development. Focusing on the initial shape the aggregate forms on the surface, we find that the degree of spreading of an aggregate on a surface can play an important role in determining its eventual fate during biofilm development. Specifically, initially spread aggregates perform better when competition with surrounding unaggregated bacterial cells is low, while initially rounded aggregates perform better when competition with surrounding unaggregated cells is high. These contrasting outcomes are governed by a trade-off between aggregate surface area and height. Our results provide new insight into biofilm formation and development, and reveal new factors that may be at play in the social evolution of biofilm communities.

The presence of Pseudomonas aeruginosa in cutaneous wounds is of clinical significance and can lead to persistent infections. Manuka honey has gained ground in clinical settings due to its effective therapeutic action and broad spectrum of antibacterial activity. In this study, the effect of manuka honey on P. aeruginosa was investigated using MIC, MBC, growth kinetics, confocal microscopy, atomic force microscopy and real-time PCR. A bactericidal mode of action for manuka honey against P. aeruginosa was deduced (12 %, w/v, MIC; 16 %, w/v, MBC) and confirmed by confocal and atomic force microscopy, which showed extensive cell lysis after 60 min exposure to inhibitory concentrations of manuka honey. The inability of honey-treated cells to form microcolonies was demonstrated and investigated using Q-PCR for three key microcolony-forming genes: algD, lasR and oprF. The expression of algD increased 16-fold whereas oprF expression decreased 10-fold following honey treatment; lasR expression remained unaltered. These findings confirm that manuka honey is effective at inducing cell lysis and identify two targets, at the genetic level, that might be involved in this process. INTRODUCTIONBurns are one of the most prevalent skin traumas in modern society, and subsequent infection is common. Pseudomonas aeruginosa has been associated with burn infections for decades (McManus et al., 1985;Shankowsky et al., 1994; Tredget et al., 2004), and despite progress in treatment, infection remains the main cause of death amongst burn patients. Multidrug-resistant strains of P. aeruginosa have become commonplace due to their intrinsic resistance to antimicrobial agents (Strateva & Yordanov, 2009) and ability to acquire new antibiotic resistance mechanisms (Aloush et al., 2006). In addition to key immune defence mechanisms (Nikaido, 1994), P. aeruginosa possesses numerous virulence factors that also facilitate infection (Tredget et al., 2004). Once P. aeruginosa infections in burn wounds have established, they often become chronic, a state that is associated with the presence of bacterial biofilms (Bjarnsholt et al., 2008;James et al., 2008). Many currently used antimicrobials are ineffective against P. aeruginosa; therefore, novel therapeutic agents are sought for the successful management of wounds in burn patients.Honey is a natural product and for many centuries was held in high regard due to its antibacterial properties (Crane, 2001). Such effects have been observed against more than 80 bacterial species, including both Gram-positive and Gramnegative bacteria, and multidrug-resistant pathogens. It is for this reason that the use of honey has seen a resurgence in modern wound care management (Molan, 1992a) (for an indepth review of this topic see Cooper et al., 2009). The inherent antibacterial properties of honey are partly conferred by sugars, which account for 80 % of its weight, resulting in a high osmolarity and low water activity (Molan, 1992a). Hydrogen peroxide has been identified as the main antimicrobial component of man...