Efflux pumps are transport proteins involved in the extrusion of toxic substrates (including virtually all classes of clinically relevant antibiotics) from within cells into the external environment. These proteins are found in both Gram-positive and -negative bacteria as well as in eukaryotic organisms. 1 Pumps may be specific for one substrate or may transport a range of structurally dissimilar compounds (including antibiotics of multiple classes); such pumps can be associated with multiple drug resistance (MDR). In the prokaryotic kingdom there are five major families of efflux transporter: 2 MF (major facilitator), MATE (multidrug and toxic efflux), RND (resistance-nodulation-division), SMR (small multidrug resistance) and ABC (ATP binding cassette). All these systems utilize the proton motive force as an energy source, 3 apart from the ABC family, which utilizes ATP hydrolysis to drive the export of substrates. Recent advances in DNA technology and the advent of the genomic era have led to the identification of numerous new members of the above families, and the ubiquitous nature of efflux pumps is remarkable. Transporters that efflux multiple substrates, including antibiotics, have not evolved in response to the stresses of the antibiotic era. All bacterial genomes studied contain several different efflux pumps; this indicates their ancestral origins. It has been estimated that ∼5–10% of all bacterial genes are involved in transport and a large proportion of these encode efflux pumps. 2^, 4

Most bacteria in nature exist in aggregated communities known as biofilms, and cells within a biofilm demonstrate major physiological changes compared to their planktonic counterparts. Biofilms are associated with many different types of infections which can have severe impacts on patients. Infections involving a biofilm component are often chronic and highly recalcitrant to antibiotic therapy as a result of intrinsic physical factors including extracellular matrix production, low growth rates, altered antibiotic target production and efficient exchange of resistance genes. This review describes the biofilm lifecycle, phenotypic characteristics of a biofilm, and contribution of matrix and persister cells to biofilms intrinsic tolerance to antimicrobials. We also describe how biofilms can evolve antibiotic resistance and transfer resistance genes within biofilms. Multispecies biofilms and the impacts of various interactions, including cooperation and competition, between species on tolerance to antimicrobials in polymicrobial biofilm communities are also discussed.

Background: Foodborne outbreaks of Salmonella remain a pressing public health concern. We recently detected a large outbreak of Salmonella enterica serovar Enteritidis phage type 14b affecting more than 30 patients in our hospital. This outbreak was linked to community, national and European-wide cases. Hospital patients with Salmonella are at high risk, and require a rapid response. We initially investigated this outbreak by whole-genome sequencing using a novel rapid protocol on the Illumina MiSeq; we then integrated these data with whole-genome data from surveillance sequencing, thereby placing the outbreak in a national context. Additionally, we investigated the potential of a newly released sequencing technology, the MinION from Oxford Nanopore Technologies, in the management of a hospital outbreak of Salmonella. Results: We demonstrate that rapid MiSeq sequencing can reduce the time to answer compared to the standard sequencing protocol with no impact on the results. We show, for the first time, that the MinION can acquire clinically relevant information in real time and within minutes of a DNA library being loaded. MinION sequencing permits confident assignment to species level within 20 min. Using a novel streaming phylogenetic placement method samples can be assigned to a serotype in 40 min and determined to be part of the outbreak in less than 2 h. Conclusions: Both approaches yielded reliable and actionable clinical information on the Salmonella outbreak in less than half a day. The rapid availability of such information may facilitate more informed epidemiological investigations and influence infection control practices.