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Colistin–carbapenem combinations are synergistic in vitro against carbapenem-resistant Gram-negative bacteria. We aimed to test whether combination therapy improves clinical outcomes for adults with infections caused by carbapenem-resistant or carbapenemase-producing Gram-negative bacteria. A randomised controlled superiority trial was done in six hospitals in Israel, Greece, and Italy. We included adults with bacteraemia, ventilator-associated pneumonia, hospital-acquired pneumonia, or urosepsis caused by carbapenem-non-susceptible Gram-negative bacteria. Patients were randomly assigned (1:1) centrally, by computer-generated permuted blocks stratified by centre, to intravenous colistin (9-million unit loading dose, followed by 4·5 million units twice per day) or colistin with meropenem (2-g prolonged infusion three times per day). The trial was open-label, with blinded outcome assessment. Treatment success was defined as survival, haemodynamic stability, improved or stable Sequential Organ Failure Assessment score, stable or improved ratio of partial pressure of arterial oxygen to fraction of expired oxygen for patients with pneumonia, and microbiological cure for patients with bacteraemia. The primary outcome was clinical failure, defined as not meeting all success criteria by intention-to-treat analysis, at 14 days after randomisation. This trial is registered at ClinicalTrials.gov, number NCT01732250, and is closed to accrual. Between Oct 1, 2013, and Dec 31, 2016, we randomly assigned 406 patients to the two treatment groups. Most patients had pneumonia or bacteraemia (355/406, 87%), and most infections were caused by Acinetobacter baumannii (312/406, 77%). No significant difference between colistin monotherapy (156/198, 79%) and combination therapy (152/208, 73%) was observed for clinical failure at 14 days after randomisation (risk difference −5·7%, 95% CI −13·9 to 2·4; risk ratio [RR] 0·93, 95% CI 0·83–1·03). Results were similar among patients with A baumannii infections (RR 0·97, 95% CI 0·87–1·09). Combination therapy increased the incidence of diarrhoea (56 [27%] vs 32 [16%] patients) and decreased the incidence of mild renal failure (37 [30%] of 124 vs 25 [20%] of 125 patients at risk of or with kidney injury). Combination therapy was not superior to monotherapy. The addition of meropenem to colistin did not improve clinical failure in severe A baumannii infections. The trial was unpowered to specifically address other bacteria. EU AIDA grant Health-F3-2011-278348.

The modification of microbiota composition to a ‘beneficial’ one is a promising approach for improving intestinal as well as overall health. Natural fibres and phytochemicals that reach the proximal colon, such as those present in various nuts, provide substrates for the maintenance of healthy and diverse microbiota. The effects of increased consumption of specific nuts, which are rich in fibre as well as various phytonutrients, on human gut microbiota composition have not been investigated to date. The objective of the present study was to determine the effects of almond and pistachio consumption on human gut microbiota composition. We characterised microbiota in faecal samples collected from volunteers in two separate randomised, controlled, cross-over feeding studies (n 18 for the almond feeding study and n 16 for the pistachio feeding study) with 0, 1·5 or 3 servings/d of the respective nuts for 18 d. Gut microbiota composition was analysed using a 16S rRNA-based approach for bacteria and an internal transcribed spacer region sequencing approach for fungi. The 16S rRNA sequence analysis of 528 028 sequence reads, retained after removing low-quality and short-length reads, revealed various operational taxonomic units that appeared to be affected by nut consumption. The effect of pistachio consumption on gut microbiota composition was much stronger than that of almond consumption and included an increase in the number of potentially beneficial butyrate-producing bacteria. Although the numbers of bifidobacteria were not affected by the consumption of either nut, pistachio consumption appeared to decrease the number of lactic acid bacteria (P< 0·05). Increasing the consumption of almonds or pistachios appears to be an effective means of modifying gut microbiota composition.

Antimicrobial resistance in neonatal sepsis is rising, yet mechanisms of resistance that often spread between species via mobile genetic elements, ultimately limiting treatments in low- and middle-income countries (LMICs), are poorly characterized. The Burden of Antibiotic Resistance in Neonates from Developing Societies (BARNARDS) network was initiated to characterize the cause and burden of antimicrobial resistance in neonatal sepsis for seven LMICs in Africa and South Asia. A total of 36,285 neonates were enrolled in the BARNARDS study between November 2015 and December 2017, of whom 2,483 were diagnosed with culture-confirmed sepsis. Klebsiella pneumoniae ( n  = 258) was the main cause of neonatal sepsis, with Serratia marcescens ( n  = 151), Klebsiella michiganensis ( n  = 117), Escherichia coli ( n  = 75) and Enterobacter cloacae complex ( n  = 57) also detected. We present whole-genome sequencing, antimicrobial susceptibility and clinical data for 916 out of 1,038 neonatal sepsis isolates (97 isolates were not recovered from initial isolation at local sites). Enterobacterales ( K. pneumoniae, E. coli and E. cloacae ) harboured multiple cephalosporin and carbapenem resistance genes. All isolated pathogens were resistant to multiple antibiotic classes, including those used to treat neonatal sepsis. Intraspecies diversity of K. pneumoniae and E. coli indicated that multiple antibiotic-resistant lineages cause neonatal sepsis. Our results will underpin research towards better treatments for neonatal sepsis in LMICs. Genomic and clinical analysis of 916 bacterial isolates from neonates with sepsis in seven low- and middle-income countries (the BARNARDS study) reveals that the main species present were antimicrobial-resistant Klebsiella , Escherichia coli and Enterobacter .

Key Points Quorum sensing is a cell–cell communication process that enables bacteria to obtain information about cell density and species composition of the vicinal community and adjust their gene expression profiles accordingly. Quorum sensing involves the production, release and detection of extracellular signalling molecules known as autoinducers. Group-wide detection of autoinducers enables bacteria to collectively execute behaviours. Autoinducers are small molecules that control quorum sensing. In Gram-negative bacteria, autoinducers are often produced from S -adenosylmethionine (SAM). Autoinducers interact with specific receptors to elicit behaviours that are controlled by quorum sensing. Quorum sensing receptors are either membrane-bound histidine sensor kinases or cytoplasmic transcription factors. Autoinduction occurs when the detection of autoinducers induces the increased production of the same autoinducer molecule, forming a feed-forward regulatory loop. Other features, such as positive and negative feedback loops and small regulatory RNAs, optimize the integration of the autoinducer-encoded information and provide ideal quorum sensing dynamics. Signal integration is a process that takes place in most Gram-negative bacteria when several autoinducers and receptors work in parallel, or in series, to synchronize functions that are controlled by quorum sensing. Processes such as bioluminescence, the production of virulence factors and the formation of biofilms are controlled by quorum sensing. Quorum sensing shapes the composition of microbial communities. For example, bacterial species in the human gut microbiota produce and respond to autoinducers. There is increasing evidence that quorum sensing controls key physiological processes in the gut and may affect the virulence programmes of invading pathogens. Host cells are also known to produce autoinducer mimics. Synthetic quorum sensing modulators are molecules that agonize or antagonize quorum sensing and they are being developed as anti-virulence medicines. Distinct from traditional antibiotics, quorum sensing modulators do not affect the growth of pathogenic bacteria, but rather, disrupt their virulence programmes. Quorum sensing is used to control the behaviour of bacterial communities. In this Review, Papenfort and Bassler highlight recent discoveries about quorum sensing in Gram-negative bacteria, such as novel autoinducers and signalling networks that promote communication that ranges from intra-species to inter-kingdom. Bacteria use quorum sensing to orchestrate gene expression programmes that underlie collective behaviours. Quorum sensing relies on the production, release, detection and group-level response to extracellular signalling molecules, which are called autoinducers. Recent work has discovered new autoinducers in Gram-negative bacteria, shown how these molecules are recognized by cognate receptors, revealed new regulatory components that are embedded in canonical signalling circuits and identified novel regulatory network designs. In this Review we examine how, together, these features of quorum sensing signal–response systems combine to control collective behaviours in Gram-negative bacteria and we discuss the implications for host–microbial associations and antibacterial therapy.

The defining feature of the Gram-negative cell envelope is the presence of two cellular membranes, with the specialized glycolipid lipopolysaccharide (LPS) exclusively found on the surface of the outer membrane. The surface layer of LPS contributes to the stringent permeability properties of the outer membrane, which is particularly resistant to permeation of many toxic compounds, including antibiotics. As a common surface antigen, LPS is recognized by host immune cells, which mount defences to clear pathogenic bacteria. To alter properties of the outer membrane or evade the host immune response, Gram-negative bacteria chemically modify LPS in a wide variety of ways. Here, we review key features and physiological consequences of LPS biogenesis and modifications.Lipopolysaccharide is a key component of the Gram-negative cell envelope and functions, for example, as a permeability barrier or determinant of host immune responses. In this Review, Simpson and Trent guide us through lipopolysaccharide biogenesis and modifications and their functional and therapeutic implications.

Gram-negative bacteria possess a complex cell envelope that consists of a plasma membrane, a peptidoglycan cell wall and an outer membrane. The envelope is a selective chemical barrier 1 that defines cell shape 2 and allows the cell to sustain large mechanical loads such as turgor pressure 3 . It is widely believed that the covalently cross-linked cell wall underpins the mechanical properties of the envelope 4 , 5 . Here we show that the stiffness and strength of Escherichia coli cells are largely due to the outer membrane. Compromising the outer membrane, either chemically or genetically, greatly increased deformation of the cell envelope in response to stretching, bending and indentation forces, and induced increased levels of cell lysis upon mechanical perturbation and during L-form proliferation. Both lipopolysaccharides and proteins contributed to the stiffness of the outer membrane. These findings overturn the prevailing dogma that the cell wall is the dominant mechanical element within Gram-negative bacteria, instead demonstrating that the outer membrane can be stiffer than the cell wall, and that mechanical loads are often balanced between these structures. The outer membrane of Gram-negative bacteria is shown to be at least as stiff as the cell wall, and this property enables it to protect cells from mechanical pertubations.

Interactions between the immune system and the intestinal microbiota may play a role in coeliac disease (CD). In the present study, the potential effects of Bifidobacterium longum CECT 7347 in children with newly diagnosed CD were evaluated. A double-blind, randomised, placebo-controlled trial was conducted in thirty-three children who received a capsule containing either B. longum CECT 7347 (10 9 colony-forming units) or placebo (excipients) daily for 3 months together with a gluten-free diet (GFD). Outcome measures (baseline and post-intervention) included immune phenotype of peripheral blood cells, serum cytokine concentration, faecal secretory IgA (sIgA) content, anthropometric parameters and intestinal microbiota composition. Comparisons between the groups revealed greater height percentile increases ( P = 0·048) in the B. longum CECT 7347 group than in the placebo group, as well as decreased peripheral CD3 + T lymphocytes ( P = 0·004) and slightly reduced TNF-α concentration ( P = 0·067). Within-group comparisons of baseline and final values did not reveal any differences in T lymphocytes and cytokines in the placebo group, while decreased CD3 + ( P = 0·013) and human leucocyte antigen (HLA)-DR + T lymphocytes ( P = 0·029) and slightly reduced TNF-α concentration ( P = 0·085) were detected in the B. longum CECT 7347 group. Comparison between the groups showed that the administration of B. longum CECT 7347 reduced the numbers of the Bacteroides fragilis group ( P = 0·020) and the content of sIgA in stools ( P = 0·011) compared with the administration of placebo. Although this is a first exploratory intervention with limitations, the findings suggest that B. longum CECT 7347 could help improve the health status of CD patients who tend to show alterations in gut microbiota composition and a biased immune response even on a GFD.

Most small molecules are unable to rapidly traverse the outer membrane of Gram-negative bacteria and accumulate inside these cells, making the discovery of much-needed drugs against these pathogens challenging. Current understanding of the physicochemical properties that dictate small-molecule accumulation in Gram-negative bacteria is largely based on retrospective analyses of antibacterial agents, which suggest that polarity and molecular weight are key factors. Here we assess the ability of over 180 diverse compounds to accumulate in Escherichia coli . Computational analysis of the results reveals major differences from the retrospective studies, namely that the small molecules that are most likely to accumulate contain an amine, are amphiphilic and rigid, and have low globularity. These guidelines were then applied to convert deoxynybomycin, a natural product that is active only against Gram-positive organisms, into an antibiotic with activity against a diverse panel of multi-drug-resistant Gram-negative pathogens. We anticipate that these findings will aid in the discovery and development of antibiotics against Gram-negative bacteria. The authors use computational modelling and a set of chemically synthesized compounds to define the physicochemical properties required for small-molecule accumulation in Gram-negative bacteria. Rules for small-molecule accumulation in Gram-negative bacteria Most small molecules are unable to cross the outer membrane of Gram-negative bacteria and accumulate inside these cells, which poses a challenge for the discovery of new drugs that target Gram-negative pathogens. By examining a set of chemically diverse small molecules, Paul Hergenrother and colleagues have now defined the physicochemical properties required for small-molecule accumulation in the Gram-negative bacteria Escherichia coli . They find that small molecules containing an amine, and which are amphiphilic, rigid and have low globularity, are most likely to be successful. They then apply these guidelines to convert a compound that targets Gram-positive bacteria only into a broad-spectrum antibiotic that is active against several Gram-negative pathogens.

Peptidoglycan and almost all surface glycopolymers in bacteria are built in the cytoplasm on the lipid carrier undecaprenyl phosphate (UndP) 1 – 4 . These UndP-linked precursors are transported across the membrane and polymerized or directly transferred to surface polymers, lipids or proteins. UndP is then flipped to regenerate the pool of cytoplasmic-facing UndP. The identity of the flippase that catalyses transport has remained unknown. Here, using the antibiotic amphomycin that targets UndP 5 – 7 , we identified two broadly conserved protein families that affect UndP recycling. One (UptA) is a member of the DedA superfamily 8 ; the other (PopT) contains the domain DUF368. Genetic, cytological and syntenic analyses indicate that these proteins are UndP transporters. Notably, homologues from Gram-positive and Gram-negative bacteria promote UndP transport in Bacillus subtilis , indicating that recycling activity is broadly conserved among family members. Inhibitors of these flippases could potentiate the activity of antibiotics targeting the cell envelope. A study identifies two broadly conserved families of flippases that catalyse the transport of undecaprenyl phosphate in bacteria and could function to recycle dolichol phosphate in eukaryotes and archaea.

Silver is considered as antibacterial agent with well-known mode of action and bacterial resistance against it is well described. The development of nanotechnology provided different methods for the modification of the chemical and physical structure of silver, which may increase its antibacterial potential. The physico-chemical properties of silver nanoparticles and their interaction with living cells differs substantially from those of silver ions. Moreover, the variety of the forms and characteristics of various silver nanoparticles are also responsible for differences in their antibacterial mode of action and probably bacterial mechanism of resistance. The paper discusses in details the aforementioned aspects of silver activity.