The enemy within: phloem‐limited pathogens

The growing impact of phloem‐limited pathogens on high‐value crops has led to a renewed interest in understanding how they cause disease. Although these pathogens cause substantial crop losses, many are poorly characterized. In this review, we present examples of phloem‐limited pathogens that include intracellular bacteria with and without cell walls, and viruses. Phloem‐limited pathogens have small genomes and lack many genes required for core metabolic processes, which is, in part, an adaptation to the unique phloem environment. For each pathogen class, we present multiple case studies to highlight aspects of disease caused by phloem‐limited pathogens. The pathogens presented include Candidatus Liberibacter asiaticus (citrus greening), Arsenophonus bacteria, Serratia marcescens (cucurbit yellow vine disease), Candidatus Phytoplasma asteris (Aster Yellows Witches’ Broom), Spiroplasma kunkelii, Potato leafroll virus and Citrus tristeza virus. We focus on commonalities in the virulence strategies of these pathogens, and aim to stimulate new discussions in the hope that widely applicable disease management strategies can be found.     

Comparison of One‐Step and Two‐Step Meta‐Analysis Models Using Individual Patient Data

The problem of combining information from separate trials is a key consideration when performing a meta‐analysis or planning a multicentre trial. Although there is a considerable journal literature on meta‐analysis based on individual patient data (IPD), i.e. a one‐step IPD meta‐analysis, versus analysis based on summary data, i.e. a two‐step IPD meta‐analysis, recent articles in the medical literature indicate that there is still confusion and uncertainty as to the validity of an analysis based on aggregate data. In this study, we address one of the central statistical issues by considering the estimation of a linear function of the mean, based on linear models for summary data and for IPD. The summary data from a trial is assumed to comprise the best linear unbiased estimator, or maximum likelihood estimator of the parameter, along with its covariance matrix. The setup, which allows for the presence of random effects and covariates in the model, is quite general and includes many of the commonly employed models, for example, linear models with fixed treatment effects and fixed or random trial effects. For this general model, we derive a condition under which the one‐step and two‐step IPD meta‐analysis estimators coincide, extending earlier work considerably. The implications of this result for the specific models mentioned above are illustrated in detail, both theoretically and in terms of two real data sets, and the roles of balance and heterogeneity are highlighted. Our analysis also shows that when covariates are present, which is typically the case, the two estimators coincide only under extra simplifying assumptions, which are somewhat unrealistic in practice.     

Forecasting Pollen Concentration by a Two‐Step Functional Model

Summary A functional regression model to forecast the cypress pollen concentration during a given time interval, considering the air temperature in a previous interval as the input, is derived by means of a two‐step procedure. This estimation is carried out by functional principal component (FPC) analysis and the residual noise is also modeled by FPC regression, taking as the explicative process the pollen concentration during the earlier interval. The prediction performance is then tested on pollen data series recorded in Granada (Spain) over a period of 10 years.     

Inhibition of death receptor signaling by bacterial gut pathogens

Gastrointestinal bacterial pathogens such as enteropathogenic Escherichia coli, Salmonella and Shigella control inflammatory and apoptotic signaling in human intestinal cells to establish infection, replicate and disseminate to other hosts. These pathogens manipulate host cell signaling through the translocation of virulence effector proteins directly into the host cell cytoplasm, which then target various signaling pathways. Death receptors such as TNFR1, FAS and TRAIL-R induce signaling cascades that are crucial to the clearance of pathogens, and as such are major targets for inhibition by pathogens. This review focuses on what is known about how bacterial gut pathogens inhibit death receptor signaling to suppress inflammation and prevent apoptosis.     

The bacterial chemical repertoire mediates metabolic exchange within gut microbiomes

Microbial communities in the gut have been hypothesized to play key roles in the health of the host organism. Exploring the relationship between these populations and disease states has been a focus of the human microbiome project. However, the biological roles of the compounds produced by the gut bacteria are largely unknown. We hypothesize that these compounds act as metabolic exchange factors-mediating inter-species and intra-species interactions in the microbiome. This view is supported through this review of known bacterial metabolic exchange factors and evidence for uncharacterized metabolic exchange factors in the gut. The impact of model systems and technological developments in exploring this hypothesis are also discussed. Together, these investigations are revolutionizing our understanding of the gut microbiome-presenting the possibility of identifying new strategies for treating disease in the host.     

The forensic analysis of foodborne bacterial pathogens in the age of whole‐genome sequencing

The forensic evaluation of bacterial pathogens presents new challenges to the forensic science community. This review examines bacterial pathogens as objects of forensic comparison, focusing on their nucleic acid sequences as an important aspect of the comparison process. Because of the clonal propagation of most bacterial pathogens, a phylogenetic approach to understanding the diversity and using this understanding to address common forensic questions is explored. As a general phylogenetic framework is now employed in human mitochondrial DNA analysis, we will use the relevant concepts and approaches common in this area to develop this approach further. We also address the impact of the current ease and prevalence of whole‐genome DNA sequence analysis in the forensic comparison process.     

The specific and sensitive detection of bacterial pathogens within 4 h using bacteriophage amplification

This paper describes a novel approach, termed the 'phage amplification assay', for the rapid detection and identification of specific bacteria. The technique is based on the phage lytic cycle with plaque formation as the assay end-point. It is highly sensitive, quantitative and gives results typically within 4 h. The assay comprises four main stages : (1) phage infection of target bacterium ; (2) destruction of exogenous phage ; (3) amplification of phage within infected host and (4) plaque formation from infected host with the aid of helper bacteria. A key component of this assay is a potent virucidal agent derived from natural plant extracts, pomegranate rind extract (PRE). In combination with ferrous sulphate PRE can bring about an 11 log-cycle reduction in phage titre within 3 min. This is achieved without any injury to the infected target bacteria. Subsequently, any resulting plaques are derived only from infected target organisms. Data are presented for a range of bacterial hosts including Pseudomonas aeruginosa, Salmonella typhimurium and Staphylococcus aureus. The detection limit for Ps. aeruginosa was 40 bacteria ml⁻¹ in a time of 4 h and 600 bacteria m⁻¹ for Salm. typhimurium. Application of the principles of this technology to other bacterial genera is discussed.     

Close encounters of the type‐six kind: injected bacterial toxins modulate gut microbial composition

Bacteria of the phylum Bacteroidetes constitute a substantial portion of the human gut microbiota, including symbionts and opportunistic pathogens. How these bacteria coexist and provide colonization resistance to pathogenic strains is not well understood. In this issue of EMBO Reports, Hecht and colleagues describe a mechanism by which strains of Bacteroides fragilis compete with each other for an intestinal niche 1 . Prompted by the observation that B. fragilis populations appear to be dominated by either commensal, non‐toxigenic strains, or by enterotoxigenic, potentially pathogenic strains, the authors investigated mechanisms of competition between these two subsets. In agreement with two recent studies 2 3 , Hecht et al 1 found that competition between B. fragilis strains is dependent on a type‐6 secretion system (T6SS) apparatus, secreted effectors, and immunity genes. They identify a T6SS effector–immunity gene pair that enables a non‐toxigenic strain to competitively exclude enterotoxigenic B. fragilis, thus providing a proof of principle for the use of T6SS‐mediated killing as a therapeutic strategy to eradicate pathogenic strains.     

Survival of coliforms and bacterial pathogens within protozoa during chlorination.

The susceptibility of coliform bacteria and bacterial pathogens to free chlorine residuals was determined before and after incubation with amoebae and ciliate protozoa. Viability of bacteria was quantified to determine their resistance to free chlorine residuals when ingested by laboratory strains of Acanthamoeba castellanii and Tetrahymena pyriformis. Cocultures of bacteria and protozoa were incubated to facilitate ingestion of the bacteria and then were chlorinated, neutralized, and sonicated to release intracellular bacteria. Qualitative susceptibility of protozoan strains to free chlorine was also assessed. Protozoa were shown to survive and grow after exposure to levels of free chlorine residuals that killed free-living bacteria. Ingested coliforms Escherichia coli, Citrobacter freundii, Enterobacter agglomerans, Enterobacter cloacae, Klebsiella pneumoniae, and Klebsiella oxytoca and bacterial pathogens Salmonella typhimurium, Yersinia enterocolitica, Shigella sonnei, Legionella gormanii, and Campylobacter jejuni had increased resistance to free chlorine residuals. Bacteria could be cultured from within treated protozoans well after the time required for 99% inactivation of free-living cells. All bacterial pathogens were greater than 50-fold more resistant to free chlorine when ingested by T. pyriformis. Escherichia coli ingested by a Cyclidium sp., a ciliate isolated from a drinking water reservoir, were also shown to be more resistant to free chlorine. The mechanism that increased resistance appeared to be survival within protozoan cells. This study indicates that bacteria can survive ingestion by protozoa. This bacterium-protozoan association provides bacteria with increased resistance to free chlorine residuals which can lead to persistence of bacteria in chlorine-treated water. We propose that resistance to digestion by predatory protozoa was an evolutionary precursor of pathogenicity in bacteria and that today it is a mechanism for survival of fastidious bacteria in dilute and inhospitable aquatic environments.     

Integrated multi-scale strategies to investigate nutritional compounds and their effect on the gut microbiota

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The Evolution of Insertion Sequences within Enteric Bacteria

To identify mechanisms that influence the evolution of bacterial transposons, DNA sequence variation was evaluated among homologs of insertion sequences IS1, IS3 and IS30 from natural strains of Escherichia coli and related enteric bacteria. The nucleotide sequences within each class of IS were highly conserved among E. coli strains, over 99.7% similar to a consensus sequence. When compared to the range of nucleotide divergence among chromosomal genes, these data indicate high turnover and rapid movement of the transposons among clonal lineages of E. coli. In addition, length polymorphism among IS appears to be far less frequent than in eukaryotic transposons, indicating that nonfunctional elements comprise a smaller fraction of bacterial transposon populations than found in eukaryotes. IS present in other species of enteric bacteria are substantially divergent from E. coli elements, indicating that IS are mobilized among bacterial species at a reduced rate. However, homologs of IS1 and IS3 from diverse species provide evidence that recombination events and horizontal transfer of IS among species have both played major roles in the evolution of these elements. IS3 elements from E. coli and Shigella show multiple, nested, intragenic recombinations with a distantly related transposon, and IS1 homologs from diverse taxa reveal a mosaic structure indicative of multiple recombination and horizontal transfer events.     

New Strategy for Two‐Step Sequential Deposition: Incorporation of Hydrophilic Fullerene in Second Precursor for High‐Performance p‐i‐n Planar Perovskite Solar Cells

In p‐i‐n planar perovskite solar cells (pero‐SCs) based on methylammonium lead iodide (MAPbI₃) perovskite, high‐quality MAPbI₃ film, perfect interfacial band alignment and efficient charge extracting ability are critical for high photovoltaic performance. In this work, a hydrophilic fullerene derivative [6,6]‐phenyl‐C₆₁‐butyric acid‐(3,4,5‐tris(2‐(2‐(2‐methoxyethoxy)ethoxy)ethoxy)phenyl)methanol ester (PCBB‐OEG) is introduced as additive in the methylammonium iodide precursor solution in the preparation of MAPbI₃ perovskite film by two‐step sequential deposition method, and obtained a top‐down gradient distribution with an ultrathin top layer of PCBB‐OEG. Meanwhile, a high‐quality perovskite film with high crystallinity, less trap‐states, and dense‐grained uniform morphology can well grow on both hydrophilic (poly(3,4‐ethylenedioxythiophene)/poly(styrenesulfonic acid)) and hydrophobic (polytriarylamine, PTAA) hole transport layers. When the PCBB‐OEG‐containing perovskite film (pero‐0.1) is prepared in a p‐i‐n planar pero‐SC with the configuration of ITO/PTAA/pero‐0.1/[6,6]‐phenyl‐C₆₁‐butyric acid methyl ester/Al, the device delivers a promising power conversion efficiency (PCE) of 20.2% without hysteresis, which is one of the few PCE over 20% for the p‐i‐n planar pero‐SCs. Importantly, the pero‐0.1‐based device shows an excellent stability that can retain 98.4% of its initial PCE after being exposed for 300 h under ambient atmosphere with a high humidity, and the flexible pero‐SCs based on pero‐0.1 also demonstrate a promising PCE of 18.1%.     

Incorporating germination‐induction into decontamination strategies for bacterial spores

Oenococcus oeni is the dominant species able to cope with a hostile environment of wines, comprising cumulative effects of low pH, high ethanol and SO₂ content, nonoptimal growth temperatures and growth inhibitory compounds. Ethanol tolerance is a crucial feature for the activity of O. oeni cells in wine because ethanol acts as a disordering agent of its cell membrane and negatively affects metabolic activity; it damages the membrane integrity, decreases cell viability and, as other stress conditions, delays the start of malolactic fermentation with a consequent alteration of wine quality. The cell wall, cytoplasmic membrane and metabolic pathways are the main sites involved in physiological changes aimed to ensure an adequate adaptive response to ethanol stress and to face the oxidative damage caused by increasing production of reactive oxygen species. Improving our understanding of the cellular impact of ethanol toxicity and how the cell responds to ethanol stress can facilitate the development of strategies to enhance microbial ethanol tolerance; this allows to perform a multidisciplinary endeavour requiring not only an ecological study of the spontaneous process but also the characterization of useful technological and physiological features of the predominant strains in order to select those with the highest potential for industrial applications.     

Enteric pathogen exploitation of the microbiota-generated nutrient environment of the gut

Residing within the intestine is a large community of commensal organisms collectively termed the microbiota. This community generates a complex nutrient environment by breaking down indigestible food products into metabolites that are used by both the host and the microbiota. Both the invading intestinal pathogen and the microbiota compete for these metabolites, which can shape both the composition of the flora, as well as susceptibility to infection. After infection is established, pathogen mediated inflammation alters the composition of the microbiota, which further shifts the makeup of metabolites in the gastrointestinal tract. A greater understanding of the interplay between the microbiota, the metabolites they generate, and susceptibility to enteric disease will enable the discovery of novel therapies against infectious disease.     

The neglected intrinsic resistome of bacterial pathogens

Bacteria with intrinsic resistance to antibiotics are a worrisome health problem. It is widely believed that intrinsic antibiotic resistance of bacterial pathogens is mainly the consequence of cellular impermeability and activity of efflux pumps. However, the analysis of transposon-tagged Pseudomonas aeruginosa mutants presented in this article shows that this phenotype emerges from the action of numerous proteins from all functional categories. Mutations in some genes make P. aeruginosa more susceptible to antibiotics and thereby represent new targets. Mutations in other genes make P. aeruginosa more resistant and therefore define novel mechanisms for mutation-driven acquisition of antibiotic resistance, opening a new research field based in the prediction of resistance before it emerges in clinical environments. Antibiotics are not just weapons against bacterial competitors, but also natural signalling molecules. Our results demonstrate that antibiotic resistance genes are not merely protective shields and offer a more comprehensive view of the role of antibiotic resistance genes in the clinic and in nature.     

The role of bacterial pathogens in cancer

The association of Helicobacter pylori with gastric cancer is the best-studied relationship between a bacterial infection and cancer. Other bacterial pathogens in humans and rodents are now being recognized as potentially having a direct role in carcinogenesis. Thus, it might be possible to understand the pathogenesis and prevention of certain cancers by studying the bacterial infections associated with them, and their effects on the host. However, the mechanisms by which bacteria contribute to cancer formation are complex, and recent investigations show that they involve the interplay between chronic inflammation, direct microbial effects on host cell physiology and, ultimately, changes in tissue stem cell homeostasis.