Scavenging Bacterial Siderophores with Engineered Lipocalin Proteins as an Alternative Antimicrobial Strategy

Iron acquisition mediated by siderophores, high-affinity chelators for which bacteria have evolved specific synthesis and uptake mechanisms, plays a crucial role in microbiology and in host–pathogen interactions. In the ongoing fight against bacterial infections, this area has attracted biomedical interest. Beyond several approaches to interfere with siderophore-mediated iron uptake from medicinal and immunochemistry, the development of high-affinity protein scavengers that tightly complex the siderophores produced by pathogenic bacteria has appeared as a novel strategy. Such binding proteins have been engineered based on siderocalin—also known as lipocalin 2—an endogenous human scavenger of enterobactin and bacillibactin that controls the systemic spreading of commensal bacteria such as Escherichia coli. By using combinatorial protein design, siderocalin was reshaped to bind several siderophores from Pseudomonas aeruginosa and, in particular, petrobactin from Bacillus anthracis, none of which is recognized by the natural protein. Such engineered versions of siderocalin effectively suppress the growth of corresponding pathogenic bacteria by depriving them of their iron supply and offer the potential to complement antibiotic therapy in situations of acute or persistent infection.     

Bacterial Retention in Lipopolysaccharide Coated Silica Sand

Abstract

Wastewater reclamation has been widely practiced in agriculture. When reclaimed wastewater is used for irrigation, among other requirements, it is important that the pathogenic bacteria are removed. Consequently, technologies such as immobilization or sorption barriers have been developed. To enhance the removal efficiency, biopolymers have been introduced to amend these immobilization or sorption barriers. In this study, removal of pathogenic bacteria by lipopolysaccharide (LPS)‐amended barriers was investigated by means of laboratory column experiments. Two typical gram‐negative pathogenic bacterial strains of Escherichia coli and Pseudomonas fluoresences and one gram‐positive bacterial strain of Streptococcus mitis were selected as the model bacteria in this research. Bacterial adhesion to uncoated and LPS‐coated silica sand was correlated to their interaction free energies. Both E.coli, P. fluorescens, and S. mitis had negative interaction free energies with silica sand, demonstrating their adhesion potentials to silica sand. After LPS coating, bacterium‐sediment interaction free energies decreased (negatively increased), and consequently, bacterial retention increased. Bacterial deposition coefficient in silica sand corresponded to their interaction free energies with silica sand. This study demonstrated that bacterial retention in porous media was determined by their interactions with the sediments, which could be predicted based on independently determined bacterial and medium physicochemical surface properties.     

Development of Quorum-Based Anti-Virulence Therapeutics Targeting Gram-Negative Bacterial Pathogens

Quorum sensing is a cell density-dependent signaling phenomenon used by bacteria for coordination of population-wide phenotypes, such as expression of virulence genes, antibiotic resistance and biofilm formation. Lately, disruption of bacterial communication has emerged as an anti-virulence strategy with enormous therapeutic potential given the increasing incidences of drug resistance in pathogenic bacteria. The quorum quenching therapeutic approach promises a lower risk of resistance development, since interference with virulence generally does not affect the growth and fitness of the bacteria and, hence, does not exert an associated selection pressure for drug-resistant strains. With better understanding of bacterial communication networks and mechanisms, many quorum quenching methods have been developed against various clinically significant bacterial pathogens. In particular, Gram-negative bacteria are an important group of pathogens, because, collectively, they are responsible for the majority of hospital-acquired infections. Here, we discuss the current understanding of existing quorum sensing mechanisms and present important inhibitory strategies that have been developed against this group of pathogenic bacteria.     

Arginine Metabolism in Bacterial Pathogenesis and Cancer Therapy

Antibacterial resistance to infectious diseases is a significant global concern for health care organizations; along with aging populations and increasing cancer rates, it represents a great burden for government healthcare systems. Therefore, the development of therapies against bacterial infection and cancer is an important strategy for healthcare research. Pathogenic bacteria and cancer have developed a broad range of sophisticated strategies to survive or propagate inside a host and cause infection or spread disease. Bacteria can employ their own metabolism pathways to obtain nutrients from the host cells in order to survive. Similarly, cancer cells can dysregulate normal human cell metabolic pathways so that they can grow and spread. One common feature of the adaption and disruption of metabolic pathways observed in bacterial and cancer cell growth is amino acid pathways; these have recently been targeted as a novel approach to manage bacterial infections and cancer therapy. In particular, arginine metabolism has been illustrated to be important not only for bacterial pathogenesis but also for cancer therapy. Therefore, greater insights into arginine metabolism of pathogenic bacteria and cancer cells would provide possible targets for controlling of bacterial infection and cancer treatment. This review will summarize the recent progress on the relationship of arginine metabolism with bacterial pathogenesis and cancer therapy, with a particular focus on arginase and arginine deiminase pathways of arginine catabolism.     

细菌的泛基因组分析

细菌基因组学在不同领域的广泛应用得益于新一代测序(next generation sequencing,NGS)技术所实现的基因组及转录组测序的发展。大量的细菌基因组数据为不同方面深入了解物种内部的多样性提供了数据资源。近年来,泛基因组分析(pan-genome analysis)方法受到研究者广泛的关注。通过泛基因组分析,任何特定物种均可获得准确的基因库信息,为进一步分析细菌的基因功能、环境适应性及致病机制提供了思路。本文对泛基因组的研究策略、模型构建及其在细菌中的研究应用作一综述。     

Assessing the Antimicrobial Activity of Polyisoprene Based Surfaces

There has been an intense research effort in the last decades in the field of biofouling prevention as it concerns many aspects of everyday life and causes problems to devices, the environment, and human health. Many different antifouling and antimicrobial materials have been developed to struggle against bacteria and other micro- and macro-organism attachment to different surfaces. However the “miracle solution” has still to be found. The research presented here concerns the synthesis of bio-based polymeric materials and the biological tests that showed their antifouling and, at the same time, antibacterial activity. The raw material used for the coating synthesis was natural rubber. The polyisoprene chains were fragmented to obtain oligomers, which had reactive chemical groups at their chain ends, therefore they could be modified to insert polymerizable and biocidal groups. Films were obtained by radical photopolymerization of the natural rubber derived oligomers and their structure was altered, in order to understand the mechanism of attachment inhibition and to increase the efficiency of the anti-biofouling action. The adhesion of three species of pathogenic bacteria and six strains of marine bacteria was studied. The coatings were able to inhibit bacterial attachment by contact, as it was verified that no detectable leaching of toxic molecules occurred.     

Acid-Denatured Green Fluorescent Protein (GFP) as Model Substrate to Study the Chaperone Activity of Protein Disulfide Isomerase

Green fluorescent protein (GFP) has been widely used in several molecular and cellular biology applications, since it is remarkably stable in vitro and in vivo. Interestingly, native GFP is resistant to the most common chemical denaturants; however, a low fluorescence signal has been observed after acid-induced denaturation. Furthermore, this acid-denatured GFP has been used as substrate in studies of the folding activity of some bacterial chaperones and other chaperone-like molecules. Protein disulfide isomerase enzymes, a family of eukaryotic oxidoreductases that catalyze the oxidation and isomerization of disulfide bonds in nascent polypeptides, play a key role in protein folding and it could display chaperone activity. However, contrasting results have been reported using different proteins as model substrates. Here, we report the further application of GFP as a model substrate to study the chaperone activity of protein disulfide isomerase (PDI) enzymes. Since refolding of acid-denatured GFP can be easily and directly monitored, a simple micro-assay was used to study the effect of the molecular participants in protein refolding assisted by PDI. Additionally, the effect of a well-known inhibitor of PDI chaperone activity was also analyzed. Because of the diversity their functional activities, PDI enzymes are potentially interesting drug targets. Since PDI may be implicated in the protection of cells against ER stress, including cancer cells, inhibitors of PDI might be able to enhance the efficacy of cancer chemotherapy; furthermore, it has been demonstrated that blocking the reductive cleavage of disulfide bonds of proteins associated with the cell surface markedly reduces the infectivity of the human immunodeficiency virus. Although several high-throughput screening (HTS) assays to test PDI reductase activity have been described, we report here a novel and simple micro-assay to test the chaperone activity of PDI enzymes, which is amenable for HTS of PDI inhibitors.     

A century of bacteriophage research and applications: impacts on biotechnology,health, ecology and the economy!

About a century ago, bacteriophages or phages – viruses that infect bacteria – were discovered and reported in the scientific literature. This review aims at a comprehensive survey of bacteriophage discovery, research and applications since the 1920s, and its impact on molecular biology, biotechnology, health, ecology and the economy. Phage therapy has been proven a valuable asset to deal with pathogenic bacterial infections since the early 1920s, and has been practiced ever since, especially in the former Soviet Union and in Eastern Europe. The Western world remained sceptical and resorted to the widespread use of antibiotics since the 1940s. Now that antibiotic resistance among pathogenic bacteria has spread alarmingly and few really novel antibiotic compounds are in the pipeline, renewed attention is being directed to the use of phages as antibacterial agents in medicine. Because of this renewed interest in phage therapy in the Western world, novel applications with phages are being pursued in the human health, environmental and the agri-food sectors. This review will focus on (1) the history of early phage use and its successes and problems, (2) the study of phages as important tools in the development of molecular biology and biotechnology, (3) current developments in phage research including the use of phage endolysins for use in antibacterial treatment, (4) phage production systems including undesirable phage contamination of industrial fermentation processes based on bacteria, (5) recent applications in phage therapy and in phage-based control, and (6) the roles of phages in nature and in the human gut. © 2018 Society of Chemical Industry     

Phage therapy and phage control: to be revisited urgently!!

Phage therapy has been a valuable asset since the 1920s to deal with pathogenic bacterial infections, and has been practised ever since, especially in the former Soviet Union and in eastern Europe. The western world remained sceptical and resorted to the widespread use of antibiotics as soon as they became available. Now that antibiotic resistance among pathogenic bacteria has spread alarmingly and no really new antibiotic compounds are in the pipeline, renewed attention should be directed to the use of phages as antimicrobials in medicine as well as in the health and the agro‐food sector. The history of early phage use, its successes and problems and the current developments in phage research, production and applications in phage control and therapy are critically reviewed in this context. © 2013 Society of Chemical Industry     

Recent Advances in Inhibitors of Bacterial Fatty Acid Synthesis Type II (FASII) System Enzymes as Potential Antibacterial Agents

Bacterial infections are a constant and serious threat to human health. With the increase of multidrug resistance of clinically pathogenic bacteria, common antibiotic therapies have been less effective. Fatty acid synthesis type II (FASII) system enzymes are essential for bacterial membrane lipid biosynthesis and represent increasingly promising targets for the discovery of antibacterial agents with new mechanisms of action. This review highlights recent advances in inhibitors of bacterial FASII as potential antibacterial agents, paying special attention to the activities, mechanisms, and structure–activity relationships of those inhibitors that mainly target β‐ketoacyl‐ACP synthase, β‐ketoacyl‐ACP reductase, β‐hydroxyacyl‐ACP dehydratase, and enoyl‐ACP reductase. Although inhibitors with low nanomolar and selective activity against various bacterial FASII have entered clinical trials, further research is needed to expand upon both available and yet unknown scaffolds to identify new FASII inhibitors that may have antibacterial potential, particularly against resistant bacterial strains.     

Quorum Quenching Enzymes and Their Application in Degrading Signal Molecules to Block Quorum Sensing-Dependent Infection

With the emergence of antibiotic-resistant strains of bacteria, the available options for treating bacterial infections have become very limited, and the search for a novel general antibacterial therapy has received much greater attention. Quorum quenching can be used to control disease in a quorum sensing system by triggering the pathogenic phenotype. The interference with the quorum sensing system by the quorum quenching enzyme is a potential strategy for replacing traditional antibiotics because the quorum quenching strategy does not aim to kill the pathogen or limit cell growth but to shut down the expression of the pathogenic gene. Quorum quenching enzymes have been identified in quorum sensing and non-quorum sensing microbes, including lactonase, acylase, oxidoreductase and paraoxonase. Lactonase is widely conserved in a range of bacterial species and has variable substrate spectra. The existence of quorum quenching enzymes in the quorum sensing microbes can attenuate their quorum sensing, leading to blocking unnecessary gene expression and pathogenic phenotypes. In this review, we discuss the physiological function of quorum quenching enzymes in bacterial infection and elucidate the enzymatic protection in quorum sensing systems for host diseases and their application in resistance against microbial diseases.     

儿童呼吸道易感病原菌多重PCR检测方法的建立

目的建立儿童呼吸道5种易感病原菌多重PCR(Multiplex PCR,mPCR)检测方法,并应用该方法分析北京地区儿童呼吸道易感病原菌菌群分布情况。方法分别设计针对肺炎链球菌(Streptococcus pneumoniae,SP)自溶素基因(ply)、肺炎克雷伯杆菌(Klebsiella pneumoniae,KP)16S核糖体RNA基因(16S rRNA)、金黄色葡萄球菌(Staphylococcus aureus,SA)热核酸酶基因(nuc)、表皮葡萄球菌(Staphylococcus epidermidis,SE)特异性保守基因(SeS)、流感嗜血杆菌(Haemophilus influenzae,HI)16S rRNA基因的特异性引物,建立mPCR检测方法;验证该方法的特异性及敏感性;并采用该方法对北京地区128例呼吸道感染患儿和80名健康儿童咽拭子或痰标本进行检测,并与传统培养法检测结果进行比对。结果 mPCR法能对同一样品中5种病原菌的DNA模板同时进行PCR扩增;该方法检测5种病原菌之间及与呼吸道其他易感病原之间无交叉反应,敏感性可达5×10-10μg/μl;mPCR法检测128例呼吸道感染患儿的菌群分布情况为:HI 24.22%、SP 15.63%、KP 4.69%,SA 2.34%,未检出SE,与培养法相比,两种检测方法的总符合率为78.91%;感染患儿与健康儿童相比,HI和SP的感染率差异有统计学意义(P<0.01)。结论建立了从呼吸道标本中直接检测多病原儿童易感病原菌的mPCR检测方法,该方法特异性强、敏感性高,为临床正确诊断和治疗细菌感染性疾病提供了参考。     

Recent Advances in Bacteria Identification by Matrix-Assisted Laser Desorption/Ionization Mass Spectrometry Using Nanomaterials as Affinity Probes

Identifying trace amounts of bacteria rapidly, accurately, selectively, and with high sensitivity is important to ensuring the safety of food and diagnosing infectious bacterial diseases. Microbial diseases constitute the major cause of death in many developing and developed countries of the world. The early detection of pathogenic bacteria is crucial in preventing, treating, and containing the spread of infections, and there is an urgent requirement for sensitive, specific, and accurate diagnostic tests. Matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) is an extremely selective and sensitive analytical tool that can be used to characterize different species of pathogenic bacteria. Various functionalized or unmodified nanomaterials can be used as affinity probes to capture and concentrate microorganisms. Recent developments in bacterial detection using nanomaterials-assisted MALDI-MS approaches are highlighted in this article. A comprehensive table listing MALDI-MS approaches for identifying pathogenic bacteria, categorized by the nanomaterials used, is provided.     

Rhamnolipids: Production in bacteria other than Pseudomonas aeruginosa

Rhamnolipids produced by Pseudomonas aeruginosa are the most studied biosurfactants due to their potential applications in a wide variety of industries and the high levels of their production. However, even though these biosurfactants are already produced at an industrial scale, the fact that P. aeruginosa is an opportunistic pathogen impose a restriction for its large scale production due to the intrinsic health hazard of the process. Other bacterial species that have been reported to be rhamnolipid producers are the pathogens Burkholderia mallei and B. pseudomallei, and recently the non‐pathogenic B. thailandensis. This short review presents information on rhamnolipid production by bacteria different from P. aeruginosa, as well as some approaches that have been taken to produce rhamnolipids using non‐pathogenic bacteria by genetic engineering of different bacteria. The low frequency of occurrence of rhamnolipid production among natural isolates that are not P. aeruginosa or Burkholderia, as well as the absence of orthologs of the genes involved in rhamnolipid synthesis (rhl genes) among the hundreds of sequenced bacterial genomes, suggest that the rare reported cases of these type of rhamnolipid‐producing bacteria have acquired this trait through horizontal gene transfer either from P. aeruginosa or from a member of Burkholderia.     

炼油废水COD降解菌固态菌剂的制备及性能考察

考察了几种吸附载体对炼油废水COD降解菌的吸附效果,确定以麦麸作为该菌种的吸附载体。固态菌剂保存在5℃条件下存活菌数较高,添加脱氧剂对菌种存活有促进作用。固态菌剂保存1 a后,菌种浓度仍能达到8.3×10~8CFU/g,比液态保存菌种高4.15×10~3倍。以保存期为1 a之内的固态菌剂降解炼油废水,其效果与新培养的液态菌种相当,COD去除率均在82%以上;当保存时间1 a时,固态菌剂对炼油废水的降解效果有所下降。     

Natural Quorum Sensing Inhibitors – Small Molecules,Big Messages

Due to the increasing emergence of antibiotic resistant bacterial strains within the past few decades, bacterial infections in general – and hospital-borne infections, in particular – have become increasingly difficult to fight. It is therefore crucial to find new strategies to fight pathogenic bacteria. Targeted inhibition of quorum sensing (QS) presents a promising alternative. QS is a cell density-dependent signaling pathway used for intra- and interspecies coordination of gene expression. In many bacteria, pathogenic phenotypes, as well as the expression of virulence factors, are under the control of QS regulons. A closer look at natural quorum sensing inhibitors may be helpful to identify potent compounds that can be used as alternatives to antibiotics. Moreover, it will also provide insight into the interactions between species that compete for the same habitat and resources. This review aims to summarize our current knowledge concerning natural QS inhibitors, as a starting point for the design and synthesis of new therapeutics to treat or prevent bacterial infections.     

Phage‐mediated transfer of virulence genes

Bacteriophages as accessory genetic elements play a crucial role in the dissemination of genes and the promotion of genetic diversity within bacterial populations. Such horizontal transfer of DNA is critical in the emergence of new pathogenic organisms, through the dissemination of genes encoding virulence factors such as toxins, adhesins and agressins. Phages can transfer genes that are not necessary for bacteriophage persistence and are generally recognised by their ability to convert their host bacteria to new phenotypes. This phenomenon is known as phage conversion. If such converting genes encode for virulence factors, the consequences of phage infection may include increased virulence of the host bacteria, and the conversion of a non‐pathogenic strain to a potentially dangerous pathogen. A number of virulence factors in bacteria causing diseases in plants, animals and humans are encoded by converting phages, the vast majority of which are temperate as opposed to lytic in nature. © 2001 Society of Chemical Industry     

Synthesis,PASS-Predication and in Vitro Antimicrobial Activity of Benzyl 4-O-benzoyl-α-l-rhamnopyranoside Derivatives

Benzyl α-l-rhamnopyranoside 4, obtained by both conventional and microwave assisted glycosidation techniques, was subjected to 2,3-O-isopropylidene protection to yield compound 5 which on benzoylation and subsequent deprotection of isopropylidene group gave the desired 4-O-benzoylrhamnopyranoside 7 in reasonable yield. Di-O-acetyl derivative of benzoate 7 was prepared to get newer rhamnopyranoside. The structure activity relationship (SAR) of the designed compounds was performed along with the prediction of activity spectra for substances (PASS) training set. Experimental studies based on antimicrobial activities verified the predictions obtained by the PASS software. Protected rhamnopyranosides 5 and 6 exhibited slight distortion from regular ¹C₄ conformation, probably due to the fusion of pyranose and isopropylidene ring. Synthesized rhamnopyranosides 4–8 were employed as test chemicals for in vitro antimicrobial evaluation against eight human pathogenic bacteria and two fungi. Antimicrobial and SAR study showed that the rhamnopyranosides were prone against fungal organisms as compared to that of the bacterial pathogens. Interestingly, PASS prediction of the rhamnopyranoside derivatives 4–8 were 0.49 < P_a < 0.60 (where P_a is probability ‘to be active’) as antibacterial and 0.65 < P_a < 0.73 as antifungal activities, which showed significant agreement with experimental data, suggesting rhamnopyranoside derivatives 4–8 were more active against pathogenic fungi as compared to human pathogenic bacteria thus, there is a more than 50% chance that the rhamnopyranoside derivative structures 4–8 have not been reported with antimicrobial activity, making it a possible valuable lead compound.     

Antifungal and antibacterial effects of 1‐monocaprylin on textile materials

The efficacy of a new antimicrobial treatment of textile materials based on the use of 1‐monocaprylin against various species of saprophytic moulds (Alternaria alternate CCM F‐397, Aspergillus niger ATCC 16404, Mucor racemosus CCM 8190, Penicillium ochrochloron CCM F‐158, Trichoderma viridae CCM F‐728), pathogenic moulds (Epidermophyton floccosum CCM 8339, Trichophyton mentagrophytes ATCC 9533, Trichophyton rubrum DSMZ 4167), pathogenic yeasts (Candida albicans ATCC 10231, Candida parapsilosis CCM 8260), Gram‐negative bacteria (Escherichia coli ATCC 11229, Klebsiella pneumoniae ATCC 4352) and Gram‐positive bacteria (Staphylococcus aureus ATCC 6853) was investigated. The testing was carried out according to DIN EN ISO 20645 disc‐diffusions test, using a 2‐layer method. The results showed that 1‐monocaprylin effectively killed all tested bacterial strains and pathogenic microorganisms with an exception of saprophytic moulds only, which were partially resistant. Textiles treated with 1‐monocaprylin reduced the growth of pathogenic, potentially dangerous microorganisms frequently occurring for example on the feet and in the shoes. Practical applications: To prevent microbial contamination leading to degradation of textile materials, various antimicrobial agents aimed at killing or suppressing of microorganism growth are applied. Among others, also MAGs belong to safe and efficient antimicrobial agents. Their application in antimicrobial treatment of textiles may be a suitable alternative to commercially used antimicrobial agents, as these endogenous lipid substances are present almost in all animal and plant tissues and are harmless to human body. The present study has confirmed that MAGs possess a suitable inhibitory activity when applied on textiles and are capable of hindering and even suppressing growth of bacteria and moulds that may occur during storage and use of textiles. The obtained results can be closely related to potential industrial applications of MAGs as effective agents for antimicrobial textiles and lining and insole materials for footwear, including those designed for diabetics.