中药单体抑制金黄色葡萄球菌生物被膜形成的研究进展

金黄色葡萄球菌是一种临床上十分常见的病原细菌,其在医疗器械和植入物上形成的生物被膜赋予了金黄色葡萄球菌极强的抗生素耐药性,导致相关慢性感染难以彻底根除,从而对人类健康产生极大的威胁。天然产物来源的活性单体成分在抑制金黄色葡萄球菌生物被膜形成方面发挥了重要的作用,为抗细菌生物被膜研究提供了新策略。围绕金黄色葡萄球菌生物被膜形成过程中的分子调控机制,以及中药单体的抑制机制展开介绍,旨在为抑制细菌生物被膜感染的中医药基础研究提供理论参考。     

十二烷基苯磺酸钠对引发内置物感染表皮葡萄球菌生物被膜的作用

【背景】随着医用内置物的广泛使用,由表皮葡萄球菌生物被膜导致的医院获得性感染不断增多,目前鲜见关于表面活性剂针对表皮葡萄球菌生物被膜作用的报道。【目的】通过研究阴离子型表面活性剂十二烷基苯磺酸钠(sodium dodecyl benzene sulfonate,SDBS)分别对ATCC 35984 (产膜表皮葡萄球菌标准株)生物被膜的清除、生物被膜内细菌代谢和形成生物被膜的关键物质多糖胞间黏附素(polysaccharide intercellular adhesion,PIA)产生的影响,为临床使用SDBS防治由表皮葡萄球菌生物被膜引起的相关感染提供可靠的理论及实践依据。【方法】利用XTT减低法,评价SDBS对ATCC 35984已形成生物被膜的清除效率及对生物被膜内细菌代谢的影响;激光共聚焦显微镜观察SDBS对生物被膜作用的效果;采用刚果红培养基观察SDBS对PIA产生的影响。【结果】浓度为256、128、64、32、16 mg/L的SDBS在作用6、12、24 h时,对ATCC 35984的生物被膜均有显著的清除效率(P<0.01);浓度为32 mg/L时对生物被膜内细菌的...     

c-di-AMP调控细菌生物被膜的形成

细菌生物被膜是细菌持续性致病的重要机制。研究细菌生物被膜的形成和发展可为顽固性细菌感染防治提供新的思路与策略。环二腺苷酸c-di-AMP(Cyclic diadenosine monophosphate)是继c-di-GMP之后在细菌中新发现的一种核苷酸第二信使分子。研究发现,c-di-AMP参与调节细菌多种生理功能,包括细菌生长代谢、生物被膜形成、细胞壁的合成以及细菌毒力因子等。本文综述了c-di-AMP参与调控细菌生物被膜形成的不同方式及其分子机制。鉴于c-di-AMP在调控细菌生物被膜中的重要性,其可作为抗细菌生物被膜感染新药研发的潜在靶点。     

细菌生物被膜分散及分子调控机制研究进展

生物被膜分散(Biofilm Dispersal)是生物被膜发展后期细菌响应营养物、低浓度的一氧化氮、D-氨基酸、自诱导肽(Autoinducing Peptide,AIP)、酰基高丝氨酸内酯(Acyl Homoserine Lactones,AHL)、腺苷三磷酸(Adenosine Triphosphate,ATP)等信号变化而做出的一种程序性反应,有利于细菌从恶劣的生物被膜内部环境中脱离出来寻找新的定殖位点。此外,由生物被膜引起的细菌短暂的抗生素耐受性在分散过程中会恢复正常水平,这有助于治疗由致病菌引起的难治愈的生物被膜相关疾病。目前生物被膜分散的相关研究正处于起步阶段,本文希望通过综述生物被膜分散现象、信号分子及调控机制,可以更好地了解细菌生物被膜分散对于防控病原微生物和应用有益微生物的重要意义。     

细菌显微追踪技术在生物被膜中的应用

细菌生物被膜是粘附于物体表面的由细菌细胞及其胞外物质组成的复杂膜样物聚集体,具有很强的耐药性和免疫逃逸能力。生物被膜内细菌的代谢活性、运动状态等与浮游细菌有明显区别。近年来,先进的显微成像技术结合新型图像处理方法,在研究细菌的运动、生理等方面发挥了重要作用。本文围绕生物被膜,概述了细菌显微追踪技术在其研究中的应用。主要从细菌的运动方式和生物被膜形成过程的调控两方面出发,介绍了在单细胞水平上利用该技术研究生物被膜的进展,包括细菌的游泳、蹭行、群集运动和多种信号通路调控下生物被膜的形成过程等,并展望了该技术在生物被膜其他相关研究领域的应用前景。     

细菌生物被膜基质的研究进展

细菌生物被膜(biofilm)附着在生物或者非生物表面,由细菌及其分泌的糖、蛋白质和核酸等多种基质组成的细菌群落,是造成病原细菌持续性感染、毒力和耐药性的重要原因之一.细菌的生物被膜基质由复杂的胞外聚合物(extracellular polymeric substances,EPS)构成,影响生物被膜的结构和功能.本文...     

葡萄球菌生物被膜的基因调控机制研究进展

摘要:细菌的耐药性问题是目前医学临床面临的严峻问题,其中细菌生物被膜的形成是引起细菌持续性感染的主要致病机制之一。细菌生物被膜的形成过程十分复杂,受多种因子和多基因的共同调控,且不同的因子和基因在生物被膜形成的不同阶段所起的作用不同。本文重点对引起院内感染的主要致病菌葡萄球菌的生物被膜形成的基因调控机制,以及药物抑制葡萄球菌生物被膜的研究现状进行综述,旨在为解决医学临床中存在的细菌感染,研制抗生物被膜药物和疫苗等提供参考。     

植物提取物及其活性成分抑制细菌生物被膜的研究进展北大核心CSCD

大量研究报道生物被膜细菌对抗生素的耐药性是浮游菌的10–1 000倍,据报道细菌生物被膜是80%以上细菌感染的罪魁祸首,对医疗保健领域构成了严峻的挑战。植物提取物及其活性成分对细菌生物被膜有明显的抑制作用,包括减少生物被膜量、生物被膜活菌数以及清除已经成熟的生物被膜等。该文对这些有效的植物提取物及其活性成分进行了总结,并分析了其抗细菌生物被膜的作用机制。旨在为防治细菌生物被膜感染的植物类药物的开发提供参考。     

铜绿假单胞菌生物被膜组成及其受群体感应系统和c-di-GMP调控的研究进展

作为人类条件性感染的前三大病原菌之一的铜绿假单胞菌,是一种革兰氏阴性细菌,对免疫功能低下和囊性纤维化患者可以造成严重和持续性感染。造成这种持续感染的原因主要是由于细菌接收外界信号后,在自身调控网络的协同作用下,会依附于固体表面,并产生胞外多糖、基质蛋白和胞外DNA等大分子物质形成高度结构化的膜状复合物将自身包裹形成生物被膜群体结构。生物被膜可以有效帮助细菌定殖、提高细菌对抗菌物质和宿主免疫反应的抵抗能力、促进群落细菌的细胞-细胞之间的信号交流等,是临床治疗中病原菌慢性感染和反复感染最重要的原因之一。本篇综述重点介绍了铜绿假单胞菌生物被膜的各组成成分及其在生物被膜形成中的重要功能,并进一步阐述了群体感应系统(las、rhl、pqs与iqs)和c-di-GMP对铜绿假单胞菌生物被膜形成的调控作用。通过本篇综述可以更清晰地了解细菌生物被膜形成和调控的过程,为开发新的治疗生物被膜感染策略提供帮助。     

柱[5]芳烃衍生物对细菌抗菌活性及生物被膜抑制的研究进展北大核心

病原体的耐药性很强,其生物被膜(biofilm,BF)的形成是导致耐药性的主要原因之一。生物被膜一旦形成,根除难度很大,会导致患者持久性感染,引发多种慢性疾病,并给全球医疗体系带来沉重负担。柱芳烃(pillararenes)是一类具有独特柱状结构的新型大环化合物,由于其在构建功能化和生物活性材料开发中的潜在应用引起人们广泛的关注。此外,它们在预防和控制抗生素耐药性(antimicrobial resistance,AMR)方面具有广阔的应用前景。本文综述了柱[5]芳烃衍生物对细菌病原菌的抗菌活性,并进一步揭示其在抗菌活性中的抑菌机制,尤其是对生物被膜的抑制作用。在此基础上,探索新的抑菌杀菌策略,用非传统药物以解决抗生素耐药性问题,以期为开发新的抗菌剂防控生物被膜或治疗细菌感染提供理论依据。     

Evaluation of the Enterococcus faecalis Biofilm-Associated Virulence Factors AhrC and Eep in Rat Foreign Body Osteomyelitis and In Vitro Biofilm-Associated Antimicrobial Resistance

Enterococcus faecalis can cause healthcare-associated biofilm infections, including those of orthopedic devices. Treatment of enterococcal prosthetic joint infection is difficult, in part, due to biofilm-associated antimicrobial resistance. We previously showed that the E. faecalis OG1RF genes ahrC and eep are in vitro biofilm determinants and virulence factors in animal models of endocarditis and catheter-associated urinary tract infection. In this study, we evaluated the role of these genes in a rat acute foreign body osteomyelitis model and in in vitro biofilm-associated antimicrobial resistance. Osteomyelitis was established for one week following the implantation of stainless steel orthopedic wires inoculated with E. faecalis strains OG1RF, ΩahrC, and ∆eep into the proximal tibiae of rats. The median bacterial loads recovered from bones and wires did not differ significantly between the strains at multiple inoculum concentrations. We hypothesize that factors present at the infection site that affect biofilm formation, such as the presence or absence of shear force, may account for the differences in attenuation in the various animal models we have used to study the ΩahrC and ∆eep strains. No differences among the three strains were observed in the planktonic and biofilm antimicrobial susceptibilities to ampicillin, vancomycin, daptomycin, linezolid, and tetracycline. These findings suggest that neither ahrC nor eep directly contribute to E. faecalis biofilm-associated antimicrobial resistance. Notably, the experimental evidence that the biofilm attachment mutant ΩahrC displays biofilm-associated antimicrobial resistance suggests that surface colonization alone is sufficient for E. faecalis cells to acquire the biofilm antimicrobial resistance phenotype.     

植入物动物感染模型在葡萄球菌生物膜研究中的应用与进展

葡萄球菌生物膜引起的持续性感染及耐药性问题一直是临床治疗的难题,围绕生物膜形成分子机制的研究成为防治葡萄球菌生物膜相关感染的关键。建立葡萄球菌感染动物模型有利于研究体内生物膜形成、扩散、致病机制及药物的体内抗生物膜效果评估等。然而,动物体内生物膜形成的影响因素多,如动物种类、植入材料、接种部位、感染剂量、观察时间及评估方法等均会影响体内生物膜形成。结合本课题研究,系统地总结了近40年来葡萄球菌生物膜感染动物模型,重点综述动物模型的建立方法、适用范围及优缺点,为葡萄球菌生物膜感染的防治提供理论依据。     

A Systems-Level Approach for Investigating Pseudomonas aeruginosa Biofilm Formation

Prevention of the initiation of biofilm formation is the most important step for combating biofilm-associated pathogens, as the ability of pathogens to resist antibiotics is enhanced 10 to 1000 times once biofilms are formed. Genes essential to bacterial growth in the planktonic state are potential targets to treat biofilm-associated pathogens. However, the biofilm formation capability of strains with mutations in these essential genes must be evaluated, since the pathogen might form a biofilm before it is eliminated. In order to address this issue, this work proposes a systems-level approach to quantifying the biofilm formation capability of mutants to determine target genes that are essential for bacterial metabolism in the planktonic state but do not induce biofilm formation in their mutants. The changes of fluxes through the reactions associated with the genes positively related to biofilm formation are used as soft sensors in the flux balance analysis to quantify the trend of biofilm formation upon the mutation of an essential gene. The essential genes whose mutants are predicted not to induce biofilm formation are regarded as gene targets. The proposed approach was applied to identify target genes to treat Pseudomonas aeruginosa infections. It is interesting to find that most essential gene mutants exhibit high potential to induce the biofilm formation while most non-essential gene mutants do not. Critically, we identified four essential genes, lysC, cysH, adk, and galU, that constitute gene targets to treat P. aeruginosa. They have been suggested by existing experimental data as potential drug targets for their crucial role in the survival or virulence of P. aeruginosa. It is also interesting to find that P. aeruginosa tends to survive the essential-gene mutation treatment by mainly enhancing fluxes through 8 metabolic reactions that regulate acetate metabolism, arginine metabolism, and glutamate metabolism.     

Mycobacterium avium Possesses Extracellular DNA that Contributes to Biofilm Formation,Structural Integrity,and Tolerance to Antibiotics

Mycobacterium avium subsp. hominissuis is an opportunistic pathogen that is associated with biofilm-related infections of the respiratory tract and is difficult to treat. In recent years, extracellular DNA (eDNA) has been found to be a major component of bacterial biofilms, including many pathogens involved in biofilm-associated infections. To date, eDNA has not been described as a component of mycobacterial biofilms. In this study, we identified and characterized eDNA in a high biofilm-producing strain of Mycobacterium avium subsp. hominissuis (MAH). In addition, we surveyed for presence of eDNA in various MAH strains and other nontuberculous mycobacteria. Biofilms of MAH A5 (high biofilm-producing strain) and MAH 104 (reference strain) were established at 22°C and 37°C on abiotic surfaces. Acellular biofilm matrix and supernatant from MAH A5 7 day-old biofilms both possess abundant eDNA, however very little eDNA was found in MAH 104 biofilms. A survey of MAH clinical isolates and other clinically relevant nontuberculous mycobacterial species revealed many species and strains that also produce eDNA. RAPD analysis demonstrated that eDNA resembles genomic DNA. Treatment with DNase I reduced the biomass of MAH A5 biofilms when added upon biofilm formation or to an already established biofilm both on abiotic surfaces and on top of human pharyngeal epithelial cells. Furthermore, co-treatment of an established biofilm with DNase 1 and either moxifloxacin or clarithromycin significantly increased the susceptibility of the bacteria within the biofilm to these clinically used antimicrobials. Collectively, our results describe an additional matrix component of mycobacterial biofilms and a potential new target to help treat biofilm-associated nontuberculous mycobacterial infections.     

Biofilms: strategies for metal corrosion inhibition employing microorganisms

Corrosion causes dramatic economic loss. Currently widely used corrosion control strategies have disadvantages of being expensive, subject to environmental restrictions, and sometimes inefficient. Studies show that microbial corrosion inhibition is actually a common phenomenon. The present review summarizes recent progress in this novel strategy: corrosion control using beneficial bacteria biofilms. The possible mechanisms may involve: (1) removal of corrosive agents (such as oxygen) by bacterial physiological activities (e.g., aerobic respiration), (2) growth inhibition of corrosion-causing bacteria by antimicrobials generated within biofilms [e.g., sulfate-reducing bacteria (SRB) corrosion inhibition by gramicidin S-producing Bacillus brevis biofilm], (3) generation of protective layer by biofilms (e.g., Bacillus licheniformis biofilm produces on aluminum surface a sticky protective layer of γ-polyglutamate). Successful utilization of this novel strategy relies on advances in study at the interface of corrosion engineering and biofilm biology.     

Staphylococcal biofilm disassembly

Staphylococcus aureus and Staphylococcus epidermidis are a frequent cause of biofilm-associated infections that are a tremendous burden on our healthcare system. Staphylococcal biofilms exhibit extraordinary resistance to antimicrobial killing, limiting the efficacy of antibiotic therapy, and surgical intervention is often required to remove infected tissues or implanted devices. Recent work has provided new insight into the molecular basis of biofilm development in these opportunistic pathogens. Extracellular bacterial products, environmental conditions, and polymicrobial interactions have all been shown to influence profoundly the ability of these bacteria to colonize and disperse from clinically relevant surfaces. We review new developments in staphylococcal biofilm disassembly and set them in the context of potential strategies to control biofilm infections.     

Unique Biofilm Signature,Drug Susceptibility and Decreased Virulence in Drosophila through the Pseudomonas aeruginosa Two-Component System PprAB

Bacterial biofilm is considered as a particular lifestyle helping cells to survive hostile environments triggered by a variety of signals sensed and integrated through adequate regulatory pathways. Pseudomonas aeruginosa, a Gram-negative bacterium causing severe infections in humans, forms biofilms and is a fantastic example for fine-tuning of the transition between planktonic and community lifestyles through two-component systems (TCS). Here we decipher the regulon of the P. aeruginosa response regulator PprB of the TCS PprAB. We identified genes under the control of this TCS and once this pathway is activated, analyzed and dissected at the molecular level the PprB-dependent phenotypes in various models. The TCS PprAB triggers a hyper-biofilm phenotype with a unique adhesive signature made of BapA adhesin, a Type 1 secretion system (T1SS) substrate, CupE CU fimbriae, Flp Type IVb pili and eDNA without EPS involvement. This unique signature is associated with drug hyper-susceptibility, decreased virulence in acutely infected flies and cytotoxicity toward various cell types linked to decreased Type III secretion (T3SS). Moreover, once the PprB pathway is activated, decreased virulence in orally infected flies associated with enhanced biofilm formation and dissemination defect from the intestinal lumen toward the hemolymph compartment is reported. PprB may thus represent a key bacterial adaptation checkpoint of multicellular and aggregative behavior triggering the production of a unique matrix associated with peculiar antibiotic susceptibility and attenuated virulence, a particular interesting breach for therapeutic intervention to consider in view of possible eradication of P. aeruginosa biofilm-associated infections.     

Ellagic acid derivatives from Rubus ulmifolius inhibit Staphylococcus aureus biofilm formation and improve response to antibiotics

Background

Biofilms contribute to the pathogenesis of many forms of Staphylococcus aureus infection. Treatment of these infections is complicated by intrinsic resistance to conventional antibiotics, thus creating an urgent need for strategies that can be used for the prevention and treatment of biofilm-associated infections.

Methodology/Principal Findings

This study demonstrates that a botanical natural product composition (220D-F2) rich in ellagic acid and its derivatives can limit S. aureus biofilm formation to a degree that can be correlated with increased antibiotic susceptibility. The source of this composition is Rubus ulmifolius Schott. (Rosaceae), a plant used in complementary and alternative medicine in southern Italy for the treatment of skin and soft tissue infections. All S. aureus clonal lineages tested exhibited a reduced capacity to form a biofilm at 220D-F2 concentrations ranging from 50–200 µg/mL, which were well below the concentrations required to limit bacterial growth (530–1040 µg/mL). This limitation was therapeutically relevant in that inclusion of 220D-F2 resulted in enhanced susceptibility to the functionally-distinct antibiotics daptomycin, clindamycin and oxacillin. Testing with kidney and liver cell lines also demonstrated a lack of host cell cytotoxicity at concentrations of 220D-F2 required to achieve these effects.

Conclusions/Significance

These results demonstrate that extract 220D-F2 from the root of Rubus ulmifolius can be used to inhibit S. aureus biofilm formation to a degree that can be correlated with increased antibiotic susceptibility without toxic effects on normal mammalian cells. Hence, 220D-F2 is a strong candidate for development as a botanical drug for use in the prevention and treatment of S. aureus biofilm-associated infections.     

A novel bacteriophage cocktail reduces and disperses Pseudomonas aeruginosa biofilms under static and flow conditions

Pseudomonas aeruginosa is an opportunistic human pathogen that forms highly stable communities – biofilms, which contribute to the establishment and maintenance of infections. The biofilm state and intrinsic/acquired bacterial resistance mechanisms contribute to resistance/tolerance to antibiotics that is frequently observed in P. aeruginosa isolates. Here we describe the isolation and characterization of six novel lytic bacteriophages: viruses that infect bacteria, which together efficiently infect and kill a wide range of P. aeruginosa clinical isolates. The phages were used to formulate a cocktail with the potential to eliminate P. aeruginosa PAO1 planktonic cultures. Two biofilm models were studied, one static and one dynamic, and the phage cocktail was assessed for its ability to reduce and disperse the biofilm biomass. For the static model, after 4 h of contact with the phage suspension (MOI 10) more than 95% of biofilm biomass was eliminated. In the flow biofilm model, a slower rate of activity by the phage was observed, but 48 h after addition of the phage cocktail the biofilm was dispersed, with most cells eliminated (> 4 logs) comparing with the control. This cocktail has the potential for development as a therapeutic to control P. aeruginosa infections, which are predominantly biofilm centred.