The in vitro Antimicrobial Activity and Chemometric Modelling of 59 Commercial Essential Oils against Pathogens of Dermatological Relevance

This study reports on the inhibitory concentration of 59 commercial essential oils recommended for dermatological conditions, and identifies putative compounds responsible for antimicrobial activity. Essential oils were investigated for antimicrobial activity using minimum inhibitory concentration assays. Ten essential oils were identified as having superior antimicrobial activity. The essential oil compositions were determined using gas chromatography coupled to mass spectrometry and the data analysed with the antimicrobial activity using multivariate tools. Orthogonal projections to latent structures models were created for seven of the pathogens. Eugenol was identified as the main biomarker responsible for antimicrobial activity in the majority of the essential oils. The essential oils mostly displayed noteworthy antimicrobial activity, with five oils displaying broad‐spectrum activity against the 13 tested micro‐organisms. The antimicrobial efficacies of the essential oils highlight their potential in treating dermatological infections and through chemometric modelling, bioactive volatiles have been identified.     

肝病住院患者抗菌药物使用调查

目的了解肝病住院患者抗菌药物的使用情况。方法对62例肝病住院患者抗菌药物的应用进行调查。结果抗菌药物的使用率为25.81％;单用占75．00％;主要抗菌药物为哌拉西林、阿莫西林和头孢曲松;用药途径以静脉以及口服 静脉为主;以治疗性用药为主,预防用药在住院患者中的总使用率为4．84％;细菌标本送检率仅为6．25％。结论减少预防性用药,严格掌握抗菌药物的使用可降低抗菌药物的应用率;对于复数耐药菌感染应采取联合用药;提高细菌标本的送检率对于抗菌药物的合理应用十分重要;加强临床医师的抗菌药物知识更新和公众普及教育。     

抗菌肽改良设计及抗炎作用的研究进展

抗菌肽因其具有广谱抗菌活性、不容易引起抵抗性,被认为是先天免疫系统对抗微生物感染的多功能工具。然而,天然抗菌肽存在抗菌活性低、稳定性低、溶血性高等问题,使其较难应用于临床,所以研究人员对抗菌肽进行改良设计以期获得更高抗菌活性、更低溶血活性的新型抗菌肽。另外,天然抗菌肽作为一类免疫效应因子而被发现,其表现出的抑菌、免疫调节、内毒素中和等作用,使得研究人员对抗菌肽在抗炎作用的研究表现出极大的兴趣。就抗菌肽的药物设计方法及抗炎作用机制进行综述。     

九香虫抗菌肽CcAMP1的分离纯化和抗菌活性检测

【目的】从药用昆虫九香虫 Coridius chinensis 中分离纯化抗菌肽,为进一步开发九香虫抗菌肽资源及深入挖掘九香虫的药用功能奠定基础。【方法】用大肠杆菌Escherichia coli 和金黄色葡萄球菌 Staphylococcus aureus 混合物作诱导源刺激九香虫产生抗菌肽,对血淋巴进行提取、凝胶过滤层析、固相萃取及反相色谱纯化,活性组分经质谱测定。对分离得到的这种抗菌肽进行人工合成,并进行抗菌活性检测。【结果】本研究获得一种九香虫抗菌肽CcAMP1,由17个氨基酸残基组成,分子量为1 997.37 u,带1个正电荷,表面有5个疏水氨基酸。对人工合成的CcAMP1进行抗菌活性检测表明,该抗菌肽与九香虫血淋巴一样对金黄色葡萄球菌等革兰氏阳性菌和大肠杆菌等革兰氏阴性菌都有较好的抗菌活性,且对革兰氏阴性菌的抗菌活性更强。【结论】从九香虫中分离得到具有较强抗菌活性的阳离子抗菌肽CcAMP1,有较大的开发利用价值。     

A colorimetric assay for rapid screening of antimicrobial peptides

The increased resistance of various bacteria toward available antibiotic drugs has initiated intensive research efforts into identifying new sources of antimicrobial substances. Short antibiotic peptides (10-30 residues) are prevalent in nature as part of the intrinsic defense mechanisms of most organisms and have been proposed as a blueprint for the design of novel antimicrobial agents. Antimicrobial peptides are generally believed to kill bacteria through membrane permeabilization and extensive pore-formation. Assays providing rapid and easy evaluation of interactions between antimicrobial membrane peptides and lipid bilayers could significantly improve screening for substances with effective antibacterial properties, as well as contribute to the elucidation of structural and functional properties of antimicrobial peptides. Here we describe a colorimetric sensor in which particles composed of phospholipids and polymerized polydiacetylene (PDA) lipids were shown to exhibit striking color changes upon interactions with antimicrobial membrane peptides. The color changes in the system occur because of the structural perturbation of the lipids following their interactions with antimicrobial peptides. The assay was also sensitive to the antibacterial properties of structurally and functionally related peptide analogs.     

Diversity of antimicrobial peptides and their mechanisms of action

Antimicrobial peptides encompass a wide variety of structural motifs. Many peptides have alpha-helical structures. The majority of these peptides are cationic and amphipathic but there are also hydrophobic alpha-helical peptides which possess antimicrobial activity. In addition, some beta-sheet peptides have antimicrobial activity and even antimicrobial alpha-helical peptides which have been modified to possess a beta-structure retain part of their antimicrobial activity. There are also antimicrobial peptides which are rich in a certain specific amino acid such as Trp or His. In addition, antimicrobial peptides exist with thio-ether rings, which are lipopeptides or which have macrocyclic Cys knots. In spite of the structural diversity, a common feature of the cationic antimicrobial peptides is that they all have an amphipathic structure which allows them to bind to the membrane interface. Indeed, most antimicrobial peptides interact with membranes and may be cytotoxic as a result of disturbance of the bacterial inner or outer membranes. Alternatively, a necessary but not sufficient property of these peptides may be to be able to pass through the membrane to reach a target inside the cell. The interaction of these peptides with biological membranes is not just a function of the peptide but is also modulated by the lipid components of the membrane. It is not likely that this diverse group of peptides has a single mechanism of action, but interaction of the peptides with membranes is an important requirement for most, if not all, antimicrobial peptides.     

全基因组测序与生物信息学分析在细菌耐药性研究中的应用

随着耐药细菌的大量出现及广泛传播,细菌耐药性成为全球备受关注的问题。耐药细菌的特征如耐药基因、毒力因子、质粒分型等以及不同菌株间亲缘关系对于细菌耐药性流行病学及分子生物学的研究有着十分重要的意义。但是传统的技术手段如聚合酶链式反应(Polymerase chain reaction,PCR)和脉冲场凝胶电泳(Pulsed field gel electrophoresis,PFGE)等得到的结果不够全面且精确度低,对于现有的研究存在很大的局限性。全基因组测序技术(Whole genome sequencing,WGS)和生物信息学分析(Bioinformatics analysis)由于能够快速详尽地得到耐药细菌的特征,也能更加精细地判断不同菌株间的进化关系,逐渐成为更加有效的技术手段,为耐药性研究提供了有效的帮助。因此,文中系统地介绍全基因组测序分析流程中的各个步骤,主要包括文库构建、平台测序以及后期数据分析三大方面的不同方法和其相应的特点,期望相关研究人员对此能够有更全面的了解,并得到一定的帮助。     

The chemistry and applications of antimicrobial polymers: a state-of-the-art review

Microbial infection remains one of the most serious complications in several areas, particularly in medical devices, drugs, health care and hygienic applications, water purification systems, hospital and dental surgery equipment, textiles, food packaging, and food storage. Antimicrobials gain interest from both academic research and industry due to their potential to provide quality and safety benefits to many materials. However, low molecular weight antimicrobial agents suffer from many disadvantages, such as toxicity to the environment and short-term antimicrobial ability. To overcome problems associated with the low molecular weight antimicrobial agents, antimicrobial functional groups can be introduced into polymer molecules. The use of antimicrobial polymers offers promise for enhancing the efficacy of some existing antimicrobial agents and minimizing the environmental problems accompanying conventional antimicrobial agents by reducing the residual toxicity of the agents, increasing their efficiency and selectivity, and prolonging the lifetime of the antimicrobial agents. Research concerning the development of antimicrobial polymers represents a great a challenge for both the academic world and industry. This article reviews the state of the art of antimicrobial polymers primarily since the last comprehensive review by one of the authors in 1996. In particular, it discusses the requirements of antimicrobial polymers, factors affecting the antimicrobial activities, methods of synthesizing antimicrobial polymers, major fields of applications, and future and perspectives in the field of antimicrobial polymers.     

南极沉积物来源抗菌细菌的筛选及抑菌物质的鉴定

【背景】科学研究表明,由于南极环境条件特殊,微生物资源丰富,有望筛选出功效显著的抗菌微生物。【目的】以黄瓜枯萎病致病菌木贼镰刀菌(Fusarium equiseti)为指示菌,从南极沉积物中分离筛选具有拮抗作用的细菌菌株并对其抑菌物质进行初步鉴定。【方法】应用平板对峙法和琼脂扩散法分别对样品和发酵液进行初筛和复筛,筛选出对F. equiseti抑菌效果最强的菌株,基于形态学、生理生化、分子生物学分析,对该菌株进行鉴定。之后,对目标菌株发酵上清中的抑菌物质进行抑菌谱研究,并对其抑菌成分进行温度和pH的稳定性检测,通过硫酸铵沉淀的方法初步鉴定发酵液中的抑菌物质。【结果】从南极沉积物样品中共分离纯化出62株细菌,有5株具有较好的抑菌效果,其中抑菌效果最强的菌株鉴定为枯草芽孢杆菌斯氏亚种(Bacillus subtilis subsp.spizizenii),命名为JYM35。抑菌谱检测结果显示,菌株JYM35对丝瓜枯萎病致病菌层生镰刀菌(Fusarium proliferatum)和辣椒枯萎病致病菌F. equiseti具有较强的抑菌效果,对长豆褐腐病致病菌笄霉属(Choanephora)有较明显的拮抗作用,对水产致病菌副溶血弧菌(Vibrio parahaemolyticus)和溶藻弧菌(V.alginolyticus)也有一定的拮抗作用。菌株JYM35发酵上清中所含的抑菌物质热稳定性强且耐碱不耐酸,硫酸铵沉淀后可初步确定其抑菌物质隶属蛋白类。【结论】菌株JYM35是一株产蛋白类活性物质的广谱型抑菌菌株,对枯萎病致病菌的拮抗作用最强。因此具有较好的开发利用价值。     

Expression, purification and antimicrobial activity of puroindoline A protein and its mutants

Wheat puroindoline proteins, PINA and PINB, play key roles in determining wheat grain hardness as well as in defending the plant against pathogens. PINA has much greater membrane-binding property and antimicrobial activity because it contains more tryptophan residues in the unique tryptophan-rich domain (TRD). In order to obtain proteins with higher antimicrobial activity, mutants of PINA containing two or three copies of TRD, designated ABBC and ABBBC, respectively, were constructed and expressed in E. coli Rosetta-gami (DE3). Metal affinity chromatography was used to purify the soluble affinity-tagged recombinant proteins. The secondary structures of the recombinant proteins were predicted by the online program Protein Homology/analog Y Recognition Engine v2.0 and experimentally assessed using circular dichroism. Minimum inhibition concentration tests and fluorescence microscope analyses were employed to evaluate the antimicrobial activities of the mutants. The results showed that the purified recombinant ABBC was correctly folded and presented significantly higher antimicrobial activities against E. coli and S. aureus than wild-type PINA, suggesting its potential use as an antimicrobial agent. The results also confirmed that TRD is a determinant of the antimicrobial activity of PINA and demonstrated that it is feasible to enhance the antimicrobial activity of PINA by adding one copy of TRD.     

Anti-infection peptidomics of amphibian skin

Peptidomics and genomics analyses were used to study an anti-infection array of peptides of amphibian skin. 372 cDNA sequences of antimicrobial peptides were characterized from a single individual skin of the frog Odorrana grahami that encode 107 novel antimicrobial peptides. This contribution almost triples the number of currently reported amphibian antimicrobial peptides. The peptides could be organized into 30 divergent groups, including 24 novel groups. The diversity in peptide coding cDNA sequences is, to our knowledge, the most extreme yet described for any animal. The patterns of diversification suggest that point mutations as well as insertion, deletion, and "shuffling" of oligonucleotide sequences were responsible for the diversity. The diversity of antimicrobial peptides may have resulted from the diversity of microorganisms. These diverse peptides exhibited both diverse secondary structure and "host defense" properties. Such extreme antimicrobial peptide diversity in a single amphibian species is amazing. This has led us to reconsider the strong capability of innate immunity and molecular genetics of amphibian ecological diversification and doubt the general opinion that 20-30 different antimicrobial peptides can protect an animal because of the relatively wide specificity of the peptide antibiotics. The antimicrobial mechanisms of O. grahami peptides were investigated. They exerted their antimicrobial functions by various means, including forming lamellar mesosome-like structures, peeling off the cell walls, forming pores, and inducing DNA condensation. With respect to the development of antibiotics, these peptides provide potential new templates to explore further.     

Sensitivity of environmental microorganisms to antimicrobial agents.

The sensitivity of different microorganisms, considered as typical representatives of the microflora of soil and water, was established to evaluate the influence of the nonmedical use of antimicrobial agents on bacterial ecology. Only seven strains, six chemoorganotrophs and one chemolithotroph, could be considered as relatively sensitive to the 21 antimicrobial compounds tested. The other 29 microorganisms may be regarded as resistant to most antimicrobial agents. Streptomyces were sensitive to high concentrations of active substances. Broad-spectrum antibiotics showed an effect on environmental bacteria similar to that on human pathogens. Cephalothin stimulated the growth of a Chlorella sp. From these experiments, it appears that spilled antimicrobial agents have little chance of causing an alteration in the microbial ecology.     

Antimicrobial activity of the C-terminal of the major royal jelly protein 4 in a honeybee (Apis cerana)

The protein component of honeybee royal jelly (RJ) is constituted by major royal jelly proteins (MRJPs). The Asiatic honeybee (Apis cerana) MRJP-4 (AcMRJP4) exhibits antimicrobial activities. In this study, we identified the antimicrobial activity of AcMRJP4-15, which is a hydrophilic peptide with 88 amino acid residues in the C-terminal of AcMRJP4 that contains a high content of Asn and positively charged amino acids. Recombinant AcMRJP4-15, which is expressed as a 15-kDa peptide in baculovirus-infected insect cells, induced structural damage to the cell walls of bacteria, fungi, and yeast. Interestingly, the antimicrobial activity of AcMRJP4-15 was greater than that of AcMRJP4, demonstrating that the antimicrobial activity of AcMRJP4 was due in large part to the C-terminal. Our data suggest that AcMRJP4-15 can function as an effective antimicrobial agent.     

Structure-activity relationships of antimicrobial peptides from the skin of Rana esculenta inhabiting in Korea

The anuran (frogs and toads) skin is a rich source of antimicrobial peptides that can be developed therapeutically. We searched the skin secretions of Korean Rana esculenta for antimicrobial peptides, and isolated two cationic peptides with antimicrobial activity and little hemolytic activity: a 46-residue peptide of the esculentin-1 family and a 24-residue peptide of the brevinin-1 family. Their sequences showed some differences from the esculentins-1 and brevinins-1 of European Rana esculenta, indicating that sequence diversification of anuran skin antimicrobial peptides can arise from differences in habitat as well as from species differences. The 46-residue peptide named esculentin-1c had broad antimicrobial activity, while the 24-residue peptide named brevinin-1Ed exhibited limited activity. The solution structure of brevinin-1Ed was in good agreement with that of other brevinin-1-like peptides, with an amphipathic alpha-helix spanning residues 3-20, stabilized in membrane-mimetic environments. The weak bioactivity of brevinin-1Ed was attributable to the unusual presence of an anionic amino acid in the middle of the helical hydrophilic face. This report contributes to world-wide investigations of the structure-activity relationships and evolutional diversification of anuran-skin antimicrobial peptides.     

芳香植物精油的抗菌性及在动物生产中的应用

芳香植物精油为具特征性气味的挥发性油状液体, 是从芳香植物中提取的一种重要次生代谢物质。芳香植物精油的抗菌活性由其化学成分和浓度决定, 其中酚类、含氧萜类和萜烯类在抗菌方面表现出较强的活性。芳香植物精油的抗菌机制主要涉及脂肪酸外膜的改变、细胞质膜的损坏、质子动力的消耗、代谢物及离子泄露。在畜牧业生产体系中, 抗生素的无序使用不仅可能引发“超级细菌”的产生, 其残留亦会造成畜产品不安全和环境污染。芳香植物精油作为一种天然植物抗菌剂, 毒性较低且无残留, 作为饲料添加剂可用于维持动物机体的健康, 有望成为重要的抗生素替代品。该文阐述了芳香植物精油的活性成分、抗菌作用机制及其在动物生产中的应用, 为抗菌机理研究和新技术开发利用提供了理论依据。     

Preservation of antimicrobial properties of complement peptide C3a, from invertebrates to humans

The human anaphylatoxin peptide C3a, generated during complement activation, exerts antimicrobial effects. Phylogenetic analysis, sequence analyses, and structural modeling studies paired with antimicrobial assays of peptides from known C3a sequences showed that, in particular in vertebrate C3a, crucial structural determinants governing antimicrobial activity have been conserved during the evolution of C3a. Thus, regions of the ancient C3a from Carcinoscorpius rotundicauda as well as corresponding parts of human C3a exhibited helical structures upon binding to bacterial lipopolysaccharide permeabilized liposomes and were antimicrobial against gram-negative and gram-positive bacteria. Human C3a and C4a (but not C5a) were antimicrobial, in concert with the separate evolutionary development of the chemotactic C5a. Thus, the results demonstrate that, notwithstanding a significant sequence variation, functional and structural constraints imposed on C3a during evolution have preserved critical properties governing antimicrobial activity.     

Honeybee (Apis cerana) major royal jelly protein 4 exhibits antimicrobial activity

Major royal jelly proteins (MRJPs) are important protein components of bee royal jelly (RJ) and exhibit various biological and pharmacological activities. The antimicrobial activities of the royalisin and the jelleines contained within MRJP 1 and MRJP 2 in RJ have been elucidated. However, the antimicrobial effects of other MRJPs remain largely unknown. In this study, we demonstrated the antimicrobial activity of the Asiatic honeybee (Apis cerana) MRJP 4 (AcMRJP4). Recombinant AcMRJP4 was expressed as a 63-kDa protein in baculovirus-infected insect cells. We examined the antimicrobial activity of recombinant AcMRJP4 against bacteria, fungi, and yeast. The mechanisms underlying the antimicrobial activity of AcMRJP4 were assessed using western blot analysis, immunofluorescence staining, and scanning electron microscopy. Recombinant AcMRJP4 bound to the cell walls of bacteria, fungi, and yeast and induced structural damage in the microbial cell walls. AcMRJP4 has an antimicrobial role and exhibits a broad spectrum of antimicrobial activities against bacteria, fungi, and yeast. We demonstrated that AcMRJP4 functions as an antimicrobial agent with activity against bacteria, fungi, and yeast. Together, our data identified a novel function of MRJP 4 as an antimicrobial agent.     

Antimicrobial activity of ultrasound-assisted solvent-extracted spices

AIMS: The objective of this research was to determine the antimicrobial activity of conventional and high-intensity ultrasound-assisted (HI-US) solvent-extracted Thai spices, including ginger (Zingiber officinale Rose), fingerroot (Bosenbergia pandurata Holtt) and turmeric (Curouma longa Linn). METHODS AND RESULTS: Extracts were obtained using hexane, isopropanol and a 7 : 3 isopropanol : hexane mixture as solvents with and without HI-US. The antimicrobial activity of the extracts was assayed against four strains each of Listeria monocytogenes and Salmonella Typhimurium DT 104 using an agar dilution assay. Application of HI-US did not alter antibacterial activity against S. Typhimurium, but antilisterial activity of some HI-US spice extracts decreased. Solvent type affected antimicrobial efficacy of extracts with hexane producing the least antimicrobial activity. Fingerroot extracted with isopropanol-hexane and without HI-US had the best antilisterial effect while HI-US-isopropanol fingerroot extract had the greatest antimicrobial efficacy against S. Typhimurium. CONCLUSIONS: Application of HI-US reduced time of extraction to 5 min, compared with the 24 h required for conventional extraction and maintained antimicrobial activity against Salmonella but slightly reduced activity against Listeria. SIGNIFICANCE AND IMPACT OF THE STUDY: HI-US in combination with proper solvent selection may offer a new tool to optimize extraction of spice essential oil for use as antimicrobial agents, and reduce processing time and costs.     

Structure-Activity Relationship of a U-Type Antimicrobial Microemulsion System

The structure-activity relationship of a U-type antimicrobial microemulsion system containing glycerol monolaurate and ethanol at a 1∶1 mass ratio as oil phase and Tween 20 as surfactant were investigated along a water dilution line at a ratio of 80∶20 mass% surfactant/oil phase, based on a pseudo-ternary phase diagram. The differential scanning calorimetry results showed that in the region of up to 33% water, all water molecules are confined to the hydrophilic core of the reverse micelles, leading to the formation of w/o microemulsion. As the water content increases, the water gains mobility, and transforms into bicontinuous in the region of 33–39% water, and finally the microemulsion become o/w in the region of above 39% water. The microstructure characterization was confirmed by the dynamic light scattering measurements and freeze-fracture transmission electron microscope observation. The antimicrobial activity assay using kinetics of killing analysis demonstrated that the microemulsions in w/o regions exhibited relatively high antimicrobial activity against Escherichia coli and Staphylococcus aureus due to the antimicrobial oil phase as the continuous phase, while the antimicrobial activity started to decrease when the microemulsions entered the bicontinuous region, and decreased rapidly as the water content increased in the o/w region, as a result of the dilution of antimicrobial oil droplets in the aqueous continuous phase.     

Isolation and identification of antimicrobial components from the epidermal mucus of Atlantic cod (Gadus morhua)

The epidermal mucus of fish species has been found to contain antimicrobial proteins and peptides, which is of interest in regard to fish immunity. An acidic extract from the epidermal mucus of the Atlantic cod (Gadus morhua) was found to exhibit antimicrobial activity against Bacillus megaterium, Escherichia coli and Candida albicans. This activity varied significantly when salt was added to the antimicrobial assay, and was eliminated by pepsin digestion. No lysozyme activity was detected in the extract. By using weak cationic exchange chromatography together with reversed-phase chromatography, and monitoring the antimicrobial activity, we have isolated four cationic proteins from the mucus extract. Using N-terminal and C-terminal amino acid sequence analysis, together with MS, the antimicrobial proteins were identified as histone H2B (13 565 Da), ribosomal protein L40 (6397 Da), ribosomal protein L36A (12 340 Da) and ribosomal protein L35 (14 215 Da). The broad spectra of antimicrobial activities in the cod mucus and the characterization of four antimicrobial polypeptides suggest that mucus compounds contribute to the innate host defence of cod.