Metabolic profiling as a tool for prioritizing antimicrobial compounds

Metabolomics is an analytical technique that allows scientists to globally profile low molecular weight metabolites between samples in a medium- or high-throughput environment. Different biological samples are statistically analyzed and correlated to a bioactivity of interest, highlighting differentially produced compounds as potential biomarkers. Here, we review NMR- and MS-based metabolomics as technologies to facilitate the identification of novel antimicrobial natural products from microbial sources. Approaches to elicit the production of poorly expressed (cryptic) molecules are thereby a key to allow statistical analysis of samples to identify bioactive markers, while connection of compounds to their biosynthetic gene cluster is a determining step in elucidating the biosynthetic pathway and allows downstream process optimization and upscaling. The review focuses on approaches built around NMR-based metabolomics, which enables efficient dereplication and guided fractionation of (antimicrobial) compounds.     

Metabolite induction via microorganism co-culture: A potential way to enhance chemical diversity for drug discovery

Microorganisms have a long track record as important sources of novel bioactive natural products, particularly in the field of drug discovery. While microbes have been shown to biosynthesize a wide array of molecules, recent advances in genome sequencing have revealed that such organisms have the potential to yield even more structurally diverse secondary metabolites. Thus, many microbial gene clusters may be silent under standard laboratory growth conditions. In the last ten years, several methods have been developed to aid in the activation of these cryptic biosynthetic pathways. In addition to the techniques that demand prior knowledge of the genome sequences of the studied microorganisms, several genome sequence-independent tools have been developed. One of these approaches is microorganism co-culture, involving the cultivation of two or more microorganisms in the same confined environment. Microorganism co-culture is inspired by the natural microbe communities that are omnipresent in nature. Within these communities, microbes interact through signaling or defense molecules. Such compounds, produced dynamically, are of potential interest as new leads for drug discovery. Microorganism co-culture can be achieved in either solid or liquid media and has recently been used increasingly extensively to study natural interactions and discover new bioactive metabolites. Because of the complexity of microbial extracts, advanced analytical methods (e.g., mass spectrometry methods and metabolomics) are key for the successful detection and identification of co-culture-induced metabolites.     

A fungal scent from the cheese

Assembly of microbial communities is shaped by various physical and chemical factors deriving from their environment, including other microbes inhabiting the certain niche. In addition to direct cell–cell contacts, primary and secondary metabolites impact the growth of microbial community members. Metabolites might act as growth-promoting (e.g., cross-feeding), growth-inhibiting (e.g., antimicrobials) or signalling molecules. In multi-species microbial assemblies, secreted metabolites might influence specific members of the community, altering species abundances and therefore the functioning of these microcosms. In the current issue, Cosetta and colleagues describe a unique volatile metabolite-mediated cross-kingdom interaction that shapes the cheese rind community assembly. The study paves the way of our understanding how fungus-produced volatile compounds promote the growth of a certain bacterial genus, a principal connection between community members of the cheese rind.     

Biologically active metabolites of marine actinobacteria

Data on the chemical structures and biologic activities of metabolites of obligate and facultative marine actinobacteria published between 2000 and 2007 are reviewed. The structural features of five groups of metabolites related to macrolides and compounds containing lactone, quinone, and diketopiperazine residues; cyclic peptides; alkaloids; and compounds of combined nature are discussed. The review shows the large chemical diversity of metabolites of actinobacteria isolated from marine ecotopes. In addition to metabolites identical to those previously isolated from terrestrial actinobacteria, marine actinobacteria produce compounds not found in other natural sources, including microorganisms. Probably, the biosynthesis of new chemotypes of bioactive compounds by marine actinobacteria is related to the chemical adaptation of microorganisms to the marine environment. The review emphasizes the importance of chemical studies of metabolites produced by marine actinobacteria. These studies will provide new data on marine microbial producers of biologically active compounds and the chemical structures and biologic activities of new natural lowmolecular-weight bioregulators.     

Microbial biotransformation: a tool for drug designing

For centuries microbial biotransformation has proved to be an imperative tool in alleviating the production of various chemicals used in food, pharmaceutical, agrochemical and other industries. In the field of pharmaceutical research and development, biotransformation studies have been extensively applied to investigate the metabolism of compounds (leads, lead candidates, etc.) using animal models. The microbial biotransformation phenomenon is then commonly employed in comparing metabolic pathways of drugs and scaling up the metabolites of interest discovered in these animal models for further pharmacological and toxicological evaluation. Microorganisms can conveniently afford drugs difficult obtained via synthesis. The plethora of reported microbial biotransformations along with its added benefits has already invoked further research in bioconversion of novel and structurally complex drugs. This review alternatively discusses the prospect of microbial biotransformation studies as a significant element ameliorating drug discovery and design in terms of cost-effectiveness, environment protection and greater structural diversity as compared to animal models used to study metabolism. To explicate the microbial biotransformation paradigm in drug designing 3 main areas in this aspect have been analyzed: 1—lead expansion: obtaining pharmacologically improved metabolites from bioactive molecules; 2—biosynthesis of precursors/intermediates involved in the production of bioactive molecules; 3—resolution of racemic mixture to obtain enantiomers possessing different pharmacological profiles.     

基于微生物共培养的隐性基因簇激活策略

微生物次级代谢产物是药物先导化合物的重要源泉之一。随着测序技术的迅猛发展,越来越多的微生物基因组得以测序完成。伴随着测序技术的进步,生物信息学也得到了快速发展。基因组序列分析发现,链霉菌和丝状真菌等微生物中存在大量的已知的或未知的次级代谢物生物合成基因簇(secondary metabolite-biosynthetic gene clusters,SM-BGCs)。然而,在实验室培养条件下大部分基因簇无法表达或表达量很低,导致难以发现这些基因簇所对应的代谢产物,人们将这类基因簇称为“隐性基因簇”或“沉默基因簇”。通过调节基因簇中特异调控基因或基因簇外全局性调控基因的表达,对代谢途径的定向改造,以及将基因簇导入异源宿主等策略,能够激活部分隐性基因簇的表达。通过激活隐性基因簇的表达,能够发现通过常规实验室培养无法获得的具有独特生物活性的新结构代谢产物,成为创新药物的重要来源之一。然而,这些基因簇激活策略都严重依赖于对特定菌株或宿主的遗传操作。近年来,通过模拟自然混合培养中微生物间相互作用,开发了通过混合特定微生物菌株在厌氧或好氧条件下激活隐性基因簇的方法,称之为共培养激活策略。这种策略不...     

Are antibiotics naturally antibiotics?

Antibiotics have been used for more than 50 years and are the cornerstone of infectious disease treatment; in addition, these low-molecular-weight bioactive compounds have been applied to many other therapeutic purposes. However, there is almost no information on the evolutionary biology or ecology of naturally occurring low-molecular-weight compounds. The large number of different structural types and the extremely broad range of biological activities of organic molecules produced by microbes raise many questions concerning their roles in nature. Recent evidence for the enormous complexity of microbial populations in the environment favors the notion that the principal roles of small molecules in microbial ecology are cell–cell communication and not antibiosis.     

成像质谱在微生物代谢产物研究中的应用

微生物代谢产物的结构和功能多样,对相邻微生物和环境会产生重要影响。传统的天然产物分离方法不能系统全面地监测单一或混合微生物样品中代谢物的合成和释放模式。成像质谱能够同时可视化观察从单一微生物菌落到复杂微生物群落的多个代谢产物的时空分布,可以用于发现重要的生物活性分子,观察微生物菌落的代谢交流,以及跟踪微生物之间相互竞争过程中代谢物的修饰等方面的研究。本文综述了成像质谱在微生物代谢产物研究中的最新进展,展望了该技术的应用前景。     

The environment,microbes and bioremediation: microbial activities modulated by the environment

Microorganisms in nature are largely responsible for the biodegradation and removal of toxic and non-toxic chemicals. Many organisms are also known to have specific ecological niches for proliferation and colonization. The nature of the environment dictates to a large extent the biodegradability of synthetic compounds by modulating the evolutionary processes in microorganisms for new degradative genes. Similarly, environmental factors often determine the extent of microbial gene expression by activating or repressing specific gene or sets of genes through a sensory signal transduction process. Understanding how the environment modulates microbial activity is critical for successful bioremediative applications.     

Secondary metabolites as endogenous effectors of microbial cytodifferentiation

The present survey covers the regulatory role of microbial secondary metabolites and related compounds as endogenous signals of cell differentiation of the producing organisms. Several antibiotics have been shown to exert autoregulatory effects on differentiation-associated functions. The mechanisms of self-protection of the producing cells against the autotoxicity of secondary metabolites are discussed in terms of an integral part of the modulation of signal strength. As a further topic, the review deals with the hormone-like interference of particular metabolites with differentiating cells. Conclusive discussion concerns the potential use of microbial signal molecules either as tools for directed manipulations of product syntheses or as pharmaceutics.     

Secondary metabolites of soil <Emphasis Type="Italic">Bacillus</Emphasis> spp.

Bacillus species produce secondary metabolites that are the object of natural product chemistry studies. The wide structural variability of these compounds has attracted the curiosity of chemists and their biological activities have inspired the pharmaceutical industry to search for lead structures in microbial extracts. Screening of microbial extracts reveals the large structural diversity of natural compounds with broad biological activities, such as antimicrobial, antiviral, immunosuppressive, and antitumor activities, that enable the bacterium to survive in its natural environment. These findings widen the potential industrial importance of Bacillus spp., particularly of B. thuringiensis, beyond insecticidal usage and may help explain the role of Bacillus spp. in the soil ecosystem.     

Functional characterization and taxonomic classification of novel gammaproteobacterial diversity in sponges

Sponges harbour exceptionally diverse microbial communities, whose members are largely uncultured. The class Gammaproteobacteria often dominates the microbial communities of various sponge species, but most of its diversity remains functional and taxonomically uncharacterised. Here we reconstructed and characterised 32 metagenome-assembled genomes (MAGs) derived from three sponge species. These MAGs represent ten novel species and belong to seven orders, of which one is new. We propose nomenclature for all these taxa. These new species comprise sponge-specific bacteria with varying levels of host specificity. Functional gene profiling highlights significant differences in metabolic capabilities across the ten species, though each also often exhibited a large degree of metabolic diversity involving various nitrogen- and sulfur-based compounds. The genomic features of the ten species suggest they have evolved to form symbiotic interaction with their hosts or are well-adapted to survive within the sponge environment. These Gammaproteobacteria are proposed to scavenge substrates from the host environment, including metabolites or cellular components of the sponge. Their diverse metabolic capabilities may allow for efficient cycling of organic matter in the sponge environment, potentially to the benefit of the host and other symbionts.     

Construction of a microbial natural product library for chemical biology studies

The RIKEN Natural Products Depository (NPDepo) is a public depository of small molecules. Currently, the NPDepo chemical library contains 39,200 pure compounds, half of which are natural products and their derivatives. In order to reinforce the uniqueness of our chemical library, we have improved our strategies for the collection of microbial natural products. Firstly, a microbial metabolite fraction library coupled with an MP (microbial products) plot database provides a powerful resource for the efficient isolation of microbial metabolites. Secondly, biosynthetic studies of microbial metabolites have enabled us to not only access ingenious biosynthetic machineries, but also obtain a variety of biosynthetic intermediates. Our chemical library contributes to the discovery of molecular probes for increasing our understanding of complex biological processes and for eventually developing new drug leads.     

An ecological perspective of microbial secondary metabolism

Bacteria and fungi produce a remarkable array of bioactive small molecules. Many of these have found use in medicine as chemotherapies to treat diseases ranging from infection and cancer to hyperlipidemia and autoimmune disorders. The applications may or may not reflect the actual targets for these compounds. Through careful studies of microbes, their associated molecules and their targets, a growing understanding of the ecology of microbial secondary metabolism is emerging that exposes the central role of secondary metabolites in many complex biological systems.     

Microbial technologies for the discovery of novel bioactive metabolites

Soil microbes represent an important source of biologically active compounds. These molecules present original and unexpected structure and are selective inhibitors of their molecular targets. At Biosearch Italia, discovery of new bioactive molecules is mostly carried out through the exploitation of a proprietary strain collection of over 50000 strains, mostly unusual genera of actinomycetes and uncommon filamentous fungi. A critical element in a drug discovery based on microbial extracts is the isolation of unexploited groups of microorganisms that are at the same time good producers of secondary metabolites. Molecular genetics can assist in these efforts. We will review the development and application of molecular methods for the detection of uncommon genera of actinomycetes in soil DNA and for the rapid dereplication of actinomycete isolates. The results indicate a substantial presence in many soils of the uncommon genera and a large diversity of isolated actinomycetes. However, while uncommon actinomycete strains may provide an increased chance of yielding novel structures, their genetics and physiology are poorly understood. To speed up their manipulation, we have developed vectors capable of stably maintaining large segments of actinomycete DNA in Escherichia coli and of integrating site specifically in the Streptomyces genome. These vectors are suitable for the reconstruction of gene clusters from smaller segment of cloned DNA, the preparation of large-insert libraries from unusual actinomycete strains and the construction of environmental libraries.     

微生物共培养在新活性化合物挖掘中的研究进展

微生物次级代谢产物结构新颖、活性多样,一直以来都是药物及其先导化合物的重要来源。但近年来在普通微生物纯培养中发现的新活性物质不断减少,重复发现逐渐增加;而微生物共培养因更加接近微生物生长的自然环境,在药物及其先导物挖掘方面发挥着越来越重要的作用。本文分析了近5年来利用真菌vs真菌、真菌vs细菌共培养挖掘新活性化合物的研究现状,以及研究中存在的问题。     

海洋动物来源微生物的共培养研究进展

近年来,由于一些新疾病的发生和细菌耐药性的出现,微生物来源次级代谢产物的筛选重复率越来越高,微生物一些代谢基因在现有实验室条件下无法表达,所以需要发现新的微生物资源,同时找到激活微生物代谢产物基因的方法.海洋动物体内蕴含着大量的共附生微生物资源,可以产生很多具有生物活性的化合物,是潜在的药用资源.本文综述了近年来海洋动...     

Metagenomic approaches to exploit the biotechnological potential of the microbial consortia of marine sponges

Natural products isolated from sponges are an important source of new biologically active compounds. However, the development of these compounds into drugs has been held back by the difficulties in achieving a sustainable supply of these often-complex molecules for pre-clinical and clinical development. Increasing evidence implicates microbial symbionts as the source of many of these biologically active compounds, but the vast majority of the sponge microbial community remain uncultured. Metagenomics offers a biotechnological solution to this supply problem. Metagenomes of sponge microbial communities have been shown to contain genes and gene clusters typical for the biosynthesis of biologically active natural products. Heterologous expression approaches have also led to the isolation of secondary metabolism gene clusters from uncultured microbial symbionts of marine invertebrates and from soil metagenomic libraries. Combining a metagenomic approach with heterologous expression holds much promise for the sustainable exploitation of the chemical diversity present in the sponge microbial community.     

Changes in phytotoxicity of polycyclic aromatic hydrocarbons in the course of microbial degradation

Phytotoxicity of six polycyclic aromatic hydrocarbons (PAHs) and their 16 oxidized derivatives that may be microbial metabolites arising in the course of PAH degradation was determined using an express test with the seedlings of sorghum (Sorghum bicolor L. Moench) and alfalfa (Medicago sativa L.). It was shown that germinating capacity is the least informative characteristic and the most useful parameter is development of seedlings during 3 days in the presence of compounds under investigation. Among unsubstituted compounds, toxicity in respect to seedlings decreased in the series fluorene > phenanthrene > anthracene. Chrysene, fluoranthene, and pyrene stimulated shoot development. It was found that some of the metabolites produced as a result of microbial degradation of phenanthrene (9,10-phenanthrenequinone, 1-hydroxy-2-naphthoic and benzoic acids) are more toxic for plants than starting PAH molecules. The obtained results are important for understanding rhizosphere processes associated with phytoremediation technique.