Isolation, identification and characterization of potentially novel antibiotics from rhizospheric bacteria without detectable in vitro resistance

从根际细菌中分离、鉴定和表征潜在的新型抗生素，且体外未检测到耐药性

• 批准号: 10581945
• 负责人: ANDRE O HUDSON
• 金额: $ 10万
• 依托单位: ROCHESTER INSTITUTE OF TECHNOLOGY
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-09-15 至 2024-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10581945
• 关键词: Accounting; Acinetobacter; Anti-Bacterial Agents; Antibiotic Resistance; Antibiotics; Bacteria; Bacterial Antibiotic Resistance; Biological Assay; Cessation of life; Chemicals; Clinical; Crude Extracts; Data; Databases; Development; Disease Resistance; Drug resistance; Epidemic; Escherichia coli; Future; Gene Cluster; Goals; In Vitro; Individual; Infection; Intermediate resistance; Lead; Mass Spectrum Analysis; Morbidity - disease rate; Multi-Drug Resistance; Multiple Bacterial Drug Resistance; Multiple drug resistant Mycobacteria Tuberculosis; Pathway interactions; Persons; Poverty; Production; Property; Proteins; Pseudomonas aeruginosa; Reporting; Research; Resistance; Sampling; Solid; Staphylococcus aureus; Students; Time; Vancomycin; World Health Organization; base; clinically relevant; combat; experimental study; gene product; genome annotation; genome sequencing; human mortality; methicillin resistant Staphylococcus aureus; microorganism interaction; multi-drug resistant pathogen; novel; pathogenic bacteria; public health relevance; resistant strain; response; whole genome

ABSTRACT Globally, approximately 2.8M people are infected with bacteria that are deemed resistant to clinically relevant antibiotics an annual basis. Of these infections, 700,000 individuals will die with the United Sates accounting for 5% (35,000) of these deaths. In the 2019 report, The World Health Organization states that if no action is take, drug resistant diseases could cause 10 million deaths each year by 2050 and that by 2030, antibiotic resistance could force up to 24 million people into extreme poverty. There is an urgent need for the development of novel antibiotics to combat the drastic rise in the number of antibiotic resistant bacteria. In particular, there are few molecules with broad spectrum activity against multidrug resistant pathogens. One of the bottlenecks that is impeding the further development of novel antibiotics is the lack of identified gene clusters producing them, which could lead to optimized heterologous production. The overarching goal of this proposal is to identify and characterize the biosynthetic gene clusters and products from these anabolic pathways that responsible for the production of antibiotics, which in crude extracts were shown to be active in killing multi-drug resistant pathogens. The proposed research is significant since antibiotic resistance to clinically relevant antibiotics is increasing across different types of pathogenic bacteria, and particularly, broad-spectrum options to treat multidrug resistant strains are currently limited. The PI and his students recently isolated and identified several strains of rhizospheric bacteria, which possess broad-spectrum antibiotic activity. These initial experiments were performed using a culture-based approach, which screened for strains competing against each other on solid media, leveraging the chemical ecological concept of competition between bacterial species from the same environmental sample. One of the strains, Exiguobacterium sp RIT 594 produces a cocktail of antibiotic compounds (in response to another isolate Acinetobacter sp RIT 592). The antibiotic activity produce a >6 log reductions in three relevant resistant clinical isolates: (1) the epidemic strain MRSA USA300 (FPR3757), (2) NDM-producing E. coli (MCR1_NJ) and (3) VIM-producing P. aeruginosa (AR-0266) in 24 h time kill assays. Importantly, the preliminary data show that the emergence of in vitro resistance was not detected in the time kill assays. Subsequent analysis of crude extracts using LC-MS provided evidence and identified more than 200 novel compounds that are unknown in the mass spectrometry databases. Whole genome sequencing and annotation, revealed 8 putative gene clusters that contained a large number of uncharacterized proteins that are predicated to produce a variety of secondary metabolites that are putatively endowed with antibiotic properties.

抽象的 在全球范围内，大约280万人感染了被认为抗临床相关的细菌 抗生素年度。在这些感染中，有70万人将死于联合国会计 这些死亡中有5％（35,000）。在2019年的报告中，世界卫生组织指出，如果没有任何行动 服用，耐药性疾病可能每年到2050年每年造成1000万次死亡，到2030年，抗生素 抵抗可能会迫使多达2400万人陷入极端贫困。迫切需要 新型抗生素的发展，以打击抗生素耐药细菌数量的急剧上升。在 特别是，很少有针对多药耐药病原体具有广谱活性的分子。之一 阻碍新抗生素进一步发展的瓶颈缺乏鉴定的基因 产生它们的簇，可能导致优化的异源产生。总体目标 建议是识别和表征这些合成代谢的生物合成基因簇和产物 负责产生抗生素的途径，在粗提取物中被证明是活跃的 杀死多药耐药病原体。拟议的研究很重要，因为抗生素耐药性 临床相关的抗生素正在增加不同类型的致病细菌，尤其是 目前，广泛的治疗多药抗性菌株的宽光谱选项受到限制。 PI和他的学生 最近分离并鉴定了几种具有广谱的根际细菌菌株 抗生素活性。这些最初的实验是使用基于培养的方法进行的，该方法筛选了 对于在固体媒体上相互竞争的菌株，利用化学生态概念 来自同一环境样本的细菌物种之间的竞争。其中之一， Exigubacterium sp RIT 594产生抗生素化合物的鸡尾酒（响应另一种分离株） ACINETOBACTER SP RIT 592）。抗生素活性在三种相关抗性中产生> 6 log降低 临床分离株：（1）流行菌株MRSA USA300（FPR3757），（2）ndm产生大肠杆菌（MCR1_NJ） （3）在24小时杀死测定中产生VIM生产的铜绿假单胞菌（AR-0266）。重要的是，初步数据 表明在时间杀伤测定中未检测到体外抗性的出现。随后的分析 使用LC-MS的原油提取物提供了证据，并确定了200多种新型化合物 在质谱数据库中未知。整个基因组测序和注释，揭示了8个假定 基因簇包含大量未表征的蛋白质，这些蛋白质可产生A 多种二级代谢产物，这些代谢物具有抗生素特性。