Converting staphylococcal pathogenicity islands from malevolence to benevolence

将葡萄球菌致病岛从恶意转变为仁慈

• 批准号: 10536664
• 负责人: Richard P. Novick
• 金额: $ 76.55万
• 依托单位: NEW YORK UNIVERSITY SCHOOL OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-01-21 至 2024-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10536664
• 关键词: Abscess; Address; Adsorption; Animals; Anti-Bacterial Agents; Antibiotic Resistance; Antibiotics; Bacteria; Bacteriophage Typing; Bacteriophages; Beneficence; Binding; CRISPR/Cas technology; Cell Wall; Cessation of life; Chromosomes; Clinical; Clinical Trials; Cloning; Clustered Regularly Interspaced Short Palindromic Repeats; Cytolysis; DNA; Development; Disease; Dose; Drug Metabolic Detoxication; Effectiveness; Engineering; Ensure; Frequencies; Future; Genes; Genetic; Genus staphylococcus; Goals; Implant; In Vitro; Infection; Island; Life; Listeria monocytogenes; Lung; Methods; Mus; Operative Surgical Procedures; Organism; Pathogenicity Island; Pharmaceutical Preparations; Plants; Plasmids; Preparation; Prevention; Probability; Procedures; Process; Production; Promoter Regions; Resistance; Resistance development; Sepsis; Series; Specificity; Staphylococcal Infections; Staphylococcus Phages; Staphylococcus aureus; Subcutaneous abscess; Superantigens; Surface; System; Teichoic Acids; Testing; Therapeutic; Therapeutic Agents; Therapeutic Uses; Time; Toxic Shock Syndrome; Toxin; Variant; Vertebral column; Virulence; Virulence Factors; Work; design; experimental study; genetic element; immunogenicity; in vivo; intraperitoneal; methicillin resistant Staphylococcus aureus; nanoliposome; novel; particle; preservation; prevent; programs; receptor; remediation; restriction enzyme; subcutaneous

Project summary/Abstract Staphylococcus aureus, causes a wide variety of life-threatening infections, many of which cannot be treated effectively, owing to antibiotic resistance. Consequently, there is an urgent need for new ways to treat these infections, which annually cause some 18,000 deaths in the US. We have developed a novel non-antibiotic method, for treating staphylococcal infections based on the highly mobile staphylococcal pathogenicity islands (SaPIs). The SaPIs are ~15 kb genetic elements that are stably inserted in the staph chromosome but can be induced by “helper” phages to excise and replicate. The replicated SaPI DNA is packaged in infectious phage- like particles which are released upon phage-induced lysis. The SaPIs carry and disseminate genes encoding superantigen toxins and other virulence factors. Instead of working on the prevention of SaPI spread, we hit upon the idea of exploiting SaPI spread by converting these agents of disease into agents of therapy – antibacterial drones (ABDs). To create the ABDs, we have re-engineered the SaPIs, deleting their natural cargo (toxin genes), increasing their packaging capacity from 15 to >40 kb, and inserting antibacterial modules. We have also modified the helper phage so that ABD particles are produced in the absence of functional phage. The ABD particles are administered to an infected animal (or plant), where they attach to the infecting bacteria, insert their DNA, express their antibacterial cargo genes and thus abrogate the infection. As proof of principle, we have begun by incorporating into ABDs either CRISPR/cas9 or CRISPR/dcas9 modules with spacers targeting a chromosomal gene or the promoter region of a global virulence regulator, respectively. Preliminary studies have shown that the CRISPR/cas9-containing ABD kills S. aureus in vitro by DNA cleavage, blocks the development of a subcutaneous S. aureus abscess, and rescues mice given a lethal dose of S. aureus intraperitoneally. The CRISPR/dcas9 containing ABD blocks the expression of staphylococcal virulence in vitro and blocks the formation of a subcutaneous abscess in vivo. This proposal outlines our program to develop the ABD system and validate our underlying hypothesis that SaPIs can be converted to versatile and fully effective anti-staphylococcal therapeutic agents. There are 3 specific aims. In Aim I we will construct and test ABDs designed to treat the wide array of infections caused by S. aureus. In Aim II, we will focus on expanding the host range of the ABDs to target diverse S. aureus strains. In Aim III, we will encapsidate the ABD particles for increased efficacy, test for ABD resistance and immunogenicity, and refine our procedures for producing and preserving high titer ABD preparations.

项目摘要/摘要 金黄色葡萄球菌会引起多种威胁生命的感染，其中许多无法治疗 有效地，由于抗生素耐药性。因此，迫切需要新的方法来对待这些 感染，每年在美国造成约18,000人死亡。我们已经开发了一种新颖的非抗生素 方法，用于基于高流动的葡萄球菌致病岛治疗葡萄球菌感染 （SAPI）。 SAPI是〜15 kb的遗传元素，稳定地插入了葡萄球菌中，但可以是 由“助手”噬菌体引起的体验和复制。复制的SAPI DNA包装在感染性噬菌体中 就像在噬菌体引起的裂解时释放的颗粒一样。 SAPI携带并传播编码的基因 超抗原毒素和其他病毒因素。我们没有努力预防SAPI传播，而是击中 通过将这些疾病的药物转化为治疗药物来利用SAPI扩散的想法 - 抗菌无人机（ABDS）。为了创建ABD，我们重新设计了SAPI，删除了它们的自然 货物（毒素基因），将其包装能力从15 kb增加到40 kb，并插入抗菌模块。 我们还修改了辅助噬菌体，以便在没有功能的情况下产生ABD颗粒 噬菌体。 ABD颗粒被施用到感染的动物（或植物）上，并将其附着在感染上 细菌，插入其DNA，表达其抗菌货物基因，从而消除感染。作为证明 原则，我们已经将与ABDS纳入ABDS CRISPR/CAS9或CRISPR/CAS9模块一起开始 分别针对染色体基因或全球病毒调节剂的启动子区域的垫片。 初步研究表明，含CRISPR/CAS9的ABD在体外杀死金黄色葡萄球菌。 乳沟，阻止皮下s。金黄色葡萄球菌的发育，并反应于致死剂量的小鼠 腹膜内金黄色葡萄球菌。包含ABD的CRISPR/DCAS9阻止了葡萄球菌的表达 体外毒力并阻止体内皮下脓肿的形成。该提议概述了我们的 开发ABD系统并验证我们的基本假设的程序，即SAPI可以转换为 多功能且充分有效的抗稳定球菌治疗剂。有3个具体目标。在目标上，我会 构建和测试ABD，旨在处理金黄色葡萄球菌引起的广泛感染。在AIM II中，我们将 专注于将ABD的寄主范围扩展到靶向潜水的金黄色葡萄球菌菌株。在AIM III中，我们将 封装ABD颗粒以提高效率，测试ABD耐药性和免疫原性，并完善 我们生产和保存高滴定ABD准备的程序。

项目成果

Antibacterial particles and predatory bacteria as alternatives to antibacterial chemicals in the era of antibiotic resistance.

• DOI: 10.1016/j.mib.2021.09.016
• 发表时间: 2021-12
• 期刊: Current opinion in microbiology
• 影响因子: 5.4
• 作者: Novick RP

Dynamics of Antibacterial Drone Establishment in Staphylococcus aureus: Unexpected Effects of Antibiotic Resistance Genes.

• DOI: 10.1128/mbio.02083-21
• 发表时间: 2021-12-21
• 期刊: mBio
• 影响因子: 6.4
• 作者: Dhasmana N;Ram G;McAllister KN;Chupalova Y;Lopez P;Ross HF;Novick RP
• 通讯作者: Novick RP

A regulatory cascade controls Staphylococcus aureus pathogenicity island activation.

• DOI: 10.1038/s41564-021-00956-2
• 发表时间: 2021-10
• 期刊: Nature microbiology
• 影响因子: 28.3
• 作者: Haag AF;Podkowik M;Ibarra-Chávez R;Gallego Del Sol F;Ram G;Chen J;Marina A;Novick RP;Penadés JR
• 通讯作者: Penadés JR