Bacteriophage to treat multidrug‐resistant UTI in Persons with Spinal Cord Injury

我们的总体目的是针对脊髓损伤(SCI)患者面临的两个紧迫的临床问题。这个 第一个问题是尿路感染(UTI)的持续发病率以及菌血症和败血症的死亡率。 由尿路中的病原体引起的。尿路感染和相关并发症是常见的 对脊髓损伤(SCI)患者的健康甚至生存构成威胁。第二个临床困境 脊髓损伤患者正经历着来自多重耐药细菌的不成比例的伤害，特别是 大肠埃希氏菌是最常见的泌尿系病原体。我们计划开发噬菌体，这种病毒可以杀死特定的 细菌菌株，作为治疗SCI中耐多药大肠杆菌UTI和脓毒症的药物。噬菌体有很多 表明它们将在这一角色中发挥作用的特性，包括对抗生素没有交叉耐药性，能力 攻击生物膜，体外进化，并在感染部位作为自我更新剂。噬菌体疗法 耐药尿路细菌在脊髓损伤人群中有潜在的应用，超越了UTI治疗，也适用于 退伍军人一般，如用于泌尿外科手术前的膀胱绝育。噬菌体疗法 与VHA抗生素管理工作保持一致，并可能减轻艰难梭菌经历的负担 退伍军人。我们在这里提出的关于脊髓损伤最常见的尿路病原体--大肠杆菌--的工作将 随后将其应用于其他SCI尿路病原体，制成广谱有效的噬菌体鸡尾酒。项目 目的：我们将开发针对大肠杆菌的噬菌体，这些噬菌体对多种脊髓损伤特异的E。 在导管相关性尿路感染(CAUTI)和/或 败血症。目的1确定噬菌体在小鼠感染模型中对耐药大肠杆菌的杀灭效果 CAUTI(1A)、菌血症(1B)，并与感染大肠杆菌病原体所对的抗生素组合 抗性(1C)。目标2是开发和测试一组有效对抗当代大肠杆菌分离株的噬菌体。 来自患有脊髓损伤的退伍军人。在目标2A中，我们将积极收集患有脊髓损伤的退伍军人的大肠杆菌临床分离株，创造 一个SCI特有的细菌库。在目标2B中，我们将收集并实验产生具有广泛的 宿主范围对抗大肠杆菌，创建了一个测序噬菌体库，覆盖了80%以上的SCI大肠杆菌菌株。在……里面 目标2C我们将从我们的噬菌体库中创造出噬菌体鸡尾酒，并在小鼠身上测试鸡尾酒的效果 大鼠脊髓损伤致急性尿路感染模型的建立。目标3是检查抗噬菌体大肠埃希菌在 CAUTI的治疗和确定如何克服噬菌体耐药性。目标3A：确定大肠杆菌的持久性 噬菌体治疗后细菌对噬菌体产生抗药性。目标3BB：确定是否需要补充剂量的 在治疗后的几天内，噬菌体可以防止大肠杆菌的复发。方法：我们建议研究噬菌体。 在体外和体内，使用我们建立的小鼠CAUTI和菌血症模型来实现这三个目标。在这些 我们将跟踪微生物结果和临床结果的实验。微生物的结果包括 细菌和噬菌体的定量计数，[噬菌体和/或处理前后的微生物群多样性 抗生素和尿管相关生物膜的图像。]我们的临床结果旨在 总结一下人类的经历，特别是日常生活活动的能力。我们有两个 临床结果的测量：疾病严重程度指数(评估健康)和行为评分(评估 动物是否仍在进行例行活动)。目标1中的实验将使用细菌(E.Coli JJ2528)和 从我们现有的收藏中提取噬菌体(HP3)，这样工作就可以立即开始。在这份提案的最后， 我们将有一个专门针对脊髓损伤的细菌库，我们还将开发、表征和测试 体内有一个噬菌体文库，可以有效地裂解80%以上的SCI分离株。我们的团队聚集在一起 在临床感染、流行病学、脊髓损伤临床管理、脊髓损伤研究、 完成这项工作的微生物学、病毒学、[元基因组学、生物膜成像]和动物研究。

Our overall purpose is to take aim at two urgent clinical problems for persons with spinal cord injury (SCI). The first problem is ongoing morbidity from urinary tract infections (UTI) and mortality from bacteremia and sepsis arising from pathogens in the urinary tract. Urinary tract infections and associated complications are a common threat to the wellbeing and even survival of persons with spinal cord injury (SCI). The second clinical dilemma is the disproportionate harm persons with SCI are experiencing from multidrug -resistant bacteria, particularly Escherichia coli, the most common urinary pathogen. We plan to develop bacteriophage, viruses that kill specific strains of bacteria, as a treatment for multidrug-resistant E. coli UTI and sepsis in SCI. Phage have numerous properties that suggest they will be effective in this role, including lack of cross resistance with antibiotics, ability to attack biofilms, evolvability in vitro, and being a self-renewing agent at the site of infection. Phage therapy for resistant urinary organisms has potential applications in the SCI population beyond UTI treatment and also for the Veteran population in general, such as for bladder sterilization prior to urologic procedures. Phage therapy aligns with VHA antibiotic stewardship efforts and may alleviate the burden of Clostridium difficile experienced by Veterans. The work we propose here with the most common urinary pathogen in SCI, E. coli, would subsequently be applied to the other SCI urinary pathogens to create broadly effective phage cocktails. Project Objectives: We will develop E. coli-specific phages that are efficacious against a broad range of SCI-specific E. coli isolates from our VA hospital in a mouse model of catheter-associated urinary tract infection (CAUTI) and/or sepsis. Aim 1 is to determine the efficacy of phage in killing antibiotic-resistant E. coli in murine infection models of CAUTI (1A), bacteremia (1B), and in combination with antibiotics to which the infecting E. coli pathogen is resistant (1C). Aim 2 is to develop and test a bank of phages effective against contemporary E. coli isolates from Veterans with SCI. In Aim 2A, we will actively collect E. coli clinical isolates from Veterans with SCI, creating an SCI-specific bacterial strain bank. In Aim 2B, we will collect and experimentally generate phages with broad host range against E. coli, creating a bank of sequenced phage that cover over 80% of the SCI E. coli strains. In Aim 2C we will create phage cocktails from our phage bank, and test the efficacy of the cocktails in the mouse model of CAUTI induced by SCI E. coli. Aim 3 is to examine the emergence of phage-resistant E. coli during treatment of CAUTI and determine how to overcome phage resistance. Aim 3A: Determine if E. coli persistence after phage treatment results from bacterial resistance to phage. Aim 3Bb: Determine if a “recharge” dose of phage several days into therapy prevents recurrence of E. coli. Methods: We propose to study bacteriophage in vitro and in vivo using our established models of mouse CAUTI and bacteremia for all three aims. In these experiments we will track microbiological outcomes and clinical outcomes. The microbiological outcomes include quantitative counts of bacteria and phage, [microbiome diversity before and after treatment with phage and/or antibiotics, and images of urinary catheter-associated biofilms.] Our clinical outcomes are intended to recapitulate the human experience, particularly the ability to perform activities of daily living. We have two measures of clinical outcomes: the disease severity index (assesses health) and the behavioral score (assesses whether animals still perform routine activities). The experiments in Aim 1 will use bacteria (E. coli JJ2528) and phage (HP3) from our existing collections, so that the work can begin immediately. At the end of this proposal, we will have a bacterial strain bank specific to SCI, and we also will have developed, characterized, and tested in vivo a library of phage that are effective at lysing over 80% of these SCI isolates. Our team brings together complementary expertise in clinical infections, epidemiology, SCI clinical management, SCI research, microbiology, virology, [metagenomics, biofilm imaging,] and animal studies to accomplish the work proposed.