NeoChip for specific and rapid identification of congenital CMV and neonatal HSV infections on minimal sample volume

NeoChip 用于以最少的样本量特异性快速识别先天性 CMV 和新生儿 HSV 感染

• 批准号: 10539056
• 负责人: Shelley M Lawrence
• 金额: $ 56.74万
• 依托单位: UNIVERSITY OF UTAH
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-01-01 至 2025-06-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10539056
• 关键词: Address; Advocate; Antibiotics; Antibodies; Antimicrobial Resistance; Antiviral Agents; Antiviral resistance; Base Sequence; Biological; Biological Assay; Birth; Blinded; Blood; Blood Volume; Blood specimen; California; Child; Clinical; Clinical Research; Congenital herpes simplex; Consumption; Culture Techniques; Cytomegalovirus; DNA; Data; Databases; Detection; Diagnosis; Diagnostic; Disabled Persons; Disease; Dyes; Early Intervention; Early identification; Enrollment; Epidemiology; Etiology; Evaluation; Failure; Fingerprint; Fluorescence; Funding; Genome; Genotype; Goals; Gold; Health; Hour; Immunoglobulin G; Immunoglobulin M; Individual; Infant; Infection; Investigation; Laboratories; Life; Machine Learning; Measures; Microbe; Modality; Modernization; Molecular; Mothers; Neonatal; Neonatal Screening; Newborn Infant; Nucleic Acid Amplification Tests; Nucleotides; Outcome; Pathogen detection; Pathogenicity; Patients; Perinatal; Polymerase Chain Reaction; Predictive Value; Pregnant Women; Prevalence; RNA; Reaction; Reporting; Research; Research Personnel; Resistance; Resolution; Risk; Saliva; Sampling; Sensitivity and Specificity; Simplexvirus; Spottings; Symptoms; Techniques; Technology; Testing; Therapeutic; Therapeutic Intervention; Time; TimeLine; Tissues; Training; Translating; Tube; United States; Universities; Update; Urine; Variant; Viral; Viral Load result; Virus Diseases; accurate diagnosis; antimicrobial; artificial intelligence algorithm; base; biobank; clinical database; clinical outcome measures; clinically relevant; congenital cytomegalovirus; cost; detection limit; detection method; diagnosis standard; diagnostic biomarker; diagnostic tool; digital; disease prognosis; interdisciplinary approach; intervention program; machine learning algorithm; melting; microbial; multidisciplinary; neonatal infection; neonate; next generation sequencing; offspring; pathogen; pathogen genomics; pathogenic virus; point of care; postnatal; prevent; prospective; rapid diagnosis; resistance gene; screening program; single molecule; transmission process; viral detection

Project Summary Congenital cytomegalovirus (cCMV) and neonatal herpes simplex virus (nHSV) impose major health threats on neonates. Although CMV and HSV are lifelong infections with periods of latency and reactivation, most maternal infections remain undiagnosed due to nonspecific or absent clinical symptoms. In the United States, a child is permanently disabled by cCMV infection every hour, even though 9 of 10 infants are asymptomatic after birth and remain undiagnosed. Invasive nHSV, on the other hand, is a rare neonatal infection that presents with a broad range of clinical symptoms, including those that may be life-threatening. Viral culture and DNA detection by polymerase chain reaction (PCR) have become the “Gold Standard” for the diagnosis of cCMV and nHSV infection, despite poor sensitivity of PCR assays in neonates and time consuming culture techniques (up to 5-7 days). Universal genotyping of pathogen genomic sequences using High Resolution Melt (U-HRM) provides a simple, low cost, rapid, and modern alternative to viral cultures and PCR techniques. By measuring the fluorescence of an intercalating dye as PCR-amplified pathogen DNA or RNA fragments are heated and disassociate, sequence defined melt curves, or “fingerprints”, are generated with single-nucleotide resolution in a closed-tube reaction. These unique microbial “fingerprints” are then automatically identified and quantified using machine learning technology, with an accuracy of 99-100% on minimal blood volume (1 mL), in a platform called NeoChip. Presently, we have established unique signature melt curves for 40 bacterial species and antimicrobial resistance genes that commonly infect neonates. Additionally, NeoChip has been expanded to distinguish individually amplified melt curve signatures for multiple pathogen identification and quantification, as required for polymicrobial infection. In this proposal, we will build out NeoChip’s comprehensive database by incorporating clinical strains of CMV and HSV with actionable antiviral resistance genes. Because NeoChip identifies variances in nucleic acid sequences, individual differentiation and quantification of CMV and HSV strains are possible. We will also translate the NeoChip for specific and rapid diagnosis of cCMV and nHSV infection in a large prospective clinical study of pregnant women and their offspring(s), as well as directly compare the platform to standard quantitative nucleic acid test (QNAT) assays, IgG/IgM antibody testing, and clinical outcome measures for statistical concordance (predictive value). Finally, we will validate and translate NeoChip for cCMV detection and clinical correlation using dried blood spot (DBS) samples for incorporation into standard universal newborn screening programs. NeoChip’s goal is to provide an accurate and valid test for the timely diagnosis of pathogen etiology (viral, bacterial, and fungal) in a single test with efficacy on broad tissue matrices and capacity to inform microbial resistance, thereby facilitating early administration of targeted antimicrobials and therapeutics. This proposal directly addresses the funding call by applying a multidisciplinary approach to address the biomedical challenges of rapidly and accurately diagnosing cCMV and nHSV to facilitate disease prognostication and early therapeutic or intervention programs.

项目概要 先天性巨细胞病毒 (cCMV) 和新生儿单纯疱疹病毒 (nHSV) 对人类健康构成重大威胁 新生儿。尽管 CMV 和 HSV 是终生感染，具有潜伏期和重新激活期，但大多数孕产妇感染 由于非特异性或缺乏临床症状，感染仍未得到诊断。在美国，一个孩子是 尽管每 10 名婴儿中有 9 名在出生后没有症状，但每小时都会因 cCMV 感染而永久残疾，并且 仍未确诊。另一方面，侵袭性 nHSV 是一种罕见的新生儿感染，其表现广泛 一系列临床症状，包括可能危及生命的症状。病毒培养和 DNA 检测 聚合酶链反应（PCR）已成为诊断cCMV和nHSV感染的“金标准”， 尽管新生儿 PCR 检测的灵敏度较差且培养技术耗时（长达 5-7 天）。 使用高分辨率熔解 (U-HRM) 对病原体基因组序列进行通用基因分型提供了一种简单、 病毒培养和 PCR 技术的低成本、快速、现代替代方案。通过测量荧光 当 PCR 扩增的病原体 DNA 或 RNA 片段被加热并解离时插入染料，测序 定义的熔解曲线或“指纹”是在闭管反应中以单核苷酸分辨率生成的。 然后使用机器学习自动识别和量化这些独特的微生物“指纹” 技术，在名为 NeoChip 的平台上，对最小血量 (1 mL) 的准确度为 99-100%。目前， 我们为 40 种细菌和抗菌素耐药基因建立了独特的特征熔解曲线， 常感染新生儿。此外，NeoChip 已得到扩展，可以区分单独扩增的熔解 根据多种微生物感染的需要，用于多种病原体识别和定量的曲线特征。 在这个提案中，我们将通过纳入CMV的临床毒株来建立NeoChip的综合数据库 和具有可操作的抗病毒抗性基因的HSV。因为 NeoChip 可以识别核酸的变异 CMV 和 HSV 毒株的序列、个体分化和定量是可能的。我们也会翻译 NeoChip 在一项大型前瞻性临床研究中用于特异性、快速诊断 cCMV 和 nHSV 感染 孕妇及其后代，以及直接将平台与标准定量核酸进行比较 酸性测试 (QNAT) 测定、IgG/IgM 抗体测试和临床结果测量以实现统计一致性 （预测值）。最后，我们将使用 NeoChip 验证和翻译用于 cCMV 检测和临床相关性 干血斑 (DBS) 样本，用于纳入标准通用新生儿筛查计划。新芯片的 目标是提供准确有效的检测，以便及时诊断病原体病因（病毒、细菌和细菌） 真菌）在一次测试中对广泛的组织基质具有功效并且能够告知微生物耐药性，从而 促进早期施用靶向抗菌药物和治疗方法。该提案直接针对 通过应用多学科方法来筹集资金，以快速准确地解决生物医学挑战 诊断 cCMV 和 nHSV，以促进疾病预测和早期治疗或干预计划。