Exploration of new approaches to microbial typing and phenotypic characterization based on Fourier transform infrared and high-resolution magic-angle spinning NMR spectroscopy and the development of active packaging employing natural antimicrobials

拟议的Discovery Grant研究的第一个方面将针对基于傅里叶变换红外(FTIR)光谱、高分辨率魔角旋转(HRMAS)核磁共振和质谱学的差异来探索密切相关细菌之间的区别性质，重点是它们在微生物分型、疫情检测和生物标记物发现方面的应用。在我们的工作中，正在利用这些能力来开发快速鉴定耐抗生素细菌的方法，如耐甲氧西林金黄色葡萄球菌(MRSA)和万古霉素耐药肠球菌(VRE)，这是医疗保健相关感染的最常见原因之一，并鉴定新出现的多重耐药临床酵母菌，如金黄色念珠菌。这项工作是通过与加拿大卫生部、魁北克临床监测机构和魁北克多家医院合作进行的。关于食源性疾病，目前正在与加拿大卫生部和加拿大食品检验局(CFIA)的研究人员合作，通过全基因组测序和FTIR光谱分析，研究致病性大肠杆菌致病类型(与食源性疾病有关)的遗传变异性对其毒力的影响。我们的工作首先需要从微生物中获取大量的光谱数据集，以创建包含大量FTIR、1H和31P HRMAS核磁共振和特征良好的菌株的质谱图(MS)的光谱数据库，并通过结合多变量统计分析技术应用特征选择算法来提取识别光谱信息。拟议研究的一个主要目标是开发新的策略，旨在阐明特定细菌菌株之间光谱差异背后的特定生化差异。预计拟议的研究结果将不仅在我们的微生物鉴定和分类工作中推进生物标记物的发现，而且还将在快速发展的微生物分型领域推动生物标记物的发现，并可能有助于开发新的抗生素，以治疗因医院暴发而导致的抗生素医院感染水平的上升。拟议研究的第二个方面将侧重于探索将选定的多糖组合作为可生物降解薄膜的组成部分的蛋白质，并在薄膜中嵌入天然抗菌剂，以减缓新鲜和最低加工食品的微生物生长。建议的研究重点是应用红外成像显微镜来研究天然抗菌剂在生物可降解薄膜中的分布、迁移速率，以增强其在活性包装中的潜在用途。*****

The first facet of the proposed Discovery Grant research will be directed toward the exploration of the discriminatory nature between closely related bacterial strains based on differences in their Fourier transform infrared (FTIR) spectra, high-resolution magic angle spinning (HRMAS) NMR and mass spectrometry with a focus on their application to microbial typing, outbreak detection and biomarker discovery. In our work, these capabilities are being exploited to develop rapid methods for identification of antibiotic-resistant bacteria such as methicillin-resistant Staphylococcus aureus (MRSA) and vancomycin-resistant enterococci (VRE), which are among the most prevalent causes of healthcare-associated infections, and identification of emerging multi-drug resistant clinical yeast isolates such as Candida auris. This work is being conducted through collaborative studies with Health Canada, the Quebec clinical surveillance agency and multiple hospitals across Quebec. With regards to foodborne illness, ongoing efforts, in collaboration with researchers at Health Canada and the Canadian Food Inspection Agency (CFIA), is aimed at studying the effect of genetic variability of pathogenic E. coli pathotypes (associated with foodborne illness) to its virulence by whole genome sequencing and FTIR spectroscopy. Our work initially entails acquisition of a large spectral datasets from of microorganisms for the creation of spectral databases containing large numbers of FTIR, 1H and 31P HRMAS NMR and mass spectrometry (MS) spectra of well-characterized strains and extracting the discriminatory spectral information by applying feature-selection algorithms in conjunction with multivariate statistical analysis techniques. A major objective of the proposed research is to develop new strategies directed toward elucidation of the specific biochemical differences underlying the spectral differences among particular bacterial strains. It is anticipated that the results of the proposed research will advance biomarker discovery not only in our work on microbial identification and classification but also in the rapidly evolving field of microbial typing and may aid in the development of new antibiotics that can treat the rising levels of antibiotic nosocomial infections resulting in hospital outbreaks. The second facet of the proposed research will focus on the exploration of proteins in combination of selected polysaccharides as components of biodegradable films and embedding the films with natural antimicrobials to retard microbial growth of fresh and minimally processed foods. The proposed research focuses on the application of infrared imaging microscopy to study the distribution, migration rates of natural antimicrobials in biodegradable films in relation to enhancing their potential use in active packaging. *****