美洲大蠊肠道细菌中抗菌肽的发现与成药性评价

Bacterial resistance caused by antibiotic abuse has become a huge threat to human health, which result in the urgent need for new antibiotics. As active ingredients for the original immunity of organisms, antibacterial peptides have become the most promising source of antibiotics because of their broad-spectrum, high-efficiency antibacterial properties and their inability to cause drug resistance. Bacteria and insects are the most important sources of antimicrobial peptides. However, the complex structure of insect antibacterial peptides make it difficult to produce in the drug development, and the exploration of bacterial antimicrobial peptides faces the dilemma of how to find new sources for separation. Considering that the intestine is a special environment with few studies, abundant bacteria and competitive competition, this project creatively combines insects and bacteria, and the Periplaneta americana is chosen as object for its harsh living environment. The technologies including modern molecular biology, chromatography, mass spectrometry and microbiology are used to explore and determine the antimicrobial peptides in its intestinal bacteria under the comprehensive tracking of activity and structure, The discoved lead compounds were synthesized by solid phase peptide synthesis and their antibacterial activity, toxicity were evaluated for further development. The accomplishment of this progect will not only enrich the understanding about intestinal bacteria of Periplaneta americana, but also provide new strategy and lead compounds for the development of antibiotics.

抗生素滥用导致的细菌耐药，已成为威胁人类健康的重要隐患，寻找新型抗生素成为迫在眉睫的问题。抗菌肽作为生物体原始免疫的反应物质，因具有广谱高效抗菌特性，且不易引起耐药性，已成为最有潜力的抗生素来源。细菌和昆虫是抗菌肽最主要的来源，然而昆虫抗菌肽结构复杂后期生产困难，而细菌抗菌肽的发现面临着如何寻找新的分离源的困境。考虑到肠道是一个研究较少，细菌种类丰富，生存竞争激烈的特殊环境。本项目创新性的将昆虫和细菌二者结合，以生活环境恶劣的美洲大蠊为载体。通过活性和结构综合追踪的方式，采用现代分子生物学、色谱、质谱和微生物学等相关技术深入挖掘其肠道细菌中的抗菌肽并确定其结构。通过多肽固相合成技术解决活性先导化合物来源问题的基础上，对其抗菌活性、毒性等成药性参数进行初步评价，为进一步的开发奠定基础。本研究的开展不仅将丰富对美洲大蠊肠道细菌的认识，同时可以为抗生素的开发提供新的思路和先导化合物。

抗生素滥用导致的细菌耐药，已成为威胁人类健康的重要隐患，寻找新型抗生素成为迫在眉睫的问题。特殊生境和激烈生存竞争中的微生物是新型抗生素发掘的重点方向。基于此本项目以生存环境恶劣的美洲大蠊为研究对象，选取具有激烈生存竞争环境的肠道为微生物分离源，通过稀释涂布法和平板划线法共分离获得185株肠道微生物。以6种病原细菌为指示菌，采用交叉条纹法、对峙培养法和牛津杯法筛选出具有抗菌活性的肠道微生物共85株，具体包含细菌28株、真菌23株和放线菌34株。进一步对具有良好抗菌活性的12株放线菌和3株真菌进行了菌种鉴定，从而构建了具有抗菌活性的美洲大蠊肠道微生物菌种资源库。本项目进一步选择抗菌谱较广且对耐甲氧西林的金黄色葡萄球菌具有明显抗菌活性的三株链霉菌进行培养条件优化和大规模发酵，并从其中一株链霉菌的次生代谢产物中分离鉴定了4个化合物，其中新化合物1个。随后采用二倍稀释法，对这些化合物的抗菌活性进行了系统研究。考虑到很多抗生素也具有抗肿瘤活性，本项目还对分离所得放线菌提取物的细胞毒活性进行了筛选，并发现6株具有显著抗肿瘤活性的菌株。本项目的研究结果表明美洲大蠊肠道内不仅含有丰富的微生物种质资源，且这些特境微生物具有广泛的抗菌和抗肿瘤活性代谢产物。本项目的完成不仅丰富了对美洲大蠊肠道细菌的认识，同时为新型抗生素和抗肿瘤药物的开发提供新的思路和先导化合物。