Exploitation of Burkholderia bacteria as novel antibiotic producers using a genome mining approach

利用新的发现工具和产生菌来鉴定抗药抗生素迫切需要发现和开发新的抗生素来治疗由超级细菌引起的耐药感染。我们今天使用的大多数抗生素都是由一种土壤细菌——链霉菌产生的。卡迪夫大学的研究人员发现，另一种被称为伯克霍尔德氏菌的细菌能产生新型抗生素，杀死对耐甲氧西林金黄色葡萄球菌（MRSA）等现有抗生素产生耐药性的超级细菌。为了进行临床和商业开发，必须确定抗生素的化学结构，以确定它们是否是新化合物。通过与华威大学化学系的专家合作，我们已经确定了一种伯克霍尔德菌抗生素的完整结构，并确定了两种新型抗生素，角斗士碱和越南霉素的部分结构。纯化和确定抗生素的化学结构需要时间和大量的专业知识，因此研究人员现在使用一种名为基因组挖掘的新工具来发现抗生素。这种方法获取产生抗生素的细菌的整个遗传密码（基因组序列），然后在其中搜索与产生新型抗生素有关的基因。Wellcome Trust Sanger Institute （WTSI）是细菌基因组测序的先驱，并拥有名为太平洋生物科学测序的新技术，该技术非常适合在单序列运行中快速确定细菌基因组。与WTSI的研究人员合作，我们对产生剑兰碱的菌株的基因组进行了测序，并能够识别出与这种抗生素生产有关的基因。利用这些额外的序列信息，我们现在可以确定角斗士碱的完整结构，这是将抗生素推向临床开发所需要的。总的来说，我们的初步研究表明，伯克氏菌是一种尚未开发的、有希望的新抗生素来源，可以对抗多重耐药的超级细菌。为了确定伯克氏菌是否能像链霉菌一样成功地产生我们今天使用的大多数抗生素，我们将使用基因组挖掘作为一种新的伯克氏菌抗生素发现工具，目的是：1。利用Pacific Biosciences测序技术对24株产抗生素伯克氏菌进行基因组测序。首先，为了进一步研究我们的抗mrsa抗生素越霉素，我们将对产生这种抗生素的伯克霍尔德氏菌进行测序。另外23种能杀死当前耐药细菌的菌株将从我们的大量收集中选出。总的来说，24种产生抗生素的伯克霍尔德菌基因组将为基因组挖掘和新抗生素的发现创造一个独特的DNA文库。通过对其基因组和生产途径的化学和遗传分析，全面鉴定了剑兰素和越南霉素的结构。利用已知具有新型抗药活性的24株菌株的基因组，开展伯克霍尔德菌抗生素能力的独特基因组挖掘调查。上述拨款的前两个目标将创建基因组挖掘数据，并培训资助的研究人员掌握实施这种发现抗生素的新方法所需的所有技能。我们还将研究在这些伯克氏菌基因组中编码新型抗生素的“沉默途径”是否可以打开4。我们研究的最后一个目标是准备一个由五种新型抗生素组成的小组，作为对抗耐药性的药物推进临床前开发。除了剑兰和越南霉素，我们还将利用基因组挖掘的信息完成另外三种新型伯克氏菌抗生素的结构和活性分析。

USING NOVEL DISCOVERY TOOLS AND PRODUCER BACTERIA TO IDENTIFY RESISTANCE-BUSTING ANTIBIOTICSThere is an urgent need to discover and develop new antibiotics to treat resistant infections caused by superbugs. Most of the antibiotics we use today are produced by one group of soil bacteria, the Streptomyces. Researchers at Cardiff University discovered that another group of bacteria, known as Burkholderia, produce novel antibiotics that kill superbugs that have become resistant to current antibiotics such as Methicillin Resistant Staphylococcus aureus (MRSA). To enable clinical and commercial development, the chemical structures of antibiotics must be determined to see if they are novel compounds. Via collaboration with experts at the University of Warwick's Department of Chemistry, we have determined complete structures for one Burkholderia antibiotic, enacyloxin IIa, and partially determined the structures of two novel antibiotics, gladiolin and vietnamycin. Purifying and determining the chemical structures of antibiotics takes time and considerable expertise, therefore researchers are now using a new tool for antibiotic discovery called genome mining. This approach takes the entire genetic code (genome sequence) of an antibiotic producing bacterium and then searches it for genes involved in the production of novel antibiotics. The Wellcome Trust Sanger Institute (WTSI) are pioneers in bacterial genome sequencing and have new technology called Pacific Biosciences sequencing, which is ideally suited to rapid bacterial genome determination in a single sequence run. In collaboration with researchers at WTSI, we sequenced the genome of the gladiolin producing strain and were able to identify the genes involved in production of this antibiotic. Using this extra sequence information, we can now determine the full structure of gladiolin which is needed to take the antibiotic forward into clinical development. Overall, our preliminary research has shown that Burkholderia are an untapped, promising source of new antibiotics to combat multidrug-resistant superbugs. To determine if Burkholderia can be as successful as the Streptomyces bacteria which produce the majority of the antibiotics we use today, we will use genome mining as a new Burkholderia antibiotic discovery tool, and aim to: 1. Genome sequence 24 antibiotic producing Burkholderia using the Pacific Biosciences sequencing technology. Firstly, to advance characterization of vietnamycin, our anti-MRSA antibiotic, the Burkholderia strain producing this antibiotic will be sequenced. 23 more strains which kill current drug resistant bacteria will then be selected from our large collection. In total, the 24 antibiotic producing Burkholderia genomes will create a unique library of DNA for genome mining and discovery of new antibiotics.2. Fully identify the structure for gladiolin and vietnamycin, using a combination of chemistry and genetic analysis of their genomes and production pathways.3. Carry out a unique genome mining survey of Burkholderia antibiotic capacity using the genomes of the 24 strains known to have novel resistance-busting activity. The first two goals of the grant described above will create the genome mining data and also train the researchers on the grant in all the skills necessary to implement this new approach to antibiotic discovery. We will also investigate if "silent pathways" which encode novel antibiotics within the genomes of these Burkholderia can be switched on.4. The last objective of our research will be to prepare a panel of five novel antibiotics to take forward into preclinical development as drugs to combat resistance. In addition to gladiolin and vietnamycin, we will use the information from genome mining to complete the structure and activity analysis of three further novel Burkholderia antibiotics.