Genetics of the switch from predatory to axenic growth in the living antibiotic Bdellovibrio bacteriovorus

蛭弧菌是一种小型的、对人类友好的捕食性细菌，它会侵入其他病原菌的细胞（比如那些让老年人感染压疮和糖尿病溃疡的细菌，以及感染农作物的细菌）并杀死它们。蛭弧菌对人类、动物或植物细胞没有活性，但它们天生擅长杀死其他细菌。致病细菌没有简单的细胞表面受体用于蛭弧菌附着和入侵，因此它们不容易对蛭弧菌产生抗性（与传统抗生素的情况不同）蛭弧菌在20世纪60年代被发现，最近我们和其他人一直在研究它们的基因和基因组，并试图了解它们如何对人类有用。我们建议它们可以用于乳膏或喷雾剂中，用于人类感染的伤口，也可以喷洒在受感染的作物上，以杀死感染的作物腐烂细菌。蛭弧菌几乎是唯一的捕食性细菌，大多数细菌生长在食物上，不侵入活菌。蛭弧菌实际上有一个3.8Mb的大基因组（我们在2004年帮助测序），并获得了像正常细菌一样生长的基因，以及捕食性的基因。为了有效地杀死病原体，蛭弧菌必须失去像普通细菌一样在食物中正常生长的能力，因为这种能力使它们能够在伤口的血清中，汁液中和植物周围的土壤中生长，而不需要是捕食性的并杀死引起感染的目标致病菌。因此，我们项目的重点是研究它们如何非捕食性地生长（如所谓的HI菌株），我们将研究出哪些基因编码这些特性，并试图在不损害捕食性生长（我们想用它来杀死病原体）的情况下将它们重新编码。以前的科学家在1992年发表了一篇很好的论文，表明当蛭弧菌作为HI以非捕食性方式生长时，基因组中称为命中位点的一个基因发生了变化，这可能是部分原因，但2001年的另一篇论文表明，并非所有以非捕食性方式生长的蛭弧菌都在这个位点发生了变化，所以在这个过程中还有更重要的基因需要发现。目前还没有人知道hit基因座的作用以及它如何控制HI的生长--我们想在这里做这件事，看看如果我们生产活的抗生素，HI菌株的遗传稳定性如何。我们已经进行了以前的研究，表明当蛭弧菌捕食性生长和作为HI生长时，基因组中的所有基因都被打开;有很多基因差异，有些基因是专门打开的，只是为了作为HI生长。我们希望研究这些基因的作用，特别是那些调节基因组开启的基因--我们也许能够删除其中一个调节基因，关闭整个HI生长能力，留下一个专性捕食的蛭弧菌作为活的抗生素。我们也已经能够偶然分离出一种突变的蛭弧菌，我们发现它不能作为HI生长，因此它是专性捕食性的。这种菌株在从复杂分子中获得铁以供生长方面存在缺陷，但可以在掠夺性生长模式中生长良好，可能是因为它能够通过消化猎物细菌的蛋白质来从猎物细菌中释放铁。因此，该菌株表明，我们将能够敲除蛭弧菌中控制HI生长的基因，而不会消除捕食性生长的能力。在我们的项目中，我们希望对不能作为HI生长的铁清除突变体中表达的基因进行分析，以试图了解它的问题所在。长期的想法是将蛭弧菌开发为人类，动物和植物疾病的真实的新疗法，这是重要的第一步之一

Bdellovibrio are small, human-friendly, predatory bacteria which invade the cells of other pathogenic bacteria (such as ones that give old people infections in pressure sores and diabetic ulcers, and bacteria that infect crop plants) and kill them. Bdellovibrio have no activities against human, animal or plant cells but they are naturally good at killing other bacteria. Pathogenic bacteria do not have simple cell surface receptors for Bdellovibrio attachment and invasion and so it is not easy for them to develop resistance to the Bdellovibrio (unlike the situation with conventional antibiotics) Bdellovibrio were discovered in the 1960s and recently we and others have been researching their genes and genomes and trying to understand how they may be useful to humans. We propose that they can be used in a cream or a spray to apply to infected wounds in humans and also to be sprayed on infected crop plants to kill off infections crop rot bacteria. Bdellovibrio are almost unique in being predatory bacteria, most bacteria grow on foods and do not invade live bacteria. Bdellovibrio actually have a large 3.8Mb genome (which we helped to sequence in 2004) and have got both the genes to grow like normal bacteria, and the genes to be predatory.To be efficient at killing pathogens Bdellovibrio will have to lose the ability to grow normally on food like regular bacteria, because that ability allows them to grow in serum in wounds and in sap and on soil around plants, without needing to be predatory and kill the target pathogenic bacteria that are causing the infection. So, the point of our project is to work out how they grow non-predatorily (as so-called HI strains), we will work out which genes code for these properties and try to inactivate them, without impairing predatory growth (which we want to use to kill the pathogens). Previous scientists have published one nice paper in 1992 showing that one gene, in a region of the genome called the hit locus, has changes within it when the Bdellovibrio are growing non predatorily as HIs and that this might be partly responsible, but another paper in 2001 showed that not all Bdellovibrio that were growing non-predatorily had changes in this locus, so there are more important genes to find in the process too. No-one has yet found out what the hit locus gene does and how it might control HI growth- we want to do this here and to see how genetically stable HI strains may be if we produce as living antibiotics. We have carried out previous research that has shown all the genes switched on from the genome when the Bdellovibrio are growing predatorily and when they are growing as HIs; there are lots of gene differences and some are specifically switched on just to grow as HIs. We want to investigate what these genes do, especially some that regulate the switching on of groups of genes- we may be able to delete one of these regulator genes and switch off the whole HI growing abilities, leaving an obligately predatory Bdellovibrio for use as a living antibiotic. We have also already been able to isolate a mutant Bdellovibrio by chance, that we found to be unable to grow as an HI and which is therefore obligately predatory. This strain has a defect in obtaining iron for growth from complex molecules, yet can grow fine in the predatory growth mode, probably because it is able to free the iron from prey bacteria by digesting their proteins. Thus this strain shows that we will be able to knock out genes in Bdellovibrio that control HI growth without abolishing the ability to grow predatorily. In our project we want to profile the genes that are expressed in our iron-scavenging mutant that can't grow as an HI, to try to understand what is wrong with it. The long term idea is to develop Bdellovibrio as a real new treatment for human, animal and plant disease and this is one of the important first steps