Time - Scale Pan Genome Analysis of Horizontal Gene Transfer in heterogeneous Bacteria Pool
Time-Scale Pan Genome Analysis of Horizontal Gene Transfer in Heterogeneous Bacteria Pool
Jung, Young Kwang and Jung, Woo Chul
UNIST Genomics 2016
Introduction
In genomic classes, our professor tells us a lot of interesting things about eukaryotic genomes such as leopard, whale, tiger, human genomes. But I think we had no chance to learn about the bacterial genomes. So we`ll introduce prokaryotic genome in this paper and also give our question about that.
Pan genome is prokaryotic genomes(include bacteria and archaea) which has variation in species or strains such as E. coil, streptococcus agalactiae. It is composed of 3 components : core genome, accessory genome, and specific genes.
Core genome is a set of essential genes in bacterial species. The genome has basic function to maintain the organism such as replication, translation and homeostasis. It is common genome in the species, so it is very important to survive. Therefore by natural selection, the core genome is not changed.
But the accessory genome has many variations in the bacteria. Because of these variations in the accessory genome, it can make possible to adapt different environments such as soil, water, acidic hot springs, radioactive waste and etc. Through these different adaption to many habitats, the bacteria can have different life styles like pathogen or resistance to antibiotics.
Specific genes can be distributed as two types: species - specific genes and strain - specific genes. Species - specific genes are genome about species. Only single species has the species - specific genes compared with other species. And Strain - species genes are genome about strains. Only one strain has the strain - specific genes compared with other strains. So specific genes show the characteristics of the species or strains.
Gene transfer is a phenomena that transfer genes from one to another. It is composed of two kinds : vertical gene transfer and horizontal gene transfer. Vertical gene transfer is a transferring genes from first generation to next generation. And horizontal gene transfer is transferring genes between different species or same species. In fact, we think horizontal gene transfer can make more mutations and can change more the composition of pan genome than vertical gene transfer, so we`ll focus on the horizontal gene transfer.
The horizontal gene transfer is composed of 3 parts: conjugation, transduction, transformation. To explain about the concept of the transfer, we need to know the term, donor cell and recipient cell. Donor cell is the cell that gives DNA to another. And recipient cell is the cell that receives the DNA from donor cell.
Conjugation occurs when donor cell meets recipient cell and makes linkage, called pilius. And through the pilius, genes of donor cell transfer into recipient cell.
Transduction is the gene transfer between bacteria and bacteriophage. When bacteriophage invade the bacteria, the phage makes its DNA from the bacteria. And the virus(donor) that has bacterial DNA transfer the gene of bacteria into another bacteria(recipient).
Transformation occurs by lysis of the bacteria. When a bacterium is broken out by the lysis, the gene of the bacteria diffuse out and can be free. So the free bacterial DNA(donor) go into another bacteria(recipient).
We predict relation between pan genome and horizontal gene transfer, but actually there is no clear relation between them in our research papers. So our question is like this: how can pan genome change by the horizontal gene transfer. For the question, we`ll introduce our hypothetic experiment in this paper.
Method
We make an assumption that can occur horizontal gene transfer. There are recipient bacteria A which can have resistance to antibiotics a, not b and there are another bacteria B which can have resistance to antibiotics b, not a and can occur horizontal gene transfer.
1. Prepare a plate for the bacteria A
2. Put the bacteria B into the plate of A
3. Check the time for every 30 minutes, and treat both antibiotics a and b into the plate according to the times.
4. Count the number of colonies and analyze the results in the plate.
Experiment
Prepare natural bacterial mixture We are supposing acquisition of substrate bacteria from the nature, for example, water sample from Ga-mak pond or micro-biome sample from human gut. The reason why we choose this way is that there is possibility that a certain bacterial species reacts to gene transferring situation differentially in heterogeneously constituted situation in nature, compared with one in isolated species sample in vitro. More specifically, we presume that one species can induce more or less gene transfer between other species in a certain context due to some reason other than population effect.
Horizontal gene transfer screening experiment First, we would set control group experiment for which we prepare a separated bacterial sample from natural mixture. We need separation process to carry out HGT experiment on each distinct species in the sample, so that we apply the introduced method to each of the segregated species samples with the same time scale. Likewise, we can also prepare experimental group which consists of heterogeneous species as a mixed sample. We track the viability of each species in recipient bacteria mixture using specific antibody which the species have resistance against. The result from control and experimental group will show trend line of relation between number of viable colonies and time variation of antibiotic treatment. We expect, moreover, that the trend might be either positively or negatively affected in experimental group under heterogeneously constituted condition.
Genome sequencing To see how this difference of these phenotype is related to the genome, we investigate the genome of those distinct samples. There might be three different version genome for a single species each from the linage isolated from natural mixture, the recipient linage after HGT experiment of the isolate species, and the recipient linage after HGT experiment of the heterogeneous species pool. With the sequenced data we can construct a pan genome from the genome group of different species, or also from the group of genome from single species in different contexts. Either way, we can investigate change of the gene set due to gain from HGT and maybe loss of some gene related to HGT reception after HGT screening. Moreover, there might be several genes related to inter-species effect on HGT of each species in natural bacterial pool.
Discussion
Dynamics of horizontal gene transfer There might be a case that lateral gene transfer on one species is affected by existence of another species. More specifically, a certain gene may encode for proteins related to interactions between them. Some of the gene are presumed to be related to the velocity of the transfer. We expect that we would see how the gene set is affected by horizontal gene transfer, and make a database for further investigation in which we can use assess the vulnerability of novel bacterial sample that inhabit in a certain environment.
Limitation What we described so far is based on our hypothetical experimental design. We see there are a lot of limitations existing and few of them are significant as we should consider alternative design at most. When we separate each species from the natural pool of bacteria, biggest problem reported is that most of them are not viable on separated condition in a laboratory. We should choose several species which are already reported as able to be cultured in isolated form, as a result, this experiment cannot reflect actual dynamics in a nature state. Even if we could isolate a certain species, obtaining antibiotics against which they are resistant specifically is another problem. We should prepare at least two distinct antibiotics each for donor and recipient bacteria. One more thing to consider is that cell division can affect the observation of recipient bacterial cell.
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