Project description:Bdellovibrio bacteriovorus 109J is a predatory bacterium which lives by predating on other Gram-negative bacteria to obtain the nutrients it needs for replication and survival. Here, we evaluated the effects a bacterial signaling molecule called diffusible signaling factor (DSF) have on B. bacteriovorus 109J transcriptomics.
Project description:Bdellovibrio bacteriovorus is a Gram-negative bacterium that is a pathogen of other Gram-negative bacteria, including many bacteria which are pathogens of humans, animals and plants. As such Bdellovibrio has potential as a biocontrol agent, or living antibiotic. B. bacteriovorus HD100 has a large genome and it is not yet known which of it encodes the molecular machinery and genetic control of predatory processes. We have tried to fill this knowledge-gap using mixtures of predator and prey mRNAs to monitor changes in Bdellovibrio gene expression at a timepoint of early-stage prey infection and prey killing in comparison to control cultures of predator and prey alone and also in comparison to Bdellovibrio growing axenically (in a prey-or host independent “HI” manner) on artificial media containing peptone and tryptone. From this we have highlighted genes of the early predatosome with predicted roles in prey killing and digestion and have gained insights into possible regulatory mechanisms as Bdellovibrio enter and establish within the prey bdelloplast. Approximately seven percent of all Bdellovibrio genes were significantly up-regulated at 30 minutes of infection- but not in HI growth- implicating the role of these genes in prey digestion. Five percent were down-regulated significantly, implicating their role in free-swimming, attack-phase physiology. This study gives the first post- genomic insight into the predatory process and reveals some of the important genes that Bdellovibrio expresses inside the prey bacterium during the initial attack. Keywords: Transcriptional analysis
Project description:Bdellovibrio bacteriovorus is a Gram-negative bacterium that is a pathogen of other Gram-negative bacteria, including many bacteria which are pathogens of humans, animals and plants. As such Bdellovibrio has potential as a biocontrol agent, or living antibiotic. B. bacteriovorus HD100 has a large genome and it is not yet known which of it encodes the molecular machinery and genetic control of predatory processes. We have tried to fill this knowledge-gap using mixtures of predator and prey mRNAs to monitor changes in Bdellovibrio gene expression at a timepoint of early-stage prey infection and prey killing in comparison to control cultures of predator and prey alone and also in comparison to Bdellovibrio growing axenically (in a prey-or host independent âHIâ manner) on artificial media containing peptone and tryptone. From this we have highlighted genes of the early predatosome with predicted roles in prey killing and digestion and have gained insights into possible regulatory mechanisms as Bdellovibrio enter and establish within the prey bdelloplast. Approximately seven percent of all Bdellovibrio genes were significantly up-regulated at 30 minutes of infection- but not in HI growth- implicating the role of these genes in prey digestion. Five percent were down-regulated significantly, implicating their role in free-swimming, attack-phase physiology. This study gives the first post- genomic insight into the predatory process and reveals some of the important genes that Bdellovibrio expresses inside the prey bacterium during the initial attack. Keywords: Transcriptional analysis 3 replicates of attack phase cells and 3 replicates of 30 minutes post-infection cells were analysed on individual arrays. Replicate 3 was normalized separately.
Project description:Bdellovibrio is a Gram-negative bacterium that preys upon other Gram-negative bacteria, including many pathogens, and as such has potential as a biocontrol agent. Little is known of the molecular and genetic control of Bdellovibrio’s attack upon its prey and of the nature of the HI phenotype. Here, we apply microarray technology to monitor changes of gene expression during the initial stages of prey infection to determine which predatory genes are important in this stage and to gain insight into possible regulatory mechanisms controlling the predation process. Comparison to gene expression during HI growth reveals a “predatosome” of genes specifically upregulated during predation and implicates some of those important in HI growth.
Project description:Bdellovibrio is a Gram-negative bacterium that preys upon other Gram-negative bacteria, including many pathogens, and as such has potential as a biocontrol agent. Little is known of the molecular and genetic control of Bdellovibrioâ??s attack upon its prey and of the nature of the HI phenotype. Here, we apply microarray technology to monitor changes of gene expression during the initial stages of prey infection to determine which predatory genes are important in this stage and to gain insight into possible regulatory mechanisms controlling the predation process. Comparison to gene expression during HI growth reveals a â??predatosomeâ?? of genes specifically upregulated during predation and implicates some of those important in HI growth. 3 replicates of attack phase cells and 3 replicates of Host-Independent grown cells were analysed on individual arrays.
Project description:The experimental project studied a MIDAS adhesin minus mutant of predatory bacterium B. bacteriovorus.The predatory bacterium normally invades and lives inside E.coli bacteria, rounding them up to form a two-bacterial structure, called a bdelloplast, and killing the E.coli from the inside. However the MIDAS mutant predator failed to invade in 10% of cases due to one of its (many) attachment/invasion mechanisms being absent. We enriched and purified the 10% of bdelloplasts which did not have an invaded predator inside, by Percoll gradient centrifugation. Although these bdelloplasts did not have an invaded predator they were still rounded and dead. We sent the bdelloplast sample for total protein content analysis at the Oxford Advanced Proteomics Facility. We found that although the bdelloplasts areE.coli cells they also contain secreted Bdellovibrio proteins that normally an invading wild type Bdellovibrio is known to secrete into their prey, during invasion. This suggests that a short-lived failed attachment allowed the Bdellovibrio to secrete in predatory proteins , even though it failed to enter the E.coli, and that those predatory proteins alone were enough to round and kill it.
Project description:Bdellovibrio bacteriovorus HD100 is a predatory bacterium which attacks a wide range of gram negative bacterial pathogens and is proposed to be a potential living antibiotic. In the current study, we evaluated the effects of indole, a bacterial signaling molecule commonly produced within the gut, on the predatory ability of B. bacteriovorus HD100. Indole significantly delayed predation on E. coli MG1655 and S. enterica KACC 11595 at physiological concentrations (0.25 to 1 mM) and completely inhibited predation when present at 2 mM. Microscopic analysis revealed that indole blocked the predator from attacking the prey. Furthermore, indole was not toxic to the predator but slowed down its motility. Microarray and RT-qPCR analyses confirmed this as the gene group showing the greatest down-regulation in the presence of 1 and 2 mM indole was flagellar assembly and motility genes. Aside from this group, indole also caused a wide spectrum changes in gene expression including the general down-regulation of genes involved in ribosome assembly and RNA translation. Furthermore, indole addition to the predatory culture after the entrance of B. bacteriovorus into the prey periplasm slowed down bdelloplast lysis. In conclusion, indole is an important gut-related signaling molecule that can have significant impacts on the predation efficiency and predator behavior. These findings should be taken into consideration especially if B. bacteriovorus is to be applied as a probiotic or living antibiotic.
Project description:Bdellovibrio bacteriovorus HD100 is a predatory bacterium which attacks a wide range of gram negative bacterial pathogens and is proposed to be a potential living antibiotic. In the current study, we evaluated the effects of indole, a bacterial signaling molecule commonly produced within the gut, on the predatory ability of B. bacteriovorus HD100. Indole significantly delayed predation on E. coli MG1655 and S. enterica KACC 11595 at physiological concentrations (0.25 to 1 mM) and completely inhibited predation when present at 2 mM. Microscopic analysis revealed that indole blocked the predator from attacking the prey. Furthermore, indole was not toxic to the predator but slowed down its motility. Microarray and RT-qPCR analyses confirmed this as the gene group showing the greatest down-regulation in the presence of 1 and 2 mM indole was flagellar assembly and motility genes. Aside from this group, indole also caused a wide spectrum changes in gene expression including the general down-regulation of genes involved in ribosome assembly and RNA translation. Furthermore, indole addition to the predatory culture after the entrance of B. bacteriovorus into the prey periplasm slowed down bdelloplast lysis. In conclusion, indole is an important gut-related signaling molecule that can have significant impacts on the predation efficiency and predator behavior. These findings should be taken into consideration especially if B. bacteriovorus is to be applied as a probiotic or living antibiotic. Bdellovibrio bacteriovorus HD100 was incubated for 30 min at 30°C in HEPES buffer supplemented with 0,1, and 2 mM indole. RNA was then extracted from each sample and purified. 100 ng of RNA from each sample were used for microarray experiment. For zero and 1 mM indole treatments, three independant samples were tested while for 2 mM indole treatment, two samples were tested. A total of 8 arrays were used.