Dataset Information

Analysis of abrB Expression during the Infectious Cycle of Bacillus thuringiensis Reveals Population Heterogeneity.

ABSTRACT: Using the model host/pathogen pair Galleria mellonella/Bacillus thuringiensis, we have shown that these bacteria could kill their insect host, survive in its cadaver and form spores by sequentially activating virulence, necrotrophism and sporulation genes. However, the population isolated from the cadavers was heterogeneous, including non-sporulating cells in an unknown physiological state. To characterize these bacteria, we used a transcriptional fusion between the promoter of a gene expressed during early exponential growth (abrB) and a reporter gene encoding a destabilized version of GFP, in combination with a fluorescent reporter of the necrotrophic state. The composition of the bacterial population during infection was then analyzed by flow cytometry. We showed that the PabrB promoter was activated in the population that had turned on the necrotrophic reporter, suggesting a re-entry into vegetative growth. Strikingly, the cells that did not go through the necrotrophic state did not activate the PabrB promoter and appear as a dormant subpopulation. We propose a new model describing the B. thuringiensis cell types during infection.

SUBMITTER: Ben Rejeb S

PROVIDER: S-EPMC5732988 | biostudies-literature | 2017

REPOSITORIES: biostudies-literature

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Analysis of abrB Expression during the Infectious Cycle of Bacillus thuringiensis Reveals Population Heterogeneity.

Ben Rejeb Samia S Lereclus Didier D Slamti Leyla L

Frontiers in microbiology 20171212

Using the model host/pathogen pair Galleria mellonella/Bacillus thuringiensis, we have shown that these bacteria could kill their insect host, survive in its cadaver and form spores by sequentially activating virulence, necrotrophism and sporulation genes. However, the population isolated from the cadavers was heterogeneous, including non-sporulating cells in an unknown physiological state. To characterize these bacteria, we used a transcriptional fusion between the promoter of a g ...[more]

PMID: 29312181

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Project description:UNLABELLED:Bacillus thuringiensis (Bt) is armed to complete a full cycle in its insect host. During infection, virulence factors are expressed under the control of the quorum sensor PlcR to kill the host. After the host's death, the quorum sensor NprR controls a necrotrophic lifestyle, allowing the vegetative cells to use the insect cadaver as a bioincubator and to survive. Only a part of the Bt population sporulates in the insect cadaver, and the precise composition of the whole population and its evolution over time are unknown. Using fluorescent reporters to record gene expression at the single-cell level, we have determined the differentiation course of a Bt population and explored the lineage existing among virulent, necrotrophic, and sporulating cells. The dynamics of cell differentiation were monitored during growth in homogenized medium, biofilm formation, and colonization of insect larvae. We demonstrated that in the insect host and in planktonic culture in rich medium, the virulence, necrotrophism, and sporulation regulators are successively activated in the same cell. In contrast, in biofilms, activation of PlcR is dispensable for NprR activation and we observed a greater heterogeneity than under the other two growth conditions. We also showed that sporulating cells arise almost exclusively from necrotrophic cells. In biofilm and in the insect cadaver, we identified an as-yet-uncharacterized category of cells that do not express any of the reporters used. Overall, we showed that PlcR, NprR, and Spo0A act as interconnected integrators to allow finely tuned adaptation of the pathogen to its environment. IMPORTANCE:Bt is an entomopathogen found ubiquitously in the environment and is a widely used biopesticide. Studies performed at the population level suggest that the infection process of Bt includes three successive steps (virulence, necrotrophism, and sporulation) controlled by different regulators. This study aimed to determine how these phenotypes are activated at the cellular level and if they are switched on in all cells. We used an insect model of infection and biofilms to decipher the cellular differentiation of this bacterium under naturalistic conditions. Our study reveals the connection and lineage existing among virulent, necrotrophic, and sporulating cells. It also shows that the complex conditions encountered in biofilms and during infection generate great heterogeneity inside the population, which might reflect a bet-hedging strategy to ameliorate survival. These data generate new insights into the role of regulatory networks in the adaptation of a pathogen to its host.

Project description:Borrelia burgdorferi is a zoonotic pathogen whose maintenance in nature depends upon an infectious cycle that alternates between a tick vector and mammalian hosts. Lyme disease in humans results from transmission of B. burgdorferi by the bite of an infected tick. The population dynamics of B. burgdorferi throughout its natural infectious cycle are not well understood. We addressed this topic by assessing the colonization, dissemination and persistence of B. burgdorferi within and between the disparate mammalian and tick environments. To follow bacterial populations during infection, we generated seven isogenic but distinguishable B. burgdorferi clones, each with a unique sequence tag. These tags resulted in no phenotypic changes relative to wild type organisms, yet permitted highly sensitive and specific detection of individual clones by PCR. We followed the composition of the spirochete population throughout an experimental infectious cycle that was initiated with a mixed inoculum of all clones. We observed heterogeneity in the spirochete population disseminating within mice at very early time points, but all clones displayed the ability to colonize most mouse tissues by 3 weeks of infection. The complexity of clones subsequently declined as murine infection persisted. Larval ticks typically acquired a reduced and variable number of clones relative to what was present in infected mice at the time of tick feeding, and maintained the same spirochete population through the molt to nymphs. However, only a random subset of infectious spirochetes was transmitted to naïve mice when these ticks next fed. Our results clearly demonstrate that the spirochete population experiences stochastic bottlenecks during both acquisition and transmission by the tick vector, as well as during persistent infection of its murine host. The experimental system that we have developed can be used to further explore the forces that shape the population of this vector-borne bacterial pathogen throughout its infectious cycle.

Dataset Information

Analysis of abrB Expression during the Infectious Cycle of Bacillus thuringiensis Reveals Population Heterogeneity.

Publications

Analysis of <i>abrB</i> Expression during the Infectious Cycle of <i>Bacillus thuringiensis</i> Reveals Population Heterogeneity.

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