Affiliation:
1. Department of Biological Safety, Federal Institute for Risk Assessment, Berlin, Germany
2. Max Planck Institute for Molecular Genetics, Berlin-Dahlem, Germany
3. Laboratory of Applied Analytical and Environmental Chemistry, Humboldt-Universitaet zu Berlin, Berlin, Germany
Abstract
ABSTRACT
Bdellovibrio bacteriovorus
HD100 is an obligate predatory bacterium that attacks and invades Gram-negative bacteria. The predator requires living bacteria to survive as growth and replication take place inside the bacterial prey. It is possible to isolate mutants that grow and replicate outside prey bacteria. Such mutants are designated host or prey independent, and their nutritional requirements vary. Some mutants are saprophytic and require prey extracts for extracellular growth, whereas other mutants grow axenically, which denotes the formation of colonies on complete medium in the absence of any prey components. The initial events leading to prey-independent growth are still under debate, and several genes may be involved. We selected new mutants by three different methods: spontaneous mutation, transposon mutagenesis, and targeted gene knockout. By all approaches we isolated mutants of the
hit
(host interaction) locus. As the relevance of this locus for the development of prey independence has been questioned, we performed whole-genome sequencing of five prey-independent mutants. Three mutants were saprophytic, and two mutants could grow axenically. Whole-genome analysis revealed that the mutation of a small open reading frame of the
hit
locus is sufficient for the conversion from predatory to saprophytic growth. Complementation experiments were performed by introduction of a plasmid carrying the wild-type
hit
gene into saprophytic mutants, and predatory growth could be restored. Whole-genome sequencing of two axenic mutants demonstrated that in addition to the
hit
mutation the colony formation on complete medium was shown to be influenced by the mutations of two genes involved in RNA processing. Complementation experiments with a wild-type gene encoding an RNA helicase, RhlB, abolished the ability to form colonies on complete medium, indicating that stability of RNA influences axenic growth.
Publisher
American Society for Microbiology
Subject
Molecular Biology,Microbiology
Cited by
34 articles.
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