Control of predation and development in Bdellovibrio bacteriovorus

Faculty mentor: Dr. John Tudor

Research interests:  Study of the developmental cycle of the obligatory predacious bacterium Bdellovibrio

Background:

Members of the genus Bdellovibrio are a diverse group of predatory Gram-negative bacteria that  exhibit  a unique biphasic life cycle. The several diverse members of this group share many characteristics with other predatory and pathogenic bacteria.  In the attack phase,  bdellovibrios exist as small,  highly motile, curved rods found  in  a  variety of macroenvironments.  The highly motile, attack-phase bdellovibrios are propelled by a single, polar, sheathed flagellum. The life cycle of the bdellovibrios  alternates between the motile, non-growing attack-phase, spent in a variety of macroenvironments, and the growth phase, spent within the periplasm of a susceptible  bacterium.  Most strains are capable of preying upon Gram-negative enteric bacteria and others with similar LPS structure. All wild type Bdellovibrio isolated have been obligately predacious, having an absolute requirement for prey cells in order to reproduce.  My laboratory has been examining the nature of this Bdellovibrio-prey cell interaction for a number of years.  Understanding this unique interaction is important as a model system of prokaryotic differentiation and cell-cell interactions.  It could also have broad biological significance and serve as a model for understanding the molecular basis of the obligate interaction between some medically important bacteria and their hosts.

The attachment of the Bdellovibrio cell to its prey following the initial collision proceeds in two stages.  Initial attachment is reversible and non-specific, whereas the second stage of attachment is specific and irreversible.  A few minutes following irreversible attachment, the bdellovibrio moves through the outer membrane and peptidoglycan of its prey, leaving its detached flagellum behind.  My lab, as well as others have shown that a variety of enzymatic activities are released, which are directed against the prey cell envelope.  Penetration results in the almost immediate death of the invaded cell.  Penetration of the prey’s outer envelope is followed immediately by the transformation of the prey cell into an osmotically-stable growth chamber, termed a bdelloplast.   Bdellovibrio macromolecular synthesis and cellular growth appear to begin between 45 and 60 minutes following the initial attack. The bdelloplast serves as a unique chamber specifically modified by the Bdellovibrio to support its highly efficient intracellular growth phase. Macromolecular synthesis and DNA replication within the growing Bdellovibrio cell is confined to the intraperiplasmic growth-phase and is initiated from the uptake of monomeric units derived from the degrading prey cell cytoplasm that have leaked into the periplasmic space. We have shown that nutrients from the prey cell protoplast are acquired from the periplasm by the translocation of outer membrane porins into the cytoplasmic membrane of the bdelloplast.

Growth of the  intraperiplasmic  bdellovibrio  proceeds with the elongation of  the invading attack cell into a long aseptate spiral-shaped filament. When the bdelloplast  is exhausted of nutrients,  the elongated spiral cell fragments into individual attack-phase  bdellovibrios,  each with a newly synthesized polar sheathed-flagellum.   A lytic enzyme is then induced that breaks down the modified bdelloplast peptidoglycan from the inside, resulting in lysis of the bdelloplast  and release of the progeny bdellovibrios. The length of the intraperiplasmic growth-phase is approximately three and one-half hours at 30°C for B. bacteriovorus  growing on E. coli as prey.  When grown axenically, prey-independent strains of Bdellovibrio follow the same pattern of growth characterized by elongation of motile cells into non-motile spiral-shaped filaments and fragmentation into individual motile progeny.  None of the details regarding the signaling processes involved, DNA replication during elongation of  the periplasmic filamentous cell,   segregation of chromosomes into attack cells, cell division or gene expression and regulation are known.

Current Research Direction:

My laboratory is now focusing on identifying and characterizing the genetic control of the bdellovibrio developmental cycle.  In the past two to three years, the genomes of three bdellovibrio strains have been sequenced, and offer a great opportunity to further study the genetics, physiology, and biochemistry of these unique predators.  One obvious approach would be examining the transcriptional activity of these predators throughout their life cycle in order to assign specific functions related to the predatory life style to individual genes.  This approach, along with analysis of sequenced genomes, would provide valuable information concerning temporal gene expression during differing phases of development.

 While the use of genomic analysis and global expression patterns are powerful tools, they are limited in defining gene function, especially in regard to predicted genes with little or no detectable similarity to known genes. My laboratory has recently developed a protocol that permits the introduction of transposons into facultatively predaceous bdellovibrios, and the subsequent screening of large numbers of transposon-insertion mutants for predation using two different facultative strains of B. bacteriovorus 109J. These facultative predators thus have the potential for use as a system for identifying genes essential for predatory-growth but dispensable for prey independent growth. A facultative mutant defective in a gene required only for predatory growth (such as a mutant unable to express the enzymes required for prey cell penetration) would still be viable and could be grown in complex medium, but not on prey cells. We believe this is a very fruitful approach for the identification of genes necessary for predaceous growth only.  Using this technique, students in the lab have isolated and sequenced fifteen predatory genes. Expression analysis of these suspected predatory genes can easily be analyzed by looking at temporal gene expression using the real-time polymerase chain reaction (RT-PCR) or Northern blots. 

The ongoing research agenda for my laboratory is to continue isolating new predatory mutants and analyzing expression of essential predatory genes with a view of identifying function of gene products important in the bdellovibrio developmental cycle.

Contact information:

Dr. John Tudor 
Email: jtudor@sju.edu
Mailing address:
Biology Department
5600 city Ave
Philadelphia, PA 19131
Phone: 610-660-1821
Fax 610-660-1832