Field of Science

#Microtwjc 1 - bacterial cell shape and pathogenesis

This is new ground for me as a virologist, writing about bacterial genetics and pathogenesis. But it's for a good cause - the cause of Microbiology Twitter Journal Club or #microtwjc. Set up by Zoonotica and encompassing a number of other microbiologists. This post serves background reading for the discussion on this paper on Tuesday the 8th May at 20:00 BST.

This paper aims to determine the role of bacterial shape/structure in how micro-organisms cause disease. I don't think they actually succeed in doing this as it's going to be a pretty complicated picture. But it does highlight a number of interesting points in bacterial genetics and pathogenesis, especially for an organism in which very little is really known.


Two very different structures - ultimately a single genetic change. But how?



They use Campylbacter Jenjuni as a model for this as the bacteria has a peculiar structure (corkscrew) that has been linked - but not proven - to be an adaptation to life in an animals gut. This bacterium is a commensal organism of chickens and is found throughout it's GI tract and often it can infect and cause disease in other animals, like humans. 


We have to first realise that it's structure must have been adapted to it's lifestyle. Not necessarily to burrowing through gut epithelia but it could have functions in immune evasion, cell division and cell attachment etc. We haven't been able to test these different theories because before this paper no genetic determinants were known to govern C.jejuni cell shape. 


It's all because of this guy: pgp1


Using an assay to detect changes in peptidoglycan structures, the group identified one particular gene:  cjj81176_1344 which they now called pgp1. This gene, when mutated to be non-functional, caused chemical changes in the bacterium's cell wall as picked up by their fluorescence assay. The rest of the paper is characterising this one gene.


This gene encoded a protein which is very highly conserved among helical and vibroid bacteria. Pgp1 is an enzyme and in particular it is a metallocarboxypeptidase. This means it cleaves the bonds between amino acids at one side (the carboxy-terminus) and not the amino end. Using bioinformatic tools they think this protein is found in the cell periplasm and could have a role in cell wall maintenance/building and hence shape.


To test if this gene had a role in structure and pathogenesis, they knocked it out and knocked it in to a number of C.jenjuni isolates. They use these bacteria in in vitro and in vivo assays, such as: morphology, motility, biofilm formation, chemical analysis, host colonisation and innate immune activation. I've summarised their results below:


1) under the electron microscope the bacteria look different when you knock pgp1 out. They look bulkier and are not twisted/helical. I'm not sure if they are longer/shorter or have increased/decreased surface area. These things are important but it isn't clear.


2) in vitro it has slight motility and biofilm formation differences. 


3) When they overexpressed the protein it had the same effect as when they removed it. Suggesting that it isn't simply an off/on issue.


4) Chemically, their cell walls looked pretty different. It looks like they have the same kinds of peptidoglycan structures in them but have different quantities of each component. Which is indicative that it is the ratios of each component that effects cell shape. This is important because not only do the cells look different, they have different chemical structures and could therefore 'look' different to the host.


Peptidoglycan differences when you remove the pgp1 gene. Seems like they have the same kinds of structures, only different amounts.


5) In chickens, this bacteria's host, it has a significant colonisation defect when the protein is knocked out.


6) There are slight innate immunity differences when they looked at human immunity. Not chicken.


Pathogenesis - colonisation differences in chickens and differences in innate immunity in a human system.


Conclusions:


Linking a single gene to cell shape and then to bacterial life cycle is pretty complicated. This gene seems to control the relative amounts of peptidoglycan components in the bacterial cell wall and this in turn influences cell shape/motility/biofilm/colonisation/immune response. Linking the change in shape directly to these factors, in my opinion, seems premature as this bacteria now has essentially a completely different cell wall (and other chemical changes possibly) which could have a number of influences on many biological processes. 


But this is the first step in an interesting story of the role of bacterial shape/structure in life cycle. This is probably why it focusses on the characterisation of this system. I don't doubt that the structure plays a role in pathogenesis but it is likely to be a very complicated and convoluted story.


Future experiments I would do: rule out the chance that changes simply in chemical structure of the cell wall could effect immune recognition or even motility etc. Or even changes in cell size/surface area.


Perform targeted knock out of specific regions of pgp1 protein to determine exactly how this molecule governs cell shape and possibly other functions.


Delve into chicken immunology, no point confusing things using a chicken colonisation assay followed by human innate immunity.


Knock this gene out in other helical/vibroid bacteria. Does it still function?


ResearchBlogging.orgFrirdich, E., Biboy, J., Adams, C., Lee, J., Ellermeier, J., Gielda, L., DiRita, V., Girardin, S., Vollmer, W., & Gaynor, E. (2012). Peptidoglycan-Modifying Enzyme Pgp1 Is Required for Helical Cell Shape and Pathogenicity Traits in Campylobacter jejuni PLoS Pathogens, 8 (3) DOI: 10.1371/journal.ppat.1002602

1 comment:

  1. I didn't see this post before, but this is great coverage of the paper! I was really excited by the shape change, it's always great when a gene knockout has such a clear phenotypic change.

    ReplyDelete

Markup Key:
- <b>bold</b> = bold
- <i>italic</i> = italic
- <a href="http://www.fieldofscience.com/">FoS</a> = FoS