Do bacteria communicate using nanotubes?

Until Friday I'm at the Microbial Population Genetics Gordon Conference, a black hole from which no tweets or blog posts must emerge.  Well, that's an exaggeration - we just can't tweet or post about the conference.  But someone at last night's excellent poster session drew my attention to a remarkable paper published in February, and now that I've read it I'm going to write about it here.

The paper is titled Intercellular nanotubes mediate bacterial communication.  It's by Gyanendra Dubey and Sigal Ben-Yehuda (Hebrew University) and appeared in Cell 144:590-600.  (It's behind a paywall here, but you can find a pdf of it here at OpenWetWare).The authors found that Bacillus subtilis cells form tubular connections through which small molecules, proteins and plasmids can pass from one cell to another.  Although the authors' ideas about bacterial communication and cooperation appear to owe more to Sesame Street than to evolutionary theory, this is a very surprising and important finding.

The paper first reports that, when a mixture of B. subtilis cells with and without GFP were mixed on an agar-solidified medium,  GFP- cells lying next to GFP+ cells gradually acquired GFP.  The same transfer happened if some cells instead were preloaded with a different fluorophore, one known to not pass through cell membranes.  If the cells contained different chromosomally encoded antibiotic resistance proteins, some cells became transiently resistant to both antibiotics.  And if some cells contained an antibiotic-resistance gene on a plasmid, some of the plasmid-free cells acquired the plasmid.  Plasmid acquisition was not blocked by DNase I and did not occur when a sub-inhibitory concentration of the detergent SDS was added to the mixed cells or when free plasmid DNA was added to plasmid-free cells.

The obvious next step was to look at the cells with electron microscopy.  This showed tubular connections between cells, which the authors unfortunately chose to call 'nanotubes'.  (Unfortunately because nanotube is a very well established term for tubular filaments of fullerene

How the authors prepared the cells for EM is probably important.  Normally cells are first suspended in liquid and then placed on an EM grid, but this might disrupt any cell connections.  So instead the authors grew cells on agar medium for 3 hr, and then placed EM grids on top of the cells.  They let the cells grow for 3 more hr and then lifted the grids and the attached cells from the agar.  It would have been good to have repeated this unconventional procedure using cells that were already dead when placed on the plate, or otherwise incapacitated so they couldn't actively form connections.  This would show that the apparent connections aren't just coming from the agar. 

What's bizarre about this result is that nobody has ever observed these connections before, but the authors don't discuss why the connections wouldn't have been discovered by earlier B. subtilis researchers. It's true that researchers may not have used exactly these conditions.  However the connections must have formed quite quickly in their experiments, as evidence of transfer was seen within 15 minutes, so I would think that anyone looking at cell behaviour under a microscope would have noticed that cells became stuck together. 

It could be that microbiologists have overlooked this because bacteria aren't usually co-cultivated or agar surfaces.  But growth on surfaces is the norm for bacteria in natural environments.  If connecting tubes and molecular transfer were this ubiquitous, phages and plasmids would be much more uniformly distributed both within and between species.  Evolutionary processes would also be very different; mutant phenotypes would blend when different strains made contact.

Overall I'm dubious. The data looks OK, but the phenomenon doesn't make biological or evolutionary sense.

***A few more details I picked up on rereading the paper:

• The frequency of plasmid transfer after >4 hr of co-cultivation was only 10^-7.  That's very low, given the apparently high transfer rate of other molecules.
• Transfer was blocked by levels of SDS that didn't affect cell growth - this suggests that the tubes do not have the same envelope as the cell bodies.