Sunday, December 6, 2009

Very hard to happen upon aerophiles!

I'm writing this on my Blackberry so I can't check the aerophile literature right now! But here goes anyway:
I found out recently that the biodensity of airborne microorganisms, in colony forming units (cfu), is about:
1 cfu per 10l at ground level.
If same biodensity at altitude, allowing for lower air density:
Say 0.01 bar
1 cfu per 1000l
This is the same as a clean room :-O

How much air at this density would a detector 10cm2 square have to travel through to detect one colony?

10cm3 is 1l

Since 1 cfu per 1000l expected:
So, 1000 x 10cm cube=10,000cm
Or 100m air needs to be travelled through to detect 1 cfu at altitude
So 10cfu per km

U2 aircraft have flown similar transects of the high atmosphere.
A 1000km U2 flight could therefore theoretically detect about 10000cfu.
But only a single colony was detected during such a flight in real life using a similarly sized detector, suggesting that at the culturing conditions used in the experiment is question, biodensity is about 10000 less than at ground level, even correcting for altitude (and actually U2s fly slightly lower and atmospheric pressure is somewhat higher than 0.01bar). And that's assuming the single colony seen wasn't a ground contaminant.

If it was 10,000 less biodense then on the ground, then this means (allowing for altitude) not 1 cfu per 1000l but 1cfu per 10 million l !

But balloons fly far less far. If one opened a sampling port at final altitude, this suggests you'd need to allow the balloon to drift 1000km - OR pump 10 million L through your sampler.

Say you have a 50ml syringe pipetting up and down. 20 strokes is 1L

You would therefore need approximately 200 million strokes to detect one cfu. If each return stroke took 10s, this is 2 billion seconds
This is about 60 years!!

A long duration drifting balloon, flying thousands of kilometres, therefore seems more feasible.

One must also however be aware of sample port size.
The above calculations are for a 100cm2 detector (eg 10x10cm)
If your sampler had only a 1cm2 aperture, this would obviously be a hundred times less. Then the balloon would have to drift for 100,000km to detect 1cfu!

Could a large volume pump be developed, for instance using bellows?

Alternatively, could the sampler area be massively increased?
For instance, imagine a Massive Array of Inexpensive Detectors, MAID - a ribbon 10cm by 1km perhaps. That's 10,000 times the surface area of the original 100cm2 detector concept.
Then you would only have to drift for 100m to detect one cfu!

Of course there may be organisms that are hard to culture but which can still be detected by RT-PCR for instance. But these calculations certainly set an upper pessimistic boundary of what effort is needed to biodetect at altitude.

How big a cavity is 10 million litres? Could you perhaps sample this big a volume using the interior of the balloon?
10 million litres is 10,000 cubic metres
4/3 pi (r)cubed
So 30000 = 4 pi (rA)cubed
So 7500 = pi (r)cubed
So 24000 = (r)cubed
So the cube root or 24000 = r in metres
This is a radius of about 29m
So, a balloon 58m across has a volume of 10 million litres
Is this too big? How much does the canopy weigh? I have no idea.
One could in theory fly the canopy with helium to altitude, then rapidly deflate it and fill it with ambient air. If it was fully sealed, the canopy would then rapidly descend and collapse upon itself since the exterior air would be getting much denser. You would be left with a crumpled canopy with a volume, at ground level, of about 100,000l - a precious sample of high-altitude air of a sufficient amount to possibly replicate the U2 experiments.

Please feel free to pick this apart/add references! I can just about believe the U2 results from this (so much air sampled along the transect) but now have doubts about any bioprospecting results claimed from high altitude balloons so far, unless they drifted for long distances (1000s km) before recovery. Both of these types of experiments involved culturing on the ground, so as always ground contamination, or contamination of the balloon before flight or by aerophiles in the troposphere, remains a possibility.
Our efforts to carry out experiments at altitude negate this BUT how do we sample enough air? Must we fly on a long duration balloon now?
One cfu literally means one single bacterium landed on the detector and was able to divide and grow into a visible colony. In turn this would literally mean only single copies of key genes to be detected by us by RT-PCR etc. In short, it doesn't matter how sensitive your assay is if the odd bacteria you are chasing in millions of litres doesn't happen upon your detector - otherwise there is nothing to detect!

Long duration? MAIDs? (Microfluidic?) Collapsed canopies? Any thoughts gratefully received!


1 comment: