The Nemesis Project

I’ve repeatedly heard that every academic, at some stage in their career, has that one project that just won’t behave itself and become a nice little publishable package. Experimental results lead to more questions than answers. Reviewers say the ides are interesting but are unconvinced by the conclusions or have issues in the reliability of the method. You're rejected but encouraged to re-submit. The whole thing drags on for years, quietly ticking away in the background, while research avenues with more promise for short-term gain are chased instead. But the nemesis project never dies, it stays there at the back of your head. Too much time and effort has already been invested, you’re in too deep to give up on it now.

My own mini-version of this has just ended. My nemesis project has just been published in Astrobiology (link, and link to non-paywalled pre-print) 3 years after my supervisor scribbled down a ‘cool idea which won’t take much time to test’ (I may be paraphrasing there, it was a long time ago). This is not my usual post-publication summary of my work, I will hopefully write that soon, instead this is a story of how much behind the scenes failure can have gone into one, small, relatively insignificant, successfully published paper.

The idea for the project came out of the PlanetaryProtection of the Outer Solar System project, which I have written about before (link). In one of the early meetings back in 2016 my supervisor, clearly paying full attention to whatever was being discussed at the time, scribbled a vague experimental idea onto a scrap of paper. This idea was to see if we could use a well-established environmental sampling technique (solid phase micro extraction, SPME) to test spacecraft hardware surfaces for organic contamination.  Now this was interesting as organic contamination is a big issue in planetary protection, we don’t want to send dirty spacecraft with highly sensitive instruments to the (currently) pristine icy moons of the outer solar system. We’d end up only detecting muck from Earth and so either getting all excited over nothing, misinterpreting it for evidence of alien life, or, a real interesting extra-terrestrial signal would be missed, lost in background noise from the contaminants. Current methods employed for detecting contaminants on surfaces tend to be time consuming, complicated and may involve multiple solvents being used in the process – themselves potential contaminants.

The premise was therefore simple: Get some stainless steel to use as a budget stand in for a spacecraft surface, contaminate it, see if SPME (coupled with GC-MS) is sensitive enough to detect contaminants at the levels of cleanliness required for life-detection missions.

The first version of this study just involved leaving some stainless steel L-shaped brackets (bought from a hardware store) out in the lab to collect fallout contamination from the air and also handling them with and without gloves to see if they picked up anything detectable from hand transfer.

To be scientifically valid a study like this must be reproducible, so many repetitions of everything being tested are needed, simple, but monotonous, time consuming work – perfect for an undergraduate summer internship! Georgios, a 2^nd year undergraduate and now co-author on the final paper, gave up 6 weeks of his summer in 2017 to do this, creating loads of data for me to work up afterwards. Now initially we thought this first version of the study was pretty good, however the reviewers had other thoughts.

Reject but encourage re-submission.

The issues basically boiled down to our method being a bit woolly and bullsh#t (again, paraphrasing). How could we know what was on the surface to detect and therefore how sensitive the method was if we hadn’t specifically contaminated it ourselves at a known concentration? We’d basically skipped the proof of concept stage and gone straight to real-word testing (well as real as you can get without a real spacecraft)...So yeah, fair enough.

Not having scope to dedicate 6 weeks of lab time to completely redo the experimental side of this study myself, the project had to get shelved until the following summer (2018) when I could get a second student, Yuting, who was keen to get some experience in the mind-numbing, soul destroying boredom of repetitive lab work.

In the meantime, I took the reviewer comments, which despite being rather critical were all very valid and helpful, and developed a whole new method for testing the sensitivity of this technique. This was to be much more scientific, creating a whole range of solutions of astrobiologically-relevant contaminants to contaminate a surface with much better-defined properties (although it was still basically just a steel nut).  

Once again, the student project seemed a success. Yuting produced a shed load of nice replicate data over the summer, which I turned into a completely new manuscript. None of the data set from Georgios’s original experimental run even made it into the new work, and after a few weeks of tidying up and writing we re-submitted.

Again, however, the reviewers didn’t quite agree, while they did think the method was now (mostly) sound, they didn’t agree the results were as promising as we did and wanted more and better data. Almost annoyingly this wasn’t a rejection this time, there was now a time pressure involved with a re-submission deadline. I could have ignored it and waited another year, but there was an end in sight, a way to kill this thing. Jon just needed to work some magic to tweak the mass spec settings to decrease the noise and make the data more convincing. Unfortunately, this meant I now had to repeat all the experimental work with the new settings myself, replicating a whole summer student project in about 3 weeks.

This was not fun, but it worked.

Now at the end of it all, it is clear that without the multiple knock backs and the intermediary time periods to just think about how to improve the methodology, this study would’ve been pretty rubbish. This is definitely a case where the review process has greatly improved an original idea and has shown me that rejections don’t always have to be a bad thing, they can be an opportunity. However, this only took 3 years to take down, I’m not sure how I’d feel if this had grown into some 5 or 10 year, or even career-spanning, monster.

Maybe that’ll be the next quick project…

Washing samples to detect 'Martian' organic matter

The beginning of this year seemed to be a good time for our group getting papers accepted and the third of those that snuck in is now online in Astrobiology:

Effects of Oxygen-Containing Salts on the Detection of Organic Biomarkers on Mars and in Terrestrial Analogue Soils. Wren Montgomery,Elizabeth A. Jaramillo, Samuel H. Royle, Samuel P. Kounaves, DirkSchulze-Makuch and Mark A. Sephton. (2019) Astrobiology

Once again, we’ve been looking at the troublesome effects of perchlorates on the detection of organic molecules, only this time we’ve used the Atacama Desert as a stand-in for Mars.

I’ve discussed the ‘perchlorate problem’ on Mars numerous times on this blog as it’s what I’ve been working on for the last couple of years (although not any more, watch this space). In short, oxygen-rich salts (of which perchlorate is probably the most problematic) are present in the Martian soil, we also expect there to be organic matter present (note, organic does not necessarily mean biological, see previous rant). With Mars lander missions, such as Curiosity, we attempt to detect the organic matter in the soil by heating the samples up in an oven to break down large molecules into smaller (more volatile) fragments that we can detect. However, those salts also break down when heated, releasing oxygen. This oxygen causes the organic matter to combust and any interesting organic molecules ‘burn up’ and are lost as carbon dioxide and carbon monoxide gases. This explains the difficulty that Mars missions have had in detecting organic matter on the surface, there has only been a single successful detection, which I’ll come back to later.

Mars today is a hyper-arid environment. While there may have been flowing rivers, lakes and even seas on the surface in the past, nowadays any liquid water anywhere near the surface is very rare. This allows the build-up of perchlorates and other salts which are highly soluble and would otherwise be washed away, which is why they are rare on Earth. This is where the Atacama Desert comes in as, recent floods notwithstanding, this is one of the driest places on Earth and so is one of the few places where perchlorates are present in the soil in significant amounts. This, combined with the low abundance of organic matter in the desert soil, due to its inhospitability to most life, makes the Atacama a (relatively) easy option for testing out ideas about Mars.

A rather Martian looking dawn in the Atacama Desert

The whole premise of this study can essentially be boiled down to: if these perchlorates are so soluble, can we just wash them out of our samples to allow us to detect the trace amounts of organic matter that we previously could not detect?

The answer, it turns out, is yes.

When the desert soil samples were initially analysed, by heating them in a similar fashion to what is carried out on Mars, showed little or no evidence of any organic matter being present.

Sub-samples of those soils were then well washed in very pure water, filtered and then dried. Unsurprisingly, analysis of the water showed that it had dissolved most of the soluble salts from the soils and it did not appear to have washed away any organic matter (which is mostly insoluble in water).

Once dried, analyse of these leached (washed) soil samples now allowed the detection of a variety of organic molecules. The molecules detected were indicative of the presence of cynobacteria (algae) that are known to be able to grow even in the dry desert.

This was, all-round, a pretty good result!

This potential for the problematic salts to be washed away has some pretty exciting implications for our search for organic matter on Mars. If we want to get around this ‘perchlorate problem’ we can either:

1. Wash our Martian samples with water. This, however, introduces a whole host of issues. Do we take water to Mars, it’s pretty heavy and we risk creating a nice, wet habitable environment for any Earth microbes that have hitched a ride, a major issue for planetary protection. Do we produce water on Mars by melting water-ice or extracting it from hydrated minerals, again, this would probably upset the mission’s Planetary Protection Officer (yes, this is a real job).

Buried ice exposed on steep slopes could be a useful water source

2. Go look for areas with evidence of ‘recent’ water activity on Mars were the salts will already have been leached away for us. This is the ‘easy’ option and what may have already happened accidentally. A rock unit called the lower Murray mudstone is the one place on Mars where evidence of complex organic matter has been found so far, co-incidentally this unit also has one of the lowest concentrations of perchlorate yet measured on Mars. There is evidence that, after the mudstones were deposited and buried, fluids flowed through the rock. These fluids could have leached away any soluble salts originally present, leaving being the insoluble organic matter, making it easier to detect. Areas with evidence of current or more recent water activity, such as above near-surface aquifers and near exposed and melting water-ice could also be promising areas to check out, however, these will also present planetary protection issues if there is liquid water available to support life.

Mineral veins show evidence of fluid flow, these fluids may have 'washed away' the soluble salts

This was actually a project that Wren had been trying to get published for a while now and the whole organic matter: perchlorate ratio paper we published last year actually originated as a response to reviewer’s comment to one of the early drafts of this current work. Happily, the two studies agree with each other, and NASA’S detection of organic matter which was announced while we were working on the re-write, pretty nicely (which is always good).

Scientists blew up a piece of Mars, you’ll be amazed what they found out!

Or: Indigenous Organic-Oxidized Fluid Interactions in the Tissint Mars Meteorite, Jaramillo EA, Royle, SH, Claire MW, Kounaves SP and Sephton MA. (2019). GRL

Apologies for the clickbait title, it seems as though this is now compulsory with all space-related scientific writing. I did blow up a piece of Mars though and I did find stuff out, you can read all about it in a paper I co-authored which is now published in GRL. Alternatively, here is a summary, it is massively biased towards the organic geochemistry side of things as that is the bit I did (and the only bit I really understand).

On the 18^th July 2011 a meteorite was observed exploding over the desert in Morocco. Over the following few months fragments were collected near the village of Tissint, which the meteorite has been named after. Very few meteorites have actually been found so soon after impact, usually they have sat around on Earth for many years. In this time they become contaminated, they sit in dirt, water flows through them and (Earth) microbes make them their home. Previous studies have shown that the Tissint meteorite is actually a piece of Mars; Martian igneous rocks that were formed around 600 million years ago were blasted off into space by a large meteorite impact on the surface of that planet around 1 million years ago. This means that the Tissint meteorite gives us a unique opportunity to explore the geochemical processes happening in the Martian crust, with the freshest samples we’re going to get until (if) Mars Sample Return happens.

the fragment of Tissint used in the study

Elizabeth, the lead author, had collected samples from the fall site and analysed them, along with fragments of the meteorite to compare their salt contents as part of her PhD. This had raised a few questions that needed answering, so I was drafted in to pyrolyze (i.e. flash heat/blow up) the same samples to see what organic molecules were present and what they could add to the story.

Tiny bits of Mars ready for me to grind up and pyrolyze

As I have mentioned before in my posts, organic does not equal biological. We were not looking for aliens here, organic molecules are just those molecules that contain carbon and hydrogen. While they are the ‘building blocks’ of life, they also form from non-biological process, including in space and in hot fluid (hydrothermal) systems deep underground. As such they are common in meteorites and comets and we expect to find them on Mars.

Looking at the organic content had 2 purposes: 

1. We wanted to know if the meteorite showed signs of becoming contaminated in the short period of time before it was found (biologically sourced contaminants from Earth-bugs finding their way into the samples would be pretty easy to spot);

2. If no signs of contamination then we hoped to identify actual Martian organic molecules and see what they could tell us about ancient Mars.

Neither the inorganic salt content nor the organic molecular compositions of Tissint suggested that the meteorite was contaminated (Yay!). While I detected evidence of microbial life in the soil samples, these hadn’t got into the meteorite. What I did detect in the meteorite were simple aromatic (ring-shaped) organic molecules, some of which contained sulphur. This was quite exciting as these were the same kinds of compounds that have been found in other Martian meteorites AND on the surface of Mars by the Curiosity Rover. Suggesting that we had detected actual Marian organic matter!

Organic molecules found on Mars (by Eigenbrode et al. (2018)), we also found various thiophenes, alkylbenzene, chlorobenzene and napthalene in Tissint, along with other sulphur-bearing compounds 

The mixture of salts and organic molecules within Tissint suggest that less than 600 million years ago, recent in Mars-terms, an oxygen-rich, salty water-brine flowed through the near-surface crustal rocks of Mars. Electrochemical reactions likely formed both the organic molecules and the salts in this fluid as reactions occurred on the surfaces of variously charged minerals.

As the composition of this fluid was similar to seawater and, there was a readily available source of organic carbon as a food source, this could have produced a temporarily habitable environment near the surface of Mars during the late Amazonian period. This is much later than the surface of Mars was potentially habitable and although we don't go as far as saying anything actually lived in it, it could have done....