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[Music]

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where's Michael Morrison I'm kind of

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gonna go back to it doctor to Lucas was

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kind of talking about at lunch with

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doing experiments that we kind of do on

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the ground but in a low orbit

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environment to kind of see how this

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microgravity environment affects what

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we're doing however instead of doing

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protein crystals I'm actually looking at

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how bacteria grow and adapt in this

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unique environment that can't be found

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on earth I'm instead of some background

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on how life adapts to space probably the

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best studied organism that we know of

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for her space are humans we've had a

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continuous presence in space for the

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last 70 and a half years and that's just

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if you consider the International Space

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Station before that there was a Space

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Station mer Skylab Apollo Vostok and so

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we've had about sixty years of studying

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how humans adapt to spaceflight and what

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we found as he pointed out was that

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astronauts lose bone density and muscle

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mass in space but they also experience

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this fluid redistribution because we've

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evolved with gravity constantly pulling

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down on our bodies so we have mechanisms

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mechanisms in our bodies to help push

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this fluid back up to kind of keep our

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blood circulating and everything kind of

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working in a nice harmonious manner and

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this dysregulation we are fluids when

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you get into space because gravity is no

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longer pointing down you can stress the

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body out which can cause this regulation

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of the immune system which has been seen

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in astronauts coming back from space as

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flare-ups of different dormant viruses

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such as epstein-barr and herpes and

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these astronauts we send up there are

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you know very active healthy individuals

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that we send up there for example right

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this photo right here is of this Oh what

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is it click okay well little the top

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button okay so this is a astronaut tim

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peake and this photo he was competing in

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the London Marathon where he actually

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set the record for the fastest off world

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marathon time at 3 at 3 hours and 35

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minutes and 12 seconds and so if we're

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you know seeing these effects and

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astronauts who are really physically fit

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what would we see in it were humans

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myself who a marathon would probably

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kill so this is kind of astronauts kind

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of idea but you know these are macro you

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know big organisms my focus is more on

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the prokaryotes because we see keep

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sending humans to space but prokaryotes

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go with us everywhere we go they're

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central for our immune system our

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digestion and does it affect these

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prokaryotes and it actually does what

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we've observed is that prokaryotes have

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faster growth rates in space and it can

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reach fine cell density higher final

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cell densities they some experiments

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also suggest that antibiotic resistance

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increases in space as well as appearance

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and there's not things you want if the

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immune system becomes dysregulated big

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concern for NASA for sending astronauts

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past low-earth orbit because if you send

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them out there they're halfway to Mars

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they get sick you can't bring them back

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you said we have to figure this out kind

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of thing and last what they what kind of

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prokaryotes is this alteration and

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biofilm formation biofilms do have some

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role in chronic infections and

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antibiotic resistance but NASA's noticed

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that they affect the systems on the

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space station as well such as the space

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station mer it actually clogged the

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water reclaiming system with a biofilm

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and they had it completely read change

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out the entire system and also they find

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biofilms on the surface of this capsules

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which can actually corrode material and

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that's not something you want again

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pathway to space all sudden your machine

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you're starting to corrode what you can

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do about it

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so this is something that NASA is very

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interested in and that's kind of what my

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lab wants to look into kind of figuring

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out what's going on here and some kind

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of molecular like responses we can kind

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of predict and kind of treat so as the

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Lucas kind of point out spaceflight

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hardware is important so for us we use

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the biological research in canisters or

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brick space flight hardware they contain

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the petri dish that we air deposited

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vasila settler spores which are a soil

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bacteria that are non pathogenic know

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which astronauts whatsoever and the

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spores are dormant and so when we send

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them up we can inject media into them

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they become you know viable growing

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cells and so we can affect of what they

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do just in space without having to worry

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about the launch and the return and down

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here there's an actual chamber for the

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media that can be stored in until we're

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ready to inoculate the petri dish and we

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used a rich media TS

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that has protein and glucose as carbon

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sources as well as about twenty ten

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percent glycerol so we could actually

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freeze our samples and bring them back

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to earth and they would still be in this

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kind of state of hey we're still in

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space even though they're on the ground

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white when we follow them and so the

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actual timeline for our mission which

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became known as Brook 21 because it was

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the 21st brick mission launched in April

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2015 dr. the National Space Station they

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inoculated the samples in space right

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here and they roiled to grow at ambient

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space station temperature approximately

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23 degrees Celsius for about 25 hours

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after which they were frozen in minus 80

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degrees and shipped back to earth where

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you know me and my lab were able to get

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our hands on and I saw actually

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processing the samples and because space

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is limited we wanted to do a lot with a

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little

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and so this is kind of everything we

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wanted to do with them and you can kind

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of see we've kind of looked at the

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antibiotic profiles and the mutation

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rates and everything I'm going to focus

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mostly on the transcriptome how they're

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actually adapting today but I guess it's

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a little late but Josh Lee had a student

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in our lab was actually giving a poster

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on this yesterday

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pretty sure if you still are interested

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in that Josh Ling Han would be happy to

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talk to you about mutation frequencies

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in space so they're my little plug for

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him but from here on I'm mostly talking

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about the transcriptome and how the gene

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expression is affected by space so how

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do we do transcriptomics we get the

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samples back they're frozen we thaw them

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in the lab and we immediately extract

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the RNA so we have all the RNA that was

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up in space before it's degraded as well

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as the ground samples that we flew are

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that we didn't fly that we're sitting at

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NASA while the ground all the

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spaceflight samples were flying and we

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then sequenced that RNA to get it you

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know figure out what the actual code is

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and then we can do differential

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expression analysis on those sequences

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so differential expression analysis I

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take the reads and I filter them for

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quality making sure that you know we

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actually have good reads or reads aren't

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you know messed up or anything and then

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I line them to the genome and every

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single sample we had had about 50

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million reads per sample and we had in

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equals 3 for Brick 21 so that's total of

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6 samples 3 per condition and we use a

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machine to do this because I

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you want to graduate eventually and not

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have film and then once we align them to

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the genome we count how many sequences

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are there for every single gene so we

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can get kind of a quantitative number of

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how much was this gene being expressed

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in space compare and you know compared

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to on the ground and then we statistics

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we use actually two packages Lima into

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ec2 to determine if it's you know higher

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or lower in space compared to ground in

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order for a gene to be consider

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differentially expressed it had to have

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at least a two-fold change in the number

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of counts and between flight and ground

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with a p-value of point zero one and

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since we were looking about four

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thousand four hundred transcripts we did

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correct from local testing bias using

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Benjamin Hochberg and all that stuff and

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so we're pretty confident with our you

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know genes we get and then we do

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functional analysis to kind of figure

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out what are those genes actually doing

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and do they kind of group into different

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pathways so we can kind of try to target

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pathways not just individual genes so we

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did this on Brick 21 and we found 293

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genes that were differentially expressed

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between flight and ground so there's

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about 177 that were higher in space

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flight and about 116 that were higher in

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the ground samples and from this we did

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you know our functional annotation try

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figure out what was going on and we

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found about 10 enriched KEGG pathways

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KEGG pathways are just kind of submitted

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but pathways such as biotin metabolism

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arginine we actually had a couple of

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non-ribosomal peptide biosynthesis

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pathways going on and it's an actually

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metabolism just to kinda name a few but

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while we were analyzing this data nasa

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was generous enough to let our lab kind

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of participate on a second space flight

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experiment which became those brook 23

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where we flew the exact same hardware

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exact same strain exact same media and

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this was very important because to this

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point no transcriptome for bacteria has

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ever been duplicated in space so this

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was kind of a nice idea to say can we

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see the same results if we duplicate the

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sensation of the gold standard of

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science and however adhering the kind of

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development and planning stage instead

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of rowing it for 20-25 hours we they

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grew these for 26 hours that's the only

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difference between these two we actually

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wanted to look at stationary phase which

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the first one for an exponential phase

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to kind of see like differences in fit

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growth phases but if there's an effect

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by spaceflight we should still see genes

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up regardless of what

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Faye's it's in so can we kind of use

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these two different phases to pull out

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genes that spaceflight effects

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continuously and so these flew about a

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year after the first ones and then so we

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did you know transcriptome profiling the

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same way as the first one and I love

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this photo because these are actually

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our samples in space and they're

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floating I love looking at there's no

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strings attached and so we found about

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255 genes in the 23 samples that were

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differentially expressed so feel a few

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less than brick 20 where I'm about the

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same number and we found about seven and

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rich pathways

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among these 255 genes this is all great

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and everything but you know how do they

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compare to one another that was the you

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know the big question and so I want to

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know what the variation between the

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different samples were you know do are

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their samples clustered together are

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they completely over the map and so I

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ran a principal component analysis and

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what you can kind of see is if you look

275
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at the first principal component you see

276
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that the sample is kind of clustered

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between brick 21 and brick 23 and have

278
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about 51 percent of variation between

279
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all the samples and because we grew them

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at different state growth phases this

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isn't uncommon because they should have

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completely different gene expressions

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when they're you know rapidly growing an

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exponential phase that's when they're

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dormant in stationary phase but if you

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look over at the second principal

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component you see that about 21 percent

288
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of the variants between all the genes

289
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can be explained by grouping the ground

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samples together and the flight samples

291
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together so we do see this effect of

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spaceflight even though our examples

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were going to this different face so we

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can actually pull out maybe some

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effective spaceflight even though it's

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it's not the dominant source of our

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variants and so we then want to see can

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we actually see this at the actual gene

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level do we have new genes that are

300
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differentially expressed in both

301
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experiments and we found 91 genes that

302
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were differentially expressed in both

303
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experiments so yeah about put and so 55

304
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of these genes were upregulated in

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spaceflight which include our biotin

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metabolism biotin uptake genes as well

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as our biofilm biosynthesis genes as I

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mentioned before you know NASA is very

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interested in these biofilm by since the

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scheme and then we found 36 that

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regulating the ground control and these

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are national metabolism into component

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systems an actually metabolism is a bit

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of a misleading because it was really

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just the nitrate reductase genes which

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are expressed in low oxygen conditions

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and this is we also saw some other

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fermentation genes that are also

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expressed in low oxygen conditions so we

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think most of the genes it looks like

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most the genes that are down regulated

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were due to maybe a difference in oxygen

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between the flight and ground sample but

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the flight samples going back to those

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you know we didn't see this biofilm

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bison for this gene which we're kind of

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like wanting to dive in a bit more into

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like okay we see them up masses kind of

329
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interest in these you know what do we

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actually see with our gene counts and I

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know probably most of you do not

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understand what these four little

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radiations are so I will be I will kind

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of dive into that all these genes are

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the genes required to make biofilms in

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bacillus unless the organism that we're

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using so any available that aren't

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familiar with biofilms the most simple

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and probably when you associate with the

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most is plaque on your teeth it's a

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community of cells that kind of grouped

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together in a protein sugar matrix that

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kind of shared nutrients kind of help

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each other out and this protects them

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from invaders antibiotics and a bunch of

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other environmental stresses so you know

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bacteria love to grow in these biofilms

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and they're made up of like I said EXO

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polysaccharide or EPS sugars there's a

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biofilm surface layer protein and then

351
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you also have a protein scaffold that

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kind of holds them together as well

353
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within the sugar matrix so we have all

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of our genes here and if you look at you

355
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know this cut off of it this is a log to

356
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scale so this is a two-fold change

357
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there's only two genes that don't have a

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two-fold change between fly and grounds

359
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so almost all of our biofilm

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biosynthesis genes are upregulated in

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spaceflight and so this is the first

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molecular evidence that we have that

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points towards spaceflight you know

364
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promoting biofilm biosynthesis now we

365
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don't have a you know reason why this is

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right now but you know something that

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we're hoping to look into in the future

368
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with more spaceflight missions you know

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and more regular tests on regulations so

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it's kind of with that in the conclusion

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we

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fine about 91 genes that are

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differentially expressed and these 91

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genes we're gonna look further into

375
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because these could be good candidates

376
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for how space is affecting bacteria like

377
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what are these you know we'd knock out

378
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these 91 genes one of them how does it

379
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affect space why does it have a effect

380
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at all you know kind of dive in further

381
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also look further into biofilm

382
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biosynthesis we do see the first

383
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molecular kind of evidence for this

384
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increased production in space and it

385
00:14:00,430 --> 00:13:58,639
appears that at least for the hardware

386
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we're using the available of nitrogen

387
00:14:04,090 --> 00:14:02,930
oxygen is different between flight and

388
00:14:06,400 --> 00:14:04,100
ground and that's something that we are

389
00:14:10,389 --> 00:14:06,410
discussing with NASA on kind of making

390
00:14:12,280 --> 00:14:10,399
adjustments to the hardware for that but

391
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you know science is in a one-man show so

392
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I have a lot of people I want to thank

393
00:14:18,430 --> 00:14:17,029
primarily my boss and Patricia Fajardo

394
00:14:20,079 --> 00:14:18,440
who actually helped with a lot of

395
00:14:21,220 --> 00:14:20,089
processing and integration the

396
00:14:23,650 --> 00:14:21,230
integration of the spaceflight

397
00:14:25,960 --> 00:14:23,660
experiments the BRIC 21 BRIC 23 teams

398
00:14:27,400 --> 00:14:25,970
who you know helped integration with the

399
00:14:29,350 --> 00:14:27,410
preparing of the samples and everything

400
00:14:31,810 --> 00:14:29,360
and the astronauts who actually ran my

401
00:14:35,650 --> 00:14:31,820
experiments in space that you know they

402
00:14:39,100 --> 00:14:35,660
wouldn't let me do and so with that

403
00:14:43,060 --> 00:14:39,110
oh and NASA for funding me and and with

404
00:14:47,620 --> 00:14:46,139
[Applause]

405
00:14:57,340 --> 00:14:47,630
thank you Michael

406
00:15:04,549 --> 00:15:02,569
do you have ideas about why biotin is

407
00:15:05,749 --> 00:15:04,559
upregulated is it a general stress

408
00:15:07,939 --> 00:15:05,759
response do you think there's a certain

409
00:15:10,609 --> 00:15:07,949
protein that is a cofactor that's

410
00:15:12,169 --> 00:15:10,619
hindered in some way like so yeah we're

411
00:15:14,389 --> 00:15:12,179
working fused about that little bit we

412
00:15:16,609 --> 00:15:14,399
don't know for sure because it's you

413
00:15:19,850 --> 00:15:16,619
it's a B vitamin it's used in the

414
00:15:22,100 --> 00:15:19,860
biosynthesis of some fatty acids and

415
00:15:24,169 --> 00:15:22,110
branched chain amino acids and buccaneer

416
00:15:25,729 --> 00:15:24,179
genesis but we looked at all the genes

417
00:15:27,710 --> 00:15:25,739
involved in those pathways and none of

418
00:15:35,780 --> 00:15:27,720
more differentially expressed so we have

419
00:15:38,660 --> 00:15:35,790
no clue why they're up regarding the

420
00:15:40,609 --> 00:15:38,670
about film production do you think that

421
00:15:43,160 --> 00:15:40,619
a my absolutely to do with form sensing

422
00:15:45,859 --> 00:15:43,170
signal regulation with volatility in the

423
00:15:48,259 --> 00:15:45,869
og so there is some thought about

424
00:15:50,509 --> 00:15:48,269
motility in space we didn't see any

425
00:15:52,639 --> 00:15:50,519
motility genes upregulated but we did

426
00:15:55,160 --> 00:15:52,649
see a quorum sensing gene surfactant

427
00:15:57,499 --> 00:15:55,170
that was up regulated in space so that

428
00:15:59,030 --> 00:15:57,509
probably one reason why we're seeing the

429
00:16:02,539 --> 00:15:59,040
increase in point of information but we

430
00:16:04,069 --> 00:16:02,549
don't necessarily know you know why that

431
00:16:06,309 --> 00:16:04,079
coursing gene would be higher in space

432
00:16:09,019 --> 00:16:06,319
but we did see one Qantas engine

433
00:16:11,970 --> 00:16:09,029
okay let's thank Michael again I don't