We got a Christmas present from listener Anna: a small plastic tube full of dead flies. They've recently been infesting the hospital where she works. She wants us to figure out what they are, and what caused the infestation. Can DNA crack the case? Plus, the return of Gins & Genes...
In this episode
01:12 - Will it sequence: a tube of flies
Will it sequence: a tube of flies
Ed Farnell, Illumina
With the gauntlet thrown down, Phil Sansom went to visit gene company Illumina. He brought the tube full of dead flies to scientist Ed Farnell - someone who originally worked on a Naked Scientists project back in 2017...
Ed - I think it was during the middle of the horse lasagna crisis, I think we're going to call it. Wouldn't it be cool if we took a sausage, which is something that traditionally, you know, it's harder to tell what's gone into that because everything's minced up and we sequenced it and we had a look to see what species we could find in there.
Phil - Did you find horse?
Ed - We didn't find any horse. No, but we found all the things you might expect to find in a butcher shop. So we found some lamb, we found some chicken, we found some beef. And when we looked really, really hard, we even managed to find some human DNA as well, probably from the butcher who was preparing the sausages.
Phil - Well, let me present you with your new sequencing task. And I've got to say it is less nice than a sausage.
Ed - Yeah, go on.
Phil - Alright, here it comes.
Ed - Oh man. Okay. Yeah, no, that is actually quite disgusting. So I kind of made a bold statement that I might not find it that gross, but these are some really, really big flies.
Phil - Isn't it gross?
Ed - Yeah. They're pretty hairy. They look pretty, pretty grim.
Phil - Have you ever worked with anything like this before?
Ed - With whole flies? No. So to take it from the fly itself is really interesting.
Phil - Is it going to be tricky?
Ed - Yeah. So they're going to present a couple of challenges that we've been thinking about. One thing is how are we going to detect the bacteria and how are we going to detect the bacteria against the host? What most people will do is they will smear the flies all over some agar plates and they’ll grow the bacteria. The problem with that is this sort of culture can introduce a bias whereby only certain types of bacteria will grow on certain growth media and actually there's a smarter method we can use now with sequencing. So we're going to try two different experiments with these flies. I think with some of them, what we'll try doing is just mashing up the entire fly. Then we'll get all of the fly DNA and all of the bacteria that's in the fly: on the outside of the fly, on the inside of the fly, in its mouth parts, in its stomach.
Phil - You're going to get a lot of fly.
Ed - We're going to get an awful lot of fly and also bear in mind the fly and the bacteria will be mixed all up together. So this would be a whole metagenomic sequencing. And the way we combat having a lot of fly is by doing a lot of sequencing.
Phil - And what's metagenomic?
Ed - Metagenomic. So that's looking at the bacterial population, the whole bacterial population within the fly.
Phil - Oh, so not just the fly, the extra stuff there?
Ed - It's looking at the extra stuff. So what we can do is we can extract all that DNA,, say "Oh this matches to fly, we'll get rid of that. We're not interested in it." Everything that's left, we can take that and start aligning it to bacterial and viral genomes and see what we get.
Phil - What's the risk that you just get overwhelmed by fly, fly, fly, and you never get bacteria?
Ed - So there is some risk, but I say we can sequence very, very deeply and you'd be surprised how much bacteria there are in things. Even in humans, like for example, if you've ever done one of the tests like 23 and me or ancestry where you do a spit tube, those spit tubes can be almost 50% bacteria from the inside of your mouth.
Phil - Okay. You said first option was mash up the flies. Second option?
Ed - So the second option is just to dunk the flies in a special chemical that is normally part of the DNA extraction procedure and we're just going to leave them there and dissolve basically everything on the outside of them and hopefully not too much fly. So I'm not sure how that's going to work out. This is the bit where it gets a bit experimental because we don't really know how long we're going to have to leave them in there so we don't dissolve too much fly. But we do still collect everything that's on the outside of them and then we just proceed with the rest of our DNA extraction from there to get hopefully what is most of the things on the surface of the fly.
Phil - Perfect.
Ed - And if both of them work the nice thing here is we'll have a nice comparison. It's like is the bacteria population on the outside of the fly different to the bacterial population on the whole fly? So you know, we get an idea of maybe there's something different in the stomach to the outside.
Phil - Oh, that's interesting.
Ed - I just thought about it now, yeah, we will have a kind of differential. I mean it's not perfect. I'm sure those of you listening will be able to pick lots of holes in the idea, but I think we'll be able to do something a little bit more interesting with that. Maybe, depending on how it works out. I mean, it's science. We're trying this for the first time, so we'll see how it works out.
Phil - Other parts of the challenge that we were originally given by our contact at Addenbrooke's: these flies probably came from something that died in the eaves, some sort of horrible squirrel or possum or something...
Ed - That just got a magnitude more disgusting but carry on.
Phil - But do you think you could find that based on what's in the flies' stomachs?
Ed - That's a really good question. I don't know enough about fly biology to know how old these flies are, what might still be left in their stomachs, how recently the flies came from the eaves. I mean we can but look and see what happens.
Phil - I'd be really excited if you found squirrel in there. I mean, disgusted.
Ed - Disgusted and excited! Yeah, that would be a unique challenge, but we'll give it a go.
Phil - Before I left, Ed put the tube of flies in Illumina’s freezer. The one that looks like a bank safe, and keeps things inside around minus eighty degrees C. And that was it. For two very festive weeks I left them in Ed’s capable hands.
06:36 - Fly contain multitudes: a hidden world
Fly contain multitudes: a hidden world
Zenobia Lewis, University of Liverpool
Do flies really have even smaller bugs living on them, let alone inside them? Zen Lewis is a scientist at the University of Liverpool who researches what's called their microbiota - and she's found that it has strange, unexpected effects on the way they behave. Phil Sansom found out more...
Zen - Definitions in this field are still somewhat contentious, but for me, the microbiome represents the genetic material comprising all of the microbes that are found on animals. It's somewhat different from what is also known as the microbiota. So that's the actual organisms themselves, the ecosystem of microbes, for example, on an animal.
Phil - When it comes to flies, they're very small already. Do they have little microbes living on them as well?
Zen - They do on and inside them. Microbes are found on pretty much everything. If you think about humans, some data came out a few years ago that suggested that there are 10 times as many microbial cells on a human compared to actual human cells. So we really are made of microbes in a way, and the fly is no different.
Phil - What organisms have flies got on them and inside them?
Zen - We've known for some decades now, from the 70s, that many, many insect species including flies, have what's called endosymbionts. Those are microbes that are found actually within the cells of the host species and they're found in flies too. More recently, we've started to examine how what's called commensal bacteria, microbes in the gut, for example, can impact on the host fly. Some bacteria is beneficial in terms of things like digestive processes, for example.
Phil - So are they breaking down food that the flies' bodies can't by themselves?
Zen - Well, let me give you an example. So something myself and my collaborators, Anne Lizé and Chloe Heys, have been working on recently is a fruit fly called Drosophila sechellia that is only found in the Seychelle islands. And it's evolved to be able to feed on a tropical fruit called Morinda or Noni. Now Noni is actually super, super toxic and pretty much kills anything else that goes near it. And yet this fly has evolved to be able to break down, to use this, this food source. And we've got some preliminary data which suggests that the microbes that these flies have in the gut are what in evolutionary history would have allowed this fly to adapt to this novel poisonous food source.
Phil - Wow. So what generally controls what kinds of bacteria are in there? Is it usually stuff like that, where it's some part of their evolution back in the past?
Zen - Yes. Evolutionary past certainly. Organisms also accrue microbes from the food that they eat, fecal matter that kind of is deposited with the egg as it leaves the female reproductive tract, and also in some species, microbes are literally transferred between individuals. So in some species of termites, older individuals will, in a nutshell, kind of feed younger individuals with the contents of their guts, which then allows them to be able to break down wood I think, I'm not a termite expert, which they wouldn't be able to do so unless they received those microbes from their nest mates.
Phil - Just to jump in here - and how do they receive those microbes from their nest-mates? Butt to mouth, my friend. Butt to mouth.
Phil - Insects are disgusting.
Zen - But they're also very, very cool.
Phil - So does that mean with all these different ways of getting microbes, there are thousands that you might find in one fly?
Zen - Many insects, including flies, actually have quite simple microbial ecosystems. So they tend to be characterised by two, three, maybe four main groups of microbes. And you tend to see similar groups of microbial organisms in the fly.
Phil - If the flies don't have the right composition or if they don't have gut bacteria at all, do weird things start to happen?
Zen - They do indeed. Some of my own work we've been examining kin recognition in fruit flies.
Phil - This is kin, like you tell who your brothers and sisters are?
Zen - Yeah, exactly. In the fly that we've been working with, melanogaster, they're unable to tell if another individual is closely related to them or not, and that's a bit dangerous because, you know, if you mate with a brother or sister, you're potentially going to have poor quality offspring. However, if we disrupt the gut bacteria, which we do very simply by lacing the food with antibiotics, the flies then become able to recognize kin.
Phil - That is so strange.
Zen - It's very, very weird.
Phil - What do you think is going on?
Zen - Well, what we think is that because in flies they have this kind of scent signal, you know, like pheromones in a way basically, that signal is produced by these small organs called oenocytes. Now the functioning of the oenocyte operates via cues that come from the gut. So we think that by disturbing the bacteria, knocking out the bacteria, we're changing the action of the gut, which in turn changes the action of the oenocytes which are producing the scent signal. And so flies are giving off different signals compared to if they were normal.
Phil - Is this generally the kind of thing that people find when they look at these microbiota, that they have these weird effects that you wouldn't expect at first glance that are actually quite important?
Zen - That's what we're increasingly finding. I mean previously, a lot of research that was done about microbes was from a pathogenic perspective. So microbes causing disease and it's only now that we're starting to see that actually they have much, much wider impacts on their host's behaviour, biology and indeed evolution.
14:45 - Gins & Genes: to catch a carcass
Gins & Genes: to catch a carcass
Ed Farnell, Illumina; Eva Higginbotham, University of Cambridge; Matt Routledge, Addenbrookes Hospital; Will Lowe, Cambridge Distillery
A listener has - kindly - sent in somewhere between ten and twenty dead flies. She's challenged us to do a high-tech fly autopsy: to sequence whatever DNA we could find, and figure out what the flies are. And what were they eating before they died? The results are in - and it's time for Gins & Genes...
Eva - Hi, I'm Eva Higginbotham and I'm a PhD student at the Department of Zoology.
Ed - Hi, I'm Ed Farnell. I'm a product support scientist at Illumina.
Matt - Hi there, I'm Matt Routledge. I'm an infectious disease doctor at Addenbrooke's hospital.
Phil - Welcome everybody. It's great to have you here for Gins & Genes. Will, what are we going to be drinking today?
Will - Today we're going to have a look at anty gin, which is a gin literally made from ants.
Phil - From ants.
Will - Yes. Certain ants have certain flavours about them. It's the formic acid that these ants use to both defend themselves and communicate, has a sort of lemongrassy flavour for us. I'll let you taste it and see what you think.
Eva - Ooh, yeah.
Ed - Okay. Yeah, yeah. Lemongrass, I guess.
Matt - Yeah. Wow. Yeah.
Phil - I don't like it.
Will - Well look, four out of five's not bad. So why don't you try it with the tonic as well?
Phil - Bottoms up everyone. So the reason we're all here: Matt, you work in the lab that had these flies, right?
Matt - Yeah. So I've got to admit, I've got slightly a vested interest in finding out what potentially I may have been inadvertently exposed to by these flies.
Phil - Was it a big infestation?
Matt - It wasn't as if we were batting them off all the time, but there were plenty. We have no idea why they got in. We don't know where they came from.
Phil - Okay. The field is open. Ed, we gave them to you. What did you do next?
Ed - I took one fly and basically dunked it in an extraction buffer to get the DNA out, but tried not to disturb the fly too much so we would just get the things on the outside. And then with another fly I mashed it up really well, which is quite disgusting, and then dissolved that all in lysis buffer. So we have this comparison of potentially what's in the fly and what's on the fly.
Phil - Did either of them work?
Ed - We were really lucky, I guess. Both of our extractions worked on the first attempt. One of the first interesting things is, we thought, we'd compare our different extraction methods and see what we have. So for the exterior-only fly, our fly that we dunked in the buffer, we found that was actually enriched for an epibiont signature - which is a word I did not know until today - but that means basically things that are on the outside of stuff. Which was really, really good! We found a signature for the outside of things on the outside of our fly. And we also find it didn't contain an insect intestinal signature. So we managed also not to get the outsides of our fly into our exterior extraction. And on the opposite side we found that the whole-mash fly had both the epibiont and the intestinal insect signatures.
Eva - If you were specifically looking at just gut bacteria, then the ideal scenario would be of course to dissect that out of many, many flies, and your hands would get really tired, and you would hate yourself by the end of it. But you'd want to dissect them all out and then just run your experiments on that and your extractions on that.
Ed - I found that my 15-year-old dissection skills weren't quite up to the job,
Eva - But it is really promising that you actually found your controls worked, right. You found that there were species of bacteria that should be on the outside, and they were there, and there are species of bacteria should be on the inside, and they were also there. So that's great.
Phil - So what fly did we give you?
Ed - So the flies that we had were Calliphora vicina. Common name for it is the urban bluebottle blowfly.
Phil - Calliphora vicina. I put the name to Zen Lewis from the University of Liverpool.
Zen - I don't know much about that, but I think it's one of the blowflies and I believe it's one of the species that's used in forensics to work out how long it's been since a body became a body. I'm not sure that anyone has examined what the core microbiota is, but I have read some work which suggested that because the flies feed on cadavers, they actually pick up bacteria from the cadavers that they eat.
Phil - This episode doesn’t get any less disgusting, does it? But at least this is promising news. If the flies pick up the things they eat, can the DNA sequencing figure out their last meal? Back to Gins & Genes.
Ed - When we compared the whole fly and the exterior fly samples, we actually found that chicken DNA and a species of fish were enriched in the sample that had come from the mashed up fly rather than the exterior, which might point us towards what they'd been feeding on.
Matt - Okay, right. So at least now we know what to go and look for. Probably some chicken wing or fish finger or something behind the radiator somewhere.
Phil - Who brings in chicken wings or fish fingers for packed lunch?
Matt - I don't know. I mean...
Phil - Is it you?
Matt - It's not me! No, no.
Phil - How big was the trace of chicken and fish in there?
Ed - So it wasn't very high. And I think this sort of comes around to some of the caveats of our experiment. So we would have needed another the fly that we'd taken from a completely different environment, and have also extracted that, and that would have controlled for any DNA that was in our environment when we were doing the extraction in our labs, or indeed just giving us a background of what's normal in a fly.
Phil - Plus do you know how long ago necessarily the fly would have had to have eaten the fish or chicken?
Eva - The flies that I know about are actually a little bit different from the flies we're talking about here. So I work on Drosophila melanogaster, which is a fruit fly, and they're quite a lot smaller I think than the blowflies you're talking about. But if fruit flies are anything to go by, they actually have a quite complex digestive process. Their stomach is split into a midgut, a hind gut, and a foregut; but the midgut itself is split into five sections as well, and actually each of those have their own version of a microbiome and bacterial signature. I don't know if that means that if a fly ate a little bit of chicken, it's going to hang around for a long time; but it does go to show that actually we think of these insects as being really, really simple, but they have in some ways a more complex digestive process than we do.
Phil - Did you find anything else in there that might've been hiding or living inside their digestive system?
Ed - We were actually really surprised that there weren't more bacteria. We kind of expected flies... you think of them as being fairly disgusting, living in rubbish dumps; actually the proportion of bacterial DNA was really, really low within the samples. But we did find a couple... one of the most abundant bacteria found was actually Cryptosporidium muris. That can cause diarrhoea in immune-compromised individuals. So that's really not nice, and was there in actually a fairly decent quantity.
Matt - In clinical practice, Cryptosporidium we generally think of as a gut pathogen, and we quite commonly pick it up when we take stool samples from patients. And in the main it doesn't cause anyone any significant harm. It's only usually patients who've got HIV, or they're on kind of immunosuppressive therapies, that we worry about. Cryptosporidium.
Phil - So this might be something common to a lot of flies that live around people?
Matt - Yeah, I would have thought so. I mean, it's probably quite common for most people to have some Cryptosporidium infection at some point in their lives. And for most people it's pretty probably asymptomatic, and they don't notice any symptoms.
Phil - Okay. What's next?
Ed - We then also went looking for some familiar-sounding bacteria, things that we might know of and be more familiar with. So we did find some Salmonella in there, which we all know can be quite unpleasant causes vomiting. And we also found Pseudomonas aeruginosa, which is a really horrible bacteria that can affect wounds, and it's really unpleasant.
Phil - What do you say to that Matt? Are those bacteria that you're familiar with, and what do they do?
Matt - Salmonella is probably the one that most people have heard of. And there's lots of different types of Salmonella. The most common thing that Salmonella causes is like a gastroenteritis, particularly associated with eating under-cooked eggs. People think that the actual egg itself is infected with the Salmonella, but in actual fact the Salmonella sits on the outside of the egg; because as I'm sure some of your listeners are aware, birds have a shared cloaca, where both their GI tract and their reproductive tract drain into. So when the chicken's passing the egg it gets contaminated with the Salmonella that lives in the gut, and so the Salmonella is all sitting on the outside of the egg. And so the moment you crack it, you inoculate the egg with the Salmonella. And if it's not cooked properly, you eat it, it gives you gastroenteritis. Pseudomonas aeruginosa, so that's an interesting bug. It forms biofilms on things, so it's incredibly sticky. It's a big problem in intensive care units. It's also a bug that's incredibly naturally resistant to lots of commonly used antibiotics. So I'm slightly worried that I may have been exposed to now Salmonella and Pseudomonas aeruginosa.
Phil - Anything else from the results?
Ed - We did a bit of a sanity check and we went down out list looking at things that we thought sounded reasonable, or that we recognised the names of. And actually what we found was: the levels of these two bacteria in the flies were lower than bald eagle, wolf, and minke whale. So we suspect that there could well be some false positives in there.
Phil - What has this fly been eating!
Ed - So this is a bit of a cautionary tale about using metagenomics. You're always... the method uses some reasonably complex bioinformatics to try and fit things, you're looking at very small fragments of DNA and trying to align them against wider genomes. And you want to do it with enough stringency so you don't call too many false positives, weird things like bald eagle and minke whale, but without causing, you know, without losing too much data. So I think this is a reflection of how far we've got down the list, you know, before we get to something weird.
Phil - Right. Good sequencing job. What are you expecting next Ed?
Ed - I have no idea. I think it'd be really interesting to know what else people would like to see sequenced. We have these sequencets waiting for experiments to be done. So let's see what we can find.
Phil - Right, good job everyone. Your work is done. Let's have a drink.
24:36 - What is dark chocolate sneezing?
What is dark chocolate sneezing?
Listener Amy asks this question, and Eva Higginbotham has been looking into it...
Eva - So I've had a little read around to see what I can find out about this dark chocolate sneezing, and it's actually a version of something called the photic sneeze reflex, where you walk outside from a dark room into a light room and you immediately sneeze uncontrollably, between one and sometimes up to forty sneezes. Now the interesting thing is, the dark chocolate sneezing is a type of photic sneeze reflex, despite the fact that it's not actually caused by photons, that's where the 'photic' comes from. It actually gets put into kind of a catch-all term, photic sneeze reflex, to mean the light sneezing, or some people have sneezing caused by eating things that are spicy, or things like mint or grapefruit. Basically what's happening is your body is having a reaction to a strong stimulus.
There are lots of different theories about why it happens, but essentially it's a different kind of sneeze, right? Because a normal sneeze happens because you have some sort of irritation in your nose, and this is more just a nerve misfiring in your face producing a sneeze. So one of the reasons this is a really interesting thing to talk about today is that, number one, it's not that well studied, despite the fact that it affects really quite a large proportion of the population - I mean, I've seen numbers anywhere between 11 and 35%, which is quite a lot of people; but also because it's kind of a rare example of something we see out in the world that has kind of a really interesting genetic reason. It's a heritable thing; so therefore, if your parents sneeze when they see light, eat dark chocolate, eat mint, eat grapefruit, whatever, it means that you yourself are more likely to sneeze. And it's a dominant trait, which means that if you have one parent who has it, then you are 50% likely to have it.
Now, in terms of the specifics of the genetics, that's actually less well understood. I've seen online a whole range of different things. Some places say it's something to do with chromosome 2. 23andme actually, the company that will sequence your genome if you give them a sample - for some money - found 54 different markers that they associate with photic sneezing. The fact is, we just don't quite know at the moment. But my favourite fact that I discovered while looking this up is the name of the syndrome that is given to people who have photic sneezing; and it's called Autosomal Compelling Helio-Opthalmic Outburst, which stands for ACHOO syndrome. Which I think is perfect.
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