Thinking Inside The Box: Insights Into One Of The World’s Deadliest Venoms

By Christie Wilcox | June 2, 2015 8:00 am
One of the deadliest venoms in the world can be found in the flimsy tentacles of this box jellyfish.

One of the deadliest venoms in the world can be found in the flimsy tentacles of this box jellyfish.

When you get right down to it, box jellyfish are little more than goo. The majority of their volume is mesoglea, a non-living, jello-like substance, which is sandwiched between two thin tissue layers. They have no teeth to bite with, no claws to scratch with — none of the weaponry we generally think of when we imagine a ruthless predator. Yet these boneless, brainless boxies are among the deadliest animals on Earth. The box jellyfish Chironex fleckeri can kill a full grown man in less than five minutes, and the venom it wields in its tentacles contains of some of the most rapid, potent toxins in the world.

Exactly what those toxins are, though, has remained somewhat of a mystery. Scientists have been trying to determine the composition of box jelly venom for decades, but have only uncovered some of its potent constituents. And while there’s still more to learn, last week, a research team from Queensland, Australia published the most extensive analysis of Chironex venom proteins to date, revealing some of the diverse arsenal that these gelatinous killers are equipped with.

Box jellies, like other members of the phylum Cnidaria, are armed with stinging cells along their tentacles. In each is a structure called a nematocyst which contains the venom and a harpoon-like structure on a biological thread. When nematocysts are triggered, their harpoon shoot out at speeds that can exceed 40 miles per hour, creating as much penetrative force as some bullets. Victims of box jellies can be hit with millions of these tiny stinging cells in a matter of seconds, causing large, painful welts. In severe cases, the venom causes systemic effects, including acute cardiovascular collapse and death in a matter of minutes or more delayed but potentially deadly symptoms — a condition known as Irukandji Syndrome.

“Despite the economic and medical impact that this jellyfish has on Australia (and similar species world-wide) we know very little about what is exactly in the venom,” explained Jason Mulvenna, Team Head of Infectious Disease and Cancer at QIMR Berghofer Medical Research Institute and coauthor of the paper published in BMC Genomics. The team’s goal was to perform the most in-depth analysis of Chironex venom to date, producing both a proteome (a library of the proteins present) and a transcriptome (a library of which genes are expressed).

This combination of approaches has become more and more popular in recent years as technological advances have opened the doors to faster and easier genetic sequencing as well as more precise protein determination. While they could have used either the genetic or proteomic approaches to look at venom proteins, the combination was particularly powerful. Transcriptomes tell you which genes are actively being expressed, but it can be hard to tell which of those genes are actually acting as venom toxins and which are involved in day-to-day cellular maintenance. Similarly, while proteins can be directly sequenced, it can be difficult to make sense of those kinds of data without genetic information, and there is no published genome for Chironex. So using both approaches was key to the team’s success.

From the genetic side, they constructed a tentacle transcriptome using next generation sequencing. This is done by separating out all of the messenger RNA sequences (or ‘transcripts’) — the first step on the pathway from gene to protein. They then chopped these into small pieces and sequenced them. Much like recreating a book from 10-word sentence fragments, the team was able to use special computer programs to align the little pieces, eventually creating a library of the expressed genes.

Then it was time for the protein side. To construct a ‘proteome’, the team had to isolate venom from the jelly tentacles. Unlike snakes or spiders, jellyfish cannot be ‘milked’ — so the team had to separate nematocysts from fresh tentacles, make them “sting”, and then separate the venom excreted from the capsule itself. That venom product could then be separated into individual components and identified using gel electrophoresis, liquid chromatography, and mass spectrometry.

The tentacles yielded over 20,000 predicted protein sequences that the researchers compared with known proteins in the UniProt database to identify potential toxins. They ended up with 179 likely toxins from ten different protein families. Their tandem proteomic analyses specifically identified 13 of these that were in venom in relatively high abundance: seven proteases, four of which were metalloproteinases, an alpha-macroglobulin domain containing protein, two peroxiredoxin toxins, two CRISP proteins and a turripeptide-like protease inhibitor. Another study from earlier this year had similarly constructed a transcriptome, and both research groups found metalloproteases and protease inhibitors.

The team also found new variations of the known cnidarian pore-forming proteins, or porins, that have been detected in a diverse set of species. Four were known from Chironex fleckeri when the scientists began their quest — the team found evidence for fifteen different variants.

“We now know that there is a whole bunch of unique toxins, only found in jellyfish, that may explain why the box jellyfish is one of the most venomous creatures known to man,” said Mulvenna.

“Now we know what is in the jellyfish venom we can do two things; we can start coming up with novel treatments for jellyfish stings that directly target the proteins we identified in the venom; and we can start seeing if these novel toxins are useful to us medically,” explained Mulvenna. More targeted therapeutics are a welcome idea, as previous studies have questioned whether box jelly antivenom is effective. A recent study showed that a specific blocker of the porin toxins worked far better than the antivenom in an animal model of envenomation, for example. Now that there are new targets to consider, scientists may be able to create a better treatment regime that stops the venom’s most deadly activities.

Perhaps ironically, those deadly activities may also be harnessed for good. More and more, scientists are mining venomous animals for novel pharmaceuticals. After all, these animals have had millions of years to hone the actions of their toxins — jellies, for example, have existed for some 600 million years, tweaking their venom along the way. Though the process of drug discovery is long and difficult (and thus any box jelly-derived drugs would be years if not decades away from production), the database of toxins created in this study can now be screened for useful activities like anti-cancer properties.

While the combined approach allowed for these novel discoveries, there is still much more to be done. “The upside of using this technique is speed and cost — we can generate a transcriptome quickly and use proteomics to refine and correct it,” explained Mulvenna. However, the methods used can overlook potent toxins that are present in lower concentrations, and thus further study is needed to detect and sequence the proteins that are less abundant.

In addition, correctly assembling the entire transcriptome without a genome as a guide is a bit like trying to piece together hundreds of pages of text after they’ve gone through a shredder. Mistakes stand out when the short sequences are mapped back to the constructed full-length ones; in this case 44% of the short sequences didn’t match, which Mulvenna says is “a result of errors in the assembly.” This suggests that even with the dual approach, there are proteins being produced that we still don’t know about.

The study design also is only able to detect protein toxins. Many venoms are complex chemical cocktails with diverse toxins. But for now, the team has more than enough to work with.

“Now the fun starts as we start working on individual proteins to find out what they do and why they are so potent,” Mulvenna said.

 

Citation (Open Access!): Brinkman, Diane L., Xinying Jia, Jeremy Potriquet, Dhirendra Kumar, Debasis Dash, David Kvaskoff, and Jason Mulvenna. “Transcriptome and venom proteome of the box jellyfish Chironex fleckeri.” BMC Genomics 16, no. 1 (2015): 407. DOI: 10.1186/s12864-015-1568-3

Note: on Sunday, I went off on bad science coverage of this paper, saying that I was so disgusted by the awful reporting that I couldn’t even write the post I had wanted to about the study itself. This is that post. 

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  • Tom

    After your previous post slamming coverage of this paper in particular, for focusing on how it might have applications to antivenom or cancer drugs, you talked to the author about how this research can lead to more effective antivenoms and novel medical applications; you specifically note the possibility of cancer treatments.

    This wasn’t not the focus of your post, and you gave some nuance to it, but then again, you’re not writing for the same audience that gets their science news from the Australian Broadcasting Corporation.

    • http://christiewilcox.com/ Christie Wilcox

      The reason that the other coverage was so horrendous is that it made antivenoms the entire purpose of the study, when the point of the study was discovery and learning more about these animals whose venoms have remained so elusive. Had treatments and cancer drugs been mentioned as nuanced, hypothetical follow-ups (as they are in this), I would not have been so bothered. Just because ABC has a broader audience doesn’t mean it’s ok to completely misrepresent the research and its novelty/importance.

      (To be clear: I didn’t talk about making “more effective antivenoms” because that’s not something this research will lead to — treatments other than antivenoms are different. Antivenom = antibodies, treatments = everything else that might act to inhibit toxins or their effects)

      • Tom

        Apologies for my incorrect use of “antivenom”.

        Everything is a trade. In order to get ABC’s audience to click on a story, it needs a hook, and that hook has to be something that is seen as relevant to their readers. The general public is unlikely to care about knowledge for its own sake. (Politically-driven funding is also subject to this.) Treating people who have been stung by a dangerous animal or curing cancer are good (although the latter is overused) hooks, and the articles did go into some detail about what actually WAS done, and got quotes from the author of the study (maybe reluctantly, although, again, he gave similar quotes to you) about the potential for new treatments and drugs. And now a bunch of people – future scientists, voters, people that need something to talk about at cocktail parties – are aware that scientists in Australia are doing work with box jellies. Would they know that, otherwise? Is the net benefit of them being somewhat aware of that information versus them being somewhat unclear on the details and applications positive? Beyond the headlines, I don’t think the articles were so egregious as to make the answer obviously no. And with regard to headlines and ledes, you’re not above spicing things up a little, yourself, finding a racy hook to tempt readers to click.

        I guess I have a hard time understanding just WHY this bothers you so much. This wasn’t, “hey, they could do better.” This was, “they utterly failed and I’m so mad I can’t focus on anything else.”

        • http://christiewilcox.com/ Christie Wilcox

          “The general public is unlikely to care about knowledge for its own sake”

          Bull. Journalists write stories about all kinds of science that has no medical value just because the science and the animals are cool. Snake venoms can be talked about without purely bringing in antivenom. Why the need to reduce box jellies to antivenom? That’s not hooking the public by what they care about – it’s telling the public that’s the only thing they should care about. It’s the journalists job to show why people should care about a topic, not use cheap and inaccurate ploys to get clicks. Saying everything will lead to a cancer cure erodes the public’s trust – science journalists can only make so many promises without delivering on overhyped claims before people start to think either journalists don’t know what they’re talking about, or worse, that scientists are full of it.

          And the articles were that bad – they implied we have no antivenom now! They said it was the first time anyone has looked at the venom! These are straight up factual errors, and the posts are littered with them.

  • Jason Mulvenna

    I was the senior author on the paper and this was my first publication that has picked up any significant media attention – I mostly work on parasites and parasite-induced cancers which doesn’t have a lot of relevance to the typical Australian.

    It was an interesting thing to go through and it was good to get some press coverage and family, friends and colleagues all saw what research is occurring in my lab. On the other hand I have also been congratulated by non-scientist family friends for finding a cure for cancer! I appreciate that the media needs an angle for their reports but when they inspire such a misleading conception of what a publication is about the damage done, overall, probably outweighs the positive impact of the publicity. I agree with Christie and think efforts to ‘sex up’ basic research with outlandish claims undervalues the importance of basic research and eventually will undermine the public’s trust in science.

    One off topic example is the media’s reporting on the health effects of alcohol. Some of my friends have a very keen interest in this topic and sensationalist reports stating either that x drinks a day will kill you or that x drinks a day protects you from heart disease – which may or may not have been supported by the study – just confuses everyone. They ask me as their pet scientist but I’m confused too. I know that this is often scientists inflating the significance of their results for press coverage but maybe the media needs to employ science reporters who are able to evaluate this type of thing (probably not likely in the current environment).

    The other issue is just sloppy reporting – there is an antivenom and I specifically stated that there is in interviews, I discussed potential *new* treatments for stings based on the molecular biology of the toxins that were discovered (not anti-venoma) and ‘cancer hope’ (in a lot of other headlines) is not supported by what I said which was that we sometimes throw these toxins on cancer cells to see if they have some anticancer effect.

    All in all it was nice getting some attention and at an individual level it did more good than bad for my research but overall I think reporting like this is bad for science.

    • Overburdened_Planet

      Have you tried contacting the magazine or the author to make the corrections that you want?

  • Frank H Little

    Does anybody know why the box jellyfish has developed and preserved such an extensive and varied venom kit?

  • nik

    Amazing what can be achieved in 600 million years!
    I wonder why it hasn’t developed intelligence?
    Or has it?

  • Shane Wegner

    I’m picturing someday when humanity collectively has a mighty digital catalog of all relevant proteins. I can see we are a long way from that though! In some ways, space really ISN’T the final frontier.

  • Enrique Goban

    We did a research assignment in 2012 looking at nero toxins and were looking to the box jellyfish as a potential donor program. We identified types 3 and 5C as possible candidates before funding curtailed any further research. “Fun” times, for sure. Those are some potent combinations.

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About Christie Wilcox

Dr. Christie Wilcox is a science writer based in the greater Seattle area. Her bylines include National Geographic, Popular Science, and Quanta. Her debut book, Venomous, released August 2016 (Scientific American/FSG Books). To learn more about her life and work, check out her webpage or follow her on Twitter, Google+, or Facebook.

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