A living beetle in more detail than you have ever seen one before

February 1, 2015 • 12:45 pm

by Matthew Cobb

Take a look at this video. It’s a scanning electron microscope (SEM) study of a beetle. Nothing amazing there, except you’ll soon notice that the beetle is ALIVE. This is quite astonishing, because SEM is done in a vacuum and involves completely dehydrating the sample and then covering it with a conductive metal (often gold). None of that here.

This tour de force is the work of a group of Japanese researchers, and it has just been published online in the Proceedings of the Royal Society (£££, sadly).

In fact, I can’t read the article, even with my Uni password, VPN and the rest. So I can’t tell you anything about the beetle in question (can any reader ID it?), how long it survived, whether the process is lethal or not, or indeed anything more than what’s given in the abstract:

Although extremely useful for a wide range of investigations, the field emission scanning electron microscope (FE-SEM) has not allowed researchers to observe living organisms. However, we have recently reported that a simple surface modification consisting of a thin extra layer, termed ‘NanoSuit’, can keep organisms alive in the high vacuum (10−5 to 10−7 Pa) of the SEM. This paper further explores the protective properties of the NanoSuit surface-shield. We found that a NanoSuit formed with the optimum concentration of Tween 20 faithfully preserves the integrity of an organism’s surface without interfering with SEM imaging. We also found that electrostatic charging was absent as long as the organisms were alive, even if they had not been coated with electrically conducting materials. This result suggests that living organisms possess their own electrical conductors and/or rely on certain properties of the surface to inhibit charging. The NanoSuit seems to prolong the charge-free condition and increase survival time under vacuum. These findings should encourage the development of more sophisticated observation methods for studying living organisms in an FE-SEM.

How they put the Nanosuit around the beetle, how long it survived, and all the rest will have to wait until whatever glitch there is at the Royal Society has been fixed (or the University has paid its bill) and I’m able to read it.

A previous paper by the same group gives some more detail about the NanoSuit:

Most multicellular organisms can only survive under atmospheric pressure. The reduced pressure of a high vacuum usually leads to rapid dehydration and death. Here we show that a simple surface modification can render multicellular organisms strongly tolerant to high vacuum. Animals that collapsed under high vacuum continued to move following exposure of their natural extracellular surface layer (or that of an artificial coat-like polysorbitan monolaurate) to an electron beam or plasma ionization (i.e., conditions known to enhance polymer formation). Transmission electron microscopic observations revealed the existence of a thin polymerized extra layer on the surface of the animal. The layer acts as a flexible “nano-suit” barrier to the passage of gases and liquids and thus protects the organism. Furthermore, the biocompatible molecule, the component of the nano-suit, was fabricated into a “biomimetic” free-standing membrane. This concept will allow biology-related fields especially to use these membranes for several applications.

 This breakthrough might be particularly useful for studying the activity of structures at the surface of organisms; the  comment in the abstract about the lack of electrostatic charging in living specimens is particularly intriguing. We already know that bees and plants can interact through electrostatic charges, and it may be that this forms an important aspect to arthropod ecology.

Yasuharu Takaku, Hiroshi Suzuki, Isao Ohta, Takami Tsutsui, Haruko Matsumoto, Masatsugu Shimomura, Takahiko Hariyama (2015) ‘A ‘NanoSuit’ surface shield successfully protects organisms in high vacuum: observations on living organisms in an FE-SEM’ Proceedings of the Royal Society B.
h/t @ziyatong on Tw*tter

30 thoughts on “A living beetle in more detail than you have ever seen one before”

  5. I doubt the wee beetle was happy, and neither was I. How frustrating this video is, I found myself getting agitated and yelling out loud “NO, stop there! Wait, go back, what was that?! Focus there!!! URGGG!!!” Guess I’m too much of a control freak to enjoy. and whereas “normal” people talk about the cars and big houses they’d buy if they won the lottery, to have a SEM with my dream lotto winnings…

    well, anyway, another nice little advance in microscopy, standing on the shoulders of (extremely tiny) giants.

  6. From the paper:

    (a) Experimental organisms
    Fourth-instar larvae of the mosquito Culex pipiens molestus
    (approx. 6 mm in body length), collected from puddles and maintained
    in the same water they had come from, were used. In order
    to exclude any effects of the water, they were transferred to distilled
    water at 24+18C 2 days prior to the experiment, with
    distilled water changes every 12 h. The larvae were rinsed
    thoroughly in distilled water 1 h before experiments began.
    Adult specimens of the beetle Lilioceris merdigera were collected
    from Hamamatsu City, Japan. The animals were
    maintained in the laboratory at 20+18C and fed daily. Individuals
    chosen for the experiment were rinsed thoroughly in
    distilled water 1 h prior to experimentation.
    Specimens of the amphipod Talitrus saltator were collected on a
    sandy beach in southern Tuscany (428460 N, 11860 E; Albegna,
    Grosseto, Italy), transported to the laboratory in plastic boxes containing
    wet sand and maintained in an aquarium containing sand
    moistened with artificial seawater. They were fed weekly with dry
    fish food placed on blotting paper. The experimental animals were
    rinsed thoroughly in distilled water 1 h before experiments began.</blockquoteY

    2. How did they ensure the beasts stayed on the sample holder? I wouldn’t want a bug running loose inside my SEM. Did they use glue?

  7. It would be interesting how they confirmed the beetle was alive during the scanning.

    Did they take out the beetle afterwards and re-humidify it?

    Just wondering.

    1. I assume the process is not reversible. The beetle is going to die either way, but shrink-wrapping it first allows more time to observe before it suffocates.

    2. I think they took the movement you see early on as evidence of being alive. It seemed alive at least at the beginning, though not very lively.

    1. Bristles on the underside of its feet. They make a very high surface area contact with things they walk on, so they can climb up glass, possibly by vanderWaals forces.

  9. Folding up their legs like that, and pressing their feet together means it is, well, kinda dieing. This Gary Larson cartoon immediately comes to mind. That will be be in I hope another 60 years.

    I think the beetle is a chrysomelid. These are the very large and common family of ‘leaf beetles’. Besides ladybugs, most of the beetles you see out in the open, on plants, are leaf beetles. The way to tell is their tarsal segments (the segments on their feet) look like they have 4 segments, but they actually have a small 5th segment. You can see that early in the clip.

  10. Interesting … wrap a small insect in plastic and put it into the high vacuum chamber. I wonder how long the insects live; they won’t be able to breathe in the vacuum nor will they be able to breathe when they’re removed due to the polymer coating. I also wonder why SEM rather than AFM – are the structures of interest too small for AFM or do the investigators want the high scan rate?

    1. AFMs are cantilever based – basically a needle that has to reach close to the surface, controlled by bulky servos – so are generally used on planar surfaces. I don’t know if there are such that have been adapted to look at non-planar objects.

      On the other hand, a scanning electron microscope scans it beams over the subject and collects the secondary electrons at a convenient location. Hence the focal point is the beam, and you can see beyond corners. [A SEM image should in principle never be interpreted as a 2D optical image, but as a differently distorted 3D representation. Of course we tend to accept the usual interpretation…]

      The SEM scan ability should be valuable for investigating these bugs, as they are for other complicated structures.

      1. “[A SEM image should in principle never be interpreted as a 2D optical image, but as a differently distorted 3D representation. Of course we tend to accept the usual interpretation…]”

        Huh, I don’t think I knew that. Thanks. I suppose to notice a difference from regular projective geometry you’d need to be looking at a high-relief but very regular structure.

  13. I think the title should be changed to a more accurate “Dying beetles observed with good resolution and no electrostatic charging”

  14. I would think it would be hard to keep it in focus most of the time.

    Looks like the beetle was dying, even if alive. On the other hand we have had some mild success imaging objects without coating them (sometimes forgetting) and we can still get good images, not always, but sometimes.

    Even if alive when begun, the beam has got to have at least 0.1 uA or more of current…that fellow is getting some good juice: thermally and radiologically.

  16. I remember seeing the result before, if not these remarkable videos. Here is an article that tells a little more about the discovery, and of survival times. Jerry’s fruit-fly figures prominently [bold by me]:

    “The scientists tested a number of different bug larvae in high vacuums in a scanning electron microscope and soon noticed something strange was happening. While most of the bug larvae shriveled up and died — just as a human would if he or she hopped out of a spaceship without wearing an astronaut’s spacesuit — the fruit-fly larvae seemed able to survive for 60 minutes in what should have been unbearable conditions.

    The researchers studied the critters’ surfaces and found that, when exposed to the electron beams, the extracellular substances secreted onto the fruit fly’s body formed a protective layer of polymers that acted like a thin astro-suit, a few tens of nanometers thick. Fruit fly larvae that didn’t get the electron-beam treatment died in the vacuum. Clearly, the radiation was key to getting those secretions to fuse together into armor — a little like 3D-printing a suit straight onto the bugs’ bodies.

    Other species of bug larvae don’t excrete those substances, however — so the electron beams won’t do any good. Instead, the scientists immersed the larvae in a 1% solution of Tween 20 (a nontoxic compound used in biological experiments) and then subjected the bugs to electron or plasma irradiation. The Tween 20 would act like the fruit-fly secretions and form the ultra-thin body armor.

    All the creatures treated in this way survived the vacuum, including the flatworm Dugesia japonica, the ant Pristomyrmex punctatus and a sand-hopper called Talitrus saltator. They even seemed to grow up into normal, well adjusted adults, in spite of that early trauma of dealing with the void.”

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