Old collection scale insect slides and the power of confocal microscopy

Following a tweet I saw today on Plecoptera slide collection, I became nostalgic of scale insect collections.

Scale insect descriptions and identification are done using mounted material of stained insect cuticle. Because both males and females don’t have very sclerotized cuticle, they become transparent and outer structures are very easy to observe with a light compound microscope.

An example here, we have wax scale insects on a leaf (top left). After removing the inside to keep only cuticle for staining and mounting, we can have a better view of micro structures (top right). This helps making descriptions illustrated by diagrams (bottom).


In the past, male samples were not mounted or if they were, the quality of mounts was not as good as female samples. This is mostly because males being ephemeral, they were not used for identification or description. There are however quite a few old slide mounted males in collections and especially for species that are difficult to find, they are in my opinion, quite valuable to study for phylogenetic purposes.

When I was working on a phylogenetic study of scale insects, I spent a lot of time examining male samples in old collections. Knowing that males are so rare compared to females, finding an old slide mounted male of a rare species was like finding a treasure.

For a couple of years, I focused on fossils and male morphology of the ensign scale insects (Ortheziidae).  So I found slide mounts for Ortheziidae males but some of them couldn’t be observed clearly because the cuticle was not cleared properly. Using a traditional light compound microscope, it is just impossible to look at details of setae and secretion pores.

During my master’s degree, I worked in the Natural History Museum in Paris, and my supervisor showed how to unmount samples, and reprocess them for clearing and staining. But these Ortheziidae males were rare so I tried another alternative: confocal scanning laser microscopy.

So here it is:


A lot of surface structure were easily revealed at 633 nm. This is not surprising as normally, staining is made with fuchsin acid, its emission wavelength is 630 nm.

So how much detail can we actually see?

These are abdomens of mounted samples. Setae and secretion pores are easy to see.Screen Shot 2018-04-16 at 19.56.45

When you look more closely, it is actually quite incredible how much detail can be captured! Screen Shot 2018-04-16 at 19.57.23

I think confocal imaging is a great non-invasive option to work on old collection samples, especially ones that are slide mounted, obscured and rare. Although not every place has a confocal microscope (especially museums), the American Museum of Natural History is equipped with a Zeiss LSM710 which was really handy!

Do you know any confocal images of old collection slides? I would love to see them!




Insect-fungus symbiosis: a scale insect feeds her own roof!

Scale insects are often associated to fungus. Females secreting honeydew trigger sooty mold growth and harm the host-plant. Induced undesirable sooty mold on plants makes scale insects some of the most damaging phytophagous insects.  For instance, Cryptococcus fagisuga, a felt scale insect living on beech trees is associated to a fungus called Neonectria coccinea that will grow from the holes created by the insect’s puncture hole from which it feeds. Ultimately, the mold growth becomes more lethal to the tree than the insect. This is so true that I remember my Master mentor telling me that beech trees have almost disappeared from Paris because of this species!

Sooty mold are also harmful to scale insects, as the female secreting honeydew are sessile and they would literally die in their own moldy poop. This is when ants comes into play as essential mutualistic animals: by profiting from scale insect honeydew droplets as a food source, ants naturally clean their secretive waste, preventing sooty mold growth.

Images showing the fungus structure in which the mealybug lives in (left) and details of fungus mealybug interaction (right) . Source: modified from Gavrilov (2017)

In an extraordinary situation, a species of scale insects from the mealybug family (Pseudococcidae) has taken advantage of mold and developed a unique symbiosis with a fungus species. Orbuspedum machinator, a newly described species from bamboo twigs of Thailand, has legless females and uses hyphae of a fungus Capnodium species to make herself a roof. In exchange, the mealybug secretes honeydew and feed the fungus.

A win win situation.


Hypothetical scheme of the formation of a fungal domicile by Orbuspedum machinator gen. et sp. nov. (Gavrilov 2017)



Open access: Gavrilov (2017) An amazing symbiosis between an animal and a fungus in a new species of legless mealybug (Insecta: Pseudococcidae)


10th International Conference on Juvenile Hormone

The 10th International Conference on Juvenile Hormone will be held at Tsukuba (not far from Tokyo) between June 9th and 13th.

This conference takes place every four years or so. The last one was in 2007 though, and this one was supposed to happen in 2011 in Japan but was cancelled because of the earthquake (Dr Minakuchi, pers. com.).

This will be my first conference outside the field of systematics/phylogeny/scale insects, in which I will present our preliminary results on scale insect sexual dimorphism linked with Juvenile Hormone.

Five big names in the field of JH will give plenary lectures: Xavier Belles, Marek Jindra, Reddy Palli, Alexander Raikhel and Lynn Riddiford.

Additionally, there are some exciting talks I am looking forward to, such as the action of JH in insect embryos (J.W. Truman), JH and aphid polyphenism (T. Miura)  etc.. There seems to have quite a diverse range of study organisms as well, which is exciting!

For a glance at next week program -> here

The giant scale Drosicha corpulenta: on uncovering its biology

Photo credit: Isabelle Vea
Photo credit: Isabelle Vea

May 4, 2014

Walking back from lunch and discussing with my boyfriend and a friend visiting from Kobe, I was looking down and spotted a large bug crawling on the ground. I first thought it was a pill bug with a very strange color, but half a second later, I kneeled and realized it was a giant scale insect, Drosicha corpulenta! What a sight, I have never seen this genus alive and was elated to be able to observe them walking. They were many on the ground, probably because it was very windy. We collected a couple of them.

May 7, 2014

Morning: I tried to look for the tree where they were feeding and found a few different tree species infested. I could not find any males.

After lunch: Next to my lab, I noticed a group of trees, and immediately was shocked to see the large infestation of the same scale insects. I pass by this group of trees everyday since I arrived in Nagoya University, how come I haven’t seen them before? Immediately, I try to evaluate the extant of the infestation. They are almost on every single tree. Trying to look for a male, I finally find one hanging on a tree trunk. End of afternoon: I go back to look for more males. I will find a total of 4 in the following days.

I kept a dozen females and the 4 males in a box for a few days. This video shows a male walking on a female.

The males died after 3-4 days of frantic copulation.

Photo of a male, dropping dead after his last female encounter. The endophallus (= penis) is out. This is the first time I see a scale insect penis (I have only seen slide preparations and fossilized ones). In a few scale insect families, the endophallus looks like this, and can be longer, with many spikes covering it. Other families such as the soft scales or mealybugs don’t have an endophallus. Notebook 4-8 May 10, 2014

With a colleague we bought a persimmon tree so we could collect and observe some females. Drosicha corpulenta feeds on many species and is considered a plant pest in Japan. Here is the list of plant species they have been found feeding on.

May 11, 2014

I collected a few females and put them in the soil of our persimmon tree. Half an hour later, I checked the tree and a few of them had climbed up the branches and settled to feed.

May 12, 2014

I checked the persimmon tree and the females are still feeding.

May 15, 2014

I went back to the infestation spot to collect some more individual for the persimmon tree. Most of the females are GONE. Where they were gathering by dozens, nothing is left. I was puzzled at first. What did happen? I thought that the gardeners of the university wiped out the branches of all trees, but this would be unlikely as even the oak higher branches don’t have scale insects anymore. Also, the branches really looked clean. So my alternative hypothesis is that the females marched down to the ground to lay eggs at the same time! Alternatively, on my persimmon tree, the females that were feeding on May 12 are also gone. I need to look at the soil and check my theory. According to Scalenet, the reference database for scale insects, there is an account on their biology in:

Kuwana, S.I. 1922. Studies on Japanese Monophlebinae. Contribution I: The genus WarajicoccusBulletin of Agriculture and Commerce, Imperial Plant Quarantine Station, Yokohama 1: 1-58. 

I went to the agriculture library of my building and could only find Contribution II of the journal…

May 20, 2014

The university  library managed to find a copy of the publication! This study from 1922 provides a biological account of the genus Drosicha (then Warajicoccus) additionally to morphological descriptions of all stages (juveniles, female and male). Beautiful hand drawn plates are also presented at the end of the booklet. Kuwana confirms that when the adult females are ready to lay eggs, they migrate to the ground and hide among tree leaves and other litter matter.

To be continued….

The scale insect that took three years to grow

If you want to see the adult of this scale insect species, you’ve better come to Sapporo, Hokkaido in Japan the right year. And if my calculations are accurate, it should be in the summer 2015!

Most famous scale insects are usually dangerous plant pests, attacking ornamental plants in your garden, or making fruits unaesthetic to be sold (although still edible in general…). Those species have at least one full generation per year, sometimes two and in greenhouses can make several generations a year.

But it is completely different for Xylococcus japonicus Oguma. In 1919, Dr Oguma from the Hokkaido Imperial University (now Hokkaido University) published a detailed study on the biology of X. japonicus. He received a branch of alder (Alnus japonica) in the spring of 1913, where there was, imbedded in plant tissues, a mysterious scale insect. Its presence was only betrayed by some white projections coming out of the branches.

Dr Oguma was wondering why the scale insect seemed to only attack the tree’s old tissue and did not seem to affect the youngest parts of the branches. He then followed the development of the insect for three entire years before understanding why.

This is how it all happened between 1913 and 1916:

Notebook 4-5

Summer 1913: Dr Oguma was not in Hokkaido and when he came back, he only found dead adults. Argh, too bad, maybe next year?

Spring 1914: the eggs laid by the adult females at the end of the summer 1913 finally hatched. Right after, they sought shelter within the cracks of alder’s bark, inserted they sucking mouthparts to feed and started to secrete wax protection.

Fall 1914: after a whole summer of gorging themselves with alder juice, the first-instars finally molted to second-instar nymphs.

Winter 1915: the second-instar hibernated.

Spring 1915: in May, the insects became active again, producing more wax filaments and becoming larger

Summer 1915: they were still in second-instar stage throughout the summer. They had become so large compared to the cracks they have been hiding in that they could not move an inch anymore.

Fall 1915: stuck in the cracks, still not molting..

Winter 1916: hibernation happened, and still stuck in cracks.

Spring 1916: the second molting event happened. Yay! third-instar nymphs!

Summer 1916: by August, females and males (though still third instar nymphs) were distinguishable. At the end of August, the females molted to their young adult form but the males entered their quiescent pupal stage (different from an actual insect pupa but still called pupa, confusing…). The pupal stage lasted only one week.

Reproduction: the female being completely stuck in bark cracks waits for a male (he moves and is completely different from the female – see post on sexual dimorphism soon!) with her posterior popped out from the crack. Copulation only happens in the morning. Although the male, after a minute of copulation goes away looking for more females, the freshly copulated females hides herself back in the crack and only pops her butt out again the next morning. Because adult males only live one day, the next morning always is a new male.

Fall 1916: the female started to lay eggs and by the end of September, she was found dead, her body acting as a cover to protect the eggs.

And this was the generation of Xylococcus japonicus between 1913 and 1916!

The time of development explained why only older tree branches were affected by the scale insect as one generation only feeds on the same tissue for 3 years. The affected branches are severely damaged and often fall off. Maybe, if this species had produced yearly generations, the host plant would have been easily been decimated. Because this scale insect seems to be very picky on which tree juice to drink (only feeding on alder), could it have developed this 3-year-long generation to give enough time for the host plant to recover?

I have made my calculations and if right, the next adults should emerge in August 2015, 33 generations and almost 100 years after the adults observed by Dr Oguma in 1916.

Well, I am not sure if I will still be in Japan then, but it is definitely worth looking for the scales this summer up in Hokkaido and bring some photos of second instar nymphs to show you!

To be continued…

Reference: Oguma, K. 1919. A new scale-insect, Xylococcus alni, on alder, with special reference to its metamorphosis and anatomy. Journal of the College of Agriculture, Hokkaido Imperial University 8: 77-109.