Through the Looking-Glass

Well, the mystery presented in my previous post was solved within an hour of my posting it, but before I get to that, let me back up and chronicle the adventures Julia and I have had in moth dissection so far. As I mentioned previously, we started out by practicing on some specimens I collected for a class over 15 years ago; thanks to the professor’s eagerness to be done with his position at UVM and move on to his new one at UC Berkeley, all of the collection data were thrown away and all we know is that I collected these moths somewhere in New England (mostly in Burlington, VT, but I know I got some in Massachusetts and Maine). In this post I’ll just show the males; females are an entirely different matter, and we’ll deal with them some other day.

On February 4, Julia dissected the first batch, beginning with this noctuid—which, because the wings haven’t been spread properly, I’m thinking I actually found dead on a windowsill in our house at some point and just stuck it on a pin and put it in the box of unimportant specimens from grad school in case I had a use for it at some point. [Edit: It’s a “bicolored sallow,” Sunira bicolorago. I did in fact find it dead on a windowsill in the living room last January but had neglected to label it.]

That day Julia took photos of her dissections by holding her phone up to one of the microscope eyepieces. For this one and the next, she used a stain called “Orange G” instead of Eosin Y, and no Chlorazol Black. Just experimenting. Here are the genitalia of the moth before the coverslip was added…

…and after. As you can see, the angle at which you’re viewing the genitalia, and the way in which they happen to be smooshed when slide mounted, has a big effect on their appearance. For this reason, these days permanently slide-mounting genitalia is frowned upon, and the alternative is to remove them from the slide when done examining them and put them in a tiny vial in some viscous fluid (a lot of people use glycerin; Terry Harrison uses lactic acid because it helps to neutralize the potassium hydroxide, which otherwise can continue to slowly dissolve the genitalia for years and may ultimately destroy the specimen), which is then placed on the pin together with the rest of the moth.

Next up was this substantially smaller moth, a crambid (subfamily Crambinae, and I think tribe Crambini), which despite the unspread wings I definitely did collect for that class. [Edit: It’s Agriphila ruricolellus, the “lesser vagabond sod webworm.”]

Julia’s notes for this one say “very hard to spread,” referring in this case to the valvae of the genitalia, which are normally clamped together and have to be spread apart to examine and photograph. I guess because it didn’t turn out too well, she didn’t take a photo, so here’s a quick one I took just now using our new DSLR-to-microscope adapter. The lack of any bubbles in this one may be due to the fact that she used Hoyer’s mounting medium instead of lactic acid, or maybe she was just lucky. We haven’t found any correlation between how we put on the cover slip and how many bubbles there are, other than that any attempt to improve the situation always makes it worse.

The next contestant was this fairly similar crambine… [Edit: Chrysoteuchia topiarius, the “cranberry girdler.”]

…with completely different genitalia. (Back to Eosin Y, lactic acid, and phone camera for this one.)

On February 6, Julia dissected three more males. The first was this moth that I had labeled as a tortricid when I took the class, but I’m pretty sure it’s actually a gelechioid of some sort. [Edit: Yes, it’s Depressariidae: Machimia tentoriferella, the “gold-striped leaftier.”]

The genitalia are just in the tip of that wee abdomen. In this next photo I’ve included the abdomen along with the removed genitalia (with phallus detached) so you can see the relative size.

And here’s the same photo cropped down to just the genitalia.

The second moth of the day was this one, which I’d labeled as a noctuid… moths with a wingspan of more than 1 cm or so really aren’t my thing, so please chime in if you know better. [Edit: it’s a notcuoid, but the family is Erebidae: Palthis asopialis, the “faint-spotted palthis.”]

The phallus is too far away on the slide to include in the same photo with the rest of the genitalia, so here are two separate shots. (This was another “Orange G” one).

And Julia’s last male dissection to date was this pyralid, which I did a terrible job of pinning. I’m still no good at it, which is why Julia gets to pin all of the moths around here.

It’s the Indianmeal Moth (Plodia interpunctella), a common pest of dry stored foods, and I must have grabbed it from my kitchen. These moths were abundant in the house where I lived in Burlington, due to one of my housemates essentially moving in with his girlfriend and leaving a cupboard full of neglected packaged foods.

Not Julia’s best work, but the genitalia do match what’s shown here if you squint your eyes just right.

This is the other thing that was mounted on the slide… I wasn’t sure if it included the phallus, but I guess the object at the bottom of the photo pretty much matches the shape of the phallus shown at the above link.

On February 12, it was my turn to give dissection a shot. I started with this big ol’ hemlock looper (Geometridae: Lambdina fiscellaria). Yes, it’s pinned upside down.

Not having a phone, I tried taking photos with the Olympus TG-4 held up to one of the eyepieces.

Based on the example shown here, the asymmetrical claspers (?) are typical of this species.

After dissecting a female of a different geometrid species, I tried this gelechioid, which is the same species as Julia’s #4 (they were collected together and have the same wing pattern and genitalia). [Edit: Machimia tentoriferella again.]

For another example of how the appearance of the genitalia can change depending on how you look at them, notice how before I flattened this with the coverslip there is a pair of “lips” directed straight at the camera near the top of the structure…

…and after adding the coverslip, those “lips” are smooshed downward.

Since the valve on the right was folded, I took off the coverslip and tried again. My second try was a big improvement, except for all the darn bubbles.

I tried one more time and miraculously ended up with fewer bubbles, as well as more widely spread valvae (not necessarily an improvement, but just to point out that the orientation of the valvae is of no significance when it comes to trying to identify species).

On February 17, the DSLR-to-microscope adapter had arrived in the mail, and since I hadn’t ruined any (male) specimens so far, I decided to try out a couple of unknowns and see if I could actually use genitalia to identify them. First up (being the biggest) was the willow leaf-tying crambid I wrote about last time.

To summarize how my ID attempt went, I tried Munroe’s (1976) key to the genera of Pyraustinae and didn’t reach a satisfying conclusion, other than that my moth clearly was not one of the species illustrated in that publication. I posted a link to my blog post on Twitter and a couple of moth-y Facebook groups, along with a cry for help, and about 45 minutes later Chris Grinter replied “Looks like a great match to Framinghamia helvalis“, with a link to this image in the Moth Photographers Group North American Lepidoptera Genitalia Library. He was clearly right, and early the next morning Steven Whitebread and Aaron Hunt independently told me the same thing.

The main problem was that this moth is in the subfamily Spilomelinae instead of Pyraustinae! It had occurred to me to check before I got started, since Munroe distinguishes the two groups right upfront.

Pyraustinae: “Praecinctorium weakly bilobed, the lobes parallel and longitudinal, diverging at an angle ventrolaterad from tip of praecinctorium proper; forewing of male with straplike frenulum-hook arising from costa near base, in addition to the retinaculum (a group of stiff scales arising farther back on the wing and also helping hold the frenulum in place) (Forbes, 1926: 331-332); valve of male genitalia almost always with basally directed clasper, one or more of its basally or dorsally directed lobes usually with conspicuous setae or erect scales; bursa of female genitalia almost always with rhomboidal or mouth-shaped, spinulose, transversely keeled signum.”

Spilomelinae: “Praecinctorium strongly bilobed, the lobes transverse and often projecting visibly beyond each side of base of abdomen; male with retinaculum but no frenulum-hook; valve of male genitalia with its clasper, when present, directed distad, rarely with any obvious basally directed lobe, and without conspicuous erect setae or scales; signum of female various, but not rhomboidal.”

Isn’t it nice how the only figure reference is to something in another publication from 50 years earlier? My eyes sort of glazed over as I read the the first two characters (the praecinctorium turns out to be a structure on the abdomen that is specific to Crambidae), and I thought, “well, the tip of what I’m calling the clasper is pointed downward, and those guys who told me it’s a pyraustine probably know what they’re talking about…”

The other problem was that Munroe (1976) did not cover the subfamily Spilomelinae, and to this day there still is not a monograph covering the North American species of this group. Munroe did, however, illustrate the genitalia of Framinghamia helvalis in a 1951 publication. The genus got its name from the fact that the type specimen was collected in Framingham, Massachusetts. F. helvalis is the only known species.

Steven Whitebread pointed out that I could have identified this moth by searching the Mass Moths website for species of Crambidae known to feed on willow (and known to occur in Massachusetts, like all species on the site). There turn out to be just two, and the other looks nothing like F. helvalis (and there is just one record of it from Massachusetts). F. helvalis has not been documented on Nantucket before, but has been reared from willow on Martha’s Vineyard.

Of course, wanting to put a name on the moth was only part of why I dissected it. I also wanted experience using genitalia for identification, and to that end, here is an edited version of the diagram I included in my last post, with corrections in red (many thanks to Chris Grinter, Jim Hayden, and JoAnne Russo for their feedback).

It turns out that the genus Framinghamia is characterized by not having an uncus. And apparently “distal” refers to the end of the aedoeagus/aedeagus/phallus that is distal with respect to the moth’s head, not distal with respect to the rest of the genitalia (if you look back at my previous post, you’ll see that the phallus was pointing the other way when in situ). Jim Hayden informed me that the pointy thing that appears in Tony Thomas’ image right behind where the uncus would be (and also shown in Munroe’s 1951 drawing) is the anal tube, which is supported by the subscaphium (the sclerotized ventral wall). It is often removed from dissections because it is easily torn, not informative, and full of poop.

Having successfully dissected this moth, and having not yet gotten bogged down in trying to identify it, I moved on to my first leafminer, which was a Phyllonorycter (Gracillariidae) that Mike Palmer had reared from black cottonwood (Salicaceae: Populus trichocarpa) in Montana in 2017. The leaf mines are shown here.

I chose this moth because I have several specimens from the same rearing (so it was okay if I wrecked one), because the Salicaceae-feeding Phyllonorycter species are among the only North American gracillariids for which genitalia illustrations have been published (Davis & Deschka 2001), and because the key to species is based entirely on male genitalia.

To sex gracillariids I don’t go looking for tiny bristles on the leading edges of the undersides of the hindwings; I just look at the tip of the abdomen. The first specimen I looked at was a female, but the second was a male:

As with the Framinghamia, I soaked this moth’s abdomen in Chlorazol Black for an hour or two instead of just using the red stain. Here is the result (maybe more black than needed, but better too much stain than too little if I want to be able to actually see anything; the Eosin Y hadn’t had much of an effect):

Did I mention that this moth had a wingspan of 8 mm and the abdomen was only 2 mm long? So this thing I was now left with was less than 1 mm long, and I needed to spread those narrow valvae open somehow. Here’s a side view of the same thing, with the tiny tiny detached phallus floating off to the right:

Looking at this second view, I was confused, because there seemed to be three layers of things to deal with. I interpreted those two skinny things to the left and front as the valvae, and the long, central structure as the tegumen, which left me scratching my head about that last thing to the right. In Davis & Deschka’s illustrations, in addition to the detached phallus there is another part that is illustrated separately: the eighth abdominal sternite. I thought this sternite would be part of the soft “pelt” from which I had removed the genitalia, but the illustrated sternite for the species I seemed to have was very similar in shape to this object, so I removed it and here’s what I was left with:

With this view, it is clear that what I had just removed was in fact the tegumen (oops), and what I had thought was the tegumen was actually the pair of valvae (which I never did succeed in spreading apart, but it no longer seems necessary). The good news is, it is possible to identify the species from the above photos. Using Davis & Deschka’s key (which, unlike Munroe’s key, refers to helpful figures!), all I have to do is observe that the valvae are symmetrical, that they are longer than the aedoeagus (and gradually taper to a “slender, simple, often sinuate apex”), and that the vinculum is Y-shaped, abruptly tapering to an attenuate apex, and I know I’ve got Phyllonorycter nipigon. The genitalia of my moth match the illustration for that species well. I’m still not clear what those outer two wispy things are, but that horseshoe-shaped structure (the transtilla?) is supposed to be oriented the other way, and it seems like those wispy things may be attached to it in such a way that they would be pointing the other direction if I managed to flip the horseshoe thing to match the illustration, so maybe they are some extraneous structure that was supposed to stay with the rest of the abdomen when I did the dissection. Once again, I welcome input from anyone who knows the answers!

[Edit: Thanks to Aaron Hunt and JoAnne Russo for all the IDs!]