Guest lecture by John Pearse and Janet Leonard, from Long Marine Laboratory, University of California-Santa Cruz. They did research on Banana Slugs for several years and talked about their findings in the class.
John talked about habitat, color, general characteristics, predators, and taxonomy of hermaphrodites. Janet talked mostly about sexual behavior of slugs.

• Habitat : Banana Slugs are slimy, spineless gastropods which live on forest floor in moist places. They are part of animals of forest of North West. They are also found in conifer forest, open forest, and unexpected habitats like UC Davis reserve dry area, and also in Springs. Pacific grove has lots of ice plants in which Banana Slugs live.
• Diet : Some slugs are predators. They eat other slugs, snails and earthworms. Banana Slugs eat feces. They are fond of mushrooms, but it is not their major diet. In labs they are fed lettuce.
• Color : They are mostly yellow in color, but they are also found in brown, and black colors.
• Predators : Raccoons, garter snakes, ducks, geese and salamanders eat banana slugs.
• Taxonomy : Banana Slug belongs to genus Ariolimax. Three species are included in these genus : A. californicus, A. columbianus, and A. dolichophallus. These species have opening either on top or on the right side called as pneumostome. Everything is done through this opening. Mead classified A.californicus and A.dolichophallus as two separate species based on their penis and vaginal muscle size. A.californicus has a huge vaginal muscle and penis, whereas A.dolichophallus has thinner vaginal muscle. Although the species of Ariolimax family are morphologically apart, they are not distinct at molecular level.
• Reproductive morphology : Slugs are hermaphrodites (have both male and female organs), but they also get involved in mating. They have a ovisperm duct, and possess a complex mechanism to keep the external and internal sperm separate. They have vesicle to store sperm. They have vaginal muscle and some species have large vaginal muscle. Have a penis sheath with penis inside it.

• Sexual selection : These species can be distinguished through genitalia. In 1985 it was hypothesized that the genitalia distinguishes species and sub-species, and hence it is probably a feature of sexual selection. At molecular level all the slugs are indistinct.
• Sexual behavior : Slugs get involved in elaborate mating and sperm is exchanged through genitalia. They often get involved in apophallation, a mechanism where the mating partners chew off their penis. Ariolimax stramineus has a blunt penis and no biting or chewing of penis is observed. A.californicus and A.dolichophallus show apophallation. 5 in 100 copulations end in apophallation. Although apophallation is not that often but it happens.
• Copulation and egg laying habits : Copulation starts during foggy nights of July, August, and September. Egg laying starts when it begins to rain sometime during October, November, and December. Slugs lay eggs in moist places under things like leaves. In labs they lay eggs on surfaces at 18 - 19 degrees celsius. Low pressure seems to be a stimulus for egg laying. On an average 75 eggs are laid in a week, and it takes 7 weeks for hatching. A.dolichophallus lays big eggs, and A.californicus lays tiny eggs. A.dolichophallus is the youngest to copulate at about 6 months and youngest to lay eggs at about 10-1/2 months, and the life span is about 7 years.

The slides from the presentation of John and Janet can be dowloaded here,http://compbio.soe.ucsc.edu/slugGenome/ .

Additional notes from Jonathan follow.

John originally worked with sea urchins and sea stars primarily. Janet joined the lab about twelve years ago to study marine slugs. She was interested in hermaphrodite mating behavior. Banana slugs' rare behavior of apophallation became a research focus. No one had really studied banana slugs [academically] since the forties. Alice Bryant Harper (Aptos naturalist, works with Santa Cruz Museum of Natural History) wrote The Banana Slug (1988), the best book on them.

Despite Chancellor Sinsheimer's desire to keep the sea lion as the UCSC mascot, students voted 95% for the banana slug.

There are just two complete mollusc genomes, [California sea hare and giant owl limpet], and neither are very complete.

Banana slugs habitat is very diverse. Though often found in conifer forests and considered an animal of the Northwest (first found in Washington or Oregon), they've been found in drier habitats: San Diego, Napa's McLaughlin Reserve (by small springs), abandoned rice patties in the Sacramento River Delta, oceanside iceplants in Pacific Grove. High variation in the numbers you'll see on any day/site: none or dozens.

They eat feces, hemlock, poison oak, mushrooms (reported but John has not seen), sorrel, ferns, ice plants, humus soil. In the lab they eat hamburger, cat food, apples, beans, zucchini, mushrooms, yams, lettuce and milk.

Colors may camouflage them, e.g. dead leaves often turn bright yellow, the color of species in Santa Cruz and the SF Bay area. In other areas you'll find spotted slugs – but they may be a different species.

There predators may include [seemed uncertain] garter snakes, salamanders and newts, birds and some small mammals. It is possible that some specific carnivorous snails and slugs eat banana slugs.

Aphallarion buttoni originally thought to be a different species because no penes were found when dissected (late 19th century). However, a Stanford professor later found some with penes and so sent students into the field to study. They observed apophallation. That was the end of buttoni as a separate taxon. It became Ariolimax columbianus.

All banana slugs have an opening on the right side of the “head” for defecation, breathing, and copulation. The only way to distinguish species is by dissection of the genitalia. [See slide *Ariolimax Arilimax columbians genitalia* for overview of genitalia.] The gonad has a mix of testes and ovaries, and they can play both roles at same time curing copulation. How is sperm kept separate during copulation? It is not necessarily. They can fertilize themselves.

And aphalon are born without a penis [sometimes?].

Ariolimax Meadarion californicus is found in San Mateo county. Santa Cruz has dolichophallus. [See slide comparing their genitalia.] Mead thoought dolichophallus and californicus were sufficiently different to be a separate species.

A collaborator in Belgium has been sequencing banana slug mitochrondrial DNA. They see at least five clades but cannot yet connect them. ~“Morphologically distinct and molecularly distinct are not the same thing.” [See slide.]

Interestingly the distribution of the salamander genus Ensatia is similar to that of banana slug [dolichophallus? – see slide]. Is this a remnant of five million years ago when there were islands in the Monterey Bay? Morphologically distinct but molecularly [mito. DNA] indistict suggests recent change.

Janet's interest is in sex selection. Eberhard's hypothesis: [That we can ] classif[y] based on genitalia (as done with insects, spiders, etc.) suggests the importance of sex selection. Genitalia differences in nearby counties are not explicable by natural selection, e.g. how would NS explain [improved fitness by a different vaginal muscle in the same geographical area.]

The talk focused on courtship behavior: much effort/expense in banana slug mating.

Ariolimax stramineus courtship: antiparallel alignment of slugs is standard. They line up right sides of their heads until the alignment allows copulation. The first copulation takes place in about twenty minutes with subsequent occuring over about two hours. No apophallation in the one shown on film.

Mating film for brachyphallus: This is one of the three under Ariolimax Meadarian. Note the initial biting and head swinging which seem to cause no damage. The biting helps line up the head regions. Banana slugs cannot reverse so the cirlcing helps get them get into position for mating.

Mating film for californicus: Unilateral copulation (after two hours biting, head swinging). It is hard to tell which plays the role of male and female but Janet thinks they alternate over a copulation session.

Mating film for dolicophallus: Example of apophallation (by the first to withdraw). How costly is this for a hermaphrodite? It is pretty rare: 5 out of 100 copulations end in apophallation.

Why does apophallation occur and when? They have observed it only nine times and never by virgins. Is it done as retaliation if one partner does not give any sperm? Do they run out, and perhaps later in life focus on egg laying?

Note in the table comparing copulations that dolicophallus and californicus are indistinguishable by mitochondrial DNA.

The rapid morphological changes are among the evidence for sex selection, as well as the high cost of courtship/mating. There is some evidence for sperm competition.

Soon we should have based on microsatellite data (nuclear DNA) …

It is unknown how many chromosomes they have, though chromosome variation is usually not seen till “quite high” taxonomic levels.

Their egg laying habits in the field are uncertain. Perhaps under leaflitter. They do not dig holes in moist soil like garden snails. In the lab at 19^C, they take seven weeks to hatch, sometimes two to three weeks longer. The record for a clutch is seventy-five eggs (but this could have been multiple clutches since they don't check every day). Egg size varies. Dollicophallus eggs are almost the size of jellybelly and weigh up to half a gram. For californicus they're usually under 0.3g.

Egg laying starts in fall and copulation is associated with foggy nights (late summer, Santa Cruz). Eggs are laid October through December when the rains come and usually finished by February. Hypothesis: Low pressure systems trigger egg laying. (This is based on lab observations, and makes sense since dehydration is the highest mortality source. You want to lay eggs at the start of the rainy season in a moist place.)

They have observed copulation for slugs as young as six months and egg laying as young as ten.

How long do they live? No one knows but perhaps two to three years. Some have lived thirty months in the lab (when sacrificed).