Why hermaphrodites sleep around: Polyandry offsets costs of traumatic mating and inbreeding in sea slugs with a weaponized penis John O. Martin1 & Patrick J. Krug2 1 - University of California, Irvine 2- California State University, Los Angeles pkrug@calstatela.edu

Why hermaphrodites sleep around: Polyandry offsets costs of traumatic mating and inbreeding in sea slugs with a weaponized penis John O. Martin1 & Patrick J. Krug2 1 - University of California, Irvine 2- California State University, Los Angeles pkrug@calstatela.edu 2

Sexual selection and hermaphrodites Some simulataneous hermaphrodites mediate conflict over sex roles via reciprocal mating and ‘sperm trading’ Herms lack male-display traits, but often have male-advantage traits for traumatic mating, removing pre-copulatory female choice ‘penis fencing’ in flatworms Costs of tissue repair may lower fitness through female function while benefiting male function

SEM courtesy of Sandra Millen Alderia modesta

SEM courtesy of Sandra Millen

How to optimize effort through female role? Theory predicts ndividuals should reallocate effort to egg production when beneficial, but that energy could come from 3 sources: 1. sex allocation (SA) from sperm to eggs penis - Local Sperm Competition hypothesis predicts that SA will favor the male role in larger mating groups, when sperm competition shifts from being within-ejaculate to among rival ejaculates egg/sperm effort density

How to optimize effort through female role? Theory predicts ndividuals should reallocate effort to egg production when beneficial, but that energy could come from 3 sources: 1. sex allocation (SA) from sperm to eggs penis 2. borrow from future reproduction, lay more eggs in short-term vs. eggs laid Mon Wed Fri Mon Wed Fri if role is favored now

How to optimize effort through female role? Theory predicts ndividuals should reallocate effort to egg production when beneficial, but that energy could come from 3 sources: 1. sex allocation (SA) from sperm to eggs penis 2. borrow from future reproduction 3. allocate energy from wound repair to egg production sex wounds “I feel fine” “Ow”

Benefits of polyandry Females can receive indirect, genetic benefits for offspring when clutches have multiple paternity (polyandry), and: a) your 1st mating partner was just kind of a loser (sleep around = find generally superior male; additive benefits) b) your 1st mate was not a good genetic match for you (he carries the same harmful recessive allele) c) full siblings (similar) compete for the same resources in the womb or egg mass; half-siblings compete less, develop more efficiently or reach a larger size d) fluctuating environment favors more variable offspring as a bet-hedge against unpredictable conditions

Research questions 1. In Alderia modesta and A. willowi, does egg production or offspring fitness benefit from polyandry?  do clutches with multiple sires have larger larvae, or improved hatching success, due to genetic variability?  have slugs evolved to discern when he-she has had more than one mate, and produce more eggs or bigger eggs? polyandrous clutch Mon Wed Fri

Experimental design 1: Benefits of polyandry “Oh great, you again.” (A) monogamy treatment (8 reps) - pairs of field-collected slugs held together for 2 weeks over last 4 days, scored: 1) # eggs per slug per day 2) % hatching success per clutch 3) mean offspring size at hatching (5 larvae per clutch measured) - all slugs weighed; mean wet weight per replicate group included as covariate “Stop poking me.”

A C B D A C D B ‘swinger’ slugs: (B) polyandry treatment - sets of four “swinger slugs” held as 2 pairs, for 2 weeks - swapped partners every 2 days (so each slug had 3 mates) Two trials run for A. willowi; one trial for A. modesta A C B D after 2 days.. A C D B swap

Results 1A: Alderia willowi polyandry # eggs per slug per day # eggs per slug per day monandry wet weight Polyandrous slugs laid ~50% more eggs than monogamous slugs treatment × weight, P = 0.019 trial × weight, P = 0.017

Results 1A: Alderia willowi Mean % hatching success was higher for polyandrous clutches treatment × trial, P = 0.006  effect was strong in Trial 1, weak in Trial 2 % hatching success Mean offspring size at hatching was also significantly greater for polyandrous clutches (F1,18 = 35.65; P = 0.00001)  polyandry resulted in higher offspring fitness, and triggered greater short-term maternal investment

Results 1B: Alderia modesta Polyandrous slugs laid ~30% more eggs than monogamous F1,16 = 4.84, P = 0.042 (no weight effect or interaction) # eggs per slug per day  trend towards higher hatching success for polyandrous clutches (F1,14 = 3.79; P = 0.07)  no effect of polyandry or weight on offspring size

Research questions 1. In Alderia modesta and A. willowi, does egg production or offspring fitness benefit from polyandry? 2. Do benefits of polyandy offset costs of traumatic mating in A. willowi? Prior results showed decreased egg production in larger mating groups (Llaban 2010); could reflect either: (i) SA favoring male role; or (ii) costs of mating  Do slugs recognize polyandry, and adaptively shift energy back to egg production? reallocate energy from eggs to wound repair

Experimental design 2: Cost of mating Field-collected slugs were individually isolated for 2 weeks, except for one to three daily mating bouts of 30 min - monogamy: always paired with same partner - polyandry: rotated daily among 3 partners  eggs masses then collected for 4 d, scoring: - egg production per slug per day - % hatching success - body size (covariate) mating frequency low medium high # of mating bouts per day 1 2 3

Results 2: Cost of mating monogamy: egg production declined as mating frequency increased - cost of mating # eggs per slug per day polyandry: egg production unaffected by mating frequency - opposite of expected SA to male role in larger groups mating bouts / day treatment × mating frequency, P < 0.00001

Results 2: Cost of mating monogamy: egg production declined as mating frequency increased - cost of mating? - predicted SA to male role? # eggs per slug per day polyandry: egg production unaffected by mating frequency mating bouts / day explanation: slugs detect polyandrous conditions, reallocate at least some energy from wound repair to short-term egg production

Research questions 1. In Alderia modesta and A. willowi, does egg production or offspring fitness benefit from polyandry? 2. Do benefits of polyandy offset costs of traumatic mating in A. willowi? 3. Do benefits of polyandy offset inbreeding depression in A. willowi? inbreeding may be a risk early in annual colonization events

Experimental design - exp.2 Experimental design 3: Inbreeding sire Crosses (n = 5 replicates) established clutches of full and half sibs -- slugs reared from different replicate crosses were non-relatives (r = 0) -- reared in pairs (monogamy) or pairs of pairs (polyandry) to reproductive maturity -- after 2 weeks, scored egg production + hatching success dams half sibs (r = 0.25) full sibs (r = 0.5)

Results 3: Polyandry vs. inbreeding half sibs Polyandrous half-sibs laid 10x more eggs than monogamous slugs - stronger response than non-relatives or full sibs full sibs # eggs per slug per day coefficient of relatedness (r) treatment × relatedness, P = 0.004

Results 3: Polyandry vs. inbreeding Hatching success was low for all monogamous slugs Offspring of half sibs, but not full sibs, benefited from polyandry  possibly too little diversity among full sibs for any genetic benefits of % hatching success coefficient of relatedness (r) treatment × relatedness, P = 0.037 treatment × relatedness × weight, P = 0.023

Conclusions Indirect genetic benefits to offspring select for recognition of, and response to, multiple paternity -- slugs acting in female role may steal from future reproduction (or longevity) to capitalize on short-term benefits of polyandry If this response is adaptive, then benefits of polyandry must offset costs of traumatic mating Partially inbred slugs responded most strongly to polyandry -- polyandry may offset costs of monogamy and inbreeding Similar procedure as Exp. 2

in monandrous situations, smaller (non-relative and half-sibs) slugs have higher % dev., where the opposite is true for full siblings

Bateman’s principle & sexual selection Bateman’s Principle holds that males are under sexual selection to optimize offspring quantity (# of mating opportunities), whereas females optimize quality of sperm (genetic contribution to kids) one male can fertilize 10 females one female must choose from among 10 males  leads to familiar array of male display traits, and female choice behaviors... in gonochorists (two sexes)

Female choice: the paradox of cheating A wealth of studies show in most species, females are very picky about which male gets to father their offspring Females may choose: - dominant male - male with traits that are honest indicators of offspring fitness - males with traits that exploit innate preferences One would therefore predict that once paired up with their chosen male, females would tend to be faithful

Cheaters: everybody sucks Male promiscuity is easily understood: maximize quantity But even in “monogamous” species, female promiscuity turns out to be common  in over 70% of bird species, females cheat on their partner  in many species, 20-50% of clutches have more than one biological father (but only one male raises the chicks) - up to 85% cheating in Superb Fairy Wren (M. cyaneus) - only 9% cheating in Shag (P. aristotelis) Polyandry is the term for mating with multiple males

in Panama, the giant false limpet S. gigas forms mated pairs on rocks