Successful mating and hybridisation in two closely related flatworm species despite significant differences in reproductive morphology and behaviour

Abstract

We would like to thank Tim Janicke and Georgina Rivera Ingraham for help with collecting 22 specimens of the study species M. janickei, Steven A. Ramm for providing access to the 23 images of the M. lignano stylets from their study, Jeremias Brand for help with imaging the 24 hybrid morphology, Nikolas Vellnow and Axel Wiberg for helpful discussions, and Gudrun 25 Viktorin, Lukas Zimmerman, Jürgen Hottinger, Urs Stiefel and Daniel Lüscher for technical 26 support. Abstract 33Speciation is usually a gradual process, in which reproductive barriers between two species 34 accumulate over time. Reproductive traits, like genital morphology and mating behaviour, are 35 some of the fastest diverging characters and can serve as reproductive barriers. The free-living 36 flatworm Macrostomum lignano, an established model for studying sex in hermaphrodites, 37 and its congener M. janickei are closely related, but differ substantially in their male 38 intromittent organ (stylet) morphology. Here, we examine whether these morphological 39 differences are accompanied by differences in behavioural traits, and whether these could 40 represent barriers to successful mating and hybridization between the two species. Our data 41shows that the two species differ in many aspects of their mating behaviour, with M. janickei 42 having a five-fold longer copulation duration, copulating less frequently, and having a longer 43 and more delayed suck behaviour (a postcopulatory behaviour likely involved in sexual 44 conflict). Interestingly, and despite these significant morphological and behavioural 45 differences, the two species mate readily with each other in heterospecific pairings, often 46 showing behaviours of intermediate duration. Although both species have similar fecundity in 47 conspecific pairings, the heterospecific pairings revealed clear postmating barriers, as only 48 few heterospecific pairings produced F1 hybrids. These hybrids had a stylet morphology that 49 was intermediate between that of the parental species, and they could successfully backcross 50 to both parental species. Finally, in a mate choice experiment we tested if the worms 51 preferentially mated with conspecifics over heterospecifics, since such a preference could 52 represent a premating barrier. Interestingly, the experiment showed that the nearly two-fold 53 higher mating rate of M. lignano caused it to mate more with conspecifics, leading to 54 assortative mating, while M. janickei ended up mating more with heterospecifics. Thus, while 55 the two species can hybridize, the mating rate differences could possibly lead to higher fitness 56 costs for M. janickei compared to M. lignano. 57 4 Keywords 58 mating behaviour, simultaneous hermaphrodite, mate preference, reproductive isolation, 59 hybridization, genitalia, reproductive barriers, premating barriers, postmating barriers, free-60 living flatworms 61 62 63The biological species concept defines species as groups of individuals that interbreed in 64 nature to produce viable and fertile offspring (Mayr 1...

Faster rates of molecular sequence evolution in reproduction-related genes and in species with hypodermic sperm morphologies

“…Macrostomum are simultaneous hermaphrodites, producing sperm and eggs at the same time, and several species from this genus, particularly the main model species M. lignano (Ladurner et al 2005;Wudarski et al 2020), are used to study mating behaviour, sexual selection, and sexual conflict (e.g. Schärer et al 2005;Ramm et al 2015;Marie-Orleach et al 2017;Giannakara et al 2016;Giannakara and Ramm 2017;Patlar et al 2020;Singh et al 2020aSingh et al , 2020b. In hermaphrodites, sexual conflict is expected to occur not just over the mating and remating rates, but also over the sex roles that the partners assume during copulation, with the male (or "sperm donor") role generally thought to be preferred over the female (or "sperm recipient") role (Charnov 1979;Michiels 1998;Schärer et al 2015).…”

Genome and Karyotype Reorganization after Whole Genome Duplication in Free-Living Flatworms of the Genus Macrostomum

“…However, we should note that the revealed B chromosomes open a new possibility for analysis of amplified DNA repeats in M. mirumnovem. Microdissected DNA libraries can be generated from B chromosomes and then used for NGS sequencing, as was done previously for the Bs in some species of different taxa [45][46][47]. The study of B chromosomes in M. mirumnovem, including the generation of microdissected libraries and their sequencing is in progress.…”

Self-fertilization, but not mating strategy, predicts the evolution of sex allocation plasticity in a hermaphroditic flatworm genus

“…We show that M. lignano and M. janickei, which are sibling species capable of hybridization (Singh et al 2020a), exhibit differences in SA plasticity in response to the strength of LSC (although the 95% confidence intervals of two of the six M. lignano studies overlap somewhat with that of M. janickei). Variation in SA plasticity in species with similar reproductive biology could stem from different environmental conditions experienced by the species.…”