Population Genomics and Phylogeography of a Clonal Bryophyte With Spatially Separated Sexes and Extreme Sex Ratios

Abstract

The southern Appalachian (SA) is one of the most biodiversity−rich areas in North America and has been considered a refugium for many disjunct plant species, from the last glacial period to the present. Our study focuses on the SA clonal hornwort, Nothoceros aenigmaticus J. C. Villarreal & K. D. McFarland. This hornwort was described from North Carolina and is widespread in the SA, growing on rocks near or submerged in streams in six and one watersheds of the Tennessee (TR) and Alabama (AR) Rivers, respectively. Males and female populations occur in different watersheds, except in the Little Tennessee (TN) River where an isolated male population exists ca. 48 km upstream from the female populations. The sex ratio of 1:0 seems extreme in each population. In this study, we use nuclear and organellar microsatellites from 250 individuals from six watersheds (seven populations) in the SA region and two populations from Mexico (23 individuals). We, then, selected 86 individuals from seven populations and used genotyping by sequencing to sample over 600 bi-allelic markers. Our results suggest that the SA N. aenigmaticus and Mexican plants are a nested within a clade of sexual tropical populations. In the US populations, we confirm an extreme sex ratio and only contiguous US watersheds share genotypes. The phylogenetic analysis of SNP data resolves four clusters: Mexican populations, male plants (Little Pigeon and Pigeon river watersheds) and two clusters of female plants; one from the Little Tennessee and Hiwassee Rivers (TR) and the other from the Ocoee (TR) and Coosa (AR) Rivers. All clusters are highly differentiated (Fst values over 0.9). In addition, our individual assignment analyses and PCAs reflect the phylogenetic results grouping the SA samples in three clades and recovering males and female plants with high genetic differentiation (Fst values between 0.5 and 0.9 using microsatellites and bi-allelic markers). Our results point to Pleistocene events shaping the biogeographical pattern seen in US populations. The extreme sex ratio reflects isolation and highlights the high vulnerability of the populations in the SA.

Current and historical factors drive variation of reproductive traits in unisexual mosses in Europe: A case study

“…Despite some alternative processes that could also contribute to the tendency above (e.g., current climatic differences between long-and short-term persistent populations, or differences in their phylogeographic history), this result is remarkable and has been suggested for other bryophytes based on genetic data and considering the glacial periods of the Pleistocene (Alonso-García et al, 2020). Since our "young populations" have been potentially established since the LGM (from 19 000 years BP onwards), the persistence of such a historical signal suggests that shifts in sex ratios could operate at the scale of thousands of years (Cronberg, 2000;Alonso-García et al, 2020;Bisang et al, 2020;Blackstock, 2020). Recent evidence by Hedenäs et al (2021), supports the hypothesis of a gradual shift in population sex ratio from balanced toward skewed sex ratios.…”

“…The rationale behind this approach is that populations resulting from postglacial recolonization were expected to be younger than populations in glacial refugia. Thus, we expected a highly skewed distribution of phenotypic sex ratio (i.e., a larger proportion of females) in glacial refugia due to the isolation and the concomitant longer period to accumulate the effects of any sex‐specific competitive advantage (i.e., sex‐specific differences in vegetative growth), than in areas that had been recolonized during the current interglacial period (Alonso‐García et al, 2020). Glacial refugia were estimated assuming that the current northern limit of the species distribution reflects its thermal limit.…”

“…In turn, we predicted that expressed females should be consistently larger than expressed males due to higher pre‐zygotic resource allocation into sexual reproduction for males, and, as a result, relatively higher female investment into vegetative growth ( H 4). Sex‐specific differences in reproductive investment, leading to higher pre‐zygotic female vegetative growth rates, may also contribute to phenotypic sex ratio variation. If H4 held true, we expected to find a historical signal in the broad‐scale geographic pattern of phenotypic sex ratio variation (Alonso‐García et al, 2020). Accordingly, recently founded populations (i.e., after post‐glacial recolonization) were expected to show more variable sex ratios than long‐term persistent populations (i.e., glacial refugia), wherein we expected higher female biases due to the longer drift in sex ratio via asexual reproduction ( H 5).…”

“…Interestingly, unisexual bryophyte populations studied so far have been characterized by either low or highly variable sex expression levels (e.g., Bowker et al, 2000;Bisang & Hedenäs, 2005;Stark et al, 2005a) and strongly biased phenotypic sex ratios, usually toward females (Stark, 2002;Bisang & Hedenäs, 2005;Stark et al, 2005aStark et al, , 2010. Consequently, these populations typically rely on asexual reproduction (During, 1979;Newton & Mishler, 1994;Frahm, 2007, but see Crawford et al, 2009); when this occurs through clonal propagation and/or fragmentation, it could increase the spatial segregation of sexes, and hamper the chances of sexual reproduction (Alonso-García et al, 2020). Nonetheless, clones can also become more strongly intermixed over time, increasing the chance for sex expressing male and female clones to grow within fertilization distance (Cronberg, 2002;Hedenäs et al, 2021).…”

The consequences of mating system and dispersal potential on the genetic structure of leaf-inhabiting bryophyte metapopulations in a fragmented Amazonian landscape

“…In bryophytes, as observed herein for R. flaccida , asexual reproduction plays an important role in maintaining genetic diversity over time by preventing local genotype extinction and assuring successful sexual reproduction (Alvarenga et al, 2016). Evidence from unisexual bryophytes found that genetic diversity is maintained when species stakes on asexual reproduction for their survival (Pohjamo et al, 2008;Wang et al, 2013;Holá et al, 2015;Escolástico-Ortiz et al, 2023), but contrasting evidence was found for a hornwort species (Alonso-Garcia et al, 2020).…”