First record of a sea anemone of genus Bunodosoma with two oral discs: a clue to asexual reproduction?

: Different reproducti ve strategies (both sexual and asexual) that may occur in individuals of the same species may explain the success of sea anemones (Cnidaria:Actiniaria) in colonizing different ecosystems around the planet. Here we provide the fi rst record of a Bunodosoma specimen with two oral discs and discuss the possibility of asexual reproduction in this genus. One individual of Bunodosoma cangicum was found in Carneiros beach (Pernambuco state, Brazil) with two fully individualized oral discs but with column and pedal disc still connected. Our fi nding may represent a clue to longitudinal fi ssion in Bunodosoma . Yet, despite the number of different studies carried out so far, the absence of data regarding asexual reproduction in Bunodosoma suggests that it might be rare in this genus, occurring only in sporadic cases such as recovery from injury.


INTRODUCTION
Sea anemones (Cnidaria: Actiniaria) are a successful group in terms of spatial distribution (Fautin et al. 2013).They have colonized waters in different latitudes and are common in shallow and deep waters (González-Muñoz et al. 2016, Sumida et al. 2016).One strategy that may have facilitated such success is the ability to reproduce asexually, as it is rather common among Actiniaria (Bocharova & Kozevich 2011) and may provide adaptive advantages to marine invertebrates, like population growth, maintenance of adapted genotypes and resistance to desiccation (Ottaway 1973, McGovern 2003, Oliveira & Gomes 2005).
Although asexual reproduction does not favor genetic variability, it seems to be an important mean by which animals compete for space and respond to fi ne-scale environmental heterogeneity (Williams 1975a, Sherman & Ayre 2008).Sea anemones reproduce asexually in a diversity of modes, including longitudinal fi ssion, pedal laceration, transversal fi ssion, and tentacle detachment (Bocharova & Kozevich 2011).Longitudinal fi ssion seems to be the most common in Actiniidae, as it occurs in Anthopleura and Anemonia (Shick 1991, Gomes et al. 2003).The genus Bunodosoma Verrill, 1899 is present worldwide and although belonging to Actiniidae, has no record of asexual reproduction.Here we provide the fi rst record of a Bunodosoma cangicum with two oral discs and discuss asexual reproduction in the genus.Belém & Preslercravo, 1973, was found in September 2018, at Carneiros beach in Pernambuco state, Brazil (08°42'15.6"S,035°04'43.3"W),a rocky beach with a daily regime of two low and two high tides.At low tide, B. cangicum individuals can be easily observed fixed on consolidated substrates in tide pools, sharing space with different organisms such as algae turfs, corals, zoanthids, crustaceans and echinoderms.Water temperature was approximately 31°C and the salinity was 37 at the time of observation.

One individual of Bunodosoma cangicum
Due to the position on the reef, we were unable to remove the individual from the rock.Therefore, to verify that it was undergoing a longitudinal fission process, we removed the debris that were partially covering its column and induced contraction via mechanical contact, to visualize column more clearly.Since B. cangicum has non-adhesive vesicles on its column, removal of sediment was simple and did not harm the individual.

RESULTS
The observed individual of B. cangicum had two fully individualized oral discs with two mouths and complete cycles of tentacles.Both the column and the pedal disc were still fully connected.Therefore, this suggests that the individualization of the two future specimens (i.e.parental and clone) must begin at the distal portion of the individual.Since the oral discs were approximately the same size, it was not possible to recognize the parental organism (Figure 1).

DISCUSSION
Our finding suggests the possibility of asexual reproduction in Bunodosoma, which is until now, unknown to occur in this genus.Although the presence of two oral discs in an individual could be the result of recovery process from an injury (Hand & Uhlinger 1995), this could also be related to asexual reproduction by longitudinal fission.Previous studies have reported anatomical irregularities and external scars in representatives of Bunodosoma (e.g.Daly 2004, Barragán et al. 2019), which indicated that asexual reproduction could occur in the genus.
Currently, there is little evidence on the phylogenetic signal of longitudinal fission within Acnitiinidae or any genera of the family.Geller et al. (2005) proposed that in Anthopleura, longitudinal fission has arisen independently at least four times.However, Anthopleura was regarded as a very problematic genus and the most recent study have shown that it is in fact paraphyletic (Daly et al. 2017).Therefore, the potencial homoplasy of longitudinal fission found by Geller et al. (2005) may be an artifact of the sampling or the method.Therefore, until the relationships within the genus are properly understood, evolutive studies regarding the occurrence of asexual reproduction in Anthopleura may not be considered conclusive.That is the same case for Bunodosoma, which was also found to be paraphyletic by Daly et al. (2017).This result along with the little data regarding the biology of Bunodosoma species hinders any attempt to understand the existence of a phylogenetical signal of asexual reproduction in the genus.Even though our finding is a strong indication on the occurrence of longitudinal fission, at least in B. cangicum, we cannot imply further based on the current scenario.
Longitudinal fission is considered an efficient method of asexual reproduction, because daughters can perform the functions of an adult right after separation (Bocharova & Kozevich 2011).Like other asexual processes, longitudinal fission contributes to population increase and maintenance when occurring frequently (Ottaway 1973).If this is the case for B. cangicum, it is not possible to infer frequency based on a single observation.However, such behavior may favor species in environments with oscillating conditions, like intertidal zones, where this strategy could maintain the genotype that is already established in the environment (e.g.Williams 1975b, Sherman & Ayre 2008).
Among species that occur in Brazil, longitudinal fission has been described in detail for Bellactis ilkalyseae Dube, 1983, andAnthopleura varioarmata by Belém &Monteiro (1981), a species later identified as Anthopleura dalyae González-Muñoz, Garese & Acuña, 2019.In both cases, it started at the pedal disc (proximal end) and resulted in individuals of slightly different sizes (Belém & Monteiro 1981, Oliveira & Gomes 2005).In the individual of B. cangicum observed, fission was not yet concluded but the individualized oral discs were of similar size, indicating that the separation seems to begin at the oral disc (distal end).Among sea anemones, fission usually begins at the proximal end, but for some species such as Metridium senile (Linnaeus, 1761) the process begins at the distal end (Bocharova & Kozevich 2011).
Despite its abundance along the Brazilian coast there are few studies on B. cangicum and various aspects of its basic biology are still unknown (Gomes et al. 1998, Beneti et al. 2019).Further studies including field observations, histological data and in-lab induced injuries to observe regeneration process are required to confirm the occurrence of asexual reproduction in Bunodosoma.Recording information on reproductive strategies of species of superfamily Actinioidea will enable us to test whether these characters are informative to understand the evolution of asexual reproduction withing Actiniidae.This may help to clarify taxonomic issues, especially in paraphyletic genera like Bunodosoma (Daly et al. 2017).

Figure 1 .
Figure 1.Individual of Bunodosoma cangicum during the process of longitudinal fission.a: Both clones with open oral disc.b: After mechanical contact, individual two (i2) contracted, while individual one (i1) kept oral disc fully open.As can be seen, both individuals are still connected in the middle column (arrow).Scale bars: 50mm.