Evolutionary morphology of ligophorus spp. (monogeneadactylogyridae): a geometric morphometrics approach

Resumen

Ligophorus spp. (Monogenea: Dactylogyridae) are common parasites of grey mullets from the Mediterranean Sea, the Black Sea and Sea of Azov (Sarabeev et al. 2013), which display remarkable morphological variation in their attachment organ. Thus, they offer an excellent model for investigating to which extent the phenotypic variability of the attachment organs among congeners is related to host specificity, environmental components, host-parasite coevolution and phylogeny (Vignon et al. 2011). In fact, monogeneans in general provide good models for studying morphological variation in attachment organs to test evolutionary constraints on shape (Šimková et al. 2001). Geometric morphometrics is currently considered as an ideal tool to tackle these problems (Slice, 2007), but it has been hardly applied to Monogenea and other parasites at large. In this thesis, we developed a geometric morphometrics framework, combined with multivariate statistics, to explore intra e interspecific morphological variations in shape and size haptoral structures in 14 species of Ligophorus from 6 grey mullet species. In addition, we evaluated, in the context of phylogenetically independent contrasts (PIC), the evolutionary modularity and morphological integration between units in the haptor (i.e. roots and points) across species. The modules in the haptor structures were identified, providing first insight into how these structures are morphologically organized, how they vary and how they evolve. Consequently, we explored to which extend modular structure in the anchors of Ligophorus were accounted for adaptive and phylogenetic factors acting at different levels. The following specific objectives were addressed: 1) To analyze the variability in shape and size of the dorsal and ventral anchors of Ligophorus cephali from Mugil cephalus, by means of geometric morphometrics and multivariate statistics, to assess the morphological integration between anchors and between roots and points. 2) To describe a new species of Ligophorus from M. cephalus from the Yucatán Peninsula, Mexico and to update the zoogeography of Ligophorus spp. in the light of current evidence for a complex of cryptic species. 3) To determine whether variation in the anchor shapes in 14 Ligophorus spp. is modular and integrated after evaluation of four hypotheses of modularity at both, adaptive and evolutionary levels. 4) To assess phylogenetic signal in form of ventral and dorsal anchors of 14 species of Ligophorus occurring on grey mullets from the Mediterranean, the Black Sea and Sea of Azov, to establish whether similarity in anchor form is explained by convergence or shared evolutionary history. As a synthesis of this PhD study, the following findings and conclusions were drawn: This study documented for first time the phenotypic plasticity and morphology of haptoral structures in Ligophorus cephali on Mugil cephalus. The pattern of shape variation observed was similar in ventral and dorsal anchors, with narrow and elongated anchors and short anchors. Interestingly, localised shape variation was much higher in the dorsal anchors, which is in line with the higher residual variation associated with dorsal anchors in the shape models. Moreover, in the size models the residual variation of the dorsal anchors was much higher than those of ventral anchors. In addition, we demonstrated that random effects (gill section × host individual) were an important determinant of shape in ventral, but no in dorsal anchors, and size models of dorsal and ventral anchors were clearly different. These differences between dorsal and ventral anchors in both shape and size perhaps reflect different functional roles in the attachment to the gills. In addition, the morphology in Ligophorus of haptoral structures dissimilar in shape and size revealed that curvature of the dorsal anchor can vary sharply, suggesting that the dorsal anchor/bar complex is more mobile than ventral one in this genus. This suggests a tighter control of the shape and size of ventral hanchors to fit the characteristics of the individual host microenvironment. We observed high morphological integration in shape between ventral and dorsal anchors in L. cephali suggesting strong coordination in the parts of the haptoral structure which reveals that the shapes are not independent characters and could be driven by host specificity. In addition we observed integration within parasite anchors (point and roots) in both anchors. This high level of morphological integration indicates a concerted action between anchors and suggests that a large fraction of the observed phenotypic variation does not compromise the functional role of anchors as levers. We found that gill arch was an important determinant of anchor shape and size in L. cephali. The phenotypic plasticity in anchor morphology can reflect the ability of individuals of this species to colonise a new host, responding quickly to varying environmental conditions. In order to address diversity within this genus in other geographical areas, a new species of Ligophorus, Ligophorus yucatanensis, from the gills of the flathead M. cephalus from México was described. This species was differenciated from all other species of Ligophorus by the morphology of the accessory piece of the copulatory organ. In addition the new species was distinguished by the morphology of the haptoral ventral bar and the distal end of the vaginal duct. In addition, the ventral anchors were shorter than those of all other species of Ligophorus reported in the Gulf of Mexico. The new species resembles more closely species from the Mediterranean Sea and off the coast of the Northwestern Pacific than those recorded in South and North America. Ligophorus yucatanensis was included into Entity 4 (Western North Atlantic) according to zoographic records of Ligophorus spp. from the M. cephalus species complex. We tested four different hypotheses of modularity in the haptoral anchors of 14 monogeneans species of Ligophorus evaluated by geometric morphometrics and phylogenetically independent contrasts (PIC). The roots and points represented two modules in the dorsal and ventral anchors, but modularity was not statistically supported when parasite phylogeny was accounted for, which indicated convergent evolution probably related to host characteristics and gill morphology. Moreover, PIC revealed medial and lateral modules in ventral anchors only. In contrast we found evidence for ventral and dorsal anchors pairs forming two modules, supporting the notion that they play different functional roles. Integration between all identified modules was strong. So, there is modular structure in the anchors of Ligophorus spp. accounted by adaptive and phylogenetic factors acting at different levels, and ventral and dorsal anchors evolve as integrated modules with specific roles in the attachment. The patterns of morphological change in haptoral anchors were interpreted to reconstruct the dynamics of the evolutionary processes and visualized as paths from ancestors to descendants through the phylomorphospace. The tests performed for phylogenetic signal provided strong evidence for evolutionary processes playing a major role in determining the shape and, to a lesser degree, the size of the haptoral anchors. The position in the phylomorphospace of distantly related species co-occurring on a given host species were very different: L. confusus and L. imitans parasitizing Liza ramada represent different clades and the anchors fell far apart in the shape and size morphospaces. L. szidati and L. vanbenedenii co-occurring on Liza aurata were placed in different clades, differed in shape and size of the dorsal anchors. So, no clear evidence for homoplasy in the Ligophorus spp. studied was found. In several cases, members of clades that occur in the same host species showed similar anchor forms (L. cephali-L. mediterraneus on M. cephalus, L. llewellyni and L. pilengas on Liza haematocheila) or similar shapes (L. acuminatus and L. minimus on Liza saliens). These clades probably resulted from several intra-host duplication events and their morphological similarities point to occurrence of phylogenetic constrains on anchor form. The phylogenetic position of L. imitans showed affinities with species found on Lz. saliens, suggesting that its occurrence on Lz. ramada represents a host-switch. The adaptation to a new host did not impose strong changes in haptoral anchor morphology and supports the notion of phylogeny being a major determinant of anchors morphology in Ligophorus. Thus, the variation of shape and size of ventral and dorsal anchors in 14 species of Ligophorus is largely determined by common descent and shared evolutionary history. Although homoplasy dictated by adaptations to the host or to specific gill microhabitats could not be ruled out completely, its role seemed less important.