Morphological and molecular characterization of brown-banded broodsacs and metacercariae of Leucochloridium (Trematoda: Leucochloridiidae) parasitizing the semi-slug Omalonyx unguis (Succineidae) in Argentina.

Trematodes of the genus Leucochloridium exhibit an unusual transmission strategy among mollusks (intermediate host). The fully developed sporocyst, housing encysted metacercariae, displays vivid coloration and rhythmic activity in the snail's tentacle, mimicking insect larvae. These strategies attract insectivorous birds, their final hosts, thereby increasing the chances of completing their life cycle. In South America, the reports of adults and larval stages of Leucochloridium are scarce. Brown-banded broodsac of Leucochloridium sp. were obtained from Omalonyx unguis collected in a shallow lake from Corrientes Province, Argentina. Here, we morphologically characterized the larval stages (broodsac and metacercaria), identified the parasite through DNA sequences from nuclear 28S-rRNA (28S) and the mitochondrial cytochrome c oxidase I (COI) genes, and explored its evolutionary affinities with the Leucochloridium species available in GenBank. The present broodsac displays brown bands, with a yellowish background in the first two-thirds and yellowish-white in the last third. Based on morphological comparisons, the broodsac and metacercaria described in this study could not be conclusively categorized under any known South American species of Leucochloridium. In relation to the phylogenetic reconstructions, Leucochloridium sp. consistently clustered with L. perturbatum, and species delimitation analyses resulted in recognized Leucochloridium sp. from Argentina as a distinct species. The DNA sequences obtained in this study constitute the first genetic data generated for sporocyst broodsacs in South America. Future studies, incorporating morphology, genetic, and biological data, will be essential for both species identification and the elucidation of leucochloridiid diversity in the region.

Hidden diversity in waterfall environments: The genus Acrorbis (Gastropoda: Planorbidae) from the Upper-Paraná Atlantic Forest.

High-energy freshwater environments such as rapids and waterfalls in the Upper-Paraná Atlantic Forest are home to highly endemic minute freshwater snails of the genus Acrorbis. Only one species, Acrorbis petricola, is currently included within this genus, whose geographical distribution is restricted to three known populations, one in Brazil and the other two in Argentina. Because of habitat specificity and limited geographical distribution, the species is considered vulnerable in Argentina and endangered in Brazil. In this work, we identify five new populations of A. petricola in southern Upper-Paraná Atlantic Forest, exclusively found on waterfalls from the Misiones Province, Argentina. Based on these populations and on specimens of one of the two historical populations from the Misiones Province, we explored the morphological features of shells and reproductive system of specimens from each location and provide the first molecular data on the species. We used DNA sequences from cytochrome c oxidase subunit I (COI) and 16S-rRNA genes to investigate the molecular diversity, genetic distances and genealogical relationships among populations. We verified the existence of intra- and interpopulation morphological variability, with the greatest variation being found in spire, spiral sculpture, penis sheath, flagella, prostatic diverticula and bursa copulatrix. We found interpopulation genetic diversity, with no intrapopulation variation, and identified six geographically structured genetic lineages with maximum genetic distances of up to 2.3%. Different combinations of morphological characters with the same genetic background within each locality were observed. The finding of new populations genetically differentiated not only broadens the known distribution of the species, but also illustrates that waterfall environments in the Atlantic Forest harbour a hidden diversity of Acrorbis that still remains to be discovered. This scenario suggests a complex evolutionary history that needs to be unveiled and taken into account for future development of conservation strategies in this endemic genus.

The exotic jumping snail Ovachlamys fulgens (Gude, 1900) (Gastropoda: Helicarionidae) in urban areas of the Upper-Paraná Atlantic Forest.

The exotic land gastropod Ovachlamys fulgens (Gude, 1900), aka the jumping snail, was registered in Eldorado city, Misiones province, constituting the first documented record of that species in Argentina-and for all of South America as well. Identities of the individuals were confirmed through morphological examination, and by a DNA sequencing of the cytochrome oxidase subunit I gene. DNA sequences for the mitochondrial 16S-rRNA and the nuclear 18S-rRNA genes were also obtained from the morphologically-confirmed specimens to gain insights into the genetic background of this species in Argentina, and as additional markers for enhancing the rapidity of identification by governmental authorities who are responsible for managing the presence of exotic species. Different size classes suggested that the species had successfully reproduced and become established in the locality. Ovachlamys fulgens is emphasized in the literature as being a serious pest to orchids. Misiones province harbors a high species richness of orchids, representing about one half of the known species in Argentina. Owing to the use of the orchid as an ornamental plant, the commercial trade could have facilitated the introduction of the species into Argentina. Control measures, preventive education, and surveillance are needed to prevent the further spread of this snail.

Insights into the Evolutionary History of an Extinct South American Freshwater Snail Based on Historical DNA.

Highly oxygenated freshwater habitats in the High Paraná River (Argentina-Paraguay) were home to highly endemic snails of the genus Aylacostoma, which face extinction owing to the impoundment of the Yacyretá Reservoir in the 1990s. Two species, A. chloroticum and A. brunneum, are currently included in an ongoing ex situ conservation programme, whereas A. guaraniticum and A. stigmaticum are presumed extinct. Consequently, the validity and affinities of the latter two have remained enigmatic. Here, we provide the first molecular data on the extinct A. stigmaticum by means of historical DNA analysis. We describe patterns of molecular evolution based on partial sequences of the mitochondrial 12S ribosomal RNA gene from the extinct species and from those being bred within the ex situ programme. We further use this gene to derive a secondary structure model, to examine the specific status of A. stigmaticum and to explore the evolutionary history of these snails. The secondary structure model based on A. stigmaticum revealed that most polymorphic sites are located in unpaired regions. Our results support the view that the mitochondrial 12S region is an efficient marker for the discrimination of species, and the extinct A. stigmaticum is recognized here as a distinct evolutionary genetic species. Molecular phylogenetic analyses revealed a sister group relationship between A. chloroticum and A. brunneum, and estimated divergence times suggest that diversification of Aylacostoma in the High Paraná River might have started in the late Miocene via intra-basin speciation due to a past marine transgression. Finally, our findings reveal that DNA may be obtained from dried specimens at least 80 years after their collection, and confirms the feasibility of extracting historical DNA from museum collections for elucidating evolutionary patterns and processes in gastropods.