The high biodiversity of benthic organisms in a coastal ecosystem revealed by an integrative approach / A alta biodiversidade de organismos bentônicos em um ecossistema costeiro revelada por uma abordagem integrativa

Abstract Increasing habitat modification and species loss demand consistent efforts to describe and understand biodiversity patterns. The BIOTA/FAPESP Program was created in this context and it has been a successful initiative to promote studies on biodiversity and conservation in Brazil. The BIOTA/Araçá is an interdisciplinary project that provided a detailed evaluation of the biodiversity of Araçá Bay, a coastal seascape located on the North coast of the state of São Paulo, Southeast Brazil. The bay encompasses multiple habitats, such as beaches, mangroves, rocky shores, and a tidal flat, and provides important ecosystem services. Unfortunately, the bay is the subject of complex social-environmental conflicts that oppose economic, social, and environmental demands (i.e., the expansion of neighboring harbor activities vs. small-scale artisanal fisheries and protection of biodiversity). The present study presents a survey of the benthic species occurring in the different habitats of Araçá Bay, including data obtained during the BIOTA/Araçá project and previous assessments of the area. The benthic species play an important role in marine environments and studying the diversity of these organisms that live associated with the bottom is indispensable for comprehending the environment's functioning. The macrofauna, meiofauna, and microorganisms associated with soft and hard bottom were listed, and additional information, such as the habitat and geographical distribution, were provided for each species. The checklist includes 826 species, almost 70% recorded during the BIOTA/Araçá project. The most speciose taxa were the annelids (225 spp.), mollusks (194 spp.), and crustaceans (177 spp.). Seven benthic species are endemic to Araçá Bay, 14 are considered threatened, and seven are economically exploited. Furthermore, the bay is the type locality of many taxa, and 11 new benthic species were described based on specimens sampled during the project. This project shows the importance of Araçá Bay as a unique biologically rich environment and highlights the need for conservation efforts in light of the current threats.

Resumo O aumento da modificação dos habitats e da perda de espécies demanda esforços consistentes para descrever e compreender os padrões de biodiversidade. O programa BIOTA/FAPESP foi criado nesse contexto e é uma iniciativa de sucesso para promover estudos em biodiversidade e conservação no Brasil. O BIOTA/Araçá é um projeto interdisciplinar que promoveu uma avaliação detalhada da biodiversidade da Baía do Araçá, um ecossistema costeiro localizado ao Norte do estado de São Paulo, Sudeste do Brasil. A baía engloba múltiplos habitats, tais como praias, manguezais, costões rochosos, e uma planície de maré, e também fornece importantes serviços ecossistêmicos. Infelizmente, a baía está sujeita à conflitos sócio-ambientais complexos que contrastam demandas econômicas, sociais e ambientais (i.e. a expansão das atividades do porto vizinho vs. a pesca artesanal de pequena escala e a proteção da biodiversidade). O presente estudo apresenta um levantamento das espécies bentônicas que ocorrem nos diferentes habitats da Baía do Araçá, incluindo dados obtidos durante o projeto BIOTA/Araçá e de investigações realizadas anteriormente na área. As espécies bentônicas desempenham um papel importante no ambiente marinho, e estudar a diversidade desses organismos que vivem associados ao fundo é indispensável para compreender o funcionamento do meio ambiente. A macrofauna, meiofauna, e microorganismos associados aos fundos consolidado e inconsolidado foram listados, e informações adicionais foram fornecidas para cada espécie, tais como a distribuição geográfica e nos habitats. O checklist inclui 826 espécies, quase 70% registradas durante o projeto BIOTA/Araçá. Os taxa mais especiosos foram os anelídeos (225 spp.), moluscos (194 spp.), e crustáceos (177 spp.). Entre as espécies bentônicas listadas, sete são endêmicas da Baía do Araçá, 14 são consideradas ameaçadas de extinção, e sete são exploradas economicamente. A baía é a localidade tipo de vários taxa, e 11 novas espécies bentônicas foram descritas com base em espécimes amostrados durante o projeto. Este projeto mostra a importância da Baía do Araçá como um ambiente de riqueza biológica única e demonstra a necessidade de esforços para a sua conservação considerando as atuais ameaças.

Evolutionary dynamics of whole-body regeneration across planarian flatworms.

Regenerative abilities vary dramatically across animals. Even amongst planarian flatworms, well-known for complete regeneration from tiny body fragments, some species have restricted regeneration abilities while others are almost entirely regeneration incompetent. Here, we assemble a diverse live collection of 40 planarian species to probe the evolution of head regeneration in the group. Combining quantification of species-specific head-regeneration abilities with a comprehensive transcriptome-based phylogeny reconstruction, we show multiple independent transitions between robust whole-body regeneration and restricted regeneration in freshwater species. RNA-mediated genetic interference inhibition of canonical Wnt signalling in RNA-mediated genetic interference-sensitive species bypassed all head-regeneration defects, suggesting that the Wnt pathway is linked to the emergence of planarian regeneration defects. Our finding that Wnt signalling has multiple roles in the reproductive system of the model species Schmidtea mediterranea raises the possibility that a trade-off between egg-laying, asexual reproduction by fission/regeneration and Wnt signalling drives regenerative trait evolution. Although quantitative comparisons of Wnt signalling levels, yolk content and reproductive strategy across our species collection remained inconclusive, they revealed divergent Wnt signalling roles in the reproductive system of planarians. Altogether, our study establishes planarians as a model taxon for comparative regeneration research and presents a framework for the mechanistic evolution of regenerative abilities.

Phylogenetic relationships of the Geoplaninae land planarians (Platyhelminthes, Tricladida) assessed with a total evidence approach, with the description of a new species of Gigantea.

Several studies have focused on the phylogenetic relationships within the Geoplaninae land planarians (Tricladida). In those studies, ancient phylogenetic relationships remained obscure. In this work, the phylogeny of Geoplaninae is assessed through three different datasets, namely morphological, molecular, and both datasets combined, i.e, a total evidence approach (TE). The data matrix consisted of six DNA regions, including a newly developed marker (DOM5), and a morphological matrix with 37 characters. The study produced the best-resolved hypothesis so far for the phylogeny of Geoplaninae, although ancient clades still remain elusive. The effect of the morphological data on the TE tree topology and clade support is seemingly negligible. The phylogenetic tree also suggests that most of the diagnostic morphological characters of the genera are homoplastic, while unambiguous unique synapomorphies can characterize some supra-generic informal groupings.

Morphology of parenchymally implanted foreign bodies indicates copulatory wounding in a planarian.

Traumatic mating, that is, copulation that involves wounding the partner's body, is a widespread phenomenon that is particularly prevalent in hermaphroditic animals. Traumatic mating is generally a collateral side effect of diverse strategies (from physical anchorage to injection of substances to manipulate the partner), but the trauma could also be adaptive by itself if it delays remating by the injured partner. In the Tricladida (the clade of planarian flatworms), reciprocal sperm transfer is often assumed to occur by means of a 'regular' nontraumatic copulation, that is, insertion of the penis through the partner's gonopore and deposition of the ejaculate into its genital atrium, with subsequent sperm migration to the oviducts. However, while studying the anatomy of Brazilian land planarians for taxonomic purposes, we found foreign bodies, reminiscent of spermatophores, implanted within the parenchyma of Choeradoplana albonigra (Riester, 1938). Herein, we describe and illustrate several lines of morphological evidence indicating that these foreign bodies likely represent a novel case of intragenital copulatory wounding (e.g., structural and histochemical similarity to land planarians spermatophores; implantation at the level of the gonopore; vestiges of rupture of the genital atrium's wall), corroborating that traumatic mating is pervasive and underreported in Metazoa. We also propose two different hypotheses to explain such copulatory wounding, viz., that it concerns (1) a regular mating strategy or (2) an accidental effect of the copulation. In any event, this land planarian may prove useful as a novel, noninsect terrestrial model organism to investigate the evolution of traumatic mating.

Revision and description of six species of Choeradoplana (Platyhelminthes, Tricladida), with an emendation to the genus.

Living representatives of the Neotropical genus Choeradoplana Graff, 1896 (Geoplaninae, Tricladida, Platyhelminthes) are easily recognized by the typical shape of the head which is laterally expanded, rolled-up, and ventrally provided with two glandular cushions. In this study, the morphology and phylogeny (cytochrome C oxidase subunit I gene) of several species of land planarians are taxonomically investigated. Four of the six species studied are new to science, namely: Ch. eudoxiae Silva & Carbayo, sp. nov., Ch. claudioi Lago-Barcia & Carbayo, sp. nov., Ch. onae Lago-Barcia & Carbayo, sp. nov., and Ch. riutortae Lago-Barcia & Carbayo, sp. nov. The species Choeradoplana albonigra and Ch. eudoxiae deviate from the usual body shape pattern in that the head does not present lateral expansions nor glandular cushions, becoming indistinguishable from its sister genus Cephaloflexa. Pseudogeoplana tristriata (Schultze & Müller, 1857) is also redescribed from a newly collected specimen and was discovered to be a member of Choeradoplana. Graff (1899) also studied another specimen that was considered to be conspecific with P. tristriata; however, in this new it is concluded that it is not conspecific but rather a new species. The name Pseudogeoplana aevipandemiae Lago-Barcia & Carbayo, sp. nov. is suggested for Graff's specimen.

Cataloguing Prof. Eudóxia M. Froehlich's wet collection of land planarians (Platyhelminthes, Geoplanidae): a bounty of type specimens of 78 species.

Zoologist Prof. Eudóxia Maria Froehlich (1929*-2015†) kept an uncatalogued personal collection of land planarians in wet medium. We catalogued their contents. Glass jars contained a variable number of small vials, each with one or several labeled specimens. We transcribed the species names and field data, when available, written on card labels to a table. A total of 1772 vials in 80 jars, contained over 2700 specimens. Subsequently, we cross checked field data with that of the type specimens. As a result, we identified, with different degrees of certainty, type specimens of 78 species. These types compose a very significant potential source of additional morphological details of the species to conform with current standards in the taxonomy of the group.

Five new pseudocryptic land planarian species of Cratera (Platyhelminthes: Tricladida) unveiled through integrative taxonomy.

BACKGROUND: Cratera is a genus of land planarians endemic to the Brazilian Atlantic forest. The species of this genus are distinguished from each other by a series of external and internal characters, nonetheless they represent a challenging taxonomic issue due to the extreme alikeness of the species analysed in the present work. To resolve these difficulties, we have performed morphological analyses and used three nuclear markers (ribosomal 18S and 28S, Elongation Factor, a new anonymous marker named Tnuc813) and two mitochondrial fragments (Cytochrome c oxidase subunit I gene, and a fragment encompasing NADH deshydrogenase subunit 4 gene, trnF and the beginning of the Cytochrome c oxidase I gene) in an integrative taxonomic study. METHODS: To unveil cryptic species, we applied a molecular species delimitation approach based on molecular discovery methods, followed by a validation method. The putative species so delimited were then validated on the basis of diagnostic morphological features. RESULTS: We discovered and described four new species, namely Cratera assu, C. tui, C. boja, and C. imbiri. A fifth new species, C. paraitinga was not highly supported by molecular evidence, but was described because its morphological attributes are unique. Our study documents for the genus Cratera the presence of a number of highly similar species, a situation that is present also in other genera of land planarians. The high number of poorly differentiated and presumably recent speciation events might be explained by the recent geological history of the area.

Areas of endemism of land planarians (Platyhelminthes: Tricladida) in the Southern Atlantic Forest.

Areas of endemism (AoE) are the main study units in analytical biogeographic methods, and are often defined as an area with two or more endemic species living in them, presenting substantial congruence among their range limits. We explored the distribution of land planarians (Geoplanidae, Platyhelminthes) across the southern region of the Brazilian Atlantic forest (from the state of Rio de Janeiro, to the state of Rio Grande do Sul) utilizing DaSilva's et al. (2015) protocol. We used two methods, Endemicity Analysis (EA), and Geographical Interpolation of Endemism (GIE). We identified nine AoE of terrestrial flatworms in the Southern Atlantic forest. Performance of the methodologies is discussed. These AoE of land planarians can be explained through vicariance events combined with their physiological and ecological own limitations. Interestingly, these AoE are congruent with fine-scale approaches such as that with harvestmen. Most land planarians have revealed to present a very small distributional range evidencing their potential as a good model for fine-scale studies of AoE.

X-ray microcomputed tomography applied to the taxonomic study of rare material: redescriptions of seven of Schirch's Brazilian species of land planarians (Geoplanidae, Platyhelminthes).

In 2016, the type-material of ten of the 15 Brazilian land planarians (Platyhelminthes, Tricladida, Geoplanidae) described by Schirch (1929) was discovered deposited in the Museu Nacional do Rio de Janeiro (MNRJ). Schirch only described the external morphology of these species, all originally placed in the genus Geoplana. By the 1930s and 1950s Geoplana itatiayana, G. plana, and G. rezendei underwent taxonomic revision based on the study of non-type specimens. The remaining 12 species also underwent a taxonomic revision but only based on the literature. Current names of these species are Geoplana goettei, Pseudogeoplana arpi, Ps. blaseri, Ps. bonita, Ps. bresslaui, Ps. cardosi, Ps. doederleini, Ps. lumbricoides, Ps. obscura, Ps. riedeli, Ps. theresopolitana, and Ps. wetzeli. The species Geoplana maximiliani sensu Schirch (1929) was renamed as Ps. schirchi Ogren & Kawakatsu, 1990. The present study reports a taxonomic revision of seven of Schirch's species using type material, namely Obama itatiayana, Pasipha plana, Pseudogeoplana arpi, Ps. bresslaui, Ps. doederleini, Ps. schirchi and Ps. wetzeli. Additional specimens of some of these species were also examined. Morphological data from histological preparations and from virtual sections were obtained through a non-destructive technique of X-ray computed microtomography (µCT). This approach resulted in the preservation of the entire body of at least one type-specimen of each species, and the holotype of Ps. bresslaui. Conspecificity of O. itatiayana and P. plana was confirmed, as previously reported in the literature. It is also proposed that Ps. bresslaui belongs to the genus Paraba, while the other species should remain in Pseudogeoplana, since type-specimens are either immature, poorly preserved or simply lost.

A catalogue of Eudóxia M. Froehlich's collection of planarians (Platyhelminthes) with emphasis on type specimens on histological slides.

Leading taxonomist Prof. Eudóxia Maria Froehlich (1929*-2015†) kept an uncatalogued personal collection of a large set of histological glass slides of land planarians. Many of these slides carry insufficient written information for identification purposes. 2287 slides of her collection were examined under the microscope and compared with relevant literature. We found 1633 slides bearing type material of a total of 119 nominal species, mostly collected in the Neotropical region. Identification of 148 slides (belonging to 16 species) as type material is suspected but not confirmed. Type material represents 110 triclad species. There is also type material of two Polycladida, two Rhabdocoela, three Proseriata, one Prolecitophora, and one Acoelomorpha species. The 119 nominal species were described by Ernest Marcus, Eveline du Bois Reymond-Marcus, Diva Diniz Corrêa, Cláudio Gilberto Froehlich, and Eudóxia Maria Froehlich. While keeping the slides in their original location in cupboards, the catalogue provides a quick means of retrieving desired slides. Slides were deposited in the Museu de Zoologia da Universidade de São Paulo.

Taxonomic revision of four species of the Neotropical land planarian genera Cratera and Geoplana (Platyhelminthes, Tricladida) with a description of two new species and an emendation of Cratera.

The Brazilian land planarians Cratera crioula, C. joia, Geoplana hina, and G. taxiarcha (Platyhelminthes, Tricladida, Geoplanidae) are revised taxonomically from type material and additional specimens. Geoplana hina sensu Carbayo et al. (2013) was found to be an undescribed species and therefore is described and named as Cratera picuia sp. n. A new species of the genus is also described and named as Cratera arucuia sp. n. G. hina and G. taxiarcha are transferred to Cratera. The most remarkable morphological feature of Cratera-a dilated terminal portion of the ejaculatory duct-is either absent, inconspicuous, or variable in C. hina, C. joia, C. picuia sp. n., and C. arucuia sp. n. Based on the monophyletic status of Cratera inferred elsewhere, an emendation of the genus is here proposed to encompass the morphological variation observed in the genus.

A new species of the land planarian Issoca sheds light on the polyphyletic status of the genus (Platyhelminthes, Tricladida, Geoplaninae).

A new species of the genus Issoca (Platyhelminthes, Tricladida, Geoplaninae) is described. Issoca assanga sp. n. presents the diagnostic features of the genus, with the exception of the relative position of the subneural parenchymal muscle layer with the cephalic retractor muscle, which are overlapped in the type species of the genus but are intersected in the new species. Rather than a polymorphic character, the relative position of these muscle layers might reflect the polyphyletic status of the genus.

Micro-computed tomography scan and virtual histological slide data for the land planarian Obama otavioi (Platyhelminthes).

BACKGROUND: We investigated whether images obtained through X-ray micro-computed tomography (µCT) can be used in conjunction with traditional methods for morphological studies of soft-bodied land planarians. µCT is non-invasive and provides true-to-scale three-dimensional imagery at high resolution. We compared µCT-based images of a recently described land planarian species of Obama otavioi (Platyhelminthes) with those obtained from light microphotography of histological sections, most of which were also digitized at high magnification. FINDINGS: The specimens studied were collected in 2012. Subsequent µCT-based images of the stained body of a paratype show nearly all morphological features provided by traditional histology, with the exception of particularly minute structures, smaller than 5 µm, such as the sensory pits and single muscle fibers, which are best visible on traditional histological sections. Because the technique is non-destructive, the scanned specimen is preserved without damage. The raw and derivative µCT data and virtual histological sections are freely available in GigaDB. CONCLUSIONS: The µCT datasets of these stained soft-bodied organisms reveal images of external and internal structures that support previous taxonomic studies. This technique can be particularly important for non-destructively revealing internal details of whole museum specimens at a faster rate than histology alone. High-resolution virtual histological slides also allow further searches for new, previously unstudied morphological features. The use of X-ray equipment with higher resolution can enable smaller sensory organ and muscle fiber details to be seen. The image sets, µCT-based images and digitized histological slides can be disseminated without the constraints of specimen loans.

Evolution of flatworm central nervous systems: Insights from polyclads.

The nervous systems of flatworms have diversified extensively as a consequence of the broad range of adaptations in the group. Here we examined the central nervous system (CNS) of 12 species of polyclad flatworms belonging to 11 different families by morphological and histological studies. These comparisons revealed that the overall organization and architecture of polyclad central nervous systems can be classified into three categories (I, II, and III) based on the presence of globuli cell masses -ganglion cells of granular appearance-, the cross-sectional shape of the main nerve cords, and the tissue type surrounding the nerve cords. In addition, four different cell types were identified in polyclad brains based on location and size. We also characterize the serotonergic and FMRFamidergic nervous systems in the cotylean Boninia divae by immunocytochemistry. Although both neurotransmitters were broadly expressed, expression of serotonin was particularly strong in the sucker, whereas FMRFamide was particularly strong in the pharynx. Finally, we test some of the major hypothesized trends during the evolution of the CNS in the phylum by a character state reconstruction based on current understanding of the nervous system across different species of Platyhelminthes and on up-to-date molecular phylogenies.

Anatomical deviation of male organs of land planarians from Rio de Janeiro, Brazil, with description of two new species of Cratera (Platyhelminthes, Tricladida).

Two new land planarian species, collected in the State of Rio de Janeiro, Brazil, are described. Their external aspect is similar to that of Imbira marcusi Carbayo et al., 2013 and Pseudogeoplana theresopolitana (Schirch, 1929), respectively. The analysis of the internal organs, however, revealed they belong to the genus Cratera. The male copulatory organs of one species is very different from any other geoplaninid, for the penis papilla holds a large, distal cavity receiving the ejaculatory duct and, furthermore, the papilla projects vertically downwards from the roof of the male atrium. Thus we consider it as a new species, Cratera cuarassu sp. nov. The second species differs from its congeners in that the dorsal insertion of the penis papilla is anterior to the ventral one, and in that the female atrium is narrowed in the anterior portion. The species was found in the type locality of Pseudogeoplana theresopolitana (Schirch, 1929) and compares well with it in the external features. However, since its internal organs are unknown and the type material of the species is seemingly lost, we describe it as Cratera anamariae Carbayo, sp. nov.

Catálogo dos "Turbellaria" (Platyhelminthes) do Estado de São Paulo / Checklist of "Turbellaria" (Platyhelminthes) from São Paulo State, Brazil

Mediante consulta bibliográfica e de bases de dados encontramos registros para o estado de São Paulo de 312 espécies de turbelários (incluindo Acoelomorpha, novo filo) de ambientes marinhos, límnicos e terrestres. Em 1999 haviam sido registradas aproximadamente 333 espécies; o número maior deve ser atribuído a espécies posteriormente sinonimizadas e, provavelmente, à precisão menor dos cômputos anteriores. Os dois únicos pesquisadores especialistas do grupo, trabalhando no estado, estimam, porém, um número muito maior. Há três coleções científicas abrigando turbelários, quase exclusivamente, terrestres.

A recent survey of the literature and databases on turbellarian fauna from the State of São Paulo, Brazil, yielded a total of 312 species (including Acoelomorpha, a new phylum) inhabiting marine, as well as freshwater, and terrestrial habitats. In 1999, approximately 333 species were registered. This higher number is related to a number of species synonymized thereafter, and probably related to lower accuracy of prior accounts. Nonetheless, the only two taxonomists studying this animal group in the State estimated a much higher actual number of species. In the State there are three scientific collections containing turbellarians, almost exclusively from terrestrial habitats.

Estado do conhecimento dos macroturbelários (Platyhelminthes) do Brasil: [revisão] / State of knowledge of the macroturbellarians (Platyhelminthes) from Brazil: [review]

Nas últimas décadas tem crescido o esforço internacional para conhecer a biodiversidade mundial visando a criação de políticas públicas de conservação. O Brasil vem participando deste esforço, como evidenciam as recentes publicações sobre o estado do conhecimento de vários táxons de organismos. Os macroturbelários (Polycladida e Tricladida, Platyhelminthes) fazem parte da fauna mundialmente menos estudada. Neste trabalho inventariamos por regiões a diversidade da fauna brasileira conhecida de macroturbelários e comparamos esta diversidade com a do resto do mundo. Analisamos também a atividade dos taxonomistas brasileiros do grupo no contexto mundial. Os resultados mostram que praticamente a totalidade das 246 espécies nominais descritas procede das regiões Sudeste e Sul. O número de espécies deverá aumentar significativamente quando forem feitas novas coletas, tanto em biomas e regiões ainda pouco ou nada explorados quanto nas áreas já amostradas. O Brasil está, relativamente a outros países, bem provido de especialistas. No entanto, em face da grande diversidade estimada, as amostragens deveriam ocorrer concomitantemente com a formação de novos especialistas.

In the last decades, international efforts have increased, with the aim of acquiring greater knowledge on worldwide biodiversity and to propose adequate conservation policies. Brazil has joined in these efforts, as is shown by recent publications on the state of knowledge of several taxa of organisms. The macroturbellarians (Polycladida and Tricladida, Platyhelminthes) are part of one of the less studied faunal groups, not only in Brazil but also in other regions of the world. In the present study we inventoried the knowledge on the diversity of Brazilian macroturbellarians and make a comparison with that from the rest of the world. We also analyze the accomplishments of Brazilian taxonomists dealing with macroturbellarians, with regard to the world context. Our results show that almost all of the 246 described species are from the South-eastern and Southern regions. The number of species will increase significantly when new samples are undertaken in biomes and regions still little or non-sampled, as well as in already explored areas. Brazil is relatively well provided with specialists in relation to other countries. Nonetheless, in view of the high numbers of estimated diversity, new samples should concur with the academic formation of new taxonomists.