Molecular diversity within the genus Laeonereis (Annelida, Nereididae) along the west Atlantic coast: paving the way for integrative taxonomy

The polychaete genus Laeonereis (Annelida, Nereididae) occurs over a broad geographic range and extends nearly across the entire Atlantic coast of America, from the USA to Uruguay. Despite the research efforts to clarify its diversity and systematics, mostly by morphological and ecological evidence, there is still uncertainty, mainly concerning the species Laeonereis culveri, which constitutes an old and notorious case of taxonomic ambiguity. Here, we revised the molecular diversity and distribution of Laeonereis species based on a multi-locus approach, including DNA sequence analyses of partial segments of the cytochrome c oxidase subunit I (COI), 16S rRNA, and 28S rRNA genes. We examined Laeonereis specimens collected from 26 sites along the American Atlantic coast from Massachusetts (USA) to Mar del Plata (Argentina). Although no comprehensive morphological examination was performed between different populations, the COI barcodes revealed seven highly divergent MOTUs, with a mean K2P genetic distance of 16.9% (from 6.8% to 21.9%), which was confirmed through four clustering algorithms. All MOTUs were geographically segregated, except for MOTUs 6 and 7 from southeastern Brazil, which presented partially overlapping ranges between Rio de Janeiro and Sao Paulo coast. Sequence data obtained from 16S rRNA and 28S rRNA markers supported the same MOTU delimitation and geographic segregation as those of COI, providing further evidence for the existence of seven deeply divergent lineages within the genus. The extent of genetic divergence between MOTUs observed in our study fits comfortably within the range reported for species of polychaetes, including Nereididae, thus providing a strong indication that they might constitute separate species. These results may therefore pave the way for integrative taxonomic studies, aiming to clarify the taxonomic status of the Laeonereis MOTUs herein reported.This work was supported by the FAPESP (Grants n~ 2011/50317-5, 2015/25623-6, 2017/06167-5, 2018/10313-0) and CNPq through a productivity grant to A.C.Z.A (301551/2019-7). Marcos AL Teixeira was supported by a PhD fellowship (SFRH/BD/131527/2017) from FCT. Pedro Vieira was supported by a Post-Doctoral Fellowships (BPD1/next-sea/2018, NORTE-01-0145-FEDER-000032). Filipe Costa and the University of Minho's contribution was supported by the strategic program UID/BIA/04050/2013 POCI-010145-FEDER-007569. Victor C Seixas was supported by a Post-Doctoral Fellowship sponsored by CAPES-PNPD (88882.316714/2019-01). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript

How oogenesis analysis combined with dna barcode can help to elucidate taxonomic ambiguities: A polychaete study-based approach

Polychaetes are common in coastal and estuarine environments worldwide and constitute one of the most complex groups of marine invertebrates. The morpho-physiology of the female reproductive system (FRS) can be understood by using histological tools to describe reproductive cycle and gametogenesis paths and, among other purposes, aiming to identify and differentiate polychaete species. However, this histology-based approach is rarely combined with molecular tools, which is known to accurately delimitate species. In the same way, the description and understanding of oogenesis and vitellogenesis paths within polychaetes are lacking for most families, narrowing the range of its utility. Therefore, the present study aims to describe the oogenesis in three polychaete species common and abundant on the South American Atlantic coast (Laeonereis culveri, Scolelepis goodbodyi and Capitella biota) and investigate the utility of reproductive features and gametogenesis as a relevant associate knowledge to discriminate species, particularly useful for putative cryptic species, integrated with morphological and molecular data. In a first attempt, the results obtained herein allow the authors to describe two new subtypes of oogenesis, dividing it in extraovarian oogenesis type I and II and intraovarian type I and II. The results also demonstrate that the following histological characters of the FRS can be relevant for the separation of related species: a) oogenesis type, b) occurrence or absence of a true ovary, c) ovary tissue organization, d) type of accessory cells present, and e) oocyte morphology. Additionally, these histological features of FRS, when compared with correlated species studied under this scope, converge with the genetic data. The analysis of cytochrome oxidase I (COI) barcode sequences differentiates between North and South American Atlantic populations of L. culveri (16.78% genetic distance), while in S. goodbodyi and C. biota it discriminates them from their congeneric species. These results highlight theOs poliquetas são comuns em ambientes costeiros e estuarinos em todo o mundo e constituem um dos grupos mais complexos de invertebrados marinhos. A morfo-fisiologia do sistema reprodutor feminino (FRS) pode ser compreendida por meio de ferramentas histológicas para identificar e diferenciar estes anelídeos. No entanto, essa abordagem histológica raramente é combinada com ferramentas moleculares, amplamente conhecidas por delimitar espécies congenéricas ou crípticas com maior precisão. Do mesmo modo, a descrição e o entendimento da oogênese e vitelogênese dentre os poliquetas, para a maioria das famílias, é ainda limitado. Portanto, o presente estudo tem como objetivo descrever a oogênese em três espécies de poliquetas comuns e abundantes na costa sul-americana (Laeonereis culveri, Scolelepis goodbodyi e Capitella biota) e investigar a utilidade das características reprodutivas e da gametogênese como um conhecimento associado relevante para discriminar espécies, particularmente útil para espécies crípticas putativas, integradas a dados morfológicos e moleculares. Os resultados aqui obtidos permitiram descrever dois novos subtipos de oogênese, dividindo-a em oogênese extra-ovariana dos tipos I e II e intra-ovariana dos tipos I e II. Os resultados também demonstram que os seguintes caracteres histológicos do FRS podem ser relevantes para a separação de espécies relacionadas: a) tipo de oogênese, b) presença ou ausência de um ovário verdadeiro, c) organização tissular ovariana, d) tipo de células acessórias presentes e, e) morfologia do ovócito. Além disso, essas características histológicas do FRS, quando comparadas às espécies correlatas estudadas sob esse escopo, convergem com os dados genéticos separando espécies putativas e congenéricas. As análises com DNA barcode demonstraram que em L. culveri é possível diferenciar as populações atlânticas Norte e Sul-americanas (16,78% de distância genética), enquanto para S. goodbodyi e C. biota fica evidente sua distinção com espécies congenéricas. Esses resultados destacam a importância da abordagem com múltiplas ferramentas e mostram que tanto a histologia quanto a histo-fisiologia do FRS e o DNA barcode podem ser usados para identificar e discriminar espécies crípticas e potencialmente crípticas, o que geralmente não é possível quando se utilizam apenas caracteres morfológicos. Além disso, esses caracteres também podem ser úteis na diferenciação de espécies relacionadas e / ou populações geograficamente distintas desses poliquetas.The authors would like to thank IB/UNICAMP, IO/USP and CEBIMar/USP for providing logistic support. In addition, the authors would like to thank the CBMA and the IB-S for the technical support. This work was supported by the FAPESP (Grants no 2011/50317-5, 2015/25623-6, 2017/06167-5) and CNPq through a productivity grant to A.C.Z.A (306534/2015-0). M.A.L.T was supported by a PhD fellowship (SFRH/BD/131527/2017) from FCT. P.E.V. was supported by a Post-Doctoral Fellowships (BPD1/next-sea/2018, NORTE-01-0145-FEDER-000032). F.O.C. and the University of Minho contribution was supported by the strategic programme UID/BIA/04050/2013 POCI-01-0145-FEDER-007569

Revealing the diversity of the green Eulalia (Annelida, Phyllodocidae) species complex along the European coast, with description of three new species

The green phyllodocids Eulalia clavigera and E. viridis are a known European pseudo-cryptic complex, but questions about its distribution and evidence of additional lineages in previous studies call for an investigation of the real diversity within the complex. We analyze DNA sequences (mtCOI-5P, ITS, and 28S rRNA) of different populations of E. clavigera from intertidal and subtidal marine waters along the North East Atlantic, Mediterranean Sea, the Azores and Webbnesia (Madeira, Savage islands and Canaries), and populations of E. viridis from the Scandinavia. This provided compelling evidence for the existence of six additional divergent evolutionary lineages, three of the most abundant being described here as new species: Eulalia feliciae sp. nov., intertidal and unique to the Western Mediterranean, Eulalia madeirensis sp. nov., subtidal and unique to the Madeira Island (Portugal), and Eulalia xanthomucosa sp. nov., mostly subtidal and occurring in the British Isles and southern France. Complementary morphometric analyses showed that E. feliciae sp. nov. and E. madeirensis sp. nov. formed two independent morphometric clusters, while E. xanthomucosa sp. nov. often overlapped with E. clavigera sensu stricto (s. s.), although being unique in showing a yellow coloration and parapodial cirri on median segments larger in relation to its body size. Recent biotechnological findings based on “E. clavigera” specimens highlight the importance of formally describing cryptic complexes, since each lineage chemistry might be unique and may have a range of distinct effects and applications.This study was supported by the project ATLANTIDA–Platform for the monitoring of the North Atlantic Ocean and tools for the sustainable exploitation of the marine resources, with the reference NORTE-01–0145-FEDER-000040, co-financed by the European Regional Development Fund (ERDF), through Programa Operacional Regional do Norte (NORTE 2020). Thanks are due, for the financial support to CESAM (UIDB/50017/2020 + UIDP/50017/2020), to Portuguese Foundation for Science and Technology and Ministry of Education and Science (FCT/MEC) through national funds, and the co-funding by the FEDER, within the PT2020 Partnership Agreement and Compete 2020. Marcos A. L. Teixeira was supported by a PhD grant from FCT co-financed by ESF (SFRH/BD/131527/2017) and from the DNAqua-Net STSM grant “Rich and hidden biodiversity not yet barcoded in the Canary archipelago (Spain) as an opportunity to enrich the DNA barcode reference library for European polychaetes,” under the EU Cost action CA15219–Developing new genetic tools for bio-assessment of aquatic ecosystems in Europe. Pedro E. Vieira was supported by national funds through the Portuguese Foundation for Science and Technology (FCT, I.P.) in the scope of the project (early detection and monitoring of non-indigenous species in coastal ecosystems based on high-throughput sequencing tools, PTDC/BIA-BMA/29754/2017). Ascensão Ravara was supported by national funds, through FCT, I.P., in the scope of the framework contract foreseen in the numbers 4, 5, and 6 of the article 23, of the Decree-Law 57/2016, of August 29, changed by Law 57/2017, of July 19. Arne Nygren was supported by the Norwegian Taxonomy Initiative [https://www.biodiversity.no/Pages/135523] (Cryptic polychaete species in Norwegian waters, knr 49–13, pnr 70184228), the Swedish Taxonomy Initiative [https://www.artdatabanken.se/en/the-swedish-taxonomy-initiative/] (Polychaete species complexes in Swedish waters, dnr 140/07 1.4 and 166/08 1.4), and Kungliga Fysiografiska sällskapet Nilsson-Ehle donationerna [https://www.fysiografen.se/sv/]

Molecular and morphometric analyses identify new lineages within a large Eumida (Annelida) species complex

We report on two new lineages of the Eumida sanguinea complex from Great Britain and describe one of them as a new species using a multilocus approach, including the mitochondrial DNA COI-5P and the nuclear markers ITS (ITS1, 5.8S rRNA and ITS2) and 28S rRNA. The molecular analysis placed Eumida mackiei sp. nov. in a monophyletic clade with 19.1% (COI), 10.1% (ITS) and 1.7% (28S) mean distance to its nearest neighbour. Molecular diagnoses were also applied to nine lineages within the E. sanguinea complex. This was complemented with morphometric data employing multivariate statistical analysis and the incorporation of statistical dissimilarities against three other described species from the complex. Eumida mackiei sp. nov. can be distinguished from E. notata and E. maia by the larger distance between the eyes and differences in morphometric proportions mainly in the dorsal and ventral cirri as well as in the prostomial appendages. E. sanguinea sensu stricto failed to produce a cluster of its own in the morphometric analysis, probably due to juvenile bias. Integrative taxonomy provided strong evidence to formally describe a new cryptic species that can now be used in biomonitoring or other relevant ecological research.UA - Universidade de Aveiro(70184228)

The curious and intricate case of the European Hediste diversicolor (Annelida, Nereididae) species complex, with description of two new species

Past molecular studies using mtDNA sequences and alloenzymes signalled the existence of at least two cryptic species within the Hediste diversicolor morphotype, in European coasts. However, to this day, no new species descriptions have been made. In this study, we identified five completely sorted lineages using a multi-locus approach, including the mitochondrial DNA cytochrome oxidase I gene (COI-5P) and the nuclear markers ITS2 rRNA and 28S rRNA. The molecular data were complemented with morphometric measurements examined through multivariate statistical analysis and the incorporation of statistical dissimilarities. Apart from the Baltic Sea, where three of the lineages occur in sympatry, Hediste diversicolor comprises four deeply divergent allopatric lineages in the rest of Europe. They group populations from the NE Atlantic and part of the Western Mediterranean Sea; from the Tyrrhenian Sea; from the Adriatic and Ionian Sea; and, lastly, from the Caspian, Black and the northern Aegean Seas. The lineage from the Ionian Sea revealed low genetic distances compared with the one from the Adriatic Sea and lacked enough specimens for the morphometric analysis, preventing further conclusions about its independent status. Three independent morphometric clusters were identified mainly based on worm size, the distance between the anterior and posterior eyes, parapodia proportions and the length of several prostomial appendages. Two sympatric lineages present in the Baltic Sea, showed evidence of possible hybridization and lacked significant PCA morphometric variation between them. The two remaining lineages were formally described as new species, namely Hediste pontii sp. nov. (Adriatic Sea) and Hediste astae sp. nov. (northern Aegean, Caspian and Black Seas). These new species can now be formally recognized and used in biomonitoring or other relevant ecological studies. Finally, a neotype is defined for H. diversicolor, whose usage is restricted to the NE Atlantic lineage.UA - Universidade de Aveiro(70184228

Reappraisal of the hyperdiverse Platynereis dumerilii (Annelida: Nereididae) species complex in the Northern Atlantic, with the description of two new species

Morphologically similar species are often overlooked but molecular techniques have been effective in signalling potential hidden diversity, boosting the documentation of unique evolutionary lineages and ecological diversity. Platynereis dumerilii and Platynereis massiliensis are part of a recognised species complex, where differences in the reproductive biology have mainly been highlighted to date. Analyses of DNA sequence data (COI, 16S rDNA and D2 region of the 28S rDNA) of populations of the apparent morphotype of P. dumerilii obtained from a broader sampling area along European marine waters, including the Azores and Webbnesia islands (Madeira and Canaries), provided compelling evidence for the existence of at least 10 divergent evolutionary lineages. Complementing the genetic data, morphological observations of the better represented lineages revealed two major groups with distinctive paragnath patterns. Two new Platynereis species were erected: P. nunezi sp. nov., widespread in the Azores and Webbnesia islands, and P. jourdei sp. nov., restricted to the western Mediterranean. The new combination P. agilis is also proposed for Nereis agilis, previously unaccepted for one of the lineages present both in the Northeast Atlantic and western Mediterranean. Platynereis dumerilii is redescribed based on topotypic material. However, uncertainty in the identity of P. massiliensis due to the original brief description and the absence of type and topotypic material prevents the unequivocal assignment to the lineage assumed in this and previous studies. The remaining five lineages are represented by only a few small specimens with morphological features poorly preserved and were therefore not described in this study.This study was supported by the project Next generation monitoring of coastal ecosystems in a scenario of global change (NextSea, NORTE-01-0145-FEDER-000032), under the PORTUGAL 2020 Partnership Agreement, through the European Regional Development Fund (ERDF). Thanks are due, for the financial support of CESAM – Centro de Estudos do Ambiente e do Mar (UIDB/50017/2020+UIDP/50017/2020), to Portuguese Foundation for Science and Technology and Ministry of Education and Science (FCT/MEC) through national funds, and the co-funding by the FEDER, within the PT2020 Partnership Agreement and Compete 2020. The research leading to these results also received partial funding, from the European Union’s Horizon 2020 research and innovation programme under grant agreement No 730984, ASSEMBLE Plus project (application n. 8229, 4th CALL, "Crypticism in the marine realm: DNA barcode-based outlook into selected invertebrate taxa of the Eastern Mediterranean"). Marcos AL Teixeira was supported by a PhD grant from FCT co-financed by ESF (SFRH/BD/131527/2017) and from the DNAqua-Net STSM grant "Rich and hidden biodiversity not yet barcoded in the Canary archipelago (Spain) as an opportunity to enrich the DNA barcode reference library for European polychaetes", under the EU Cost action CA15219 - Developing new genetic tools for bioassessment of aquatic ecosystems in Europe. Pedro Vieira work was supported by national funds through the Portuguese Foundation for Science and Technology (FCT, I.P.) in the scope of the project (Early detection and monitoring of non-indigenous species in coastal ecosystems based on high-throughput sequencing tools, PTDC/BIA-BMA/29754/2017). Ascensão Ravara was funded by national funds, through FCT, I.P., in the scope of the framework contract foreseen in the numbers 4, 5 and 6 of the article 23, of the Decree-Law 57/2016, of August 29, changed by Law 57/2017, of July 19. Financial support to Arne Nygren from the Norwegian Taxonomy Initiative [http://www.biodiversity.no/Pages/135523] (Cryptic polychaete species in Norwegian waters, knr 49-13, pnr 70184228), the Swedish Taxonomy Initiative [https://www.artdatabanken.se/en/the-swedish-taxonomy-initiative/] (Polychaete species complexes in Swedish waters, dnr 140/07 1.4 and 166/08 1.4), and Kungliga Fysiografiska sällskapet Nilsson-Ehle donationerna [https://www.fysiografen.se/sv/]. Financial support to Torkild Bakken from the Norwegian Taxonomy Initiative project Polychaetes in Norwegian Ports project no. 70184238

Relevância dos conhecimentos, habilidades e métodos instrucionais na perspectiva de estudantes e profissionais da área contábil: estudo comparativo internacional

Com o incentivo dos estudos de Albrecht e Sack (2000), Francisco e Kelly (2002) e com base na Taxonomia de Bloom (1956), o objetivo deste trabalho consiste em comparar a percepção de estudantes de cursos de Ciências Contábeis em Instituições de Ensino Superior (IES) brasileiras e profissionais da Contabilidade no Brasil quanto aos conhecimentos, habilidades e métodos de ensino-aprendizagem considerados como mais importantes para a atuação do contador no mercado de trabalho. O estudo contou com uma amostra de 1.710 sujeitos: 769 estudantes matriculados em cursos de graduação em Ciências Contábeis em IES brasileiras e 941 contadores registrados nos Conselhos Regionais de Contabilidade de várias regiões brasileiras, que responderam ao questionário desenvolvido por Lin, Xiong e Liu (2005) sobre a importância atribuída pelos respondentes aos quesitos conhecimento, habilidades e métodos de ensino-aprendizagem. Os resultados encontrados evidenciaram maiores níveis de importância percebida pelos profissionais (quando comparados aos estudantes) nos quesitos investigados. Comparativamente com a China e com os EUA, os escores dos profissionais brasileiros são sempre maiores para as três dimensões (conhecimentos, habilidades e métodos). Os estudantes brasileiros em comparação com os chineses, de forma geral, também atribuem maior importância para as três dimensões analisadas. Os elevados escores apontados pelos respondentes brasileiros merecem atenção por parte dos gestores das IES, levando em conta a intensidade das mudanças de perfil dos jovens estudantes e a velocidade das alterações do mercado profissional. Destaque-se que esses resultados devem ser mais explorados, analisando-se as suas razões, inferindo-se que o contexto atual da Contabilidade no Brasil, decorrente do processo global de harmonização das normas internacionais, é cenário propício para explicações a esse respeito