Identification of the taxonomic status of Scinax nebulosus and Scinax constrictus (Scinaxinae, Anura) based on molecular markers / Identificação do status taxonômico de Scinax nebulosus e Scinax constrictus (Scinaxinae, Anura), baseado em marcadores moleculares

Abstract The validation of many anuran species is based on a strictly descriptive, morphological analysis of a small number of specimens with a limited geographic distribution. The Scinax Wagler, 1830 genus is a controversial group with many doubtful taxa and taxonomic uncertainties, due a high number of cryptic species. One example is the pair of species Scinax constrictus and Scinax nebulosus, which share a similar morphology. Scinax constrictus is restricted to the Brazilian Cerrado savanna, while S. nebulosus is widely distributed throughout northern South America. Despite the validation of many anuran species, discriminations based only on morphological traits is quite difficult due to the high conservative morphology of some groups. In this context, the present study uses mitochondrial and nuclear genes to provide a more consistent diagnosis and test the validity of S. constrictus as a distinct species from S. nebulosus, as well as evaluate the position of these taxa within the Scinax genus. The topologies obtained herein uphold the monophyletic status of Scinax based on all molecular markers assessed in this study, in all analytical approaches, with high levels of statistical support.

Resumo A validação de muitas espécies de anuros é baseada em uma análise morfológica e descritiva de um pequeno número de espécimes com uma distribuição geográfica limitada. O gênero Scinax Wagler, 1830 é um grupo controverso com muitos táxons duvidosos e incertezas taxonômicas devido ao grande número de espécies crípticas. Um exemplo são as espécies, Scinax constrictus e Scinax nebulosus, que compartilham uma morfologia similar. Scinax constrictus é restrito à savana do Cerrado brasileiro, enquanto S. nebulosus é amplamente distribuído pelo norte da América do Sul. Apesar da validação de muitas espécies de anuros, a discriminação baseada apenas em características morfológicas é bastante difícil, devido à alta morfologia conservadora de alguns grupos. Neste contexto, o presente estudo utiliza genes mitocondriais e nucleares para fornecer um diagnóstico mais consistente e para testar a validade de S. constrictus como uma espécie distinta de S. nebulosus, bem como avaliar a posição destes táxons dentro do gênero Scinax. As topologias obtidas confirmaram o status monofilético de Scinax com base em todos os marcadores moleculares, em todas as abordagens analíticas, com altos níveis de suporte estatístico.

A new Hyphessobrycon (Characiformes: Characidae) of the Hyphessobrycon heterorhabdus species-group from the lower Amazon basin, Brazil

A new species of Hyphessobrycon belonging to the Hyphessobrycon heterorhabdus species-group from the lower rio Tapajós, state of Pará, Brazil, is described. The new species is allocated into the Hyphessobrycon heterorhabdus species-group due to its color pattern, composed by an anteriorly well-defined, horizontally elongated humeral blotch that becomes diffuse and blurred posteriorly, where it overlaps with a conspicuous midlateral dark stripe that becomes blurred towards the caudal peduncle and the presence, in living specimens, of a tricolored longitudinal pattern composed by a dorsal red or reddish longitudinal stripe, a middle iridescent, golden or silvery longitudinal stripe, and a more ventrally-lying longitudinal dark pattern composed by the humeral blotch and dark midlateral stripe. It can be distinguished from all other species of the group by possessing humeral blotch with a straight or slightly rounded ventral profile, lacking a ventral expansion present in all other species of the group. The new species is also distinguished from Hyphessobrycon heterorhabdus by a 9.6% genetic distance in the cytochrome c oxidase I gene. The little morphological distinction of the new species when compared with its most similar congener, H. heterorhabdus, indicates that the new species is one of the first truly cryptic fish species described from the Amazon basin.(AU)

Uma nova espécie de Hyphessobrycon pertencente ao grupo Hyphessobrycon heterorhabdus é descrita da região do baixo rio Tapajós, estado do Pará, Brasil. A nova espécie é incluída no grupo Hyphessobrycon heterorhabdus devido ao seu padrão de coloração, composto por uma mancha umeral alongada, anteriormente bem definida, que se torna difusa e borrada posteriormente, onde se sobrepõe a uma conspícua faixa escura médio-lateral que se torna borrada próxima ao pedúnculo caudal, e pela presença, em exemplares vivos, de um padrão longitudinal tricolor, composto por uma faixa longitudinal vermelha ou avermelhada dorsal, uma faixa média iridescente dourada ou prateada e, mais ventralmente, o padrão longitudinal escuro composto pela faixa escura médio-lateral e mancha umeral. A espécie pode ser distinguida das outras espécies pertencentes ao grupo por possuir uma mancha umeral com região ventral retilínea ou levemente arredondada, sem uma expansão ventral presente nas demais espécies do grupo. A espécie também se diferencia de Hyphessobrycon heterorhabdus por uma distância genética de 9,6% no gene citocromo c oxidase I. A sutil diferença morfológica da nova espécie quando comparada ao seu congênere mais similar, H. heterorhabdus, indica que a nova espécie é uma das primeiras espécies de peixes verdadeiramente crípticas descritas da Bacia Amazônica.(AU)

Molecular species delimitation of the genera Anodus, Argonectes, Bivibranchia and Micromischodus (Ostariophysi: Characiformes)

ABSTRACT A good taxonomic assessment of specimens is an essential task to many biological studies and DNA data have provided additional sources of information to assist in the disentanglement of taxonomic problems among living organisms, as has been the case of some taxa of the megadiverse Neotropical ichthyofauna. Here we assessed all valid species in the Neotropical freshwater fish genera Anodus, Argonectes, Bivibranchia and Micromischodus of the family Hemiodontidae to establish molecular species boundaries among them. All species delimitation methods defined exactly only one MOTU for Anodus elongatus, Argonectes longiceps, A. robertsi, Bivibranchia bimaculata, B. notata, B. velox, and Micromischodus sugillatus, resulting in total congruence between nominal species and MOTUs for these seven taxa. The three species having discordant results across analyses: Anodus orinocensis, Bivibranchia fowleri, and Bivibranchia simulata, matched more than one MOTU per species in some methods, meaning that cryptic diversity may exist within these taxa. Overall, this great correspondence among morphological and molecular boundaries for thae species analysed seem to be indicative of a reasonably stable taxonomy within these Hemiodontidae genera.

RESUMO Uma avaliação taxonômica adequada dos espécimes é uma tarefa essencial para muitos estudos biológicos, e os dados de DNA têm fornecido fontes adicionais de informações para auxiliar no desemaranhamento de muitos grupos taxonômicos, como tem sido o caso de alguns táxons da megadiversa ictiofauna Neotropical. Aqui examinamos todas as espécies válidas dos gêneros de peixes Neotropicais de água doce Anodus, Argonectes, Bivibranchia e Micromischodus (família Hemiodontidae) para estabelecer limites moleculares entre elas. Todos os métodos de delimitação definiram exatamente apenas uma MOTU para Anodus elongatus, Argonectes longiceps, A. robertsi, Bivibranchia bimaculata, B. notata, B. velox e Micromischodus sugillatus, resultando em congruência total entre espécies nominais e MOTUs para estes sete táxons. As três espécies com resultados divergentes entre as análises, a saber, Anodus orinocensis, Bivibranchia fowleri e Bivibranchia simulata, corresponderam a mais de um MOTU por espécie em alguns métodos, mostrando que pode existir uma diversidade críptica dentro desses taxa. No geral, esta grande correspondência entre os limites morfológicos e moleculares para as espécies analisadas parece indicar uma taxonomia relativamente estável para esses gêneros de Hemiodontidae.

Contributions to the taxonomy of Trachelyopterus (Siluriformes): comparative cytogenetic analysis in three species of Auchenipteridae

Auchenipteridae is divided into subfamilies Centromochlinae and Auchenipterinae. Parauchenipterus is included in the latter and is subject of taxonomic discussions concerning its validation or synonymization with Trachelyopterus. Herein, three species from two hydrographic basins were cytogenetically analyzed: Parauchenipterus striatulus from Doce River and two sympatric species, P. galeatus and Trachelyopterus coriaceus, from the Araguaia River. Diploid number of 58 chromosomes was verified for all species, but P. striatulus has different karyotype formula from the others. The three species have heterochromatin located in terminal regions of almost all chromosomes and in pericentromeric region on acrocentric chromosomes. Simple NORs was verified on a subtelocentric chromosome for all species. 5S rDNA sites were detected in three submetacentric chromosome pairs in P. striatulus; in a metacentric chromosome pair and submetacentric pair in T. coriaceus; and in one metacentric chromosome pair in P. galeatus. The similarities found in the karyotypes of the three species suggest the existence of only one genus, Trachelyopterus; therefore, our data refutes the validation of Parauchenipterus. Moreover, the differences in 5S rDNA distribution in P. galeatus in comparison with other populations already studied, indicate the existence of a new taxonomic unit, which suggests a species complex in P. galeatus.(AU)

Auchenipteridae é dividida nas subfamílias Centromochlinae e Auchenipterinae. Parauchenipterus encontra-se incluído na última e tem sido alvo de discussões relacionadas com a problemática taxonômica de validação ou sinonimização com Trachelyopterus. Foram analisadas citogeneticamente três espécies de duas bacias hidrográficas: Parauchenipterus striatulus do rio Doce, P. galeatus e Trachelyopterus coriaceus, simpátricas do rio Araguaia. Todas as espécies analisadas apresentaram número diploide de 58 cromossomos, com diferença na fórmula cariotípica de P. striatulus. A heterocromatina foi localizada nas regiões terminais de quase todos os cromossomos e na região pericentromérica nos cromossomos acrocêntricos das três espécies. AgNORs e DNAr 18S detectaram RONs simples em um par de cromossomos subtelocêntricos nas três espécies. DNAr 5S foi detectado em três pares de cromossomos submetacêntricos em P. striatulus; em um par de cromossomos metacêntricos e um par submetacêntrico em T. coriaceus; e em apenas um par de cromossomos metacêntricos em P. galeatus. As semelhanças encontradas nos cariótipos das três espécies analisadas indicam a existência de somente Trachelyopterus (não validação de Parauchenipterus) e a diferença encontrada na distribuição de DNAr 5S de P. galeatus em relação às outras populações já estudadas sugere a existência de uma nova unidade taxonômica, portanto P. galeatus compreende um complexo de espécies.(AU)

Integrative taxonomy reveals a new species of pacu (Characiformes: Serrasalmidae: Myloplus) from the Brazilian Amazon

Pacus of the genus Myloplus represent a formidable taxonomic challenge, and particularly so for the case of M. asterias and M. rubripinnis, two widespread and common species that harbor considerable morphological diversity. Here we apply DNA barcoding and multiple species discovery methods to find candidate species in this complex group. We report on one well-supported lineage that is also morphologically and ecologically distinct. This lineage represents a new species that can be distinguished from congeners by the presence of dark chromatophores on lateral-line scales, which gives the appearance of a black lateral line. It can be further diagnosed by having 25-29 branched dorsal-fin rays (vs. 18-24), 89-114 perforated scales from the supracleithrum to the end of hypural plate (vs. 56-89), and 98-120 total lateral line scales (vs. 59-97). The new species is widely distributed in the Amazon basin, but seems to have a preference for black- and clearwater habitats. This ecological preference and black lateral line color pattern bears a striking similarity to the recently described silver dollar Metynnis melanogrammus.(AU)

Pacus do gênero Myloplus representam um desafio taxonômico formidável, e particularmente o caso de M. asterias e M. rubripinnis, duas espécies amplamente distribuídas e comuns que abrigam uma considerável diversidade morfológica. Aplicamos aqui a tecnologia do DNA barcoding e múltiplos métodos de descoberta de espécies para encontrar possíveis espécies novas nesse grupo complexo. Registramos uma linhagem bem suportada que também é distinta morfológica e ecologicamente. Essa linhagem representa uma nova espécie que pode ser distinguida das demais congêneres por apresentar cromatóforos escuros nas escamas da linha lateral que conferem uma aparência de linha lateral preta. Ela pode ser adicionalmente diagnosticada por ter 25-29 raios ramificados na nadadeira dorsal (vs. 18-24), 89-114 escamas perfuradas do supracleitro até o final da placa hipural (vs. 56-89) e 98-120 escamas totais na linha lateral (vs. 59-97). A nova espécie é amplamente distribuída na bacia Amazônica, mas aparentemente possui preferência por habitats de água preta e clara. A preferência ecológica e o padrão de colorido escuro da linha lateral consistem em semelhanças impressionantes com o silver dólar recém descrito Metynnis melanogrammus.(AU)

How oogenesis analysis combined with DNA barcode can help to elucidate taxonomic ambiguities: a polychaete study-based approach / Como análises de oogênese combinadas com DNA barcode podem elucidar ambiguidades taxonômicas: uma abordagem baseada em estudos com poliquetas

Abstract: 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 the importance of multi-tool approach and shows that both FRS histology and histo-physiology, and DNA barcoding can be used to identify and discriminate cryptic species, which is usually not possible when using morphological characters. Besides, these characters may also be useful in differentiating related species, and/or geographically distinct populations among polychaetes.

Resumo: Os 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.

Morphological and molecular differences between the Amphiamerican populations of Antillesoma (Sipuncula: Antillesomatidae), with the description of a new species

Introduction: The sipunculans are a group of marine invertebrates that have been little studied in the tropical eastern Pacific (TEP). Antillesoma antillarum is a species belonging to the monospecific family Antillesomatidae, considered widely distributed in tropical and subtropical localities across the globe. Objective: The main objective of this work was to examine the morphological and molecular differences between specimens from both coasts of tropical America to clarify the taxonomy of this species. Methods: We examined the morphology with material from the Mexican Caribbean and southern Mexican Pacific. To perform molecular analyses, two sequences of the COI molecular marker were obtained from specimens collected in Panteón Beach, Oaxaca, southern Mexican Pacific, and compared with four sequences identified as A. antillarum in GenBank, all of them from different localities. A phylogenetic reconstruction was performed with the maximum likelihood method and genetic distances were calculated with the Kimura 2P model and compared to reference values. Results: The phylogenetic analysis revealed three different lineages of Antillesoma that are well supported by bootstrap values: Antillesoma antillarum sensu stricto from the Caribbean Sea and Florida; a sister group to the one represented by our samples from the Mexican Pacific; and a third group from Thailand. Conclusion: Based on morphological traits and molecular data, Antillesoma mexicanum sp. nov. is described from the Mexican Pacific, differing from A. antillarum in the trunk papillae, color patterns and, additionally, the specimens from the Caribbean attain significantly bigger trunk sizes than the ones Pacific.

Introducción: Los sipúnculos son un grupo de gusanos marinos sin segmentación poco estudiados en el Pacífico oriental tropical (POT). Antillesoma antillarum es una especie perteneciente a la familia monoespecífica Antillesomatidae la cual se consideraba que se distribuía ampliamente en distintas localidades tropicales y subtropicales del mundo. Objetivo: El objetivo principal del trabajo fue examinar las diferencias morfológicas y moleculares entre ejemplares de ambas costas de América. Métodos: Para el análisis morfológico se revisó material del Caribe mexicano y del Pacífico sur de México. Para los análisis moleculares se obtuvieron secuencias del marcador mitocondrial COI de ejemplares de A. mexicanum sp. nov. de la playa de Panteón en Puerto Ángel, Oaxaca del Pacífico sur de México; también se incluyeron cuatro secuencias de GenBank de A. antillarum de diferentes localidades para la comparación filogenética con el método de Máxima Verosimilitud. Se calcularon las distancias genéticas con el modelo Kimura 2P y fueron comparadas con valores de referencia. Resultados: El análisis filogenético evidenció tres linajes diferentes: Antillesoma antillarum sensu stricto del Mar Caribe y Florida, el grupo hermano representado por nuestra recolecta en el Pacífico mexicano y un tercer grupo de Tailandia. Conclusión: Basados en datos morfológicos y moleculares, Antillesoma mexicanum sp. nov. fue descrita para el Pacífico mexicano, que difiere de A. antillarum en las papilas del tronco, el patrón de coloración y, adicionalmente, los ejemplares del Caribe fueron significativamente más grandes que los del Pacífico mexicano.

Comparative cytogenetics of Astyanax (Teleostei: Characidae) from the upper Paraguay basin

Astyanax is one of the most abundant and diverse taxa of fishes in the Neotropical region. In order to increase the amount of cytogenetic information for Astyanax as well as to exhibit data to subsidize future taxonomic studies, this work analyzed three species of Astyanax: two species are cryptic, and are here reported to live in syntopy (A. abramis and A. lacustris); the first karyotype description for A. pirapuan is also presented. Cytogenetic analyzes reveal a diploid number of 2n=50 chromosomes for three species, yet with differences in their karyotype morphology. The physical mapping of 18S rDNA showed up to thirteen sites in A. pirapuan and two in A. abramis and A. lacustris. The physical mapping of 5S rDNA has proven to be an effective marker for the characterization of species of Astyanax studied in this work.(AU)

Astyanax é um dos táxons mais representados e diversos na região Neotropical. Com o intuito de aumentar as informações citogenéticas para Astyanax e apresentar dados que possam subsidiar estudos taxonômicos futuros, este trabalho traz uma análise citogenética de três espécies de Astyanax: duas espécies consideradas crípticas, aqui reportadas em sintopia (A. abramis e A. lacustris) e a primeira descrição cariotípica de A. pirapuan. As análises citogenéticas revelaram 2n=50 cromossomos para as três espécies, com diferença na morfologia cariotípica de cada uma. Foram observados apenas dois sítios de rDNA 18S em A. abramis e A. lacustris e até 13 para A. pirapuan. O mapeamento físico do rDNA 5S se mostrou como um marcador efetivo para a caracterização das espécies de Astyanax abordadas neste estudo.(AU)

Phylogenetic analysis reveals cryptic species diversity within minute intestinal fluke, Stellantchasmus falcatus Onji and Nishio, 1916 (Trematoda, Heterophyidae)

Objective To examine the taxonomic boundaries in the Stellantchasmus populations in different hosts and reveal the cryptic speciation of the Stellantchasmus in Thailand based on both morphological and molecular approaches. Methods Stellantchasmus falcatus (S. falcatus) s.l. was collected from different hosts throughout Thailand. The worms were examined and measured. The nuclear ITS2 gene and the mitochondrial COI gene were used to investigate the S. falcatus s.l. from Dermogenus pusillus and Liza subviridis. Results Stellantchasmus was one of food-borne trematode that widely distribute throughout Thailand. The comparison of S. falcatus s.l. from Dermogenus pusillus and Liza subviridis indicated a genetic divergence of nuDNA with 3.6% and mtDNA with 19.3%, respectively. Morphological characteristics were fairly different in the ratio of body size, length of prepharynx, and ratio of organ size. Conclusions Our results provide initial evidence that S. falcatus s.l. from different hosts tend to be a different species based on both molecular and morphological characters. Cryptic species complexes are generally found among parasites that tend to have large populations and/or rapid evolution. The degree of genetic diversity existing would suggest the practice of targeted regimes to design and minimize these lections of anthelmintic resistance in the future.

Phylogenetic analysis reveals cryptic species diversity within minute intestinal fluke, Stellantchasmus falcatus Onji and Nishio, 1916 (Trematoda, Heterophyidae)

OBJECTIVE@#To examine the taxonomic boundaries in the Stellantchasmus populations in different hosts and reveal the cryptic speciation of the Stellantchasmus in Thailand based on both morphological and molecular approaches.@*METHODS@#Stellantchasmus falcatus (S. falcatus) s.l. was collected from different hosts throughout Thailand. The worms were examined and measured. The nuclear ITS2 gene and the mitochondrial COI gene were used to investigate the S. falcatus s.l. from Dermogenus pusillus and Liza subviridis.@*RESULTS@#Stellantchasmus was one of food-borne trematode that widely distribute throughout Thailand. The comparison of S. falcatus s.l. from Dermogenus pusillus and Liza subviridis indicated a genetic divergence of nuDNA with 3.6% and mtDNA with 19.3%, respectively. Morphological characteristics were fairly different in the ratio of body size, length of prepharynx, and ratio of organ size.@*CONCLUSIONS@#Our results provide initial evidence that S. falcatus s.l. from different hosts tend to be a different species based on both molecular and morphological characters. Cryptic species complexes are generally found among parasites that tend to have large populations and/or rapid evolution. The degree of genetic diversity existing would suggest the practice of targeted regimes to design and minimize these lections of anthelmintic resistance in the future.

A new species of Astyanax (Ostariophysi: Characidae) from the headwaters of the arheic Río Sucuma, Catamarca, Northwestern Argentina

A new species of Astyanax is described from the arheic system of Río Sucuma, in Catamarca, Northwestern Argentina. The new species is distinguished from its congeners by a combination of characters including the presence of a broad vertical humeral spot, absence of maxillary teeth, and absence of circuli in posterior field of scales. Furthermore, this species is distinguished by the orbital diameter, head length, branched anal-fin rays, perforated lateral-line scales, transverse scales, dentary teeth with abrupt decrease in size. This species is, to date, the single known fish endemic of Río Sucuma basin.(AU)

Una nueva especie de Astyanax, se describe del Sistema arreico del Río Sucuma, en Catamarca, Noroeste de Argentina. Esta nueva especie se distingue de sus congéneres por una combinación de caracteres que incluyen la presencia de una gran mancha humeral, verticalmente elongada, ausencia de diente en el maxilar, ausencia de circuli en el campo posterior de las escamas, además se distingue por un ojo relativamente pequeño, longitud de la cabeza, radios ramificados en la aleta anal; escamas perforadas en la línea lateral; escamas transversas; dentario con dientes que decrecen abruptamente. Esta especie es, a la fecha, el único pez endémico para la cuenca de Río Sucuma.(AU)

Identification of the taxonomic status of Scinax nebulosus and Scinax constrictus (Scinaxinae, Anura) based on molecular markers

Abstract The validation of many anuran species is based on a strictly descriptive, morphological analysis of a small number of specimens with a limited geographic distribution. The Scinax Wagler, 1830 genus is a controversial group with many doubtful taxa and taxonomic uncertainties, due a high number of cryptic species. One example is the pair of species Scinax constrictus and Scinax nebulosus, which share a similar morphology. Scinax constrictus is restricted to the Brazilian Cerrado savanna, while S. nebulosus is widely distributed throughout northern South America. Despite the validation of many anuran species, discriminations based only on morphological traits is quite difficult due to the high conservative morphology of some groups. In this context, the present study uses mitochondrial and nuclear genes to provide a more consistent diagnosis and test the validity of S. constrictus as a distinct species from S. nebulosus, as well as evaluate the position of these taxa within the Scinax genus. The topologies obtained herein uphold the monophyletic status of Scinax based on all molecular markers assessed in this study, in all analytical approaches, with high levels of statistical support.

Resumo A validação de muitas espécies de anuros é baseada em uma análise morfológica e descritiva de um pequeno número de espécimes com uma distribuição geográfica limitada. O gênero Scinax Wagler, 1830 é um grupo controverso com muitos táxons duvidosos e incertezas taxonômicas devido ao grande número de espécies crípticas. Um exemplo são as espécies, Scinax constrictus e Scinax nebulosus, que compartilham uma morfologia similar. Scinax constrictus é restrito à savana do Cerrado brasileiro, enquanto S. nebulosus é amplamente distribuído pelo norte da América do Sul. Apesar da validação de muitas espécies de anuros, a discriminação baseada apenas em características morfológicas é bastante difícil, devido à alta morfologia conservadora de alguns grupos. Neste contexto, o presente estudo utiliza genes mitocondriais e nucleares para fornecer um diagnóstico mais consistente e para testar a validade de S. constrictus como uma espécie distinta de S. nebulosus, bem como avaliar a posição destes táxons dentro do gênero Scinax. As topologias obtidas confirmaram o status monofilético de Scinax com base em todos os marcadores moleculares, em todas as abordagens analíticas, com altos níveis de suporte estatístico.

Can sibling species of the Drosophila willistoni subgroup be recognized through combined microscopy techniques?

ABSTRACT In several arthropod groups, male genitalia is the most important feature for species identification, especially in cryptic species. Cryptic species are very common in the Drosophila genus, and the Neotropical Drosophila willistoni species group is a good example. This group currently includes 24 species divided into three subgroups: alagitans, bocainensis and willistoni. There are six sibling species in the willistoni subgroup – D. willistoni, D. insularis, D. tropicalis, D. equinoxialis, D. pavlovskiana and D. paulistorum, which is a species complex composed of six semispecies – Amazonian, Andean-Brazilian, Centroamerican, Interior, Orinocan and Transitional. The objective of this study was to characterize male genitalia of the willistoni subgroup, including the D. paulistorum species complex, using scanning electron microscopy and light microscopy. We also tried to contribute to the identification of these cryptic species and to add some comments about evolutionary history, based on male genitalia characters. Despite being cryptic species, some differences were found among the siblings, including the Drosophila paulistorum semispecies.

Cryptic species and genetic structure in Didemnum granulatum Tokioka, 1954 (Tunicata: Ascidiacea) from the southern Brazilian coast / Espécies crípticas e estrutura genética em Didemnum granulatum Tokioka 1954 (Tunicata: Ascidiacea) na costa sul do Brasil

Didemnum granulatum is a colonial fouling ascidian that lives in subtidal substrates, worldwide. It exhibits two morphotypes, orange and beige. In this study, we verified if the color morphotypes and/or the spatial distribution of specimens in different islands might be associated to patterns of genetic structure of a single species, or if they represent distinct cryptic species. Specimens were collected in four islands, along the coast of the Santa Catarina state. A segment of 490 bp from the mitochondrial gene cytochrome c oxidase subunit 1 (COI) was amplified from 45 samples. Twenty-one haplotypes were identified. The total haplotype diversity (0.912) and the total nucleotide diversity (0.044) were high. The global Fst of the populations analyzed was 0.97, with most of the variation occurring between orange and beige groups (82.19%). The variation found between populations within groups was 15.37%, and 2.45% within populations. Haplotype networks and the neighbor-joining tree showed clear genetic divergence between individuals of distinct colors, and between the islands. These evidences strongly support the presence of a complex of two cryptic species for D. granulatum occupying the studied area. Both species were also highly genetically structured between islands, suggesting that the conservation process of these populations is complex.

Didemnum granulatum é uma ascídia colonial incrustante de substratos consolidados infralitorâneos, e de ampla distribuição mundial. Variação na pigmentação é comum em invertebrados marinhos, e morfotipos de D. granulatum das cores laranja e bege foram detectados na região. Neste estudo, nós verificamos se os morfotipos de cor e/ou a distribuição espacial dos espécimens nas diferentes ilhas podem estar associados aos padrões de estruturação genética de uma única espécie, ou se eles representam espécies crípticas distintas. Os espécimens foram coletados em quatro ilhas ao longo da costa do estado de Santa Catarina. Um total de 45 amostras tiveram amplificados um segmento de 490pb do gene mitocondrial citocromo oxidase subunidade I (COI). Vinte e um haplótipos foram identificados. A diversidade haplotípica total (0.912) e a diversidade nucleotídica total (0.044) foram altas. O Fst global das populações analisadas foi 0.97, e a maior parte da variação ocorreu entre os grupos laranja e bege (82.19%). A variação encontrada entre as populações dentro dos grupos foi 15.37%, e 2.45% dentro das populações. A rede de haplótipos e a árvore de Neighbor-joining mostraram nítidas divergências genéticas entre os indivíduos de cores distintas, e entre as ilhas. Tais evidências sugerem a presença de um complexo de duas espécies crípticas de D. granulatum na região. Ambas as espécies foram fortemente estruturadas geneticamente entre as ilhas, o que reforça a necessidade de que tais atributos sejam considerados em medidas de conservação e proteção do ambiente marinho, mais especificamente nestas ilhas.

Cryptic species of Paracoccidioides brasiliensis: impact on paracoccidioidomycosis immunodiagnosis

We aimed to evaluate whether the occurrence of cryptic species of Paracoccidioides brasiliensis, S1, PS2, PS3 and Paracoccidioides lutzii, has implications in the immunodiagnosis of paracoccidioidomycosis (PCM). Small quantities of the antigen gp43 were found in culture filtrates of P. lutzii strains and this molecule appeared to be more variable within P. lutzii because the synonymous-nonsynonymous mutation rate was lower, indicating an evolutionary process different from that of the remaining genotypes. The production of gp43 also varied between isolates belonging to the same species, indicating that speciation events are important, but not sufficient to fully explain the diversity in the production of this antigen. The culture filtrate antigen AgEpm83, which was obtained from a PS3 isolate, showed large quantities of gp43 and reactivity by immunodiffusion assays, similar to the standard antigen (AgB-339) from an S1 isolate. Furthermore, AgEpm83 was capable of serologically differentiating five serum samples from patients from the Botucatu and Jundiaí regions. These patients had confirmed PCM but, were non-reactive to the standard antigen, thus demonstrating an alternative for serological diagnosis in regions in which S1 and PS2 occur. We also emphasise that it is not advisable to use a single antigen preparation to diagnose PCM, a disease that is caused by highly diverse pathogens.

Investigación y conservación de la biodiversidad en Perú: importancia del uso de técnicas modernas y procedimientos administrativos eficientes

Este comentario resalta la importancia que el análisis de secuencias de ADN tiene en los estudios de biodiversidad y la necesidad de mejorar los procedimientos administrativos concernientes a las investigaciones de biodiversidad en el Perú. La rápida pérdida de la biodiversidad y recursos naturales del Perú justifican la urgencia de apoyar aquellas investigaciones que ayuden a identificar, describir y caracterizar a la biodiversidad en la brevedad posible, para que se puedan tomar las debidas medidas de conservación y mitigación. Enfatizamos la importancia del uso de técnicas modernas dentro del modelo de estudio de la taxonomía integradora, incluyendo el estudio de los procesos evolutivos asociados a áreas con mayor diversidad y endemismo y los efectos del cambio climático sobre la biodiversidad peruana. Asimismo, es esencial que las entidades gubernamentales encargadas de evaluar los planes, solicitudes y requisitos asociados a las investigaciones de la biodiversidad del Perú reconozcan y apoyen el enfoque de la taxonomía integradora. El uso de técnicas modernas dentro del modelo de la taxonomía integradora, junto a procedimientos administrativos eficientes, puede convertirse en la mejor herramienta para proteger la biodiversidad peruana.

This commentary focuses on the importance that DNA sequence analysis has in biodiversity studies and the need for improving administrative procedures regarding biodiversity research in Peru. Given the rapid loss of biodiversity and natural resources in many areas of Peru, research aimed at identifying, characterizing, and describing biodiversity is a priority for developing appropriate conservation and mitigation strategies. For current and future research on Peruvian biodiversity and conservation, we emphasize the importance of using an integrative taxonomy approach, including the study of evolutionary processes associated with areas of high biodiversity and endemism. Additionally, it is essential that governmental institutions responsible for reviewing permit applications and research plans, focusing on biodiversity research in Peru, recognize and support studies that incorporate an integrative taxonomy approach. The use of modern techniques in the integrative taxonomy model in conjunction with efficient administrative procedures will be the best method for protecting Peruvian biodiversity.

Variación morfométrica intra e interespecífica entre poblaciones de Odontophrynus (Anura: Cycloramphidae) del área central de Argentina / Intraand inter-specific morphometric variation between Odontophrynus populations (Anura: Cycloramphidae) of central Argentina

Morphometric analyses are particularly important, and for many years they have supported evolutionary and ecological phenomena, and have been useful for the classification of new species, mainly to the lowest taxonomic levels. In order to assess the degree of sexual dimorphism, the intra-specific morphometric variation and the inter-specific morphological differences, we performed morphometric analyses of two morphologically cryptic species, Odontophrynus cordobae (diploid) and O. americanus (tetraploid). For this, we measured 15 morphometric variables on 211 individuals from 18 localities of Córdoba province. We found sexual dimorphism in six and three parameters in O. cordobae and O. americanus, respectively. Diploid and tetraploid males significantly differed in six morphometric variables. Discriminant Function Analysis (DFA) including all populations correctly classified a 76.37% of individuals within their respective species. DFA based on four groups (1- allopatric O. cordobae, 2-syntopic O. cordobae; 3- allopatric O. americanus; 4- syntopic O. americanus) accurately classified a 73.45% of individuals within their respective groups. Allopatric individuals of each species and, allopatric and syntopic individuals within each species were clearly segregated. Syntopic individuals of O. cordobae and O. americanus showed the lowest percentages of correct classification. Morphometric differences between sympatric diploids and tetraploids were not greater than those observed in allopatry. These findings deviate from the expected results under the hypothesis of character displacement, and they suggest that external morphological characters would not have a major influence on the recognition and choice of conspecific males by females.

El análisis morfométrico ha servido como apoyo para explicar fenómenos ecológicos y evolutivos y para la clasificación de nuevas especies. Nosotros realizamos el análisis morfométrico de dos especies morfológicamente crípticas con el objeto de evaluar el grado de diferenciación intra e interespecífica. Se midieron 15 variables sobre 211 individuos pertenecientes a 18 localidades del centro de Argentina. Se encontró dimorfismo sexual en seis variables morfométricas en Odontophrynus cordobae y en tres variables en O. americanus. Se encontraron diferencias significativas entre especies en seis variables. El análisis discriminante incluyendo los individuos de todas las poblaciones relevadas para O. cordobae y O. americanus, mostró una clasificación positiva de los individuos dentro de su respectiva especie del 76.37%. En el análisis discriminante realizado con base en cuatro grupos (O. cordobae alopátrico y sintópico y O. americanus alopátrico y sintópico) se obtuvo una reclasificación de los individuos dentro de cada grupo del 73.45%. Las diferencias morfométricas entre especies fueron mayores en sintopía. Los resultados se alejan de los resultados esperados bajo la hipótesis del desplazamiento de caracteres y podrían sugerir que los caracteres morfológicos externos no tendrían una influencia importante en el reconocimiento y elección de machos coespecíficos por parte de las hembras.

Molecular-based rapid inventories of sympatric diversity: A comparison of DNA barcode clustering methods applied to geography-based vs clade-based sampling of amphibians.

Molecular markers offer a universal source of data for quantifying biodiversity. DNA barcoding uses a standardized genetic marker and a curated reference database to identify known species and to reveal cryptic diversity within wellsampled clades. Rapid biological inventories, e.g. rapid assessment programs (RAPs), unlike most barcoding campaigns, are focused on particular geographic localities rather than on clades. Because of the potentially sparse phylogenetic sampling, the addition of DNA barcoding to RAPs may present a greater challenge for the identification of named species or for revealing cryptic diversity. In this article we evaluate the use of DNA barcoding for quantifying lineage diversity within a single sampling site as compared to clade-based sampling, and present examples from amphibians. We compared algorithms for identifying DNA barcode clusters (e.g. species, cryptic species or Evolutionary Significant Units) using previously published DNA barcode data obtained from geography-based sampling at a site in Central Panama, and from clade-based sampling in Madagascar. We found that clustering algorithms based on genetic distance performed similarly on sympatric as well as clade-based barcode data, while a promising coalescent-based method performed poorly on sympatric data. The various clustering algorithms were also compared in terms of speed and software implementation. Although each method has its shortcomings in certain contexts, we recommend the use of the ABGD method, which not only performs fairly well under either sampling method, but does so in a few seconds and with a user-friendly Web interface.

Aproximación a la filogenia de Spodoptera (Lepidoptera: Noctuidae) con el uso de un fragmento del gen de la citocromo oxidasa I (COI) / Approach to Spodoptera (Lepidoptera: Noctuidae) phylogeny based on the sequence of the cytocrhome oxydase I (COI) mitochondrial gene

The genus Spodoptera includes 30 species of moths considered important pests worldwide, with a great representation in the Western Hemisphere. In general, Noctuidae species have morphological similarities that have caused some difficulties for assertive species identification by conventional methods. The purpose of this work was to generate an approach to the genus phylogeny from several species of the genus Spodoptera and the species Bombyx mori as an out group, with the use of molecular tools. For this, a total of 102 S. frugiperda larvae were obtained at random in corn, cotton, rice, grass and sorghum, during late 2006 and early 2009, from Colombia. We took ADN samples from the larval posterior part and we analyzed a fragment of 451 base pairs of the mitochondrial gene cytochrome oxydase I (COI), to produce a maximum likelihood (ML) tree by using 62 sequences (29 Colombian haplotypes were used). Our results showed a great genetic differentiation (K2 distances) amongst S. frugiperda haplotypes from Colombia and the United States, condition supported by the estimators obtained for haplotype diversity and polymorphism. The obtained ML tree clustered most of the species with bootstrapping values from 73-99% in the interior branches; with low values also observed in some of the branches. In addition, this tree clustered two species of the Eastern hemisphere (S. littoralis and S. litura) and eight species of the Western hemisphere (S. androgea, S. dolichos, S. eridania, S. exigua, S. frugiperda, S. latifascia, S. ornithogalli and S. pulchella). In Colombia, S. frugiperda, S. ornithogalli and S. albula represent a group of species referred as “the Spodoptera complex” of cotton crops, and our work demonstrated that sequencing a fragment of the COI gene, allows researchers to differentiate the first two species, and thus it can be used as an alternative method to taxonomic keys based on morphology. Finally, the ML tree did not cluster S. frugiperda with S. ornithogalli, suggesting that both species do not share the same recent ancestral even though they coexist in cotton. We suggest sequencing other genes (mitochondrial and nuclear) to increase our understanding of this genus evolution.

En este trabajo se secuenció un fragmento de 451pb del gen mitocondrial de la citocromo oxidasa I (COI) en 62 secuencias del género Spodoptera y una secuencia de Bombix mori (grupo externo). Los resultados mostraron gran diferenciación genética (distancia K2) entre los haplotipos de Spodoptera frugiperda de Colombia y Estados Unidos, según los estimadores de diversidad haplotípica, diversidad y polimorfismo nucleotídicos calculados. Un árbol de ML agrupó las especies con valores de bootstrap entre 73-99% en las ramas internas. No obstante algunas ramas presentaron bajos valores de bootstrap. Este árbol formó un grupo constituido por las especies del hemisferio oriental (S. littoralis y S. litura) y también agrupó las especies localizadas en el hemisferio occidental (S. androgea, S. dolichos, S. eridania, S. exigua, S. frugiperda, S. latifascia, S. ornithogalli y S. pulchella). Esto demuestra que el árbol agrupó las especies con base en su origen geográfico. Contrariamente, el árbol no agrupó a S. frugiperda con S. ornithogalli, demostrando que a pesar de que ambas coexisten en el cultivo de algodón, no comparten un ancestro común reciente. En Colombia, estas especies forman parte del “complejo Spodoptera” del algodón, y nuestros resultados demuestran que la secuenciación de este gen permite diferenciarlas sin necesidad del uso de claves taxonómicas de sus estadios larvales. Este trabajo es una aproximación a la filogenia de este género, por lo cual la inclusión de más genes (mitocondriales y nucleares) son necesarios para futuros trabajos.

Karyotype description of possible new species of the Hypostomus ancistroides complex (Teleostei: Loricariidae) and other Hypostominae / Descrição cariotípica de possíveis novas espécies do complexo Hypostomus ancistroides e outros Hypostominae

Cytogenetic analyses were performed in four species of the Hypostominae subfamily, three from Hypostomus (Hypostomini) genus and Rhinelepis aspera (Rhinelepini). Three populations of Hypostomus ancistroides were analyzed, which had 2n=68 chromosomes, but presented different karyotype formulas. Hypostomus regani and H. strigaticeps, both from Ivaí river, showed 2n=72 chromosomes with two distinct cytotypes. In turn, R. aspera of the upper Paraná river basin presented 2n=54 chromosome. Multiple Nucleolar Organizer Regions (NORs) have been evidenced by silver nitrate staining in species of Hypostomus and single NOR in R. aspera. The observed variation in the chromosome number and the marked variability in karyotype formulas and NORs reveal a certain amount of karyotype variation in the genus Hypostomus suggesting the probable existence of cryptic species with independent chromosome traits. Therefore, our data can be of great value in discriminating species and understanding their chromosomal evolution.

Foram analisadas três populações de peixes identificadas como Hypostomus ancistroides, as quais apresentaram 2n=68 cromossomos, com distintas fórmulas cariotípicas. Hypostomus regani e H. strigaticeps, ambas do rio Ivaí, apresentaram 2n=72 cromossomos com citótipos distintos. Rinelepis aspera da bacia do alto rio Paraná apresentou 2n=54 cromossomos. Nucleolar Organizer Regions (NORs) múltiplas foram evidenciadas por nitrato de Prata para as espécies do gênero Hypostomus e NOR simples para R. aspera. A variação de número cromossômico observada, como também a acentuada variação nas fórmulas cariotípicas e nas NORs, são discutidas, sugerindo a existência de possíveis espécies crípticas com caracteres cromossômicos independentes. Portanto, nossos dados podem ser de grande valia na discriminação das espécies e no entendimento de sua evolução cromossômica.