Convergent and environmentally associated chromatic polymorphism in Bryconops Kner, 1858 (Ostariophysi: Characiformes: Iguanodectidae).

Bryconops Kner, 1858, includes two well defined subgenera based on morphological evidence, with each containing at least one species (B. (Bryconops) caudomaculatus and B. (Creatochanes) melanurus) with a very wide distribution, within which regional populations present color variations. To test if phenotypic variation is related to cladogenetic events, we performed tests for phylogenetic independence and determined the strength of convergence for color characters in relation to water type, as the variation between clear, black and white waters is considered to be one of the major driving forces in the evolution of Amazonian fishes. Color characters for fins above the median line of the body were generally found to be independent from phylogeny and the Wheatsheaf test strongly supports convergence of the dorsal fin color between populations of species in the same type of water, with a similar trend suggested for the color of the dorsal lobe of the caudal fin. This means that simple color characters cannot necessarily be relied upon for taxonomic revisions of the genus as local phenotypic variants may represent environmentally determined plasticity or convergent evolution. Further studies are required to determine the validity of these characters.

Metapopulation dynamics of the migratory fish Prochilodus lineatus (Characiformes: Prochilodontidae) in a lotic remnant of the Grande River, Southeastern Brazil

River impoundments for electricity generation lead to environmental changes which severely affect fish migration and species richness. However, little is known about their effect on the genetic structure and population dynamics downstream from the reservoir. Here, we analyzed a set of ten microsatellite loci of Prochilodus lineatus, an important South American migratory fish. Specimens (n = 150) were sampled from five sites in a remnant lotic system that includes sections of the Grande, Pardo and Mogi Guaçu rivers, southeastern Brazil. The data showed that all microsatellites were polymorphic with the allele number per locus ranging from 5 to 32, and genetic diversity (H e ) varied from 0.74 to 0.80. Indices of genetic differentiation and Bayesian analysis showed a significant genetic structure and three genetic clusters inhabiting this river system. An asymmetric gene flow suggests source-sink metapopulation dynamics from tributaries (genetic source) to the main river (genetic sink). A genetic cluster that was not detected in the upper Mogi and Pardo rivers tributaries may indicate there is a "trapped gene pool" downstream from the Porto Colômbia dam. Thus, here we provide new insights into the genetic structure and population dynamics of a migratory fish species in a highly dammed river basin.(AU)

Represamento de rios para geração de eletricidade levam a mudanças ambientais que afetam severamente a migração de peixes e riqueza de espécies. No entanto, pouco se sabe sobre seu efeito na estrutura genética e dinâmica populacional a jusante de reservatórios. Aqui, analisamos um conjunto de dez loci de microssatélites de Prochilodus lineatus, um importante peixe migratório sul-americano. Os espécimes (n = 150) foram amostrados em cinco locais de um sistema lótico remanescente que inclui seções dos rios Grande, Pardo e Mogi Guaçu, sudeste do Brasil. Os dados mostraram que todos microssatélites eram polimórficos com o número de alelos por locus variando de 5 a 32 e diversidade genética (H e ) variou de 0,74 a 0,80. Índices de diferenciação genética e análise de agrupamento baseada em modelo bayesiano indicou a presença de três agrupamentos genéticos habitando este sistema fluvial. Um fluxo gênico assimétrico sugere dinâmica metapopulacional de fonte-sumidouro dos tributários (fonte genética) para o rio principal (sumidouro genético). Um agrupamento genético que não foi detectado nos tributários rio Mogi e rio Pardo parecem indicar que há um "trapped gene pool" a jusante da represa de Porto Colômbia. Assim, nós provemos aqui novos conhecimentos sobre a estrutura genética e dinâmica populacional de uma espécie de peixe migratório em um rio altamente fragmentado por barramentos.(AU)

New 12S metabarcoding primers for enhanced Neotropical freshwater fish biodiversity assessment.

The megadiverse Neotropical fish fauna lacks a comprehensive and reliable DNA reference database, which hampers precise species identification and DNA based biodiversity assessment in the region. Here, we developed a mitochondrial 12S ribosomal DNA reference database for 67 fish species, representing 54 genera, 25 families, and six major Neotropical orders. We aimed to develop mini-barcode markers (i.e. amplicons with less than 200 bp) suitable for DNA metabarcoding by evaluating the taxonomic resolution of full-length and mini-barcodes and to determine a threshold value for fish species delimitation using 12S. Evaluation of the target amplicons demonstrated that both full-length library (565 bp) and mini-barcodes (193 bp) contain enough taxonomic resolution to differentiate all 67 fish species. For species delimitation, interspecific genetic distance threshold values of 0.4% and 0.55% were defined using full-length and mini-barcodes, respectively. A custom reference database and specific mini-barcode markers are important assets for ecoregion scale DNA based biodiversity assessments (such as environmental DNA) that can help with the complex task of conserving the megadiverse Neotropical ichthyofauna.

Genetic diversity and aquaculture conservation for a threatened Neotropical catfish

Due to the ecological importance of Lophiosilurus alexandri, the present work evaluated its genetic representativeness by comparing wild stocks to broodstocks that were kept at three restocking hatcheries along the São Francisco River. A total of 97 samples were genotyped for newly developed microsatellite markers. Low levels of genetic diversity (average alleles number of 4.2 alleles) were detected in all cases, being more severe in captive groups. Significant pairwise FST and DEST values, Structure, and DAPC analyses showed that wild animals were structured in two groups, and a third group was formed by captive animals, evidencing the need to adopt genetic criteria to retain genetic diversity in the hatcheries. For this reason, three full-sib families were constructed to select the best relatedness estimator for L. alexandri and establish a cut-off value aimed to avoid full-sibling matings in the hatcheries. Two estimators, Wang (RW) and Lynch & Li (RLL), were accurate in reflecting the relatedness level for full-sibs in this species. According to them, less than 50% of the potential breeding matings in the three hatcheries are advisable. The innate low diversity of L. alexandri highlights the importance of minimizing inbreeding and retaining genetic diversity towards the species recovery.(AU)

Devido à importância ecológica de Lophiosilurus alexandri, o presente trabalho avaliou sua representatividade genética, comparando estoques selvagens com plantéis de reprodutores de três larviculturas ao longo do Rio São Francisco. Noventa e sete amostras foram genotipadas com marcadores microssatélites recém-desenvolvidos. Baixos níveis de diversidade genética (número médio de alelos de 4,2) foram detectados em todos os casos, sendo mais severo no cativeiro. Os valores de FST e DEST par a par, as análises do Structure e DAPC mostraram a estruturação dos animais selvagens em dois grupos, e um terceiro formado pelas larviculturas, evidenciando a necessidade de adoção de critérios genéticos para retenção da diversidade genética no cativeiro. Por essa razão, três famílias de irmãos completos foram construídas para selecionar o melhor estimador de parentesco para a espécie e estabelecer os valores mínimos de corte para evitar cruzamentos indesejados. Dois estimadores, Wang (RW) e Lynch & Li (RLL), foram eficientes em refletir as relações de parentesco para irmãos completos nessa espécie. Segundo eles, menos de 50% dos potenciais cruzamentos são recomendáveis nas três larviculturas. A baixa diversidade genética inerente ao L. alexandri destaca a importância de minimizar a consanguinidade e evitar perda de diversidade genética, visando a recuperação da espécie.(AU)

Hidden Diversity Hampers Conservation Efforts in a Highly Impacted Neotropical River System.

Neotropical Rivers host a highly diverse ichthyofauna, but taxonomic uncertainty prevents appropriate conservation measures. The Doce River Basin (DRB), lying within two Brazilian threatened hotspots (Atlantic Forest and Brazilian Savanna) in south-east Brazil, faced the worst ever environmental accident reported for South American catchments, due to a dam collapse that spread toxic mining tailings along the course of its main river. Its ichthyofauna was known to comprise 71 native freshwater fish species, of which 13 endemic. Here, we build a DNA barcode library for the DRB ichthyofauna, using samples obtained before the 2015 mining disaster, in order to provide a more robust biodiversity record for this basin, as a baseline for future management actions. Throughout the whole DRB, we obtained a total of 306 barcodes, assigned to 69 putative species (with a mean of 4.54 barcodes per species), belonging to 45 genera, 18 families, and 5 orders. Average genetic distances within species, genus, and families were 2.59, 11.4, and 20.5%, respectively. The 69 species identified represent over 76% of the known DRB ichthyofauna, comprising 43 native (five endemic, of which three threatened by extinction), 13 already known introduced species, and 13 unknown species (such as Characidium sp., Neoplecostomus sp., and specimens identified only at the sub-family level Neoplecostominae, according to morphological identification provided by the museum collections). Over one fifth of all analyzed species (N = 16) had a mean intraspecific genetic divergence higher than 2%. An integrative approach, combining NND (nearest neighbor distance), BIN (barcode index number), ABGD (automatic barcode gap discovery), and bPTP (Bayesian Poisson Tree Processes model) analyses, suggested the occurrence of potential cryptic species, species complex, or historical errors in morphological identification. The evidence presented calls for a more robust, DNA-assisted cataloging of biodiversity-rich ecosystems, in order to enable effective monitoring and informed actions to preserve and restore these delicate habitats.

Revealing Hidden Diversity of the Underestimated Neotropical Ichthyofauna: DNA Barcoding in the Recently Described Genus Megaleporinus (Characiformes: Anostomidae).

Molecular studies have improved our knowledge on the neotropical ichthyofauna. DNA barcoding has successfully been used in fish species identification and in detecting cryptic diversity. Megaleporinus (Anostomidae) is a recently described freshwater fish genus within which taxonomic uncertainties remain. Here we assessed all nominal species of this genus using a DNA barcode approach (Cytochrome Oxidase subunit I) with a broad sampling to generate a reference library, characterize new molecular lineages, and test the hypothesis that some of the nominal species represent species complexes. The analyses identified 16 (ABGD and BIN) to 18 (ABGD, GMYC, and PTP) different molecular operational taxonomic units (MOTUs) within the 10 studied nominal species, indicating cryptic biodiversity and potential candidate species. Only Megaleporinus brinco, Megaleporinus garmani, and Megaleporinus elongatus showed correspondence between nominal species and MOTUs. Within six nominal species, a subdivision in two MOTUs was found, while Megaleporinus obtusidens was divided in three MOTUs, suggesting that DNA barcode is a very useful approach to identify the molecular lineages of Megaleporinus, even in the case of recent divergence (< 0.5 Ma). Our results thus provided molecular findings that can be used along with morphological traits to better define each species, including candidate new species. This is the most complete analysis of DNA barcode in this recently described genus, and considering its economic value, a precise species identification is quite desirable and fundamental for conservation of the whole biodiversity of this fish.

Analysis of propagule pressure and genetic diversity in the invasibility of a freshwater apex predator: the peacock bass (genus Cichla)

An important step in invasive biology is to assess biological variables that could be used to predict invasion success. The study of genetics, evolution, and interactions of invasive and native species in invaded ranges provides a unique opportunity to study processes in population genetics and the capability of a species' range expansion. Here, we used information from microsatellite DNA markers to test if genetic variation relates to propagule pressure in the successful invasion of an apex predator (the Amazonian cichlid Cichla) into Southeastern Brazilian River systems. Invasive populations of Cichla have negatively impacted many freshwater communities in Southeastern Brazil since the 1960s. Reduction of genetic variation was observed in all invasive populations for both Cichla kelberi (CK) and Cichla piquiti (CP). For instance, heterozygosity was lower in the invasive range when compared to native populations from the Amazon basin (CP HE = 0.179/0.44; CK HE = 0.258/0.536 respectively). Therefore, despite the successful invasion of Cichla in southeast Brazil, low genetic diversity was observed in the introduced populations. We suggest that a combination of factors, such as Cichla's reproductive and feeding strategies, the "evolutionary trap" effect and the biotic resistance hypothesis, overcome their depauperete genetic diversity, being key aspects in this apex predator invasion. Uma importante etapa na biologia da invasão é acessar variáveis biológicas que podem predizer o sucesso de invasão. O estudo da genética, evolução e interações entre invasores e espécies nativas no ambiente invadido pode prover uma oportunidade única para o estudo dos processos em genética de populações e a capacidade de uma espécie ampliar seu habitat. Nesse trabalho, nos utilizamos dados de marcadores de DNA microssatélites para testar se a variação genética é relacionada a pressão de propágulo na invasão bem sucedida do predador de topo (o ciclídeo Amazônico Cichla) nos rios do Sudeste Brasileiro. Populações invasoras de Cichla vem impactando negativamente diversas comunidades de água doce no Sudeste brasileiro deste 1960. A redução da variação genética foi observada em todas populações invasoras, tanto para Cichla kelberi (CK) como Cichla piquiti (CP). Por exemplo, a heterozigose foi menor no ambiente invadido quando comparada com as populações nativas da bacia Amazônica (CP HE = 0.179/0.44; CK HE = 0.258/0.536 respectivamente). Assim, apesar do sucesso da invasão de Cichla no sudoeste do Brasil, baixa diversidade genética foi observada nas populações introduzidas. Nós sugerimos que uma combinação de fatores, como as estratégias reprodutivas de Cichla, o efeito de "armadilha evolutiva" e a hipótese de resistências biótica superam o efeito que a diversidade genética depauperada exerce, sendo aspectos-chave na invasão desse predador de topo de cadeia.

Genetic diversity patterns of Haemonchus placei and Haemonchus contortus populations isolated from domestic ruminants in Brazil.

Parasitic nematodes of the genus Haemonchus infect a range of ruminant hosts and are of major veterinary and economic importance. In this study, the genetic variability of seven isolates of Haemonchus placei and Haemonchus contortus was evaluated using the mitochondrial gene cytochrome oxidase subunit I and the nuclear gene b-tubulin isotype 1. A total of 156 specimens were obtained from cattle, sheep, goat and buffalo herds raised on commercial properties from the southern and southeastern regions of Brazil and identified to the species level by sequencing of the nuclear internal transcribed spacer 2. Thirty-four percent of the specimens were identified as H. placei and 66% as H. contortus. Cattle were the preferred hosts for H. placei, whereas H. contortus was most frequent in the other three ruminant species. Analysis of genetic differentiation between isolates revealed that high rates of gene flow are operating among populations of both nematode species, including among those from different ruminant host species. Populations of H. placei were less polymorphic and presented a lower frequency of single nucleotide polymorphisms associated with benzimidazole resistance compared with H. contortus. In line with the low amount of genetic structure observed among isolates, alleles of b-tubulin 1 associated with benzimidazole resistance were present at relatively high frequencies of 5­20% in isolates of H. contortus from farms that never used this class of anthelmintic. The results presented here are consistent with the hypothesis of multiple origins of alleles associated with benzimidazole resistance, with the trade of animals among properties acting as the main factor promoting the spread of anthelmintic resistance.

Deep barcode divergence in Brazilian freshwater fishes: the case of the São Francisco River basin.

BACKGROUND AND AIMS: The application of DNA barcoding as a global standard for fish identification is probing diverse worldwide realms (Nearctic, Australian and the Neotropics) and environments (e.g. marine and freshwater). Comparing the patterns of sequence divergence among conspecific and congeneric taxa between realms can provide valuable information on recent evolutionary histories of lineages as barcode data accumulates. MATERIALS AND METHODS: Herein, we have analyzed over 100 species (around 50%) of the Neotropical fish fauna from the São Francisco River, in southeast Brazil. Our aims were to test the performance of DNA barcoding in this biodiversity-rich region, and to compare patterns of genetic divergence with previous studies. RESULTS: The mean Kimura two-parameter distances within species, genera, families, orders, and classes were 0.5, 10.6, 21.0, 22.7, and 24.4%, respectively, with 100% of the species examined successfully differentiated by barcoding. With the exception of Astyanax bimaculatus lacustris, Piabina argentea, and Bryconamericus stramineus, all other species yield a single, cohesive cluster of barcode sequences. The average 'nearest-neighbor distance' was 11.12%, 21-fold higher than the mean within species distance of around 0.54%. In a few instances, deep lineage divergences among conspecifics (up to 10%) and congenerics (up to 22.9%) taxa were revealed. CONCLUSIONS: Reflecting possible cases of cryptic speciation and the deeper phylogeographic history of São Francisco fish fauna, with some higher clades extending back into the late Cretaceous and Cenozoic (90 mya), when much of the diversification of the Neotropical region apparently took place. In addition, barcodes also highlighted misidentifications and helped to document range extensions for known species.

DNA barcoding unveils a high rate of mislabeling in a commercial freshwater catfish from Brazil.

BACKGROUND AND AIMS: Molecular markers have contributed to species authentication by flagging mislabeling and the misidentification of commercial landings. Such tools are of great value since the market substitution of fish of lower value for highly commercialized species is expected to become more pronounced due to a shortage of natural stocks. MATERIALS AND METHODS: Here we report on the molecular identification 4results from processed fish products (i.e. fillets) and whole fishes sold in Brazilian markets under the common name surubim (Pseudoplatystoma spp.). RESULTS: DNA barcoding revealed the incorrect labeling of around 80% of all samples analyzed, with mislabeling being more pronounced within fillets rather than whole fish. CONCLUSION: To our knowledge, this is the first report correlating the rate of fraud with processed fish products. The establishment of an official list of acceptable common names for freshwater fish and seafood is urgently needed in Brazil for further trade regulations to take place.