Mitochondrial DNA diversity and maternal origins of Pakistani donkey / Diversidade de DNA mitocondrial e origens maternas do burro do Paquistão

Domestic donkey plays a key role as a draft animal in rural economy of Pakistan where its population is increasing every year. The complete mtDNA control region of forty randomly sampled donkeys was PCR- amplified and sequenced bi-directionally using specific primers. Distinct mtDNA haplotypes obtained in the current study (KY446001−KY446011) were subjected to haplotype (h) and nucleotide diversity (π) measures using DnaS as well as to phylogenetic, Network, and AMOVA analyses. There were a total 27 polymorphic sites present within 11 unique mtDNA haplotypes from the studied 40 animals from different regions. Neighbor-joining network and median-joining network both illustrated the splitting of all these haplotypes into two well-defined Nubian and Somali lineages, confirming African maternal origin of Pakistani domestic donkey. Diversity parameters h (0.967± 0.037) and π (0.02917± 0.00307) were found to reveal high levels of genetic diversity in Pakistani donkeys. AMOVA demonstrated only 1% of genetic differences between two mtDNA maternal lineages, pointing to lack of population substructure in Pakistani donkeys as is the case with worldwide domestic donkey population. Pakistani donkeys have African maternal origin and high levels of mtDNA diversity. High genetic diversity may be due to non-selective breeding and heteroplasmy. We herein provide the first report on mtDNA diversity of control region in Pakistani domestic donkey.

O burro doméstico possui um papel fundamental como animal de tração na economia rural do Paquistão, onde a população desse animal está aumentando a cada ano. A região de controle de mtDNA completa de 40 burros amostrados aleatoriamente foi ampliada por PCR e sequenciada bidirecionalmente por intermédio de primers específicos. Haplótipos distintos de mtDNA obtidos no estudo atual (KY446001 − KY446011) foram submetidos a medidas de haplótipo (h) e diversidade de nucleotídeos (π) por meio de DnaS, bem como análises filogenéticas, de rede e AMOVA. Havia um total de 27 sítios polimórficos presentes em 11 haplótipos de mtDNA exclusivos dos 40 animais estudados de diferentes regiões. A rede de união de vizinhos e a rede de união mediana ilustram a divisão de todos esses haplótipos em duas linhagens núbias e somalis bem definidas, confirmando a origem materna africana do burro doméstico do Paquistão. Os parâmetros de diversidade h (0,967 ± 0,037) e π (0,02917 ± 0,00307) revelaram altos níveis de diversidade genética em burros paquistaneses. AMOVA demonstrou apenas 1% de diferenças genéticas entre as duas linhagens maternas de mtDNA, apontando a falta de subestrutura populacional em burros paquistaneses, como é o caso da população mundial de burros domésticos. Os burros paquistaneses têm origem materna africana e altos níveis de diversidade de mtDNA. A alta diversidade genética pode ser por causa da reprodução não seletiva e de heteroplasmia. Aqui, fornecemos o primeiro relatório sobre a diversidade do mtDNA da região de controle em burros domésticos do Paquistão

Intravitreal injection of mitochondrial DNA induces cell damage and retinal dysfunction in rats

BACKGROUND: Retinal neurodegeneration is induced by a variety of environmental insults and stresses, but the exact mechanisms are unclear. In the present study, we explored the involvement of cytosolic mitochondrial DNA (mtDNA), resulting in the cGAS-STING dependent inflammatory response and apoptosis in retinal damage in vivo. METHODS: Retinal injury was induced with white light or intravitreal injection of lipopolysaccharide (LPS). After light-or LPS-induced injury, the amount of cytosolic mtDNA in the retina was detected by PCR. The mtDNA was isolated and used to transfect retinas in vivo. WB and real-time PCR were used to evaluate the activation of cGAS-STING path-way and the levels of apoptosis-associated protein at different times after mtDNA injection. Retinal cell apoptosis rate was detected by TUNEL staining. Full-field electroretinography (ERG) was used to assess the retinal function. RESULTS: Light injury and the intravitreal injection of LPS both caused the leakage of mtDNA into the cytoplasm in retinal tissue. After the transfection of mtDNA in vivo, the levels of cGAS, STING, and IFN-ß mRNAs and the protein levels of STING, phosph-TBK1, phospho-IRF3, and IFN-ß were upregulated. mtDNA injection also induced the activation of caspase 3 and caspase 9. BAX and BAK were increased at both the mRNA and protein levels. The release of cytochrome c from the mitochondria to the cytosol was increased after mtDNA injection. The wave amplitudes on ERG decreased and retinal cell apoptosis was detected after mtDNA injection. CONCLUSIONS: Cytosolic mtDNA triggers an inflammatory response. It also promotes apoptosis and the dysfunction of the retina.

CO1-Based DNA barcoding for assessing diversity of Pteropus giganteus from the state of Azad Jammu Kashmir, Pakistan / Código de barras de DNA à base de CO1 para avaliar a diversidade de Pteropus giganteus do estado de Azad Jammu e Caxemira, Paquistão

Abstract The flying fox (Pteropus giganteus) also familiar with the name of the greater Indian fruit Bat belongs to the order Chiroptera and family Pteropodidae. Current research emphasis on the DNA barcoding of P. giganteus in Azad Jammu Kashmir. Bat sequences were amplified and PCR products were sequenced and examined by bioinformatics software. Congeneric and conspecific, nucleotide composition and K2P nucleotide deviation, haplotype diversity and the number of haplotypes were estimated. The analysis showed that all of the five studied samples of P. giganteus had low G contents (G 19.8%) than C (27.8%), A (25.1%) and T (27.3%) contents. The calculated haplotype diversity was 0.60% and the mean intraspecific K2P distance was 0.001% having a high number of transitional substitutions. The study suggested that P. giganteus (R=0.00) do not deviate from the neutral evolution. It was determined from the conclusion that this mtDNA gene is a better marker for identification of Bat species than nuclear genes due to its distinctive characteristics and may serve as a landmark for the identification of interconnected species at the molecular level and in the determination of population genetics.

Resumo A raposa-voadora (Pteropus giganteus), também conhecida como morcego indiano, pertence à ordem dos Chiroptera e à família Pteropodidae. A presente pesquisa dá ênfase ao código de barras de DNA de P. giganteus em Azad Jammu e Caxemira. Sequências genéticas dos morcegos foram amplificadas, e os produtos de PCR foram sequenciados e examinados por software de bioinformática. De espécies congenérica e coespecífica, foram estimados composição nucleotídica e desvio de nucleotídeos K2P, diversidade de haplótipos e número de haplótipos. A análise mostrou que todas as cinco amostras estudadas de P. giganteus apresentaram baixos teores de G (19,8%) em comparação com C (27,8%), A (25,1%) e T (27,3%). A diversidade de haplótipos calculada foi de 0,60%, e a distância média intraespecífica de K2P foi de 0,001%, com um elevado número de substituições transicionais. O estudo sugeriu que P. giganteus (R = 0,00) não se desviou da evolução neutra. É possível concluir que o gene mtDNA é um marcador favorável para identificação de espécies de morcegos do que genes nucleares por causa de suas características distintivas e pode servir como um marco para a identificação de espécies interconectadas em nível molecular e para a determinação genética de populações.

Genetic diversity in two threatened species of guitarfish (Elasmobranchii: Rhinobatidae) from the Brazilian and Argentinian coasts: an alert for conservation

The guitarfishes Pseudobatos horkelii and Pseudobatos percellens meet the criteria for threatened status as Critically Endangered (CR) and Endangered (EN), respectively. Both species occur in the Southern Atlantic Ocean. Considering the lack of data on the genetic structure of these species, the present study evaluated the genetic variability and population structure of the P. horkelii and P. percellens in the southern region of Brazil and the northern coast of Argentina, based on sequences of mitochondrial DNA, Control Region (D-loop). Samples of P. horkelii (n = 135) were analyzed in six localities situated in Northern Argentina, along the Brazilian states' coast. The mean of nucleotide diversity was 0.0053, the ΦST was 0.4277 and demographic analysis of P. horkelii suggests the existence of stability of the populations, with D = 0.9929, FS = 2.0155, SSD = 0.0817, R = 0.2153. In P. percellens (n = 101) were analyzed from six Brazilian localities along the coast of Santa Catarina, Paraná, and São Paulo. The mean nucleotide diversity was 0.0014 and ΦST value of 0.2921, the demographic analysis indicates a high migration rate of P. percellens among the localities evaluated, with D = 0.5222, FS = 0.3528, SSD = 0.01785, R = 0.3890.(AU)

As raias violas Pseudobatos horkelii e Pseudobatos percellens, são listados como "Criticamente em Perigo" (CR) e "Em Perigo" (EN), respectivamente. Ambas as espécies ocorrem no Sul do Oceano Atlântico. Considerando a falta de dados sobre a estrutura genética dessas espécies, o presente estudo avaliou a variabilidade genética e a estrutura populacional de P. horkelii e P. percellens na região sudeste do Brasil e litoral norte da Argentina, com base em sequências de DNA mitocondrial, região de controle (D-loop). Amostras de 135 indivíduos de P. horkelii analisados em seis localidades, situadas no norte da Argentina e ao longo da costa dos estados brasileiros. A média da diversidade nucleotídica foi de 0.0053, o índice ΦST foi de 0.4277 e a análise demográfica de P. horkelii, indicou a existência de estabilidade das populações, com D = 0.9929, Fus = 2.0155, SSD = 0.0817, R = 0.2153. Em 101 exemplares de P. percellens, foram analisados em seis localidades brasileiras ao longo do litoral de Santa Catarina, Paraná e São Paulo. A diversidade nucleotídica média foi de 0.0014 e o valor ΦST de 0.2921, a análise demográfica indicou uma alta taxa de migração de P. percellens entre as localidades analisadas, com D = 0.5222, FS = 0.3528, SSD = 0.01785, R = 0.3890.(AU)

Hydrography rather than lip morphology better explains the evolutionary relationship between Gymnogeophagus labiatus and G. lacustris in Southern Brazil (Cichlidae: Geophagini)

Gymnogeophagus labiatus and G. lacustris have been long recognized as sister species exhibiting different ecological requirements. Gymnogeophagus labiatus occurs in rock bottom rivers in the hydrographic basins of Patos Lagoon (HBP) and Tramandaí River (HBT), while G. lacustris is exclusive from sand bottom coastal lagoons of the HBT. In this study, we used molecular markers, morphological measurements and data from nuptial male coloration to investigate the evolutionary relationship between these species in each hydrographic basin. We found, for all data sets, a closer relationship between G. labiatus and G. lacustris from the HBT than between G. labiatus populations from HBT and HBP. In particular, lip area had a large intraspecific plasticity, being uninformative to diagnose G. lacustris from G. labiatus. Molecular clock-based estimates suggest a recent divergence between species in the HBT (17,000 years ago), but not between G. labiatus from HBP and HBT (3.6 millions of years ago). Finally, we also found a divergent G. labiatus genetic lineage from the Camaquã River, in the HBP. These results show that the current taxonomy of G. labiatus and G. lacustris does not properly represent evolutionary lineages in these species.(AU)

Gymnogeophagus labiatus e G. lacustris vêm sendo consideradas espécies irmãs que possuem diferentes exigências ecológicas. Gymnogeophagus labiatus ocorre em rios de fundo de pedra nas bacias hidrográficas da Laguna dos Patos (HBP) e do rio Tramandaí (HBT), enquanto G. lacustris é exclusivo da HBT, ocorrendo em lagoas costeiras de fundo de arenoso. Nesse estudo, foram usados marcadores moleculares, medidas morfológicas e dados sobre a coloração nupcial em machos para investigar a relação evolutiva entre estas espécies em cada bacia hidrográfica. Para todos os conjuntos de dados foi observada uma relação mais próxima entre G. labiatus e G. lacustris da HBT do que entre as populações de G. labiatus da HBP e HBT. Em particular, a área do lábio teve uma grande plasticidade intraespecífica, não sendo informativa para diagnosticar G. lacustris de G. labiatus. Estimativas baseadas no relógio molecular sugeriram uma divergência recente entre as espécies da HBT (17.000 anos atrás), mas não entre as populações de G. labiatus da HBP e HBT (3,6 milhões de anos atrás). Finalmente, também foi encontrada uma linhagem genética de G. labiatus divergente no rio Camaquã, na HBP. Esses resultados mostram que a taxonomia atual de G. labiatus e G. lacustris não representa adequadamente as linhagens evolutivas nessas espécies.(AU)

Técnicas de substituição de DNA mitocondrial para reduzir a transmissão da Síndrome de Leigh / Mitochondrial DNA replacement techniques to reduce Leigh syndrome transmission

A Síndrome de Leigh (SL) é uma doença neuro-metabólica congênita, que faz parte do grupo das encefalopatias fatais, com progressão e morte dentro de 2 anos, em média. A SL é causada por mutações no DNA que causam alterações na geração de ATP celular pelas mitocôndrias. As mitocôndrias contêm seu próprio DNA (mtDNA) e, ao contrário do DNA nuclear, o mtDNA é herdado somente da mãe. Mulheres portadores de mutações causadoras da SL podem vivenciar experiências muito tristes ao tentarem realizar o sonho da maternidade. As técnicas de substituição de mtDNA mutado com mtDNA saudável de doadora, oferecem a essas mulheres a possibilidade de terem uma criança geneticamente relacionada sem a SL. O desenvolvimento e a aplicação clínica de terapias de substituição de mtDNA já são uma realidade, tendo o primeiro bebê gerado a partir da técnica nascido em 2016. Mas será que essas técnicas são seguras? Neste trabalho, revisamos a SL e algumas técnicas de substituição de mtDNA já aplicadas em humanos, que envolvem a transferência de pronúcleos de zigotos ou de fuso acromático de oócitos. Concluímos que, apesar dos resultados promissores, ainda é cedo para assegurar a aplicabilidade clínica de técnicas de substituição de mtDNA em seres humanos. (AU)

Leigh syndrome (SL) is a congenital neurometabolic disease included in the group of fatal encephalopathies, with progression and death within 2 years on average. SL is caused by mutations in the DNA that cause changes in the generation of cellular ATP by mitochondria. Mitochondria contain their own DNA (mtDNA) and, unlike nuclear DNA, mtDNA is inherited only from the mother. Women with SL mutations may experience mournful situations when attempting to fulfill the dream of motherhood. Techniques for replacing mutant mtDNA with healthy donor mtDNA provide these women with the possibility of having a genetically related child without SL. The development and clinical application of mtDNA replacement therapies is a reality, and the first baby generated using the technique was born in 2016. However, are these techniques safe? In this article, we review SL and some mtDNA replacement techniques that have been used in humans, which involve zygote pronuclear transfer or oocyte spindle transfer. We conclude that, despite the promising results, it is too early to ensure that mtDNA replacement techniques are clinically applicable to humans. (AU)

Association of whole mtDNA, an NADPH G11914A variant, and haplogroups with high physical performance in an elite military troop

Physical performance is a multifactorial and complex trait influenced by environmental and hereditary factors. Environmental factors alone have been insufficient to characterize all outstanding phenotypes. Recent advances in genomic technologies have enabled the investigation of whole nuclear and mitochondrial genome sequences, increasing our ability to understand interindividual variability in physical performance. Our objective was to evaluate the association of mitochondrial polymorphic loci with physical performance in Brazilian elite military personnel. Eighty-eight male military personnel who participated in the Command Actions Course of the Army were selected. Total DNA was obtained from blood samples and a complete mitochondrial genome (mtDNA) was sequenced using Illumina MiSeq platform. Twenty-nine subjects completed the training program (FINISHED, 'F'), and fifty-nine failed to complete (NOT_FINISHED, 'NF'). The mtDNA from NF was slightly more similar to genomes from African countries frequently related to endurance level. Twenty-two distinct mtDNA haplogroups were identified corroborating the intense genetic admixture of the Brazilian population, but their distribution was similar between the two groups (FST=0.0009). Of 745 polymorphisms detected in the mtDNA, the position G11914A within the NADPH gene component of the electron transport chain, was statistically different between F and NF groups (P=0.011; OR: 4.286; 95%CI: 1.198-16.719), with a higher frequency of the G allele in group F individuals). The high performance of military personnel may be mediated by performance-related genomic traits. Thus, mitochondrial genetic markers such as the ND4 gene may play an important role on physical performance variability.

Quantification of mitochondrial DNA damage and copy number in circulating blood of patients with systemic sclerosis by a qPCR-based assay

Abstract Background: Although not fully understood, oxidative stress has been implicated in the pathogenesis of different autoimmune diseases such as systemic sclerosis. Accumulating evidence indicates that oxidative stress can induce mitochondrial DNA (mtDNA) damage and variations in mtDNA copy number (mtDNAcn). Objective: The aim of this study was to explore mtDNAcn and oxidative DNA damage byproducts in peripheral blood of patients with systemic sclerosis and healthy controls. Methods: Forty six patients with systemic sclerosis and forty nine healthy subjects were studied. Quantitative real-time PCR used to measure the relative mtDNAcn and the oxidative damage (oxidized purines) of each sample. Results: The mean mtDNAcn was lower in patients with systemic sclerosis than in healthy controls whereas the degree of mtDNA damage was significantly higher in cases as compared to controls. Moreover, there was a negative correlation between mtDNAcn and oxidative DNA damage. Study limitations: The lack of simultaneous analysis and quantification of DNA oxidative damage markers in serum or urine of patients with systemic sclerosis and healthy controls. Conclusion: These data suggest that alteration in mtDNAcn and increased oxidative DNA damage may be involved in the pathogenesis of systemic sclerosis.

A Century after! Rediscovery of the ancient catfish Diplomystes Bleeker 1858 (Siluriformes: Diplomystidae) in coastal river basins of Chile and its implications for conservation

The ancient catfish family Diplomystidae, with seven species endemic to rivers of southern South America, represents one of the oldest branches of the diverse order Siluriformes. With most species endangered, new reports of these species become extremely valuable for conservation. Currently, it is assumed that Diplomystes species inhabit only Andean (large) basins, and that they are extinct from coastal (small) basins from which their presence have not been recorded since 1919. Here, we document new records of the family Diplomystidae in the Laraquete and Carampangue basins, two coastal basins from the Nahuelbuta Coast Range, Chile, with no previous reports. This finding represents the rediscovery of the genus in coastal basins in more than a Century. Based on analysis of mitochondrial DNA sequences, the collected specimens were found to be closely related to Diplomystes nahuelbutaensis from the Andean Biobío Basin, but sufficiently differentiated to suggest that coastal basin populations are a different management unit. These populations are important because, contrary to previous thoughts, they prove these catfish can survive in small river networks, providing unique opportunities for research and conservation. The conservation category of Critically Endangered (CE) is recommended for the populations from the Laraquete and Carampangue basins.(AU)

La familia de bagres Diplomystidae, con siete especies endémicas de ríos del sur de Sudamérica, es uno de los linajes mas antiguos del diverso orden Siluriformes. Al estar la mayoría de las especies amenazadas, nuevos registros de éstas son extremadamente valiosos para su conservación. Actualmente, se ha asumido que los Diplomystidos se distribuyen solo en cuencas Andinas (más grandes), y que sus especies estarían extintas en cuencas de menor tamaño como las costeras, sin registros desde 1919. En este trabajo documentamos la familia Diplomistidae en las cuencas de Carampangue y Laraquete, dos cuencas costeras de la Cordillera de Nahuelbuta, Chile, lo que representa el primer registro de esta familia en estas cuencas costeras. Además, este hallazgo representa el re-descubrimiento de la familia en cuencas costeras después de un siglo. Sobre la base de análisis de ADN mitocondrial, los especímenes colectados se relacionaron más cercanamente con poblaciones de la especie Diplomystes nahuelbutaensis presente en la cuenca del Biobío. Sin embargo, existen diferencias genéticas suficientes entre las poblaciones costeras y las del Biobío para justificar su separación como unidad de manejo distinta. Estas poblaciones costeras son importantes porque demuestran que los Diplomístidos pueden sobrevivir en cuencas de pequeño tamaño, ofreciendo oportunidades únicas para su investigación y conservación. Se recomienda la categoría de conservación En Peligro Critico de Extinción (CR) para las poblaciones de las cuencas Laraquete y Carampangue.(AU)

Molecular phylogenetics of Black Cobra (Naja naja) in Pakistan

Background: Snakes are found on every continent in the world except Antarctica, and on smaller land masses. Being ecologically important, they also cause a large number of bites, leading to millions of deaths. Mitochondrial and nuclear gene sequences are being used to identify, characterize, and infer genetic biodiversity among different snake species. Furthermore, phylogenetics helps in inferring the relationships and evolutionary histories among these species. Black cobra is one of the four most venomous snakes in Pakistan. Four mitochondrial (ND4, Cytochrome b, 12S rRNA, and 16S rRNA) and four nuclear (C-mos, RAG-1, BDNF, and NT3) genes were used to trace diversity and infer the phylogenetic relationship of black cobra in Pakistan. Results: Almost similar phylogenies were obtained through maximum likelihood and Bayesian inference, showing two species of cobra in Pakistan, namely, black cobra (Naja naja) and brown cobra (Naja oxiana). All Naja species were divided into three clades: black cobra (N. naja) and brown cobra (N. oxiana) cladding with different species of Naja; N. naja (Pakistan) cladding with N. naja from Nepal; and N. oxiana showed close relationship with Naja kaouthia from Thailand and Naja siamensis from Thailand. Conclusion: It was confirmed genetically that there are two cobra species in Pakistan, i.e., black and brown cobras. This study will help in not only genetic conservation but also developing anti-venom against snake species.

Whole sequencing of the mitochondrial genome of breast cancer tissue in Mexican-mestizo postmenopausal women with different body mass index

Abstract Background Mitochondrial and oxidative stress has been related to obesity and breast cancer being this cancer more frequent and more aggressive in postmenopausal women with obesity. Objective The objective of this study was to investigate whether Mexican-Mestizo postmenopausal women with breast cancer and obesity present different somatic mutations in the mitochondrial DNA (mtDNA) when compared to women with normal body mass index (BMI). Subjects and Methods We included six Mexican-Mestizo postmenopausal women bearing breast cancer and who underwent mastectomy or breast-conserving surgery. BMI was determined in each case. Patients’ genomic DNA was isolated from blood leukocytes and tumor tissue samples. Whole mtDNA sequence was determined by MitoChip v2.0 mitochondrial resequencing array, and data were analyzed using the GeneChip Sequence Analysis Software. Tumor mtDNA sequence was compared with matched leukocyte mtDNA sequence. Results Three women had a normal BMI and three presented obesity. Overall, we found 64 genetic variants: 53.1% were somatic mutations and 46.9% were polymorphisms; 44.1% were in the non-coding region and 55.9% were in genes that encode for mitochondrial proteins. Among the somatic mutations, 67.7% were in patients with normal BMI and 32.3% in patients with obesity. Conclusions We did not find a higher frequency of mitochondrial somatic mutations in postmenopausal women with breast cancer and obesity compared to those with normal BMI. However, results could be due to the small number of women studied.

Molecular identification and histological aspects of Renicola sloanei (Digenea: Renicolidae) in Puffinus puffinus (Procellariiformes): a first record / Identificação molecular e aspectos histológicos de Renicola sloanei (Digenea: Renicolidae) em Puffinus puffinus (Procellariiformes): primeiros registros

Abstract Renicolids are parasites that inhabit the renal tubules and ureters of molluscivorous and piscivorous birds. Puffinus puffinus is a migratory seabird that was identified as the definitive host of Renicola spp. Studies focusing on the renicolid species and the resulting renal lesions are valuable for their association with causes of stranding in seabirds. The aim of this study was to identify the renicolid trematodes and evaluate the histological findings in two P. puffinus stranded on the coast of Paraná state, Brazil. The parasites were evaluated by histologic, ultrastructural and molecular assays, while tissue changes were analyzed by histologic methods. The morphological and morphometrical characteristics of the parasites, along with polymerase chain reaction and sequencing assays (ribosomal and mitochondrial regions), identified the species as Renicola sloanei. The results also suggest that this helminth can be the adult form of Cercaria pythionike. The dilation of collecting ducts was the main histological finding in the kidneys. In conclusion, R. sloanei was identified, and for the first time, P. puffinus was described as a host of this digenean inducing mild renal changes.

Resumo Renicolídeos são parasitos que habitam túbulos renais e ureteres de aves que se alimentam de moluscos e peixes. Puffinus puffinus, ave marinha migratória, foi registrada como hospedeiro definitivo de Renicola spp. Estudos relacionados com as espécies de renicolídeos e as lesões renais resultantes são importantes para o entendimento das causas de óbito de aves marinhas. O objetivo deste estudo foi identificar os trematódeos renicolídeos e avaliar os achados histológicos em dois P. puffinus encalhados no litoral do Estado do Paraná, Brasil. Os parasitos foram avaliados por ensaios histológicos, ultraestruturais e moleculares, enquanto as alterações teciduais foram analisadas por métodos histológicos. As características morfológicas e morfométricas dos parasitos, juntamente com a reação em cadeia da polimerase e sequenciamento (regiões ribossomal e mitocondrial), identificaram a espécie como Renicola sloanei. Os resultados também sugerem que este helminto pode ser a forma adulta de Cercaria pythionike. A dilatação dos ductos coletores foi o principal achado histológico renal. Em conclusão, R. sloanei foi identificado, e pela primeira vez P. puffinus foi descrito como hospedeiro deste digenético induzindo alterações renais discretas.

Multiloci analyses suggest synonymy among Rhomboplites, Ocyurus and Lutjanus and reveal the phylogenetic position of Lutjanus alexandrei (Lutjanidae: Perciformes)

Lutjanidae comprises 21 genera and 135 species widespread throughout Atlantic, Indian and Pacific oceans. Nonetheless, the phylogenetic relationships of Lutjaninae remain uncertain. Furthermore, phylogenetic hypotheses for Lutjanus alexandrei, an endemic species from northeastern Brazilian coast, in Lutjanidae are absent so far. Therefore, we carried out multiloci analyses, combining both mitochondrial and nuclear DNA sequences in Lutjaninae species from Western Atlantic focusing on the controversial relationships among Lutjanus, Rhomboplites, and Ocyurus. Besides, we determined the phylogenetic position and dated the origin of L. alexandrei. The phylogenetics trees based on the 4.4 kb for 11 species corroborated the synonym among Lutjanus and the putative monotypic genera. For the dating of L. alexandrei, another nucleotide dataset (3.0 kb; 40 species) validated the genetic identity of this species that diverged from the sister taxon L. apodus between 2.5 - 6.5 Mya, probably as a result of the barrier caused by the muddy outflow from Orinoco and Amazon rivers along the coastal zone. This report is the most robust multiloci analysis to confirm the synonymy of the three genera of Lutjaninae from Western Atlantic and the first reliable inference about the phylogenetic relationships and origin of L. alexandrei.(AU)

A Família Lutjanidae compreende 21 gêneros e 135 espécies, distribuídas ao longo dos oceanos Atlântico, Índico e Pacífico. As relações filogenéticas dos Lutjaninae são incertas. Além disso, a espécie Lutjanus alexandrei, endêmica da costa nordeste do Brasil, não foi inclusa em nenhuma hipótese filogenética até o presente. Assim, realizamos uma análise integrando DNA mitocondrial e nuclear para espécies de Lutjaninae do Atlântico Ocidental, direcionada para a controversa relação entre Lutjanus, Rhomboplites e Ocyurus. Além disso, alocamos filogeneticamente L. alexandrei e datamos sua origem. As árvores filogenéticas baseadas em 4.4 kb de 11 espécies corroboraram a sinonímia entre os monotípicos e Lutjanus. Para a datação de L. alexandrei, outro banco de nuclueotídeos foi analisado (3.0 kb; 40 espécies), validando geneticamente a espécie e a colocando como irmã de L. apodus, da qual se separou entre 2.5 - 6.5 Mya, o que provavelmente foi provocado pela faixa enlameada na região costeira, influenciada pelas descargas dos rios Amazonas e Orinoco, que funciona como barreira. Este trabalho representa a mais robusta análise multiloci direcionada para a sinonimização dos três gêneros de Lutjaninae e a primeira hipótese filogenética a propor um posicionamento e origem para L. alexandrei.(AU)

Frecuencia de heteroplasmia en las regiones hipervariables HVI y HVII del ADN Mitocondrial en una muestra de la población de Maracaibo, Venezuela / Frequency of heteroplasmy in HVI and HVII mitocondrial genome regions in a population sample of Maracaibo,Venezuela

Justificación:El estudio de los polimorfismos de las regiones hipervariables I y II (HVI y HVII) del ADN mitocondrial (ADNmt) se ha convertido en una herramienta invaluable para la ciencia forense, ya que enalgunas ocasionesun determinadoindividuopuedepresentar más de un tipo de ADNmitocondrial,este fenómeno es conocido como Heteroplasmia. Lacoexistencia de dos o más poblaciones de ADNmt puedeocurrir enuna sola mitocondria, célula oindividuo, lo que puede aumentar la complejidad en la interpretación de los resultados de las experticias forenses. Objetivos:Analizar la frecuencia de la heteroplasmia en las regiones HVI y HVII del genoma mitocondrialen una muestra de la población de Maracaibo. Metodología:Seseleccionaron al azar 50 muestras de ADN de la población de Maracaibo, las regiones hipervariables se amplificaron mediantereacción en cadena de la polimerasa, posteriormente se secuenciaron mediante método de Sanger y los fragmentos se separaron por electroforesis capilar, se reportaron las diferencias con respecto a la secuencia de referencia de Cambridge. Resultados: El 26% de las muestras presentaron heteroplasmia en la región HVI, el 52%en la región HVII.Conclusiones:El hecho deaparecer laheteroplas-miaen una determinadasecuencianoinválida el uso del análisis del ADN mitocondrial con fines forenses, dependiendo de la complejidad del caso a peritar,la heteroplasmia puede ser de gran ayuda...(AU)

RNA editing analysis of ATP synthase genes in the cotton cytoplasmic male sterile line H276A

BACKGROUND: Pollen development is an energy-consuming process that particularly occurs during meiosis. Low levels of adenosine triphosphate (ATP) may cause cell death, resulting in CMS (cytoplasmic male sterility). DNA sequence differences in ATP synthase genes have been revealed between the N- and S-cytoplasms in the cotton CMS system. However, very few data are available at the RNA level. In this study, we compared five ATP synthase genes in the H276A, H276B and fertile F1 (H276A/H268) lines using RNA editing, RNA blotting and quantitative real time-PCR (qRT-PCR) to explore their contribution to CMS. A molecular marker for identifying male sterile cytoplasm (MSC) was also developed. RESULTS: RNA blotting revealed the absence of any novel orf for the ATP synthase gene sequence in the three lines. Forty-one RNA editing sites were identified in the coding sequences. RNA editing showed that proteins had 32.43% higher hydrophobicity and that 39.02% of RNA editing sites had proline converted to leucine. Two new stop codons were detected in atp6 and atp9 by RNA editing. Real-time qRT-PCR data showed that the atp1, atp6, atp8, and atp9 genes had substantially lower expression levels in H276A compared with those in H276B. By contrast, the expression levels of all five genes were increased in F1 (H276A/H268). Moreover, a molecular marker based on a 6-bp deletion upstream of atp8 in H276A was developed to identify male sterile cytoplasm (MSC) in cotton. CONCLUSIONS: Our data substantially contributes to the understanding of the function of ATP synthase genes in cotton CMS. Therefore, we suggest that ATP synthase genes might be an indirect cause of cotton CMS. Further research is needed to investigate the relationship among ATP synthase genes in cotton CMS.

Lutzomyia longipalpis (Diptera: Psychodidae) Argentina-Bolivia border: new report and genetic diversity

American visceral leishmaniasis (AVL) has two main scenarios of transmission as follows: scattered cases in rural areas and urban outbreaks. Urban AVL is in active dispersion from the northeastern border of Argentina-Paraguay-Brazil to the South. The presence of Lutzomyia longipalpis was initially reported in urban environments in the northwestern border of the country. The presence of Lu. longipalpis, environmental variables associated with its distribution, and its genetic diversity were assessed in Salvador Mazza, Argentina, on the border with Bolivia. The genetic analysis showed high haplotype diversity, low nucleotide diversity, and low nucleotide polymorphism index. We discuss the hypothesis of an expanding urban population with introgressive hybridisation of older haplogroups found in their path in natural forest or rural environments, acquiring a new adaptability to urban environments, and the possibility of changes in vector capacity.

Síndrome de leigh: a propósito de um caso clínico com mutação no dna mitocondrial / Leigh syndrome: a case report with a mitochondrial dna mutation

RESUMO Objetivo: A síndrome de Leigh é uma doença neurodegenerativa com incidência de 1:40.000 nados-vivos. Apresenta ampla heterogeneidade clínica, bioquímica e genética, mas com alterações neuropatorradiológicas homogêneas. Não existe tratamento específico, e o prognóstico é reservado. O objetivo deste estudo foi familiarizar os profissionais de saúde com a doença. Descrição do caso: Menina de 16 meses, com hipotonia axial e atraso do desenvolvimento psicomotor. Dos exames realizados: cariótipo, potenciais auditivos evocados e avaliação oftalmológica normais; presença de hiperlactacidemia e hipocitrulinemia. Após a realização de ressonância magnética cerebral sob anestesia, observou-se agravamento da hipotonia com necessidade de internação por episódios de cianose/apneia. O eletroencefalograma não mostrou atividade epileptiforme. A neuroimagem revelou hipersinal lenticular bilateral com lesão do putâmen e do globo pálido esquerdo. Encontrou-se a mutação 8993T>G (MT-ATP6) no DNA mitocondrial. Comentários: De 10 a 30% dos doentes com síndrome de Leigh apresentam mutações do DNA mitocondrial. A descompensação com agravamento neurológico após intervenção anestésica está descrita e, nesse caso, apoiou o diagnóstico. Importante alertar para casos semelhantes, com diminuição de exames invasivos para diagnóstico.

ABSTRACT Objective: Leigh syndrome is a neurodegenerative disorder with an incidence of 1:40,000 live births. It presents wide clinical, biochemical, and genetic heterogeneity, but with homogenous neuropatoradiological alterations. There is no specific treatment, and the prognosis is reserved. This case report aimed familiarize health professionals with the disease. Case Description: A 16-month-hold girl who was followed in outpatient clinic due to axial hypotonia and delayed psychomotor development. Karyotype, auditory evoked potentials and ophthalmologic evaluation were normal. Evidence of hyperlactacidemia and hypocitrullinemia was detected in the patient. After performing brain magnetic resonance under anesthesia, hypotonia got worse, and the patient was hospitalized after an episode of cyanosis and apnea. The electroencephalogram showed no epileptiform activity. Neuroimaging revealed bilateral lenticular hyperintensity, especially in the putamen and in the left globus pallidus regions. Molecular analysis revealed an 8993T>G (MT-ATP6) mutation in the mitochondrial DNA. Comments: Between 10 and 30% of individuals with Leigh syndrome have mitochondrial DNA mutations. The decompensation after anesthetic intercurrences is typically associated with neurological deterioration and, in this case, increased the diagnosis suspicion. It is important to alert for similar cases and to reduce invasive diagnostic tests if the diagnosis is suspected.

Variabilidad genética de Aedes aegypti en el departamento de Sucre, Colombia, mediante el análisis de la secuencia de nucleótidos del gen mitocondrial ND4 / Genetic variability of Aedes aegypti in the department of Sucre, Colombia, by analysis of the nucleotide sequence of the mitochondrial ND4 gene

Resumen Introducción. Aedes aegypti es la especie de mosquito de mayor relevancia en América por transmitir los virus del dengue, del Zika, del chikungunya y de la fiebre amarilla. Tanto factores ecológicos como el control químico, pueden influir en la composición genética de las poblaciones de Ae. aegypti, por lo cual es necesaria su caracterización. Objetivo. Determinar la variabilidad genética de las poblaciones de Ae. aegypti en cuatro municipios del departamento de Sucre, Colombia. Materiales y métodos. Larvas de Ae. aegypti, recolectadas en los municipios de Sincelejo, Sampués, Corozal y Guaranda del departamento de Sucre, fueron criadas en laboratorio hasta el estado adulto. Como marcador genético, se utilizó un segmento del gen mitocondrial ND4, que codifica para la subunidad 4 de la enzima NADH-deshidrogenasa. El análisis genético incluyó la estimación de parámetros de diversidad de nucleótidos, haplotipos, de estructura genética y de flujo de genes. Resultados. Se obtuvieron 108 secuencias parciales de 357 nucleótidos y cuatro haplotipos de nucleótidos del gen ND4 de Ae. aegypti. Se encontró una diferenciación genética significativamente alta entre las poblaciones de Sampués y Guaranda mediante el índice de fijación (F ST =0,59467), las de Sincelejo y Sampués (F ST = 0,25637), y las de Corozal y Guaranda (F ST = 0,22237). Se evidenció un gran flujo de genes (Nm=infinito) entre las poblaciones de Sincelejo y Corozal. Conclusión. Existen diferencias genéticas entre las poblaciones del mosquito Ae. aegypti de los municipios del departamento de Sucre. Se registra la presencia de un nuevo haplotipo del gen mitocondrial ND4 de Ae. aegypti en Colombia, el cual fue detectado en el municipio de Sincelejo.

Abstract Introduction. Aedes aegypti is the most important mosquito species in America for the transmission of viruses of dengue, Zika, Chikungunya and yellow fever. Ecological factors as well as chemical controls can affect the genetic composition of Ae. aegypti populations, which is why its genetic characterization is necessary. Objective. To determine the genetic variability of Ae. aegypti populations in four municipalities of Sucre department, Colombia. Materials and methods. Larvae of Ae. aegypti, collected in the municipalities of Sincelejo, Sampués, Corozal and Guaranda, Sucre department, were reared under laboratory conditions to adult stage. A segment of the mitochondrial ND4 gene which codes for the subunit 4 of the enzyme NADH-dehydrogenase was used as genetic marker. The genetic analysis included the estimation of parameters of nucleotide and haplotype diversity, genetic structure and gene flow. Results. One hundred and eight partial sequences of 357 nucleotides and four nucleotide haplotypes of the ND4 gene of Ae. aegypti were obtained. A significantly high genetic differentiation was found between the Sampués and Guaranda populations (F ST =0.59467),Sincelejo and Sampués (F ST =0.25637), and Corozal and Guaranda (F ST =0.22237). A high gene flow (Nm=infinite) was observed among the populations of Sincelejo and Corozal. Conclusion. There are genetic differences between the Ae. aegypti populations from the municipalities of Sucre department. The presence of a new haplotype of the mitochondrial ND4 gene of Ae. aegypti in Colombia was recorded, detected in the municipality of Sincelejo.

Diversidad del ADN mitocondrial en restos óseos prehispánicos / Mitochondrial DNA diversity in prehispanic bone remains on the eastern Colombian Andes

Resumen Introducción. El ADN antiguo que se extrae de los restos óseos humanos permite analizar la composición genética de las poblaciones precolombinas y determinar las dinámicas poblacionales que dieron origen a la diversidad de las poblaciones contemporáneas. Objetivo. Determinar la diversidad genética y la relación con otras comunidades contemporáneas y antiguas de América, de los restos óseos asociados al Templo del Sol en Sogamoso, Colombia. Materiales y métodos. Se analizaron 13 individuos pertenecientes al periodo precolombino muisca (siglos IX-XVI d. C.), provenientes de los alrededores del Templo del Sol en Sogamoso, Boyacá, Andes orientales colombianos. Se amplificó el ADN mitocondrial (ADNmt) y se determinaron los polimorfismos de la longitud de los fragmentos de restricción (Restriction Fragment Length Polymorphism, RFLP) para los cuatro haplogrupos amerindios (A, B, C y D). Además, se amplificaron y analizaron los marcadores autosómicos, incluida la amelogenina, y los marcadores de los polimorfismos de repeticiones cortas en tándem (Short Tandem Repeat, STR) del cromosoma Y. Resultados. El haplogrupo A fue el linaje mitocondrial más frecuente en esta población, seguido de los haplogrupos B y C; no se detectó el haplogrupo D. Los análisis de variación genética indicaron una diversidad semejante a la de las poblaciones pertenecientes a la familia lingüística chibcha, contemporánea en Colombia y Centroamérica. Se logró hacer la determinación molecular del sexo de los individuos estudiados y compararla con los datos osteológicos. Con una sola excepción, los datos bioantropológicos y moleculares concordaron. Conclusiones. Estos resultados aportan nuevos elementos a la hipótesis del origen centroamericano de los grupos chibchas del altiplano cundiboyacense con base en marcadores genéticos, y permitieron establecer el sexo y las relaciones de parentesco.

Abstract Introduction: DNA extracted from ancient human bones allows to analyze the genetic makeup of preColumbian populations and to determine the dynamics that gave rise to the diversity of contemporary populations. Objective: To determine the genetic diversity of skeletal remains associated with the Templo del Sol (Sun Temple) and their relationship with other contemporary and ancient communities of America. Materials and methods: We analyzed 13 individuals belonging to the pre-Columbian Muisca Period (IX-XVI centuries AD) from the vicinities of the Templo del Sol (Sun Temple) (Sogamoso, Boyacá) in the eastern Colombian Andes. Mitochondrial DNA was amplified and RFLPs were performed in order to type the four traditional Amerindian haplogroups (A, B, C and D). In addition, autosomal markers including amelogenin and Y-chromosome STRs were amplified. Results: Among the observed mitochondrial lineages, haplogroup A was the most frequent, followed by haplogroups B and C; no evidence of haplogroup D was found. The genetic variation analysis indicated a similar diversity of pre-Columbian Muiscas to that of contemporary populations belonging to the Chibcha linguistic family from Colombia and Central America. Molecular sexing was accomplished and it was compared to osteological data. With only one exception, anthropological and molecular data were consistent. Conclusions: Our results contribute new genetic elements supporting the hypothesis of Central American origin of the Chibcha groups of the Cundiboyacense plateau, and allowed sex typing and kinship evaluations.