Recent Y chromosome divergence despite ancient origin of dioecy in poplars (Populus).

All species of the genus Populus (poplar, aspen) are dioecious, suggesting an ancient origin of this trait. Despite some empirical counter examples, theory suggests that nonrecombining sex-linked regions should quickly spread, eventually becoming heteromorphic chromosomes. In contrast, we show using whole-genome scans that the sex-associated region in Populus trichocarpa is small and much younger than the age of the genus. This indicates that sex determination is highly labile in poplar, consistent with recent evidence of 'turnover' of sex-determination regions in animals. We performed whole-genome resequencing of 52 P. trichocarpa (black cottonwood) and 34 Populus balsamifera (balsam poplar) individuals of known sex. Genomewide association studies in these unstructured populations identified 650 SNPs significantly associated with sex. We estimate the size of the sex-linked region to be ~100 kbp. All SNPs significantly associated with sex were in strong linkage disequilibrium despite the fact that they were mapped to six different chromosomes (plus 3 unmapped scaffolds) in version 2.2 of the reference genome. We show that this is likely due to genome misassembly. The segregation pattern of sex-associated SNPs revealed this to be an XY sex-determining system. Estimated divergence times of X and Y haplotype sequences (6-7 Ma) are much more recent than the divergence of P. trichocarpa (poplar) and Populus tremuloides (aspen). Consistent with this, in P. tremuloides, we found no XY haplotype divergence within the P. trichocarpa sex-determining region. These two species therefore have a different genomic architecture of sex, suggestive of at least one turnover event in the recent past.

A 34K SNP genotyping array for Populus trichocarpa: design, application to the study of natural populations and transferability to other Populus species.

Genetic mapping of quantitative traits requires genotypic data for large numbers of markers in many individuals. For such studies, the use of large single nucleotide polymorphism (SNP) genotyping arrays still offers the most cost-effective solution. Herein we report on the design and performance of a SNP genotyping array for Populus trichocarpa (black cottonwood). This genotyping array was designed with SNPs pre-ascertained in 34 wild accessions covering most of the species latitudinal range. We adopted a candidate gene approach to the array design that resulted in the selection of 34 131 SNPs, the majority of which are located in, or within 2 kb of, 3543 candidate genes. A subset of the SNPs on the array (539) was selected based on patterns of variation among the SNP discovery accessions. We show that more than 95% of the loci produce high quality genotypes and that the genotyping error rate for these is likely below 2%. We demonstrate that even among small numbers of samples (n = 10) from local populations over 84% of loci are polymorphic. We also tested the applicability of the array to other species in the genus and found that the number of polymorphic loci decreases rapidly with genetic distance, with the largest numbers detected in other species in section Tacamahaca. Finally, we provide evidence for the utility of the array to address evolutionary questions such as intraspecific studies of genetic differentiation, species assignment and the detection of natural hybrids.

Higher differentiation among subspecies of the house mouse (Mus musculus) in genomic regions with low recombination.

In the early stages of reproductive isolation, genomic regions of reduced recombination are expected to show greater levels of differentiation, either because gene flow between species is reduced in these regions or because the effects of selection at linked sites within species are enhanced in these regions. Here, we study the patterns of DNA sequence variation at 27 autosomal loci among populations of Mus musculus musculus, M. m. domesticus, and M. m. castaneus, three subspecies of house mice with collinear genomes. We found that some loci exhibit considerable shared variation among subspecies, while others exhibit fixed differences. We used an isolation-with-gene-flow model to estimate divergence times and effective population sizes (N(e) ) and to disentangle ancestral variation from gene flow. Estimates of divergence time indicate that all three subspecies diverged from one another within a very short period of time approximately 350,000 years ago. Overall, N(e) for each subspecies was associated with the degree of genetic differentiation: M. m. musculus had the smallest N(e) and the greatest proportion of monophyletic gene genealogies, while M. m. castaneus had the largest N(e) and the smallest proportion of monophyletic gene genealogies. M. m. domesticus and M. m. musculus were more differentiated from each other than either were from M. m. castaneus, consistent with greater reproductive isolation between M. m. domesticus and M. m. musculus. F(ST) was significantly greater at loci experiencing low recombination rates compared to loci experiencing high recombination rates in comparisons between M. m. castaneus and M. m. musculus or M. m. domesticus. These results provide evidence that genomic regions with less recombination show greater differentiation, even in the absence of chromosomal rearrangements.

Rash, fever and proteinuria after amoxicillin in a SLE patient.

We report a case of severe type IV hypersensitivity reaction to amoxicillin, which occurred in a person with a 12-year history of SLE. The present case illustrates the wide differential diagnosis in a SLE patient who presents with an allergic drug reaction. The attribution of the presenting symptoms to the underlying SLE and/or to the drugs used to treat SLE and coexisting conditions is a major challenge.

Reduced introgression of the Y chromosome between subspecies of the European rabbit (Oryctolagus cuniculus) in the Iberian Peninsula.

The role of the Y chromosome in speciation is unclear. Hybrid zones provide natural arenas for studying speciation, as differential introgression of markers may reveal selection acting against incompatibilities. Two subspecies of the European rabbit (Oryctolagus cuniculus) form a hybrid zone in the Iberian Peninsula. Previous work on mitochondrial DNA (mtDNA), Y- and X-linked loci revealed the existence of two divergent lineages in the rabbit genome and that these lineages are largely subspecies-specific for mtDNA and two X-linked loci. Here we investigated the geographic distribution of the two Y chromosome lineages by genotyping two diagnostic single nucleotide polymorphisms in a sample of 353 male rabbits representing both subspecies, and found that Y chromosome lineages are also largely subspecies-specific. We then sequenced three autosomal loci and discovered considerable variation in levels of differentiation at these loci. Finally, we compared estimates of population differentiation between rabbit subspecies at 26 markers and found a surprising bimodal distribution of F(ST)values. The vast majority of loci showed little or no differentiation between rabbit subspecies while a few loci, including the SRY gene, showed little or no introgression across the hybrid zone. Estimates of population differentiation for the Y chromosome were surprisingly high given that there is male-biased dispersal in rabbits. Taken together, these data indicate that there is a clear dichotomy in the rabbit genome and that some loci remain highly differentiated despite extensive gene flow following secondary contact.

Correlação entre flexibilidade das articulações glenoumerais e coxofemorais e o desempenho funcional de idosas fisicamente ativas / Association between flexibility of the glenohumeral and hip joints and functional performance in active elderly women

OBJETIVO: Investigar a relação entre a flexibilidade da flexão e extensão das articulações glenoumerais (GU) e coxofemorais (CF) e o desempenho funcional (DF) de idosas funcionalmente independentes e fisicamente ativas. MÉTODOS: Determinou-se em 22 voluntárias (idade=70±6 anos) seis conjuntos de amplitudes de movimentos por goniometria ativo-assistida (ADM) na flexão e extensão das GU e CF. O DF foi determinado pelos testes: velocidade de caminhada habitual (VCH) e máxima (VCM); levantar e sentar em cadeira (LSC); Timed up and Go Test (TUGT); vestir blusa (VBL); subir degraus (SE); levantar do decúbito dorsal (LDD); pegar moeda no solo (PMS); teste de caminhada de seis minutos (TC6M). As associações entre as variáveis ADM e o DF foram testadas por técnicas de correlação simples e múltipla. RESULTADOS: Houve correlações significantes (p<0,05) entre as ADM de CF e os testes LSC (r=0,42 e r=0,45), SE (r=0,52 e r=0,53) e TC6M (r=0,58 e r=0,59) (lados direito e esquerdo, respectivamente). A correlação múltipla ratificou esses resultados (r²=0,51; p<0,05), indicando que 51 por cento da variância nos testes deveu-se à ADM de CF. Não houve associações significantes entre as ADMs de GU e os testes de DF. CONCLUSÕES: Verificou-se associação significante entre a flexibilidade ativo-assistida de CF e alguns testes específicos de DF. Nenhuma relação foi identificada para ADM de GU. Estudos adicionais são necessários para elucidar as relações entre flexibilidade passiva de diferentes grupos articulares e a funcionalidade de idosos.

OBJECTIVE: To investigate the relationship between flexibility of flexion and extension of the glenohumeral and coxofemoral joints and functional performance among physically active and functionally independent elderly women. METHODS: Six sets of range of motion (ROM) measurements relating to flexion and extension of the glenohumeral and coxofemoral joints were determined in 22 volunteers (age 70±6 years), using assisted-active goniometry. Functional performance was measured using the following tests: normal walking speed (NWS); maximum walking speed (MWS); sit-to-stand test (SST); timed up and go test (TUGT); putting on a blouse (PBL); going up stairs (GUS); rising from dorsal decubitus (RDD); picking up a coin from the floor (PCF); and 6-minute walk test (6WT). The relationships between the ROM variables and functional performance were tested using simple and multiple regression techniques. RESULTS: There were significant correlations (p<0.05) between coxofemoral ROM and the SST (r=0.42 and r=0.45), GUS (r=0.52 and r=0.53) and 6WT (r=0.58 and r=0.59) (right and left sides, respectively). The multiple regression ratified the results (r²=0.51; p<0.05), thus indicating that coxofemoral ROM accounted for 51 percent of the variance in the tests. There were no significant correlations between the glenohumeral ROMs and the functional performance tests. CONCLUSIONS: There was a significant association between assisted-active flexibility of the coxofemoral joint and some specific functional performance tests. No relationship involving glenohumeral ROM was identified. Additional studies are needed in order to elucidate the relationships between passive flexibility of different joint groups and functional performance in elderly people.

Efeitos de um programa de treinamento resistido com volume e intensidade moderados e velocidade elevada sobre o desempenho funcional de mulheres idosas / Effects of a programme of resistance training with volume and intensity moderates and high speed on functional performance of old women

A tensão muscular pode ser influenciada pela velocidade de movimento, sendo um componente importante da prescrição do treinamento da força em idosos. Esse estudo observou os efeitos de 12 semanas de treinamento resistido (TR), realizado com intensidade e volume moderados e velocidade elevada, porém não explosiva, sobre a força/potência muscular (FM) e o desempenho funcional (DF). Para tal, 24 idosas (68,7±9 anos) fisicamente ativas, foram igualmente distribuídas em grupos controle (GCO) e experimental (GEX). O GEX realizou, 2 X por semana durante 12 semanas, de série única de 10 a 15 repetições de exercícios com intensidade relativa de 50 a 70% de 1RM e fase concêntrica das contrações fixada em menos de 1 seg. O grupo controle não praticou exercícios de força. A FM foi aferida pelo teste de 1RM nos exercícios leg press horizontal (LPH) e flexão dos joelhos na cadeira (CF). Mediu-se o DF através do tempo para realizar as seguintes tarefas: a) caminhada de 10 metros (C10); b) levantar da posição ajoelhada (LPA); c) levantar e sentar de uma cadeira (LSC). Não havia diferenças entre os grupos na linha de base. Ao fim da intervenção, a ANOVA de duas entradas para medidas repetidas seguida da verificação post-hoc de Fisher (p<0,05), demonstrou não haver diferenças significativas para GCO. No GEX foram observados incrementos significativos em todos as medidas de FM (LPH, p<0,0001; CF, p<0,0001) e DF (CAM10, p<0,0001; LSC, p=0,004; LPA, p=0,005). Conclui-se que idosas fisicamente ativas podem ter melhoria significativa de FM e DF em decorrência de TR realizado com intensidade e volume moderados e velocidade elevada.

It has been suggested that lower extremity muscle power and strength are important for physical function in older adults. However, the majority of studies investigating the effects of training on muscle power are based on isokinetic exercises and few studies have determined the effects of isoinertial resistance training (RT) on muscle power and strength and functional performance in older adults. The purpose of the present study was to evaluate the effects of a 12 weeks RT in muscle strength and functional performance in healthy, active, older women. Twentyfour independent and active older women (aged 60-85 years) were assigned either to a control group (CG, n=12), or to a RT group (RTG, n=12). The RTG trained 10 exercises for whole body with one set of 10-15 repetitions at 50 to 70% of 1-repetition maximum (1-RM) released with high-velocity (concentric phase ≤1sec), 2 days per week for 12 weeks. The CG did not perform strength exercises. Both groups were evaluated in 1-RM lower body strength in horizontal leg press (HLP) and knee flexion (KF), maximal walking speed (MWS); kneel rise time (KRT); fivetimes-sit-to-stand test (FTSST) before and after the training period. There was no significant difference between groups in base-line (p<.05). After the 12 weeks, the 2-way ANOVA showed inter and intragroups significant differences (p<.05). RTG significantly increased the results for all 1RM tests: HLP (p<.0001); KF (p<.0001) and physical performance: MWS (p<.0001); KRT (p=.005); FTSST (p=.004). In conclusion, strength training of moderate volume/intensity and high-velocity can promote gains in tasks related with lower extremity muscle power in elder women.

High levels of nucleotide diversity in the European rabbit (Oryctolagus cuniculus) SRY gene.

We have sequenced 2,388 bp of the European rabbit sex determining region Y (SRY) gene. These data provide a 10-fold increase in the coverage of the Y chromosome in this species, including the entire open reading frame of the SRY, the polyadenylation signal, and two repetitive sequences in the 5' -region. A survey of 2021 bp of this gene in eight domestic breeds and four wild individuals revealed a total of nine single nucleotide polymorphisms and one indel, defining two deeply divergent lineages. The resulting estimation of nucleotide diversity (pi=1.34 x10(-3)) is very high when compared with other species, but no variability was detected among the domestic breeds. This study represents a first step in the characterization of the European rabbit Y chromosome and its variability. These sequences can be used in additional phylogeographical analyses of the European rabbit and other Leporid species, as well as in evolutionary studies of sex determination and the Y chromosome in wild species.

Entry of African swine fever virus into Vero cells and uncoating.

African swine fever virus (ASFV) enters Vero cells by adsorptive endocytosis [Valdeira, M.L., Geraldes, A., 1985. Morphological study on the entry of African swine fever virus into cells, Biol Cell. 55, 35-40]. Electron microscopy of a lysosomotropic drug-controlled penetration indicated that this step takes place in the endosomes, after fusion between the viral envelope and the limiting membrane of the endosome. Inhibition studies with colcemid, cytochalasin B, sodium azide, dinitrophenol, lysosomotropic weak bases, and the ionophore monensin, showed that the virus uptake is largely independent of cytoskeletal and lysosomal function, but dependent on oxidative phosphorylation. Some protease inhibitors inhibited viral replication at an early step, indicating that the initiation of infection depends on a viral proteolytic cleavage.

Evidence for an acid phosphatase in African swine fever virus.

An acid phosphatase activity has been detected in purified preparations of African swine fever virus. Purified viral cores obtained after Nonidet-P40 and 2-mercaptoethanol treatment of the virus retained the activity as assayed with nitrophenyl phosphate as substrate at pH 5. Enzyme cytochemistry by electron microscopy showed that the acid phosphatase activity is localized mainly inside the core of the virion. The molecular weight and the isoelectric point of the virus acid phosphatase activity confirmed that it was distinct from the host cellular enzyme.

Effect of chloroquine on African swine fever virus infection.

When present during the whole infective cycle, the lysosomotropic drug, chloroquine, inhibited cytopathic changes and production of African swine fever virus (ASFV) in Vero cells. This inhibition decreased when the drug was added from 1 h to 4 h after infection. Chloroquine had no effect on the virus nor on viral adsorption and internalization. Electron microscopy showed that, in the presence of the drug, the virions were retained in large vacuoles having a lysosomal appearance. This inhibition was fully reversible, even when the drug was removed as late as 72 h after infection. The results support the hypothesis that ASFV enters the cells by adsorptive endocytosis and not by fusion with the plasma membrane.

Morphological study on the entry of African swine fever virus into cells.

The early interactions between African swine fever virus (ASFV) and monkey kidney cells in culture, and the effect of chloroquine were studied by electron microscopy. Our results indicate that ASFV uptake occurs by endocytosis: after attachment to the cell surface, the virions were seen in coated pits and were internalized by endocytosis in endosomes and finally in lysosomes. Virions in coated vesicles were never seen. All these steps were completed in about 15 min. Direct penetration of viruses through the plasma membrane was never observed. In order to elucidate the participation of an acidic intracellular compartment in the penetration of ASFV, we studied the effect of chloroquine, a lysosomotropic drug known to increase the pH of acidic intracellular vacuoles and to inhibit ASFV infection. In the presence of this drug there were no apparent alterations on binding, endocytosis and intracellular distribution of the viral particles. The main effect of chloroquine was to retain the virions in lysosomes. When the drug was removed from the medium, the viral particles disappeared and images of binding of viral membranes with the membranes of the intracellular vacuoles were obtained, suggesting that the inhibited step is the uncoating of the virus. Viral fusion with the plasma membrane was obtained when the medium was acidified to pH 5-6. These results suggest that ASFV enters the cells by adsorptive endocytosis and that the uncoating process takes place intracellularly in a way similar to that described for Semliki Forest virus and other enveloped viruses.