Initial description of the phylogeography, population structure and genetic diversity of Atlantic spotted dolphins from Brazil and the Caribbean, inferred from analyses of mitochondrial and nuclear DNA

We provide initial information regarding the population structure and genetic diversity of Stenella frontalis from the Caribbean and southeastern Brazil from analyses of mitochondrial control region sequences and sequences from the first intron of the α-lactalbumin gene. Comparisons with previously described S. frontalis sequences showed a high number of haplotypes shared between populations throughout their distribution range. High diversity was found for southeastern Brazil and Caribbean samples, and population structure analyses indicate significant differentiation among population units at the FST level, but not at the ΦST level. Significant differentiation at the FST level was found between the Caribbean population unit and all other populations units. These results suggest historical or present connectivity between the Azores and Madeira and the southeastern Brazil groups and population differentiation between the Caribbean and southeastern Brazil, supporting the notion of two separate stocks in the waters around the Atlantic coast of South America. © 2013 Elsevier Ltd.

Genetic characterisation of populations of the critically endangered Goliath grouper (Epinephelus itajara, Serranidae) from the Northern Brazilian coast through analyses of mtDNA

The Goliath grouper (Epinephelus itajara) is one of the most endangered species of fish of the subfamily Epinephelinae. Slow to develop and mature, and dependent on mangrove habitats for breeding, the species also suffers intense harvesting, which has reduced drastically in numbers in many areas. To contribute to the understanding of the characteristics of E. itajara populations, we conducted a molecular genetics study of the species, focusing on populations from the Northern Brazilian coast. The mtDNA control region (D-loop) of 116 individuals from five localities (Bragança, Ajuruteua, Parnaíba, Fortaleza and Natal) was analysed, and a sequence of 499 base pairs identified. Analyses of the sequences indicated that genetic variability was generally lower in E. itajara than in other endangered species of the genus. AMOVA found no significant grouping structure among the populations. Nested Clade Analysis revealed a significant association between genetic variability and geographic distribution among only three populations (Ajuruteua, Parnaíba and Natal). Genetic diversity was higher in populations from the Amazon region, which may be related to the better conservation of mangrove habitats in this area. Therefore, the present study could be used for the implementation of conservation and management measures in order to protect and consolidate these populations.

Cluster and meta-analyses of genetic parameters for feed intake traits in growing beef cattle

A data set based on 50 studies including feed intake and utilization traits was used to perform a meta-analysis to obtain pooled estimates using the variance between studies of genetic parameters for average daily gain (ADG); residual feed intake (RFI); metabolic body weight (MBW); feed conversion ratio (FCR); and daily dry matter intake (DMI) in beef cattle. The total data set included 128 heritability and 122 genetic correlation estimates published in the literature from 1961 to 2012. The meta-analysis was performed using a random effects model where the restricted maximum likelihood estimator was used to evaluate variances among clusters. Also, a meta-analysis using the method of cluster analysis was used to group the heritability estimates. Two clusters were obtained for each trait by different variables. It was observed, for all traits, that the heterogeneity of variance was significant between clusters and studies for genetic correlation estimates. The pooled estimates, adding the variance between clusters, for direct heritability estimates for ADG, DMI, RFI, MBW and FCR were 0.32 +/- 0.04, 0.39 +/- 0.03, 0.31 +/- 0.02, 0.31 +/- 0.03 and 0.26 +/- 0.03, respectively. Pooled genetic correlation estimates ranged from -0.15 to 0.67 among ADG, DMI, RFI, MBW and FCR. These pooled estimates of genetic parameters could be used to solve genetic prediction equations in populations where data is insufficient for variance component estimation. Cluster analysis is recommended as a statistical procedure to combine results from different studies to account for heterogeneity.

Genetic diversity of Leishmania amazonensis strains isolated in northeastern Brazil as revealed by DNA sequencing, PCR-based analyses and molecular karyotyping

Abstract Background Leishmania (Leishmania) amazonensis infection in man results in a clinical spectrum of disease manifestations ranging from cutaneous to mucosal or visceral involvement. In the present study, we have investigated the genetic variability of 18 L. amazonensis strains isolated in northeastern Brazil from patients with different clinical manifestations of leishmaniasis. Parasite DNA was analyzed by sequencing of the ITS flanking the 5.8 S subunit of the ribosomal RNA genes, by RAPD and SSR-PCR and by PFGE followed by hybridization with gene-specific probes. Results ITS sequencing and PCR-based methods revealed genetic heterogeneity among the L. amazonensis isolates examined and molecular karyotyping also showed variation in the chromosome size of different isolates. Unrooted genetic trees separated strains into different groups. Conclusion These results indicate that L. amazonensis strains isolated from leishmaniasis patients from northeastern Brazil are genetically diverse, however, no correlation between genetic polymorphism and phenotype were found.

Comparative DNA sequence and methylation analyses of orthologous genes in humans and non-human primates: Genetic and epigenetic footprints of evolution

Welche genetische Unterschiede machen uns verschieden von unseren nächsten Verwandten, den Schimpansen, und andererseits so ähnlich zu den Schimpansen? Was wir untersuchen und auch verstehen wollen, ist die komplexe Beziehung zwischen den multiplen genetischen und epigenetischen Unterschieden, deren Interaktion mit diversen Umwelt- und Kulturfaktoren in den beobachteten phänotypischen Unterschieden resultieren. Um aufzuklären, ob chromosomale Rearrangements zur Divergenz zwischen Mensch und Schimpanse beigetragen haben und welche selektiven Kräfte ihre Evolution geprägt haben, habe ich die kodierenden Sequenzen von 2 Mb umfassenden, die perizentrischen Inversionsbruchpunkte flankierenden Regionen auf den Chromosomen 1, 4, 5, 9, 12, 17 und 18 untersucht. Als Kontrolle dienten dabei 4 Mb umfassende kollineare Regionen auf den rearrangierten Chromosomen, welche mindestens 10 Mb von den Bruchpunktregionen entfernt lagen. Dabei konnte ich in den Bruchpunkten flankierenden Regionen im Vergleich zu den Kontrollregionen keine höhere Proteinevolutionsrate feststellen. Meine Ergebnisse unterstützen nicht die chromosomale Speziationshypothese für Mensch und Schimpanse, da der Anteil der positiv selektierten Gene (5,1% in den Bruchpunkten flankierenden Regionen und 7% in den Kontrollregionen) in beiden Regionen ähnlich war. Durch den Vergleich der Anzahl der positiv und negativ selektierten Gene per Chromosom konnte ich feststellen, dass Chromosom 9 die meisten und Chromosom 5 die wenigsten positiv selektierten Gene in den Bruchpunkt flankierenden Regionen und Kontrollregionen enthalten. Die Anzahl der negativ selektierten Gene (68) war dabei viel höher als die Anzahl der positiv selektierten Gene (17). Eine bioinformatische Analyse von publizierten Microarray-Expressionsdaten (Affymetrix Chip U95 und U133v2) ergab 31 Gene, die zwischen Mensch und Schimpanse differentiell exprimiert sind. Durch Untersuchung des dN/dS-Verhältnisses dieser 31 Gene konnte ich 7 Gene als negativ selektiert und nur 1 Gen als positiv selektiert identifizieren. Dieser Befund steht im Einklang mit dem Konzept, dass Genexpressionslevel unter stabilisierender Selektion evolvieren. Die meisten positiv selektierten Gene spielen überdies eine Rolle bei der Fortpflanzung. Viele dieser Speziesunterschiede resultieren eher aus Änderungen in der Genregulation als aus strukturellen Änderungen der Genprodukte. Man nimmt an, dass die meisten Unterschiede in der Genregulation sich auf transkriptioneller Ebene manifestieren. Im Rahmen dieser Arbeit wurden die Unterschiede in der DNA-Methylierung zwischen Mensch und Schimpanse untersucht. Dazu wurden die Methylierungsmuster der Promotor-CpG-Inseln von 12 Genen im Cortex von Menschen und Schimpansen mittels klassischer Bisulfit-Sequenzierung und Bisulfit-Pyrosequenzierung analysiert. Die Kandidatengene wurden wegen ihrer differentiellen Expressionsmuster zwischen Mensch und Schimpanse sowie wegen Ihrer Assoziation mit menschlichen Krankheiten oder dem genomischen Imprinting ausgewählt. Mit Ausnahme einiger individueller Positionen zeigte die Mehrzahl der analysierten Gene keine hohe intra- oder interspezifische Variation der DNA-Methylierung zwischen den beiden Spezies. Nur bei einem Gen, CCRK, waren deutliche intraspezifische und interspezifische Unterschiede im Grad der DNA-Methylierung festzustellen. Die differentiell methylierten CpG-Positionen lagen innerhalb eines repetitiven Alu-Sg1-Elements. Die Untersuchung des CCRK-Gens liefert eine umfassende Analyse der intra- und interspezifischen Variabilität der DNA-Methylierung einer Alu-Insertion in eine regulatorische Region. Die beobachteten Speziesunterschiede deuten darauf hin, dass die Methylierungsmuster des CCRK-Gens wahrscheinlich in Adaption an spezifische Anforderungen zur Feinabstimmung der CCRK-Regulation unter positiver Selektion evolvieren. Der Promotor des CCRK-Gens ist anfällig für epigenetische Modifikationen durch DNA-Methylierung, welche zu komplexen Transkriptionsmustern führen können. Durch ihre genomische Mobilität, ihren hohen CpG-Anteil und ihren Einfluss auf die Genexpression sind Alu-Insertionen exzellente Kandidaten für die Förderung von Veränderungen während der Entwicklungsregulation von Primatengenen. Der Vergleich der intra- und interspezifischen Methylierung von spezifischen Alu-Insertionen in anderen Genen und Geweben stellt eine erfolgversprechende Strategie dar.

Genetic and functional analyses of cell division proteins FtsZ and FtsA in Escherichia coli

In the current model for bacterial cell division, the FtsZ protein forms a ring that marks the division plane, creating a cytoskeletal framework for the subsequent action of other essential division proteins such as FtsA and ZipA. The putative protein complex ultimately generates the division septum. The essential cell division protein FtsZ is a functional and structural homolog of eukaryotic tubulin, and like tubulin, FtsZ hydrolyzes GTP and self-assembles into protein filaments in a strictly GTP-dependent manner. FtsA shares sequence similarity with members of the ATPase superfamily that include actin, but its actual function remains unknown. To test the division model and elucidate functions of the division proteins, this dissertation primarily focuses on the analysis of FtsZ and FtsA in Escherichia coli. ^ By tagging with green fluorescent protein, we first demonstrated that FtsA also exhibits a ring-like structure at the potential division site. The localization of FtsA was dependent on functional FtsZ, suggesting that FtsA is recruited to the septum by the FtsZ ring. In support of this idea, we showed that FtsA and FtsZ directly interact. Using a novel E. coli in situ assay, we found that the FtsA-FtsZ interaction appears to be species-specific, although an interspecies interaction could occur between FtsA and FtsZ proteins from two closely related organisms. In addition, mutagenesis of FtsA revealed that no single domain is solely responsible for its septal localization or interaction with FtsZ. To explore the function of FtsA, we purified FtsA protein and demonstrated that it has ATPase activity. Furthermore, purified FtsA stimulates disassembly of FtsZ polymers in a sedimentation assay but does not affect GTP hydrolysis of FtsZ. This result suggests that in the cell, FtsA may function similarly in regulating dynamic instability of the FtsZ ring during the cell division process. ^ To elucidate the structure-function relationship of FtsZ, we carried out thorough genetic and functional analyses of the mutagenized FtsZ derivatives. Our results indicate that the conserved N-terminal domain of FtsZ is necessary and sufficient for FtsZ self-assembly and localization. Moreover, we discovered a critical role for an extreme C-terminal domain of FtsZ that consists of only 12 residues. Truncated FtsZ derivatives lacking this domain, though able to polymerize and localize, are defective in ring formation in vivo as well as interaction with FtsA and ZipA. Alanine scanning mutagenesis of this region pinpointed at least five residues necessary for the function of FtsZ. Studies of protein levels and protein-protein interactions suggested that these residues may be involved in regulating protein stability and/or FtsZ-FtsA interactions. Interestingly, two of the point mutants exhibited dominant-negative phenotypes. ^ In summary, results from this thesis work have provided additional support for the division machinery model and will contribute to a better understanding of the coordinate functions of FtsA and FtsZ in the cell division process. ^

Genetic susceptibility, cigarette smoking, and wood dust exposure in lung cancer: Case control analyses

This case control study was conducted to assess the association between lung cancer risk, mutagen sensitivity (a marker of cancer susceptibility), and a putative lung carcinogen, wood dust exposure. There were 165 cases (98 African-Americans, 67 Mexican-Americans) with newly diagnosed, previously untreated lung cancer, and 239 controls, frequency-matched on age, sex, and ethnicity.^ Mutagen sensitivity ($\ge$1 break/cell) was associated with a statistically significant elevated risk for lung cancer (odds ratio (OR) = 4.1, 95% confidence limits (CL) = 2.3,7.2). Wood dust exposure was also a significant predictor of risk (OR = 2.8, 95% CL = 1.2,6.6) after controlling for smoking and mutagen sensitivity. When stratified by ethnicity, wood dust exposure was a significant risk factor for African-Americans (OR = 4.0, 95% CL = 1.4,11.5), but not for Mexican-Americans (OR = 1.5, 95% CL = 0.3,7.1). Stratified analysis suggested a greater than multiplicative interaction between wood dust exposure and both mutagen sensitivity and smoking.^ The cases had significantly more breaks on chromosomes 4 and 5 than the controls did with ORs of 4.9 (95% CL = 2.0, 11.7) and 3.9 (95% CL = 1.6, 9.3), respectively. Breaks at 4p14, 4q27, 4q31, 5q21-22, 5q31, and 5q33 were significantly more common in lung cancer patients than in controls. Lung cancer risk had a dose-response relationship with breaks on chromosomes 4 and 5. Cigarette smoking had a strong interaction with breaks on chromosomes 2, 4, and 5.^ In a molecular cytogenetic study, using chromosome painting and G-banding, we showed that: (1) the proportion of chromosome 5 abnormalities surviving as chromosome-type aberrations remained significantly higher in cells of lung cancer cases (14%) than in controls (5%) (P $<$ 0.001). However, no significant differences were detected in chromosome 4 abnormalities between cases and controls; (2) the proportion of chromosome 5q13-22 abnormalities was 5.3% in the cases and 0.7% in the controls (P $<$ 0.001). 5q13-22 regions represented 40% of all abnormalities on chromosome 5 in the cases and only 14% in the controls.^ This study suggests that mutagen sensitivity, wood dust exposure, and cigarette smoking were independent risk factors for lung cancer, and the susceptibility of particular chromosome loci to mutagenic damage may be a genetic marker for specific types of lung cancer. ^

A genetic algorithm for slope stability analyses with concave slip surfaces using custom operators

Heuristic methods are popular tools to find critical slip surfaces in slope stability analyses. A new genetic algorithm (GA) is proposed in this work that has a standard structure but a novel encoding and generation of individuals with custom-designed operators for mutation and crossover that produce kinematically feasible slip surfaces with a high probability. In addition, new indices to assess the efficiency of operators in their search for the minimum factor of safety (FS) are proposed. The proposed GA is applied to traditional benchmark examples from the literature, as well as to a new practical example. Results show that the proposed GA is reliable, flexible and robust: it provides good minimum FS estimates that are not very sensitive to the number of nodes and that are very similar for different replications

Genetic and comparative analyses reveal an alternative secondary structure in the region of nt 912 of Escherichia coli 16S rRNA.

Mutations at position 912 of Escherichia coli 16S rRNA result in two notable phenotypes. The C-->U transition confers resistance to streptomycin, a translational-error-inducing antibiotic, while a C-->G transversion causes marked retardation of cell growth rate. Starting with the slow-growing G912 mutant, random mutagenesis was used to isolate a second site mutation that restored growth nearly to the wild-type rate. The second site mutation was identified as a G-->C transversion at position 885 in 16S rRNA. Cells containing the G912 mutation had an increased doubling time, abnormal sucrose gradient ribosome/subunit profile, increased sensitivity to spectinomycin, dependence upon streptomycin for growth in the presence of spectinomycin, and slower translation rate, whereas cells with the G912/C885 double mutation were similar to wild type in these assays. Comparative analysis showed there was significant covariation between positions 912 and 885. Thus the second-site suppressor analysis, the functional assays, and the comparative data suggest that the interaction between nt 912 and nt 885 is conserved and necessary for normal ribosome function. Furthermore, the comparative data suggest that the interaction extends to include G885-G886-G887 pairing with C912-U911-C910. An alternative secondary structure element for the central domain of 16S rRNA is proposed.

Fine-scale spatial genetic correlation analyses reveal strong female philopatry within a brush-tailed rock-wallaby colony in southeast Queensland

We combine spatial data on home ranges of individuals and microsatellite markers to examine patterns of fine-scale spatial genetic structure and dispersal within a brush-tailed rock-wallaby (Petrogale penicillata) colony at Hurdle Creek Valley, Queensland. Brush-tailed rock-wallabies were once abundant and widespread throughout the rocky terrain of southeastern Australia; however, populations are nearly extinct in the south of their range and in decline elsewhere. We use pairwise relatedness measures and a recent multilocus spatial autocorrelation analysis to test the hypotheses that in this species, within-colony dispersal is male-biased and that female philopatry results in spatial clusters of related females within the colony. We provide clear evidence for strong female philopatry and male-biased dispersal within this rock-wallaby colony. There was a strong, significant negative correlation between pairwise relatedness and geographical distance of individual females along only 800 m of cliff line. Spatial genetic autocorrelation analyses showed significant positive correlation for females in close proximity to each other and revealed a genetic neighbourhood size of only 600 m for females. Our study is the first to report on the fine-scale spatial genetic structure within a rock-wallaby colony and we provide the first robust evidence for strong female philopatry and spatial clustering of related females within this taxon. We discuss the ecological and conservation implications of our findings for rock-wallabies, as well as the importance of fine-scale spatial genetic patterns in studies of dispersal behaviour.

Why are there so few genetic markers available for coral population analyses?

Coral reefs are in serious decline, and research in support of reef management objectives is urgently needed. Reef connectivity analyses have been highlighted as one of the major future research avenues necessary for implementing effective management initiatives for coral reefs. Despite the number of new molecular genetic tools and the wealth of information that is now available for population-level processes in many marine disciplines, scleractinian coral population genetic information remains surprisingly limited. Here we examine the technical problems and approaches used, address the reasons contributing to this delay in understanding, and discuss the future of coral population marker development. Considerable resources are needed to target the immediate development of an array of relevant genetic markers coupled with the rapid production of management focused data in order to help conserve our globally threatened coral reef resources.

Development of microsatellite markers in Cannabis Sativa for fingerprinting and genetic relatedness analyses

Microsatellite markers were developed for Cannabis sativa L. (marijuana) to estimate the level of polymorphism, usefulness for DNA typing (genotype identification), and to measure the genetic relationships between the different plants. Twelve different oligonucleotide probes were used to screen an enriched microsatellite library of Cannabis sativa in which 49% of the clones contained microsatellite sequences. Characterization of microsatellite loci in Cannabis revealed that GA/CT was the most abundant class of isolated microsatellites representing 50% overall. Eleven polymorphic SSR markers were developed, derived from dinucleotide motifs and eight from trinucleotide motifs. A total of 52 alleles were detected averaging 4.7 alleles/locus. The expected heterozygosity of the eleven loci ranged between 0.368 and 0.710 and the common probability of identical genotypes was 1.8 x 107. The loci identified 27 unique profiles of the 41 Cannabis samples. The eleven microsatellite markers developed in this study were found to be useful for DNA fingerprinting and for assessing genetic relationships in Cannabis.

Molecular Genetic Variability Of Commercial And Wild Accessions Of Passion Fruit (passiflora Spp.) Targeting Ex Situ Conservation And Breeding.

Passiflora species are distributed throughout Latin America, and Brazil and Colombia serve as the centers of diversity for this genus. We performed cross-species amplification to evaluate 109 microsatellite loci in 14 Passiflora species and estimated the diversity and genetic structure of Passiflora cincinnata, Passiflora setaceae and Passiflora edulis. A total of 127 accessions, including 85 accessions of P. edulis, a commercial species, and 42 accessions of 13 wild species, were examined. The cross-species amplification was effective for obtaining microsatellite loci (average cross-amplification of 70%). The average number of alleles per locus (five) was relatively low, and the average diversity ranged from 0.52 in P. cincinnata to 0.32 in P. setacea. The Bayesian analyses indicated that the P. cincinnata and P. setacea accessions were distributed into two groups, and the P. edulis accessions were distributed into five groups. Private alleles were identified, and suggestions for core collections are presented. Further collections are necessary, and the information generated may be useful for breeding and conservation.

Comparative Study Of Transgenic And Non-transgenic Maize (zea Mays) Flours Commercialized In Brazil, Focussing On Proteomic Analyses.

Genetically modified foods are a major concern around the world due to the lack of information concerning their safety and health effects. This work evaluates differences, at the proteomic level, between two types of crop samples: transgenic (MON810 event with the Cry1Ab gene, which confers resistance to insects) and non-transgenic maize flour commercialized in Brazil. The 2-D DIGE technique revealed 99 differentially expressed spots, which were collected in 2-D PAGE gels and identified via mass spectrometry (nESI-QTOF MS/MS). The abundance of protein differences between the transgenic and non-transgenic samples could arise from genetic modification or as a result of an environmental influence pertaining to the commercial sample. The major functional category of proteins identified was related to disease/defense and, although differences were observed between samples, no toxins or allergenic proteins were found.