Evaluation of APOE polymorphisms and the risk for age-related macular degeneration in a Southeastern Brazilian population

This study aimed to evaluate the role of APOE polymorphisms (rs429358 and rs7412) in the risk of age-related macular degeneration in a sample of the Southeastern Brazilian population. Seven hundred and five unrelated individuals were analyzed, 334 with age-related macular degeneration (case group), and 371 without the disease (control group). In the case group, patients were further stratified according to disease phenotypes, divided into dry and wet age-related macular degeneration, and non-advanced and advanced age-related macular degeneration. APOE polymorphisms (rs429358 and rs7412) were evaluated through polymerase chain reaction and direct sequencing. In the comparison of cases vs. controls, none of the associations reached statistical significance, considering the Bonferroni-adjusted P-value, although there was a suggestive protection for the E3/E4 genotype (OR = 0.626; P-value = 0.037) and E4 carriers (OR = 0.6515; P-value = 0.047). Statistically significant protection for both the E3/E4 genotype and E4 carriers was observed in the comparisons: advanced age-related macular degeneration vs. controls (OR = 0.3665, P-value = 0.491 × 10⁻³ and OR = 0.4031, P-value = 0.814 × 10⁻³, respectively), advanced age-related macular degeneration vs. non-advanced age-related macular degeneration (OR = 0.2529, P-value = 0.659 × 10⁻⁴ and OR = 0.2692, P-value = 0.631 × 10⁻⁴, respectively). In the comparison of wet age-related macular degeneration vs. control, protection was statistically significant only for E3/E4 (OR = 0.4052, P-value = 0.001). None of the comparisons demonstrated any significant association for E2 genotypes or E2 carriers in age-related macular degeneration risk in this study. Findings suggest a protective role of the E4 haplotype in the APOE gene in the risk for advanced and wet forms of age-related macular degeneration, in a sample of the Brazilian population. To our knowledge, this is the first Brazilian study to show the association between APOE polymorphisms and age-related macular degeneration.     

Confirmation of a gene for multiple sclerosis (MS) to chromosome region 19q13.3

To identify the chromosomal localizations of the multiple sclerosis (MS) genes, we conducted a genomewide linkage analysis using eighteen affected families. A MS gene is linked to markers located in the 19q13.3 region (multipoint lod-score = 2.1). Apolipoprotein E (ApoE) gene, located in this region, is an excellent candidate gene for MS because the ApoEe4 allele is acting as a severity allele in the disease.     

Evaluation of the viability and germination of tempisque [<i>Sideroxylon capiri</i> (A.DC.) Pittier Sapotaceae]

The tempisque (Sideroxylon capiri) is a tree native to Mexico used by the rural population for housing construction, poles and hedges, as fuel (wood) and also for fodder and ornamental purposes, among others. It is considered an endangered species. In order to contribute to its preservation and sustainable management, it was considered important to determine the proportion of viable seeds, the loss of viability due to storage period and the germination process by applying pregerminative treatments. We found that freshly collected seeds showed 100% viability, which decreased to 0% after 5 months of storage. According to the cumulative germination significant differences between treatments (p≤0.01) were found. It was observed that seeds can accelerate their time of germination with the previous exposure of 24 h in water at room temperature. The soaking treatment in water for 24 h at room temperature obtained final germination of 55%, while with the control 39% was reached. Soaking in hydrogen peroxide and scarification were the treatments with lower germination percentage (33 and 23%, respectively). To get a higher percentage of germinated seeds in a short time, it is necessary to give a soaking treatment in water for 24 h before sowing.     

Compensatory substitutions and the evolution of the mitochondrial 12S rRNA gene in mammals

12S ribosomal RNA (rRNA) gene sequences from a suite of mammalian taxa (13 placentals, 4 marsupials, 1 monotreme), for which phylogenetic relationships are well established based on independent criteria, were employed to study the evolution of this gene. Phylogenetic analysis of 12S sequences produces a phylogeny that agrees with expectations. Base composition provides evidence for directional symmetrical substitution pressure in loops; in stems, base composition is much more even. Rates of nucleotide substitution are lower in stems than loops. Patterns of nucleotide substitution show an overall preference for transitions over transversions, with this difference more profound in stems than loops. Among different transversion pathways, there is a wide range of transformation frequencies. An analysis of compensatory substitutions shows that there is strong evidence for their occurrence and that a weighting factor of 0.61 should be applied in phylogenetic analyses to account for the dependence of mutations at stem positions relative to positions where changes are independent. Among stem variables (i.e., stem length, interaction distance, substitution rates, G+C content, and the percentage of bases that are paired), several significant correlations were discovered, but stem length and interaction distance are uncorrelated with other variables.      

Pregnancies following long luteal phases in southern white rhinoceros (Ceratotherium simum simum)

All extant species in the Rhinocerotidae family are experiencing escalating threats in the wild, making self-sustaining captive populations essential genetic reservoirs for species survival. Assisted reproductive technologies (ARTs) will become increasingly important for achieving and maintaining ex situ population sustainability and genetic diversity. Previous reports have shown that a large proportion of captive southern white rhinoceros (SWR) females are irregularly cyclic or acyclic, and that cycling females display two different estrous cycle lengths of approximately 30 or 70 days. It has been suggested that the longer estrous cycle length is infertile or subfertile, as no term pregnancies have been observed following long cycles. Here we report the achievement of two pregnancies following long luteal phases, using ovulation induction and artificial insemination with either fresh or frozen-thawed semen. One female SWR conceived on the first insemination attempt and gave birth to a live offspring. A second female conceived twice in consecutive long cycles although the first embryo was resorbed by 33 days post-insemination. A pregnancy from this female's second insemination is ongoing with expected parturition in November 2019. Whether prolonged estrous cycles in SWR are subfertile or infertile in natural breeding situations remains unclear. However, our findings demonstrate that the application of ARTs following prolonged cycles can result the successful establishment of pregnancies in SWR. Therefore, with ARTs, female SWR otherwise considered nonreproductive due to long estrous cycles may still have the potential for representation and contribution to the ex situ population.     

1‐Undecene from Pseudomonas aeruginosa is an olfactory signal for flight‐or‐fight response in Caenorhabditis elegans

Animals possess conserved mechanisms to detect pathogens and to improve survival in their presence by altering their own behavior and physiology. Here, we utilize Caenorhabditis elegans as a model host to ask whether bacterial volatiles constitute microbe‐associated molecular patterns. Using gas chromatography–mass spectrometry, we identify six prominent volatiles released by the bacterium Pseudomonas aeruginosa. We show that a specific volatile, 1‐undecene, activates nematode odor sensory neurons inducing both flight and fight responses in worms. Using behavioral assays, we show that worms are repelled by 1‐undecene and that this aversion response is driven by the detection of this volatile through AWB odor sensory neurons. Furthermore, we find that 1‐undecene odor can induce immune effectors specific to P. aeruginosa via AWB neurons and that brief pre‐exposure of worms to the odor enhances their survival upon subsequent bacterial infection. These results show that 1‐undecene derived from P. aeruginosa serves as a pathogen‐associated molecular pattern for the induction of protective responses in C. elegans.     

Mutagenicity of the mycotoxin diacetoxyscirpenol on somatic and germ cells of mice

Genotoxicity of diacetoxyscirpenol (DAS) was studied on laboratory mice after intraperitoneal injection with single and repeated doses. DAS was administrated at three different dose levels (0.5, 0.75, and 1.0 mg/kg body weight). The study was conducted on both somatic and germ cells additional to the sperm morphology analysis. DAS treatment resulted in a significant reduction (P<0.01) in mitotic activity at all levels of doses tested, confirming that DAS is a potent protein and DNA synthesis inhibitor. At somatic cells (bone marrow) both structural and numerical chromosome abnormalities were observed. Single dose treatment showed significant abnormalities only with high dose treatment. In contrast, at repeated dose similar abnormalities were also observed with some significance but no systematic relation between the administrated dose and abnormalities ratio could be settled. In germ cells (testicles), structural and numerical abnormalities were also observed. In general, the frequencies of scored abnormalities at germ cells were lower than that the somatic cells. Sperm count test revealed a decrease in the number of released sperm after toxin treatment. Abnormalities of sperm shape (head and tail) were observed, confirming the positive correlation between cytogenetic damage and sperm abnormality. The results also proved that DAS is a very toxic mycotoxin, in addition to inducing chromosomal abnormalities, it causes a severe inhibition of DNA synthesis which subsequently affects the cell cycle and cell division. A good system for good harvesting practice and good food technology can lower the risk for the consumers.     

Patterns of differentiation and hybridization in North American wolflike canids, revealed by analysis of microsatellite loci

Genetic divergence and gene flow among closely related populations are difficult to measure because mutation rates of most nuclear loci are so low that new mutations have not had sufficient time to appear and become fixed. Microsatellite loci are repeat arrays of simple sequences that have high mutation rates and are abundant in the eukaryotic genome. Large population samples can be screened for variation by using the polymerase chain reaction and polyacrylamide gel electrophoresis to separate alleles. We analyzed 10 microsatellite loci to quantify genetic differentiation and hybridization in three species of North American wolflike canids. We expected to find a pattern of genetic differentiation by distance to exist among wolflike canid populations, because of the finite dispersal distances of individuals. Moreover, we predicted that, because wolflike canids are highly mobile, hybrid zones may be more extensive and show substantial changes in allele frequency, relative to nonhybridizing populations. We demonstrate that wolves and coyotes do not show a pattern of genetic differentiation by distance. Genetic subdivision in coyotes, as measured by theta and Gst, is not significantly different from zero, reflecting persistent gene flow among newly established populations. However, gray wolves show significant subdivision that may be either due to drift in past Ice Age refugia populations or a result of other causes. Finally, in areas where gray wolves and coyotes hybridize, allele frequencies of gray wolves are affected, but those of coyotes are not. Past hybridization between the two species in the south-central United States may account for the origin of the red wolf.      

In silico single strand melting curve: a new approach to identify nucleic acid polymorphisms in Totiviridae

Background

The PCR technique and its variations have been increasingly used in the clinical laboratory and recent advances in this field generated new higher resolution techniques based on nucleic acid denaturation dynamics. The principle of these new molecular tools is based on the comparison of melting profiles, after denaturation of a DNA double strand. Until now, the secondary structure of single-stranded nucleic acids has not been exploited to develop identification systems based on PCR. To test the potential of single-strand RNA denaturation as a new alternative to detect specific nucleic acid variations, sequences from viruses of the Totiviridae family were compared using a new in silico melting curve approach. This family comprises double-stranded RNA virus, with a genome constituted by two ORFs, ORF1 and ORF2, which encodes the capsid/RNA binding proteins and an RNA-dependent RNA polymerase (RdRp), respectively.

Results

A phylogenetic tree based on RdRp amino acid sequences was constructed, and eight monophyletic groups were defined. Alignments of RdRp RNA sequences from each group were screened to identify RNA regions with conserved secondary structure. One region in the second half of ORF2 was identified and individually modeled using the RNAfold tool. Afterwards, each DNA or RNA sequence was denatured in silico using the softwares MELTSIM and RNAheat that generate melting curves considering the denaturation of a double stranded DNA and single stranded RNA, respectively. The same groups identified in the RdRp phylogenetic tree were retrieved by a clustering analysis of the melting curves data obtained from RNAheat. Moreover, the same approach was used to successfully discriminate different variants of Trichomonas vaginalis virus, which was not possible by the visual comparison of the double stranded melting curves generated by MELTSIM.

Conclusion

In silico analysis indicate that ssRNA melting curves are more informative than dsDNA melting curves. Furthermore, conserved RNA structures may be determined from analysis of individuals that are phylogenetically related, and these regions may be used to support the reconstitution of their phylogenetic groups. These findings are a robust basis for the development of in vitro systems to ssRNA melting curves detection.

Electronic supplementary material

The online version of this article (doi:10.1186/1471-2105-15-243) contains supplementary material, which is available to authorized users.