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Table of contents: 2019

Research Article

Passiflora cristalina, Passiflora miniata and Passiflora coccinea are wild species with similar floral characteristics, especially color and floral structure, as well as the color of their fruits. Due to their similarities, mainly the floral characteristics, these species are often confused in the field. Given that hybridization is commonplace between Passiflora species in the same region, hybrids could result from crosses involving mainly P. coccinea. We examined genetic distance and possible hybrid nature across P. cristalina, P. miniata, and P. coccinea, via ISSR and SSR markers. Genomic DNA was extracted from leaf samples of five Passiflora species (P. cristalina, P. coccinea, P. miniata, P. setacea, and P. edulis), the latter two being used as witness species. Following quantification, the amplification conditions were tested and optimized. Eighteen ISSR primers presented satisfactory amplification products, with 81 bands being amplified and 99% polymorphism. Through genetic distance and cluster analysis, P. cristalina and P. coccinea were found to be genetically close, while P. miniata remained in an isolated cluster, nevertheless with low dissimilarity with P. cristalina. Twenty-three SSR primers were tested, of which 18 were polymorphic. There was a high transferability rate, 95.65%, demonstrating that genetic proximity between tax is directly related to successful transferability. The main coordinates, genetic distance and cluster analyses showed a clear separation of species presenting similar floral characteristics (P. cristalina, P. coccinea, and P. miniata) from the remaining ones used as controls (P. edulis and P. setacea). SSR markers identified P. cristalina as a possible natural hybrid between P. miniata and P. coccinea.

Genet. Mol. Res. 18(1): GMR18040
DOI: 10.4238/gmr18040
Research Article

Worldwide, several women become victims of rape every day. Many of those women are also murdered, with their bodies sometimes being found in an advanced state of decomposition, resulting in loss of evidence important to criminal investigations. Diptera is one of the main orders associated with human body decomposition. Fly species that belong to the family Calliphoridae are usually scavengers and are frequently found on decomposing bodies, thereby playing an important role in forensic entomology. The recovery and genotyping of human Y-STR DNA from the gastrointestinal contents of the calliphorid Chrysomya albiceps larvae has promising applications in the investigation of sexual crimes, such as rape, and in cases of murder and abandonment of the victim’s body, which may be found in a state of decomposition. We studied this species of fly with the aim of supporting such investigations. After establishment of a colony, larvae were fed with decomposing human semen mixed in ground bovine meat (1 mL per 200 g beef). Larvae (10–15) were collected every 24 h and kept in 70% ethanol, to give a total of 96 larvae obtained after eight days of decomposition. The digestive system of each larva was resected. Molecular typing was conducted, which comprised sample extraction, quantification, amplification, and capillary electrophoresis with 16 STR loci from the Y chromosome. We succeeded in establishing a Y-STR DNA profile, with amplification of up to 11 loci, from individual samples, or up to 15 loci, when a combination of samples corresponding to the time-points 48, 72, 120, 144, and 192 h was used.

 

Genet. Mol. Res. 18(1): GMR18189
DOI: 10.4238/gmr18189
Research Article

The mangaba, Hancornia speciosa, (Apocynaceae) is a fruit tree native to Brazil with predominantly extractivist production. The fruit can be consumed in natura; however, it is widely consumed as frozen pulp and ice cream produced by agro-industry companies. We evaluated the genetic diversity of 213 individuals that make up the mangaba GeneBank of Embrapa Tabuleiros Costeiros, using nine microsatellite markers (SSR). A total of 147 alleles were identified, with a mean of 16 alleles per locus; 100% polymorphism was observed among accessions. Reliability of the result was verified based on stress (0.042) and correlation (0.988) values. The alleles presented a high frequency of heterozygosity (He > Ho). The Fst (0.22) and f (0.07) values indicated moderate population structure, with great diversity within accessions. Bayesian analysis indicated the most adequate grouping with k = 2. The Unweighted Pair Group Method analysis showed three distinct groups according to similarity. The BI accession had the best genetic structure. The PM5/GX2, CN1/CN9, G18/PA1, JA14/JA15, and OI8/OI9 pairs of individuals are the closest genetically. We conclude that the Mangaba GeneBank has high diversity; this knowledge is relevant to develop strategies for the management of these genetic resources.

 

Genet. Mol. Res. 18(1): GMR18108
DOI: 10.4238/gmr18108
Research Article

Pepper has considerable genetic diversity and versatility. Knowledge of the genetic control of traits in peppers is of great importance for breeding programs given the large variety of types, sizes, colors and flavors. To this end, we examined the inheritance of seedling and plant traits in ornamental pepper (Capsicum annuum). The experiment was conducted in a greenhouse in Areia, Paraíba, Brazil. Seven ornamental pepper accessions (C. annuum) belonging to the Federal University of Paraiba’s Germplasm Bank were investigated: UFPB001, UFPB004, UFPB77.3, UFPB099, UFPB134, UFPB137 and UFPB390. Morphoagronomic characterization was performed based on Capsicum descriptors, and 12 quantitative traits were evaluated in seedlings and plants. The data were subjected to variance analysis and subsequent diallel analysis performed according to Hayman's method. The t statistic was used to test the adequacy of the additive-dominance model. The traits seedling height, hypocotyl diameter, cotyledon leaf length, plant height, bifurcation height, leaf length and width and chlorophyll a and b are in agreement with the additive-dominant model. Correlations were positive and significant for seedling height (0.470) and hypocotyl diameter (0.885). Cotyledonary leaf length and width showed negative and significant values of -0.088 and -0.669, respectively. The correlations were positive for the following traits: plant height, stem diameter, first bifurcation height, canopy diameter, leaf length and chlorophyll b, with values ranging from 0.094 to 0.965. Leaf width and chlorophyll a exhibited negative r correlation values. In the genetic parameters estimate, the positive r correlation for most of the traits indicates that the recessive alleles were generally responsible for the increase in these traits. Genetic gains for plant traits in ornamental peppers are possible using breeding programs. The parents UFPB001 and UFPB134 exhibited the highest concentration of favorable alleles for size traits and are indicated for selection for continued improvement programs.

Genet. Mol. Res. 18(1):
DOI: 10.4238/gmr18120
Research Article

Many studies have explored variability to select cattle with high genetic potential for economic interest traits. Genetic variability is a powerful tool to improve production indexes in cattle, as it also is associated with variations in meat and carcass quality traits. We made a Genome-Wide Association Study of beef cattle of Bos indicus origin, in particular Nelore animals, to identify regions and genes associated with carcass quality, by examining phenotypic and genotypic data from 909 animals. Several genes in associated regions were observed to have above 1% of the portion of explained genetic variance explained: for hot carcass weight, genes LRGUK, TRIM24, SVOPL, TEX37, CA10, OXSR1; for ribeye area, genes TWIST2, SFXN1, CMYA5, CPQ and MRS2; for backfat thickness, genes OR2S2, 5S_rRNA, LOC100299372, LOC523083, LOC532403, LOC613441, SNORA69 and ITGA9; and for marbling, genes EMCN, LNX1, EIF5, SNORA28 and DSC3. The various genomic regions associated with small effects show the complexity of these phenotypes and that they do not depend only on the effects of a few genes to determine their variations.

 

 

Genet. Mol. Res. 18(1): GMR18226
DOI: 10.4238/gmr18226
Research Article

The objective of this study was to investigate, using path analysis, the genotypic correlations of primary and secondary production components as well as their decomposition into direct and indirect effects on the yield of coffee crops subjected to programed cycle pruning (PCP). Twenty-two Conilon coffe genotypes belonging to the breeding program developed by Incaper were subjected to PCP and 17 morpho-agronomic traits were measured in new branches. The traits plagiotropic branch length and number of plagiotropic leaves were discarded to eliminate collinearity problems. Path analysis was efficient in identifying the traits with greatest influence on yield. The most important traits linked to yield were number of orthotropic branches and orthotropic branch dry matter, while the secondary traits were orthotropic branch diameter, orthotropic internode length, and length of the plagiotropic branch internode. To increase coffee crop yield, producers should select genotypes that produce an intermediate number of orthotropic branches, which have a higher dry matter yield. Subsequently, they should select genotypes with larger orthotropic branch diameters, and among these, those which have shorter orthotropic and plagiotropic internode lengths.

 

Genet. Mol. Res. 18(1): GMR18114
DOI: 10.4238/gmr18114
Research Article

The parasitic protozoan Perkinsus marinus (Perkinsidae) is known to infect marine bivalves; unfortunately, treatment options are quite limited. The parasite is associated with mass mortalities worldwide and it requires notification to the World Organization for Animal Health (OIE). In Brazil, since the first report of P. marinus infecting Crassosstrea rhizophorae in the state of Paraíba in 2013, populations of oysters have been subject to continuous surveillance programs by the Ministry of Agriculture, Livestock and Food Supply for OIE-listed pathogens. Here, we report the first official case of P. marinus detection in native Crassosstrea sp. and in Crassostrea gigas from southern Brazil by PCR followed by sequencing of amplified fragments of the rDNA ITS region. For a better understanding of the epidemiology of P. marinus, we studied the parasite’s phylogenetic intraspecific variability based on its rDNA NTS region by comparisons of our isolates with other isolates from two Mexican regions on the Pacific coast. All Brazilian isolates clustered together with the Mexican isolates. As expected, high similarities were seen between all Brazilian isolates. Additional studies on P. marinus genotyping using new genomic target tools should be conducted for a better understanding of this parasite’s epidemiology.

Genet. Mol. Res. 18(1):
DOI: 10.4238/gmr18015
Research Article

Genipap (Genipa americana - Rubiaceae) is native to tropical America, occurring in Brazil in practically all biomes, except in the Pampas. It has socioeconomic importance, mainly due to the use of fruits in the manufacture of sweets and juices besides the medicinal properties of its leaves. We evaluated the genetic diversity of 73 individuals from 15 natural populations in the state of Sergipe using ISSR markers. The choice of areas was made randomly and genetic material was extracted from young leaves. PCR analysis using nine markers generated 113 fragments, which were used to estimate genetic diversity. The Shannon index was 0.36 and the similarity of individuals varied between 0.11 and 0.81. The individuals Salgado 3 (SAL3) and Salgado 4 (SAL4) were the most similar (0.81) and seven pairs had low similarity (0.11). The genetic distance the UPGMA analysis, which divided individuals into two distinct groups. The level of genetic variability found allows differentiation between genotypes that can be used to enrich the Genipap Active GermPlasm Bank in Sergipe, and the information generated will be useful for the conservation of these genetic resources and in future breeding programs for this species.

 

Genet. Mol. Res. 18(1): GMR18119
DOI: 10.4238/gmr18119
Research Article

The soybean crop is undoubtedly important to not only Brazil but also for most parts of the globe, as economic and social dependency upon this crop becomes evident. However, the scenario of the soybean crop production has faced challenges with environmental changes, which have escalated the incidence of pests. Due to the abundance of stink bugs, especially Euschistus heros in tropical regions, they can considerably impact the productivity of the crop compromising total grain yield and seed quality. Therefore, this research was aimed to evaluate soybean lines with desirable agronomic traits under high natural infestation of Euschistus heros and to access the genotypic and phenotypic correlations between important agronomic traits for soybean breeding. Twenty-three soybean F8 lines developed by the Soybean Breeding Program of Federal University of Uberlândia and four cultivars (Msoy 8527, UFUS Xavante, Msoy 8787, and UFUS Milionária) were evaluated under randomized complete block design with three repetitions during the growing season of 2015/2016 and 16 agronomic traits were accessed. There was genetic variability for all traits but Asian soybean rust severity at 1% probability level through F test. For all traits in the study but the total number of pods and Asian soybean rust severity, the coefficient of genotypic determination was superior to 70%, indicating that the most part of phenotypic variability was due to genetic differences among the genotypes in study. Grain yield was demonstrated to be a useful trait for indirect selection of soybean genotypes with resistance to brown stink bugs. Also, early cycle genotypes, heavy seeds and a higher number of pods revealed a positive correlation with grain yield over the same conditions. The lines G1, G2, and G24 are very promising genotypes as they have shown valuable agronomic traits for stink bug resistance.

Genet. Mol. Res. 18(1): GMR18182
DOI: 10.4238/gmr18182
Research Article

The DMRT3 gene has been described as the main gene influencing gait in horses; however, most studies have examined its effects by comparing information from trotted and gaited individuals. Within the Brazilian Mangalarga Marchador breed, animals can present two types of gaits (marcha batida and marcha picada), which differ in the movement form – diagonal (marcha batida) or lateral (marcha picada). We examined a possible association of the g.22999655C>A DMRT3 SNP with the type of gait, in this two-gaits breed. The relationship between individuals was calculated for the two phenotypic groups, as well as the inbreeding coefficient and its trend over generations. The allelic and genotypic frequencies were estimated based on genotype data from 310 horses (205 marcha batida; 105 marcha picada).Two threshold models were used to determine the proportion of phenotypic variance explained by DMRT3 genotypes. The average coancestry for animals with marcha batida was 0.04 ± 0.00, whereas in marcha picada it was 0.05 ± 0.01 (higher than the population mean of 0.04 ± 0.00). The inbreeding coefficient was 1.71% and 3.48% for marcha batida and marcha picada animals, respectively. Effects of the DMRT3 gene ranged from 9.94 to 10.77% (as a proportion of phenotypic variance) when included as an uncorrelated random effect and as a covariable, respectively. The genotypic frequencies of the DMRT3 and the patterns of kinship and inbreeding could be a result of selection in this breed, resulting in statistical confusion between the DMRT3 effects and population structure.

 

Genet. Mol. Res. 18(1): GMR18217
DOI: 10.4238/gmr18217
Research Article

Atherosclerosis is a multifactorial chronic inflammatory disease that occurs in response to endothelial aggression. Systemic arterial hypertension is the main risk factor for the formation of atheromas, increasing the risk of cardiovascular diseases. Several genes are involved in atherogenesis and hypertension. We analyzed polymorphisms of candidate genes that potentially participate in processes related to this pathology, including G894T and T786C of eNOS, as well as GSTT1 and GSTM1 in 167 hypertensive patients and 100 controls. Blood samples were from patients attended at the Angiogenesis/Vascular Surgery and Cardiology Department of the Angiogyn clinic in Goiania. There was significant prevalence of the genotype GT (76%) and the mutant allele T (56%) of the T786C (eNOS) polymorphism in the patients. For the polymorphism T786C (eNOS), the heterozygote genotype (TC) was found in 58% of the samples; allele C was found in 61%, but there was no significant difference compared to controls. The GSTT1 genotype was found in 84% and GSTM1 was found in 73%; for both their predominance was significant. There are many possible explanations for how these polymorphisms affect the development of atherosclerosis and hypertension, but more studies are necessary for their elucidation.

 

Genet. Mol. Res. 18(1): GMR18089
DOI: 10.4238/gmr18089
Research Article

DNA-assisted selection can be applied to vegetal species in the seed stage; however, little is known about the effect of seed fractionation on the physiological quality and viability of the seedlings or the effectiveness of DNA extraction from seed pieces. We evaluated the efficiency of pre-germinative genotypic screening by DNA markers from manually cut partial seeds of rice, beans and maize. Tests to evaluate PCR amplification and physiological quality were performed. We observed that the Sarkosyl method was efficient to extract DNA from a ½ fraction of the rice seeds and ¼ of the bean and maize seeds, generating good quality SSR-PCR products. The physiological quality of the rice seeds cut in half and the bean and maize seeds remaining fraction of ¾ of the original seed provided a high germination percentage. The method is effective for simultaneously genotyping and germinating plants from a single seed, since DNA extracted from these fractions of seeds can be used for studies with DNA markers, while the remaining portions can be used for seedling production.

 

Genet. Mol. Res. 18(1): GMR18130
DOI: 10.4238/gmr18130
Research Article

The envelope E2 glycoprotein of classical swine fever virus (CSFV) is an immunodominant protein which neutralizes the virus. Previous trials to eradicate CSFV have been expensive, inefficient and time-consuming process without complete success. In contrast to DNA transfection into cultured cells, the efficacy of gene transduction in in vivo organ is very low because of the presence of cortical laminar structures and multilayered cells followed by quality and quantity of DNA. Therefore, to eradicate CSFs, developing an effective and inexpensive vaccine targeting E2 glycoprotein is important. Here, we reported using different quantity of DNA with lipofectamine-2000 reagents that could markedly enhance the effectiveness of gene transfer in particular experiment while we are looking for long term development of animal vaccine and an alternative strategy for large scale production of CSFV E2 glycoprotein using baculovirus (bac-to-bac) system in silkworm, Bombyx mori, L.  Our results show successful expression of E2 glycoprotein in BmN cell lines and silkworm larvae. The direct injection of recombinant rBacmid/BmNPV/ E2 DNA with lipofectamine-2000 reagent infecting the silkworm larvae are varied in different groups and clear symptoms of infection were found and polyhedrons were counted by hemocytometer in individual and different batch. Confocal and electronic microscopy further revealed the expressed polyhedral, followed by SDS-PAGE and western blot further supporting our data. Our study provides an alternative strategy to produce large scale protein against CSFV. Current work to purify the E2 protein for elucidating its structure and development of vaccine is underway.

Genet. Mol. Res. 18(1): GMR18084
DOI: 10.4238/gmr18084
Research Article

Cotton is the most widely utilized natural fiber in the world. Brazil is currently one of the world’s largest cotton producers. Cotton crops are cultivated in all regions of the country, especially in the Cerrado biome. Studies of genotype x environment (GxE) interactions evaluate the adaptability and stability of cotton genotypes. Adaptability and stability evaluations help understand genotype responses to environmental stimuli and the predictability of genotypes in their response to environmental oscillations. We examined the effect of the genotype x environment interaction on cotton yield and fiber characteristics and compared artificial neural networks (ANNs) with conventional methods for assessing adaptability and stability of colored-fiber cotton genotypes. The experiment was conducted at the experimental farm of Universidade Federal de Uberlândia, during four crop years. Twelve genotypes of colored-fiber cotton were evaluated. The experimental design was randomized complete blocks with three replicates. Seed cotton yield was evaluated. The GxE interaction was analyzed by the F-test at α = 0.05. Adaptability, stability, and the factors of the decomposed GxE interaction were analyzed by the Eberhart and Russell, Centroid and ANN methods. The GxE interaction was significant for the variable seed cotton yield, demonstrating differences in genotype behavior among environments. The interactions were predominantly complex. There was concordance between Eberhart and Russsell and ANN analyses. Genotypes UFUJP-02 and UFUJP-17 were responsive to environmental stimuli; they had high predictability, in addition to high fiber yield. The ANN method reliably evaluated adaptability compared with Eberhartand Russel and Centroid methods.

 

Genet. Mol. Res. 18(1): GMR18104
DOI: 10.4238/gmr18104
Research Article

Human papillomavirus infection is the main risk factor for cervical cancer. Other risk factors include smoking and genetic susceptibility. Glutathione-S-transferases are enzymes involved in tobacco carcinogen metabolism, and genes encoding these enzymes are highly polymorphic. We compared the frequencies of GSTM1 and GSTT1 null polymorphisms in women with cervical cancer and in a control group, as well as to determine possible associations between such polymorphisms, cigarette smoking and the prognosis of cervical cancer. The series comprised 135 cervical cancer patients and 100 women without cancer. Genotypes were investigated by PCR. The results were compared using the Chi-square test or Fisher's exact test, and survival analysis by Kaplan-Meier test and Log-rank. Among the cases, the frequency of GSTM1 gene null polymorphism was 22.2%, and for the GSTT1 gene it was 48.5%. Among the controls, the frequency of the GSTM1 gene null polymorphism was 45.0%, while for GSTT1 it was 56.0%. A significant association was found between smoking and cervical cancer (P = 0.0062; OR = 2.16). Differently from GSTM1, the GSTT1 null polymorphism was not associated with cervical cancer risk in this study. The GSTT1 null genotype was significantly associated with worse prognosis. The overall survival rate for the cervical cancer group was 78.5%, and when stratified by genotypes, survival was higher in patients presenting at least one of the alleles, GSTT1 or GSTM1, indicating a higher risk of death for those presenting dual nullity (P = 0.031; RR = 2.458).

Key words:
Genet. Mol. Res. 19(1): GMR18067
DOI: 10.4238/gmr18067

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