ABSTRACT. Thirty-seven 45 X Turner syndrome patients with confirmed peripheral blood lymphocyte karyotype were initially selected to determine the origin of the retained X chromosome and to correlate it with their parents’ stature. Blood samples were available in 25 families. The parental origin of the X chromosome was determined in 24 informative families through the analysis of the exon 1 - CAG repeat variation of the androgen receptor gene. In 70.8% of the cases, the retained X chromosome was maternal in origin and 29.2% was paternal. When we classified the patients according to maternal (Xm) or paternal (Xp) X chromosome, there was a positive correlation between patients’ and maternal heights only in the Xm group. There was no correlation with paternal height in either group, and a significant correlation with target height was only observed in the Xm group. In conclusion, maternal height is the best variable correlating with the height of 45 X Turner syndrome patients who retain the maternal X chromosome, suggesting a strong influence of genes located on the maternal X chromosome on stature.

ABSTRACT. Within the Meliponini, a widely distributed group of stingless bees, Melipona rufiventris has been considered as a single, cohesive species. Recently, analysis of morphological characters led to the splitting of this species into two species, M. mondury and M. rufiventris. The former occurs in the Atlantic Rain Forest ranging from Santa Catarina to Bahia States, while the latter is found in other parts of Brazil. We used PCR + RFLP to identify genetic marker patterns of the mtDNA between these species. Nine mtDNA regions were amplified and digested with four restriction enzymes (EcoRI, EcoRV, HindIII, and HinfI). Six species-specific restriction sites were identified for M. mondury and M. rufiventris with all enzymes, except for HindIII. The molecular data agree with the morphological classification.

ABSTRACT. Anaplasmosis is a bovine intraerythrocytic disease caused by the bacterium Anaplasma marginale; it causes significant economic losses in tropical and subtropical regions, worldwide. The msp4 gene of an A. marginale strain isolated in Paraná, Brazil, was amplified by PCR and sequenced; its cloning into the pET102/D-TOPO^® vector produced an msp4-6xHis-V5-HP thioredoxin fusion gene construct. This recombinant clone was over-expressed in Escherichia coli BL21(DE-3); the expressed fusion protein was found almost entirely in the insoluble form (inclusion bodies) in the cell lysate. The inclusion bodies were solubilized with urea and the recombinant protein was purified by Ni-NTA column and dialyzed. This method produced a relatively high yield of rMSP4, which was used to immunize rabbits. The deduced amino acid sequence encoded by MSP4 showed 99% homology to A. marginale isolates from Florida, USA, and from Minas Gerais, Brazil. Both rMSP4 and native MSP4 were recognized by post-immunization rabbit serum, showing that rMSP4 has conserved epitopes. As antigenicity was preserved, rMSP4 might be useful for the development of vaccine against anaplasmosis.

ABSTRACT. Twenty-five RAPD loci and 6 isozyme loci were studied to characterize the genetic variability of natural populations of Anthonomus grandis from two agroecosystems of Brazil. The random-amplified polymorphic DNA data disclosed a polymorphism that varied from 52 to 84% and a heterozygosity of 0.189 to 0.347. The index of genetic differentiation (G_ST) among the six populations was 0.258. The analysis of isozymes showed a polymorphism and a heterozygosity ranging from 25 to 100% and 0.174 to 0.277, respectively. The genetic differentiation (F_ST) among the populations obtained by isozyme data was 0.544. It was possible to observe rare alleles in the populations from the Northeast region. The markers examined allowed us to distinguish populations from large-scale, intensive farming region (cotton belts) versus populations from areas of small-scale farming.

ABSTRACT. In Nysius californicus (family Lygaeidae, subfamily Orsillinae), a pest commonly known as the seed bug, the chromosome complement is 2n = 16 (12A + 2m + XY), testes are formed by seven seminiferous tubules covered by an orange-colored membrane, and spermatogenesis is cystic. At prophase, sex chromosomes are heteropycnotic and autosomes usually show a chiasma. At metaphase, sex chromosomes along with microchromosomes may be seen located at the center of a ring formed by the remaining autosomes. A characteristic specific of N. californicus was the presence of nucleolar material observed from the cystic cell to the completely differentiated spermatozoon. Variations in size, shape and location of the nucleolar material occur during this process, denoting a variable degree of activity in the different stages.

ABSTRACT. Telomerase activity is responsible for telomere maintenance and is believed to be crucial in most immortal cells and cancer cells; however, its clinicopathological significance in gastric cancer remains to be clarified. The aim of the present study was to assess whether malignant progression of gastric adenocarcinoma correlates with telomerase activity. We also investigated the correlation between telomerase activity and histopathological findings. We examined telomerase activity in tumor specimens and adjacent normal tissues from 43 patients with gastric adenocarcinoma. Telomerase activity was measured quantitatively by the TRAPEZE Gel Based Telomerase Detection Kit. Approximately 98% of the tumor tissues were telomerase positive, but telomerase activity was detected not only in tumor tissues but also in normal gastric mucosa. Although telomerase activity was found to be higher in tumor samples than normal tissue for each subject, we could not find a general cut-off level for telomerase activity in gastric adenocarcinoma. In addition, telomerase activity was not correlated with tumor invasion, lymph node involvement and histological stage. Our results support the idea that telomerase reactivation is a common event in gastric adenocarcinoma and it is not related to histopathological parameters. Since it is difficult to set a cut-off level for this type of cancer, we suggest that the prognostic utility of telomerase assay has not yet reached the clinic in terms of predicting outcome for patients with gastric adenocarcinoma. For the assessment of gastric carcinoma, telomerase activity should be evaluated in both tumor and normal tissues, because normal gastric mucosa samples show appreciable telomerase activity.

ABSTRACT. The purpose of the present study was to determine the effects of the steroidal plant hormone, 24-epibrassinolide (BL), on the mitotic index and growth of onion (Allium cepa) root tips. The classical Allium test was used to gather and quantify data on the rate of root growth, the stages of mitosis, and the number of mitoses in control and BL-treated groups of onions. Low doses of BL (0.005 ppm) nearly doubled the mean root length and the number of mitoses over that of controls. Intermediate doses of BL (0.05 ppm) also produced mean root lengths and number of mitoses that were significantly greater than those of the controls. The highest dose of BL (0.5 ppm) produced mean root lengths and number of mitoses that were less than control values, but the differences were not statistically significant. Examination of longitudinally sectioned root tips produced relatively similar results. This study confirms the suppositions of previous authors who have claimed that exogenously applied BL can increase the number of mitoses in plants, but failed to show cytogenetic data. This is the first report detailing the effects of BL on chromosomes and the cell cycle.

ABSTRACT. In the present study, we report on the case of a 43-year-old male patient seeking for fertility assistance, who showed a seminal analysis and testicular biopsy of complete azoospermia. Peripheral blood culture for chromosome studies revealed a karyotype of 46 chromosomes with a ring-Y-chromosome that lost the long arm heterochromatin. Molecular analysis of genomic DNA from the patient detected the presence of the sex-determining region of the Y-chromosome (SRY) but the complete absence of regions involved in spermatogenesis (AZFa, AZFb, AZFc). Several molecular markers distributed along the Y-chromosome were tested through PCR amplification, and a breakpoint was established close to the centromere, predicting the deletion of the growth control region, in agreement with the short stature observed in this patient. All results obtained through molecular cytogenetic characterization are in accordance with the clinical features observed in this patient.

ABSTRACT. Records of Holstein cows from the Dairy Records Processing Center at Raleigh, NC were edited to obtain three data sets: 65,720 first, 50,694 second, and 65,445 later lactations. Correlations among yield traits and somatic cell score were estimated with three different models: 1) bovine somatotropin (bST) administration ignored, 2) bST administration as a fixed effect and 3) administration of bST as part of the contemporary group (herd-year-month-bST). Heritability estimates ranged from 0.13 to 0.17 for milk, 0.12 to 0.20 for fat, 0.14 to 0.16 for protein yields, and 0.08 to 0.09 for somatic cell score. Estimates were less for later than first lactations. Estimates of genetic correlations among yields ranged from 0.35 to 0.85 with no important differences between estimates with the 3 models. Estimates for lactation 2 agreed with estimates for lactation 1. Estimates of genetic correlations for later lactations were generally greater than for lactations 1 and 2 except between milk and protein yields. Estimates of genetic correlations between yields and somatic cell score were mostly negative or small (-0.45 to 0.11). Estimates of environmental correlations among yield traits were similar with all models (0.77 to 0.97). Estimates of environmental correlations between yields and somatic cell score were negative (-0.22 to -0.14). Estimates of phenotypic correlations among yield traits ranged from 0.70 to 0.95. Estimates of phenotypic correlations between yields and somatic cell score were small and negative. For all three data sets and all traits, no important differences in estimates of genetic parameters were found for the two models that adjusted for bST and the model that did not.

ABSTRACT. Records of Holstein cows were used to examine how different models account for the effect of bovine somatotropin (bST) treatment on genetic evaluation of dairy sires for yield traits and somatic cell score. Data set 1 included 65,720 first-lactation records. Set 2 included 50,644 second-lactation records. Set 3 included 45,505 records for lactations three, four and five. Estimated breeding values (EBV) of sires were with three different animal models. With Model 1, bST administration was ignored. With Model 2, bST administration was used as a fixed effect. With Model 3, administration of bST was used to define the contemporary group (herd-year-month of calving-bST). Correlations for EBV of 1,366 sires with treated daughters between pairs of the three models were calculated for milk, fat and protein yields and somatic cell score for the three data sets. Correlations for EBV of sires between pairs of models for all traits ranged from 0.971 to 0.999. Fractions of sires with bST-treated progeny selected in common (top 10 to 15%) were 0.94 and usually greater for all pairs of models for all traits and parities. For this study, the method of statistical adjustment for bST treatment resulted in a negligible effect on genetic evaluations of sires when some daughters were treated with bST and suggests that selection of sires to produce the next generation of sires and cows might not be significantly affected by how the effect of bST is modeled for prediction of breeding values for milk, fat and protein yields and somatic cell score.

ABSTRACT. The ability to detect nuclear damage is an important tool for the development of sperm preservation methods. We used the acridine orange test (AOT) and the terminal deoxynucleotidyl transferase-mediated dUDP nick-end labeling (TUNEL) assay to assess the DNA status of sperm cells preserved with different lyophilization media. The AOT did not detect any differences between different lyophilization media. However, differences in DNA integrity were observed among treatments with the TUNEL assay, suggesting that TUNEL is a more sensitive method to evaluate sperm DNA. The use of TCM 199 and 10% FCS as a lyophilization medium resulted in 14% of the cells with DNA fragmentation in TUNEL test. The AOT indicated only 4% of the cells with chromatin damage, with this same treatment, with no significant differences when compared to the other treatments. The degree of DNA fragmentation was negatively related to fertilizing potential, as sperm DNA damage was inversely correlated with pro-nucleus formation. The TUNEL assay was found to be an efficient method to detect DNA damage in sperm, and it could be used as a tool to predict male fertility.

ABSTRACT. This paper presents a novel approach to the problem of splice site prediction, by applying stochastic grammar inference. We used four grammar inference algorithms to infer 1465 grammars, and used 10-fold cross-validation to select the best grammar for each algorithm. The corresponding grammars were embedded into a classifier and used to run splice site prediction and compare the results with those of NNSPLICE, the predictor used by the Genie gene finder. We indicate possible paths to improve this performance by using Sakakibara’s windowing technique to find probability thresholds that will lower false-positive predictions.

ABSTRACT. In Brazil, the center of cassava origin, cassava landraces have acquired through their domestication a large diversity in relation to many economic traits such as high content of carotenoids and excellent palatability among other characters. One of these clones, which has been grown by indigenous Brazilian farmers and is now being maintained in the University of Brasília gene bank, showed a high level of lycopene content (5 mg/kg viz. a viz. zero in common cultivars, and 12-20 mg/kg in tomato, a lycopene-rich vegetable). A second landrace called UnB 400 had a high content of b-carotene, which reached 4 mg/kg.

ABSTRACT. The goal of the present study was to determine if simple methods, especially hot saline solution (HSS) and MspI and HaeIII restriction endonucleases, which do not require special equipments, may be helpful in studies of genetic variability in the lady beetle, Cycloneda sanguinea. The HSS method extracted the heterochromatin region, suggesting that it is composed mostly of DNA rich in A-T base pairs. However, the X and y chromosomes were resistant to HSS banding. These bands facilitated the identification of each chromosome. In this study, we used the restriction endonucleases with different G-C base target sequences: MspI C/GGC and HaeIII GG/CC. The use of restriction enzyme MspI did not show an effect on the autosomal chromosomes. On the other hand, the sex pair showed a pale staining, to help in the recognition of these chromosomes. HaeIII produced characteristic bands which were identified all along the chromosomes, facilitating the identification of each chromosome. Based on these results, we can consider the heterochromatin being heterogeneous. The findings obtained here, using different chromosomal banding techniques, may be useful in the identification of intraspecific chomosome variability, specifically in Coccinellidae (Coleoptera) chromosomes, even without special equipment.

ABSTRACT. Perna perna is the most important cultivated mussel of Santa Catarina, Brazil, sustaining an important economic input for many local families. Natural stocks of P. perna are depleted by the extraction of adults and seeds for consumption and culture. The aim of the present study was to use the microsatellite locus pms-2 to study the variation of the genetic composition and diversity between natural and cultured stocks in samples of 2001 and 2005 from Penha, Santa Catarina. DNA was extracted from adductor muscle by Chelex/proteinase-K and phenol/chloroform protocols. Amplification by polymerase chain reaction was performed using specific primers for analyzing the pms-2 locus. Polymerase chain reaction products were submitted to vertical denatured 6% polyacrylamide gel electrophoresis and horizontal 2% agarose gel electrophoresis, and visualized by silver staining and ethidium bromide, respectively. Allele diversity and heterozygote deficiency were higher for samples of 2005 than for those of 2001. No significant genetic differentiation was found between natural and cultured stocks of 2001 by the c² test, but G² (likelihood ratio) detected slight differences (I = 0.949; c², P = 0.147; G², P = 0.046), while cultured and natural stocks of 2005 were very different (I = 0.798, P = 0.006). Between the years of 2001 and 2005, a large change in genetic composition was observed (I = 0.582; P 0.001). Although nothing is known about natural changes in the genetic composition of this species with time, the results suggest a strong impact of human activities on natural stocks of P. perna, which is expected to be related to heavy extraction and farming.

ABSTRACT. Cytological analysis of microsporogenesis in 72 popcorn plants, comprising nine from the original population (CMS-43, S₀) and 63 from seven cycles of self-fertilization (S₁ to S₇), one plant of S₀ generation (plant 2) was identified with B chromosomes. The number of B chromosomes varied from two to three in the same anther. The pattern of chromosome pairing and meiotic behavior of Bs were similar to those found in other plant species. The presence of B chromosomes did not affect chiasma frequency and chiasma distribution in A chromosomes. This is the first report of B chromosomes in popcorn.

ABSTRACT. Suppressor of cytokine signaling 2 (SOCS2) is a negative regulator of growth hormone signaling. The deletion of SOCS2 in mice results in a 30-50% increase in post-natal growth. In an effort to identify polymorphisms in the SOCS2 gene that may be associated with body size in dogs, we characterized the canine SOCS2 gene and analyzed its genetic diversity among small and large dog breeds. The study was carried out on a total of 520 dogs from 66 different breeds. Dogs were classified as large or small based on height and weight as determined by their respective American Kennel Club breed standards. The SH2 and SOCS domains of the canine SOCS2 gene were sequenced in 32 dogs from different breeds. Only one non-synonymous sequence variant (DQ415457:g.326G>T) was detected which corresponds to an amino acid change (Asp127Tyr). All samples were genotyped by PCR/RFLP and the allele frequencies were determined for each dog breed. The T allele was distributed primarily among European large dog breeds with a gene frequency ranging from 0.72 to 0.04. The nature of the nucleotide change and the effect on the protein together with the finding of a QTL related to body size in the same CFA15 region by other researchers suggest canine SOCS2 as a potential candidate gene for body size in dogs. Future studies will be needed to clarify the role of the 326G>T polymorphism and its interaction with genes like growth hormone and insulin-like growth factor 1.

ABSTRACT. The putative eukaryotic translation initiation factor 5A (eIF5A) is an essential protein for cell viability and the only cellular protein known to contain the unusual amino acid residue hypusine. eIF5A has been implicated in translation initiation, cell proliferation, nucleocytoplasmic transport, mRNA decay, and actin polarization, but the precise biological function of this protein is not clear. However, eIF5A was recently shown to be directly involved with the translational machinery. A screen for synthetic lethal mutations was carried out with one of the temperature-sensitive alleles of TIF51A (tif51A-3) to identify factors that functionally interact with eIF5A and revealed the essential gene YPT1. This gene encodes a small GTPase, a member of the rab family involved with secretion, acting in the vesicular trafficking between endoplasmatic reticulum and the Golgi. Thus, the synthetic lethality between TIF51A and YPT1 may reveal the connection between translation and the polarized distribution of membrane components, suggesting that these proteins work together in the cell to guarantee proper protein synthesis and secretion necessary for correct bud formation during G1/S transition. Future studies will investigate the functional interaction between eIF5A and Ypt1 in order to clarify this involvement of eIF5A with vesicular trafficking.

ABSTRACT. Duffy blood group genotype was studied in 95 unrelated subjects from four African-Brazilian communities of the Amazon region: Trombetas, Pitimandeua, Curiaú, and Mazagão Velho. Genotyping was performed using an allele-specific primer polymerase chain reaction technique for determining the three major alleles at FY blood group, and as expected, FY*O allele was the most common one, with frequencies ranging from 56.4% in Mazagão Velho to 72.2% in Pitimandeua, whereas the FY*O/FY*O genotype was found with frequencies between 32.3% in Mazagão Velho and 58.8% in Curiaú. Genotype and allele distributions in the four Amazonian communities are consistent with a predominantly African origin with some degree of local differentiation and admixture with people of Caucasian ancestry and/or Amerindians. These results reveal that the impact of the FY*O/FY*O genotype on the transmission and endemicity of the vivax malaria deserves to be investigated in full detail in an attempt to identify the contribution of host biological factors and explain the non-homogeneous prevalence of malaria in the region expressed by its different levels of exposure.

ABSTRACT. Five published DNA extraction protocols were compared for their ability to produce good quality DNA from fresh and herbarium leaves of several species of the genus Dalbergia. The leaves of these species contain high amounts of secondary metabolites, which make it difficult to perform a clean DNA extraction and thereby interfering with subsequent PCR amplification. The protocol that produced the best DNA quality in most of the Dalbergia species analyzed, utilizes polyvinylpyrrolidone to bind the phenolic compounds, a high molar concentration of NaCl to inhibit co-precipitation of polysaccharides and DNA, and LiCl for removing RNA by selective precipitation. The DNA quality of herbarium specimens was worse than that for fresh leaves, due to collecting conditions and preservation of samples. We analyzed 54 herbarium specimens, but the recovered DNA allowed successful PCR amplification in only eight. For the genus Dalbergia, the herbarium is an important source of material for phylogenetic and evolutionary studies; due to the occurrence of the different species in various geographical regions in Brazil, it is difficult to obtain fresh material in nature. Our results demonstrated that for Dalbergia species the methods used for the collection and preservation of herbarium specimens have a mayor influence on DNA quality and in the success of phylogenetic studies of the species.

ABSTRACT. The present study was carried out to estimate both (co)variance components and genetic parameters for frame scores obtained using two methods (FRAME_GMA and FRAME_BIF) as well as phenotypic and genetic correlations with traits such as weaning weight, weight gain from weaning to yearling, scrotal circumference, muscle score, and an empiric index for animal classification for the Special Certificate of Identification and Production (CEIP). Data on 12,728 animals, raised in Southeastern Brazil, with ages from 490 to 610 days were analyzed. Estimates of heritability for FRAME_GMA and FRAME_BIF in multi-trait analysis were 0.28 and 0.24, respectively. Genetic correlation coefficients between frame scores and the growth trait were of medium magnitude, which indicates that genetic selection for weight resulted in undesirable responses, increasing the animals’ frames. Small changes should be expected in the frame of animals that have been submitted to a genetic selection regarding muscle score and scrotal circumference. The low magnitude of phenotypic and genetic correlation between frame scores and the empirical selection index that classifies animals for CEIP, a Brazilian official certificate that recognizes the value of seedstock that is not registered at breeders associations, but is genetically evaluated, does not indicate important responses in giving a CEIP to animals that have been directly or indirectly selected for frame. Other studies must be performed to determine estimates of the genetic parameters for frame scores in other beef cattle populations.

ABSTRACT. Acid phosphatase activity (Gömori technique) in salivary gland cells was investigated in adult insects (males and females) of four species of triatomines: Triatoma infestans, Panstrongylus megistus, Rhodnius neglectus, and Rhodnius prolixus. Binucleated cells with bulky and polyploidy nuclei were detected, with acid phosphatase activity in the heterochromatin and nucleolus, which showed the most intense response. Thus, the activity of these phosphatases during rRNA molecule transcription, possibly in the nucleolar fibrillar center, is suggested. The difference in reactivity found among salivary glands is associated with the cellular metabolism of these regions and, probably, with the biosynthesis of their different secretions. This must be essential in maintaining the hematophagy of triatomines.

ABSTRACT. Expressed sequenced tags (ESTs) were prepared to establish a baseline for molecular genetic studies of the tarnished plant bug, Lygus lineolaris (Palisot de Beauvois). The largest class of identifiable ESTs (15.2%) was from genes involved in cellular metabolic functions, including physiological processes. Twenty-seven ESTs (9.8%) were from genes associated with transcription and translation, including ribosomal genes. One hundred and forty-two of the 276 unique ESTs were from genes not previously identified from any organism. Twelve sequences appear to be associated with feeding and digestion and may be targets for pest control studies.

ABSTRACT. Cultures of adipose tissue explants are a valuable tool for studying the intracellular mechanisms involving hormones and nutrients. However, testing how fatty acids affect cells requires a carrier molecule; bovine serum albumin (BSA) has been used for this purpose. However, contaminants can alter the cellular response. Our objectives were to: 1) test BSA as a fatty acid carrier and 2) evaluate polyvinyl alcohol (PVA) as a replacement for BSA. Adipose tissue explants from nine pigs were cultured in medium 199 for 4, 12, 24, and 48 h, with the following treatments: control, PVA (100 mM PVA added) and PVA + pGH (100 mM PVA plus 0.1 mg/mL porcine growth hormone). After each culture period, explants were collected and assayed for lipogenesis. After 48 h in culture, explants were assayed for lipolysis. A preliminary study with different commercial sources and high concentrations showed that BSA affected lipogenic rates. On the other hand, there were no effects of PVA on lipid synthesis, while pGH (positive control) reduced glucose incorporation into lipids (P < 0.01) when compared to both control and PVA (P < 0.05). There was no difference between control and PVA for lipolysis rates. However, pGH increased lipolysis when compared to control (P < 0.01) and PVA (P < 0.05). We demonstrated that BSA can alter lipogenesis, which precludes its use as a carrier molecule. On the other hand, addition of PVA had no effect on lipolysis or lipogenesis. We suggest the use of PVA instead of BSA for adding bioactive fatty acids to cultures of adipose tissue.

ABSTRACT. Sequence polymorphisms in the growth hormone (GH) gene and its transcriptional regulators, Pit-1 and Prop-1, were evaluated for associations with growth and carcass traits in two populations of Brangus bulls Chihuahuan Desert Rangeland Research Center (CDRRC, N = 248 from 14 sires) and a cooperating breeding program (COOP, N = 186 from 34 sires). Polymorphisms were SNP mutations in intron 4 (C/T) and exon V (C/G) in GH, A/G in exon VI in Pit-1, and A/G in exon III in Prop-1. In the COOP population, bulls of Pit-1 GG genotype had a significantly greater percentage of intramuscular fat than bulls of the AA or AG genotype, and bulls of the Prop-1 AA genotype had significantly greater scrotal circumference than bulls of AG or GG genotypes at ~365 days of age. Also, heterozygous genotypes for the two GH polymorphisms appeared advantageous for traits of muscularity and adiposity in the COOP population. The heterozygous genotype of GH intron 4 SNP was associated with advantages in weight gain, scrotal circumference, and fat thickness in the CDRRC population. The two GH polymorphisms accounted for ³27.7% of the variation in these traits in the CDRRC population; however, R² was <5% in the COOP population. Based on haplotype analyses the two GH SNPs appeared to be in phase; the haplotype analyses also paralleled with the genotype analyses. Polymorphisms in GH and its transcriptional regulators appear to be predictors of growth and carcass traits in Brangus bulls, particularly those with heterozygous GH genotypes.