Microsatellite Genotyping of Plasmodium vivax Isolates from Pregnant Women in Four Malaria Endemic Countries

Plasmodium vivax is the most widely distributed human parasite and the main cause of human malaria outside the African continent. However, the knowledge about the genetic variability of P. vivax is limited when compared to the information available for P. falciparum. We present the results of a study aimed at characterizing the genetic structure of P. vivax populations obtained from pregnant women from different malaria endemic settings. Between June 2008 and October 2011 nearly 2000 pregnant women were recruited during routine antenatal care at each site and followed up until delivery. A capillary blood sample from the study participants was collected for genotyping at different time points. Seven P. vivax microsatellite markers were used for genotypic characterization on a total of 229 P. vivax isolates obtained from Brazil, Colombia, India and Papua New Guinea. In each population, the number of alleles per locus, the expected heterozygosity and the levels of multilocus linkage disequilibrium were assessed. The extent of genetic differentiation among populations was also estimated. Six microsatellite loci on 137 P. falciparum isolates from three countries were screened for comparison. The mean value of expected heterozygosity per country ranged from 0.839 to 0.874 for P. vivax and from 0.578 to 0.758 for P. falciparum. P. vivax populations were more diverse than those of P. falciparum. In some of the studied countries, the diversity of P. vivax population was very high compared to the respective level of endemicity. The level of inter-population differentiation was moderate to high in all P. vivax and P. falciparum populations studied.     

Effects of light and temperature on seed germination of cacti of Brazilian ecosystems

Environmental factors are used by plants as spatio‐temporal indicators of favorable conditions for seed germination. Thus, the objective of this study was to determine the effect of light and temperature on seed germination of 30 taxa of Cactaceae occurring in northeastern Brazil and to evaluate whether fluctuations in temperature are capable of altering light sensitivity. The seeds were tested for germination under two light conditions (12 h photoperiod and continuous darkness) and 10 temperature treatments: eight constant temperatures (10, 15, 20, 25, 30, 35, 40 and 45°C) and two alternating temperatures (30/20°C and 35/25°C). The species studied showed two photoblastic responses. All cacti from the Cactoideae subfamily (22 taxa) were classified as positive photoblastic (i.e., no germination in darkness), regardless of the temperature treatment used. Likewise, temperature fluctuation did not alter the seed sensitivity to light. On the other hand, the species of the Opuntioideae (five taxa) and Pereskioideae (three taxa) subfamilies are indifferent to light (i.e., germinated both in the presence and absence of light). The cacti from the areas of Caatinga and Cerrado showed an optimal germination temperature of 30°C, while the species from Atlantic Forest and Restinga areas showed an optimal germination temperature of 25°C.     

Determination of ellagic acid in oak leaves and in sheep ruminal fluid by ion-pair RP-HPLC

An isocratic ion-pair high-performance liquid chromatography (IP-RP-HPLC) method with UV detection was developed to identify and quantify ellagic acid (EA). This phenolic compound is widely distributed in the plants and is often present in the diet of ruminants. The method was validated and validation parameters were: linearity range 5-100 mg/L; correlation coefficient, 0.9995; mean recoveries (99.94 and 101.07%) and detection limit 1.4 mg/L. Method was applied for the determination of ellagic acid in oak leaves and in ruminal fluid from to a vitro ruminal system. The proposed method proved to be rapid and accurate and can be successfully used in ruminant nutrition studies.     

Towards an interactive,process‐based approach to understanding range shifts: developmental and environmental dependencies matter

Many species are undergoing distributional shifts in response to climate change. However, wide variability in range shifting rates has been observed across taxa, and even among closely‐related species. Attempts to link climate‐mediated range shifts to traits has often produced weak or conflicting results. Here we investigate interactive effects of developmental processes and environmental stress on the expression of traits relevant to range shifts. We use an individual‐based modelling approach to assess how different developmental strategies affect range shift rates under a range of environmental conditions. We find that under stressful conditions, such as at the margins of the species’ fundamental niche, investment in prolonged development leads to the greatest rates of range shifting, especially when longer time in development leads to improved fecundity and dispersal‐related traits. However, under benign conditions, and when traits are less developmentally plastic, shorter development times are preferred for rapid range shifts, because higher generational frequency increases the number of individual dispersal events occurring over time. Our results suggest that the ability of a species to range shift depends not only on their dispersal and colonisation characteristics but also how these characteristics interact with developmental strategies. Benefits of any trait always depended on the environmental and developmental sensitivity of life history trait combinations, and the environmental conditions under which the range shift takes place. Without considering environmental and developmental sources of variation in the expression of traits relevant to range shifts, there is little hope of developing a general understanding of intrinsic drivers of range shift potential.     

Changes in tropomyosin during primary culture of embryonic myocardiocytes.

We chose the Hamburger and Hamilton's stage 29 (HH 29) to investigate the expression of tropomyosin in chick myocardiocytes during 14 days on culture. Throughout 14 days of cell culture, changes in cell morphology were accompanied by a redistribution of tropomyosin in different cell compartments. We used FACScan, SDS-PAGE and densitometric analysis to quantify total cell tropomyosin and concentrations of this protein in different cell fractions. Tropomyosin was found mostly in the cytoskeletal fraction than in the cytoplasmic. When we compared the densitometric values from SDS-PAGE of cells in different stages of development we found that in HH 19, tropomyosin was more abundant in the cytoplasmic than in the cytoskeletal fraction. By HH 29, the two fractions had become inverted, and in HH 39, tropomyosin was clearly more abundant in the cytoskeletal than in the cytoplasmic fraction. In the IFI analysis, tropomyosin was found to label the Stress fiber-like structures (SFL) in different patterns depending on the area of the cell which expressed this protein.     

Novel mutations in the 3' region of the polycystic kidney disease 1 (PKD1) gene

Mutation screening in 90 unrelated ADPKD1 patients was carried out on some of the exons in the single copy area (37, 38, 39, 44, 45) using genomic PCR and SSCP. Four novel mutations were found: a 15 bp in-frame deletion in exon 39 [nt11449 (del 15)], a 2 bp deletion in exon 44 [nt12252 (del 2)], a G insertion in exon 44 [nt12290 (Ins G)], and a GTT in-frame deletion in exon 45 [nt12601 (del 3)].     

p53 Expression in circulating lymphocytes of non-melanoma skin cancer patients from an arsenic contaminated region in Mexico. A pilot study

Arsenic is a common environmental toxicant and epidemiological studies associate arsenic exposure with various pathologic disorders and several types of cancer. Skin cancers are the most common arsenic-induced neoplasias and the prevalence of skin lesions has been reported to be significantly elevated in individuals exposed to arsenic via drinking water in Mexico. Being lymphocytes the main cells used for human monitoring, we evaluated the expression of p53 protein in the lymphocytes from 44 healthy individuals and 19 samples from individuals living in a chronic arsenicism endemic region. Of the latter group, 12 individuals had non-melanoma skin cancer and 9 of them expressed p53 in the circulating lymphocytes, whereas only one of the 7 non-cancer arsenic exposed individuals expressed it. In the healthy non-arsenic exposed group only one from 44 individuals expressed the protein. These results suggest a clear relationship between non-melanoma skin cancer and p53 expression in circulating lymphocytes. p53 expression in circulating lymphocytes should be evaluated as a potential biomarker of effect or susceptibility.     

Self-incompatibility genotypes in almond re-evaluated by PCR,stylar ribonucleases,sequencing analysis and controlled pollinations

As part of the almond breeding programme at IRTA, we investigated the S genotypes of several cultivars using a combination of RNase zymograms, testcrosses, pollen-tube growth analysis and molecular identification by PCR analysis. For some of the cultivars examined, discrepancies appeared between their S alleles as reported in the literature and those found in this investigation, leading to a re-evaluation of their S genotypes. Analysis of the stylar ribonucleases (RNases), which are known to correlate with S alleles, of cvs. Achaak, Ardechoise, Desmayo Largueta, Ferrastar, Gabaix, Garbí, Glorieta, Languedoc, Primorskiy and Texas revealed inconsistencies with respect to the S₅ and S₁₀ alleles. However, PCR with the conserved primer pair AS1II/AmyC5R failed to detect any of these inconsistencies. When the S alleles from Desmayo Largueta, Gabaix, Primorskiy and Texas were sequenced, Texas and Primorskiy were found to carry the reported S₅ allele, while Desmayo Largueta and Gabaix carried a new allele, which has been tentatively denoted as S₂₅ This new S allele, previously reported to be S₁₀, was also identified in Achaak, Ardechoise and Ferrastar. The proposed new S genotypes are Achaak (S₂S₂₅), Ardechoise (S₁S₂₅), Desmayo Largueta (S₁S₂₅), Ferrastar (S₂S₂₅) and Gabaix (S₁₀S₂₅). The S alleles of Garbí, Glorieta, Languedoc, Texas and Primorskiy remain as reported in the literature. Testcrosses in the field and laboratory confirmed the new S genotypes. One cultivar (Gabaix) could be assigned to the existing cross-incompatibility group O of unique genotypes, and two new groups were established (XVI and XVII) consisting of two cultivars each. The clarification of these S alleles will be useful in almond breeding programmes and for planning new commercial orchards in the future.     

Impaired PBPC collection in patients with myeloma after high-dose melphalan

BACKGROUND: Tandem stem cell transplantation is an important treatment option for patients with myeloma and some additional tumors. In an attempt to reduce the contamination of the stem cell graft with tumor cells, patients with myeloma who entered complete remission after the first transplant underwent a second episode of mobilization to obtain progenitor cells for the second transplant. METHODS: Twenty-two patients with myeloma participated in the study. The first mobilization utilized CY, etoposide and filgrastim. The second mobilization used the same regimen, but seven patients received only filgrastim. The interval between the two collection periods was 6 months (median; range 4-9 months). The preparative regimen for the first transplant consisted of melphalan 200 mg/m(2). RESULTS: The number of total white cells collected during the two collection episodes was similar: 10.8+/-1.6 x 10(8)/kg white cells vs. 11.8+/-1.7 x 10(8)/kg white cells (P=0.63). The collected CD34(+) cell dose was much larger during the first collection: 45.2+/-8.4 x 10(6)/kg vs. 6.9+/-2.7 x 10(6)/kg (P<0.001). Similarly, the collected colony-forming unit (CFU)-GM dose was much larger during the first collection: 295.4+/-59.3 x 10(4)/kg vs. 67.3+/-21.6x10(4)/kg (P<0.001). While the CD34(+) cells collected during the two collection episodes correlated significantly (r=0.55, P<0.01); the first dose was a median of 14.9-fold larger. DISCUSSION: No laboratory parameter was able reliably to predict the results of the second collection. A second mobilization/collection episode as part of a tandem transplant approach carries a considerable risk of failing to obtain sufficient progenitor cells.