Pherogram normalization in capillary electrophoresis and micellar electrokinetic chromatography analyses in cases of sample matrix-induced migration time shifts

When analyzing bio-matrix samples using capillary electrophoresis (CE) or micellar electrokinetic chromatography (MEKC), unwanted shifts in the time axis are often observed, both between samples and standards and between samples, thus hampering identification. These shifts are caused by either or both of two sample matrix-induced effects: variations in stacking conditions (effective field strength or migration length) and variations in electroosmotic flow. Based on elementary CE principles and provided that any two peaks in the pherograms can be linked, these variations can be separately accounted and quantitatively corrected for, so that perfectly overlapping pherograms of standards and samples can be obtained after normalization. The method was validated using samples of a DNA ladder, separated in a sieving polymer. In addition, a number of data files from CE and MEKC analyses (steroids, opioids, beta-blockers, amines, and inorganic anions) previously published by other authors were successfully normalized. A freeware computer programme, CEqualizer, for normalizing ASCII files of detector signals using the method described, is available to the CE community from http: //www.ceyork.f2s.com.     

Trypanosoma cruzi: populations bearing opposite virulence induce differential expansion of circulating CD3+CD4-CD8- T cells and cytokine serum levels in young and adult rats

The JG strain is the least virulent while the CL-Brener clone is one of the most virulent Trypanosoma cruzi populations in young rats. In this study, we determined that the parasitemia peak values in CL-Brener clone-infected adult rats were 50-fold lower than in young rats and that mortality was null as compared to 45% death in young rats. Low parasitemia, milder and sustained myocarditis and myositis characterized JG infections. CL-Brener clone caused a significantly higher production of pro-inflammatory cytokines and higher expansion of CD3⁺CD4⁻CD8⁻, double-negative (DN) T cells, during the acute phase in both adult and young rats. DN T cell frequencies correlated with IFN-γ levels. These findings may explain the higher inflammation and fast acute phase resolution in CL-Brener infection. In young rats, IL-10 levels were similar in both infections. The IL-10/IFN-γ ratio was higher in JG acute infection in accordance with the milder inflammation and parasite persistence leading to a chronic phase. In conclusion, virulence and pathogenicity depend on T. cruzi ability to induce expansion of DN T cells and production of specific cytokines.     

Hybridization and low genetic diversity in the endangered Alabama red‐bellied turtle (Pseudemys alabamensis)

Pseudemys alabamensis is one of the most endangered freshwater turtle species in the United States due to its restricted geographic distribution in coastal Alabama and Mississippi. Populations of P. alabamensis are geographically isolated from one another by land and saltwater, which could act as barriers to gene flow. It is currently unknown how differentiated these populations are from one another and whether they have experienced reductions in population size. Previous work found morphological differences between Alabama and Mississippi populations, suggesting that they may be evolutionarily distinct. Other Pseudemys turtles such as P. concinna and P. floridana occur naturally within the same geographic area as P. alabamensis and are known to hybridize with each other. These more abundant species could threaten the unique genetic identity of P. alabamensis through introgression. In order to evaluate the endangered status of P. alabamensis and the level of hybridization with other species, we used mitochondrial and nuclear microsatellite markers to assess genetic variation within and among populations of P. alabamensis throughout its range and estimate admixture with co‐occurring Pseudemys species. In P. alabamensis, we found no variation in mitochondrial DNA and an excess of homozygosity in microsatellite data. Our results show genetic differentiation between Alabama and Mississippi populations of P. alabamensis, and low estimated breeding sizes and signs of inbreeding for two populations (Fowl River, Alabama and Biloxi, Mississippi). We also found evidence of admixture between P. alabamensis and P. concinna/P. floridana. Based on our results, P. alabamensis is highly endangered throughout its range and threatened by both low population sizes and hybridization. In order to improve the species’ chances of survival, focus should be placed on habitat preservation, maintenance of genetic diversity within both the Mississippi and Alabama populations, and routine population‐monitoring activities such as nest surveillance and estimates of recruitment.     

Genetic dissection of seminal root architecture in elite durum wheat germplasm

Although root architecture has been shown to play an important role in crop performance, particularly under drought conditions, no information is available on the genetic control of root traits in durum wheat, a crop largely grown in rainfed areas with low rainfall. In our study, a panel of 57 elite durum wheat accessions were evaluated under controlled conditions for root and shoot traits at the seedling stage. Significant genetic variability was detected for all the root and shoot traits that were investigated. Correlation analysis suggested that root and shoot features were only partially controlled by common sets of genes. The high linkage disequilibrium (up to 5 cM) present in the germplasm collection herein considered allowed us to use simple sequence repeat‐based association mapping to identify chromosome regions with significant effects on the investigated traits. In total, 15 chromosome regions showed significant effects on one or more root architectural features. A number of these regions also influenced shoot traits and, in some cases, plant height measured in field conditions. Major effects were detected on chromosome arms 2AL (at Xgwm294), 7AL (at Xcfa2257 and Xgwm332) and 7BL (at Xgwm577 and Xcfa2040). The accessions with the most remarkable differences in root features will provide a valuable opportunity to assemble durum wheat mapping populations well suited for ascertaining the effects of root architecture on water use efficiency and grain yield.     

Description of a New Galapagos Giant Tortoise Species (Chelonoidis; Testudines: Testudinidae) from Cerro Fatal on Santa Cruz Island

The taxonomy of giant Galapagos tortoises (Chelonoidis spp.) is currently based primarily on morphological characters and island of origin. Over the last decade, compelling genetic evidence has accumulated for multiple independent evolutionary lineages, spurring the need for taxonomic revision. On the island of Santa Cruz there is currently a single named species, C. porteri. Recent genetic and morphological studies have shown that, within this taxon, there are two evolutionarily and spatially distinct lineages on the western and eastern sectors of the island, known as the Reserva and Cerro Fatal populations, respectively. Analyses of DNA from natural populations and museum specimens, including the type specimen for C. porteri, confirm the genetic distinctiveness of these two lineages and support elevation of the Cerro Fatal tortoises to the rank of species. In this paper, we identify DNA characters that define this new species, and infer evolutionary relationships relative to other species of Galapagos tortoises.     

Stable peptide inhibitors prevent binding of lethal and oedema factors to protective antigen and neutralize anthrax toxin in vivo

The lethal and oedema toxins produced by Bacillus anthracis, the aetiological agent of anthrax, are made by association of protective antigen with lethal and oedema factors and play a major role in the pathogenesis of anthrax. In the present paper, we describe the production of peptide-based specific inhibitors in branched form which inhibit the interaction of protective antigen with lethal and oedema factors and neutralize anthrax toxins in vitro and in vivo. Anti-protective antigen peptides were selected from a phage library by competitive panning with lethal factor. Selected 12-mer peptides were synthesized in tetra-branched form and were systematically modified to obtain peptides with higher affinity and inhibitory efficiency.     

Trypanosoma cruzi: blood parasitism kinetics and their correlation with heart parasitism intensity during long-term infection of Beagle dogs

The goals of the present study were to evaluate the kinetics of blood parasitism by examination of fresh blood, blood culture (BC) and PCR assays and their correlation with heart parasitism during two years of infection in Beagle dogs inoculated with the Be-78, Y and ABC Trypanosoma cruzi strains. Our results showed that the parasite or its kDNA is easily detected during the acute phase in all infected animals. On the other hand, a reduced number of positive tests were verified during the chronic phase of the infection. The frequency of positive tests was correlated with T. cruzi strain. The percentage of positive BC and blood PCR performed in samples from animals inoculated with Be-78 and ABC strains were similar and significantly larger in relation to animals infected with the Y strain.Comparison of the positivity of PCR tests performed using blood and heart tissue samples obtained two years after infection showed two different patterns associated with the inoculated T. cruzi strain: (1) high PCR positivity for both blood and tissue was observed in animals infected with Be-78 or ABC strains; (2) lower and higher PCR positivity for the blood and tissue, respectively, was detected in animals infected with Y strains. These data suggest that the sensitivity of BC and blood PCR was T. cruzi strain dependent and, in contrast, the heart tissue PCR revealed higher sensitivity regardless of the parasite stock.     

Surveying an endangered saproxylic beetle, Osmoderma eremita, in Mediterranean woodlands: a comparison between different capture methods

Measuring population size is riddled with difficulties for wildlife biologists and managers, and in the case of rare species, it is sometimes practically impossible to estimate abundance, whereas estimation of occupancy is possible. Furthermore, obtaining reliable population size estimates is not straightforward, as different sampling techniques can give misleading results. A mark-recapture study of the endangered saproxylic beetle Osmoderma eremita was performed in central Italy by applying four independent capture methods within a study area where 116 hollow trees were randomly selected to set traps. Detection probability and population size estimates were drawn from each of these four capture methods. There were strong differences in detection probability among methods. Despite using pheromone and beetle manipulation, capture histories were not affected by trap-happiness or trap-shyness. Population size estimates varied considerably in both abundance and precision by capture method. A number of 0.5 and 0.2 adult beetles per tree was estimated using the whole data set by closed and open population models, respectively. Pitfall trap appeared the optimal method to detect the occurrence of this species. Since in the southern part of its distribution range, a single population of O. eremita is widespread in the landscape, and includes beetles from more than one hollow tree, conservation efforts should focus not only on preserving few and isolated monumental hollow trees, but should be extended to large stands.     

Maternal and neonate diatom diets impair development and sex differentiation in the copepod Temora stylifera

Development and sex differentiation in the copepod Temora stylifera was studied in the presence of maternal and larval diets of the diatoms Thalassiosira rotula and Skeletonema marinoi, either provided alone or supplemented with the control dinoflagellate Prorocentrum minimum. Both diatoms had deleterious effects on growth compared to the control when used as pure diets, inducing very low or even zero survival from hatching to adulthood. This effect was deleted when the diet was supplemented with a good food (P. minimum) only in the case of T. rotula. By contrast, with a maternal or larval diet of S. marinoi, nauplii did not pass metamorphosis even when this alga was mixed with P. minimum. Arrested development was not due to lack of feeding since early and late nauplii (NII and NV) ingested all three algae at similar rates when used as single diets, and did not show any preference when the algae were offered as mixtures. Since mortality rates with a mixed diet of S. marinoi + P. minimum were similar or even higher than those obtained with a single diet of S. marinoi, we suggest that this diatom is more toxic than nutritionally deficient for T. stlyfera development. No males were produced in cohorts reared on pure diatom diets or with a mixture of S. marinoi + P. minimum, and intermediate male:female sex ratios were obtained with the mixed T. rotula + P. minimum diet. Possibly some diatoms produce compounds such as oxylipins or new molecules that alter sex differentiation in T. stylifera.