First report of an intraerythrocytic small piroplasm in wild Iberian lynx (Lynx pardinus)

A wild injured Iberian lynx (Lynx pardinus) was taken from the Sierra Morena population. During the health check small intraerythrocytic piroplasms, morphologically indistinguishable from other feline piroplasms, were observed in Wright-Giemsa-stained blood films. Amplification by polymerase chain reaction of a portion of the 18S nuclear small subunit (NSS) rRNA gene and sequencing revealed similarity of the unknown organism with sequences obtained from Pallas's cat from Mongolia and from a domestic cat in Spain. In a retrospective (1993-2003) study of 50 Iberian lynx tissue samples, no amplifications of the 18S NSS rRNA gene of the organism were obtained. This is the first report of a naturally occurring erythroparasitemia in the Iberian lynx and the first documented case of naturally occurring piroplasm infection in a free-ranging felid from Europe.     

Reactive oxygen species and development in microbial eukaryotes

Reactive oxygen species (ROS) have been regarded as inevitable harmful by-products of aerobic metabolism. Growing evidence, however, suggests that ROS play important physiological roles. This raises questions about the pathways that different groups of organisms use to produce and sense ROS. In microbial eukaryotes, recent data show (i) increased ROS levels during cell differentiation, (ii) the existence of ROS-producing enzymes, such as NADPH oxidases (NOX), (iii) the involvement of NOX in developmental processes, and (iv) a conservation in the signal-transduction mechanisms used to detect ROS. This shows that manipulation of reactive species, as strategy to regulate cell differentiation, is ubiquitous in eukaryotes and suggests that such strategy was selected early in evolution.     

Characteristics of in vivo murine erythropoietic response to sodium orthovanadate

Current knowledge about the effects of vanadium compounds on erythropoiesis is still reduced and even contradictory. The aim of this work was to evaluate the in vivo effects of a single dose of sodium orthovanadate (OV, 33 mg/kg i.p.) on CF-1 mice in a time course study (0-8 days). Murine erythropoiesis was assessed through a combinatory of experimental approaches. Classical peripheral and bone marrow (BM) hematological parameters were determined. Erythroid maturation in blood stream and hemopoietic tissues (59Fe uptake assays), BM erythroid progenitor frequency (clonogenic assays) and erythroid crucial protein expressions for commitment and survival: GATA-1, erythropoietin receptor (Epo-R) and Bcl-xL (immunoblottings) were evaluated. Neither BM cellularities nor BM viabilities changed noticeably during the study. Peripheral reticulocytes showed a biphasic increment on days 2 and 8 post-OV. hematocrits enhanced transiently between days 2 and 4. 59Fe uptake percentages enhanced in peripheral blood nearly two-fold over control values between 4 and 8 days (p<0.01) without changes in BM and spleen. Additionally, mature erythroid BM compartments: polychromatophilic erythroblasts and orthochromatic normoblasts increased by the eighth day. BFU-E colonies remained near basal values during the whole experience, whilst CFU-E colonies raised 60% over control at 8 days post-OV (p<0.05). GATA-1 and Epo-R were significantly over-expressed from the third until the end of the experimental protocol (p<0.01). Surprisingly, Bcl-xL showed a constitutive expression pattern without changes during the experience. Experimental data let us suggest that OV does not to cause bone marrow cytotoxicity and that it accelerates maturation of BM committed erythroid precursors. Moreover, there are significant correlations among erythroid-related protein expressions: GATA-1 and Epo-R and the frequency of CFU-E. In addition, Bcl-xL expression invariance during the time course study would indicate that the stimulatory effect of OV treatment on erythropoiesis was mainly exerted on the maturation of red cell precursors rather than on the antiapoptosis of erythroid terminal progenitors.     

Correlation between molecular markers and adaptively significant genetic variation in Bromus tectorum (Poaceae), an inbreedingannual grass

Single sequence repeat (SSR) and amplified fragment length polymorphic (AFLP) molecular marker genotypes in cheatgrass (Bromus tectorum) were compared to published data on phenotypic variation in seed dormancy, vernalization requirement, and resistance to the pathogen Ustilago bullata. Several features of cheatgrass facilitated this study: it is a recent invader in the western United States, has considerable phenotypic polymorphism, and is an obligate self-pollinator. Forty self-pollinating lines from four populations common to the three phenotypic data sets were analyzed for molecular genetic variation using seven SSR loci and 31 AFLP loci. We examined correlations between distance matrices using the Mantel test for each pair of studies. The two molecular data sets were significantly correlated (r = 0.636). The AFLP markers often distinguished among several lines with identical SSR genotypes. The AFLP data were also significantly correlated with the phenotypic data (r values from 0.4640 to 0.5658), but the SSR data were much more highly correlated (r values from 0.677 to 0.844). The difference between molecular marker systems was especially notable when an outlier population from Potosi Pass, Nevada, was excluded from the analysis. These results suggest that SSR markers may be good surrogates for phenotypic traits in population genetic studies of strongly inbreeding species such as cheatgrass.     

A synthetic peptide homologous to functional domain of human IL-10 down-regulates expression of MHC class I and Transporter associated with Antigen Processing 1/2 in human melanoma cells

Tumor cells treated with IL-10 were shown to have decreased, but peptide-inducible expression of MHC class I, decreased sensitivity to MHC class I-restricted CTL, and increased NK sensitivity. These findings could be explained, at least partially, by a down-regulation of TAP1/TAP2 expression. In this study, IT9302, a nanomeric peptide (AYMTMKIRN), homologous to the C-terminal of the human IL-10 sequence, was demonstrated to mimic these previously described IL-10 effects on MHC class I-related molecules and functions. We observed a dose-dependent down-regulation of MHC class I at the cell surface of melanoma cells after 24-h treatment with IT9302. The IL-10 homologue peptide also caused a dose-dependent inhibition of the IFN-gamma-mediated surface induction of MHC class I in a melanoma cell line. We demonstrated, using Western blot and flow cytometry, that IT9302 inhibits the expression of TAP1 and TAP2 proteins, but not MHC class I H chain or low molecular protein molecules. Finally, peptide-treated melanoma cells were shown to be more sensitive to lysis by NK cells in a dose-dependent way. Taken together, these results demonstrate that a small synthetic peptide derived from IL-10 can mimic the Ag presentation-related effects mediated by this cytokine in human melanomas and increase tumor sensitivity to NK cells, which can be relevant in the designing of future strategies for cancer immune therapy.     

Gel-free mass spectrometry-based high throughput proteomics: tools for studying biological response of proteins and proteomes

Revolutionary advances in biological mass spectrometry (MS) have provided a basic tool to make possible comprehensive proteomic analysis. Traditionally, two-dimensional gel electrophoresis has been used as a separation method coupled with MS to facilitate analysis of complex protein mixtures. Despite the utility of this method, the many challenges of comprehensive proteomic analysis has motivated the development of gel-free MS-based strategies to obtain information not accessible using two-dimensional gel separations. These advanced strategies have enabled researchers to dig deeper into complex proteomes, gaining insights into the composition, quantitative response, covalent modifications and macromolecular interactions of proteins that collectively drive cellular function. This review describes the current state of gel-free, high throughput proteomic strategies using MS, including (i) the separation approaches commonly used for complex mixture analysis; (ii) strategies for large-scale quantitative analysis; (iii) analysis of post-translational modifications; and (iv) recent advances and future directions. The use of these strategies to make new discoveries at the proteome level into the effects of disease or other cellular perturbations is discussed in a variety of contexts, providing information on the potential of these tools in electromagnetic field research.     

Differential effects of strength training leading to failure versus not to failure on hormonal responses, strength, and muscle power gains.

The purpose of this study was to examine the efficacy of 11 wk of resistance training to failure vs. nonfailure, followed by an identical 5-wk peaking period of maximal strength and power training for both groups as well as to examine the underlying physiological changes in basal circulating anabolic and catabolic hormones. Forty-two physically active men were matched and then randomly assigned to either a training to failure (RF; n = 14), nonfailure (NRF; n = 15), or control groups (C; n = 13). Muscular and power testing and blood draws to determine basal hormonal concentrations were conducted before the initiation of training (T0), after 6 wk of training (T1), after 11 wk of training (T2), and after 16 wk of training (T3). Both RF and NRF resulted in similar gains in 1-repetition maximum bench press (23 and 23%) and parallel squat (22 and 23%), muscle power output of the arm (27 and 28%) and leg extensor muscles (26 and 29%), and maximal number of repetitions performed during parallel squat (66 and 69%). RF group experienced larger gains in the maximal number of repetitions performed during the bench press. The peaking phase (T2 to T3) after NRF resulted in larger gains in muscle power output of the lower extremities, whereas after RF it resulted in larger gains in the maximal number of repetitions performed during the bench press. Strength training leading to RF resulted in reductions in resting concentrations of IGF-1 and elevations in IGFBP-3, whereas NRF resulted in reduced resting cortisol concentrations and an elevation in resting serum total testosterone concentration. This investigation demonstrated a potential beneficial stimulus of NRF for improving strength and power, especially during the subsequent peaking training period, whereas performing sets to failure resulted in greater gains in local muscular endurance. Elevation in IGFBP-3 after resistance training may have been compensatory to accommodate the reduction in IGF-1 to preserve IGF availability.     

Quantitative comparison of bacterial communities in two Mediterranean sponges

Marine sponges can host in their tissues abundant and diverse bacterial communities. Lack of truly quantitative data on bacterial abundance and dynamics limits our understanding of the organization and functioning of these endobiotic communities. In this technical note, we describe a quantitative polymerase chain reaction approach to quantify the relative abundance of multiple clades of three major sponge-associated bacterial phyla: Chloroflexi, Acidobacteria, and Actinobacteria. To test our approach we used the Mediterranean sponges Spongia lamella and Aplysina aerophoba. We designed five out of the six primer sets used in our study. We tested the new primer sets for specificity and optimized their conditions. Our preliminary data showed that Spongia lamella had larger bacterial abundance than Aplysina aerophoba, except for one clade of Chloroflexi. The two Chloroflexi clades investigated in our study amplified a fraction of the Chloroflexi present in Spongia lamella and most of what is present in Aplysina aerophoba, suggesting a more diverse Chloroflexi population in Spongia lamella than in Aplysina aerophoba. This quantitative technique has a great potential to provide a rapid and robust assessment of sponge microbial target and could contribute to deciphering the complexity of these largely unknown host-symbiont interactions.