Endophytic and entomopathogenic strains of Beauveria sp to control the bovine tick Rhipicephalus (Boophilus) microplus

Pathogenicity of strains of the entomopathogenic fungus Beauveria bassiana and endophytic strains of Beauveria sp against the bovine tick Rhipicephalus (Boophilus) microplus was tested in laboratory bioassays and under field conditions. Suspensions containing 10(5), 10(7) and 10(9) conidia/mL were prepared of each fungal strain for laboratory bioassays. The ticks were maintained at 28 degrees C, 90 +/- 5% relative humidity, and the following variables were evaluated: initial female weight, egg weight, hatching percentage, reproductive efficiency, and percentage control. For tests under field conditions, a Beauveria suspension containing 10(6) conidia/mL was sprayed on tick-infested cows. After 72 h, the ticks were collected to estimate mortality under field conditions. Laboratory bioassays showed a mortality of 20 to 50% of the ticks seven days after inoculation with 10(7) Beauveria conidia/mL. Under field conditions 10(6) Beauveria conidia/mL induced 18-32% mortality. All Beauveria strains were effective in biological control of R. (Boophilus) microplus under laboratory and field test conditions. This is the first demonstration that endophytic fungi can be used for biological control of the cattle tick; this could help reduce environmental contamination by diminishing the need for chemical acaricides. Two endophytic strains were isolated from maize leaves and characterized by molecular sequencing of 5.8S rDNA ITS1 and ITS2 and morphological analyses of conidia. We found that these two endophytic Beauveria isolates, designated B95 and B157, are close to Beauveria amorpha.

Transcriptomic basis for an antiserum against Micrurus corallinus (coral snake) venom

Background: Micrurus corallinus (coral snake) is a tropical forest snake belonging to the family Elapidae. Its venom shows a high neurotoxicity associated with pre- and post-synaptic toxins, causing diaphragm paralysis, which may result in death. In spite of a relatively small incidence of accidents, serum therapy is crucial for those bitten. However, the adequate production of antiserum is hampered by the difficulty in obtaining sufficient amounts of venom from a small snake with demanding breeding conditions. In order to elucidate the molecular basis of this venom and to uncover possible immunogens for an antiserum, we generated expressed sequences tags (ESTs) from its venom glands and analyzed the transcriptomic profile. In addition, their immunogenicity was tested using DNA immunization. Results: A total of 1438 ESTs were generated and grouped into 611 clusters. Toxin transcripts represented 46% of the total ESTs. The two main toxin classes consisted of three-finger toxins (3FTx) (24%) and phospholipases A(2) (PLA(2)s) (15%). However, 8 other classes of toxins were present, including C-type lectins, natriuretic peptide precursors and even high-molecular mass components such as metalloproteases and L-amino acid oxidases. Each class included an assortment of isoforms, some showing evidence of alternative splicing and domain deletions. Five antigenic candidates were selected (four 3FTx and one PLA(2)) and used for a preliminary study of DNA immunization. The immunological response showed that the sera from the immunized animals were able to recognize the recombinant antigens. Conclusion: Besides an improvement in our knowledge of the composition of coral snake venoms, which are very poorly known when compared to Old World elapids, the expression profile suggests abundant and diversified components that may be used in future antiserum formulation. As recombinant production of venom antigens frequently fails due to complex disulfide arrangements, DNA immunization may be a viable alternative. In fact, the selected candidates provided an initial evidence of the feasibility of this approach, which is less costly and not dependent on the availability of the venom.

Detection of polymorphisms of the mtDNA control region of Caretta caretta (Testudines: Cheloniidae) by PCR-SSCP

Marine turtles are increasingly being threatened worldwide by anthropogenic activities. Better understanding of their life cycle, behavior and population structure is imperative for the design of adequate conservation strategies. The mtDNA control region is a fast-evolving matrilineal marker that has been employed in the study of marine turtle populations. We developed and tested a simple molecular tracing system for Caretta caretta mtDNA haplotypes by polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP). Using this technique, we were able to distinguish the SSCP patterns of 18 individuals of the haplotypes CC-A4, CC-A24 and CCxLO, which are commonly found in turtles sampled on the Brazilian coast. When we analyzed 15 turtles with previously unknown sequences, we detected two other haplotypes, in addition to the other four. Based on DNA sequencing, they were identified as the CC-A17 and CC-A1 haplotypes. Further analyses were made with the sea turtles, Chelonia mydas (N = 8), Lepidochelys olivacea (N = 3) and Eretmochelys imbricata (N = 1), demonstrating that the PCR-SSCP technique is able to distinguish intra-and interspecific variation in the family Cheloniidae. We found that this technique can be useful for identifying sea turtle mtDNA haplotypes, reducing the need for sequencing.

Heteroduplex formation and S1 digestion for mapping alternative splicing sites

The identification of alternatively spliced transcripts has contributed to a better comprehension of developmental mechanisms, tissue-specific physiological processes and human diseases. Polymerase chain reaction amplification of alternatively spliced variants commonly leads to the formation of heteroduplexes as a result of base pairing involving exons common between the two variants. S1 nuclease cleaves single-stranded loops of heteroduplexes and also nicks the opposite DNA strand. In order to establish a strategy for mapping alternative splice-prone sites in the whole transcriptome, we developed a method combining the formation of heteroduplexes between 2 distinct splicing variants and S1 nuclease digestion. For 20 consensuses identified here using this methodology, 5 revealed a conserved splice site after inspection of the cDNA alignment against the human genome (exact splice sites). For 8 other consensuses, conserved splice sites were mapped at 2 to 30 bp from the border, called proximal splice sites; for the other 7 consensuses, conserved splice sites were mapped at 40 to 800 bp, called distal splice sites. These latter cases showed a nonspecific activity of S1 nuclease in digesting double-strand DNA. From the 20 consensuses identified here, 5 were selected for reverse transcription-polymerase chain reaction validation, confirming the splice sites. These data showed the potential of the strategy in mapping splice sites. However, the lack of specificity of the S1 nuclease enzyme is a significant obstacle that impedes the use of this strategy in large-scale studies.

Alternative splicing enriched cDNA libraries identify breast cancer-associated transcripts

Background: Alternative splicing (AS) is a central mechanism in the generation of genomic complexity and is a major contributor to transcriptome and proteome diversity. Alterations of the splicing process can lead to deregulation of crucial cellular processes and have been associated with a large spectrum of human diseases. Cancer-associated transcripts are potential molecular markers and may contribute to the development of more accurate diagnostic and prognostic methods and also serve as therapeutic targets. Alternative splicing-enriched cDNA libraries have been used to explore the variability generated by alternative splicing. In this study, by combining the use of trapping heteroduplexes and RNA amplification, we developed a powerful approach that enables transcriptome-wide exploration of the AS repertoire for identifying AS variants associated with breast tumor cells modulated by ERBB2 (HER-2/neu) oncogene expression. Results: The human breast cell line (C5.2) and a pool of 5 ERBB2 over-expressing breast tumor samples were used independently for the construction of two AS-enriched libraries. In total, 2,048 partial cDNA sequences were obtained, revealing 214 alternative splicing sequence-enriched tags (ASSETs). A subset with 79 multiple exon ASSETs was compared to public databases and reported 138 different AS events. A high success rate of RT-PCR validation (94.5%) was obtained, and 2 novel AS events were identified. The influence of ERBB2-mediated expression on AS regulation was evaluated by capillary electrophoresis and probe-ligation approaches in two mammary cell lines (Hb4a and C5.2) expressing different levels of ERBB2. The relative expression balance between AS variants from 3 genes was differentially modulated by ERBB2 in this model system. Conclusions: In this study, we presented a method for exploring AS from any RNA source in a transcriptome-wide format, which can be directly easily adapted to next generation sequencers. We identified AS transcripts that were differently modulated by ERBB2-mediated expression and that can be tested as molecular markers for breast cancer. Such a methodology will be useful for completely deciphering the cancer cell transcriptome diversity resulting from AS and for finding more precise molecular markers.

Melatonin treatment decreases c-fos expression in a headache model induced by capsaicin

The aim of the present work was to analyze c-fos response within the trigeminal nucleus caudalis (TNC) of pinealectomized rats and animals that received intraperitoneal melatonin, after intracisternal infusion of capsaicin, used to induce intracranial trigeminovascular stimulation. Experimental groups consisted of animals that received vehicle solution (saline-ethanol-Tween 80, 8:1:1, diluted 1:50) only (VEI, n = 5); animals that received capsaicin solution (200 nM) only (CAP, n = 6); animals submitted to pinealectomy (PX, n = 5); sham-operated animals (SH, n = 5); animals submitted to pinealectomy followed by capsaicin stimulation (200 nM) after 15 days (PX + CAP, n = 7); and animals that received capsaicin solution (200 nM) and intraperitoneal melatonin (10 mg/kg) (CAP + MEL, n = 5). Control rats, receiving vehicle in the cisterna magna, showed a small number of c-fos-positive cells in the TNC (layer I/II) as well as the sham-operated and pinealectomized rats, when compared to animals stimulated by capsaicin. On the other hand, pinealectomized rats, which received capsaicin, presented the highest number of c-fos-positive cells. Animals receiving capsaicin and melatonin treatment had similar expression of the vehicle group. Our data provide experimental evidence to support the role of melatonin and pineal gland in the pathophysiology of neurovascular headaches.

Social Networks Shape the Transmission Dynamics of Hepatitis C Virus

Hepatitis C virus (HCV) infects 170 million people worldwide, and is a major public health problem in Brazil, where over 1% of the population may be infected and where multiple viral genotypes co-circulate. Chronically infected individuals are both the source of transmission to others and are at risk for HCV-related diseases, such as liver cancer and cirrhosis. Before the adoption of anti-HCV control measures in blood banks, this virus was mainly transmitted via blood transfusion. Today, needle sharing among injecting drug users is the most common form of HCV transmission. Of particular importance is that HCV prevalence is growing in non-risk groups. Since there is no vaccine against HCV, it is important to determine the factors that control viral transmission in order to develop more efficient control measures. However, despite the health costs associated with HCV, the factors that determine the spread of virus at the epidemiological scale are often poorly understood. Here, we sequenced partial NS5b gene sequences sampled from blood samples collected from 591 patients in Sao Paulo state, Brazil. We show that different viral genotypes entered Sao Paulo at different times, grew at different rates, and are associated with different age groups and risk behaviors. In particular, subtype 1b is older and grew more slowly than subtypes 1a and 3a, and is associated with multiple age classes. In contrast, subtypes 1a and 3b are associated with younger people infected more recently, possibly with higher rates of sexual transmission. The transmission dynamics of HCV in Sao Paulo therefore vary by subtype and are determined by a combination of age, risk exposure and underlying social network. We conclude that social factors may play a key role in determining the rate and pattern of HCV spread, and should influence future intervention policies.

Naturally-acquired humoral immune responses against the N- and C-termini of the Plasmodium vivax MSP1 protein in endemic regions of Brazil and Papua New Guinea using a multiplex assay

Background: Progress towards the development of a malaria vaccine against Plasmodium vivax, the most widely distributed human malaria parasite, will require a better understanding of the immune responses that confer clinical protection to patients in regions where malaria is endemic. Methods: Glutathione S-transferase (GST) and GST-fusion proteins representing the N-terminus of the merozoite surface protein 1 of P. vivax, PvMSP1-N, and the C-terminus, PvMSP1-C, were covalently coupled to BioPlex carboxylated beads. Recombinant proteins and coupled beads were used, respectively, in ELISA and Bioplex assays using immune sera of P. vivax patients from Brazil and PNG to determine IgG and subclass responses. Concordances between the two methods in the seropositivity responses were evaluated using the Kappa statistic and the Spearman's rank correlation. Results: The results using this methodology were compared with the classical microtitre enzyme-linked immnosorbent assay ( ELISA), showing that the assay was sensitive, reproducible and had good concordance with ELISA; yet, further research into different statistical analyses seems desirable before claiming conclusive results exclusively based on multiplex assays. As expected, results demonstrated that PvMSP1 was immunogenic in natural infections of patients from different endemic regions of Brazil and Papua New Guinea ( PNG), and that age correlated only with antibodies against the C-terminus part of the molecule. Furthermore, the IgG subclass profiles were different in these endemic regions having IgG3 predominantly recognizing PvMSP1 in Brazil and IgG1 predominantly recognizing PvMSP1 in PNG. Conclusions: This study validates the use of the multiplex assay to measure naturally-acquired IgG antibodies against the merozoite surface protein 1 of P. vivax.

PAKs supplement improves immune status and body composition but not muscle strength in resistance trained individuals

Mixed formula supplements are very popular among recreational and professional weightlifters. They are usually known as PAKs and they are supposed to have a synergistic effect of their different nutrients. The purpose of this study was to determine the effects of chronic (4 weeks) PAKS supplementation in combination with strength training on body composition, immune status and performance measures in recreationally trained individuals with or without PAKs supplementation. Methods: Twelve male subjects (Placebo n = 6 and PAKs supplement n = 6) were recruited for this study. The body composition, one maximum strength repetition tests and immune status were assessed before and after 4 week supplementation. Our data showed that, 4 week PAK supplementation associated with strength exercise not was effective in change strength than compared with placebo group. However, we observed that, PAK supplement was able to improve immune status and reduced body composition when compared with placebo group. These results indicate that, a mixed formula supplement is able to improve immune status and body composition but not maximum strength in recreational strength trained subjects in a 4 weeks period.

First Direct Observation of Muon Antineutrino Disappearance

This Letter reports the first direct observation of muon antineutrino disappearance. The MINOS experiment has taken data with an accelerator beam optimized for (nu) over bar (mu) production, accumulating an exposure of 1.71 x 10(20) protons on target. In the Far Detector, 97 charged current (nu) over bar (mu) events are observed. The no-oscillation hypothesis predicts 156 events and is excluded at 6.3 sigma. The best fit to oscillation yields vertical bar Delta(m) over bar (2)vertical bar = [3.36(-0.40)(+0.46)(stat) +/- 0.06(sys)] x 10(-3) eV(2), sin(2)(2 (theta) over bar) = 0.86(-0.12)(+0.11)(stat) +/- 0.01(syst). The MINOS nu(mu) and (nu) over bar (mu) measurements are consistent at the 2.0% confidence level, assuming identical underlying oscillation parameters.

Energy of the first excited state of (43)Ar

Results of proton-proton-gamma coincidence measurements using the (36)S+(9)Be reaction revealed a gamma ray of 201.27 +/- 0.16 keV that most probably corresponds to the transition between the predicted 7/2(-) first excited state to the 5/2(-) ground state of (43)Ar.

(7,9,10)Be elastic scattering and total reaction cross sections on a (12)C target

Elastic scattering angular distributions for (7)Be, (9)Be, and (10)Be isotopes on (12)C target were measured at laboratory energies of 18.8, 26.0, and 23.2 MeV, respectively. The analysis was performed in terms of optical model potentials using Woods-Saxon and double-folding form factors. Also, continuum discretized coupled-channels calculations were performed for (7)Be and (9)Be + (12)C systems to infer the role of breakup in the elastic scattering. For the (10)Be + (12)C system, bound states coupled-channels calculations were considered. Moreover, total reaction cross sections were deduced from the elastic scattering analysis and compared with published data on other weakly and tightly bound projectiles elastically scattered on the (12)C target, as a function of energy.

Elastic scattering and total reaction cross sections for the (8)Li+(12)C system

The elastic-scattering angular distribution for (8)Li on (12)C has been measured at E(LAB) = 23.9 MeV with (8)Li radioactive nuclear beam produced by the Radioactive Ion Beams in Brazil facility. This angular distribution was analyzed in terms of optical-model with Woods-Saxon and double-folding Sao Paulo potential. The roles of the breakup and inelastic channels were also investigated with cluster folding and deformed potentials, respectively, through coupled-channels calculations. The angular distribution for the proton-transfer (12)C((8)Li, (9)Be)(11)B reaction was also measured at the same energy. The spectroscopic factor for the <(9)Be|(8)Li + p > bound system was obtained and compared with shell-model calculations and with other experimental values. Total reaction cross sections for the present system were also extracted from the elastic-scattering analysis. A systematic of the reduced reaction cross sections obtained from the present and published data on (6,7,8)Li isotopes on (12)C was performed as a function of energy.

Elastic scattering and total reaction cross sections for the (8)B, (7)Be, and (6)Li+(12)C systems

Angular distributions for the elastic scattering of (8)B, (7)Be, and (6)Li on a (12)C target have been measured at E(lab) = 25.8, 18.8, and 12.3 MeV, respectively. The analyses of these angular distributions have been performed in terms of the optical model using Woods-Saxon and double-folding type potentials. The effect of breakup in the elastic scattering of (8)B + (12)C is investigated by performing coupled-channels calculations with the continuum discretized coupled-channel method and cluster-model folding potentials. Total reaction cross sections were deduced from the elastic-scattering analysis and compared with published data on elastic scattering of other weakly and tightly bound projectiles on (12)C, as a function of energy. With the exception of (4)He and (16)O, the data can be described using a universal function for the reduced cross sections.

Microwave dielectric permittivity and photoluminescence of Eu(2)O(3) doped laser heated pedestal growth Ta(2)O(5) fibers

We report the microwave dielectric properties and photoluminescence of undoped and europium oxide doped Ta(2)O(5) fibers, grown by laser heated pedestal growth technique. The effects of Eu(2)O(3) doping (1-3 mol %) on the structural, optical, and dielectric properties were investigated. At a frequency of 5 GHz, the undoped material exhibits a dielectric permittivity of 21 and for Eu(2)O(3) doped Ta(2)O(5) samples it increases, reaching up to 36 for the highest doping concentration. Nevertheless, the dielectric losses maintain a very low value. For this wide band gap oxide, Eu(3+) optical activation was achieved and the emission is observed up to room temperature. Thus, the transparency and high permittivity make this material promising for electronic devices and microwave applications. (c) 2008 American Institute of Physics.