Who attempts suicide among medical students?

OBJECTIVE: To identify factors associated with a history of suicide attempt in medical students. METHODS: A Web-based survey was sent out to a sample of medical students. A multi-predictor Poisson regression was performed to identify factors associated with a history of suicide attempt. In addition, an elastic net regularization was used to build a risk calculator to identify students at risk for attempted suicide. RESULTS: A total of 4,840 participants were included in the study. Prevalence of suicide attempts in the sample was 8.94%. Risk factors associated with past suicide attempt in the multi-predictor Poisson regression were as follows: female gender (P < 0.001); homosexuality (P < 0.001); low income (P = 0.026); bullying by university peers (P = 0.006); childhood (P = 0.001) or adult (P = 0.001) trauma; family history of suicide (P = 0.005); suicidal ideation within the last month (P < 0.001); daily tobacco use (P = 0.037); and being at severe risk for alcohol abuse (P = 0.023). Our elastic net model performed well with an AUC of 0.83. CONCLUSIONS: This study identifies a number of key factors associated with a history of suicide attempts among medical students. Future longitudinal studies should assess the causal relationship between these factors and suicide attempts. Additionally, these results demonstrate that current available data on suicide attempts among medical students can be used to develop an accurate risk algorithm.

Heterologous expression of an alternative oxidase from Moniliophthora perniciosa in Saccharomyces cerevisiae: Antioxidant function and in vivo platform for the study of new drugs against witches' broom disease.

The fungus Moniliophthora perniciosa is the causal agent of witches' broom disease (WBD), one of the most devastating diseases of cacao, the chocolate tree. Many strategies to control WBD have been tested so far, including the use of agrochemicals such as the strobilurins. Strobilurins are fungicides of the QoI family, and they are used in the control of a wide array of fungal diseases in many different crops, including cereals, field crops, fruits, tree nuts, and vegetables. These drugs act by specifically inhibiting fungal respiration at the Qo site of complex III, which is a component of the main mitochondrial respiratory chain. However, M. perniciosa is resistant to this family of chemicals. It has been postulated that this resistant phenotype is, at least in part, a result of the strong ability of this fungus to counteract the oxidative stress generated by the impairment of the main mitochondrial respiratory chain, through the activation of an alternative oxidase (Mp-AOX). To test this hypothesis, we expressed functional mitochondria-localized Mp-AOX in the model yeast Saccharomyces cerevisiae. We demonstrated that heterologous expression of Mp-AOX strongly inhibits hydrogen peroxide production by mitochondria. It also diminishes the total cell amount of oxidized glutathione (GSSG), resulting in a fifty-fold higher GSH/GSSG ratio in cells expressing Mp-AOX than in wild type cells. In addition, Mp-AOX activity decreases yeast growth rate and leads to low biomass production. Therefore, we propose the use of this heterologous expression system to direct the development of new inhibitors of fungal AOX by comparing the differences in optical density of Mp-AOX-expressing cells in the presence and absence of potential AOX inhibitors. Together, our results confirm the antioxidant role of Mp-AOX and provide an in vivo platform to be used in the screening of new fungicides based on Mp-AOX inhibition.

Feed can be a source of Campylobacter jejuni infection in broilers.

1. The aim was to determine the importance of a contaminated diet as a possible cause of Campylobacter jejuni infection in broilers. 2. This study evaluated the viability of C. jejuni in both starter and finisher diets and the interference from other mesophilic bacteria in this viability. 3. Starter and finisher samples of broiler diet were deliberately contaminated with 3 or 5 log CFU·g-1 of C. jejuni (NCTC 11351) and then maintained at two different storage temperatures (25°C or 37°C) for 3 or 5 d. 4. C. jejuni survived during this period and, when inoculated at 103 CFU·g-1, multiplied with greater proliferation at a storage temperature of 37°C. There was no relationship between the amount of mesophilic bacteria and C. jejuni viability. 5. This study highlights the importance of the diet in the epidemiology of C. jejuni in broilers.

Assessing the effect of sodium dichloroisocyanurate concentration on transfer of Salmonella enterica serotype Typhimurium in wash water for production of minimally processed iceberg lettuce (Lactuca sativa L.).

UNLABELLED: This study evaluated the impact of sodium dichloroisocyanurate (5, 10, 20, 30, 40, 50 and 250 mg l(-1) ) in wash water on transfer of Salmonella Typhimurium from contaminated lettuce to wash water and then to other noncontaminated lettuces washed sequentially in the same water. Experiments were designed mimicking the conditions commonly seen in minimally processed vegetable (MPV) processing plants in Brazil. The scenarios were as follows: (1) Washing one inoculated lettuce portion in nonchlorinated water, followed by washing 10 noninoculated portions sequentially. (2) Washing one inoculated lettuce portion in chlorinated water followed by washing five noninoculated portions sequentially. (3) Washing five inoculated lettuce portions in chlorinated water sequentially, followed by washing five noninoculated portions sequentially. (4) Washing five noninoculated lettuce portions in chlorinated water sequentially, followed by washing five inoculated portions sequentially and then by washing five noninoculated portions sequentially in the same water. Salm. Typhimurium transfer from inoculated lettuce to wash water and further dissemination to noninoculated lettuces occurred when nonchlorinated water was used (scenario 1). When chlorinated water was used (scenarios 2, 3 and 4), no measurable Salm. Typhimurium transfer occurred if the sanitizer was ≥10 mg l(-1) . Use of sanitizers in correct concentrations is important to minimize the risk of microbial transfer during MPV washing. SIGNIFICANCE AND IMPACT OF THE STUDY: In this study, the impact of sodium dichloroisocyanurate in the wash water on transfer of Salmonella Typhimurium from inoculated lettuce to wash water and then to other noninoculated lettuces washed sequentially in the same water was evaluated. The use of chlorinated water, at concentration above 10 mg l(-1) , effectively prevented Salm. Typhimurium transfer under several different washing scenarios. Conversely, when nonchlorinated water was used, Salm. Typhimurium transfer occurred in up to at least 10 noninoculated batches of lettuce washed sequentially in the same water.

The gene ICS3 from the yeast Saccharomyces cerevisiae is involved in copper homeostasis dependent on extracellular pH.

In the yeast Saccharomyces cerevisiae, many genes are involved in the uptake, transport, storage and detoxification of copper. Large scale studies have noted that deletion of the gene ICS3 increases sensitivity to copper, Sortin 2 and acid exposure. Here, we report a study on the Δics3 strain, in which ICS3 is related to copper homeostasis, affecting the intracellular accumulation of this metal. This strain is sensitive to hydrogen peroxide and copper exposure, but not to other tested transition metals. At pH 6.0, the Δics3 strain accumulates a larger amount of intracellular copper than the wild-type strain, explaining the sensitivity to oxidants in this condition. Unexpectedly, sensitivity to copper exposure only occurs in acidic conditions. This can be explained by the fact that the exposure of Δics3 cells to high copper concentrations at pH 4.0 results in over-accumulation of copper and iron. Moreover, the expression of ICS3 increases in acidic pH, and this is correlated with CCC2 gene expression, since both genes are regulated by Rim101 from the pH regulon. CCC2 is also upregulated in Δics3 in acidic pH. Together, these data indicate that ICS3 is involved in copper homeostasis and is dependent on extracellular pH.

The use of alpha-1 adrenergic blockers in children with distal ureterolithiasis: a systematic review and meta-analysis.

INTRODUCTION: Urinary lithiasis is the main urologic cause of emergency treatment in adult patient. In the past years, the incidence in children population has increased. However, literature about the use of alpha-1 adrenergic blockers in pediatric population with distal ureterolithiasis is still scarce. The drug acts by decreasing ureter contractions, especially in the distal portion, facilitating calculus expulsion. OBJECTIVE: This review has the objective to evaluate the use of alpha-1 adrenergic blockers as medical expulsive treatment in children with distal ureterolithiasis. EVIDENCE ACQUISITION: An electronic literature search was performed using the MEDLINE, COCHRANE, and LILACS databases. We further searched manually the references of the primary studies. Searches were concluded on October 4th, 2014. Articles were selected, independently and in pairs, by the respective titles and summaries. Any divergence was resolved by consensus. EVIDENCE SYNTHESIS: Alpha-1 adrenergic antagonists increased the probability of calculus expulsion by 27% (NNT=4). Calculi smaller than 5mm, increased by 33% (NNT=3). Larger than 5mm, increased by 34% (NNT=3). CONCLUSION: Alpha-1 adrenergic blocker use is related with a greater incidence of expulsion of ureteral calculi, smaller or greater than 5mm, and fewer episodes of pain when compared to ibuprofen. However it is necessary larger samples to enhance the power analysis of the expulsion of ureteral calculi larger than 5mm and the episodes of pain. PATIENT SUMMARY: This review analyzed the outcome of alpha adrenergic antagonist in children with ureteral calculi. We conclude that it is the best medicine for use, since it helps the expulsion of the stone.

Biological development of Saccharicoccus sacchari () (Hemiptera: Pseudococcidae) on sugarcane in different temperatures.

The pink sugarcane mealybug Saccharicoccus sacchari (Cockerell, 1895) (Hemiptera: Pseudococcidae) occurs practically in all sugarcane producing regions Saccharum spp. (Poaceae) causing damage. Little information is available about insect's biology under Brazilian conditions. In this work, biological development of pink sugarcane mealybug was studied at temperatures of 23 °C ± 2 °C and 28 °C ± 2 °C without photophase and relative humidity of 80%. Number and viability of eggs, incubation time, duration of the last oviposition, duration of each nymphal instar, viability of the nymphs, start of oviposition and longevity of the females were recorded. Biological development of insects was stipulated by the SAS University Edition software, version 9.4. There were differences in the life cycle of the pseudococcid at both temperatures evaluated. Females of S. sacchari had three nymphal instars and reproduce asexually. Asexual reproduction occurs in the field and under controlled conditions. By increasing the temperature increases, insect lived longer and the presence of the winged male in Brazil indicates the possibility of sexual reproduction of the species.

The implementation of portable air-cleaning technologies in healthcare settings - a scoping review.

The COVID-19 pandemic revealed opportunities to improve prevention practices in healthcare settings, mainly related to the spread of airborne microbes (also known as bioaerosols). This scoping review aimed to map methodologies used to assess the implementation of portable air cleaners in healthcare settings, identify gaps, and propose recommendations for future research. The protocol was registered in the Open Science Framework and reported following the checklist provided by the Preferred Reporting Items for Systematic Reviews and Meta-Analysis - an extension for Scoping Reviews (PRISMA-ScR) statement. The search strategy was performed in five databases and one grey literature source. At the last selection phase, 24 articles that fulfilled our inclusion criteria were summarized and disseminated. Of these, 17 studies were conducted between 2020 and 2022; one of them was a protocol of a multicentre randomized controlled trial. The outcomes measured among the studies include airborne microbe counts, airborne particle concentrations, and rate of infections/interventions. The leading healthcare settings assessed were dental clinics (28%), patient's wards (16%), operating rooms (16%), and intensive care units (12%). Most of the devices demonstrated a significant potential to mitigate the impact of bioaerosols. Although some indoor air quality parameters can influence the mechanics of aerosols, only a few studies controlled these parameters in their analyses. Future clinical research should assess the rate of infections through randomized controlled trials with long-term follow-up and large sample sizes to determine the clinical importance of the findings.

Nitrogen metabolism in maize plants submitted to drought, brassinosteroids and azospirillum.

The water deficit in particular, reduces the productivity of vegetable crops. To minimize these harmful effects on agriculture, several agronomic and physiological practices are being studied, such as the use of bacteria and water stress attenuators, such as brassinosteroids. Considering the socioeconomic relevance of corn culture and its sensitivity when exposed to water deficit, the objective of the present study was to evaluate the action of brassinosteroids and azospirillum on nitrogen metabolism in corn plants subjected to water stress conditions. The experiment was carried out in a greenhouse, in a period of 47 days, with corn plants, using the hybrid K9606 VIP3. The design was completely randomized, in a 2x2x3 factorial scheme, with six replications. The first factor corresponds to two water regimes (presence and absence of water deficit). The second corresponds to inoculation via seed of Azospirillum brasiliense and absence of inoculation. And the third corresponds to the application of three concentrations of brassinosteroids (0, 0.3 and 0.6 µM). Were determined Nitrate; nitrate reductase; free ammonium; total soluble aminoacids; soluble proteins; proline; glycine betaine and glutamine synthetase. The lack of water in plants provided a reduction in the protein and nitrate reductase contents, in leaves and roots. For ammonium, plants with water deficit inoculated at a concentration of 0.3 µM, obtained an increase of 7.16 (70.26%) and 13.89 (77.04%) mmol NH4 + .Kg-1. DM (Dry mass) on the leaf and root respectively. The water deficit in the soil provided significant increases in the concentrations of glycine betaine, nitrate, proline and aminoacids, both in the leaves and in the roots of the corn plants. On the other hand, the contents of glutamine synthetase had a reduction in both leaves and roots.

Does silicon attenuate PEG 6000-induced water deficit in germination and growth initial the seedlings corn.

Water stress limits the initial growth and development of maize mass and grain, as well as the physiological process for absorbing the amount of mineral elements. The objective was to evaluate the effect of silicon on germination and growth of corn seedlings submitted to water deficit. The experiment was carried out in the laboratory and the experimental design was completely randomized (factorial 3 × 4), with three concentrations of calcium silicate (0.0; 1.0 and 2.0 mM) and 4 solutions of PEG-6000 to simulate different osmotic potentials (0, 0; -0.3; -0.6; -0.9 MPa). Germination percentage, germination speed index (GSI), mean germination time (MGT), percentage of non-germinated and abnormal germinated, length and dry matter of shoot, root and total seedlings were evaluated. Water deficiency reduced the parameters TG, GSI and MGT. The water deficit reduce the MSPA, MSR and MST with more than 80% reduction in mass from seedlings without deficiency to seedlings with deficiency. For CPA, CR and CT there was a reduction of at least 87%, 70% and 77%, respectively, among seeds without deficiency compared to seeds submitted to deficiency. The use of silicon in corn seeds did not attenuate the stress caused by water deficit simulated by PEG-6000.

Evaluation of sperm DNA peroxidation in fertile and subfertile dogs.

Oxidative stress (OS) has been recognized as one of the most important causes of male infertility. The antioxidant activities of seminal plasma and epididymal fluid are not enough to prevent OS, which can damage sperm membranes and DNA, so antioxidant supplementation has been used as a treatment of male infertility. The aim of this experiment was to evaluate the DNA peroxidation before and after antioxidant supplementation with vitamin C and E in dogs with and without fertility problems. A total of eleven dogs were used and were divided in two groups: fertile group (G1), dogs with normal spermiogram (n = 5); subfertile group (G2): dogs with low sperm count (<20 × 10(6) sptz/ml) and/or more than 30% of total sperm pathology (n = 6). Both groups received 500 mg/day of vitamin C and 500 mg/day of vitamin E for 60 days. A semen sample was collected before (M1) and after (M2) oral supplementation. Samples were analysed for DNA peroxidation by measuring the 8-hydroxy-2'-deoxyguanosine concentration. No significant difference was observed between groups at either time. Oral supplementation with 500 mg/day of vitamin C and 500 mg/day of vitamin E did not change the DNA peroxidation in fertile and subfertile dogs.

The Influence of Cement Removal Techniques on In Situ Bacterial Adhesion and Biodegradation at the Marginal Interface of Ceramic Laminates.

OBJECTIVES: This in situ study aimed to analyze the influence of different resin cement removal techniques on bacterial adhesion and biodegradation at the marginal interface of ceramic laminates. METHODS AND MATERIALS: Eighty feldspathic ceramic (F) blocks were prepared and cemented onto bovine enamel slabs (7×2.5×2 mm). Excess cement was removed using a microbrush (MBR), a scalpel blade (SCP), or a Teflon spatula (TSP). For the biodegradation analysis, 40 disc-shaped resin cement specimens were prepared (7×1.5 mm) using a Teflon mold. The specimens were randomly allocated into two groups: (1) No finishing procedure (only Mylar strip), and (2) with finishing and polishing procedures using the Jiffy system (Ultradent, South Jordan, UT, USA) (n=20). The in situ phase consisted of using an intraoral palatal device by 20 volunteers for 7 days. Each device contained five cylindrical wells (8×3 mm), where three dental blocks and two cement specimens were included in the wells. Surface roughness (Ra) was measured using a contact profilometer. A micromorphological analysis was performed under a stereomicroscope and a scanning electron microscope. Bacterial adhesion was quantitated based on the number of colony-forming units (CFU/mL) and their biofilm development potential. RESULTS: The cement removal techniques directly affected surface roughness at the marginal interface (p<0.001), and the SCP technique produced higher mean roughness, regardless of the surface area analyzed. Surface polishing protected cement specimens from further biodegradation (p=0.148). There were no differences in CFU counts between the groups after the in situ phase (p=0.96). All specimens showed CFU with a strong ability to develop a biofilm. CONCLUSIONS: The techniques used for cement removal increased the surface roughness of ceramic laminates, particularly SCP, but they did not affect bacterial adhesion at the marginal interface. Surface polishing of the resin cement is recommended to mitigate biodegradation.

Virulence of entomopathogenic nematodes and their symbiotic bacteria, under laboratory conditions, aiming controlling Saccharicoccus sacchari (Cockerell, 1895) (Hemiptera: Pseudococcidae) on sugarcane.

Sugarcane crops Saccharum spp. (Poales: Poaceae) produces different derivatives to the world: sugar, ethanol and bioenergy. Despite the application of pesticides, insect pests still cause economic losses, among these the pink sugarcane mealybug Saccharicoccus sacchari (Cockerell, 1895) (Hemiptera: Pseudococcidae) causing direct and indirect damage to the plant. This study assess the virulence of three entomopathogenic nematodes (EPNs) species and their symbiont bacteria against the pink sugarcane mealybug, under laboratory conditions. Fourteen treatments represented by control (distilled water), Heterorhabditis bacteriophora Poinar, 1976 (HB EN01) (Rhabditida: Heterorhabditidae), Steinernema rarum (Doucet, 1986) (PAM25) and Steinernema carpocapsae Weiser, 1955 (All) (Rhabditida: Steinermatidae) at concentrations of 25, 50, 75 and 100 infective juveniles (IJs)/insect, and the standard chemical product, thiamethoxam, were assayed. In a second experiment, the bacteria Photorhabdus luminescens (Thomas and Poinar, 1979), Xenorhabdus szentirmaii Lengyel, 2005 and Xenorhabdus nematophila (Poinar and Thomas, 1965) (Enterobacterales: Morganellaceae) at 3.0 x 109 cells/ml were assessed for each treatment. Ten replications were stablished, each one counting ten females/mealybugs inside a 10 cm Petri dish, amounting 100 individuals/treatment. All treatments were kept under stable conditions (25±1 ºC, H 70±10%, in the dark). All nematodes species infected S. sacchari. Steinerma rarum (PAM25) provided the highest mortality against the pink sugarcane mealybug (79.25%), followed by H. bacteriophora (HB EN01) (58.25%) and S. carpocapsae (All) (42.50%) (P<0.001). The mortality rate caused by X. szentirmaii, P. luminescens and X. nematophila were 40, 45 and 20%, respectively. Steinerma rarum (PAM25) has conditions to be a potential agent to be incorporate into the integrated pest management in sugarcane.

Mitigation of the effects of salt stress in cowpea bean through the exogenous aplication of brassinosteroid.

Cowpea (Vigna unguiculata (L.) Walp.) is a legume widely cultivated by small, medium and large producers in several Brazilian regions. However, one of the concerns for the production of cowpea in Brazil in recent years is the low rainfall activity in these regions, which generates the accumulation of salts on the surface. The objective of this work was to evaluate the effects of salt stress on growth parameters and enzyme activity in cowpea plants at different concentrations of brassinosteroids. Experiment was developed in a greenhouse using a completely randomized experimental design in a 3 x 3 factorial scheme. The treatments consisted of three levels of brassinosteroids (0, 3 and 6 µM EBL) and three levels of salt stress (0, 50 and 100 mM NaCl). Growth factors (height, diameter and number of leaves) decreased in the saline condition. With the presence of brassinosteroid the height did not increase, but the number of leaves did, mainly in the saline dosage of 100 mM NaCl. In the variable membrane integrity, brassinosteroid was efficient in both salinity dosages, the same not happening with the relative water content, where the saline condition did not affect the amount of water in the vegetable, with the application of brassino it remained high, decreasing only at dosage 100 mM NaCl. The nitrate reductase enzyme was greatly affected in the root system even with the application of increasing doses of brassino. Therefore, brassinosteroids as a promoter of saline tolerance in cowpea seedlings was positive. The concentration of 3µM of EBL provided the most satisfactory effect in tolerating the deleterious effects of the saline condition. The same cannot be concluded for the concentration of 6µM of EBL that did not promote tolerance to some variables.

Chagas disease in Italy: breaking an epidemiological silence.

Chagas disease, a neglected tropical disease that due to population movements is no longer limited to Latin America, threatens a wide spectrum of people(travellers, migrants, blood or organ recipients,newborns, adoptees) also in non-endemic countries where it is generally underdiagnosed. In Italy, the available epidemiological data about Chagas disease have been very limited up to now, although the country is second in Europe only to Spain in the number of residents from Latin American. Among 867 at-risk subjectsscreened between 1998 and 2010, the Centre for Tropical Diseases in Negrar (Verona) and the Infectious and Tropical Diseases Unit, University of Florence found 4.2% patients with positive serology for Chagas disease (83.4% of them migrants, 13.8% adoptees).No cases of Chagas disease were identified in blood donors or HIV-positive patients of Latin American origin. Among 214 Latin American pregnant women,three were infected (resulting in abortion in one case).In 2005 a case of acute Chagas disease was recorded in an Italian traveller. Based on our observations, we believe that a wider assessment of the epidemiological situation is urgently required in our country and public health measures preventing transmission and improving access to diagnosis and treatment should be implemented.

Distribution, abundance and seasonality of scale insects in sugarcane crops in the state of São Paulo.

In the state of São Paulo, the main sugarcane producing region of the world, two species of scale insects have frequently occurred, Aclerda takahashii (Kuwana, 1932) (Hemiptera: Aclerdidae) and Saccharicoccus sacchari (Cockerell, 1895) (Hemiptera: Pseudococcidae). To map the distribution and abundance of these species, 17 sugarcane producing fields, distributed in six mesoregions in São Paulo, were evaluated in August 2017 and, January, February, June and July 2018 during the ripening phase. The study on the seasonality of these species, by the presence or absence of the scale insects during the phenological cycle of the plant, was conducted between August 2017 and July 2018 in two sugarcane producing fields in the municipality of Jaboticabal, São Paulo, Brazil. The presence of S. sacchari was found in all the analyzed locations, and A. takahashii in twelve. Both scale insects showed significant difference of infestation in the node's region of the stems during the ripening phase in one of the studied locations. The aclerdid presented significant difference by infestation in one site during the ripening phase of the plant. The pseudococcid infested a greater number of nodes in the following phases of development; vegetative, grand growth and ripening in both studied areas, but it was in one site during the ripening phase that presented the greatest difference. Although the pink sugarcane mealybug was more abundant than A. takahashii in both studies, there were no patterns of relationships between the numbers of individuals to geographical locations and temperature.

Encephalopathy and encephalitis during acute SARS-CoV-2 infection. Spanish Society of Neurology COVID-19 Registry. / Encefalopatías y encefalitis durante la infección aguda por SARS-CoV2. Registro de la Sociedad Española de Neurología SEN COVID-19.

OBJECTIVES: Since the beginning of the COVID-19 pandemic, the Spanish Society of Neurology has run a registry of patients with neurological involvement for the purpose of informing clinical neurologists. Encephalopathy and encephalitis were among the most frequently reported complications. In this study, we analyse the characteristics of these complications. PATIENTS AND METHODS: We conducted a retrospective, descriptive, observational, multicentre study of patients with symptoms compatible with encephalitis or encephalopathy, entered in the Spanish Society of Neurology's COVID-19 Registry from 17 March to 6 June 2020. RESULTS: A total of 232 patients with neurological symptoms were registered, including 51 cases of encephalopathy or encephalitis (21.9%). None of these patients were healthcare professionals. The most frequent syndromes were mild or moderate confusion (33%) and severe encephalopathy or coma (9.8%). The mean time between onset of infection and onset of neurological symptoms was 8.02 days. Lumbar puncture was performed in 60.8% of patients, with positive PCR results for SARS-CoV-2 in only one case. Brain MRI studies were performed in 47% of patients, with alterations detected in 7.8% of these. EEG studies were performed in 41.3% of cases, detecting alterations in 61.9%. CONCLUSIONS: Encephalopathy and encephalitis are among the complications most frequently reported in the registry. More than one-third of patients presented mild or moderate confusional syndrome. The mean time from onset of infection to onset of neurological symptoms was 8 days (up to 24hours earlier in women than in men). EEG was the most sensitive test in these patients, with very few cases presenting alterations in neuroimaging studies. All patients treated with boluses of corticosteroids or immunoglobulins progressed favourably.

Quantitation of non-amplified genomic DNA by bead-based hybridization and template mediated extension coupled to alkaline phosphatase signal amplification.

Klenow I polymerase activity was combined with solid phase DNA hybridization to detect non-amplified genomic DNA (gDNA) sequences from Escherichia coli. Aminopropyl-controlled pore glass surface-bound oligonucleotides were hybridized to fragmented gDNA. The template-mediated extension at the 3'-terminus of the immobilized probe was then promoted in the presence of Klenow I polymerase and digoxigenin-labeled nucleotides. Detection of the extended probes was accomplished with an anti-digoxigenin alkaline phosphatase conjugate protocol coupled to colorimetric or fluorescent detection. Using the colorimetric protocol, the proof-of-concept was established. The fluorescence-based methodology, on the other hand, provided the basis for a quantitative interpretation of the data, affording a detection limit of 5 pM gDNA.

Critical overview of the main features and techniques used for the evaluation of the clinical applicability of L-asparaginase as a biopharmaceutical to treat blood cancer.

L-asparaginase is an enzyme used as a biopharmaceutical to treat acute lymphoblastic leukemia. Several adverse effects have been related to L-asparaginase use, so the scientific community has searched for novel proteoforms of L-ASNase. However, some critical characteristics must be considered for a novel L-ASNase source to be effective as an antitumour drug. Accordingly, this article provides a critical analysis of the parameters and methods applied to estimate L-ASNase concentration, measure the L-ASNase and GLNase activities and kinetics, evaluate the enzyme preparations purity and define the antitumour activity against leukemic cells in vitro. Among the main features, the proposed new enzyme needs to present high affinity for L-asparagine; low percentage of glutaminase activity in relation to L-ASNase; high enzyme stability and half-life and mainly antileukemic activity when a low protein amount is applied. These parameters are discussed in an attempt to guide the consideration of an enzyme as a promising biopharmaceutical against ALL.

Equine seminal plasma and sperm membrane: Functional proteomic assessment.

During ejaculation, a large amount of seminal plasma proteins interact with the sperm membrane, leading to a series of biochemical and structural changes implicated in sperm function and gamete interaction. However, the roles of the majority of these proteins remain unknown. This study aimed to investigate the proteome and functionality of the major equine proteins of seminal plasma and the sperm membrane. Seminal plasma and enriched-membrane proteins (150 µg) were separated by two-dimensional gel electrophoresis, and the respective maps were analyzed. Protein identification was performed by in-gel digestion and tandem mass spectrometry (GeLC-MS/MS). Samples were also submitted to in-solution digestion (complex protein mixture) and identified by shotgun analysis by LC-MS/MS; bioinformatic tools were used to investigate protein functions. Seminal plasma and sperm membrane extract maps contained 91.0 ± 8.2 spots and 245.3 ± 11.3 spots, respectively, within the 3-10 pH range. In total, the most abundant proteins identified in 2D maps and in complex protein mixtures included 24 proteins for seminal plasma and 33 for sperm membrane extract, with a high degree of confidence (P < 0.05). Of these, HSP1, CRISP3 and KLK1E2 were the most abundant in seminal plasma; HSP1 was highly abundant in sperm membrane extract, in many isoforms, which is related to membrane destabilization and may compromise sperm preservation. HSP1-polybromo-1 interactions suggested a role in DNA stabilization. Prosaposin was identified in seminal plasma and may play a role in the fertilization process. IZUMO4, a member of the IgSF family involved in the prefertilization stages, was identified in 2D gel and MS/MS analysis of sperm membrane extract. Ten proteins of seminal plasma were found to interact with the sperm membrane and were related to binding and catalytic activities (clusterin, CRISP3, epididymal sperm-binding protein 1, kallikrein1E2, seminal plasma protein A3, and HSP1). Additionally, other identified proteins were associated with DNA integrity, capacitation and recognition of pregnancy. These findings indicate that the binding of specific proteins to the plasma membrane during ejaculation may influence sperm survival after cryopreservation and may play a role in decreasing the quality in stallions with toxic seminal plasma. Elucidation of these interactions is an important step in understanding the biological processes related to equine fertility and facilitates future investigations on the selection and application of low freezability semen strategies.