Olfactory response of Trichogramma pretiosum (Hymenoptera: Trichogrammatidae) to volatiles induced by transgenic maize.

Plants not only respond to herbivorous damage but adjust their defense system after egg deposition by pest insects. Thereby, parasitoids use oviposition-induced plant volatiles to locate their hosts. We investigated the olfactory behavioral responses of Trichogramma pretiosum Riley, 1879 (Hymenoptera: Trichogrammatidae) to volatile blends emitted by maize (Zea mays L.) with singular and stacked events after oviposition by Spodoptera frugiperda Smith, 1797 (Hymenoptera: Trichogrammatidae) moths. Additionally, we examined possible variations in gene expression and on oviposition-induced volatiles. We used a Y-tube olfactometer to test for the wasp responses to volatiles released by maize plants oviposited by S. frugiperda and not-oviposited plants. Using the real-time PCR technique (qRT-PCR), we analyzed the expression of lipoxygenase and three terpene synthases genes, which are enzymes involved in the synthesis of volatile compounds that attract parasitoids of S. frugiperda. Olfactometer tests showed that T. pretiosum is strongly attracted by volatiles from transgenic maize emitted by S. frugiperda oviposition (VTPRO 3, more than 75% individuals were attracted). The relative expression of genes TPS10, LOX e STC was higher in transgenic hybrids than in the conventional (isogenic line) hybrids. The GC-MS analysis revealed that some volatile compounds are released exclusively by transgenic maize. This study provides evidence that transgenic hybrids enhanced chemical cues under oviposition-induction and helped to increase T. pretiosum efficiency in S. frugiperda control. This finding shows that among the evaluated hybrids, genetically modified hybrids can improve the biological control programs, since they potentialize the egg parasitoid foraging, integrating pest management.

Virtual screening reveals a viral-like polymerase inhibitor that complexes with the DNA polymerase of Moniliophthora perniciosa.

The filamentous fungus Moniliophthora perniciosa is a basidiomycota that causes the witches' broom disease in cocoa trees (Theobroma cacao L.). The mitochondrial DNA polymerase of M. perniciosa (MpmitDNApol) is classified within the B family of DNA polymerases, which can be found in viruses and cellular organelles. Using virtual screening processes, accessing KEGG, PubChem, and ZINC databases, we selected the 27 best putative nucleoside viral-like polymerase inhibitors to test against MpmitDNApol. We used Autodock Vina to perform docking simulations of the selected molecules and to return energy values in several ligand conformations. Then, we used Pymol v1.7.4.4 to check the stereochemistry of chiral carbons, hydrogen bonding receptors, absence or presence of hydrogen, sub and superstructure, numbers of rings, rotatable bonds, and donor groups. We selected the Entecavir Hydrate, a drug used to control hepatitis B; subsequently AMBER 14 was used to describe the behavior of polymerase-entecavir complex after setting up 3500 ps of simulation in water at a temperature of 300 K. From the simulation, a graph of Potential Energy was generated revealing that the ligand remains in the catalytic site after 3500 ps with a final energy of -612,587.4214 kcal/mol.

Ocular experimental leishmaniasis in C57BL/10 and BALB/c mice induced by Leishmania amazonensis infection.

There are few studies on human ocular leishmaniasis found in the literature. The purpose of this study was to describe experimental ocular leishmaniasis, caused by Leishmania amazonensis evaluating two different infection routes: intravitreal and instillation in C57BL/10 and BALB/c mice. In this work all animals presented low anti-Leishmania IgM and IgG titers regardless of the infection route or mouse strain. The histopathological eye analysis showed that the mice inoculated by the intravitreal route developed more severe lesions, presenting parasites in the anterior region of the eye 60 days after infection. The C57BL/10 mice presented cells containing parasitophorous vacuoles associated with pigmented cells and inflammatory infiltrate, which included mast cells. Ninety days after infection no parasites could be found in either mouse strain, which led us to hypothesize that parasites had been eliminated. In this context, we show that both intravitreal and instillation routes were effective in promoting ocular leishmaniasis infections in C57BL/10 and BALB/c mice. There were no differences in the parasite infection between the two mouse models and it mimicked the ocular lesions described in symptomatic dogs in endemic areas of visceral leishmaniasis.

An antigenic domain of the Leishmania amazonensis nucleoside triphosphate diphosphohydrolase (NTPDase 1) is associated with disease progression in susceptible infected mice.

An antigenic conserved B domain was previously identified within nucleoside triphosphate diphosphohydrolases (NTPDases) of plants and parasites. Now, the r-potDomain B, a 6× His-tag polypeptide belonging to the conserved B domain from the potato apyrase, and synthetic peptides LbB1LJ and LbB2LJ derived from the B domain from Leishmania NTPDase 1 were used as molecular tools for studies of the Leishmania amazonensis NTPDase 1. Widespread subcellular location of the specific NTPDase 1 was detected by Western blots of promastigote fractions and ultrastructural immunocytochemical microscopy using immune sera raised against these biomolecules. In addition, the L. amazonensis-infected BALB/c mice were evaluated at 12 to 120 days after infection, which progresses showing typical nodular lesion. High antibody reactivity with either r-potDomain B, LbB1LJ, or LbB2LJ was found in L. amazonensis-infected BALB/c mice indicating the antigenicity of the B domain from NTPDase 1 isoform. The IgG1 antibody reactivity significantly increased at 90-120 days postinfection, 18- to 24-fold when compared to the 12th day, and remained elevated even at 120th after infection, coinciding with the most active stage of the disease. In contrast, significantly higher IgG2a antibody reactivity with each biomolecule was observed at 40th day, about two- to fourfold higher than those found at 12th or 20th day, and decreased along 120-day period. Apparently, the conserved B domain is capable to induce IgG2a production in early disease stages. All together, these results suggest that r-potDomain B or synthetic peptides could be molecular starting points in experimental protocols of immunotherapy and/or vaccination for leishmaniasis.

Evaluation of genetic diversity in fig accessions by using microsatellite markers.

Fig (Ficus carica L.) is a fruit of great importance worldwide. Its propagation is carried out with stem cuttings, a procedure that favors the occurrence of synonymy among specimens. Thus, molecular markers have become an important tool for studies of DNA fingerprinting, germplasm characterization, and genetic diversity evaluation in this plant species. The aim of this study was the analysis of genetic diversity among accessions of fig and the detection of synonyms among samples using molecular markers. Five microsatellite markers previously reported as polymorphic to fig were used to characterize 11 fig cultivars maintained in the germplasm bank located in Lavras, Minas Gerais. A total of 21 polymorphic DNA fragments were amplified, with an average of 4.2 alleles per locus. The average allelic diversity and polymorphic information content were 0.6300 and 0.5644, respectively, whereas the total value for the probability of identity was 1.45 x 10(-4). The study allowed the identification of 10 genotypes and 2 synonymous individuals. The principal coordinate analysis showed no defined clusters despite the formation of groups according to geographical origin. However, neighbor-joining analysis identified the same case of synonymy detected using principal coordinate analysis. The data also indicated that the fig cultivars analyzed constitute a population of individuals with high genetic diversity and a broad range of genetic variation.

Relationship between vitamin D and physical activity: systematic review and meta-analysis.

Vitamin D plays an important role in immune function and inflammation and the physical activity demonstrate relationship near to syntheses of vitamin D, considering to necessary to factors immunologics, environmental and physical. We searched five databases through February 20, 2021. Two reviewers screened the studies, collected data, assessed the risk of bias, and ranked the evidence for each outcome across the studies, independently and in duplicate. The prespecified endpoints of interest were Cardiorespiratory Fitness; Sunlight Exposure; Body Mass Index (BMI). We only included data from peer-reviewed articles in our primary analyses. In our primary analysis, there was a positive trend between serum 25(OH)D <20 ng/ml and body mass index, this result should be interpreted with caution, considering confidence intervals (RR 1.10 95% CI 0.37 to 1.83. We identified 4 high quality evidence that vitamin D levels and high physical activity required a direct relationship considering (four trials with 2,253) ; RR 0. (RR 0.0; 95% CI -0.15 to 0.15) (RR 0.59; 95% CI 0.43 to 0.75. Although the evidence available so far, from observational studies of medium quality, can be seen as showing a trend towards an association between sufficient serum levels of 25(OH)D and physical activity, this relationship has been shown. have a stimulating effect on vitamin D synthesis, the relationship of low body mass index with sufficient vitamin D levels is not based on solid evidence. We await results from ongoing studies to determine this effectiveness.

Enrichment of a continuous culture of Saccharomyces cerevisiae with the yeast Issatchenkia orientalis in the production of ethanol at increasing temperatures.

A fermentation system was continuously fed with sugar-cane syrup and operated with recycling of Saccharomyces cerevisiae cells at temperatures varying from 30 to 47 °C. The aim of the present work was to obtain and study the colonies of isolates showing elongated cells of yeasts which were sporadically observed at the end of this continuous process. Based on a sequence of assays involving methods of classical taxonomy and RAPD-PCR, two groups of isolates showing characteristics of non-Saccharomyces yeasts were identified in the yeast population where S. cerevisiae was the dominant yeast. The largest group of non-Saccharomyces yeasts, resulting from a slow proliferation over the 2 months, reached a final level of 29.6% at the end of the process. RAPD-PCR profiles obtained for the isolates of this dominant non-Saccharomyces yeast indicated that they were isolates of Issatchenkia orientalis. Pichia membranifaciens was the only species of non-Saccharomyces yeast detected together with I. orientalis but at a very low frequency. The optimum temperature for ethanol formation shown by the isolate 195B of I. orientalis was 42 °C. This strain also showed a faster ethanol formation and biomass accumulation than the thermotolerant strain of S. cerevisiae used as the starter of this fermentation process. Some isolates of I. orientalis were also able to grow better at 40 °C than at 30 °C on plates containing glycerol as carbon source. Yeasts able to grow and produce ethanol at high temperatures can extend the fermentation process beyond the temperature limits tolerated by S. cerevisiae.

Efficiency of biological control for fall armyworm resistant to the protein Cry1F.

Understanding the ecological and toxicological relationship between genetically modified cultivars (GM) and biological control agents is of great importance for discussions related to the compatability of GM cultivars and integrated management strategies for pest resistance. The present study evaluated the search behavior and predatory capacity of Orius insidiosus (Say) (Hemiptera: Anthocoridae) and Doru luteipes (Scudder) (Dermaptera: Forficulidae) on eggs and caterpillars of Spodoptera frugiperda (J. E. Smith) (Lepidoptera: Noctuidae) resistant or not to the protein Cry1F expressed in Bt corn. To determine the search time, a stopwatch was run until the capture of the first prey, predation capacity was evaluated by counting the prey remaining after 24 hours of infestation. The injuries of S. frugiperda in genetically modified and conventional corn in the presence and absence of predators was also evaluated. The predators were not able to distinguish between resistant and susceptible prey (eggs or caterpillars), given the predatory behaviour observed. There was no difference in searching time or predatory capacity between the predators for eggs and caterpillars of either resistant or susceptible S. frugiperda. In the presence of predators, the injury scores for resistant S. frugiperda on the Bt corn plants were lower. It was concluded that O. insidiosus and D. luteipes did not notice the presence of the protein Cry1F in the prey S. frugiperda, which may facilitate the combined use of GM corn and biological control in integrated management programs and for management of pest resistance.

Resistance of bmr energy sorghum hybrids to sugarcane borer and fall armyworm.

The lower lignin content in plants species with energy potential results in easier cellulose breakdown, making glucose available for ethanol generation. However, higher lignin levels can increase resistance to insect attack. The objective of this work was to evaluate the susceptibility of a bmr-6 biomass sorghum (a mutant genotype with a lower concentration of lignin) to important pests of energy sorghum, Diatraea saccharalis and Spodoptera frugiperda. Experiments were performed in the laboratory and greenhouse to evaluate the development of these pests on the biomass sorghum bmr hybrids BR007, BR008, and TX635 and their respective conventional near-isogenic genotypes (without the bmr gene). The lignin content was higher in non-bmr hybrids, but the evaluated insect variables varied between treatments, not being consistent in just one hybrid or because it is bmr or not. The lowest survival of S. frugiperda was observed in the BR008 hybrid, both bmr and non-bmr. The S. frugiperda injury scores on plants in the greenhouse were high (>7) in all treatments. For D. saccharalis, there was no difference in larval survival in the laboratory, but in the greenhouse, the BR007 hybrid, both bmr and non-bmr, provided greater survival. Due the need to diversify the energy matrix and the fact that greater susceptibility of the bmr hybrids to either pests was not found in this study, these results hold promise for cultivation of these biomass sorghum hybrids for the production of biofuels.

Different visitation frequencies of native and non-native bees to vines: how much vegetation is necessary to improve fruit production?

Pollination is provided by biodiversity and maintains global food production. We investigated the effects of vegetation cover on the abundance of floral visitor and vine (Vitis labrusca Raf.) production. We expected an increase in both floral visitor frequencies and vineyard yields with an increase in native vegetation cover in the landscape. We also investigated different scenarios of visitor abundance with and without honeybees (Apis mellifera L.). We surveyed floral visitors from ten vineyard plots with different native cover surrounding them and related both visitors and native vegetation to fruit set. Considering some of these vineyards, we compared physical and chemical traits of berries to understand how they vary according to native vegetation. Floral visitor abundance was positively related to native vegetation cover. However, considering only native bee abundance, we found a dual (hyperbolic) response. Apis mellifera (L.) Africanized was the most abundant species and had the highest number of interactions; however, when removed from the network analysis, the relationship between vineyards and native bees became more specialized. The fruit size and mass of berries differed among vineyards, as did some chemical traits related to commercial quality of fruits, such as soluble solids, pH and flavonoids. Vineyards surrounded by intermediate areas of native vegetation present a balance between resource availability from vineyards and native vegetation. Apis and non-Apis (such as flies and small bees) floral visitors, known to have different effects on vine pollination, could hypothetically provide variation in vine production and quality. Considering a near 20% native vegetation increment, there was an enhancement, on average, of ten-fold more berries per bunch, the changing physical and chemical fruit traits by vegetation increment could also increase the aggregate value of vines and the value of pollination services in the economy.

Leishmania (Leishmania) infantum/chagasi: Histopathological aspects of the skin in naturally infected dogs in two endemic areas.

In the New World, visceral leishmaniasis (VL), which is a progressive disease and frequently fatal, is caused by Leishmania (Leishmania) infantum/chagasi. It is endemic in many regions of Brazil and occasionally occurs in non-endemic regions when dogs from an endemic area are introduced. The aim of the present study is to compare different skin infection patterns of dogs from two leishmaniasis endemic areas. A histological analysis of dogs from Campo Grande, Mato Grosso do Sul state, a region where epidemic episodes are currently taking place, showed dermic inflammatory infiltrates, composed of numerous vacuolated parasitized macrophages, few lymphocytes, plasma cells and many degranulated mast cells. In the other region of the study, São Luís, Maranhão state, the skin of dogs presented a remarkable inflammatory reaction composed mainly of plasma cells, lymphocytes and very few parasites. We concluded that there is a difference in the skin lesion patterns of dogs with leishmaniasis that is directly related to the endemic area where the animals live.

Does Singular and Stacked Corn Affect Choice Behavior for Oviposition and Feed in Spodoptera frugiperda (Lepidoptera: Noctuidae)?

Little is known about the effects of genetically modified corn plants on the foraging of Spodoptera frugiperda (J.E. Smith). Therefore, this study examines whether singular herbicide-tolerant and insect-resistant plants and their stacked events interfere with food preference and oviposition of S. frugiperda. Two non-Bt corn hybrids and three Bt-hybrids, some of them with glyphosate tolerance (GT), were evaluated. Food preference of larvae and biological parameters were assessed. Oviposition preference bioassays involved choice and no choice condition in plants uninfested and previously infested by larvae in a greenhouse and in the field. The results indicate that there is no relationship between preference of larvae and adult moths. Adult females selected preferentially transgenic hybrids, while larvae selected non-Bt hybrid. Fall armyworm larvae avoid Bt-toxin-expressing leaf tissues, survived only on the non-Bt leaf tissues, and showed minor differences in other life-history traits reared on GT and non-transgenic corn leaf tissues. Female moths showed preference for transgenic plants to lay eggs, but with variable output between previously infested and uninfested plants with larvae. The fact that moths preferred Ag 3700RR2 and non-Bt hybrids for oviposition supports the refuge's strategy aiming at producing susceptible individuals. The use of this hybrid must be integrated with a program of control. The results showed also the importance of correct hybrid selection as part of insect resistance management to Bt-plants. The implications of these findings for understanding the impacts of plant-mediated cues on pest behavior in transgenic crop systems are discussed.

Crystallization, data collection and data processing of maltose-binding protein (MalE) from the phytopathogen Xanthomonas axonopodis pv. citri.

Maltose-binding protein is the periplasmic component of the ABC transporter responsible for the uptake of maltose/maltodextrins. The Xanthomonas axonopodis pv. citri maltose-binding protein MalE has been crystallized at 293 K using the hanging-drop vapour-diffusion method. The crystal belonged to the primitive hexagonal space group P6(1)22, with unit-cell parameters a = 123.59, b = 123.59, c = 304.20 A, and contained two molecules in the asymetric unit. It diffracted to 2.24 A resolution.

Resistance of bmr energy sorghum hybrids to sugarcane borer and fall armyworm / Resistência de híbridos de sorgo energia bmr à broca-da-cana-de-açúcar e lagarta-do-cartucho

Abstract The lower lignin content in plants species with energy potential results in easier cellulose breakdown, making glucose available for ethanol generation. However, higher lignin levels can increase resistance to insect attack. The objective of this work was to evaluate the susceptibility of a bmr-6 biomass sorghum (a mutant genotype with a lower concentration of lignin) to important pests of energy sorghum, Diatraea saccharalis and Spodoptera frugiperda. Experiments were performed in the laboratory and greenhouse to evaluate the development of these pests on the biomass sorghum bmr hybrids BR007, BR008, and TX635 and their respective conventional near-isogenic genotypes (without the bmr gene). The lignin content was higher in non-bmr hybrids, but the evaluated insect variables varied between treatments, not being consistent in just one hybrid or because it is bmr or not. The lowest survival of S. frugiperda was observed in the BR008 hybrid, both bmr and non-bmr. The S. frugiperda injury scores on plants in the greenhouse were high (>7) in all treatments. For D. saccharalis, there was no difference in larval survival in the laboratory, but in the greenhouse, the BR007 hybrid, both bmr and non-bmr, provided greater survival. Due the need to diversify the energy matrix and the fact that greater susceptibility of the bmr hybrids to either pests was not found in this study, these results hold promise for cultivation of these biomass sorghum hybrids for the production of biofuels.

Resumo O menor teor de lignina em espécies de plantas com potencial energético resulta na maior facilidade de quebra da celulose, disponibilizando glicose para geração de etanol. Porém, maiores teores de lignina representa um fator de resistência ao ataque de insetos. O objetivo deste trabalho foi avaliar como importantes pragas do sorgo energia, Diatraea saccharalis e Spodoptera frugiperda, se comportam quanto à alimentação e desempenho em sorgo bmr-6, um genótipo mutante com menor concentração de lignina. Foram realizados experimentos em laboratório e casa de vegetação, avaliando o desenvolvimento destas pragas nos híbridos de sorgo biomassa bmr 007, 008, TX635 e seus respectivos genótipos isogênicos convencionais (sem o gene bmr). O teor de lignina foi maior nos híbridos não bmr, mas nos parâmetros avaliados nos insetos, houve variação entre os tratamentos, não sendo consistente em apenas um híbrido e nem por ser ou não bmr. A menor sobrevivência de S. frugiperda foi verificada no híbrido BR008 tanto bmr quanto não bmr. As notas de injúria por S. frugiperda no sorgo em casa de vegetação foram altas (>7) em todos os tratamentos. Para D. saccharalis, não houve diferença significativa para a sobrevivência larval em laboratório, mas em casa de vegetação o híbrido BR007 tanto bmr quanto não bmr proporcionaram maior sobrevivência. Diante da necessidade de diversificar a matriz energética e o fato de que não foi comprovada neste estudo maior suscetibilidade dos híbridos bmr a ambas as pragas, estes resultados são promissores para o cultivo desses híbridos de sorgo biomassa para produção de biocombustíveis.

Resistance of bmr energy sorghum hybrids to sugarcane borer and fall armyworm

Abstract The lower lignin content in plants species with energy potential results in easier cellulose breakdown, making glucose available for ethanol generation. However, higher lignin levels can increase resistance to insect attack. The objective of this work was to evaluate the susceptibility of a bmr-6 biomass sorghum (a mutant genotype with a lower concentration of lignin) to important pests of energy sorghum, Diatraea saccharalis and Spodoptera frugiperda. Experiments were performed in the laboratory and greenhouse to evaluate the development of these pests on the biomass sorghum bmr hybrids BR007, BR008, and TX635 and their respective conventional near-isogenic genotypes (without the bmr gene). The lignin content was higher in non-bmr hybrids, but the evaluated insect variables varied between treatments, not being consistent in just one hybrid or because it is bmr or not. The lowest survival of S. frugiperda was observed in the BR008 hybrid, both bmr and non-bmr. The S. frugiperda injury scores on plants in the greenhouse were high (>7) in all treatments. For D. saccharalis, there was no difference in larval survival in the laboratory, but in the greenhouse, the BR007 hybrid, both bmr and non-bmr, provided greater survival. Due the need to diversify the energy matrix and the fact that greater susceptibility of the bmr hybrids to either pests was not found in this study, these results hold promise for cultivation of these biomass sorghum hybrids for the production of biofuels.

Resumo O menor teor de lignina em espécies de plantas com potencial energético resulta na maior facilidade de quebra da celulose, disponibilizando glicose para geração de etanol. Porém, maiores teores de lignina representa um fator de resistência ao ataque de insetos. O objetivo deste trabalho foi avaliar como importantes pragas do sorgo energia, Diatraea saccharalis e Spodoptera frugiperda, se comportam quanto à alimentação e desempenho em sorgo bmr-6, um genótipo mutante com menor concentração de lignina. Foram realizados experimentos em laboratório e casa de vegetação, avaliando o desenvolvimento destas pragas nos híbridos de sorgo biomassa bmr 007, 008, TX635 e seus respectivos genótipos isogênicos convencionais (sem o gene bmr). O teor de lignina foi maior nos híbridos não bmr, mas nos parâmetros avaliados nos insetos, houve variação entre os tratamentos, não sendo consistente em apenas um híbrido e nem por ser ou não bmr. A menor sobrevivência de S. frugiperda foi verificada no híbrido BR008 tanto bmr quanto não bmr. As notas de injúria por S. frugiperda no sorgo em casa de vegetação foram altas (>7) em todos os tratamentos. Para D. saccharalis, não houve diferença significativa para a sobrevivência larval em laboratório, mas em casa de vegetação o híbrido BR007 tanto bmr quanto não bmr proporcionaram maior sobrevivência. Diante da necessidade de diversificar a matriz energética e o fato de que não foi comprovada neste estudo maior suscetibilidade dos híbridos bmr a ambas as pragas, estes resultados são promissores para o cultivo desses híbridos de sorgo biomassa para produção de biocombustíveis.

The flavonoid rutin induces astrocyte and microglia activation and regulates TNF-alpha and NO release in primary glial cell cultures.

Astrocyte and microglia cells play an important role in the central nervous system (CNS). They react to various external aggressions by becoming reactive and releasing neurotrophic and/or neurotoxic factors. Rutin is a flavonoid found in many plants and has been shown to have some biological activities, but its direct effects on cells of the CNS have not been well studied. To investigate its potential effects on CNS glial cells, we used both astrocyte primary cultures and astrocyte/microglia mixed primary cell cultures derived from newborn rat cortical brain. The cultures were treated for 24 h with rutin (50 or 100 micromol/L) or vehicle (0.5% dimethyl sulfoxide). Mitochondrial function on glial cells was not evidenced by the MTT test. However, an increased lactate dehydrogenase activity was detected in the culture medium of both culture systems when treated with 100 micromol/L rutin, suggesting loss of cell membrane integrity. Astrocytes exposed to 50 micromol/L rutin became reactive as revealed by glial fibrillary acidic protein (GFAP) overexpression and showed a star-like phenotype revealed by Rosenfeld's staining. The number of activated microglia expressing OX-42 increased in the presence of rutin. A significant increase of nitric oxide (NO) was observed only in mixed cultures exposed to 100 micromol/L rutin. Enhanced TNFalpha release was observed in astrocyte primary cultures treated with 100 micromol/L rutin and in mixed primary cultures treated with 50 and 100 micromol/L, suggesting different sensitivity of both activated cell types. These results demonstrated that rutin affects astrocytes and microglial cells in culture and has the capacity to induce NO and TNFalpha production in these cells. Hence, the impact of these effects on neurons in vitro and in vivo needs to be studied.

A Leishmania (L.) amazonensis ATP diphosphohydrolase isoform and potato apyrase share epitopes: antigenicity and correlation with disease progression.

A Leishmania (Leishmania) amazonensis ATP diphosphohydrolase isoform was partially purified from plasma membrane of promastigotes by preparative non-denaturing polyacrylamide gel electrophoresis. SDS-PAGE followed by Western blots developed with polyclonal anti-potato apyrase antibodies identified diffuse bands of about 58-63 kDa, possibly glycosylated forms of this protein. By ELISA technique, a significantly higher total IgG antibody level against potato apyrase was found in serum from promastigote-infected mice, as compared to the uninfected mice, confirming both the existence of shared epitopes between the parasite and vegetable proteins, and the parasite ATP diphosphohydrolase antigenicity. By Western blotting, serum from amastigote-infected BALB/c mice recognizes both potato apyrase and this antigenic ATP diphosphohydrolase isoform isolated from promastigotes, suggesting that it is also expressed in the amastigote stage. The infection monitored along a 90-day period in amastigote-infected mice showed reactivity of IgG2a antibody in early steps of infection, while the disappearance of the IgG2a response and elevation of IgG1 antibody serum levels against that shared epitopes were associated with the progression of experimental leishmaniasis. This is the first observation of the antigenicity of a L. (L.) amazonensis ATP diphosphohydrolase isoform, and of the ability of cross-immunoreactivity with potato apyrase to differentiate serologically stages of leishmaniasis in infected mice.

Pollination and reproductive system of synchronopatric species of Cactaceae (Cactoideae) subject to interspecific flow of pollen: an example of ecological adaptation in the Brazilian Chaco.

Three synchronopatric Cactaceae species, Echinopsis rhodotricha, Harrisia balansae and Praecereus saxicola, have mostly nocturnal anthesis and similar flowers, characteristics that motivated us to perform a comparative study of reproductive ecology. Reproductive phenology was sampled monthly from December 2014 to November 2015. We describe floral biology, breeding system via pollination treatments and evaluate floral visitors from focal and filming observations. Pollen grains found on moth proboscis were compared among cactus species under light microscopy. We used fluorescent dye particles to test intra- and interspecific pollen flow. These three species have extended flowering with greater intensity in the wet season, causing high overlap. They have white and hypocrateriformis flowers that open at twilight or nightfall and last about 15 h. H. balansae seems to be self-incompatible, while E. rhodotricha presented self-compatibility. P. saxicola presented self-fertility, but most of the population seems to be self-incompatible. We suggest sphingophily for the three species, but only P. saxicola was visited by Manduca rustica (Sphingidae). However, we observed pollen grains of all three species on the proboscis of moths, especially M. rustica and M. sexta. Prolonged anthesis allowed bees (herein considered as secondary pollinators) to visit flowers of E. rhodotricha and P. saxicola. It can be concluded that the studied species share nocturnal and diurnal pollinators, suggesting interspecific pollen flow, which, however, could not be detected with fluorescent dye particles. In view of the low frequency of primary pollinators, it appears that these three species have different reproductive strategies, ensuring the fruiting and production of viable seeds.

Alkaloids from Prosopis juliflora leaves induce glial activation, cytotoxicity and stimulate NO production.

Prosopis juliflora is used for feeding cattle and humans. Intoxication with the plant has been reported, and is characterized by neuromuscular alterations and gliosis. Total alkaloidal extract (TAE) was obtained using acid/basic-modified extraction and was fractionated. TAE and seven alkaloidal fractions, at concentrations ranging 0.03-30 microg/ml, were tested for 24h on astrocyte primary cultures derived from the cortex of newborn Wistar rats. The MTT test and the measure of LDH activity on the culture medium, revealed that TAE and fractions F29/30, F31/33, F32 and F34/35 were cytotoxic to astrocytes. The EC(50) values for the most toxic compounds, TAE, F31/33 and F32 were 2.87 2.82 and 3.01 microg/ml, respectively. Morphological changes and glial cells activation were investigated through Rosenfeld's staining, by immunocytochemistry for the protein OX-42, specific of activated microglia, by immunocytochemistry and western immunoblot for GFAP, the marker of reactive and mature astrocytes, and by the production of nitric oxide (NO). We observed that astrocytes exposed to 3 microg/ml TAE, F29/30 or F31/33 developed compact cell body with many processes overexpressing GFAP. Treatment with 30 microg/ml TAE and fractions, induced cytotoxicity characterized by a strong cell body contraction, very thin and long processes and condensed chromatin. We also observed that when compared with the control (+/-1.34%), the proportion of OX-42 positive cells was increased in cultures treated with 30 microg/ml TAE or F29/30, F31/33, F32 and F34/35, with values raging from 7.27% to 28.74%. Moreover, incubation with 3 microg/ml F32, 30 microg/ml TAE, F29/30, F31/33 or F34/35 induced accumulation of nitrite in culture medium indicating induction of NO production. Taken together these results show that TAE and fractionated alkaloids from P. juliflora act directly on glial cells, inducing activation and/or cytotoxicity, stimulating NO production, and may have an impact on neuronal damages observed on intoxicated animals.