Large and variable genome size unrelated to serpentine adaptation but supportive of cryptic sexuality in Cenococcum geophilum

Estimations of genome size and its variation can provide valuable information regarding the genetic diversity of organisms and their adaptation potential to heterogeneous environments. We used flow cytometry to characterize the variation in genome size among 40 isolates of Cenococcum geophilum, an ectomycorrhizal fungus with a wide ecological and geographical distribution, obtained from two serpentine and two non-serpentine sites in Portugal. Besides determining the genome size and its intraspecies variation, we wanted to assess whether a relationship exists between genome size and the edaphic background of the C. geophilum isolates. Our results reveal C. geophilum to have one of the largest genome sizes so far measured in the Ascomycota, with a mean haploid genome size estimate of 0.208 pg (203 Mbp). However, no relationship was found between genome size and the edaphic background of the sampled isolates, indicating genetic and demographic processes to be more important for shaping the genome size variation in this species than environmental selection. The detection of variation in ploidy level among our isolates, including a single individual with both presumed haploid and diploid nuclei, provides supportive evidence for a possible cryptic sexual or parasexual cycle in C. geophilum (although other mechanisms may have caused this variation). The existence of such a cycle would have wide significance, explaining the high levels of genetic diversity and likelihood of recombination previously reported in this species, and adds to the increasing number of studies suggesting sexual cycles in previously assumed asexual fungi.     

HERITABILITY OF AEROBIC POWER OF INDIVIDUALS IN NORTHEAST BRAZIL

The objective of this study was to evaluate the genetic and environmental contribution to variation in aerobic power in monozygotic (MZ) and dizygotic (DZ) twins. The sample consisted of 20 MZ individuals (12 females and 8 males) and 16 DZ individuals (12 females and 4 males), aged from 8 to 26 years, residents in Natal, Rio Grande do Norte. The twins were assessed by a multistage fitness test. The rate of heritability found for aerobic power was 77%. Based on the results, the estimated heritability was largely responsible for the differences in aerobic power. This implies that such measures are under strong genetic influence.     

Sexual reproduction of the pentaploid,short‐styled Oxalis pes‐caprae allows the production of viable offspring

Reproduction is a key factor for the successful establishment and spread of introduced species. Oxalis pes‐caprae is a tristylous species with a self‐ and morph‐incompatibility sexual system that, in the invaded range of the western Mediterranean Basin, has been found to reproduce asexually because only the pentaploid, short‐styled morph (5x S‐morph) was introduced. The objective of this study was to test the ability of the 5x S‐morph of O. pes‐caprae to produce viable offspring in the absence of compatible mates, exploring the hypothesis that new morphs could have emerged by sexual reproduction events of the initially introduced morph. Pollen germination, pollen tube development, fruit and seed production, seed germination and offspring ploidy levels were analysed after controlled hand‐pollinations to assess self‐ and morph‐incompatibility and production of viable gametes by the 5x S‐morph. The self‐incompatibility system is still operating, but a partial breakdown in the morph‐incompatibility system combined with the production of viable gametes was observed, allowing sexual reproduction of the 5x S‐morph in the invaded range. The ability of the 5x S‐morph to reproduce sexually may have major consequences for the dynamics of invasive populations of O. pes‐caprae and could be one of the factors involved in the occurrence of new floral morphs in this invaded range.     

Generation of a human IgM monoclonal antibody directed against HLA class II molecules: a potential agent in the treatment of haematological malignancies

Major histocompatibility complex (MHC) class II molecules have been considered as a good target molecule for use in immunotherapy, because of the high expression in some lymphoma and leukaemia cells and, also, because of their restricted expression on human cells (monocytes, dendritic, B lymphocytes, thymic epithelial cells, and some cytokine-activated cells, such as T lymphocytes). We have obtained a human IgM monoclonal antibody directed against human leukocyte antigen (HLA) class II molecules, using transgenic mice carrying human Ig genes. The antibody BH1 (IgM/κ isotype) recognises HLA-class II on the surface of tumour cells from patients suffering from haematological malignancies, such as chronic and acute lymphocytic leukaemias, non-Hodgkin lymphomas and myeloid leukaemias. Interestingly, functional studies revealed that BH1 mAb recognises and kills very efficiently tumour cells from several leukaemia patients in the presence of human serum as a source of complement. These results suggest that this human IgM monoclonal antibody against HLA-class II could be considered as a potential agent in the treatment of several malignancies. Belén Díaz, Irene Sanjuan and Susana Magadán share authorship; Francisco Gambón and áfrica González–Fernández share leadership.     

The feasibility of detecting motor and sensory potentials in a sheep model

To investigate the characteristics of motor, sensory and sensory-evoked potentials (SEPs) of thoracic and lumbar roots, and demonstrate the feasibility of assessing axonal regrowth after the neurotization procedure in a sheep model. Six adult sheep were anaesthetized and placed in a sternal position. The thoracic and lumbar roots from T11 to L5 were identified at their emergence from the vertebral foramen and stimulated. Motor and sensory responses were monitored. Thoracic and lumbar roots were easily identified in all cases. Motor potentials were detected for each stimulated nerve without difficulty. The amplitudes were quite variable, ranging from 100 to 5300 microV. Sensory and SEPs were satisfactorily recorded in only three of the six animals. Sensory amplitudes also varied greatly, ranging from 25 to 120 microV. In three cases, SEPs could not be identified due to motor artefacts. The motor pathway after axonal regrowth in neurotized lumbar roots might easily be explored by proximal electric stimulation of the root, close to the sutured area. Detection of sensory and spinal cord evoked potentials might be improved by the use of curve summation techniques. Specific axonal tracing holds promise of being a useful technique for examining sensory and motor pathway recovery after neurotization in the sheep model.     

Flow cytometric and microscopic analysis of the effect of tannic acid on plant nuclei and estimation of DNA content

• Background and Aims Flow cytometry (FCM) is extensivelyused to estimate DNA ploidy and genome size in plants. In orderto determine nuclear DNA content, nuclei in suspension are stainedby a DNA-specific fluorochrome and fluorescence emission isquantified. Recent studies have shown that cytosolic compoundsmay interfere with binding of fluorochromes to DNA, leadingto flawed data. Tannic acid, a common phenolic compound, maybe responsible for some of the stoichiometric errors, especiallyin woody plants. In this study, the effect of tannic acid onestimation of nuclear DNA content was evaluated in Pisum sativumand Zea mays, which were chosen as model species. • Methods Nuclear suspensions were prepared from P. sativumleaf tissue using four different lysis buffers (Galbraith's,LB01, Otto's and Tris.MgCl₂). The suspensions were treated withtannic acid (TA) at 13 different initial concentrations rangingfrom 0·25 to 3·50 mg mL^–1. After propidiumiodide (PI) staining, samples were analysed using FCM. In additionto the measurement of nuclei fluorescence, light scatter propertieswere assessed. Subsequently, a single TA concentration was chosenfor each buffer and the effect of incubation time was assessed.Similar analyses were performed on liquid suspensions of P.sativum and Z. mays nuclei that were isolated, treated and analysedsimultaneously. FCM analyses were accompanied by microscopicobservations of nuclei suspensions. • Key Results TA affected PI fluorescence and light scatterproperties of plant nuclei, regardless of the isolation bufferused. The least pronounced effects of TA were observed in Tris.MgCl₂buffer. Samples obtained using Galbraith's and LB01 bufferswere the most affected by this compound. A newly described ‘tannicacid effect’ occurred immediately after the addition ofthe compound. With the exception of Otto's buffer, nuclei ofP. sativum and Z. mays were affected differently, with pea nucleiexhibiting a greater decrease in fluorescence intensity. • Conclusions A negative effect of a secondary metabolite,TA, on estimation of nuclear DNA content is described and recommendationsfor minimizing the effect of cytosolic compounds are presented.Alteration in light scattering properties of isolated nucleican be used as an indicator of the presence of TA, which maycause stoichiometric errors in nuclei staining using a DNA intercalator,PI.     

Design, synthesis, SAR, and biological evaluation of new 4-(phenylamino)thieno[2,3-b]pyridine derivatives

In this work, we performed the design, synthesis, and the structure-activity relationship studies of 13 new derivatives of thieno[2,3-b]pyridine. These derivatives were prepared in high yields (96-70%) and their structures were elucidated by IR, (1)H, (13)C NMR, and MS. The biological results showed some derivatives as antiparasitic agents against Giardia lamblia. Computational analysis of HOMO and LUMO energy, HOMO orbital coefficient distribution, electrostatic potential map, dipole moment, and density HOMO was performed to gain insight into the SAR aspects. This study pointed the p-methoxy substituted derivative as a leading compound for the development of new microbicidal medicines based on thieno[2,3-b]pyridine analogs.     

Quantitation of malondialdehyde in gingival crevicular fluid by a high-performance liquid chromatography-based method

Lipid peroxidation (LPO) has been associated with periodontal disease, and the evaluation of malondialdehyde (MDA) in the gingival crevicular fluid (GCF), an inflammatory exudate from the surrounding tissue of the periodontium, may be useful to clarify the role of LPO in the pathogenesis of periodontal disease. We describe the validation of a method to measure MDA in the GCF using high-performance liquid chromatography. MDA calibration curves were prepared with phosphate-buffered solution spiked with increasing known concentrations of MDA. Healthy and diseased GCF samples were collected from the same patient to avoid interindividual variability. MDA response was linear in the range measured, and excellent agreement was observed between added and detected concentrations of MDA. Samples' intra- and interday coefficients of variation were below 6.3% and 12.4%, respectively. The limit of quantitation (signal/noise=5) was 0.03 μM. When the validated method was applied to the GCF, excellent agreement was observed in the MDA quantitation from healthy and diseased sites, and diseased sites presented more MDA than healthy sites (P<0.05). In this study, a validated method for MDA quantitation in GCF was established with satisfactory sensitivity, precision, and accuracy.