Compatible and Incompatible Interactions Between Callus Tissue and Mycorrhizal or Pathogenic Fungi

Tissue cultures of Nicotiana tabacum were utilized to investigate the mechanisms associated with host specificity and non-host incompatibility in mycorrhizal and pathogenic fungi. They were tested for expression of resistance to different species of mycorrhizal fungi and to a fungal pathogen of tobacco, Thielaviopsis basicola , by monitoring the production of callose, phenolic compounds and peroxidases in dual cultures. Tobacco cells reacted to the presence of all the mycorrhizal fungi with callose deposits, whereas callose was nearly always absent in tobacco cells inoculated with their pathogen T. basicola. The broad-host range ectomycorrhizal fungi Hebeloma crustuliniforme, Lac-caria laccata and Suilhis granulatus elicited less intense responses than did Hymenoscyphus ericae. The results obtained for phenolic production and peroxidase activity were consistently similar to those obtained for callose deposition. They showed that H. ericae , an endomycorrhizal symbiont of Ericaceae, was highly incompatible with tobacco cells and that the tobacco pathogen T. basicola did not elicit strong reactions in the cells of its host. In this paper, the possibility of utilizing callus cultures as a simple model system to study both the different degrees of compatibility and the early events of recognition between mycorrhizal fungi and their host or non-host plants is discussed.     

Molecular and chromosomal evolution in anoas (Bovidae: Bubalus spec.)

As a contribution to their taxonomy, population genetic data on zoo-living anoas are reported, and a review of the history of the captive stock is provided. Four different chromosome numbers of 44, 45, 47 and 48 chromosomes have been found, respectively, when karyotyping captive anoas descending from three breeding lines. The number of chromosome arms is 60 throughout, indicating that Robertsonian rearrangements are responsible for this cytogenetic variation. An electrophoretic comparison of isozymes and blood proteins representing 21 genetic loci revealed polymorphism in seven loci: haemoglobin, glyoxalase, superoxide dismutase, phosphoglucomutase, carbonic anhydrase, glucose phosphate isomerase, and an unidentified acid serum protein. Considering the small number of founder specimens and subsequent inbreeding, allozyme variability appears fairly high in anoas. Genetic distances between zoo populations amount to 0.0505 or less. Southern blot hybridizations of restricted DNA from anoas and African buffaloes with a probe from the DRB-like region of the chimpanzee's MHC class II genes also indicate a low degree of genetic differentiation between mountain and lowland anoas. The relevance of these genetic data for the taxonomic classification of mountain and lowland anoas, and for the conservation of anoas by captive breeding is discussed.     

Antiproliferative Potential of Olneya tesota PF2 Lectin in Human Acute Monocytic Leukemia Cells

Acute monocytic leukemia is a type of myeloid leukemia that develops in monocytes. The current clinical therapies for leukemia are unsatisfactory due to their side effects and nonspecificity toward target cells. Some lectins display antitumor activity and may specifically recognize cancer cells by binding to carbohydrate structures on their surface. Therefore, this study evaluated the response of the human monocytic leukemia cell lines THP-1 to the Olneya tesota PF2 lectin. The induction of apoptosis and reactive oxygen species production in PF2-treated cells was evaluated by flow cytometry, and the lectin-THP-1 cell interaction and mitochondrial membrane potential were evaluated by confocal fluorescence microscopy. PF2 genotoxicity was evaluated by DNA fragmentation analysis via gel electrophoresis. The results showed that PF2 binds to THP-1 cells, triggers apoptosis and DNA degradation, changes the mitochondrial membrane potential, and increases reactive oxygen species levels in PF2-treated THP-1 cells. These results suggest the potential use of PF2 for developing alternative anticancer treatments with enhanced specificity.     

Proteomic fingerprinting enables quantitative biodiversity assessments of species and ontogenetic stages in Calanus congeners (Copepoda,Crustacea) from the Arctic Ocean

Species identification is pivotal in biodiversity assessments and proteomic fingerprinting by MALDI-TOF mass spectrometry has already been shown to reliably identify calanoid copepods to species level. However, MALDI-TOF data may contain more information beyond mere species identification. In this study, we investigated different ontogenetic stages (copepodids C1–C6 females) of three co-occurring Calanus species from the Arctic Fram Strait, which cannot be identified to species level based on morphological characters alone. Differentiation of the three species based on mass spectrometry data was without any error. In addition, a clear stage-specific signal was detected in all species, supported by clustering approaches as well as machine learning using Random Forest. More complex mass spectra in later ontogenetic stages as well as relative intensities of certain mass peaks were found as the main drivers of stage distinction in these species. Through a dilution series, we were able to show that this did not result from the higher amount of biomass that was used in tissue processing of the larger stages. Finally, the data were tested in a simulation for application in a real biodiversity assessment by using Random Forest for stage classification of specimens absent from the training data. This resulted in a successful stage-identification rate of almost 90%, making proteomic fingerprinting a promising tool to investigate polewards shifts of Atlantic Calanus species and, in general, to assess stage compositions in biodiversity assessments of Calanoida, which can be notoriously difficult using conventional identification methods.     

A chromosome-level genome assembly enables the identification of the follicule stimulating hormone receptor as the master sex-determining gene in the flatfish Solea senegalensis

Sex determination (SD) shows huge variation among fish and a high evolutionary rate, as illustrated by the Pleuronectiformes (flatfishes). This order is characterized by its adaptation to demersal life, compact genomes and diversity of SD mechanisms. Here, we assembled the Solea senegalensis genome, a flatfish of great commercial value, into 82 contigs (614 Mb) combining long- and short-read sequencing, which were next scaffolded using a highly dense genetic map (28,838 markers, 21 linkage groups), representing 98.9% of the assembly. Further, we established the correspondence between the assembly and the 21 chromosomes by using BAC-FISH. Whole genome resequencing of six males and six females enabled the identification of 41 single nucleotide polymorphism variants in the follicle stimulating hormone receptor (fshr) consistent with an XX/XY SD system. The observed sex association was validated in a broader independent sample, providing a novel molecular sexing tool. The fshr gene displayed differential expression between male and female gonads from 86 days post-fertilization, when the gonad is still an undifferentiated primordium, concomitant with the activation of amh and cyp19a1a, testis and ovary marker genes, respectively, in males and females. The Y-linked fshr allele, which included 24 nonsynonymous variants and showed a highly divergent 3D protein structure, was overexpressed in males compared to the X-linked allele at all stages of gonadal differentiation. We hypothesize a mechanism hampering the action of the follicle stimulating hormone driving the undifferentiated gonad toward testis.     

Effect of different doses of chromium picolinate on protein metabolism in infant rats.

This 12-day study was conducted to evaluate the effects of three different levels of dietary chromium (100, 200, and 500 microg/day) in the form of chromium picolinate (CrPic) on growth and protein use in weaned rats. No significant effect of CrPic on body weight gain, food intake, or food conversion rate was observed. Elevated doses of CrPic seemed to increase muscle mass, either by stimulating protein anabolism by activation of insulin by chromium or by lowering protein degradation. However, these effects had no repercussions on overall growth, suggesting that any anabolic effect of chromium due to the action of insulin was probably marginal.     

Conformational changes induced by cloxacillin in class a beta-lactamase from Bacillus cereus.

Class A beta-lactamases are enzymes that hydrolyse beta-lactam antibiotics such as penicillins and cephalosporins. They also hydrolyse substrate analogues such as oxacillin and cloxacillin, with a biphasic kinetic as it has been reported for Bacillus cereus beta-lactamase. A molecular model of Bacillus cereus beta-lactamase was built and the conformational changes that the substrates benzylpenicillin and cloxacilline produced in the conformation of selected regions of the protein were analyzed. This study was performed using the docking of the substrates, their tetrahedral intermediates and the corresponding acids on the active site, followed by molecular dynamic and subsequent optimisation procedures. The most important changes were produced on Tyr105 and Tyr273, when the tetrahedral intermediate of cloxacillin was docked at the active site, these amino acids are partially responsible for the stabilisation of the substrates at the active site. These changes may explain the kinetic differences observed during the hydrolysis of substrates type S and type A by beta-lactamases class A.     

Genetic Diversity of Isolates of Glomus mosseae from Different Geographic Areas Detected by Vegetative Compatibility Testing and Biochemical and Molecular Analysis

We detected, for the first time, the occurrence of vegetative incompatibility between different isolates of the arbuscular mycorrhizal fungal species Glomus mosseae. Vegetative compatibility tests performed on germlings belonging to the same isolate showed that six geographically different isolates were capable of self-anastomosing, and that the percentage of hyphal contacts leading to fusions ranged from 60 to 85%. Successful anastomoses were characterized by complete fusion of hyphal walls, protoplasm continuity and occurrence of nuclei in the middle of hyphal bridges. No anastomoses could be detected between hyphae belonging to different isolates, which intersected without any reaction in 49 to 68% of contacts. Microscopic examinations detected hyphal incompatibility responses in diverse pairings, consisting of protoplasm retraction from the tips and septum formation in the approaching hyphae, even before physical contact with neighboring hyphae. Interestingly, many hyphal tips showed precontact tropism, suggesting that specific recognition signals may be involved during this stage. The intraspecific genetic diversity of G. mosseae revealed by vegetative compatibility tests was confirmed by total protein profiles and internal transcribed spacer-restriction fragment length polymorphism profiles, which evidenced a higher level of molecular diversity between the two European isolates IMA1 and BEG25 than between IMA1 and the two American isolates. Since arbuscular mycorrhizal fungi lack a tractable genetic system, vegetative compatibility tests may represent an easy assay for the detection of genetically different mycelia and an additional powerful tool for investigating the population structure and genetics of these obligate symbionts.     

Ras-mediated cell proliferation and cell death: some clues from the interleukin-2 receptor system

Oncoproteins of the Ras family have been extensively studied because of their implication in human cancer. Their roles have been primarily assigned to the commandment of cell proliferation and suppression of apoptosis, which has also been demonstrated by the involvement of Ras activation in the signal transduction pathways triggered by most cytokine receptors. Nevertheless, the functions of Ras proteins have been extended in the last years by the findings showing that they can also act as promoters or enhancers of apoptosis in various systems and conditions. These considerations have raised the issue as to how the signals delivered by Ras are regulated and translated in terms of cellular responses, suggesting that signal complementation may direct the final fate of cells. As an example, the interleukin-2 receptor system may represent a useful model in which the meaning of Ras signals may be evaluated in terms of interactions with other simultaneous signalling events, since knowledge of the biochemical events triggered by the interaction of interleukin-2 with its cell surface receptor in lymphocytes has allowed the proposal of a complete signalling model arranged in three independent channels, one of which is mediated by Ras.This work was supported by grants from CICYT and Pharmacia-Upjohn.     

Microbiological and technological aspects of cassava-starch fermentation

The major genera found in the microflora of fermented, sour, cassava-starch were Streptococcus, Bacillus, Lactobacillus and Saccharomyces with amylase activity. Lactic acid bacteria predominated whereas the presence of moulds was not significant. No coliforms were detected. Electron microscopy showed bacteria and yeasts in contact with the starch granules and signs of erosion on the granule surface. Lactic acid was the main metabolite; no oligosaccharides, maltose or glucose were detected, indicating their rapid utilization. The degree of acidification, which correlated with the decrease in viscosity and the final quality of the product, was influenced by the variable microbial ecology.