Group I introns in the liverwort mitochondrial genome: the gene coding for subunit 1 of cytochrome oxidase shares five intron positions with its fungal counterparts.

The complete nucleotide sequence of the mitochondrial DNA (mtDNA) from a liverwort, Marchantia polymorpha, contains thirty-two introns. Twenty-five of these introns possess the characteristic secondary structures and consensus sequences of group II introns. The remaining seven are group I introns, six of which happen to interrupt the gene coding for subunit 1 of cytochrome oxidase (cox1). Interestingly, the insertion sites of one group II and four group I introns in the cox1 gene coincide with those of the respective fungal mitochondrial interns. Moreover, comparison of the four group I introns with their fungal counterparts shows that group I introns inserted at identical genomic sites in different organisms are indeed related to one another, in terms of the peptide sequences generated from the complete or fragmental ORFs encoded by these introns. At the same time, the liverwort introns turned out to be more divergent from their fungal cognates than the latter are from one another. We therefore conclude that vertical transmission from a common ancestor organism is the simplest explanation for the presence of cognate introns in liverwort and fungal mitochondrial genomes.     

Nucleotide substitutions at the -6 position in the promoter region of the factor IX gene result in different severity of hemophilia B Leyden: consequences for genetic counseling

Mutations in the promoter region of the factor IX gene result in hemophilia B Leyden, which is characterized by considerable improvement in the disease after puberty. We have found that distinct nucleotide substitutions at the -6 position in the Leyden-specific (LS) region are associated with a different severity of hemophilia B. The proband (aged 2) from one family is a severe hemophiliac with factor IX activity (F.IXC) and antigen (F.IXAg) levels less than 1.0U/dl. F.IXC and F.IXAg levels in two affected uncles are approximately 30% of normal levels. The LS region was targeted for analysis because the phenotypes suggested the inheritance of a factor IX Leyden gene. An abnormal TaqI digestion pattern was found in amplified DNA from the proband, and sequencing showed a G (-6) to C transversion that was linked to the disease in the family. In another family, two brothers (aged 8 and 9) suffer from mild hemophilia with F.IXC ranging from 7 to 10 U/dl and F.IXAg from 3 to 4 U/dl. They are the only documented members of the family with a bleeding tendency. Denaturing gradient gel electrophoresis on amplified fragments from one of the patient's genomic DNA corresponding to the 8 exons and flanking sequences of the factor IX gene suggested a defect only in a segment from the 5 region. This segment showed an altered TaqI digestion pattern, and sequencing demonstrated a G(-6) to A transition that was traced to the patients's mother and a grandmother. The different phenotypes associated with the G (-6) to A purine nucleotide transition compared with a G(-6) to C transversion provide evidence that this area is directly involved in the regulation of the human factor IX gene expression in vivo by binding of regulatory factors. The ability to predict that the conditions of a hemophilia B patient will improve with age has important implications for genetic counseling of the family. Therefore, the LS region should always be included when scanning the factor IX gene for mutations.     

Quantitative estimation of chimerism in mice using microsatellite markers

An embryonic stem cell line was established from SV129 mouse blastocysts and used to generate chimeric mice by injection into OF1 blastocysts; 18 out of the 30 resulting offspring appeared chimeric as judged from their coat color patterns, and 3 of the 13 males proved to be germ-line chimeras as they transmitted the SV129 agouti phenotype to all or part of their offspring. The degree of chimerism of these males was evaluated for different tissues using polymorphic microsatellite markers amplified by the polymerase chain reaction. It was shown that these new markers can be effectively used to quantitatively estimate levels of chimerism. The CKMM (creatine kinase, muscle) microsatellite system was used to distinguish the SV129 from the OF1 genotype. In all performed tests, the correlation between DNA ratio and signal ratio, expressed as a base 10 logarithm, was shown to exceed or equal 0.98 for known DNA ratios (SV129/OF1) ranging from 1/99 to 99/1. Linear calibration methods were used to predict the % SV129 DNA of a test sample based on the obtained signal ratio. The accuracy of the prediction was evaluated by performing repeated measurements. Differences among three repeated estimates ranged from 2 to 17% for a given sample. Microsatellite systems should be very useful to monitor chimerism involving strains that can not be discerned with coat color or biochemical markers. This will be particularly important when ES methodology becomes available in species other than mice. © 1993 Wiley-Liss, Inc.     

Partitioning the biodiversity effects on productivity into density and size components

Plant density and size — two factors that represent plant survival and growth — are key determinants of yield but have rarely been analysed explicitly in the context of biodiversity–productivity relationships. Here, we derive equations to partition the net, complementarity and selection effects of biodiversity into additive components that reflect diversity-induced changes in plant density and size. Applications of the new method to empirical datasets reveal contrasting ways in which plant density and size regulate yield in species mixtures. In an annual plant diversity experiment, overyielding is largely explained by selection effects associated with increased size of highly productive plant species. In a tree diversity experiment, the cause of overyielding shifts from enhanced growth in tree size to reduced mortality by complementary use of canopy space during stand development. These results highlight the capability of the new method to resolve crucial, yet understudied, demographic links between biodiversity and productivity.     

Growth and respiratory oxidation of reduced sulfur compounds by intact cells ofThiobacillus novellus (type strain) grown on thiosulfate

Thiobacillus novellus (type strain) was grown chemolithoautrophically on thiosulfate in batch cultures under microaerophilic conditions. Under these conditions,T. novellus grew exponentially (=0.05–0.06 h^–1). The respiratory oxidation rates of tetrathionate, thiosulfate, elemental sulfur (S^o), and sulfite were measured respirometrically with an oxygen electrode, with exponentially growing cells. Cells growing on thiosulfate as the unique energy source retain thiosulfate-oxidizing activity, S^o-oxidizing activity (SOA), and very high sulfite-oxidizing activity, but lack respiratory tetrathionate-oxidizing activity. HQNO (50 m), an inhibitor of the quinone-cytochrome b region, strongly inhibited the SOA (70%), moderately the sulfite-oxidizing activity (45%), and poorly the thiosulfate-oxidizing activity (15%), 1mm KCN totally inhibited (>89%) all respiratory activities. This study confirms that inThiobacillus novellus, as well as in otherThiobacilli, SOA is present in cells grown with thiosulfate as sole electron donor. SOA appears not to be an oxygenase; it is linked to the respiratory chain, and the electrons are probably released in the quinone-cytochrome b region.     

Elemental sulfur production during mixotrophic growth on hydrogen and thiosulfate of thermophilic hydrogen-oxidizing bacteria

In this study we measured the exogenous production and the intracellular content of elemental sulfur (S⁰) in the thermophilic sulfur-oxidizing bacteriaHydrogenobacter spp. andBacillus schlegelii during mixolithoautotrophic growth on hydrogen and thiosulfate. Under these conditions, all strains produced and released white-yellow hydrophilic S⁰ particles into the growth medium. Hydrophilic S⁰ was separated from cells by a differential low-speed centrifugation procedure. The S⁰ pellets were dried, and the S⁰ was purified by column chromatography and by thin-layer chromatography (TLC). The S⁰ TLC-band could be stained with triphenyltetrazolium chloride and piperidine procedure. Determination of intracellular S⁰ content was performed from fresh cells absolutely free of exogenous S⁰ particles. quantitation of S⁰ was performed by high-performance liquid chromatography, colorimetric thiocyanate procedure, and by UV-spectra analyses. All the strains studied, in particularB. schlegelii strains, released significant quantities of S⁰ into the growth media. In contrast, the intracellular S⁰ content was very low. Significant rhodanese activity in the presence of thiosulfate was measured with toluenepermeabilized cells and cell-free extracts ofB. schlegelii (type strain) andHydrogenobacter spp. (strain T3).     

In vivo morphogenesis and growth characteristics of phages CM (Myoviridae) virulent forRhizobium meliloti

FourRhizobium meliloti bacteriophages belonging to theMyoviridae family of tailed phages were studied. Burst sizes (50–100 virulent particles per infected cell), adsorption rates (2.6–4.1×10^–9 ml/min), and latent periods (2–4 h) made this group heterogeneous. However, these characteristics indicate an important infection ability compared with other rhizobiophages. In vivo morphogenesis of phage CM1, studied by electron microscopy, seems to have steps similar to those of some other tailed phages such as coliphages.     

In vitro study of the proteolytic activity of rumen anaerobic fungi

Abstract To better define the antigenic structure of the outer cell membranes of Legionellae, a panel of 6 monoclonal antibodies was raised against partially purified outer membranes of Legionella pneumophila serogroup 1, Corby strain. This study describes the purification and characterization of one of these monoclonal antibodies reacting with a 135-kDa protein, which was shown to be common to all 14 serogroups of Legionella pneumophila . It shows no cross-reactivity with 20 other Legionella species, or 9 other Gram-negative species tested by SDS-PAGE and Western blotting procedures. The epitope would appear to be predominantly surface exposed and, from preliminary detergent extraction studies, not peptidoglycan-associated.     

Identification of a UDP-glucosyltransferase conferring deoxynivalenol resistance in Aegilops tauschii and wheat

Aegilops tauschii is the diploid progenitor of the wheat D subgenome and a valuable resource for wheat breeding, yet, genetic analysis of resistance against Fusarium head blight (FHB) and the major Fusarium mycotoxin deoxynivalenol (DON) is lacking. We treated a panel of 147 Ae. tauschii accessions with either Fusarium graminearum spores or DON solution and recorded the associated disease spread or toxin-induced bleaching. A k-mer-based association mapping pipeline dissected the genetic basis of resistance and identified candidate genes. After DON infiltration nine accessions revealed severe bleaching symptoms concomitant with lower conversion rates of DON into the non-toxic DON-3-O-glucoside. We identified the gene AET5Gv20385300 on chromosome 5D encoding a uridine diphosphate (UDP)-glucosyltransferase (UGT) as the causal variant and the mutant allele resulting in a truncated protein was only found in the nine susceptible accessions. This UGT is also polymorphic in hexaploid wheat and when expressed in Saccharomyces cerevisiae only the full-length gene conferred resistance against DON. Analysing the D subgenome helped to elucidate the genetic control of FHB resistance and identified a UGT involved in DON detoxification in Ae. tauschii and hexaploid wheat. This resistance mechanism is highly conserved since the UGT is orthologous to the barley UGT HvUGT13248 indicating descent from a common ancestor of wheat and barley.