Clubroot resistance QTL are modulated by nitrogen input in <Emphasis Type="Italic">Brassica napus</Emphasis>

Key message

Nitrogen levels can modulate the effectiveness of clubroot resistance in an isolate- and host-specific manner. While the same QTL were detected under high and low nitrogen, their effects were altered.

Abstract

Clubroot, caused by Plasmodiophora brassicae, is one of the most damaging diseases of oilseed rape and is known to be affected by nitrogen fertilization. However, the genetic factors involved in clubroot resistance have not been characterized under nitrogen-limiting conditions. This study aimed to assess the variability of clubroot resistance under different nitrogen levels and to characterize the impact of nitrogen supply on genetic resistance factors. Linkage analyses and a genome-wide association study were conducted to detect QTL for clubroot resistance and evaluate their sensitivity to nitrogen. The clubroot response of a set of 92 diverse oilseed rape accessions and 108 lines derived from a cross between ‘Darmor-bzh’ (resistant) and ‘Yudal’ (susceptible) was studied in the greenhouse under high- and low-nitrogen conditions, following inoculation with the P. brassicae isolates eH and K92-16. Resistance to each isolate was controlled by a major QTL and a few small-effects QTL. While the same QTL were detected under both high and low nitrogen, their effects were altered. Clubroot resistance to isolate eH, but not K92-16, was greater under a low-N supply versus a high-N supply. New sources of resistance were found among the oilseed rape accessions under both low and high-N conditions. The results are discussed relative to the literature and from a crop improvement perspective.

     

Alkane composition diversity among populations of dwarf forms of Juniperus communis L.: comparison between western Europe and northern American populations

The cuticular wax composition of leaves has been analysed in three western European populations (Corsica, central Pyrenees, northern Alps) of Juniperus communis var. saxatilis Pall. (=  J. nana Willd., nom illeg.) and in one population of J. communis L. var. depressa Pursh. from North America (Sierra Nevada). Gas chromatography shows the presence of 13 alkanes in all samples ranging from C₂₃ to C₃₅ with important intraspecific polymorphism in alkane content. The dominant alkanes range from C₃₃ to C₃₅. Alkanes C₂₁ and C₂₂ were found only in Corsica and Sierra Nevada populations. Canonical discriminant analysis separated the J. communis L. var. depressa Pursh. of the population of Sierra Nevada from other populations of J. communis var. saxatilis Pall. on the basis of their higher C₃₁ content and the constant presence of C₂₁ and C₂₂ alkanes. J. communis var. saxatilis Pall. populations from the Pyrenees are close to northern Alps populations characterized by high concentrations of C₃₃, C₃₄ and C₃₅ alkanes. This paper confirms the existence of Juniperus var. saxatilis Pall. in the Pyrenees (France).  © The Linnean Society of London, Botanical Journal of the Linnean Society , 2002, 140 , 165–168.     

<i>Trichoderma</i> spp. fostering growth on <i>Capsicum chinense</i> Jacq. seedlings and antagonistic against <i>Meloidogyne incognita</i>

Fourteen native strains of Trichoderma spp. from wildand agricultural pathosystems in the state of Yucatan, Mexico, with growth-promoting ability of Capsicum chinense Jacq. seedlings were evaluated and antagonistic effect of their filtrate against second-stage juveniles (J2) of Meloidogyne incognita. The strains Th05-02 and Th27-08 showed the best significant effects on plant hight variable increments 55.57 and 47.62%, theTh07-04 with 29.48% more root length, theTh02-01 and Th07-04 isolates increased from 48.71 to 84.61% in volume radical and 53.40% of total dry biomass. Statistical analysis (p≤0.001) of Th43 and Th43-13-14 filtrates caused 100% mortality at 24 and 48h. In the test of reversibility to 24 h after replacing the filtrates Th43-13, Th43-14, TH09-06 and TH20-07 by sterile distilled water, the J2 did not recover their viability, so they were considered as the best potential strains of Trichoderma spp. with antagonistic capacity in J2 of M.incognita.     

A new ant genus from the Greater Antilles and Central America,Zatania (Hymenoptera: Formicidae), exemplifies the utility of male and molecular character systems

The ant genus Prenolepis (Hymenoptera: Formicidae) is the nominal member of the recently established Prenolepis genus‐group within the subfamily Formicinae. Our molecular phylogenetic analyses using fragments from five nuclear genes (arginine kinase, carbomoylphosphate synthase, elongation factor 1‐alpha F1, elongation factor 1‐alpha F2, wingless) and one mitochondrial gene (cytochrome oxidase I) indicate that this genus is polyphyletic. Although the majority of Prenolepis species was found to belong to the same monophyletic group (Prenolepis sensu stricto), a smaller subset of Prenolepis species, all found in either Central America or the Greater Antilles, was robustly inferred to comprise a distinct lineage that is sister to the Old World genus Paraparatrechina. Here we describe this newly discovered lineage within the larger Prenolepis genus‐group clade. The genus Zatania, gen.n. is composed of five extant species (Zatania albimaculata, Zatania cisipa, Zatania gibberosa, Zatania gloriosa, sp.n. and Zatania karstica) and one Dominican amber fossil species (Zatania electra^?, sp.n. ). These are medium‐sized ants (generally between 2.5 and 3 mm in total length) that are characterized by having long scapes and legs, and elongated mesosomata. A reliance on worker‐based taxonomy has previously prevented the discovery of this new lineage because of worker convergence consisting of various combinations of elongated mesosomata, long scapes and legs, and a constriction immediately behind the pronotum, observed in several distinct lineages within the Prenolepis genus‐group. However, we did find that male morphology complements our molecular results in revealing important diagnostic and potentially phylogenetically informative characters. Our study highlights the value for ant systematics to expand beyond its traditional foundation of worker‐based morphology and embrace character systems from other castes and molecular data.     

Heterologous expression of bacteriocin E-760 in Chlamydomonas reinhardtii and functional analysis

The use of antimicrobial peptides (AMPs) synthesized by bacteria (bacteriocins) is an alternative for combating multidrug resistant bacterial strains and their production by recombinant route is a viable option for their mass production. The bacteriocin E-760 isolated from the genus Enterococcus sp. has been shown to possess inhibitory activity against Gram-negative and Gram-positive bacteria. In this study, the expression of a chimeric protein coding for E-760 in the nucleus of C. reinhardtii was evaluated, as well as, its antibacterial activity. The synthetic gene E-760S was inserted into the genome of C. reinhardtii using Agrobacterium tumefaciens. A transgenic line was identified in TAP medium with hygromycin and also by PCR. The increment in the culture medium temperature of the transgenic strain at 35 °C for 10 minutes, increased the production level of the recombinant protein from 0.14 (Noninduced culture, NIC) to 0.36% (Induced culture, IC) of total soluble proteins (TSP); this was quantified by an ELISA assay. Recombinant E-760 possesses activity against Staphylococcus aureus in 0.34 U log, Streptococcus agalactiae in 0.48 U log, Enterococcus faecium in 0.36 U log, Pseudomonas aeruginosa in 2 U log and for Klebsiella pneumoniae, the activity was 0.07 U log. These results demonstrate that the nucleus transformation of C. reinhardtii can function as a stable expression platform for the production of the synthetic gene E-760 and it can potentially be used as an antibacterial agent.     

Optimizing Production of Antigens and Fabs in the Context of Generating Recombinant Antibodies to Human Proteins

We developed and optimized a high-throughput project workflow to generate renewable recombinant antibodies to human proteins involved in epigenetic signalling. Three different strategies to produce phage display compatible protein antigens in bacterial systems were compared, and we found that in vivo biotinylation through the use of an Avi tag was the most productive method. Phage display selections were performed on 265 in vivo biotinylated antigen domains. High-affinity Fabs (<20nM) were obtained for 196. We constructed and optimized a new expression vector to produce in vivo biotinylated Fabs in E. coli. This increased average yields up to 10-fold, with an average yield of 4 mg/L. For 118 antigens, we identified Fabs that could immunoprecipitate their full-length endogenous targets from mammalian cell lysates. One Fab for each antigen was converted to a recombinant IgG and produced in mammalian cells, with an average yield of 15 mg/L. In summary, we have optimized each step of the pipeline to produce recombinant antibodies, significantly increasing both efficiency and yield, and also showed that these Fabs and IgGs can be generally useful for chromatin immunoprecipitation (ChIP) protocols.     

SUMO modification of cell surface Kv2.1 potassium channels regulates the activity of rat hippocampal neurons

Voltage-gated Kv2.1 potassium channels are important in the brain for determining activity-dependent excitability. Small ubiquitin-like modifier proteins (SUMOs) regulate function through reversible, enzyme-mediated conjugation to target lysine(s). Here, sumoylation of Kv2.1 in hippocampal neurons is shown to regulate firing by shifting the half-maximal activation voltage (V(1/2)) of channels up to 35 mV. Native SUMO and Kv2.1 are shown to interact within and outside channel clusters at the neuronal surface. Studies of single, heterologously expressed Kv2.1 channels show that only K470 is sumoylated. The channels have four subunits, but no more than two non-adjacent subunits carry SUMO concurrently. SUMO on one site shifts V(1/2) by 15 mV, whereas sumoylation of two sites produces a full response. Thus, the SUMO pathway regulates neuronal excitability via Kv2.1 in a direct and graded manner.     

Contributions of the hippocampus and medial prefrontal cortex to energy and body weight regulation

The effects of selective ibotenate lesions of the complete hippocampus (CHip), the hippocampal ventral pole (VP), or the medial prefrontal cortex (mPFC) in male rats were assessed on several measures related to energy regulation (i.e., body weight gain, food intake, body adiposity, metabolic activity, general behavioral activity, conditioned appetitive responding). The testing conditions were designed to minimize the nonspecific debilitating effects of these surgeries on intake and body weight. Rats with CHip and VP lesions exhibited significantly greater weight gain and food intake compared with controls. Furthermore, CHip-lesioned rats, but not rats with VP lesions, showed elevated metabolic activity, general activity in the dark phase of the light-dark cycle, and greater conditioned appetitive behavior, compared with control rats without these brain lesions. In contrast, rats with mPFC lesions were not different from controls on any of these measures. These results indicate that hippocampal damage interferes with energy and body weight regulation, perhaps by disrupting higher-order learning and memory processes that contribute to the control of appetitive and consummatory behavior.     

Denitrification in field soils

Summary Recent denitrification research is reviewed to answer questions a) how much N is lost from the soil as N₂ and N₂O and b) how do agronomic practices affect this loss? The methods used to quantify denitrification are also discussed. Gaseous losses of inorganic N range between the equivalent of 0 to 20 per cent of the fertilizer N applied to arable soils and 0–7 per cent on grassland soils. Losses are greater on undrained land and also after using direct drilling to establish arable crops.Appendix 1 summarizes reported measurements of gaseous N losses.Introductory lecture     

Duplication of pilus gene complexes of Haemophilus influenzae biogroup aegyptius.

Brazilian purpuric fever (BPF) is a recently described pediatric septicemia caused by a strain of Haemophilus influenzae biogroup aegyptius. The pilus specified by this bacterium may be important in BPF pathogenesis, enhancing attachment to host tissue. Here, we report the cloning of two haf (for H. influenzae biogroup aegyptius fimbriae) gene clusters from a cosmid library of strain F3031. We sequenced a 6.8-kb segment of the haf1 cluster and identified five genes (hafA to hafE). The predicted protein products, HafA to HafD, are 72, 95, 98, and 90% similar, respectively, to HifA to HifD of the closely related H. influenzae type b pilus. Strikingly, the putative pilus adhesion, HifE, shares only 44% identity with HafE, suggesting that the proteins may differ in receptor specificity. Insertion of a mini-gammadelta transposon in the hafE gene eliminated hemadsorption. The nucleotide sequences of the haf1 and haf2 clusters are more than 99% identical. Using the recently published sequence of the H. influenzae Rd genome, we determined that the haf1 complex lies at a unique position in the chromosome between the pmbA gene and a hypothetical open reading frame, HI1153. The location of the haf2 cluster, inserted between the purE and pepN genes, is analogous to the hif genes on H. influenzae type b. BPF fimbrial phase switching appears to involve slip-strand mispairing of repeated dinucleotides in the pilus promoter. The BPF-associated H. influenzae biogroup aegyptius pilus system generally resembles other H. influenzae, but the possession of a second fimbrial gene cluster, which appears to have arisen by a recent duplication event, and the novel sequence of the HafE adhesin may be significant in the unusual pathogenesis of BPF.