Mapping of Fhb2 on chromosome 6BS: a gene controlling Fusarium head blight field resistance in bread wheat (Triticum aestivum L.)

Fusarium head blight (FHB) is one of the most important fungal wheat diseases worldwide. Understanding the genetics of FHB resistance is key to facilitate the introgression of different FHB resistance genes into adapted wheat. The objective of this project was to study the FHB resistance QTL on chromosome 6B, quantify the phenotypic variation, and qualitatively map the resistance gene as a Mendelian factor. The FHB resistant parent BW278 (AC Domain*2/Sumai 3) was used as the source of the resistance allele. A large recombinant inbred line (RIL) mapping population was developed from the cross BW278/AC Foremost. The population segregated for three known FHB resistance QTL located on chromosomes 3BSc, 5A, and 6B. Molecular markers on chromosome 6B (WMC104, WMC397, GWM219), 5A (GWM154, GWM304, WMC415), and 3BS (WMC78, GWM566, WMC527) were amplified on approximately 1,440 F_2:7 RILs. The marker information was used to select 89 RILs that were fixed homozygous susceptible for the 3BSc and 5A FHB QTLs and were recombinant in the 6B interval. Disease response was evaluated on 89 RILs and parental checks in the greenhouse and field nurseries. Dual floret injection (DFI) was used in greenhouse trials to evaluate disease severity (DS). Macroconidial spray inoculations were used in field nurseries conducted at two locations in southern Manitoba (Carman and Glenlea) over two years 2003 and 2004, to evaluate disease incidence, disease severity, visual rating index, and Fusarium-damaged kernels. The phenotypic distribution for all five-disease infection measurements was bimodal, with lines resembling either the resistant or susceptible checks and parents. All of the four field traits for FHB resistance mapped qualitatively to a coincident position on chromosome 6BS, flanked by GWM133 and GWM644, and is named Fhb2. The greenhouse-DS trait mapped 2 cM distal to Fhb2. Qualitative mapping of Fhb2 in wheat provides tightly linked markers that can reduce linkage drag associated with marker assisted selection of Fhb2 and aid the pyramiding of different resistance loci for wheat improvement.     

Parasitism of Tuta absoluta (Lepidoptera, Gelechiidae) by Pseudapanteles dignus (Hymenoptera, Braconidae) under laboratory conditions

Laboratory studies were conducted to measure selected life history traits and the functional response of the parasitoid Pseudapanteles dignus (Muesebeck), a major enemy of Tuta absoluta (Meyrick) in tomato crops in South America. Newly mated P. dignus females were individually exposed to 10 host larvae in mines for 24 h. We determined developmental time from egg to pupal formation and pupal stage duration, female adult life span, fecundity, reproductive period, daily parasitism rate, and sex ratio of offspring. For the functional response experiment, treatments consisted of six host densities: 3, 5, 7, 10, 15, 20, or 30 larvae. The number and proportion of parasitized hosts were calculated for each density. The shape of the functional response curve was analyzed by logistic regression. P. dignus females attacked hosts daily, exhibiting modest lifetime fecundity (approximately 32 parasitized hosts per female) and a female-biased offspring sex ratio. Female adult life span was 36 d. P. dignus showed a type I functional response within the range of host densities tested. We observed that females detect and parasitize the host within a wide range of densities, including low densities. The functional response curve reached an asymptote at a mean density of six hosts per day and seemed not to be egg-limited. Percent parasitism was approximately 30%. The ecological implications of the results in relation to the potential of P. dignus for the biological control of T. absoluta in tomato are discussed.     

Associations of fire ant phorids and microhabitats

We examined flight activity patterns for a guild of fire ant parasitoids in western Argentina in relationship to their host's location (mound/foraging trail) and light condition (full sun/partial sun/full shade) at different scales, from the individually sampled mound to the full day's summation for each species. We asked first whether taxa showed preferences among these conditions, and second, whether certain species and sexes might be found together more frequently than expected to by chance. All species, except the P. obtusus species complex, were significantly more likely to be found attacking ants at disturbed mounds than at paired foraging trails. The P. nocens complex and P. litoralis were more likely to be in the shade when temperatures were above the overall mean of the study (28.3 degrees C), whereas others, such as the P. obtusus complex and P. tricuspis, were more likely to be in full sun under these same conditions. Our analyses indicated that a limited set of species, particularly P. nocens with P. litoralis, and males with female P. obtusus and P. tricuspis, were more likely to be found together than expected. We also found decreasing proportions of males with increasing time of analysis. We discuss the implications of host location, metereological conditions, and sex ratios in relationship to ongoing classical biological control efforts using species of these phorids.     

Rotavirus and adenovirus in Rondônia

Acute gastroenteritis is one of the most common diseases in humans worldwide. Viral gastroenteritis is a global problem in infants and young children. In this study the incidence of diarrhea was assessed in 877 hospitalized children under five years old, over a period of 24 months and distributed in 470 cases of diarrhea and 407 age-matched group with other pathologies, as control group. Two antigen detection techniques based on enzyme immunoassay (EIA) and latex particles were used for detection of rotavirus and adenovirus. Rotavirus A was a major cause of gastroenteritis with 23.6% of cases, being 90% of these cases in young children. Adenovirus infections was detected by EIA with frequency of 6.4%. Rotavirus and adenovirus were detected in 10.1 and 1.7% of stools from control group, respectively. Interestingly, the frequency of the youngest children in the control group excreting Rotavirus A was comparable to that detected in stools from diarrheic children. We cannot rule out the existence of other enteric viruses because the etiology of 171 cases of diarrhea was not determined and active search for astrovirus and calicivirus was not done. This is the first study that shows the presence of enteric viruses in the infantile population from Western Brazilian Amazonia and it was important to help physicians in the treatment of viral gastroenteritis.     

Hepatic leukocyte recruitment in response to time-limited expression of TNF-alpha and IL-1beta

The development of chronic liver diseases is mediated by sustained hepatic inflammation. Our objective was to characterize the molecular mechanisms responsible for the hepatic inflammatory response to time-limited TNF-alpha and IL-1beta expression. C57Bl/6 mice were injected with 2 x 10(7) plaque forming units intraperitoneally of an adenoviral vector containing TNF-alpha or IL-1beta (AdTNF-alpha or AdIL-1beta). A nonreplicating adenoviral vector served as control. Four days later, under ketamine and xylazine anesthesia, the liver microvasculature was examined by intravital microscopy. In the postsinusoidal venules, leukocyte rolling increased significantly in response to both AdTNF-alpha and AdIL-1beta, compared with controls. This response was significantly reduced following injection of an anti-alpha4-integrin monoclonal antibody (MAb). Postsinusoidal rolling was further reduced to baseline following injection of an anti-P-selectin or anti-L-selectin MAb. Sinusoidal adhesion was greater in mice treated with AdIL-1beta than with AdTNF-alpha. Blocking alpha4-integrin, P-selectin, or L-selectin had no significant effect on sinusoidal or postsinusoidal adhesion. In separate experiments, we administered AdTNF-alpha or AdIL-1beta to mice deficient in ICAM-1. In ICAM-1-/- mice, postsinusoidal leukocyte rolling significantly increased following expression of IL-1beta but not TNF-alpha. AdIL-1beta- but not AdTNF-alpha-mediated sinusoidal adhesion was ICAM-1 dependent. AdTNF-alpha-induced sinusoidal adhesion was significantly reduced following 4 days of anti-MIP-2 MAb and anti-KC MAb. Prolonged expression of the cytokines TNF-alpha and IL-1beta increases hepatic leukocyte-endothelial cell interactions. Interestingly, the mechanisms through which these cytokines bring about adhesion within the sinusoids differ; AdIL-1beta sinusoidal adhesion uses an ICAM-1-dependent mechanism whereas AdTNF-alpha-mediated adhesion is ICAM-1 independent but CXC chemokine dependent.     

<Emphasis Type="Italic">Bacillus megaterium</Emphasis>—from simple soil bacterium to industrial protein production host

Bacillus megaterium has been industrially employed for more than 50 years, as it possesses some very useful and unusual enzymes and a high capacity for the production of exoenzymes. It is also a desirable cloning host for the production of intact proteins, as it does not possess external alkaline proteases and can stably maintain a variety of plasmid vectors. Genetic tools for this species include transducing phages and several hundred mutants covering the processes of biosynthesis, catabolism, division, sporulation, germination, antibiotic resistance, and recombination. The seven plasmids of B. megaterium strain QM B1551 contain several unusual metabolic genes that may be useful in bioremediation. Recently, several recombinant shuttle vectors carrying different strong inducible promoters and various combinations of affinity tags for simple protein purification have been constructed. Leader sequences-mediated export of affinity-tagged proteins into the growth medium was made possible. These plasmids are commercially available. For a broader application of B. megaterium in industry, sporulation and protease-deficient as well as UV-sensitive mutants were constructed. The genome sequence of two different strains, plasmidless DSM319 and QM B1551 carrying seven natural plasmids, is now available. These sequences allow for a systems biotechnology optimization of the production host B. megaterium. Altogether, a “toolbox” of hundreds of genetically characterized strains, genetic methods, vectors, hosts, and genomic sequences make B. megaterium an ideal organism for industrial, environmental, and experimental applications.     

A dual expression platform to optimize the soluble production of heterologous proteins in the periplasm of <Emphasis Type="Italic">Escherichia coli</Emphasis>

The functional analysis of individual proteins or of multiprotein complexes—since the completion of several genome sequencing projects—is in focus of current scientific work. Many heterologous proteins contain disulfide-bonds, required for their correct folding and activity, and therefore, need to be transported to the periplasm. The production of soluble and functional protein in the periplasm often needs target-specific regulatory genetic elements, leader peptides, and folding regimes. Usually, the optimization of periplasmic expression is a step-wise and time-consuming procedure. To overcome this problem we developed a dual expression system, containing a degP-promoter-based reporter system and a highly versatile plasmid set. This combines the differential protein expression with the selection of a target-specific expression plasmid. For the validation of this expression tool, two different molecular formats of a recombinant antibody directed to the human epidermal growth factor receptor and human 11β-hydroxysteroid dehydrogenase type 2 (11β-HSD2) were used. By application of this expression system we demonstrated that the amount of functional protein is inversely proportional to the on-line luciferase signal. We showed that this technology offers a simple tool to evaluate and improve the yield of functionally expressed proteins in the periplasm, which depends on the used regulatory elements and folding strategies.