Characterization of Quantitative Trait Loci for Leaf Rust Resistance from CI 13227 in Three Winter Wheat Populations.

Leaf rust is a widespread foliar wheat disease causing substantial yield losses worldwide. Slow-rusting is "adult plant" resistance that significantly slows epidemic development and thereby reduces yield loss. Wheat accession CI 13227 was previously characterized as having slow-rusting resistance. To validate the quantitative trait loci (QTL) and develop diagnostic markers for slow rusting resistance in CI 13227, a new population of recombinant inbred lines (RILs) of CI 13227 × Everest was evaluated for latent period (LP), final severity (FS), area under disease progress curve (AUDPC), and infection type (IT) in greenhouses and genotyped using genotyping-by-sequencing (GBS). Four QTL were identified on chromosome arms 2BL, 2DS, 3BS, and 7BL, explaining 6.82 to 28.45% of the phenotypic variance for these traits. Seven kompetitive allele specific polymorphism (KASP) markers previously reported to be linked to the QTL in two other CI 13227 populations were validated. In addition, the previously reported QLr.hwwg-7AL was remapped to 2BL (renamed QLr.hwwg-2BL) after adding new markers in this study. Phenotypic data showed that the RILs harboring two or three of the QTL had a significantly longer LP. QLr.hwwg-2DS on 2DS showed a major effect on all rust resistance traits and was finely mapped to a 2.7 Mb interval by two newly developed flanking markers from exome capture. Three disease-resistance genes and two transporter genes were identified as the putative candidates for QLr.hwwg-2DS. The validated QTL can be used as slow rusting resistance resources and the markers developed in this study will be useful for marker-assisted selection.

Identifying novel sources of resistance to wheat stem sawfly in five wild wheat species.

BACKGROUND: The wheat stem sawfly (WSS, Cephus cinctus) is a major pest of wheat (Triticum aestivum) and can cause significant yield losses. WSS damage results from stem boring and/or cutting, leading to the lodging of wheat plants. Although solid-stem wheat genotypes can effectively reduce larval survival, they may have lower yields than hollow-stem genotypes and show inconsistent solidness expression. Because of limited resistance sources to WSS, evaluating diverse wheat germplasm for novel resistance genes is crucial. We evaluated 91 accessions across five wild wheat species (Triticum monococcum, T. urartu, T. turgidum, T. timopheevii, and Aegilops tauschii) and common wheat cultivars (T. aestivum) for antixenosis (host selection) and antibiosis (host suitability) to WSS. Host selection was measured as the number of eggs after adult oviposition, and host suitability was determined by examining the presence or absence of larval infestation within the stem. The plants were grown in the greenhouse and brought to the field for WSS infestation. In addition, a phylogenetic analysis was performed to determine the relationship between the WSS traits and phylogenetic clustering. RESULTS: Overall, Ae. tauschii, T. turgidum and T. urartu had lower egg counts and larval infestation than T. monococcum, and T. timopheevii. T. monococcum, T. timopheevii, T. turgidum, and T. urartu had lower larval weights compared with T. aestivum. CONCLUSION: This study shows that wild relatives of wheat could be a valuable source of alleles for enhancing resistance to WSS and identifies specific germplasm resources that may be useful for breeding. © 2024 The Authors. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

Skim exome capture genotyping in wheat.

Next-generation sequencing (NGS) technology advancements continue to reduce the cost of high-throughput genome-wide genotyping for breeding and genetics research. Skim sequencing, which surveys the entire genome at low coverage, has become feasible for quantitative trait locus (QTL) mapping and genomic selection in various crops. However, the genome complexity of allopolyploid crops such as wheat (Triticum aestivum L.) still poses a significant challenge for genome-wide genotyping. Targeted sequencing of the protein-coding regions (i.e., exome) reduces sequencing costs compared to whole genome re-sequencing and can be used for marker discovery and genotyping. We developed a method called skim exome capture (SEC) that combines the strengths of these existing technologies and produces targeted genotyping data while decreasing the cost on a per-sample basis compared to traditional exome capture. Specifically, we fragmented genomic DNA using a tagmentation approach, then enriched those fragments for the low-copy genic portion of the genome using commercial wheat exome baits and multiplexed the sequencing at different levels to achieve desired coverage. We demonstrated that for a library of 48 samples, ∼7-8× target coverage was sufficient for high-quality variant detection. For higher multiplexing levels of 528 and 1056 samples per library, we achieved an average coverage of 0.76× and 0.32×, respectively. Combining these lower coverage SEC sequencing data with genotype imputation using a customized wheat practical haplotype graph database that we developed, we identified hundreds of thousands of high-quality genic variants across the genome. The SEC method can be used for high-resolution QTL mapping, genome-wide association studies, genomic selection, and other downstream applications.

Cloning of the broadly effective wheat leaf rust resistance gene Lr42 transferred from Aegilops tauschii.

The wheat wild relative Aegilops tauschii was previously used to transfer the Lr42 leaf rust resistance gene into bread wheat. Lr42 confers resistance at both seedling and adult stages, and it is broadly effective against all leaf rust races tested to date. Lr42 has been used extensively in the CIMMYT international wheat breeding program with resulting cultivars deployed in several countries. Here, using a bulked segregant RNA-Seq (BSR-Seq) mapping strategy, we identify three candidate genes for Lr42. Overexpression of a nucleotide-binding site leucine-rich repeat (NLR) gene AET1Gv20040300 induces strong resistance to leaf rust in wheat and a mutation of the gene disrupted the resistance. The Lr42 resistance allele is rare in Ae. tauschii and likely arose from ectopic recombination. Cloning of Lr42 provides diagnostic markers and over 1000 CIMMYT wheat lines carrying Lr42 have been developed documenting its widespread use and impact in crop improvement.

Development of the Wheat Practical Haplotype Graph database as a resource for genotyping data storage and genotype imputation.

To improve the efficiency of high-density genotype data storage and imputation in bread wheat (Triticum aestivum L.), we applied the Practical Haplotype Graph (PHG) tool. The Wheat PHG database was built using whole-exome capture sequencing data from a diverse set of 65 wheat accessions. Population haplotypes were inferred for the reference genome intervals defined by the boundaries of the high-quality gene models. Missing genotypes in the inference panels, composed of wheat cultivars or recombinant inbred lines genotyped by exome capture, genotyping-by-sequencing (GBS), or whole-genome skim-seq sequencing approaches, were imputed using the Wheat PHG database. Though imputation accuracy varied depending on the method of sequencing and coverage depth, we found 92% imputation accuracy with 0.01× sequence coverage, which was slightly lower than the accuracy obtained using the 0.5× sequence coverage (96.6%). Compared to Beagle, on average, PHG imputation was ∼3.5% (P-value < 2 × 10-14) more accurate, and showed 27% higher accuracy at imputing a rare haplotype introgressed from a wild relative into wheat. We found reduced accuracy of imputation with independent 2× GBS data (88.6%), which increases to 89.2% with the inclusion of parental haplotypes in the database. The accuracy reduction with GBS is likely associated with the small overlap between GBS markers and the exome capture dataset, which was used for constructing PHG. The highest imputation accuracy was obtained with exome capture for the wheat D genome, which also showed the highest levels of linkage disequilibrium and proportion of identity-by-descent regions among accessions in the PHG database. We demonstrate that genetic mapping based on genotypes imputed using PHG identifies SNPs with a broader range of effect sizes that together explain a higher proportion of genetic variance for heading date and meiotic crossover rate compared to previous studies.

Genomic Selection in Preliminary Yield Trials in a Winter Wheat Breeding Program.

Genomic prediction (GP) is now routinely performed in crop plants to predict unobserved phenotypes. The use of predicted phenotypes to make selections is an active area of research. Here, we evaluate GP for predicting grain yield and compare genomic and phenotypic selection by tracking lines advanced. We examined four independent nurseries of F3:6 and F3:7 lines trialed at 6 to 10 locations each year. Yield was analyzed using mixed models that accounted for experimental design and spatial variations. Genotype-by-sequencing provided nearly 27,000 high-quality SNPs. Average genomic predictive ability, estimated for each year by randomly masking lines as missing in steps of 10% from 10 to 90%, and using the remaining lines from the same year as well as lines from other years in a training set, ranged from 0.23 to 0.55. The predictive ability estimated for a new year using the other years ranged from 0.17 to 0.28. Further, we tracked lines advanced based on phenotype from each of the four F3:6 nurseries. Lines with both above average genomic estimated breeding value (GEBV) and phenotypic value (BLUP) were retained for more years compared to lines with either above average GEBV or BLUP alone. The number of lines selected for advancement was substantially greater when predictions were made with 50% of the lines from the testing year added to the training set. Hence, evaluation of only 50% of the lines yearly seems possible. This study provides insights to assess and integrate genomic selection in breeding programs of autogamous crops.

Genotyping-by-Sequencing Derived High-Density Linkage Map and its Application to QTL Mapping of Flag Leaf Traits in Bread Wheat.

Winter wheat parents 'Harry' (drought tolerant) and 'Wesley' (drought susceptible) were used to develop a recombinant inbred population with future goals of identifying genomic regions associated with drought tolerance. To precisely map genomic regions, high-density linkage maps are a prerequisite. In this study genotyping-by- sequencing (GBS) was used to construct the high-density linkage map. The map contained 3,641 markers distributed on 21 chromosomes and spanned 1,959 cM with an average distance of 1.8 cM between markers. The constructed linkage map revealed strong collinearity in marker order across 21 chromosomes with POPSEQ-v2.0, which was based on a high-density linkage map. The reliability of the linkage map for QTL mapping was demonstrated by co-localizing the genes to previously mapped genomic regions for two highly heritable traits, chaff color, and leaf cuticular wax. Applicability of linkage map for QTL mapping of three quantitative traits, flag leaf length, width, and area, identified 21 QTLs in four environments, and QTL expression varied across the environments. Two major stable QTLs, one each for flag leaf length (Qfll.hww-7A) and flag leaf width (Qflw.hww-5A) were identified. The map constructed will facilitate QTL and fine mapping of quantitative traits, map-based cloning, comparative mapping, and in marker-assisted wheat breeding endeavors.

A novel QTL associated with dwarf bunt resistance in Idaho 444 winter wheat.

KEY MESSAGE: A novel QTL, Q.DB.ui-7DS, and the PCR-based markers identified in the current study will accelerate variety development for resistance to dwarf and common bunt of wheat. Dwarf bunt [Tilletia controversa J.G. Kühn [as 'contraversa'], in Rabenhorst, Hedwigia 13: 188 (1874)] is a destructive disease of wheat (Triticum aestivum L.) that reduces grain yield and quality. A number of distinct genes conferring resistance to dwarf bunt have been used by breeding programs for nearly 100 years. However, few markers were identified that can be used in selection of dwarf bunt resistance. A recombinant inbred line (RIL) population derived from the bunt-resistant germplasm, Idaho 444 (IDO444), and the susceptible cultivar, Rio Blanco, was evaluated for phenotypic reaction to dwarf bunt inoculation in four trials in two locations (USU and USDA) over 3 years. The population was genotyped with the Diversity Arrays Technology (DArT) and the Illumina Infinium 9K iSelect marker platforms. A total of three QTL were detected, and resistant alleles were from IDO444. QTL Q.DB.ui-7DS on 7DS was determined based on the location of a DArT marker wPt-2565 (X116197), which was consistently detected and explained 32 to 56 % of phenotypic variation among the four trials. QTL Q.DB.ui-1A on 1A was detected in three Utah State University (USU) trials and explained 11-15 % of phenotypic variation. QTL Q.DB.ui-2B on 2B was detected in two USU and one United States Department of Agriculture (USDA) trials and explained up to 6 % of phenotypic variation. Two PCR-based markers were developed based on the sequence of wPt-2565 and validated in the RIL population and used in genotyping of dwarf bunt differential lines, known resistance sources, and resistant cultivars.

Distribution of Cadmium, Iron, and Zinc in Millstreams of Hard Winter Wheat (Triticum aestivum L.).

Hard winter wheat (Triticum aestivum L.) is a major crop in the Great Plains of the United States, and our previous work demonstrated that wheat genotypes vary for grain cadmium accumulation with some exceeding the CODEX standard (0.2 mg kg(-1)). Previous reports of cadmium distribution in flour milling fractions have not included high cadmium grain. This study measured the distribution of cadmium, zinc, and iron in flour and bran streams from high cadmium (0.352 mg kg(-1)) grain on a pilot mill that produced 12 flour and four bran streams. Recovery in flour was substantially greater for cadmium (50%) than for zinc (31%) or iron (22%). Cadmium, zinc, and iron in the lowest mineral concentration flour stream, representing the purest endosperm fraction, were 52, 22, and 11%, respectively, of initial grain concentration. Our results indicate that, relative to zinc and iron, a greater proportion of cadmium is stored in the endosperm, the source of white flour.

Identification of milling and baking quality QTL in multiple soft wheat mapping populations.

KEY MESSAGE: Two mapping approaches were use to identify and validate milling and baking quality QTL in soft wheat. Two LG were consistently found important for multiple traits and we recommend the use marker-assisted selection on specific markers reported here. Wheat-derived food products require a range of characteristics. Identification and understanding of the genetic components controlling end-use quality of wheat is important for crop improvement. We assessed the underlying genetics controlling specific milling and baking quality parameters of soft wheat including flour yield, softness equivalent, flour protein, sucrose, sodium carbonate, water absorption and lactic acid, solvent retention capacities in a diversity panel and five bi-parental mapping populations. The populations were genotyped with SSR and DArT markers, with markers specific for the 1BL.1RS translocation and sucrose synthase gene. Association analysis and composite interval mapping were performed to identify quantitative trait loci (QTL). High heritability was observed for each of the traits evaluated, trait correlations were consistent over populations, and transgressive segregants were common in all bi-parental populations. A total of 26 regions were identified as potential QTL in the diversity panel and 74 QTL were identified across all five bi-parental mapping populations. Collinearity of QTL from chromosomes 1B and 2B was observed across mapping populations and was consistent with results from the association analysis in the diversity panel. Multiple regression analysis showed the importance of the two 1B and 2B regions and marker-assisted selection for the favorable alleles at these regions should improve quality.

Understanding the emergence of ebola virus disease in sierra leone: stalking the virus in the threatening wake of emergence.

Since Ebola Virus Disease (EVD) was first identified in 1976 in what is now the Democratic Republic of Congo, and despite the numerous outbreaks recorded to date, rarely has an epidemic origin been identified. Indeed, among the twenty-one most documented EVD outbreaks in Africa, an index case has been identified four times, and hypothesized in only two other instances. The initial steps of emergence and spread of a virus are critical in the development of a potential outbreak and need to be thoroughly dissected and understood in order to improve on preventative strategies. In the current West African outbreak of EVD, a unique index case has been identified, pinpointing the geographical origin of the epidemic in Guinea. Herein, we provide an accounting of events that serve as the footprint of EVD emergence in Sierra Leone and a road map for risk mitigation fueled by lessons learned.

A fusion-inhibiting peptide against Rift Valley fever virus inhibits multiple, diverse viruses.

For enveloped viruses, fusion of the viral envelope with a cellular membrane is critical for a productive infection to occur. This fusion process is mediated by at least three classes of fusion proteins (Class I, II, and III) based on the protein sequence and structure. For Rift Valley fever virus (RVFV), the glycoprotein Gc (Class II fusion protein) mediates this fusion event following entry into the endocytic pathway, allowing the viral genome access to the cell cytoplasm. Here, we show that peptides analogous to the RVFV Gc stem region inhibited RVFV infectivity in cell culture by inhibiting the fusion process. Further, we show that infectivity can be inhibited for diverse, unrelated RNA viruses that have Class I (Ebola virus), Class II (Andes virus), or Class III (vesicular stomatitis virus) fusion proteins using this single peptide. Our findings are consistent with an inhibition mechanism similar to that proposed for stem peptide fusion inhibitors of dengue virus in which the RVFV inhibitory peptide first binds to both the virion and cell membranes, allowing it to traffic with the virus into the endocytic pathway. Upon acidification and rearrangement of Gc, the peptide is then able to specifically bind to Gc and prevent fusion of the viral and endocytic membranes, thus inhibiting viral infection. These results could provide novel insights into conserved features among the three classes of viral fusion proteins and offer direction for the future development of broadly active fusion inhibitors.

Enhanced Efficacy of a Codon-Optimized DNA Vaccine Encoding the Glycoprotein Precursor Gene of Lassa Virus in a Guinea Pig Disease Model When Delivered by Dermal Electroporation.

Lassa virus (LASV) causes a severe, often fatal, hemorrhagic fever endemic to West Africa. Presently, there are no FDA-licensed medical countermeasures for this disease. In a pilot study, we constructed a DNA vaccine (pLASV-GPC) that expressed the LASV glycoprotein precursor gene (GPC). This plasmid was used to vaccinate guinea pigs (GPs) using intramuscular electroporation as the delivery platform. Vaccinated GPs were protected from lethal infection (5/6) with LASV compared to the controls. However, vaccinated GPs experienced transient viremia after challenge, although lower than the mock-vaccinated controls. In a follow-on study, we developed a new device that allowed for both the vaccine and electroporation pulse to be delivered to the dermis. We also codon-optimized the GPC sequence of the vaccine to enhance expression in GPs. Together, these innovations resulted in enhanced efficacy of the vaccine. Unlike the pilot study where neutralizing titers were not detected until after virus challenge, modest neutralizing titers were detected in guinea pigs before challenge, with escalating titers detected after challenge. The vaccinated GPs were never ill and were not viremic at any timepoint. The combination of the codon-optimized vaccine and dermal electroporation delivery is a worthy candidate for further development.

Variation for resistance to kernel infection and toxin accumulation in winter wheat infected with Fusarium graminearum.

Host resistance is the main way to control Fusarium head blight (FHB) in wheat. Despite improved levels of resistance to infection and spread in vegetative tissue, the toxin deoxynivalenol (DON) can still accumulate to unacceptable concentration levels. In this study, our objectives were to assess the genetic variation for resistance to kernel infection (RKI) and resistance to toxin accumulation (RTA) and their role in controlling DON. We collected spikes with different levels of visual symptoms from each of 32 wheat genotypes and at four environments and determined DON and fungal biomass (FB) from each sample. We assessed RKI by regressing FB on the level of visual symptoms and RTA by regressing DON on FB for each genotype. Significant genetic effects were found for RKI and RTA. Some genotypes consistently had low FB in their grain despite increasing visual symptoms suggesting RKI. Additionally, some genotypes consistently had low DON in their grain despite increasing FB levels suggesting a higher RTA in these genotypes. The variation for RKI and RTA explained a significant fraction of the variation for DON among genotypes with moderate visual symptoms using independent grain samples. Although RKI and RTA were significantly correlated (r = 0.58, P = 0.05), RTA was more predictive of DON accumulation because it modeled 32 to 44% of the genotype sum of squares for DON, while only 9 to 10% were predicted using RKI. Thus, variation for RTA was important in explaining variation for DON among genotypes with acceptable levels of resistance to fungal infection and spread. This work indicates that there is a need to develop a better understanding of RTA and rapid screening methods for this trait.

Evaluation of Lassa antiviral compound ST-193 in a guinea pig model.

Lassa virus (LASV), a member of the Arenaviridae family, causes a viral hemorrhagic fever endemic to West Africa, where as many as 300,000 infections occur per year. Presently, there are no FDA-approved LASV-specific vaccines or antiviral agents, although the antiviral drug ribavirin has shown some efficacy. A recently identified small-molecule inhibitor of arenavirus entry, ST-193, exhibits submicromolar antiviral activity in vitro. To determine the antiviral utility of ST-193 in vivo, we tested the efficacy of this compound in the LASV guinea pig model. Four groups of strain 13 guinea pigs were administered 25 or 80 mg/kg ST-193, 25 mg/kg of ribavirin, or the vehicle by the intraperitoneal (i.p.) route before infection with a lethal dose of LASV, strain Josiah, and continuing once daily for 14 days. Control animals exhibited severe disease, becoming moribund between days 10 and 15 postinfection. ST-193-treated animals exhibited fewer signs of disease and enhanced survival when compared to the ribavirin or vehicle groups. Body temperatures in all groups were elevated by day 9, but returned to normal by day 19 postinfection in the majority of ST-193-treated animals. ST-193 treatment mediated a 2-3-log reduction in viremia relative to vehicle-treated controls. The overall survival rate for the ST-193-treated guinea pigs was 62.5% (10/16) compared with 0% in the ribavirin (0/8) and vehicle (0/7) groups. These data suggest that ST-193 may serve as an improved candidate for the treatment of Lassa fever.

Uncoupling GP1 and GP2 expression in the Lassa virus glycoprotein complex: implications for GP1 ectodomain shedding.

BACKGROUND: Sera from convalescent Lassa fever patients often contains antibodies to Lassa virus (LASV) glycoprotein 1 (GP1), and glycoprotein 2 (GP2); Immunization of non-human primates with viral vectors expressing the arenaviral glycoprotein complex (GPC) confers full protective immunity against a lethal challenge with LASV. Thus, the development of native or quasi native recombinant LASV GP1 and GP2 as soluble, uncoupled proteins will improve current diagnostics, treatment, and prevention of Lassa fever. To this end, mammalian expression systems were engineered for production and purification of secreted forms of soluble LASV GP1 and GP2 proteins. RESULTS: Determinants for mammalian cell expression of secreted uncoupled Lassa virus (LASV) glycoprotein 1 (GP1) and glycoprotein 2 (GP2) were established. Soluble GP1 was generated using either the native glycoprotein precursor (GPC) signal peptide (SP) or human IgG signal sequences (s.s.). GP2 was secreted from cells only when (1) the transmembrane (TM) domain was deleted, the intracellular domain (IC) was fused to the ectodomain, and the gene was co-expressed with a complete GP1 gene in cis; (2) the TM and IC domains were deleted and GP1 was co-expressed in cis; (3) expression of GP1 was driven by the native GPC SP. These data implicate GP1 as a chaperone for processing and shuttling GP2 to the cell surface. The soluble forms of GP1 and GP2 generated through these studies were secreted as homogeneously glycosylated proteins that contained high mannose glycans. Furthermore, observation of GP1 ectodomain shedding from cells expressing wild type LASV GPC represents a novel aspect of arenaviral glycoprotein expression. CONCLUSION: These results implicate GP1 as a chaperone for the correct processing and shuttling of GP2 to the cell surface, and suggest that native GPC SP plays a role in this process. In the absence of GP1 and GPC SP the GP2 protein may be processed by an alternate pathway that produces heterogeneously glycosylated protein, or the polypeptide may not fully mature in the secretory cascade in mammalian cells. The expression constructs developed in these studies resulted in the generation and purification of soluble, uncoupled GP1 and GP2 proteins from mammalian cells with quasi-native properties. The observation of GP1 ectodomain shedding from cells expressing wild type LASV GPC establishes new correlates of disease progression and highlights potential opportunities for development of diagnostics targeting the early stages of Lassa fever.

Nonstarch polysaccharides in wheat flour wire-cut cookie making.

Nonstarch polysaccharides in wheat flour have significant capacity to affect the processing quality of wheat flour dough and the finished quality of wheat flour products. Most research has focused on the effects of arabinoxylans (AX) in bread making. This study found that water-extractable AX and arabinogalactan peptides can predict variation in pastry wheat quality as captured by the wire-cut cookie model system. The sum of water-extractable AX plus arabinogalactan was highly predictive of cookie spread factor. The combination of cookie spread factor and the ratio of water-extractable arabinose to xylose predicted peak force of the three-point bend test of cookie texture.

Bacterial-based systems for expression and purification of recombinant Lassa virus proteins of immunological relevance.

BACKGROUND: There is a significant requirement for the development and acquisition of reagents that will facilitate effective diagnosis, treatment, and prevention of Lassa fever. In this regard, recombinant Lassa virus (LASV) proteins may serve as valuable tools in diverse antiviral applications. Bacterial-based systems were engineered for expression and purification of recombinant LASV nucleoprotein (NP), glycoprotein 1 (GP1), and glycoprotein 2 (GP2). RESULTS: Full-length NP and the ectodomains of GP1 and GP2 were generated as maltose-binding protein (MBP) fusions in the Rosetta strains of Escherichia coli (E. coli) using pMAL-c2x vectors. Average fusion protein yields per liter of culture for MBP-NP, MBP-GP1, and MBP-GP2 were 10 mg, 9 mg, and 9 mg, respectively. Each protein was captured from cell lysates using amylose resin, cleaved with Factor Xa, and purified using size-exclusion chromatography (SEC). Fermentation cultures resulted in average yields per liter of 1.6 mg, 1.5 mg, and 0.7 mg of purified NP, GP1 and GP2, respectively. LASV-specific antibodies in human convalescent sera specifically detected each of the purified recombinant LASV proteins, highlighting their utility in diagnostic applications. In addition, mouse hyperimmune ascitic fluids (MHAF) against a panel of Old and New World arenaviruses demonstrated selective cross reactivity with LASV proteins in Western blot and enzyme-linked immunosorbent assay (ELISA). CONCLUSION: These results demonstrate the potential for developing broadly reactive immunological assays that employ all three arenaviral proteins individually and in combination.

Barley (Hordeum vulgare L.) inositol monophosphatase: gene structure and enzyme characteristics.

The cellular myo-inositol (Ins) pool is important to many metabolic and signaling pathways in plants. Ins monophosphatase (IMPase; EC 3.1.3.25) activity is essential for the de novo synthesis of myo-Inositol (Ins), and for recycling of Ins in Ins(1,4,5)P3. However, proteins encoded by at least one family of IMP genes also have L-galactose-1-P phosphatase activity important to ascorbic acid synthesis, indicating a bifunctionality that links these two branches of carbon metabolism. As part of research into the regulation of Ins synthesis and supply during seed development, the barley IMP-1 gene and gene products were studied. The 1.4 kb barley IMP-1 promoter contains one low temperature response element (RE), two heat shock REs, one gibberellin and two auxin REs, and five sugar REs. Barley IMP-1 is expressed in all tissues assayed, and expression levels were not greatly altered by abiotic stress treatments. Reduced use of Ins for Ins P6 synthesis in developing seed of barley low phytic acid (lpa) mutants results in Ins accumulation, and IMP-1 expression is reduced in proportion to the increase in Ins level. The barley recombinant enzyme had a lower Km, indicating higher affinity, for D/L-Ins(3)P1 (Km = 9.7 microM) as compared with reported Km (Ins P1) values for other eukaryotic IMPases (43-330 microM) or with a reported Km (L-Gal-1P) of 150 microM for a kiwifruit (Actinidia deliciosa) enzyme. These and other data indicate that the barley IMP-1 gene is regulated at least in part in response to Ins metabolic needs, and that the enzyme it encodes displays catalytic properties well suited for a role in Ins synthesis, in addition to other roles as an L-gal-1-P phosphatase important to ascorbate synthesis, or as an IMPase important to Ins(1,4,5)P3 signal recycling.

New opportunities for field research on the pathogenesis and treatment of Lassa fever.

Unlike many viral hemorrhagic fevers (VHFs), Lassa fever (LF) is not a rare disease that emerges only as sporadic cases or in outbreak form. Although surveillance is inadequate to determine the true incidence, up to 300,000 infections and 5000 deaths from LF are estimated to occur yearly. The highest incidence is in the "Mano River Union (MRU) countries" of Sierra Leone, Liberia, and Guinea. Although civil unrest in this region over the past two decades has impeded capacity building and research, new-found peace in recent years presents new opportunities. In 2004, the Mano River Union Lassa Fever Network (MRU LFN) was established to assist MRU countries in the development of national and regional surveillance, diagnosis, treatment, control, and prevention of LF. Here, we review the present literature on treatment and pathogenesis of LF and outline priorities for future research in the field made possible by the improved research capacity of the MRU LFN.