Identification of genomic loci conferring broad-spectrum resistance to multiple nematode species in exotic soybean accession PI 567305.

KEY MESSAGE: Genetic analysis identified a unique combination of major QTL for resistance to important soybean nematodes concurrently present in a single soybean accession, which has not been reported earlier. An exotic soybean [Glycine max (L.) Merr.] accession, PI 567305, was reported to be highly resistant to three important nematode species, soybean cyst (SCN), root-knot (RKN), and reniform (RN) nematodes. However, genetic basis controlling broad-spectrum resistance in this germplasm has not been investigated. We report results of genetic analysis to identify genomic loci conferring resistance to these nematode species. A bi-parental population consisting of 242 F8-derived recombinant inbred lines (RILs) was developed from a cross of a nematode susceptible cultivar, Magellan, and resistant accession, PI 567305. The RILs were phenotyped for nematode resistance to three SCN HG types. They were genotyped using the Infinium SoySNP6K BeadChips and genotype-by-sequencing (GBS) methods in an attempt to evaluate the cost-effectiveness and efficiency of these two genotyping platforms. Genetic analysis confirmed the major QTL on chromosomes (Chrs) 10 and 18 with broad-spectrum resistance to the three nematodes present in this germplasm. Haplotype and copy number variation analyses of SCN resistance QTL indicated that PI 567305 has a different haplotype, which is associated with likely a unique SCN resistance mechanism different from Peking- or PI 88788-type resistance. The evaluations of both Infinium Beadchip- and GBS-based genotyping technologies provided comprehensive insights for researchers to choose a cost-effective and efficient platform for QTL mapping and for other genomic studies in soybeans.

Genetic architecture of cyst nematode resistance revealed by genome-wide association study in soybean.

BACKGROUND: Bi-parental mapping populations have been commonly utilized to identify and characterize quantitative trait loci (QTL) controlling resistance to soybean cyst nematode (SCN, Heterodera glycines Ichinohe). Although this approach successfully mapped a large number of SCN resistance QTL, it captures only limited allelic diversity that exists in parental lines, and it also has limitations for genomic resolution. In this study, a genome-wide association study (GWAS) was performed using a diverse set of 553 soybean plant introductions (PIs) belonging to maturity groups from III to V to detect QTL/genes associated with SCN resistance to HG Type 0. RESULTS: Over 45,000 single nucleotide polymorphism (SNP) markers generated by the SoySNP50K iSelect BeadChip (http// www.soybase.org ) were utilized for analysis. GWAS identified 14 loci distributed over different chromosomes comprising 60 SNPs significantly associated with SCN resistance. Results also confirmed six QTL that were previously mapped using bi-parental populations, including the rhg1 and Rhg4 loci. GWAS identified eight novel QTL, including QTL on chromosome 10, which we have previously mapped by using a bi-parental population. In addition to the known loci for four simple traits, such as seed coat color, flower color, pubescence color, and stem growth habit, two traits, like lodging and pod shattering, having moderately complex inheritance have been confirmed with great precision by GWAS. CONCLUSIONS: The study showed that GWAS can be employed as an effective strategy for identifying complex traits in soybean and for narrowing GWAS-defined genomic regions, which facilitates positional cloning of the causal gene(s).

The breeding systems of diploid and neoautotetraploid clones of Acacia mangium Willd. in a synthetic sympatric population in Vietnam.

Colchicine-induced neoautotetraploid genotypes of Acacia mangium were cloned and planted in mixture with a set of diploid clones in an orchard in southern Vietnam. Following good general flowering, open-pollinated seed was collected from trees of both cytotypes and microsatellite markers were used to determine the breeding system as characterised by the proportion of outcrosses in young seedling progeny. As predicted from the literature, the progeny of diploid clones were predominantly outcrossed (t(m) = 0.97). In contrast, the progeny of the tetraploid clones were almost entirely selfs (t(m) = 0.02; 3 of 161 seedlings assayed were tetraploid outcrosses and there were no triploids). Segregation at loci heterozygous in the tetraploid mothers followed expected ratios, indicating sexual reproduction rather than apomixis. Post-zygotic factors are primarily responsible for divergence of the breeding systems. Commonly, less than 1 % of Acacia flowers mature as a pod, and after mixed pollination, diploid outcrossed seed normally develops at the expense of selfs. Selfs of the tetraploid trees appear to express less genetic load and have a higher probability of maturing. However, this does not fully explain the observed deficiency of outcross tetraploid progeny. Presumably, there are cytogenetic reasons which remain to be investigated. In nature, selfing would increase the probability of establishment of neotetraploids irrespective of cytotype frequency in the population. Breeders need to review their open-pollinated breeding and seed production strategies. It remains to be seen whether this is an ephemeral problem, with strong fertility selection restoring potential for outcrossing over generations.

The development of an immunoassay for the detection of artemisinin compounds in urine.

We have produced monoclonal antibodies against artelinic acid and investigated the reactivity with artemisinin drugs and metabolites. Antibody F170-10 is fairly specific for artelinic acid but does bind artemisinin and artemether (3-5% cross-reactivity). Dihydroartemisinin, artesunate, and metabolites of artemisinin showed less reactivity. With this antibody, an inhibition ELISA has been set up to detect artemisinin compounds in urine. In healthy subjects who received a single oral dose of artemisinin, artemether, artesunate or dihydroartemisinin, ELISA reactivity in urine was found. This reactivity in urine paralleled the plasma concentrations of artemether and dihydroartemisinin. The results show that this immunoassay for artelinic acid can be used to detect artemisinin compounds in urine for about 8 hr after intake. With a more sensitive test, this simple method as a urine dipstick may be become useful for drug use and compliance studies in malaria-endemic areas where the artemisinin derivatives are increasingly used.

Evaluation of Soybean Resistance to Sclerotinia Stem Rot Using Reciprocal Grafting.

Sclerotinia stem rot of soybean is one of the major soybean diseases in the north central region of the United States. One disease management option is to plant cultivars that have resistance. Some sources of partial resistance have been identified, but information pertaining to the nature of resistance is limited. The objective of this study was to determine if the expression of resistance is dictated by shoots of resistant plants and if this can be altered by using resistant and susceptible soybean genotypes grafted in different shoot and rootstock combinations of self-, single-, or double-shoot grafts. After successful grafts were made, several experiments were conducted using different inoculation techniques and soybean genotypes. In one experiment, cotyledons were inoculated with a plug of fungal mycelium, plants were incubated in a mist chamber for 23 h, and plant survival was recorded over time. Based on seven grafting combinations of cross- and self-grafted plants using two soybean cultivars, grafts with NKS19-90 (partially resistant) as shoots had greater (P ≤ 0.05) plant survival at 3, 4, and 5 days after inoculation than the other graft combinations. In another experiment, a total of 17 graft combinations were generated using resistant plant introductions and two susceptible cultivars. Resistant self-grafts of the plant introductions had greater (P ≤ 0.05) plant survival (mean = 75%) than self-grafts of the susceptible cultivars (mean = 15%) at 5 days after inoculation. Inter-genotypic grafts with resistant shoots had greater (P ≤ 0.05) plant survival (mean = 65%) than those in reciprocal combinations (mean = 8%) 5 days after inoculation. A cut stem inoculation method was used to test graft combinations of one resistant and two susceptible cultivars. Grafts with susceptible shoots of cvs. Williams 82 and Asgrow 2242 had greater (P < 0.05) lesion lengths (mean = 13.2 cm) than shoots of NKS19-90 (mean = 9.2 cm) regardless of the rootstock 15 days after inoculation. In a double-graft experiment, shoots of both NKS19-90 and Williams 82 were grafted to either NKS19-90 or Williams 82 rootstocks. Regardless of the rootstock, the shoots of Williams 82 died while shoots of NKS19-90 survived. For all the experiments, resistance was greater when the grafted shoot came from a resistant source on a susceptible rootstock compared with the reciprocal combination regardless of the type of grafting technique or inoculation method.

Hypoalbuminemia increases lysophosphatidylcholine in low-density lipoprotein of normocholesterolemic subjects.

BACKGROUND: A phospholipid, lysophosphatidylcholine (LPC), is the major determinant of the atherosclerotic properties of oxidized low-density lipoprotein (LDL). Under normal circumstances most LPC is bound to albumin. We hypothesized that lipoprotein LPC concentrations are increased in hypoalbuminemic patients with the nephrotic syndrome, irrespective of their lipid levels. To test this hypothesis, we selected nephrotic and control subjects with matched LDL cholesterol levels. METHODS: Lipoproteins and the albumin-rich lipoprotein-deficient fractions were separated by ultracentrifugation and their phospholipid composition was analyzed by thin-layer chromatography. RESULTS: Nephrotic subjects (albumin 23 +/- 2 g/liter and LDL cholesterol 3.1 +/- 0.2 mmol/liter) had a LDL LPC concentration that was increased (P < 0.05) to 66 +/- 7 vs. 35 +/- 6 micromol/liter in matched controls (albumin 42 +/- 5 g/liter and LDL cholesterol 3.1 +/- 0.2 mmol/liter). LPC in very low-density lipoprotein plus intermediate-density lipoprotein (VLDL + IDL) in these subjects was also increased to 33 +/- 7 vs. 9 +/- 2 micromol/liter in controls (P < 0.05). Conversely, LPC was decreased to 19 +/- 4 micromol/liter in the albumin-containing fraction of these hypoalbuminemic patients, as compared to 46 +/- 10 micromol/liter in the controls (P < 0.05). LPC was also low (14 +/- 4 micromol/liter) in the albumin-containing fraction of hypoalbuminemic, hypocholesterolemic patients with nonrenal diseases. In hyperlipidemic nephrotic subjects (albumin 21 +/- 2 g/liter and LDL cholesterol 5.7 +/- 0.5 mmol/liter) the LPC levels in LDL and VLDL + IDL were further increased, to 95 +/- 20 and 56 +/- 23 micromol/liter, respectively (P < 0.05). CONCLUSION: These findings suggest that in the presence of hypoalbuminemia in combination with proteinuria, LPC shifts from albumin to VLDL, IDL and LDL. This effect is independent of hyperlipidemia. Increased LPC in lipoproteins may be an important factor in the disproportionate increase in cardiovascular disease in nephrotic patients with hypoalbuminemia.

Albumin restores lysophosphatidylcholine-induced inhibition of vasodilation in rat aorta.

BACKGROUND: Impairment of vasodilation by oxidized low-density lipoprotein has been attributed to lysophosphatidylcholine (LPC). Albumin avidly binds LPC. Therefore, hypoalbuminemia may directly impair vasodilation and thus contribute to increased risk of atherosclerosis in nephrotic syndrome. The addition of albumin reduces LPC in erythrocytes and endothelial cells. We hypothesized that the addition of albumin will salvage vasodilation in aortic rings previously exposed to LPC. LPC increases superoxide production and disturbs L-arginine availability. Therefore, we also decreased superoxide with a superoxide dismutase mimic, MnCl(2), and supplemented L-arginine in an attempt to restore vasodilation. METHODS: Rat aorta rings, which had been incubated with various concentrations of LPC and human serum albumin (HSA), were mounted in organ chambers. Relaxation was studied with acetylcholine (0.01 to 100 micromol/L) after precontraction with phenylephrine (CON, 0.3 micromol/L; LPC, 0.03 micromol/L). In some studies MnCl(2) or L-arginine was added to the organ chamber. RESULTS: LPC had time- and dose-dependent inhibitory effects on acetylcholine-mediated vasodilation, but no effect on nitroprusside-mediated vasodilation. Preincubation with albumin (50 or 6 g/L) could protect vasodilation against very high levels of LPC. After preincubation with LPC, the addition of albumin to the incubation salvaged vasodilation. Albumin was more effective after short LPC incubation. MnCl(2) had no specific effect on the LPC-mediated disturbance in vasodilation. L-arginine completely salvaged vasodilation at low concentrations of LPC. However, even high concentrations of L-arginine (1 mmol/L) could not improve vasodilation at LPC levels at which vasodilation was restored by albumin. CONCLUSIONS: LPC affects several pathways that inhibit vasodilation, all of which are salvaged by addition of albumin.