Use of real-time PCR to examine the relationship between disease severity in pea and <Emphasis Type="Italic">Aphanomyces euteiches</Emphasis> DNA content in roots

Aphanomyces euteiches causes severe root rot of peas. Resistance is limited in commercial pea cultivars. Real-time fluorescent PCR assay specific for A. euteiches was used to study the relationship between disease severity and pathogen DNA content in infected peas. Five pea genotypes ranging in levels of resistance were inoculated with five isolates of A. euteiches. Plants were visually rated for disease development and the amount of pathogen DNA in roots was determined using the PCR assay. The susceptible genotypes Genie, DSP and Bolero tended to have significantly more disease and more pathogen DNA than the resistant genotypes 90-2079 and PI 180693. PI 180693 consistently had less disease, while 90-2079 had the lowest amount of pathogen DNA. The Spearman correlation between pathogen DNA quantity and disease development was positive and significant (P < 0.05) for three isolates, but was not significant for two other isolates. This suggests that the real-time PCR assay may have limited application as a selection tool for resistance in pea to A. euteiches. Its utility as a selection tool would be dependent on the correlation between disease development and pathogen DNA content for a given pathogen isolate. The accuracy and specificity of the real-time PCR assay suggests considerable application for the assay in the study of mechanisms of disease resistance and the study of microbial population dynamics in plants.     

Variable response of open-pollinated seedling progeny of avocado to Phytophthora root rot

Phytophthora root rot, caused byPhytophthora cinnamomi Rands, is the most important disease of avocado (Persea americana Miller). In an attempt to identify root rot-resistant rootstocks that could ultimately be used under conditions in southern Florida, we screened open-pollinated progeny of avocado from the National Germplasm Repository in Miami. From 1996 to 1998, a total of 2,355 seedlings from 51 accessions were examined in potting mix artificially infested withP. cinnamomi. Most seedlings developed severe root rot, but tolerance was observed in some families (i.e., progeny of certain accessions). Although the most susceptible families developed mean disease ratings of up to 97% root necrosis, mean ratings for the most tolerant families were less than 60%. There was also a strong relationship between the racial background of the female parent and the tolerance of seedlings. Seedlings of the West Indian race and hybrids between it and the Guatemalan race were significantly more tolerant than those from other parents (P< 0.05). Individuals in several families developed < 50% root necrosis, the arbitrary standard of tolerance in this study. Twelve families accounted for 82% (188 of 229) of the tolerant seedlings, and only two of these did not have a West Indian or Guatemalan × West Indian pedigree. Broad-sense heritability for PRR tolerance was 0.45. This is the first report on the inheritance of PRR tolerance in avocado and on the influence of genotype and racial pedigree under controlled conditions.     

Application of non-structural protein ELISA kits in nationwide FMD surveillance in pigs to demonstrate virus circulation in Taiwan

Large scale surveillance of FMD non-structural protein (NSP) antibody in pigs was conducted to monitor for FMD virus circulation in Taiwan using Ceditest and UBI NSP ELISA kits after recurrence of FMD in 2009. A total of 53,759 serum samples were collected from pigs in the auction markets in 2009. There were 43 farms with positive FMD NSP reactors to both NSP ELISA tests in the nationwide surveillance. After tracing back, clinical examination and the NSP ELISA testing using both Ceditest and UBI on 14 follow-up serum samples from all the herds with confirmed NSP reactors in 2009, there were 4 farms classified as positive on follow-up testing criteria. In this surveillance, we have demonstrated that the NSP ELISA tests of outbreak farms followed by clinical and serological investigation could be used to detect FMD circulation in the pig population in Taiwan even while the national compulsory vaccination program is ongoing.     

Pseudomonas fluorescens contamination of a feline packed red blood cell unit and studies of canine units

Background: While screening programs have reduced the risk of infectious disease transmission by donors in human and veterinary blood banking, bacterial contamination of blood products has emerged as a major complication in human medicine. Objectives: To describe a Pseudomonas fluorescens (Pf)‐contaminated feline packed RBC (pRBC) unit and experimentally investigate Pf‐contaminated canine pRBCs. Methods: Canine pRBCs were inoculated with Pf‐rich pRBCs from the sentinel feline unit and stored at 4°C or 20°C for 72 hours. Aliquots from the pRBCs were serially evaluated by microscopy, culture, and a eubacterial 16S rRNA real‐time PCR assay. Results: One Pf‐contaminated feline unit turned black after 22 days of storage and was removed from the blood bank; a source was not found, and no other contaminated units were identified. Canine pRBCs spiked with 5 or 25 μL of the sentinel unit became culture‐ and/or 16S PCR‐positive at ≥8 hours at 20°C and 48 hours at 4°C and developed a color change at ≥24 hours. Sensitivity studies indicated that without incubation, inoculation of ≥100 μL Pf‐rich pRBCs was necessary for a positive 16S PCR test result. Conclusions: P. fluorescens grows in stored pRBCs slowly at 4°C and rapidly at 20°C. Screening of blood products for color change, estimating bacterial concentration with microscopy, and 16S PCR testing are simple and fast ways to detect bacteria in stored blood. Aseptic collection, temperature‐controlled storage, and regular visual monitoring of stored units is recommended. Discolored units should not be transfused, but examined for bacterial contamination or other blood product quality problems.     

Application of unmanned aerial systems for high throughput phenotyping of large wheat breeding nurseries

Background

Low cost unmanned aerial systems (UAS) have great potential for rapid proximal measurements of plants in agriculture. In the context of plant breeding and genetics, current approaches for phenotyping a large number of breeding lines under field conditions require substantial investments in time, cost, and labor. For field-based high-throughput phenotyping (HTP), UAS platforms can provide high-resolution measurements for small plot research, while enabling the rapid assessment of tens-of-thousands of field plots. The objective of this study was to complete a baseline assessment of the utility of UAS in assessment field trials as commonly implemented in wheat breeding programs. We developed a semi-automated image-processing pipeline to extract plot level data from UAS imagery. The image dataset was processed using a photogrammetric pipeline based on image orientation and radiometric calibration to produce orthomosaic images. We also examined the relationships between vegetation indices (VIs) extracted from high spatial resolution multispectral imagery collected with two different UAS systems (eBee Ag carrying MultiSpec 4C camera, and IRIS+ quadcopter carrying modified NIR Canon S100) and ground truth spectral data from hand-held spectroradiometer.

Results

We found good correlation between the VIs obtained from UAS platforms and ground-truth measurements and observed high broad-sense heritability for VIs. We determined radiometric calibration methods developed for satellite imagery significantly improved the precision of VIs from the UAS. We observed VIs extracted from calibrated images of Canon S100 had a significantly higher correlation to the spectroradiometer (r = 0.76) than VIs from the MultiSpec 4C camera (r = 0.64). Their correlation to spectroradiometer readings was as high as or higher than repeated measurements with the spectroradiometer per se.

Conclusion

The approaches described here for UAS imaging and extraction of proximal sensing data enable collection of HTP measurements on the scale and with the precision needed for powerful selection tools in plant breeding. Low-cost UAS platforms have great potential for use as a selection tool in plant breeding programs. In the scope of tools development, the pipeline developed in this study can be effectively employed for other UAS and also other crops planted in breeding nurseries.

     

Estimating “autotrophic” belowground respiration in spruce and beech forests: decreases following girdling

Seasonal fluxes of CO₂ from soil and the contribution of autotrophic (root + mycorrhizal) to total soil respiration (SR) were estimated for a mixed stand of European beech (Fagus sylvatica) and Norway spruce (Picea abies) in Central Europe. Mature trees of each species were girdled in August 2002 to eliminate carbohydrate allocation to roots. SR was measured at distances of 0.5, 1.0, and 1.5/2.0 m from the bole of each tree at 1–2 weeks intervals throughout the fall of 2002 and monthly during the spring and summer of 2003. The contribution of roots and mycorrhizae to total SR was estimated by the decrease in SR compared to ungirdled control trees to account for seasonal patterns evident in controls. SR decreased with soil temperature in the fall 2002 and increased again in 2003 as soil warmed. During most of the study period, SR was strongly related to soil temperature. During the dry summer of 2003, however, SR appeared to be uncoupled from temperature and was strongly related to soil water content (SWC). Mean rates of SR in beech and spruce control plots as well as root densities did not show a clear pattern with distance from the bole. SR decreased to levels below controls in beech within a few days after girdling, whereas spruce did not show a significant decrease until October 2002, 6 weeks after girdling. In both beech and spruce, decreased SR in response to girdling was greatest closest to the bole, possibly reflecting increased mycorrhizal activity close to the bole. Autotrophic respiration was estimated in beech to be as much as 50% of the total SR in the stand. The contribution of autotrophic respiration was less certain for spruce, although close to the bole, the autotrophic fraction may contribute to total SR as much as in beech. The large fraction of autotrophic respiration in total SR requires better understanding of tree level stresses that affect carbon allocation below ground.     

Activating mutations of NOTCH1 in human T cell acute lymphoblastic leukemia

Very rare cases of human T cell acute lymphoblastic leukemia (T-ALL) harbor chromosomal translocations that involve NOTCH1, a gene encoding a transmembrane receptor that regulates normal T cell development. Here, we report that more than 50% of human T-ALLs, including tumors from all major molecular oncogenic subtypes, have activating mutations that involve the extracellular heterodimerization domain and/or the C-terminal PEST domain of NOTCH1. These findings greatly expand the role of activated NOTCH1 in the molecular pathogenesis of human T-ALL and provide a strong rationale for targeted therapies that interfere with NOTCH signaling.     

Environmental persistence of 2,3,7,8-tetrachlorodibenzo-<Emphasis Type="Italic">p</Emphasis>-dioxin in soil around hardstand 7 at eglin air force base,Florida

Background and Goal  A number of global events have generated intense scientific scrutiny and public concern of polychlorinated dibenzo-p-dioxins (dioxin). DIoxins have been associated with a range of adverse health effects. 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) is recognized as among the most dangerous of the dioxin compounds, and was a contaminant found In one of the herbicides used for vegetation control during the Vietnam conflict: Agent Orange. As a result of purging spray systems and leaking drums of Agent Orange concentrate, TCDD contamination occurred in the soils surrounding Hardstand 7 at Eglin Air Force Base, Florida. This research uses a multimedia model to estimate the TCDD concentrations in surface soil around Hardstand 7 for a 70-year time Interval beginning with observed surface soil concentrations 1984. Methods  Hardstand 7 is a nearly 40-m circular concrete and asphalt aircraft parking area. The hardstand was used as a staging area for spraying equipment used to disseminate herbicide In a test area called C-52A within EAFB. Concentrated herbicide was also stored in 208-L barrels for use with the equipment, and later, for disposal. In 1984, a field investigation characterized the extent of TCDD contamination in surface soils around Hardstand 7 using a radial sampling protocol. The 1984 observed concentrations and locations was used in a multimedia model, CalTOX, as an initial source term concentrations and locations to estimate expected concentrations during the subsequent 70-years. Results  The results indicate that more than 94% of the TCDD observed in surface soils 1984 will remain after a 70-year period. Access restrictions and remediation activities at the site eliminate bar verification of the CalTOX estimates. Conclusions. TCDD is highly persistent in the soil medium and natural attenuation may not produce a significant decrease In soil concentrations. Recommendation  Active remediation actions may be required to prevent exposure to TCDD contamination surface soils. Verifying CalTOX concentration estimates an Important step that should be performed, however, the, model provides an easy to use tool to estimate TCDD surface soil contamination at herbicide storage or dispersion staging sites.