First Report of Septoria sp. Infecting Stevia rebaudiana in France and Screening of Stevia rebaudiana Genotypes for Host Resistance.

Stevia rebaudiana, for which cultivation is on the increase worldwide, accumulates acaloric intense sweeteners called steviol glycosides (SGs) in its leaves. Yields can be affected by Septoria leaf spot (SLS) caused by Septoria spp. The objectives of the research were (1) to morphologically and genetically characterize five isolates of Septoria sp. found for the first time from outbreaks of Septoria in stevia fields in Southwestern France and (2) to screen S. rebaudiana germplasm from diverse origins through an automated inoculation method using one of the isolates. Multilocus sequence typing grouped the five isolates obtained from symptomatic plants, closely related to Septoria lycopersici and Septoria apiicola. The response to Septoria sp. of 10 genotypes from different origins was assessed for disease severity (DS), either by visually scoring the symptomatic portion of the whole plants or the portion of symptomatic foliar area (PLSA) determined by image analysis, and the area under the disease progress curve (AUDPC) calculated on the basis of the disease severity rating taken 12, 15, 18, and 21 days after inoculation. No genotypes with complete resistance were identified. Moderately susceptible genotypes "Gawi" and "Esplac1" exhibited only 10 to 15% of symptomatic part on whole plant and the slowest disease development. They could be distinguished from highly susceptible ones "E8", "C", and "E161718" exhibiting up to 40% of symptomatic part on whole plant. The variability of response to Septoria sp. that exists in S. rebaudiana opens up the field of breeding strategies for the development of new cultivars for sustainable and organic S. rebaudiana production.

A retrospective evaluation of supine pelvic radiography image quality using centring points and anatomical axial rotation, including reliability of measurements (ARLEX-P STUDY).

INTRODUCTION: Pelvic radiographs are commonly used for the investigation of a variety of conditions. Comparison between examinations requires a consistent radiographic technique but variations in image quality and radiographic centring points are frequently reported in the literature. The aim of this study was to establish the amount of variation in the radiographic centring point (RCP) and pelvic axial rotation (PAR), with a secondary aim of reporting the reliability of such measures. METHODS: Using a previously acquired imaging archive, 633 adult pelvis/hip radiographs were identified on a Picture Archiving and Communication System (PACS). Radiographs with bilateral prostheses, evidence of acute pelvic trauma, projections acquired on a stretcher/trolley and those demonstrating large discontinuity between the detector and X-ray field centre were excluded. To determine centring point variation (+ values denote superior variations) and axial rotation multiple measurements were obtained from each radiograph. A video was used to train five observers and each of these reviewed ten random cases to determine inter- and intra-rater reliability. One of the five observers then performed the measurements on all remaining radiographs. RESULTS: Following exclusions 380 radiographs were evaluated. The median (IQR) RCP deviation from the inter-acetabular line was +22 (+2 to +43) mm where both iliac crests were present and -29 (-45 to -12) mm where they were not. Eleven (3%) cases demonstrate RCP variation from the midline of greater than 25 mm (no bias towards the left or right side). The median (IQR) PAR was 0.0 (-1.5 to 1.4) degrees with greater variance in PAR for male participants (p = 0.004). Almost 60% of inter-rater ICC measurements were categorised as excellent, good or moderate. CONCLUSION: Variations in RCP and PAR exist when evaluating a sample of routinely acquired pelvis radiographs. Some initial factors, such as sex and sub-examination type (full pelvis [XPEL] or low centred pelvis [XHIPB]) have been identified as having a statistical affect on variability. Further research and methods to standardise radiographic techniques is required and must be multidimensional in nature. IMPLICATIONS FOR PRACTICE: Selection of radiographic technique, including RCP, appears to influence components of the pelvis radiograph. Given the increasing clinical requirements for pelvic radiography further standardisation alongside individual optimisation is warranted.

Coatomer (COPI)-coated vesicles: role in intracellular transport and protein sorting.

Coatomer-coated vesicles have been proposed to play a role in many distinct steps of intracellular transport. Coatomer potentially plays a role in forward transport from the endoplasmic reticulum to the Golgi apparatus and through the Golgi apparatus. It may also function in retrograde transport and in the endocytic pathway. There are limitations to the various approaches used to study the role of coatomer, and looking at these helps us to better define the questions that remain to be answered.

Modifications to mobile chest radiography technique during the COVID-19 pandemic - implications of X-raying through side room windows.

INTRODUCTION: Modifications to common radiographic techniques have resulted from the challenges presented by the COVID-19 pandemic. Reports exist regarding the potential benefits of undertaking mobile radiography through side room windows. The aim of this study was to evaluate the impact on image quality and exposure factors when undertaking such examinations. METHODS: A phantom based study was undertaken using a digital X-ray room. Control acquisitions, using a commercially available image quality test tool, were performed using standard mobile chest radiography acquisition factors. Image quality (physical and visual), incidence surface air kerma (ISAK), Exposure Index (EI) and Deviation Index (DI) were recorded. Image quality and radiation dose were further assessed for two additional (experimental) scenarios, where a side room window was located immediately adjacent to the exit port of the light beam diaphragm. The goal of experimental scenario one was to modify exposure factors to maintain the control ISAK. The goal of experimental scenario two was to modify exposure factors to maintain the control EI and DI. Dose and image quality data were compared between the three scenarios. RESULTS: To maintain the pre-window (control) ISAK (76 µGy), tube output needed a three-fold increase (90 kV/4 mAs versus 90 kV/11.25 mAs). To maintain EI/DI a more modest increase in tube output was required (90 kV/8 mAs/ISAK 54 µGy). Physical and visual assessments of spatial resolution and signal-to-noise ratio were indifferent between the three scenarios. There was a slight statistically significant reduction in contrast-to-noise ratio when imaging through the glass window (2.3 versus 1.4 and 1.2; P = 0.005). CONCLUSION: Undertaking mobile X-ray examinations through side room windows is potentially feasible but does require an increase in tube output and is likely to be limited by minor reductions in image quality. IMPLICATIONS FOR PRACTICE: Mobile examinations performed through side room windows should only be used in limited circumstances and future clinical evaluation of this technique is warranted.

Chest CT in patients with shortness of breath: Comparing high pitch CT and conventional CT on respiratory artefacts and dose.

INTRODUCTION: To investigate chest respiratory artefact reduction using High Pitch Dual Source Computed Tomography (HPCT) compared to conventional CT (CCT) in symptomatic patients with shortness of breath. METHODS: Forty patients were prospectively examined on a second-generation Dual Source scanner. They were randomly divided into two groups: twenty patients underwent an experimental HPCT protocol and twenty control cases CCT protocol. Respiratory artefacts were evaluated using an ordinal score (0, 1 and 2) assigned by two readers with five and thirty years of experience. A qualitative assessment was performed using two categorical groups, group 1 = acceptable and group 2 = unacceptable. Dose Length Product (DLP) was compared. RESULTS: The two groups showed a statistical difference in artefacts reduction (p < 0.0001). HPCT demonstrated no artefacts in 82% of cases, while CCT showed no artefacts in 39% of cases. DLP showed no statistical differences (p = 0.6) with mean = 266.9 for HPCT and mean = 282.65 for CCT. HPCT provides high table speed in the z-direction allowing a high temporal resolution, which reduces respiratory artefacts during free-breathing acquisition. Despite the use of two x-ray tubes, the HPCT did not increase the dose to the patient but provided the highest images quality. CONCLUSIONS: In the emergency setting, HPCTs have been critical for achieving good image quality in uncooperative patients. IMPLICATIONS FOR PRACTICE: Acute respiratory failure is a common emergency department presentation, and the choice of high-speed acquisition CT may increase image quality.

The determinant of potyvirus ability to overcome the RTM resistance of Arabidopsis thaliana maps to the N-terminal region of the coat protein.

In Arabidopsis thaliana Columbia (Col-0) plants, the restriction of Tobacco etch virus (TEV) long-distance movement involves at least three dominant RTM (restricted TEV movement) genes named RTM1, RTM2, and RTM3. Previous work has established that, while the RTM-mediated resistance is also effective against other potyviruses, such as Plum pox virus (PPV) and Lettuce mosaic virus (LMV), some isolates of these viruses are able to overcome the RTM mechanism. In order to identify the viral determinant of this RTM-resistance breaking, the biological properties of recombinants between PPV-R, which systemically infects Col-0, and PPV-PSes, restricted by the RTM resistance, were evaluated. Recombinants that contain the PPV-R coat protein (CP) sequence in an RTM-restricted background are able to systemically infect Col-0. The use of recombinants carrying chimeric CP genes indicated that one or more PPV resistance-breaking determinants map to the 5' half of the CP gene. In the case of LMV, sequencing of independent RTM-breaking variants recovered after serial passages of the LMV AF199 isolate on Col-0 plants revealed, in each case, amino acid changes in the CP N-terminal region, close to the DAG motif. Taken together, these findings demonstrate that the potyvirus CP N-terminal region determines the outcome of the interaction with the RTM-mediated resistance.

Steric masking of a dilysine endoplasmic reticulum retention motif during assembly of the human high affinity receptor for immunoglobulin E.

Signals that can cause retention in the ER have been found in the cytoplasmic domain of individual subunits of multimeric receptors destined to the cell surface. To study how ER retention motifs are masked during assembly of oligomeric receptors, we analyzed the assembly and intracellular transport of the human high-affinity receptor for immunoglobulin E expressed in COS cells. The cytoplasmic domain of the alpha chain contains a dilysine ER retention signal, which becomes nonfunctional after assembly with the gamma chain, allowing transport out of the ER of the fully assembled receptor. Juxtaposition of the cytoplasmic domains of the alpha and gamma subunits during assembly is responsible for this loss of ER retention. Substitution of the gamma chain cytoplasmic domain with cytoplasmic domains of irrelevant proteins resulted in efficient transport out of the ER of the alpha chain, demonstrating that nonspecific steric hindrance by the cytoplasmic domain of the gamma chain accounts for the masking of the ER retention signal present in the cytoplasmic domain of the alpha chain. Such a mechanism allows the ER retention machinery to discriminate between assembled and nonassembled receptors, and thus participates in quality control at the level of the ER.

Low cytoplasmic pH inhibits endocytosis and transport from the trans-Golgi network to the cell surface.

A fibroblast mutant cell line lacking the Na+/H+ antiporter was used to study the influence of low cytoplasmic pH on membrane transport in the endocytic and exocytic pathways. After being loaded with protons, the mutant cells were acidified at pH 6.2 to 6.8 for 20 min while the parent cells regulated their pH within 1 min. Cytoplasmic acidification did not affect the level of intracellular ATP or the number of clathrin-coated pits at the cell surface. However, cytosolic acidification below pH 6.8 blocked the uptake of two fluid phase markers, Lucifer Yellow and horseradish peroxidase, as well as the internalization and the recycling of transferrin. When the cytoplasmic pH was reversed to physiological values, both fluid phase endocytosis and receptor-mediated endocytosis resumed with identical kinetics. Low cytoplasmic pH also inhibited the rate of intracellular transport from the Golgi complex to the plasma membrane. This was shown in cells infected by the temperature-sensitive mutant ts 045 of the vesicular stomatitis virus (VSV) using as a marker of transport the mutated viral membrane glycoprotein (VSV-G protein). The VSV-G protein was accumulated in the trans-Golgi network (TGN) by an incubation at 19.5 degrees C and was transported to the cell surface upon shifting the temperature to 31 degrees C. This transport was arrested in acidified cells maintained at low cytosolic pH and resumed during the recovery phase of the cytosolic pH. Electron microscopy performed on epon and cryo-sections of mutant cells acidified below pH 6.8 showed that the VSV-G protein was present in the TGN. These results indicate that acidification of the cytosol to a pH less than 6.8 inhibits reversibly membrane transport in both endocytic and exocytic pathways. In all likelihood, the clathrin and nonclathrin coated vesicles that are involved in endo- and exocytosis cannot pinch off from the cell surface or from the TGN below this critical value of internal pH.

Role of transmembrane domain interactions in the assembly of class II MHC molecules.

Evidence is presented that suggests a role for transmembrane domain interactions in the assembly of class II major histocompatibility complex (MHC) molecules. Mutations in the transmembrane domains of the class II MHC alpha or beta chains resulted in proteins that did not generate complexes recognized by conformation-dependent antibodies and that were largely retained in the endoplasmic reticulum. Insertion of the alpha and beta transmembrane domains into other proteins allowed the chimeric proteins to assemble, suggesting a direct interaction of the alpha and beta transmembrane domains. The interactions were mediated by a structural motif involving several glycine residues on the same face of a putative alpha helix.

Coatomer interaction with di-lysine endoplasmic reticulum retention motifs.

Although signals for retention in the endoplasmic reticulum (ER) have been identified in the cytoplasmic domain of various ER-resident type I transmembrane proteins, the mechanisms responsible for ER retention are still unknown. Yeast and mammalian ER retention motifs interacted specifically in cell lysates with the coatomer, a polypeptide complex implicated in membrane traffic. Mutations that affect the ER retention capacity of the motifs also abolished binding of the coatomer. These results suggest a role for the coatomer in the retrieval of transmembrane proteins to the ER in both yeast and mammals.

Anatomical and/or pathological predictors for the "incorrect" classification of red dot markers on wrist radiographs taken following trauma.

OBJECTIVE: To establish the prevalence of red dot markers in a sample of wrist radiographs and to identify any anatomical and/or pathological characteristics that predict "incorrect" red dot classification. METHODS: Accident and emergency (A&E) wrist cases from a digital imaging and communications in medicine/digital teaching library were examined for red dot prevalence and for the presence of several anatomical and pathological features. Binary logistic regression analyses were run to establish if any of these features were predictors of incorrect red dot classification. RESULTS: 398 cases were analysed. Red dot was "incorrectly" classified in 8.5% of cases; 6.3% were "false negatives" ("FNs")and 2.3% false positives (FPs) (one decimal place). Old fractures [odds ratio (OR), 5.070 (1.256-20.471)] and reported degenerative change [OR, 9.870 (2.300-42.359)] were found to predict FPs. Frykman V [OR, 9.500 (1.954-46.179)], Frykman VI [OR, 6.333 (1.205-33.283)] and non-Frykman positive abnormalities [OR, 4.597 (1.264-16.711)] predict "FNs". Old fractures and Frykman VI were predictive of error at 90% confidence interval (CI); the rest at 95% CI. CONCLUSION: The five predictors of incorrect red dot classification may inform the image interpretation training of radiographers and other professionals to reduce diagnostic error. Verification with larger samples would reinforce these findings. ADVANCES IN KNOWLEDGE: All healthcare providers strive to eradicate diagnostic error. By examining specific anatomical and pathological predictors on radiographs for such error, as well as extrinsic factors that may affect reporting accuracy, image interpretation training can focus on these "problem" areas and influence which radiographic abnormality detection schemes are appropriate to implement in A&E departments.

The Sec20/Tip20p complex is involved in ER retrieval of dilysine-tagged proteins.

Sec20p and Tip20p were previously identified as two interacting proteins involved in early steps of the secretory pathway in Saccharomyces cerevisiae. Here we describe a novel temperature-sensitive allele of TIP20 and analyze its phenotype. While sec20 and tip20 mutants exhibited a defect in forward ER-to-Golgi transport at the non-permissive temperature, both were also defective for retrieval of various dilysine-tagged proteins from the Golgi back to the endoplasmic reticulum (ER) at lower temperature. Dilysine-dependent Golgi localization of Emp47p was also defective in both mutants. These results suggest a role for the Sec20/Tip20p complex in retrieval of dilysine-tagged proteins back to the ER.

The ADP-ribosylation factor GTPase-activating protein Glo3p is involved in ER retrieval.

Retrograde transport of proteins from the Golgi to the endoplasmic reticulum (ER) has been the subject of some interest in the recent past. Here a new thermosensitive yeast mutant defective in retrieval of dilysine-tagged proteins from the Golgi back to the endoplasmic reticulum was characterized. The ret4-1 mutant also exhibited a selective defect in forward ER-to-Golgi transport of some secreted proteins at the non-permissive temperature. The corresponding RET4 gene was found to encode Glo3p, a GTPase-activating protein (GAP) specific for ADP-ribosylation factor (ARF). In vitro, the Glo3 thermosensitive mutant showed a reduced ARF1-GAP activity. The Glo3 protein belongs to a family of zinc finger proteins that may include additional ARF-GAPs. Gene deletion experiments of other family members showed that only GLO3 deletion resulted in impaired retrieval of dilysine-tagged proteins back to the ER. These results demonstrate that Glo3p is the main ARF-GAP specifically involved in ER retrieval.

Membrane sorting in the endocytic and phagocytic pathway of Dictyostelium discoideum.

To study sorting in the endocytic pathway of a phagocytic and macropinocytic cell, monoclonal antibodies to membrane proteins of Dictyostelium discoideum were generated. Whereas the p25 protein was localized to the cell surface, p80 was mostly present in intracellular endocytic compartments as observed by immunofluorescence as well as immunoelectron microscopy analysis. The p80 gene was identified and encodes a membrane protein presumably involved in copper transport. Expression of chimeric proteins revealed that the cytoplasmic domain of p80 was sufficient to cause constitutive endocytosis and localization of the protein to endocytic compartments. Dileucine- and tyrosine-based endocytic signals described previously in mammalian systems were also capable of targeting chimera to endocytic compartments. In phagocytosing cells no membrane sorting was observed during formation of the phagosome. Both p25 and p80 were incorporated non-selectively in nascent phagosomes, and then retrieved shortly after phagosome closure. Our results emphasize the fact that very active membrane traffic takes place in phagocytic and macropinocytic cells. This is coupled with precise membrane sorting to maintain the specific composition of endocytic compartments.

Development of microsatellite markers in peach [ Prunus persica (L.) Batsch] and their use in genetic diversity analysis in peach and sweet cherry ( Prunus avium L.).

We report the sequence of 41 primer pairs of microsatellites from a CT-enriched genomic library of the peach cultivar 'Merrill O'Henry'. Ten microsatellite-containing clones had sequences similar to plant coding sequences in databases and could be used as markers for known functions. For microsatellites segregating at least in one of the two Prunus F(2) progenies analyzed, it was possible to demonstrate Mendelian inheritance. Microsatellite polymorphism was evaluated in 27 peach and 21 sweet cherry cultivars. All primer pairs gave PCR-amplification products on peach and 33 on cherry (80.5%). Six PCR-amplifications revealed several loci (14.6%) in peach and eight (19.5%) in sweet cherry. Among the 33 single-locus microsatellites amplified in peach and sweet cherry, 13 revealed polymorphism both in peach and cherry, 19 were polymorphic only on peach and one was polymorphic only on cherry. The number of alleles per locus ranged from 1 to 9 for peach and from 1 to 6 on sweet cherry with an average of 4.2 and 2.8 in peach and sweet cherry, respectively. Cross-species amplification was tested within the Prunus species: Prunus avium L. (sweet cherry and mazzard), Prunus cerasus L. (sour cherry), Prunus domestica L. (European plum), Prunus amygdalus Batsch. (almond), Prunus armeniaca L. (apricot), Prunus cerasifera Ehrh. (Myrobalan plum). Plants from other genera of the Rosaceae were also tested: Malus (apple) and Fragaria (strawberry), as well as species not belonging to the Rosaceae: Castanea (chestnut tree), Juglans (walnut tree) and Vitis (grapevine). Six microsatellites gave amplification on all the tested species. Among them, one had an amplified region homologous to sequences encoding a MADS-box protein in Malus x domestica. Twelve microsatellites (29.3%) were amplified in all the Rosaceae species tested and 31 (75.6%) were amplified in all the six Prunus species tested. Thirty three (80.5%), 18 (43.9%) and 13 (31.7%) gave amplification on chestnut tree, grapevine and walnut tree, respectively.

Candidate genes and QTLs for sugar and organic acid content in peach [ Prunus persica (L.) Batsch].

The identification of genes involved in variation of peach fruit quality would assist breeders in creating new cultivars with improved fruit quality. Major genes and quantitative trait loci (QTLs) for physical and chemical components of fruit quality have already been detected, based on the peach [ Prunus persica (L.) Batsch] cv. Ferjalou Jalousia((R)) (low-acid peach) x cv. Fantasia (normally-acid nectarine) F(2) intraspecific cross. Our aim was to associate these QTLs to structural genes using a candidate gene/QTL approach. Eighteen cDNAs encoding key proteins in soluble sugar and organic acid metabolic pathways as well as in cell expansion were isolated from peach fruit. A single-strand conformation polymorphism strategy based on specific cDNA-based primers was used to map the corresponding genes. Since no polymorphism could be detected in the Ferjalou Jalousia((R)) x Fantasia population, gene mapping was performed on the almond [ Prunus amygdalus ( P. dulcis)] cv. Texas x peach cv. Earlygold F(2) interspecific cross from which a saturated map was available. Twelve candidate genes were assigned to four linkage groups of the peach genome. In a second step, the previous QTL detection was enhanced by integrating anchor loci between the Ferjalou Jalousia((R)) x Fantasia and Texas x Earlygold maps and data from a third year of trait assessment on the Ferjalou Jalousia((R)) x Fantasia population. Comparative mapping allowed us to detect a candidate gene/QTL co-location. It involved a cDNA encoding a vacuolar H(+)-pyrophosphatase ( PRUpe;Vp2) that energises solute accumulation, and QTLs for sucrose and soluble solid content. This preliminary result may be the first step in the future development of marker-assisted selection for peach fruit sucrose and soluble solid content.