Selection and analysis of differentially expressed genes in the interaction between Fusarium and Verticillium wilt pathogens and eggplant carrying the Rfo-sa1 resistance locus introgressed from S. aethiopicum

Fusarium oxysporum f. sp. melongenae (Fom) e Verticillium dahliae (Vd) sono funghi vascolari che provocano ingenti danni alle coltivazioni di melanzana sia in serra che in pieno campo. Il locus di resistenza (Rfo-sa1) a Fusarium è stato introgresso da S. aethiopicum in melanzana coltivata (Solanum melongena). Lo studio degli effetti dell’espressione della resistenza a Fusarium oxysporum sulla risposta anche agli attacchi di Verticillium dahliae è stato condotto prendendo in esame sia l’aspetto fenotipico che l’aspetto molecolare. Sono state allestite diverse prove di inoculazione fungina con cinque genotipi di melanzana, tre dei quali resistenti a Fom (All96/6x1F59, All96/6 e 305-E40) e due sensibili a Fom (1F59 e Tal1/1). I diversi genotipi sono stati inoculati con sospensione conidica di Fom, Vd e entrambi i funghi insieme con 5 differenti modalità. L’andamento della malattia è stato valutato rilevando, dopo 15, 22 e 30 giorni dall’inoculazione, l’indice di progressione della malattia lungo il fusto (IESF) e l’indice di alterazione fogliare (IAF). I risultati hanno confermato nei tre genotipi possedenti il locus Rfosa1 di resistenza a Fusarium, c’è totale assenza di sintomi in seguito ad inoculazione con Fom, e , fatto molto più importante, che sintomi nelle inoculazioni miste (Fom e Vd) sono risultati significativamente di minore intensità per IESF e IAF rispetto a quella con solo Verticillium, come osservato in lavori preliminari. Per l’individuazione e lo studio dei geni coinvolti nei meccanismi di interazione funghimelanzana è stata utilizzata la linea All96/6x1F59. Sono state applicate diverse tecniche molecolari: PCR select, qRT-PCR e microarray. E’ stato estratto l’mRNA da campioni di radice di melanzana sottoposti a inoculazione con Fom, Vd e entrambi i funghi contemporaneamente, più un’inoculazione con acqua come controllo. sono state create tre librerie sottrattive (una per ogni inoculazione considerata) a cDNA utilizzando la tecnica PCR select, e il confronto degli spot sui filtri Dot-blot ha permesso di selezionare un totale di circa 1000 cloni come differenzialmente espressi in seguito ad inoculazione. Tutti questi cloni sono stati sequenziati e classificati a livello funzionale. La selezione del gene GAPDH (gliceraldeide fosfato deidrogenasi) come gene housekeeping è stata piuttosto laboriosa. E’ stata quindi svolta la valutazione dell’espressione, mediante qRT-PCR, di cinquanta geni selezionati. Di particolare interesse sono risultati alcuni geni appartenenti alle categorie di meccanismi di difesa, trasporto e metabolismo secondario. E’ stato poi preparato e ibridato un chip microarray Combimatrix, al fine di ottenere una panoramica più ampia sui geni differenzialmente espressi in seguito a inoculazione con i due funghi. I risultati dell’array sono stati validati con la tecnica qRT-PCR. L’analisi comparativa dei dati ottenuti dalle tre tecniche utilizzate (SSH, array e qRT-PCR) ha permesso di selezionare alcuni geni differenzialmente espressi in seguito alle diverse inoculazioni. Questi geni, tutti implicati ai meccanismi di risposta ai patogeni, rappresentano il punto di partenza per l’impostazione di studi di genomica funzionale

Kaposi's sarcoma-associated herpesvirus/human herpesvirus 8-associated polyclonal body cavity effusions that mimic primary effusion lymphomas

Kaposi's sarcoma-associated herpesvirus/human herpesvirus 8 (KSHV/HHV-8) is involved in the pathogenesis of Kaposi's sarcoma (KS) and 2 lymphoproliferative diseases, primary effusion lymphoma (PEL) and the plasmablastic variant of multicentric Castleman's disease (MCD). PEL is defined as a liquid-phase lymphoma harboring monoclonal rearrangements of immunoglobulin (Ig) variable genes or, more rarely, of T-cell receptor (TCR) genes. As a rule, these elements are used for PEL diagnosis together with the demonstration of HHV-8 infection of the tumor clone. We report here a group of five patients with MCD and KS (n = 2) or MCD only (n = 2) or KS only (n = 1); subjects with KS showed cutaneous and visceral involvement. All 5 patients developed recurrent and massive HHV-8-positive body cavity effusions, often in multiple sites, and died shortly after diagnosis. Cytological examination of effusion specimens disclosed the absence of lymphoma cells in all cases. In the context of MCD or MCD and KS, the effusion cell population was found to include 5–30% of atypical lymphoid cells with plasmacytic features and reminiscent of PEL. Our observations suggest that patients with MCD and/or KS (not exclusively in association with HIV infection) may develop recurrent HHV-8-positive body cavity effusions other than PEL, although with a similar poor outcome. Their common morphologic and molecular features appear to identify a nonlymphomatous entity. The identification of HHV-8 in polyclonal effusions that mimic PEL is notable in light of the association between HHV-8 and PEL, and suggests a putative model of PEL lymphomagenesis

Coronary physiology in patients with severe aortic stenosis: Comparison between fractional flow reserve and instantaneous wave-free ratio

Background: The functional assessment of coronary artery disease (CAD) in patientswith aortic stenosis (AS) has not been validated so far, and the best strategy to physiologically investigate the relevance of coronary stenosis in this specific setting of patients remains undetermined. The aim of the study is to compare the diagnostic performance of instantaneous wave-free ratio (iFR) and fractional flow reserve (FFR) in patients with severe AS. y Methods: The functional significance of 179 coronary lesionswas investigatedwith on-line iFR and FFR measurements in 85 AS patients and comparedwith a control group formed by 167 patients (290 lesions) with stable CAD and without AS. The iFR-FFR diagnostic agreement has been tested using the conventional 0.80 FFR cut-off.Results: The correlation between iFR and FFR was similar between AS and CAD patients, aswell as the area under the curve at ROC curve analysis (0.97 vs 0.96, p= 0.88). However, using the standard iFR 0.89 threshold, the diagnostic accuracy of iFRwas significantly lower in AS comparedwith CAD (76.3% vs 86.1%, p= 0.009). According to ROC analysis, the best iFR cut-off in predicting FFR <= 0.8 was lower in AS (0.83, J= 0.82) compared with CAD (0.89, J= 0.81). Using the ROC derived cut-off of 0.83, the iFR accuracy increased significantly (91.3%, p= 0.003) while maintaining an elevated negative predictive value (95.5%).Conclusions: In the presence of severe AS, conventional iFR cut-off had lower diagnostic agreementwith FFR classification of coronary lesions compared to stable CAD patients. AS seems to influence iFR cut-off ischemic thresholds and deserves further comparative studies. (C) 2017 Elsevier B. V. All rights reserved

Physiological Versus Angiographic Guidance for Myocardial Revascularization in Patients Undergoing Transcatheter Aortic Valve Implantation

Background-Management of coronary artery disease in patients undergoing transcatheter aortic valve implantation is uncertain. Fractional flow reserve (FFR) has never been clinically validated in aortic stenosis. The study aim was to analyze the clinical outcome of FFR-guided revascularization in patients undergoing transcatheter aortic valve implantation.Methods and Results-Patients with severe aortic stenosis and coronary artery disease at coronary angiography were included in this retrospective analysis and divided in 2 groups: angiography guided (122/216; 56.5%) versus FFR-guided revascularization (94/216; 43.5%). Patients were clinically followed up and evaluated for the occurrence of major adverse cardiac and cerebrovascular events at 2-year follow-up. Most lesions in the FFR group resulted negative according to the conventional 0.80 cutoff value (111/142; 78.2%) and were deferred. The FFR-guided group showed a better major adverse cardiac and cerebrovascular event-free survival compared with the angio-guided group (92.6% versus 82.0%; hazard ratio, 0.4; 95% CI, 0.2-1.0; P=0.035). Patients with deferred lesions based on FFR presented better outcome compared with patients who underwent angio-guided percutaneous coronary intervention (91.4% versus 68.1%; hazard ratio, 0.3; 95% CI, 0.1-0.6; P=0.001).Conclusions-FFR guidance was associated with favorable outcome in this observational study in patients undergoing transcatheter aortic valve implantation. Randomized trials are needed to investigate the long-term effects of FFR-guided revascularization against angiographic guidance alone in patients with aortic stenosis

QTL Mapping in Eggplant Reveals Clusters of Yield-Related Loci and Orthology with the Tomato Genome

<div><p>In spite of its widespread cultivation and nutritional and economic importance, the eggplant (<i>Solanum melongena</i> L.) genome has not been extensively explored. A lack of knowledge of the patterns of inheritance of key agronomic traits has hindered the exploitation of marker technologies to accelerate its genetic improvement. An already established F₂ intraspecific population of eggplant bred from the cross ‘305E40’ x ‘67/3’ was phenotyped for 20 agronomically relevant traits at two sites. Up to seven quantitative trait loci (QTL) per trait were identified and the percentage of the phenotypic variance (PV) explained per QTL ranged from 4 to 93%. Not all the QTL were detectable at both sites, but for each trait at least one major QTL (PV explained ≥10%) was identified. Although no detectable QTL x environment interaction was found, some QTL identified were location-specific. Many of the fruit-related QTL clustered within specific chromosomal regions, reflecting either linkage and/or pleiotropy. Evidence for putative tomato orthologous QTL/genes was obtained for several of the eggplant QTL. Information regarding the inheritance of key agronomic traits was obtained. Some of the QTL, along with their respective linked markers, may be useful in the context of marker-assisted breeding.</p></div

QTL location (only chromosomes harboring QTL are shown).

<p>The scale shown on the <i>left</i> indicates the chromosome length in cM. Marker names are shown to the <i>right</i>; the inclusion of a superscript near a marker name indicates the presence and number of additional co-localizing markers on the Barchi et al. (2012) map. Map positions of the QTL are given on the <i>left</i> of each chromosome. The length of the vertical bars represents the QTL confidence interval. QTL shown in <i>blue</i> were detected at ML, and those in <i>red</i> at MT. Epistatic QTL are prefixed by “*” where the QTL had already been detected by MapQTL software, and by “ep” where the QTL was newly detected.</p

Inter-trait Spearman correlations assessed in the mapping population.

<p>The values in diagonal represent correlations of the same trait between the two environments. All correlations were significant (p<0.05).</p

Linkage map of eggplant and graphical representation of the QTLs detected.

<p>A) chromosomes E01 to E06; B) chromosomes E07 to E12. Marker names are shown to the <i>right</i> of each chromosome, with map distances (in cM) shown on the <i>left</i>. Map positions of the QTL identified on each chromosome (or LG) are also given on the <i>right</i>. The length of the vertical bars represents the confidence interval of the QTL (LODmax-1 interval). QTL shown in <i>blue</i> were detected only at ML, and those in <i>red</i> only at MT.</p