Modified physiology of burley tobacco plants genetically engineered to express Yb1, a functional EGY enzyme.

MAIN CONCLUSION: Transgenic overexpression of a NtEGY2 gene restores normal green color of burley tobacco plants, but does not increase nitrogen utilization efficiency beyond that exhibited by wild-type individuals. Nitrogen physiology is important in tobacco because of its role in generation of leaf yield and accumulation of nitrogen-containing alkaloids that can react with nitrosating agents in the formation of carcinogenic tobacco-specific nitrosamines. Cultivars of the burley tobacco market class are homozygous for deleterious mutant alleles at the duplicate Yb1 and Yb2 loci which have previously been associated with decreased nitrogen use and utilization efficiency; increased leaf nitrate, total nitrogen, and alkaloid levels; and reduced yields. How mutant alleles at these two loci affect these traits is not well understood. Recent characterization of the Yb1 and Yb2 genes (homologs of Arabidopsis EGY1 gene) enabled overexpression of the wild-type Yb1 allele in yb1yb1yb2yb2 plants to determine if observed unfavorable effects were due to linkage or pleiotropy, and to determine if overexpression could lead to beneficial modifications in any of these traits in transgenic plants relative to naturally-occurring wild-type genotypes. Yb1 overexpression was found to confer an agronomic benefit to yb1yb1yb2yb2 genotypes but no advantage to wild-type genotypes. RNA-Seq was used to carry out a comparative transcriptome analysis of genetically engineered and wild-type nearly isogenic lines (NILs) to gain insight on metabolic pathways affecting carbon and nitrogen metabolism that might be altered as the result of genetic variability at the Yb1 and Yb2 loci. Results indicate that complex changes in the transcriptome of tobacco can be manifested by altered expression of Yb1.

A Flexible Design Platform for Si/SiGe Exchange-Only Qubits with Low Disorder.

Spin-based silicon quantum dots are an attractive qubit technology for quantum information processing with respect to coherence time, control, and engineering. Here we present an exchange-only Si qubit device platform that combines the throughput of CMOS-like wafer processing with the versatility of direct-write lithography. The technology, which we coin "SLEDGE", features dot-shaped gates that are patterned simultaneously on one topographical plane and subsequently connected by vias to interconnect metal lines. The process design enables nontrivial layouts as well as flexibility in gate dimensions, material selection, and additional device features such as for rf qubit control. We show that the SLEDGE process has reduced electrostatic disorder with respect to traditional overlapping gate devices with lift-off metallization, and we present spin coherent exchange oscillations and single qubit blind randomized benchmarking data.

Homoeologous chromosome exchange explains the creation of a QTL affecting soil-borne pathogen resistance in tobacco.

Crop plant partial resistance to plant pathogens controlled by quantitative trait loci (QTL) is desirable in cultivar development programmes because of its increased durability. Mechanisms underlying such resistance are difficult to study. We performed RNA-seq analyses for tobacco (Nicotiana tabacum) nearly isogenic lines (NILs) with and without favourable allele(s) at Phn7.1, a major QTL influencing partial resistance to the soil-borne pathogens Phytophthora nicotianae and Ralstonia solanacearum. Based upon combined analyses of transcriptome-based sequence variation and gene expression profiles, we concluded that allelic variability at the Phn7.1 locus was likely generated from homoeologous exchange, which led to deletion of low-expressing members of the SAR8.2 gene family and duplication of high-expressing SAR8.2 genes from a different subgenome of allotetraploid tobacco. The high expression of endogenous Phn7.1-associated SAR8.2 genes was correlated with observed resistance to P. nicotianae. Our findings suggest a role for genomic rearrangements in the generation of favourable genetic variability affecting resistance to pathogens in plants.

Editing of A622 genes results in ultra-low nicotine whole tobacco plants at the expense of dramatically reduced growth and development.

Due to potential regulations that could affect nicotine levels in some tobacco products, there is interest in using genetic modification to reduce levels of this pyridine alkaloid in tobacco leaves. Enzymes coded by A622 genes have previously been indicated to be involved in one of the latter steps of tobacco alkaloid biosynthesis. Whole tobacco plants with reduced A622 activity have never been evaluated, however. We utilized CRISPR/Cas9-based editing to introduce deleterious mutations into the two A622 genes present in the Nicotiana tabacum genome. Double homozygous A622 mutant genotypes established in four recipient genotypes varying for the presence/absence of mutations in other alkaloid biosynthetic genes exhibited severely reduced nicotine accumulation in field and greenhouse experiments. A622 knockout lines exhibited lower nicotine levels than previously created genotypes with deleterious mutations in BBL genes also associated with one of the latter steps in tobacco alkaloid biosynthesis. Reduced A622 activity resulted in plants with drastically reduced growth and development, however. A622 mutant lines were later flowering and produced green leaf yields that were 60.6% lower, on average, than those for non-A622-mutated control lines. Supplementary Information: The online version contains supplementary material available at 10.1007/s11032-022-01293-w.

Analyses of diverse low alkaloid tobacco germplasm identify naturally occurring nucleotide variability contributing to reduced leaf nicotine accumulation.

Recent suggestions for mandated lowering of nicotine content in cigarettes have prompted tobacco breeders to search for N. tabacum germplasm with allelic variability contributing to low alkaloid accumulation. In this research, we phenotyped a series of 81 selected diverse tobacco introductions (TIs) to identify a sub-group with authentic low alkaloid phenotypes. We also genotyped these materials for sequences associated with the Nic1 and Nic2 loci previously reported to influence tobacco alkaloid biosynthesis. Only five low alkaloid TIs possessed previously described deletions of Ethylene Response Factor (ERF) genes at the Nic2 locus that contribute to lower alkaloid accumulation. Eleven TIs possessed an apparent deletion of ERF199, a gene recently reported to underlie the effect at the Nic1 locus. Quantitative trait locus (QTL) mapping was performed using populations derived from three selected low alkaloid TIs to possibly identify new genomic regions affecting alkaloid accumulation. A major QTL was identified on linkage group 7 in all three populations that aligned with the Nic1 locus. A newly discovered 5 bp deletion in the gene MYC2a on linkage group 5 was found to likely partially underlie the ultra-low alkaloid phenotype of TI 313. This new information is useful for tobacco breeders attempting to assemble novel genetic combinations with the potential for meeting future levels of tolerance for nicotine concentration in cigarette tobacco. Supplementary Information: The online version contains supplementary material available at 10.1007/s11032-021-01274-5.

Combined reduced expression of two gene families lowers nicotine content to ultra-low levels in cultivated tobacco.

KEY MESSAGE: Reduced expression of two gene families results in ultra-low nicotine accumulation in Nicotiana tabacum. The potential for mandated lowering of tobacco cigarette filler nicotine levels to below 0.4 mg g-1 is currently being discussed by regulatory and public health organizations. Commercial tobacco cultivars that would routinely meet this proposed standard do not currently exist. Inactivation or silencing of gene families corresponding to single enzymatic steps in the nicotine biosynthetic pathways have not resulted in tobacco genotypes that would meet this standard under conventional agronomic management. Here, we produced and evaluated under field conditions tobacco genotypes expressing an RNAi construct designed to reduce expression of the Methyl Putrescine Oxidase (MPO) gene family associated with nicotine biosynthesis. In a standard flue-cured genetic background, cured leaf nicotine levels were reduced to only 1.08 to 1.65 mg g-1. When MPO RNAi was combined with reduced Berberine Bridge Like (BBL) activity conferred by induced mutations, genotypes producing cured leaf nicotine levels slightly lower than 0.4 mg g-1 were generated. Past research has suggested that MPO activity may contribute to the biosynthesis of nornicotine in a route that does not involve nicotine. However, nornicotine was not reduced to zero in MPO-silenced plants that were also homozygous for induced mutations in known Nicotine Demethylase genes that are responsible for the vast majority of nornicotine accumulation.

Adaptation of Phytophthora nicotianae to Multiple Sources of Partial Resistance in Tobacco.

Host resistance is an important tool in the management of black shank disease of tobacco. Race development leads to rapid loss of single-gene resistance, but the adaptation by Phytophthora nicotianae to sources of partial resistance from Beinhart 1000, Florida 301, and the Wz gene region introgressed from Nicotiana rustica is poorly characterized. In greenhouse environments, host genotypes with quantitative trait loci (QTLs) conferring resistance from multiple sources were initially inoculated with an aggressive isolate of race 0 or race 1 of P. nicotianae. The most aggressive isolate was selected after each of six host generations to inoculate the next generation of plants. The race 0 isolate demonstrated a continuous gradual increase in disease severity and percentage root rot on all sources of resistance except the genotype K 326 Wz/-, where a large increase in both was observed between generations 2 and 3. Adaptation by the race 0 isolate on Beinhart 1000 represents the first report of adaptation to this genotype by P. nicotianae. The race 1 isolate did not exhibit significant increases in aggressiveness over generations but exhibited a large increase in aggressiveness on K 326 Wz/- between generations 3 and 4. Molecular characterization of isolates recovered during selection was completed via double digest restriction-site associated DNA sequencing, but no polymorphisms were associated with the observed changes in aggressiveness. The rapid adaptation to Wz resistance and the gradual adaptation to other QTLs highlights the need to study the nature of Wz resistance and to conduct field studies on the efficacy of resistance gene rotation for disease management.

Graft choice or drilling technique does not influence outcomes of ACL reconstruction in patients over forty-five.

PURPOSE: To evaluate whether graft-type and tunnel location in ACL reconstruction impact patient-reported outcomes in individuals over the age of 45. METHODS: From 2015 to 2018, patients over 45 years old undergoing primary ACL reconstruction without multi-ligamentous injuries were enrolled in an institutional registry. Baseline International Knee Documentation Committee (IKDC) subjective scores, Knee Injury and Osteoarthritis Outcome Scores (KOOS), Marx Activity Scale, and patient characteristics were collected. Follow-up occurred at a minimum of two years to obtain patient-reported outcomes. RESULTS: Of the 51 patients who qualified for the study, 44 (86.3%) patients were available at a minimum of two years after surgery date (range 24-60 months). Average age at time of surgery of the available patients was 51.6 ± 4.87 (range 45-66). Between femoral tunnel drilling methods, there were no differences in the proportion of patients achieving clinically significant improvement or post-operative outcome scores. While patients who received patellar tendon autografts were more likely to achieve clinically significant improvement in the KOOS sports subscale, there were no other differences in outcomes measures between graft types. Two patients had a retear of their graft, and an additional five patients complained of subjective instability. CONCLUSIONS: In patients over the age of 45, neither the method used to create the femoral tunnel nor the graft type used in ACL reconstruction caused a significant difference in post-operative PROMs with a minimum of two years of follow-up. LEVEL OF EVIDENCE: Therapeutic IV, Case Series.

Early onset of deep sternal wound infection after cardiac surgery is associated with decreased survival: A propensity weighted analysis.

OBJECTIVES: To compare outcomes after the development of early (≤30 days) versus delayed (>30 days) deep sternal wound infection (DSWI) after cardiac surgery. METHODS: Between 2005 and 2016, 64 patients were treated surgically for DSWI following cardiac surgery. Thirty-three developed early DSWI, while 31 developed late DSWI. The mean follow-up was 34.1 ± 32.3 months. RESULTS: Survival for the entire cohort at 1, 3, and 5 years was 93.9%, 85.1%, and 80.8%, respectively. DSWI diagnosed early and attempted medical management was strongly associated with overall mortality (hazard ratio [HR], 25.0 and 9.9; 95% confidence intervals [CIs], 1.18-52.8 and 1.28-76.5; p-value .04 and .04, respectively). Survival was 88.1%, 77.0%, 70.6% and 100%, 94.0% and 94.0% at 1, 3, and 5 years in the early and late DSWI groups, respectively (log-rank = 0.074). Those diagnosed early were more likely to have a positive wound culture (odds ratio [OR], 0.06; 95% CI, 0.01-0.69; p = .024) and diagnosed late were more likely to be female (OR, 8.75; 95% CI, 2.0-38.4; p = .004) and require an urgent DSWI procedure (OR, 9.25; 95% CI, 1.86-45.9; p = .007). Both early diagnosis of DSWI and initial attempted medial management were strongly associated with mortality (HR, 7.48; 95% CI, 1.38-40.4; p = .019 and HR, 7.76; 95% CI, 1.67-35.9; p = .009, respectively). CONCLUSIONS: Early aggressive surgical therapy for DSWI after cardiac surgery results in excellent outcomes. Those diagnosed with DSWI early and who have failed initial medical management have increased mortality.

Equivalent outcomes with minimally invasive and sternotomy mitral valve repair for degenerative mitral valve disease.

BACKGROUND: Debate continues in regard to the optimal surgical approach to the mitral valve for degenerative disease. METHODS: Between February 2004 and July 2015, 363 patients underwent mitral valve repair for degenerative mitral valve disease via either sternotomy (sternotomy, n = 109) or small right anterior thoracotomy (minimally invasive, n = 259). Survival, need for mitral valve reoperation, and progression of mitral regurgitation more than two grades were compared between cohorts using time-based statistical methods and inverse probability weighting. RESULTS: Survival at 1, 5, and 10 years were 99.2, 98.3, and 96.8 for the sternotomy group and 98.1, 94.9, and 94.9 for the minimally invasive group (hazard ratio: 0.39, 95% confidence interval [CI] 0.11-1.30, p = .14). The cumulative incidence of need for mitral valve reoperation with death as a competing outcome at 1, 3, and 5 years were 2.7%, 2.7%, and 2.7% in the sternotomy cohort and 1.5%, 3.3%, and 4.1% for the minimally invasive group (subhazard ratio (SHR) 1.17, 95% CI: 0.33-4.20, p = .81). Cumulative incidence of progression of mitral regurgitation more than two grades with death as a competing outcome at 1, 3, and 5 years were 5.5%, 14.4%, and 44.5% for the sternotomy cohort and 4.2%, 9.7%, and 20.5% for the minimally invasive cohort (SHR: 0.67, 95% CI: 0.28-1.63, p = .38). Inverse probability weighted time-based analyses based on preoperative cohort assignment also demonstrated equivalent outcomes between surgical approaches. CONCLUSIONS: Minimally invasive and sternotomy mitral valve repair in patients with degenerative mitral valve disease is associated with equivalent survival and repair durability.

Does preoperative disease severity influence outcomes in reverse shoulder arthroplasty for cuff tear arthropathy?

BACKGROUND: The degree of symptomatic disease and functional burden has been demonstrated to influence patient results and satisfaction in total hip and knee arthroplasty. Although the relationship between preoperative diagnosis and patient outcomes has been an area of study for reverse total shoulder arthroplasty (RTSA), the influence of the progression of cuff tear arthropathy (CTA) has not yet been examined. The purpose of this study was to evaluate whether preoperative radiographic disease burden and scapular geometry impact patient outcomes and satisfaction in a cohort of patients with CTA treated with RTSA. METHODS: Eighty-six patients were treated for CTA with RTSA performed by the senior author (B.G.) between September 2016 and September 2018 and were enrolled in an institutional registry. At the time of initial evaluation, the baseline American Shoulder and Elbow Surgeons (ASES) score, patient demographic characteristics, history of shoulder surgery, and presence of pseudoparalysis were collected. Radiographs were obtained to evaluate the critical shoulder angle, acromial index, and progression of CTA as assessed by Hamada grading and the Seebauer classification. Patients were contacted to reassess the ASES score and their satisfaction with the improvement in their shoulder function. RESULTS: A total of 79 patients (91.6%) were available for evaluation at a minimum of 24 months of follow-up. Multivariate logistic regression modeling revealed that scapular geometry measurements (critical shoulder angle and acromial index) and the degree of CTA (Seebauer and Hamada classifications) were not associated with worse outcomes as assessed by the ASES score. However, degenerative changes as assessed by the Hamada grade (odds ratio, 0.13 [95% confidence interval, 0.02-0.86]; P = .03) and preoperative ASES score (odds ratio, 1.04 [95% confidence interval, 1.01-1.07]; P = .008) were independently associated with higher satisfaction at 24 months of follow-up. CONCLUSION: The results indicate that patients with greater CTA disease progression did not show differing outcomes after RTSA compared with patients with milder disease. In contrast, both poorer preoperative function and degenerative changes as assessed by the Hamada classification were associated with greater satisfaction after RTSA for CTA. Given the broad spectrum of disease in CTA, there is likely a corresponding range in patient expectations that requires further study to maximize patient satisfaction.

Methicillin-resistant Staphylococcus aureus portends a poor prognosis after endovascular repair of mycotic aortic aneurysms and aortic graft infections.

OBJECTIVE: Mycotic aortic aneurysms and aortic graft infections (aortic infections [AIs]) are rare but highly morbid conditions. Open surgical repair is the "gold standard" treatment, but endovascular repair (EVR) is increasingly being used in the management of AI because of the lower operative morbidity. Multiple organisms are associated with AI, and bacteriology may be an important indication of mortality. We describe the bacteriology and associated outcomes of a group of patients treated with an EVR-first approach for AI. METHODS: All patients who underwent EVR for native aortic or aortic graft infections between 2005 and 2016 were retrospectively reviewed. Primary end points were 30-day mortality and overall mortality. The primary exposure variable was bacteria species. Logistic regression analysis was used to determine association with mortality. Kaplan-Meier survival analysis was used to estimate survival. RESULTS: A total of 2038 EVRs were performed in 1989 unique and consecutive patients. Of those, 27 patients had undergone EVR for AI. Thirteen presented ruptured (48%). Eighteen (67%) were hemodynamically unstable. Ten had a gastrointestinal bleed (37%), whereas others presented with abdominal pain (33%), fever (22%), chest or back pain (18.5%), and hemothorax (3.7%). Twenty patients had a positive blood culture (74%), with the most common organism being methicillin-resistant Staphylococcus aureus (MRSA) isolated in 37% (10). Other organisms were Escherichia coli (3), Staphylococcus epidermidis (2), Streptococcus (2), Enterococcus faecalis (1), vancomycin-resistant Enterococcus (1), and Klebsiella (1). Thirteen patients had 4 to 6 weeks of postoperative antibiotic therapy, six of whom died after therapy. Fourteen were prescribed lifelong therapy; 10 died while receiving antibiotics. On univariate analysis for mortality, smoking history (P = .061) and aerodigestive bleeding on presentation (P = .109) approached significance, whereas MRSA infection (P = .001) was strongly associated with increased mortality. On multivariate analysis, MRSA remained a strong, independent predictor of mortality (adjusted odds ratio, 93.2; 95% confidence interval, 1.9-4643; P = .023). Overall 30-day mortality was 11%, all MRSA positive. At mean follow-up of 17.4 ± 28 months, overall mortality was 59%. Overall survival at 1 year, 3 years, and 5 years was 49%, 31%, and 23%. Kaplan-Meier survival analysis demonstrated that MRSA-positive patients had a significantly lower survival compared with other pathogens (1-year, 20% vs 71%; 5-year, 0% vs 44%; P = .0009). CONCLUSIONS: In our series of AI, the most commonly isolated organism was MRSA. MRSA is highly virulent and is associated with increased mortality compared with all other organisms, regardless of treatment. Given our results, EVR for MRSA-positive AI was not a durable treatment option.

Identification and editing of a hybrid lethality gene expands the range of interspecific hybridization potential in Nicotiana.

KEY MESSAGE: Identification and inactivation of hybrid lethality genes can be used to expand the available gene pool for improvement of a cultivated crop species. Hybrid lethality is one genetic mechanism that contributes to reproductive isolation in plants and serves as a barrier to use of diverse germplasm for improvement of cultivated species. A classic example is the seedling lethality exhibited by progeny from the Nicotiana tabacum × N. africana interspecific cross. In order to increase the body of knowledge on mechanisms of hybrid lethality in plants, and to potentially develop tools to circumvent them, we utilized a transposon tagging strategy to identify a candidate gene involved in the control of this reaction. N. tabacum gene Nt6549g30 was identified to code for a class of coiled-coil nucleotide-binding site-leucine-rich repeat (CC-NBS-LRR) proteins, the largest class of plant defense proteins. Gene editing, along with other experiments, was used to verify that Nt6549g30 is the gene at the N. tabacum Hybrid Lethality 1 (NtHL1) locus controlling the hybrid lethality reaction in crosses with N. africana. Gene editing of Nt6549g30 was also used to reverse interspecific seedling lethality in crosses between N. tabacum and eight of nine additional tested species from section Suaveolentes. Results further implicate the role of disease resistance-like genes in the evolution of plant species and demonstrate the possibility of expanding the gene pool for a crop species through gene editing.

Predictors of Major Adverse Limb Events after Open Forefoot Amputation in Patients with Chronic Limb-Threatening Ischemia.

BACKGROUND: Management of patients with chronic limb-threatening ischemia (CLTI) and extensive foot necrosis presents a challenge for limb salvage. Our study evaluates preoperative risk factors that contributed to durability and efficacy of limb salvage after open transmetatarsal amputation (TMA) in patients with critical limb-threatening ischemia. METHODS: We abstracted data from patients who underwent open TMA at Los Angeles County-University of Southern California Medical Center and Keck Hospital of University of Southern California from 2009 to 2018. Multivariable logistic regression analysis, adjusting for preoperative risk factors, was used to examine predictors of major adverse limb events (MALE). The aim was to evaluate outcomes following open TMA with MALE as the primary outcome. Our hypotheses were that outcomes would be worse for patients with foot infections and renal failure. RESULTS: Forty-three open TMAs were done in 39 patients during the study period. The cohort had a mean age of 63 ± 11.6 years, 89% had a history of diabetes, 95% hypertension (HTN), 54% had end-stage renal disease (ESRD), and 26% were current smokers. MALE occurred in 39% of the cohort. Sex, race, indication, HTN, smoking status, and history of prior ipsilateral revascularization or minor amputations were not associated with MALE (P > 0.05). Multivariate logistic regression found ESRD to be an independent predictor of MALE (odds ratio 7.43, 95% confidence interval 1.12-49.17, P = 0.038) after adjusting for clinically significant covariates. CONCLUSIONS: Open TMA provides acceptable rates of limb salvage for complex patients with CLTI. ESRD is an independent risk factor for MALE following open TMA in these patients. Vigilant follow-up is essential for this morbid patient population given poorer outcomes after forefoot amputation.

Characterization of Phn15.1, a Newly Identified Phytophthora nicotianae Resistance QTL in Nicotiana tabacum.

Phytophthora nicotianae is an oomycete that causes black shank, one of the most economically important diseases affecting tobacco production worldwide. Identification and introgression of novel genetic variability affecting partial genetic resistance to this pathogen is important because of the increased durability of partial resistance over time as compared with genes conferring immunity. A previous mapping study identified a quantitative trait locus (QTL), hereafter designated as Phn15.1, with a major effect on P. nicotianae resistance in tobacco. In this research, we describe significantly improved resistance of nearly isogenic lines (NILs) of flue-cured tobacco carrying the introgressed Phn15.1 region derived from highly resistant cigar tobacco cultivar Beinhart 1000. The Phn15.1 region appeared to act in an additive or partially dominant manner to positively affect resistance. To more finely resolve the position of the gene or genes underlying the Phn15.1 effect, the QTL was mapped with an increased number of molecular markers (single-nucleotide polymorphisms) identified to reside within the region. Development and evaluation of subNILs containing varying amounts of Beinhart 1000-derived Phn15.1-associated genetic material permitted the localization of the QTL to a genetic interval of approximately 2.7 centimorgans. Importantly, we were able to disassociate the Beinhart 1000 Phn15.1 resistance alleles from a functional NtCPS2 allele(s) which contributes to the accumulation of a diterpene leaf surface exudate considered undesirable for flue-cured and burley tobacco. Information from this research should be of value for marker-assisted introgression of Beinhart 1000-derived partial black shank resistance into flue-cured and burley tobacco breeding programs.

Potential Mandated Lowering of Nicotine Levels in Cigarettes: A Plant Perspective.

The Food and Drug Administration has announced the potential for mandated lowering of nicotine levels in combustible cigarettes. The World Health Organization has recommended a lowering of cigarette filler nicotine levels to below 0.4 mg/g. To devise appropriate nicotine control strategies, regulators must consider technical feasibility, timelines for compliance, and potential impediments to implementation. Outlined here is previously unsummarized information on genetic approaches that might be used to reduce nicotine levels in cured tobacco leaves. For the benefit of regulators, altered alkaloid or toxicant profiles that might result by implementation of some of these methodologies are discussed. Also mentioned are potential licensing or regulatory impediments to use of some of the technologies per se. Implications: An understanding of technical feasibility of plant-based nicotine reduction technologies, along with the potential for corresponding alterations in alkaloid or toxicant profiles, is needed by regulators to develop effective nicotine control strategies with minimal impediments or undesirable consequences.

Genetic Control of Facultative Parthenocarpy in Nicotiana tabacum L.

Investigation of parthenocarpy, the production of fruit without fertilization, in multiple plant species could result in development of technologies for conferring seedless fruits and increased stability of fruit formation in economically important plants. We studied parthenocarpy in the model species Nicotiana tabacum L., and observed variability for expression of the trait among diverse genetic materials. Parthenocarpy was found to be partially dominant, and a single major quantitative trait locus on linkage group 22 was found to control the trait in a doubled haploid mapping population derived from a cross between parthenocarpic cigar tobacco cultivar "Beinhart 1000" and nonparthenocarpic flue-cured tobacco cultivar, "Hicks." The same genomic region was found to be involved with control of the trait in the important flue-cured tobacco cultivar, "K326." We also investigated the potential for the production of maternal haploids due to parthenogenesis in parthenocarpic tobacco seed capsules. Maternal haploids were not observed in parthenocarpic capsules, suggesting a requirement of fertilization for maternal haploid production due to parthenogenesis in N. tabacum.

Characterization of Phytophthora nicotianae Resistance Conferred by the Introgressed Nicotiana rustica Region, Wz, in Flue-Cured Tobacco.

Black shank, caused by Phytophthora nicotianae, is one of the most important diseases affecting tobacco worldwide and is primarily managed through use of host resistance. An additional source of resistance to P. nicotianae, designated as Wz, has been introgressed into Nicotiana tabacum from N. rustica. The Wz gene region confers high levels of resistance to all races, but has not been characterized. Our study found Wz-mediated resistance is most highly expressed in the roots, with only a slight reduction in stem-lesion size in Wz genotypes compared with susceptible controls. No substantial relationships were observed between initial inoculum levels and disease development on Wz genotypes, which is generally consistent with qualitative or complete resistance. Isolates of P. nicotianae adapted for five host generations on plants with the Wz gene caused higher disease severity than isolates adapted on Wz plants for only one host generation. Wz-adapted isolates did not exhibit increased aggressiveness on genotypes with other sources of partial resistance, suggesting pathogen adaptation was specific to the Wz gene. To reduce potential for pathogen population shifts with virulence on Wz genotypes, Wz should be combined with other resistance sources and rotation of varying black shank resistance mechanisms is also recommended.

Activated STING in a vascular and pulmonary syndrome.

BACKGROUND: The study of autoinflammatory diseases has uncovered mechanisms underlying cytokine dysregulation and inflammation. METHODS: We analyzed the DNA of an index patient with early-onset systemic inflammation, cutaneous vasculopathy, and pulmonary inflammation. We sequenced a candidate gene, TMEM173, encoding the stimulator of interferon genes (STING), in this patient and in five unrelated children with similar clinical phenotypes. Four children were evaluated clinically and immunologically. With the STING ligand cyclic guanosine monophosphate-adenosine monophosphate (cGAMP), we stimulated peripheral-blood mononuclear cells and fibroblasts from patients and controls, as well as commercially obtained endothelial cells, and then assayed transcription of IFNB1, the gene encoding interferon-ß, in the stimulated cells. We analyzed IFNB1 reporter levels in HEK293T cells cotransfected with mutant or nonmutant STING constructs. Mutant STING leads to increased phosphorylation of signal transducer and activator of transcription 1 (STAT1), so we tested the effect of Janus kinase (JAK) inhibitors on STAT1 phosphorylation in lymphocytes from the affected children and controls. RESULTS: We identified three mutations in exon 5 of TMEM173 in the six patients. Elevated transcription of IFNB1 and other gene targets of STING in peripheral-blood mononuclear cells from the patients indicated constitutive activation of the pathway that cannot be further up-regulated with stimulation. On stimulation with cGAMP, fibroblasts from the patients showed increased transcription of IFNB1 but not of the genes encoding interleukin-1 (IL1), interleukin-6 (IL6), or tumor necrosis factor (TNF). HEK293T cells transfected with mutant constructs show elevated IFNB1 reporter levels. STING is expressed in endothelial cells, and exposure of these cells to cGAMP resulted in endothelial activation and apoptosis. Constitutive up-regulation of phosphorylated STAT1 in patients' lymphocytes was reduced by JAK inhibitors. CONCLUSIONS: STING-associated vasculopathy with onset in infancy (SAVI) is an autoinflammatory disease caused by gain-of-function mutations in TMEM173. (Funded by the Intramural Research Program of the National Institute of Arthritis and Musculoskeletal and Skin Diseases; ClinicalTrials.gov number, NCT00059748.).

Management of patients with acute aortic syndrome through a regional rapid transport system.

OBJECTIVE: The objective of this study was to describe the outcomes of patients with acute aortic syndrome (AAS) during and after transfer to a regional aortic center by a rapid transport system. METHODS: Review of patients with AAS who were transferred by a rapid transport system to a regional aortic center was performed. Data regarding demographics, diagnosis, comorbidities, transportation, and hospital course were acquired. Severity of existing comorbidities was determined by the Society for Vascular Surgery Comorbidity Severity Score (SVSCSS). The Acute Physiology and Chronic Health Evaluation II (APACHE II) score assessed physiologic instability on admission. Risk factors associated with system-related (transfer and hospital) mortality were identified by univariate and multivariate linear regression analysis. RESULTS: During a recent 18-month period (December 2013-July 2015), 183 patients were transferred by a rapid transport system; 148 (81%) patients were transported by ground and 35 (19%) by air. Median distance traveled was 24 miles (range, 3.6-316 miles); median transport time was 42 minutes (range, 10-144 minutes). Two patients died during transport, one with a type A dissection, the other of a ruptured abdominal aortic aneurysm. There were 118 (66%) patients who received operative intervention. Median time to operation was 6 hours. Type B dissections had the longest median time to operation, 45 hours, with system-related mortality of 1.9%; type A dissections had the shortest median time, 3 hours, and a system-related mortality of 16%. Overall, system-related mortality was 15%. On univariate analysis, factors associated with system-related mortality were age ≥65 years (P = .026), coronary artery disease (P = .030), prior myocardial infarction (P = .049), prior coronary revascularization (P = .002), SVSCSS of >8 (P < .001), abdominal pain (P = .002), systolic blood pressure <90 mm Hg at sending hospital (P = .001), diagnosis of aortic aneurysm (P = .013), systolic blood pressure <90 mm Hg in the intensive care unit (P < .001), and APACHE II score >10 (P = .004). Distance traveled and transport mode and duration were not associated with increased risk of system-related mortality. Only SVSCSS of >8 (odds ratio, 7.73; 95% confidence interval, 2.32-25.8; P = .001) was independently associated with an increase in system-related mortality on multivariate analysis. CONCLUSIONS: Implementation of a rapid transport system, regardless of mode or distance, can facilitate effective transfer of patients with AAS to a regional aortic center. An SVSCSS of >8 predicted an increased system-related mortality and may be a useful metric to assess the appropriateness of patient transfer.