High preharvest donor Foxp3 mRNA level predicts late relapse of acute lymphoblastic leukaemia after haematopoietic stem cell transplantation.

OBJECTIVES: The curative effect of allogeneic haematopoietic stem cell transplantation (HSCT) for acute leukaemia is due in part to the donor T cell-mediated graft-versus-leukaemia immune reaction (GvL). Several studies have suggested that donor CD25+CD4+Foxp3+regulator T cells (Tregs) may decrease graft-versus-host disease (GvHD) without abrogating GVL. This notion may need modification in acute lymphoblastic leukaemia (ALL). METHODS: Foxp3 mRNA level was measured by qPCR in preharvest donor blood CD4+ T cells. The study comprised 45 patients with ALL in 1st or 2nd CR who received myeloablative HSCT using T-replete bone marrow grafts. RESULTS: Relapse occurred in 17 patients median 363 days after HSCT. The relapse risk was estimated by Cox univariate and multivariate proportional hazard regression. The proportionality assumption was met by analysing the preharvest donor Foxp3 mRNA level as a time-dependent covariate. Early relapse was not modified by the Foxp3 mRNA level. However, a higher Foxp3 mRNA level was associated with a significantly increased relapse risk after day 363 after transplantation, compatible with inhibition of GvL. In contrast, a higher preharvest donor CD4+ T-cell concentration was associated with reduced relapse risk. CONCLUSION: A higher preharvest donor Foxp3 mRNA level may be predictive of late ALL relapse after HSCT.

How Does Garlic Mustard Lure and Kill the West Virginia White Butterfly?

As it pertains to insect herbivores, the preference-performance hypothesis posits that females will choose oviposition sites that maximize their offspring's fitness. However, both genetic and environmental cues contribute to oviposition preference, and occasionally "oviposition mistakes" occur, where insects oviposit on hosts unsuitable for larval development. Pieris virginiensis is a pierine butterfly native to North America that regularly oviposits on an invasive plant, Alliaria petiolata, but the caterpillars are unable to survive. Alliaria petiolata has high concentrations of the glucosinolate sinigrin in its tissues, as well as a hydroxynitrile glucoside, alliarinoside. We investigated sinigrin as a possible cause of mistake oviposition, and sinigrin and alliarinoside as possible causes of larval mortality. We found that sinigrin applied to leaves of Cardamine diphylla, a major host of P. virginiensis that does not produce sinigrin, had no effect on oviposition rates. We tested the effect of sinigrin on larval performance using two host plants, one lacking sinigrin (C. diphylla) and one with sinigrin naturally present (Brassica juncea). We found no effect of sinigrin application on survival of caterpillars fed C. diphylla, but sinigrin delayed pupation and decreased pupal weight. On B. juncea, sinigrin decreased survival, consumption, and caterpillar growth. We also tested the response of P. virginiensis caterpillars to alliarinoside, a compound unique to A. petiolata, which was applied to B. oleracea. We found a significant reduction in survival, leaf consumption, and caterpillar size when alliarinoside was consumed. The 'novel weapon' alliarinoside likely is largely responsible for larval failure on the novel host A. petiolata. Sinigrin most likely contributes to the larval mortality observed, however, we did not observe any effect of sinigrin on oviposition by P. virginiensis females. Further research needs to be done on non-glucosinolate contact cues, and volatile signals that may induce P. virginiensis oviposition.

Glucosinolate-related glucosides in Alliaria petiolata: sources of variation in the plant and different metabolism in an adapted specialist herbivore, Pieris rapae.

Specialized metabolites in plants influence their interactions with other species, including herbivorous insects, which may adapt to tolerate defensive phytochemicals. The chemical arsenal of Alliaria petiolata (garlic mustard, Brassicaceae) includes the glucosinolate sinigrin and alliarinoside, a hydroxynitrile glucoside with defensive properties to glucosinolate-adapted specialists. To further our understanding of the chemical ecology of A. petiolata, which is spreading invasively in North America, we investigated the metabolite profile and here report a novel natural product, petiolatamide, which is structurally related to sinigrin. In an extensive study of North American populations of A. petiolata, we demonstrate that genetic population differences as well as developmental regulation contribute to variation in the leaf content of petiolatamide, alliarinoside, sinigrin, and a related glycoside. We furthermore demonstrate widely different metabolic fates of these metabolites after ingestion in the glucosinolate-adapted herbivore Pieris rapae, ranging from simple passage over metabolic conversion to sequestration. The differences in metabolic fate were influenced by plant ß-glucosidases, insect-mediated degradation, and the specificity of the larval gut transport system mediating sequestration.

Higher recipient pre-transplant FOXP3 mRNA expression is associated with acute leukaemia relapse after HSCT.

The effect of higher FOXP3 mRNA expression by recipient pre-transplant CD4+ T cells on leukaemia relapse was analysed in a series of 106 patients who received allogeneic haematopoietic stem cell transplantation after myeloablative conditioning with or without antithymocyte globulin (ATG) due to acute leukaemia in 1st or 2nd complete remission. FOXP3 mRNA was measured by qPCR in purified CD4+ T cells from blood obtained before conditioning. Higher FOXP3 mRNA expression was associated with an increased relapse risk when conditioning included ATG (n = 43, hazard ratio [HR] 11.0 [2.50-48.4], p = 0.00001). No effect was observed in patients not receiving ATG (HR 0.95 [0.53-1.81]).

Factors associated with the appropriate use of ultra-broad spectrum antibiotics, meropenem, for suspected healthcare-associated pneumonia.

ABSTRACT: Pneumonia is a common disease-causing hospitalization. When a healthcare-associated infection is suspected, antibiotics that provide coverage for multi-drug resistant (MDR) or extended-spectrum beta-lactamase (ESBL) bacteria are frequently prescribed. Limited data is available for guidance on using meropenem as a first-line empiric antimicrobial in hospitalized patients with risk factors for MDR/ESBL bacterial infections.This was a single-center, retrospective study designed and conducted to identify factors associated with positive cultures for MDR/ESBL pathogens in hospitalized patients with suspected healthcare-associated pneumonia.Of the 246 patients, 103 patients (41%) received meropenem. Among patients prescribed meropenem, MDR/ESBL pathogens were detected in only 20 patients (13%). Patients admitted from a skilled nursing facility/long-term acute care (SNF/LTAC) or with a history of a positive culture for MDR/ESBL pathogens were significantly associated with positive cultures of MDR/ESBL pathogens during the hospitalization (odds ratio [95% confidence intervals], 31.40 [5.20-189.6] in SNF/LTAC and 18.50 [2.98-115.1] in history of culture-positive MDR/ESBL pathogen). There was no significant difference in mortality between the 3 antibiotic groups.Admission from a SNF/LTAC or having a history of cultures positive for MDR/ESBL pathogens were significantly associated with a positive culture for MDR/ESBL pathogens during the subsequent admission. We did not detect significant association between meropenem use as a first-line drug and morbidity and mortality for patients admitted to the hospital with suspected healthcare-associated pneumonia, and further prospective studies with larger sample size are needed to confirm our findings.

Diversified glucosinolate metabolism: biosynthesis of hydrogen cyanide and of the hydroxynitrile glucoside alliarinoside in relation to sinigrin metabolism in Alliaria petiolata.

Alliaria petiolata (garlic mustard, Brassicaceae) contains the glucosinolate sinigrin as well as alliarinoside, a γ-hydroxynitrile glucoside structurally related to cyanogenic glucosides. Sinigrin may defend this plant against a broad range of enemies, while alliarinoside confers resistance to specialized (glucosinolate-adapted) herbivores. Hydroxynitrile glucosides and glucosinolates are two classes of specialized metabolites, which generally do not occur in the same plant species. Administration of [UL-(14)C]-methionine to excised leaves of A. petiolata showed that both alliarinoside and sinigrin were biosynthesized from methionine. The biosynthesis of alliarinoside was shown not to bifurcate from sinigrin biosynthesis at the oxime level in contrast to the general scheme for hydroxynitrile glucoside biosynthesis. Instead, the aglucon of alliarinoside was formed from metabolism of sinigrin in experiments with crude extracts, suggesting a possible biosynthetic pathway in intact cells. Hence, the alliarinoside pathway may represent a route to hydroxynitrile glucoside biosynthesis resulting from convergent evolution. Metabolite profiling by LC-MS showed no evidence of the presence of cyanogenic glucosides in A. petiolata. However, we detected hydrogen cyanide (HCN) release from sinigrin and added thiocyanate ion and benzyl thiocyanate in A. petiolata indicating an enzymatic pathway from glucosinolates via allyl thiocyanate and indole glucosinolate derived thiocyanate ion to HCN. Alliarinoside biosynthesis and HCN release from glucosinolate-derived metabolites expand the range of glucosinolate-related defenses and can be viewed as a third line of defense, with glucosinolates and thiocyanate forming protein being the first and second lines, respectively.

Possible evolution of alliarinoside biosynthesis from the glucosinolate pathway in Alliaria petiolata.

Nitrile formation in plants involves the activity of cytochrome P450s. Hydroxynitrile glucosides are widespread among plants but generally do not occur in glucosinolate producing species. Alliaria petiolata (garlic mustard, Brassicaceae) is the only species known to produce glucosinolates as well as a γ-hydroxynitrile glucoside. Furthermore, A. petiolata has been described to release diffusible cyanide, which indicates the presence of unidentified cyanogenic glucoside(s). Our research on A. petiolata addresses the molecular evolution of P450s. By integrating current knowledge about glucosinolate and hydroxynitrile glucoside biosynthesis in other species and new visions on recurrent evolution of hydroxynitrile glucoside biosynthesis, we propose a pathway for biosynthesis of the γ-hydroxynitrile glucoside, alliarinoside. Homomethionine and the corresponding oxime are suggested as shared intermediates in the biosynthesis of alliarinoside and 2-propenyl glucosinolate. The first committed step in the alliarinoside pathway is envisioned to be catalysed by a P450, which has been recruited to metabolize the oxime. Furthermore, alliarinoside biosynthesis is suggested to involve enzyme activities common to secondary modification of glucosinolates. Thus, we argue that biosynthesis of alliarinoside may be the first known case of a hydroxynitrile glucoside pathway having evolved from the glucosinolate pathway. An intriguing question is whether the proposed hydroxynitrile intermediate may also be converted to novel homomethionine-derived cyanogenic glucoside(s), which could release cyanide. Elucidation of the pathway for biosynthesis of alliarinoside and other putative hydroxynitrile glucosides in A. petiolata is envisioned to offer significant new knowledge on the emerging picture of P450 functional dynamics as a basis for recurrent evolution of pathways for bioactive natural product biosynthesis.

Phenylalanine derived cyanogenic diglucosides from Eucalyptus camphora and their abundances in relation to ontogeny and tissue type.

The cyanogenic glucoside profile of Eucalyptus camphora was investigated in the course of plant ontogeny. In addition to amygdalin, three phenylalanine-derived cyanogenic diglucosides characterized by unique linkage positions between the two glucose moieties were identified in E. camphora tissues. This is the first time that multiple cyanogenic diglucosides have been shown to co-occur in any plant species. Two of these cyanogenic glucosides have not previously been reported and are named eucalyptosin B and eucalyptosin C. Quantitative and qualitative differences in total cyanogenic glucoside content were observed across different stages of whole plant and tissue ontogeny, as well as within different tissue types. Seedlings of E. camphora produce only the cyanogenic monoglucoside prunasin, and genetically based variation was observed in the age at which seedlings initiate prunasin biosynthesis. Once initiated, total cyanogenic glucoside concentration increased throughout plant ontogeny with cyanogenic diglucoside production initiated in saplings and reaching a maximum in flower buds of adult trees. The role of multiple cyanogenic glucosides in E. camphora is unknown, but may include enhanced plant defense and/or a primary role in nitrogen storage and transport.