Report From the 39th Annual Meeting of the European Group for Blood and Marrow Transplantation, London, 7–10 April 2013

Antifungal prophylaxis reduces the risks of invasive fungal infection (IFI), all-cause mortality and fungal-related mortality in highly immunosuppressed haemato-oncology patients, but questions remain about the role of therapeutic drug monitoring in this patient population. There is also continuing debate concerning the benefits and risks of empirical versus pre-emptive antifungal therapy when breakthrough IFI occurs during prophylaxis. These topics were discussed during the 39th Annual Meeting of the European Group for Blood and Marrow Transplantation, held in London in April 2013.     

Phytochemical composition of Potentilla anserina L. analyzed by an integrative GC-MS and LC-MS metabolomics platform

Potentilla anserina L. (Rosaceae) is known for its beneficial effects of prevention of pre-menstrual syndrome (PMS). For this reason P. anserina is processed into many food supplements and pharmaceutical preparations. Here we analyzed hydroalcoholic reference extracts and compared them with various extracts of different pharmacies using an integrative metabolomics platform comprising GC-MS and LC-MS analysis and software toolboxes for data alignment (MetMAX Beta 1.0) and multivariate statistical analysis (COVAIN 1.0). Multivariate statistics of the integrated GC-MS and LC-MS data showed strong differences between the different plant extract formulations. Different groups of compounds such as chlorogenic acid, kaempferol 3-O-rutinoside, acacetin 7-O-rutinoside, and genistein were reported for the first time in this species. The typical fragmentation pathway of the isoflavone genistein confirmed the identification of this active compound that was present with different abundances in all the extracts analyzed. As a result we have revealed that different extraction procedures from different vendors produce different chemical compositions, e.g. different genistein concentrations. Consequently, the treatment may have different effects. The integrative metabolomics platform provides the highest resolution of the phytochemical composition and a mean to define subtle differences in plant extract formulations.     

Mechanism of Silver Nanoparticles Action on Insect Pigmentation Reveals Intervention of Copper Homeostasis

Silver nanoparticles (AgNPs), like almost all nanoparticles, are potentially toxic beyond a certain concentration because the survival of the organism is compromised due to scores of pathophysiological abnormalities past that concentration. However, the mechanism of AgNP toxicity remains undetermined. Instead of applying a toxic dose, we attempted to monitor the effects of AgNPs at a nonlethal concentration on wild type Drosophila melanogaster by exposing them throughout their development. All adult flies raised in AgNP doped food showed that up to 50 mg/L concentration AgNP has no negative influence on median survival; however, these flies appeared uniformly lighter in body color due to the loss of melanin pigments in their cuticle. Additionally, fertility and vertical movement ability were compromised due to AgNP feeding. Determination of the amount of free ionic silver (Ag⁺) led us to claim that the observed biological effects have resulted from the AgNPs and not from Ag⁺. Biochemical analysis suggests that the activity of copper dependent enzymes, namely tyrosinase and Cu-Zn superoxide dismutase, are decreased significantly following the consumption of AgNPs, despite the constant level of copper present in the tissue. Consequently, we propose a mechanism whereby consumption of excess AgNPs in association with membrane bound copper transporter proteins cause sequestration of copper, thus creating a condition that resembles copper starvation. This model also explains the cuticular demelanization effect resulting from AgNP since tyrosinase activity is essential for melanin biosynthesis. Finally, we claim that Drosophila, an established genetic model system, can be well utilized for further understanding of the biological effects of nanoparticles.     

Replication and fine mapping of asthma-associated loci in individuals of African ancestry

Asthma originates from genetic and environmental factors with about half the risk of disease attributable to heritable causes. Genome-wide association studies, mostly in populations of European ancestry, have identified numerous asthma-associated single nucleotide polymorphisms (SNPs). Studies in populations with diverse ancestries allow both for identification of robust associations that replicate across ethnic groups and for improved resolution of associated loci due to different patterns of linkage disequilibrium between ethnic groups. Here we report on an analysis of 745 African-American subjects with asthma and 3,238 African-American control subjects from the Candidate Gene Association Resource (CARe) Consortium, including analysis of SNPs imputed using 1,000 Genomes reference panels and adjustment for local ancestry. We show strong evidence that variation near RAD50/IL13, implicated in studies of European ancestry individuals, replicates in individuals largely of African ancestry. Fine mapping in African ancestry populations also refined the variants of interest for this association. We also provide strong or nominal evidence of replication at loci near ORMDL3/GSDMB, IL1RL1/IL18R1, and 10p14, all previously associated with asthma in European or Japanese populations, but not at the PYHIN1 locus previously reported in studies of African-American samples. These results improve the understanding of asthma genetics and further demonstrate the utility of genetic studies in populations other than those of largely European ancestry.     

Serial measurement of the circulating levels of tumour necrosis factor and its soluble receptors 1 and 2 for monitoring leprosy patients during multidrug treatment

Leprosy is an infectious and contagious spectral disease accompanied by a series of immunological events triggered by the host response to the aetiologic agent, Mycobacterium leprae . The induction and maintenance of the immune/inflammatory response in leprosy are linked to multiple cell interactions and soluble factors, primarily through the action of cytokines. The purpose of the present study was to evaluate the serum levels of tumour necrosis factor (TNF)-α and its soluble receptors (sTNF-R1 and sTNF-R2) in leprosy patients at different stages of multidrug treatment (MDT) in comparison with non-infected individuals and to determine their role as putative biomarkers of the severity of leprosy or the treatment response. ELISA was used to measure the levels of these molecules in 30 healthy controls and 37 leprosy patients at the time of diagnosis and during and after MDT. Our results showed increases in the serum levels of TNF-α and sTNF-R2 in infected individuals in comparison with controls. The levels of TNF-α, but not sTNF-R2, decreased with treatment. The current results corroborate previous reports of elevated serum levels of TNF-α in leprosy and suggest a role for sTNF-R2 in the control of this cytokine during MDT.     

Population Structure of Peronospora effusa in the Southwestern United States

Peronospora effusa is an obligate pathogen that causes downy mildew on spinach and is considered the most economically important disease of spinach. The objective of the current research was to assess genetic diversity of known historical races and isolates collected in 2014 from production fields in Yuma, Arizona and Salinas Valley, California. Candidate neutral single nucleotide polymorphisms (SNPs) were identified by comparing sequence data from reference isolates of known races of the pathogen collected in 2009 and 2010. Genotypes were assessed using targeted sequencing on genomic DNA extracted directly from infected plant tissue. Genotyping 26 historical and 167 contemporary samples at 46 SNP loci revealed 82 unique multi-locus genotypes. The unique genotypes clustered into five groups and the majority of isolates collected in 2014 were genetically closely related, regardless of source location. The historical samples, representing several races, showed greater genetic differentiation. Overall, the SNP data indicate much of the genotypic variation found within fields was produced during asexual development, whereas overall genetic diversity may be influenced by sexual recombination on broader geographical and temporal scales.     

Differential responses by two closely related native fishes to restoration actions

Globally, river degradation has decimated freshwater fish populations. To help reverse this trend in a southeastern Australia river, we used multiple restoration actions, including reintroduction of instream woody habitat, riparian revegetation, removal of a weir hindering fish movement, fencing out livestock, and controlling riparian weeds. We monitored the responses of native fish at the segment scale (20 km) and reach scale (0.3 km) over 7 years to assess the effectiveness of the different restoration strategies. Two closely related species, Murray cod Maccullochella peeli and trout cod Maccullochella macquariensis, increased at the restored segment compared with the control segment. However, inherent differences between river segments and low sample size hampered assessment of the mechanisms responsible for segment‐scale changes in fish abundance. In contrast, at the reach scale, only M. peeli abundance significantly increased in reaches supplemented with wood. These differential responses by 2 closely related fish species likely reflect species‐specific responses to increased habitat availability and enhanced longitudinal connectivity when the weir improved passage around a fishway. Changes in M. peeli abundance in segments supplemented with and without wood suggest an increase in carrying capacity and not simply a redistribution of individuals within the segment, facilitated the observed expansion. Our findings confirm the need to consider individual fish species' habitat preferences carefully when designing restoration interventions. Further, species‐specific responses to restoration actions provide waterway managers with precise strategies to target fish species for recovery and the potential to predict fish outcomes based on ecological preferences.     

A cysteine protease gene is expressed early in resistant potato interactions with Phytophthora infestans

A potato cysteine protease (cyp) cDNA expressed at an early stage of an incompatible interaction with Phytophthora infestans was isolated. Both the nucleotide and deduced amino acid sequences are highly homologous to those of a tomato cysteine protease, CYP1. Striking protein similarity to all known cathepsins in animals, particularly cathepsin K, was also observed. However, unlike cathepsins, a granulin binding domain is located near the carboxyl terminus of the putative CYP protein. In animals, granulins bind to receptors in the plasma membrane and signal cell growth and division. A ribonuclease protection assay demonstrated that the cyp gene is tightly regulated and is induced 15 h post inoculation with P. infestans in potato leaves either with high field resistance or in which a resistance (R) gene is activated. We conclude that a common signaling pathway is activated in each form of resistance.     

Structural basis of HP1/PXVXL motif peptide interactions and HP1 localisation to heterochromatin

HP1 family proteins are adaptor molecules, containing two related chromo domains that are required for chromatin packaging and gene silencing. Here we present the structure of the chromo shadow domain from mouse HP1beta bound to a peptide containing a consensus PXVXL motif found in many HP1 binding partners. The shadow domain exhibits a novel mode of peptide recognition, where the peptide binds across the dimer interface, sandwiched in a beta-sheet between strands from each monomer. The structure allows us to predict which other shadow domains bind similar PXVXL motif-containing peptides and provides a framework for predicting the sequence specificity of the others. We show that targeting of HP1beta to heterochromatin requires shadow domain interactions with PXVXL-containing proteins in addition to chromo domain recognition of Lys-9-methylated histone H3. Interestingly, it also appears to require the simultaneous recognition of two Lys-9-methylated histone H3 molecules. This finding implies a further complexity to the histone code for regulation of chromatin structure and suggests how binding of HP1 family proteins may lead to its condensation.