How delayed and non-adherent treatment contribute to onward transmission of malaria: a modelling study

Introduction Artemether-lumefantrine (AL) is the most widely-recommended treatment for uncomplicated Plasmodium falciparum malaria. Its efficacy has been extensively assessed in clinical trials. In routine healthcare settings, however, its effectiveness can be diminished by delayed access to treatment and poor adherence. As well as affecting clinical outcomes, these factors can lead to increased transmission, which is the focus of this study. Methods We extend a within-host model of Plasmodium falciparum to include gametocytes, the parasite forms responsible for onward transmission. The model includes a pharmacokineticpharmacodynamic model of AL, calibrated against both immature and mature gametocytes using individual-level patient data, to estimate the impact that delayed access and imperfect adherence to treatment can have on onward transmission of the parasite to mosquitoes. Results Using survey data from 7 African countries to determine the time taken to acquire antimalarials following fever increased our estimates of mean total infectivity of a malaria episode by up to 1.5- fold, compared to patients treated after 24 hours. Realistic adherence behaviour, based on data from a monitored cohort in Tanzania, increased the contribution to transmission by 2.2 to 2.4-fold, compared to a perfectly-adherent cohort. This was driven largely by increased rates of treatment failure leading to chronic infection, rather than prolonged gametocytaemia in patients who have slower, but still successful, clearance of parasites after imperfect adherence to treatment. Our model estimated that the mean infectivity of untreated infections was 29-51 times higher than that of treated infections (assuming perfect drug adherence), underlining the importance of improving treatment coverage. Conclusion Using mathematical modelling, we quantify how delayed treatment and non-adherent treatment can increase transmission compared to prompt effective treatment. We also highlight that transmission from the large proportion of infections which never receive treatment is substantially higher than those treated

CCAAT-enhancer-binding protein-beta expression in vivo is associated with muscle strength

This is a freely-available open access publication. Please cite the published version which is available via the DOI link in this record.Declining muscle strength is a core feature of aging. Several mechanisms have been postulated, including CCAAT/enhancer-binding protein-beta (C/EBP-β)-triggered macrophage-mediated muscle fiber regeneration after micro-injury, evidenced in a mouse model. We aimed to identify in vivo circulating leukocyte gene expression changes associated with muscle strength in the human adult population. We undertook a genome-wide expression microarray screen, using peripheral blood RNA samples from InCHIANTI study participants (aged 30 and 104). Logged expression intensities were regressed with muscle strength using models adjusted for multiple confounders. Key results were validated by real-time PCR. The Short Physical Performance Battery (SPPB) score tested walk speed, chair stand, and balance. CEBPB expression levels were associated with muscle strength (β coefficient = 0.20560, P = 1.03*10(-6), false discovery rate q = 0.014). The estimated handgrip strength in 70-year-old men in the lowest CEBPB expression tertile was 35.2 kg compared with 41.2 kg in the top tertile. CEBPB expression was also associated with hip, knee, ankle, and shoulder strength and the SPPB score (P = 0.018). Near-study-wide associations were also noted for TGF-β3 (P = 3.4*10(-5) , q = 0.12) and CEBPD expression (P = 9.7*10(-5) , q = 0.18) but not for CEBPA expression. We report here a novel finding that raised CEBPB expression in circulating leukocyte-derived RNA samples in vivo is associated with greater muscle strength and better physical performance in humans. This association may be consistent with mouse model evidence of CEBPB-triggered muscle repair: if this mechanism is confirmed, it may provide a target for intervention to protect and enhance aging muscle

The Second Transmembrane Domain of P2X7 Contributes to Dilated Pore Formation

Activation of the purinergic receptor P2X7 leads to the cellular permeability of low molecular weight cations. To determine which domains of P2X7 are necessary for this permeability, we exchanged either the C-terminus or portions of the second transmembrane domain (TM2) with those in P2X1 or P2X4. Replacement of the C-terminus of P2X7 with either P2X1 or P2X4 prevented surface expression of the chimeric receptor. Similarly, chimeric P2X7 containing TM2 from P2X1 or P2X4 had reduced surface expression and no permeability to cationic dyes. Exchanging the N-terminal 10 residues or C-terminal 14 residues of the P2X7 TM2 with the corresponding region of P2X1 TM2 partially restored surface expression and limited pore permeability. To further probe TM2 structure, we replaced single residues in P2X7 TM2 with those in P2X1 or P2X4. We identified multiple substitutions that drastically changed pore permeability without altering surface expression. Three substitutions (Q332P, Y336T, and Y343L) individually reduced pore formation as indicated by decreased dye uptake and also reduced membrane blebbing in response to ATP exposure. Three others substitutions, V335T, S342G, and S342A each enhanced dye uptake, membrane blebbing and cell death. Our results demonstrate a critical role for the TM2 domain of P2X7 in receptor function, and provide a structural basis for differences between purinergic receptors. © 2013 Sun et al

Measuring patient-reported outcomes: moving beyond misplaced common sense to hard science

Interest in the patient's views of his or her illness and treatment has increased dramatically. However, our ability to appropriately measure such issues lags far behind the level of interest and need. Too often such measurement is considered to be a simple and trivial activity that merely requires the application of common sense. However, good quality measurement of patient-reported outcomes is a complex activity requiring considerable expertise and experience. This review considers the most important issues related to such measurement in the context of chronic disease and details how instruments should be developed, validated and adapted for use in additional languages. While there is often consensus on how best to undertake these activities, there is generally little evidence to support such accord. The present article questions these orthodox views and suggests alternative approaches that have been shown to be effective

Macrocyclic colibactin induces DNA double-strand breaks via copper-mediated oxidative cleavage.

Colibactin is an assumed human gut bacterial genotoxin, whose biosynthesis is linked to the clb genomic island that has a widespread distribution in pathogenic and commensal human enterobacteria. Colibactin-producing gut microbes promote colon tumour formation and enhance the progression of colorectal cancer via cellular senescence and death induced by DNA double-strand breaks (DSBs); however, the chemical basis that contributes to the pathogenesis at the molecular level has not been fully characterized. Here, we report the discovery of colibactin-645, a macrocyclic colibactin metabolite that recapitulates the previously assumed genotoxicity and cytotoxicity. Colibactin-645 shows strong DNA DSB activity in vitro and in human cell cultures via a unique copper-mediated oxidative mechanism. We also delineate a complete biosynthetic model for colibactin-645, which highlights a unique fate of the aminomalonate-building monomer in forming the C-terminal 5-hydroxy-4-oxazolecarboxylic acid moiety through the activities of both the polyketide synthase ClbO and the amidase ClbL. This work thus provides a molecular basis for colibactin's DNA DSB activity and facilitates further mechanistic study of colibactin-related colorectal cancer incidence and prevention

MRI measurements of vessel calibre in tumour xenografts: comparison with vascular corrosion casting.

Vessel size index (R(v), μm) has been proposed as a quantitative magnetic resonance imaging (MRI) derived imaging biomarker in oncology, for the non-invasive assessment of tumour blood vessel architecture and vascular targeted therapies. Appropriate pre-clinical evaluation of R(v) in animal tumour models will improve the interpretation and guide the introduction of the biomarker into clinical studies. The objective of this study was to compare R(v) measured in vivo with vessel size measurements from high-resolution X-ray computed tomography (μCT) of vascular corrosion casts measured post mortem from the same tumours, with and without vascular targeted therapy. MRI measurements were first acquired from subcutaneous SW1222 colorectal xenografts in mice following treatment with 0 (n=6), 30 (n=6) or 200 mg/kg (n=3) of the vascular disrupting agent ZD6126. The mice were then immediately infused with a low viscosity resin and, following polymerisation and maceration of surrounding tissues, the resulting tumour vascular casts were dissected and subsequently imaged using an optimised μCT imaging approach. Vessel diameters were not measurable by μCT in the 200 mg/kg group as the high dose of ZD6126 precluded delivery of the resin to the tumour vascular bed. The mean R(v) for the three treatment groups was 24, 23 and 23.5 μm respectively; the corresponding μCT measurements from corrosion casts from the 0 and 30 mg/kg cohorts were 25 and 28 μm. The strong association between the in vivo MRI and post mortem μCT values supports the use of R(v) as an imaging biomarker in clinical trials of investigational vascular targeted therapies

The comparative osmoregulatory ability of two water beetle genera whose species span the fresh-hypersaline gradient in inland waters (Coleoptera: Dytiscidae, Hydrophilidae).

A better knowledge of the physiological basis of salinity tolerance is essential to understanding the ecology and evolutionary history of organisms that have colonized inland saline waters. Coleoptera are amongst the most diverse macroinvertebrates in inland waters, including saline habitats; however, the osmoregulatory strategies they employ to deal with osmotic stress remain unexplored. Survival and haemolymph osmotic concentration at different salinities were examined in adults of eight aquatic beetle species which inhabit different parts of the fresh-hypersaline gradient. Studied species belong to two unrelated genera which have invaded saline waters independently from freshwater ancestors; Nebrioporus (Dytiscidae) and Enochrus (Hydrophilidae). Their osmoregulatory strategy (osmoconformity or osmoregulation) was identified and osmotic capacity (the osmotic gradient between the animal's haemolymph and the external medium) was compared between species pairs co-habiting similar salinities in nature. We show that osmoregulatory capacity, rather than osmoconformity, has evolved independently in these different lineages. All species hyperegulated their haemolymph osmotic concentration in diluted waters; those living in fresh or low-salinity waters were unable to hyporegulate and survive in hyperosmotic media (> 340 mosmol kg(-1)). In contrast, the species which inhabit the hypo-hypersaline habitats were effective hyporegulators, maintaining their haemolymph osmolality within narrow limits (ca. 300 mosmol kg(-1)) across a wide range of external concentrations. The hypersaline species N. ceresyi and E. jesusarribasi tolerated conductivities up to 140 and 180 mS cm(-1), respectively, and maintained osmotic gradients over 3500 mosmol kg(-1), comparable to those of the most effective insect osmoregulators known to date. Syntopic species of both genera showed similar osmotic capacities and in general, osmotic responses correlated well with upper salinity levels occupied by individual species in nature. Therefore, osmoregulatory capacity may mediate habitat segregation amongst congeners across the salinity gradient

Internal genes of a highly pathogenic H5N1 influenza virus determine high viral replication in myeloid cells and severe outcome of infection in mice.

The highly pathogenic avian influenza (HPAI) H5N1 influenza virus has been a public health concern for more than a decade because of its frequent zoonoses and the high case fatality rate associated with human infections. Severe disease following H5N1 influenza infection is often associated with dysregulated host innate immune response also known as cytokine storm but the virological and cellular basis of these responses has not been clearly described. We rescued a series of 6:2 reassortant viruses that combined a PR8 HA/NA pairing with the internal gene segments from human adapted H1N1, H3N2, or avian H5N1 viruses and found that mice infected with the virus with H5N1 internal genes suffered severe weight loss associated with increased lung cytokines but not high viral load. This phenotype did not map to the NS gene segment, and NS1 protein of H5N1 virus functioned as a type I IFN antagonist as efficient as NS1 of H1N1 or H3N2 viruses. Instead we discovered that the internal genes of H5N1 virus supported a much higher level of replication of viral RNAs in myeloid cells in vitro but not in epithelial cells and that this was associated with high induction of type I IFN in myeloid cells. We also found that in vivo during H5N1 recombinant virus infection cells of haematopoetic origin were infected and produced type I IFN and proinflammatory cytokines. Taken together our data infer that human and avian influenza viruses are differently controlled by host factors in alternative cell types; internal gene segments of avian H5N1 virus uniquely drove high viral replication in myeloid cells, which triggered an excessive cytokine production, resulting in severe immunopathology

Comparison of sequencing-based methods to profile DNA methylation and identification of monoallelic epigenetic modifications.

Analysis of DNA methylation patterns relies increasingly on sequencing-based profiling methods. The four most frequently used sequencing-based technologies are the bisulfite-based methods MethylC-seq and reduced representation bisulfite sequencing (RRBS), and the enrichment-based techniques methylated DNA immunoprecipitation sequencing (MeDIP-seq) and methylated DNA binding domain sequencing (MBD-seq). We applied all four methods to biological replicates of human embryonic stem cells to assess their genome-wide CpG coverage, resolution, cost, concordance and the influence of CpG density and genomic context. The methylation levels assessed by the two bisulfite methods were concordant (their difference did not exceed a given threshold) for 82% for CpGs and 99% of the non-CpG cytosines. Using binary methylation calls, the two enrichment methods were 99% concordant and regions assessed by all four methods were 97% concordant. We combined MeDIP-seq with methylation-sensitive restriction enzyme (MRE-seq) sequencing for comprehensive methylome coverage at lower cost. This, along with RNA-seq and ChIP-seq of the ES cells enabled us to detect regions with allele-specific epigenetic states, identifying most known imprinted regions and new loci with monoallelic epigenetic marks and monoallelic expression

Tumor Endothelium Marker-8 Based Decoys Exhibit Superiority over Capillary Morphogenesis Protein-2 Based Decoys as Anthrax Toxin Inhibitors

Anthrax toxin is the major virulence factor produced by Bacillus anthracis. The toxin consists of three protein subunits: protective antigen (PA), lethal factor, and edema factor. Inhibition of PA binding to its receptors, tumor endothelium marker-8 (TEM8) and capillary morphogenesis protein-2 (CMG2) can effectively block anthrax intoxication, which is particularly valuable when the toxin has already been overproduced at the late stage of anthrax infection, thus rendering antibiotics ineffectual. Receptor-like agonists, such as the mammalian cell-expressed von Willebrand factor type A (vWA) domain of CMG2 (sCMG2), have demonstrated potency against the anthrax toxin. However, the soluble vWA domain of TEM8 (sTEM8) was ruled out as an anthrax toxin inhibitor candidate due to its inferior affinity to PA. In the present study, we report that L56A, a PA-binding-affinity-elevated mutant of sTEM8, could inhibit anthrax intoxication as effectively as sCMG2 in Fisher 344 rats. Additionally, pharmacokinetics showed that L56A and sTEM8 exhibit advantages over sCMG2 with better lung-targeting and longer plasma retention time, which may contribute to their enhanced protective ability in vivo. Our results suggest that receptor decoys based on TEM8 are promising anthrax toxin inhibitors and, together with the pharmacokinetic studies in this report, may contribute to the development of novel anthrax drugs