Intensive care antibiotic consumption and resistance patterns: a cross-correlation analysis

Background

Over recent decades, a dramatic increase in infections caused by multidrug-resistant pathogens has been observed worldwide. The aim of the present study was to investigate the relationship between local resistance bacterial patterns and antibiotic consumption in an intensive care unit in a Romanian university hospital.

Methods

A prospective study was conducted between 1st January 2012 and 31st December 2013. Data covering the consumption of antibacterial drugs and the incidence density for the main resistance phenotypes was collected on a monthly basis, and this data was aggregated quarterly. The relationship between the antibiotic consumption and resistance was investigated using cross-correlation, and four regression models were constructed, using the SPSS version 20.0 (IBM, Chicago, IL) and the R version 3.2.3 packages.

Results

During the period studied, the incidence of combined-resistant and carbapenem-resistant P. aeruginosa strains increased significantly [(gradient = 0.78, R² = 0.707, p = 0.009) (gradient = 0.74, R² = 0.666, p = 0.013) respectively], mirroring the increase in consumption of β-lactam antibiotics with β-lactamase inhibitors (piperacillin/tazobactam) and carbapenems (meropenem) [(gradient = 10.91, R² = 0.698, p = 0.010) and (gradient = 14.63, R² = 0.753, p = 0.005) respectively]. The highest cross-correlation coefficients for zero time lags were found between combined-resistant vs. penicillins consumption and carbapenem-resistant P. aeruginosa strains vs. carbapenems consumption (0.876 and 0.928, respectively). The best model describing the relation between combined-resistant P. aeruginosa strains and penicillins consumption during a given quarter incorporates both the consumption and the incidence of combined-resistant strains in the hospital department during the previous quarter (multiple R² = 0.953, p = 0.017). The best model for explaining the carbapenem resistance of P. aeruginosa strains based on meropenem consumption during a given quarter proved to be the adjusted model which takes into consideration both previous consumption and incidence density of strains during the previous quarter (Multiple R² = 0.921, p = 0.037).

Conclusions

The cross-correlation coefficients and the fitted regression models provide additional evidence that resistance during the a given quarter depends not only on the consumption of antibacterial chemotherapeutic drugs in both that quarter and the previous one, but also on the incidence of resistant strains circulating during the previous quarter.

     

Why boys will be boys: two pathways of fetal testicular androgen biosynthesis are needed for male sexual differentiation

Human sexual determination is initiated by a cascade of genes that lead to the development of the fetal gonad. Whereas development of the female external genitalia does not require fetal ovarian hormones, male genital development requires the action of testicular testosterone and its more potent derivative dihydrotestosterone (DHT). The "classic" biosynthetic pathway from cholesterol to testosterone in the testis and the subsequent conversion of testosterone to DHT in genital skin is well established. Recently, an alternative pathway leading to DHT has been described in marsupials, but its potential importance to human development is unclear. AKR1C2 is an enzyme that participates in the alternative but not the classic pathway. Using a candidate gene approach, we identified AKR1C2 mutations with sex-limited recessive inheritance in four 46,XY individuals with disordered sexual development (DSD). Analysis of the inheritance of microsatellite markers excluded other candidate loci. Affected individuals had moderate to severe undervirilization at birth; when recreated by site-directed mutagenesis and expressed in bacteria, the mutant AKR1C2 had diminished but not absent catalytic activities. The 46,XY DSD individuals also carry a mutation causing aberrant splicing in AKR1C4, which encodes an enzyme with similar activity. This suggests a mode of inheritance where the severity of the developmental defect depends on the number of mutations in the two genes. An unrelated 46,XY DSD patient carried AKR1C2 mutations on both alleles, confirming the essential role of AKR1C2 and corroborating the hypothesis that both the classic and alternative pathways of testicular androgen biosynthesis are needed for normal human male sexual differentiation.     

Geographical trends in the yolk carotenoid composition of the pied flycatcher (Ficedula hypoleuca)

Carotenoids in the egg yolks of birds are considered to be important antioxidants and immune stimulants during the rapid growth of embryos. Yolk carotenoid composition is strongly affected by the carotenoid composition of the female??s diet at the time of egg formation. Spatial and temporal differences in carotenoid availability may thus be reflected in yolk concentrations. To assess whether yolk carotenoid concentrations or carotenoid profiles show any large-scale geographical trends or differences among habitats, we collected yolk samples from 16 European populations of the pied flycatcher, Ficedula hypoleuca. We found that the concentrations and proportions of lutein and some other xanthophylls in the egg yolks decreased from Central Europe northwards. The most southern population (which is also the one found at the highest altitude) also showed relatively low carotenoid levels. Concentrations of ??-carotene and zeaxanthin did not show any obvious geographical gradients. Egg yolks also contained proportionally more lutein and other xanthophylls in deciduous than in mixed or coniferous habitats. We suggest that latitudinal gradients in lutein and xanthophylls reflect the lower availability of lutein-rich food items in the northern F. hypoleuca populations and in montane southern populations, which start egg-laying earlier relative to tree phenology than the Central European populations. Similarly, among-habitat variation is likely to reflect the better availability of lutein-rich food in deciduous forests. Our study is the first to indicate that the concentration and profile of yolk carotenoids may show large-scale spatial variation among populations in different parts of the species?? geographical range. Further studies are needed to test the fitness effects of this geographical variation.     

Replication fork stalling by bulky DNA damage: localization at active origins and checkpoint modulation

The integrity of the genome is threatened by DNA damage that blocks the progression of replication forks. Little is known about the genomic locations of replication fork stalling, and its determinants and consequences in vivo. Here we show that bulky DNA damaging agents induce localized fork stalling at yeast replication origins, and that localized stalling is dependent on proximal origin activity and is modulated by the intra-S-phase checkpoint. Fork stalling preceded the formation of sister chromatid junctions required for bypassing DNA damage. Despite DNA adduct formation, localized fork stalling was abrogated at an origin inactivated by a point mutation and prominent stalling was not detected at naturally-inactive origins in the replicon. The intra-S-phase checkpoint contributed to the high-level of fork stalling at early origins, while checkpoint inactivation led to initiation, localized stalling and chromatid joining at a late origin. Our results indicate that replication forks initially encountering a bulky DNA adduct exhibit a dual nature of stalling: a checkpoint-independent arrest that triggers sister chromatid junction formation, as well as a checkpoint-enhanced arrest at early origins that accompanies the repression of late origin firing. We propose that the initial checkpoint-enhanced arrest reflects events that facilitate fork resolution at subsequent lesions.     

The CW domain, a new histone recognition module in chromatin proteins

Post-translational modifications of the N-terminal histone tails, including lysine methylation, have key roles in regulation of chromatin and gene expression. A number of protein modules have been identified that recognize differentially modified histone tails and provide their proteins with the capacity to sense such modifications. Here, we identify the CW domain of plant and animal chromatin-related proteins as a novel module that recognizes different methylated states of lysine 4 on histone H3 (H3K4me). The solution structure of the CW domain of the Arabidopsis ASH1 HOMOLOG2 (ASHH2) histone methyltransferase provides insight into how different CW domains can distinguish different methylated histone tails. We provide evidence that ASHH2 is acting on H3K4me-marked genes, allowing for ASHH2-dependent H3K36 tri-methylation, which contributes to sustained expression of tissue-specific and developmentally regulated genes. This suggests that ASHH2 is a combined 'reader' and 'writer' of the histone code. We propose that different CW domains, dependent on their specificity for different H3K4 methylations, are important for epigenetic memory or participate in switching between permissive and repressive chromatin states.     

Membrane topology of NAADP-sensitive two-pore channels and their regulation by N-linked glycosylation

Two-pore channels (TPCs) localize to the endolysosomal system and have recently emerged as targets for the Ca(2+)-mobilizing messenger, nicotinic acid adenine dinucleotide phosphate (NAADP). However, their membrane topology is unknown. Using fluorescence protease protection assays, we show that human TPC1 and TPC2 possess cytosolic N and C termini and therefore an even number of transmembrane regions. Fluorophores placed at position 225 or 347 in TPC1, or 339 in TPC2 were also cytosolic, whereas a fluorophore at position 628 in TPC1 was luminal. These data together with sequence similarity to voltage-gated Ca(2+) and Na(+) channels, and unbiased in silico predictions are consistent with a topology in which two homologous domains are present, each comprising 6 transmembrane regions and a re-entrant pore loop. Immunocytochemical analysis of selectively permeabilized cells using antipeptide antibodies confirmed that the C-terminal tails of recombinant TPCs are cytosolic and that residues 240-254 of TPC2 prior to putative pore 1 are luminal. Both TPC1 and TPC2 are N-glycosylated with residues 599, 611, and 616 contributing to glycosylation of TPC1. This confirms the luminal position of these residues, which immediately precede the putative pore loop of the second domain. Mutation of all three glycosylation sites in TPC1 enhances NAADP-evoked cytosolic Ca(2+) signals. Our data establish essential features of the topology of two-pore channels.     

The styryl dye FM1-43 suppresses odorant responses in a subset of olfactory neurons by blocking cyclic nucleotide-gated (CNG) channels

Many olfactory receptor neurons use a cAMP-dependent transduction mechanism to transduce odorants into depolarizations. This signaling cascade is characterized by a sequence of two currents: a cation current through cyclic nucleotide-gated channels followed by a chloride current through calcium-activated chloride channels. To date, it is not possible to interfere with these generator channels under physiological conditions with potent and specific blockers. In this study we identified the styryl dye FM1-43 as a potent blocker of native olfactory cyclic nucleotide-gated channels. Furthermore, we characterized this substance to stain olfactory receptor neurons that are endowed with cAMP-dependent transduction. This allows optical differentiation and pharmacological interference with olfactory receptor neurons at the level of the signal transduction.     

Molecular basis of actin nucleation factor cooperativity: crystal structure of the Spir-1 kinase non-catalytic C-lobe domain (KIND)•formin-2 formin SPIR interaction motif (FSI) complex

The distinct actin nucleation factors of the Spir and formin subgroup families cooperate in actin nucleation. The Spir/formin cooperativity has been identified to direct two essential steps in mammalian oocyte maturation, the asymmetric spindle positioning and polar body extrusion during meiosis. Understanding the nature and regulation of the Spir/Fmn cooperation is an important requirement to comprehend mammalian reproduction. Recently we dissected the structural elements of the Spir and Fmn family proteins, which physically link the two actin nucleation factors. The trans-regulatory interaction is mediated by the Spir kinase non-catalytic C-lobe domain (KIND) and the C-terminal formin Spir interaction motif (FSI). The interaction inhibits formin nucleation activity and enhances the Spir activity. To get insights into the molecular mechanism of the Spir/Fmn interaction, we determined the crystal structure of the KIND domain alone and in complex with the C-terminal Fmn-2 FSI peptide. Together they confirm the proposed structural homology of the KIND domain to the protein kinase fold and reveal the basis of the Spir/formin interaction. The complex structure showed a large interface with conserved and positively charged residues of the Fmn FSI peptide mediating major contacts to an acidic groove on the surface of KIND. Protein interaction studies verified the electrostatic nature of the interaction. The data presented here provide the molecular basis of the Spir/formin interaction and give a first structural view into the mechanisms of actin nucleation factor cooperativity.     

Does Land-Use Intensification Decrease Plant Phylogenetic Diversity in Local Grasslands?

Phylogenetic diversity (PD) has been successfully used as a complement to classical measures of biological diversity such as species richness or functional diversity. By considering the phylogenetic history of species, PD broadly summarizes the trait space within a community. This covers amongst others complex physiological or biochemical traits that are often not considered in estimates of functional diversity, but may be important for the understanding of community assembly and the relationship between diversity and ecosystem functions. In this study we analyzed the relationship between PD of plant communities and land-use intensification in 150 local grassland plots in three regions in Germany. Specifically we asked whether PD decreases with land-use intensification and if so, whether the relationship is robust across different regions. Overall, we found that species richness decreased along land-use gradients the results however differed for common and rare species assemblages. PD only weakly decreased with increasing land-use intensity. The strength of the relationship thereby varied among regions and PD metrics used. From our results we suggest that there is no general relationship between PD and land-use intensification probably due to lack of phylogenetic conservatism in land-use sensitive traits. Nevertheless, we suggest that depending on specific regional idiosyncrasies the consideration of PD as a complement to other measures of diversity can be useful.