Genomic Analysis of Reduced Susceptibility to Tigecycline in Enterococcus faecium

Tigecycline (TIG) is approved for use for the treatment of complicated intra-abdominal infections, skin and skin structure infections, as well as pneumonia. Acquired resistance or reduced susceptibility to TIG has been observed in Gram-negative rods, has seldom been reported in Gram-positive organisms, and has not yet been reported in Enterococcus faecium. Using the serial passage method, in vitro mutant AusTig and in vitro mutants HMtig1 and HMtig2 with decreased TIG susceptibility (MICs, 0.25 μg/ml) were obtained from strains E. faecium Aus0004 and HM1070 (MICs, 0.03 μg/ml), respectively. In addition, two vancomycin-resistant E. faecium clinical isolates (EF16 and EF22) with reduced susceptibility to TIG (MICs, 0.5 and 0.25 μg/ml, respectively) were studied. Compared to the wild-type strains, the in vitro mutants also showed an increase in the MICs of other tetracyclines. An efflux mechanism did not seem to be involved in the reduced TIG susceptibility, since the presence of efflux pump inhibitors (reserpine or pantoprazole) did not affect the MICs of TIG. Whole-genome sequencing of AusTig was carried out, and genomic comparison with the Aus0004 genome was performed. Four modifications leading to an amino acid substitution were found. These mutations affected the rpsJ gene (efau004_00094, coding for the S10 protein of the 30S ribosomal subunit), efau004_01228 (encoding a cation transporter), efau004_01636 (coding for a hypothetical protein), and efau004_02455 (encoding the l-lactate oxidase). The four other strains exhibiting reduced TIG susceptibility were screened for the candidate mutations. This analysis revealed that three of them showed an amino acid substitution in the same region of the RpsJ protein. In this study, we characterized for the first time genetic determinants linked to reduced TIG susceptibility in enterococci.     

Low income,community poverty and risk of end stage renal disease

Background

The risk of end stage renal disease (ESRD) is increased among individuals with low income and in low income communities. However, few studies have examined the relation of both individual and community socioeconomic status (SES) with incident ESRD.

Methods

Among 23,314 U.S. adults in the population-based Reasons for Geographic and Racial Differences in Stroke study, we assessed participant differences across geospatially-linked categories of county poverty [outlier poverty, extremely high poverty, very high poverty, high poverty, neither (reference), high affluence and outlier affluence]. Multivariable Cox proportional hazards models were used to examine associations of annual household income and geospatially-linked county poverty measures with incident ESRD, while accounting for death as a competing event using the Fine and Gray method.

Results

There were 158 ESRD cases during follow-up. Incident ESRD rates were 178.8 per 100,000 person-years (10⁵ py) in high poverty outlier counties and were 76.3 /10⁵ py in affluent outlier counties, p trend?=?0.06. In unadjusted competing risk models, persons residing in high poverty outlier counties had higher incidence of ESRD (which was not statistically significant) when compared to those persons residing in counties with neither high poverty nor affluence [hazard ratio (HR) 1.54, 95% Confidence Interval (CI) 0.75-3.20]. This association was markedly attenuated following adjustment for socio-demographic factors (age, sex, race, education, and income); HR 0.96, 95% CI 0.46-2.00. However, in the same adjusted model, income was independently associated with risk of ESRD [HR 3.75, 95% CI 1.62-8.64, comparing the?<?$20,000 income group to the?>?$75,000 group]. There were no statistically significant associations of county measures of poverty with incident ESRD, and no evidence of effect modification.

Conclusions

In contrast to annual family income, geospatially-linked measures of county poverty have little relation with risk of ESRD. Efforts to mitigate socioeconomic disparities in kidney disease may be best appropriated at the individual level.

     

Thermotropic liquid crystals from biomacromolecules

Complexation of biomacromolecules (e.g., nucleic acids, proteins, or viruses) with surfactants containing flexible alkyl tails, followed by dehydration, is shown to be a simple generic method for the production of thermotropic liquid crystals. The anhydrous smectic phases that result exhibit biomacromolecular sublayers intercalated between aliphatic hydrocarbon sublayers at or near room temperature. Both this and low transition temperatures to other phases enable the study and application of thermotropic liquid crystal phase behavior without thermal degradation of the biomolecular components.Liquid crystals (LCs) play an important role in biology because their essential characteristic, the combination of order and mobility, is a basic requirement for self-organization and structure formation in living systems (1–3). Thus, it is not surprising that the study of LCs emerged as a scientific discipline in part from biology and from the study of myelin figures, lipids, and cell membranes (4). These and the LC phases formed from many other biomolecules, including nucleic acids (5, 6), proteins (7, 8), and viruses (9, 10), are classified as lyotropic, the general term applied to LC structures formed in water and stabilized by the distinctly biological theme of amphiphilic partitioning of hydrophilic and hydrophobic molecular components into separate domains. However, the principal thrust and achievement of the study of LCs has been in the science and application of thermotropic materials, structures, and phases in which molecules that are only weakly amphiphilic exhibit LC ordering by virtue of their steric molecular shape, flexibility, and/or weak intermolecular interactions [e.g., van der Waals and dipolar forces (11)]. These characteristics enable thermotropic LCs (TLCs) to adopt a wide variety of exotic phases and to exhibit dramatic and useful responses to external forces, including, for example, the electro-optic effects that have led to LC displays and the portable computing revolution. This general distinction between lyotropic LCs and TLCs suggests there may be interesting possibilities in the development of biomolecular or bioinspired LC systems in which the importance of amphiphilicity is reduced and the LC phases obtained are more thermotropic in nature. Such biological TLC materials are very appealing for several reasons. Most biomacromolecules were extensively characterized in aqueous environments, but in TLC phases, their solvent-free properties and functions could be investigated in a state in which no or only traces of water are present. Water exhibits a high dielectric constant and has the ability to form hydrogen bonds, greatly influencing the structure and functions of biomacromolecules or compromising electronic properties such as charge transport (12–15). Indeed, anhydrous TLC systems containing glycolipids (16–19), ferritin (20), and polylysine have been reported (21–23). However, a general approach to fabricating TLCs based on nucleic acids, polypeptides, proteins, and protein assemblies of large molecular weights such as virus particles remains elusive.Here we propose that the combination of biomaterials with suitably chosen surfactants, followed by dehydration, can be effectively applied as a simple generic scheme for producing biomacromolecular-based TLCs. We demonstrate that biological TLCs can be made from a remarkable range of biomolecules and bio-inspired molecules, including nucleic acids, polypeptides, fusion proteins, and viruses. TLC materials typically combine rigid or semirigid anisometric units, which introduce orientational anisotropy, with flexible alkyl chains, which suppress crystallization (24). In the present experiments, negatively charged biomolecules and bio-inspired molecules act as rigid parts, and cationic surfactants make up the flexible units to produce TLC phases with remarkably low LC-isotropic clearing temperatures, which is another TLC signature. Electrostatic interactions couple these rigid and flexible components into hybrid assemblies, which then order into lamellar phases of alternating rigid and flexible layers (Fig. 1) stabilized by the tendency in TLCs for rigid and flexible to spatially segregate (25).Open in a separate windowFig. 1.Proposed structures of TLCs formed by the biological building blocks complexed with surfactants, showing sketches of various lamellar phases and the corresponding phase transition temperatures (°C). The lamellar bilayer structures are made of, alternately, a sublayer of the biomacromolecules and an interdigitated sublayer of the surfactants, where the negatively charged parts of the biomolecules (e.g., phosphate groups of ssDNA and ssRNA, glutamate residues of supercharged ELPs, and N-terminal glutamate and aspartate residues of pVIII protein in phages) electrostatically interact with the cationic head groups of the surfactants. For the ssDNA–DOAB and ssRNA–DOAB smectic TLCs, the oligonucleotides are randomly orientated in the DNA (RNA) sublayers. For the ELP–DDAB complexes, in addition to the bilayer smectic phase, a modulated smectic (Sm_mod) phase is observed at lower temperature. For the phage–DOAB–DDAB lamellar structures, rodlike virus particles are embedded in a sublayer between interdigitated surfactants with additional in-plane orientational order.