Evolution in Fast Forward: a Potential Role for Mutators in Accelerating Staphylococcus aureus Pathoadaptation

Pathogen evolution and subsequent phenotypic heterogeneity during chronic infection are proposed to enhance Staphylococcus aureus survival during human infection. We tested this theory by genetically and phenotypically characterizing strains with mutations constructed in the mismatch repair (MMR) and oxidized guanine (GO) system, termed mutators, which exhibit increased spontaneous-mutation frequencies. Analysis of these mutators revealed not only strain-dependent increases in the spontaneous-mutation frequency but also shifts in mutational type and hot spots consistent with loss of GO or MMR functions. Although the GO and MMR systems are relied upon in some bacterial species to prevent reactive oxygen species-induced DNA damage, no deficit in hydrogen peroxide sensitivity was found when either of these DNA repair pathways was lost in S. aureus. To gain insight into the contribution of increased mutation supply to S. aureus pathoadaptation, we measured the rate of α-hemolysin and staphyloxanthin inactivation during serial passage. Detection of increased rates of α-hemolysin and staphyloxanthin inactivation in GO and MMR mutants suggests that these strains are capable of modifying virulence phenotypes implicated in mediating infection. Accelerated derivation of altered virulence phenotypes, combined with the absence of increased ROS sensitivity, highlights the potential of mutators to drive pathoadaptation in the host and serve as catalysts for persistent infections.     

Nitrogenase activity in the rhizosphere of sugar cane and some other tropical grasses

Summary Roots of sugar cane had considerable nitrogenase activity and produced up to 5 n moles ethylene/h/g root by the reduction of acetylene. The rhizosphere soil and soil mid-way between the cane rows also reduced acetylene.Beijerinckia indica was abundant on roots and in the soil. Nitrogenase activity was also associated with roots ofPanicum maximum,Pennisetum purpureum andCymbopogon citratus.     

Production of synthetically created phospholipase A(2) variants with industrial impact

Phospholipases A(2) (PLA(2)) play an important role for the production of lysophospholipids. Presently they are mainly obtained from porcine or bovine pancreas but these mammalian sources are not accepted in several fields of application. To make accessible a non-mammalian PLA(2) to industrial application, synthetic genes encoding PLA(2) from honey bee (Apis mellifera) with modified N-termini were constructed and expressed in Escherichia coli. While expression of the gene with an N-terminal leader sequence to direct the protein into the periplasm failed, four variants with slightly modified N-termini (I1A-PLA(2), I1V-PLA(2), His(6)-tagged PLA(2) and PLA(2) still containing the start methionine) were successfully expressed. In all cases, the PLA(2) variants were produced as inclusion bodies. Their protein content amounted to 26-35% of total cell protein. The optimized renaturation procedure and subsequent purification by cation-exchange chromatography yielded pure active enzymes in yields of 4-11 mg L(-1). The recombinant PLA(2) variants showed activities, far-UV CD and fluorescence spectra similar to the glycosylated PLA(2) isolated from the venom glands of honey bee (bv-PLA(2)). The thermodynamic stabilities of the recombinant enzymes calculated from the transition curves of guanidine hydrochloride induced unfolding were also nearly identical to the stability of bv-PLA(2). For the variant I1A-PLA(2) high-cell density fermentation in 10 L-scale using mineral salt medium was shown to increase the volumetric enzyme yield considerably.     

Novel functions of vimentin in cell adhesion, migration, and signaling

Vimentin is the major intermediate filament (IF) protein of mesenchymal cells. It shows dynamically altered expression patterns during different developmental stages and high sequence homology throughout all vertebrates, suggesting that the protein is physiologically important. Still, until recently, the real tasks of vimentin have been elusive, primarily because the vimentin-deficient mice were originally characterized as having a very mild phenotype. Recent studies have revealed several key functions for vimentin that were not obvious at first sight. Vimentin emerges as an organizer of a number of critical proteins involved in attachment, migration, and cell signaling. The highly dynamic and complex phosphorylation of vimentin seems to be a likely regulator mechanism for these functions. The implicated novel vimentin functions have broad ramifications into many different aspects of cell physiology, cellular interactions, and organ homeostasis.     

Seasonal heterotrophic flagellate and bacterial plankton dynamics in a large oligotrophic lake– Loch Ness, Scotland

1. Bacterial production in the 0–30 m water column of Loch Ness was measured using a dual labelling procedure with [³H] thymidine and [¹⁴C] leucine between May 1993 and June 1994. In most cases the uptake of the two labels did not covary, suggesting unbalanced growth. Rates of bacterial production varied from undetectable to 46.2 μg C l^–1 day^–1. Highest production coincided with the period of highest primary production, but carbon derived from this source was insufficient to meet the bacterial carbon demand, which was met by allochthonous humic inputs to the system.
2. Heterotrophic flagellate (HNAN) grazing rates, measured using fluorescently labelled bacteria, ranged between 10.3 and 24.5 bacteria cell^–1 day^–1 at temperatures between 5 and 15 °C. They removed up to 27% of the bacterial production per day.
3. Heterotrophic flagellate specific growth rates ranged from 0.043 to 0.093 h^–1 between 5 and 15 °C, giving generation times of 7.4–16.1 h.
4. bacterial and HNAN abundances were not coupled, but the highest HNAN grazing impact related to a time of high bacterial productivity.     

Chemicals eluting from disposable plastic syringes and syringe filters alter neurite growth,axogenesis and the microtubule cytoskeleton in cultured hippocampal neurons

Cultures of dissociated hippocampal neurons are often used to study neuronal cell biology. We report that the development of these neurons is strongly affected by chemicals leaching from commonly used disposable medical‐grade syringes and syringe filters. Contamination of culture medium by bioactive substance(s) from syringes and filters occurred with multiple manufacturing lots and filter types under normal use conditions and resulted in changes to neurite growth, axon formation and the neuronal microtubule cytoskeleton. The effects on neuronal morphology were concentration‐dependent and significant effects were detected even after substantial dilution of the contaminated medium. Gas chromatography‐mass spectrometry analyses revealed many chemicals eluting from the syringes and filters. Three of these chemicals (stearic acid, palmitic acid and 1,2‐ethanediol monoacetate) were tested but showed no effects on neurite growth. Similar changes in neuronal morphology were seen with high concentrations of bisphenol A and dibutyl phthalate, two hormonally active plasticisers. Although no such compounds were detected by gas chromatography–mass spectrometry, unknown plasticisers in leachates may affect neurites. This is the first study to show that leachates from laboratory consumables can alter the growth of cultured hippocampal neurons. We highlight important considerations to ensure leachate contamination does not compromise cell biology experiments.

     

Contribution of Chronic Conditions to the Disability Burden across Smoking Categories in Middle-Aged Adults,Belgium

IntroductionSmoking is considered the single most important preventable cause of morbidity and mortality worldwide, contributing to increased incidence and severity of disabling conditions. The aim of this study was to assess the contribution of chronic conditions to the disability burden across smoking categories in middle-aged adults in Belgium.MethodsData from 10,224 individuals aged 40 to 60 years who participated in the 1997, 2001, 2004, or 2008 Health Interview Surveys in Belgium were used. Smoking status was defined as never, former (cessation ≥2 years), former (cessation <2 years), occasional light (<20 cigarettes/day), daily light, and daily heavy (≥20 cigarettes/day). To attribute disability to chronic conditions, binomial additive hazards models were fitted separately for each smoking category adjusted for gender, except for former (cessation <2 years) and occasional light smokers due to the small sample size.ResultsAn increasing trend in the disability prevalence was observed across smoking categories in men (never = 4.8%, former (cessation ≥2 years) = 5.8%, daily light = 7.8%, daily heavy = 10.7%) and women (never = 7.6%, former (cessation ≥2 years) = 8.0%, daily light = 10.2%, daily heavy = 12.0%). Musculoskeletal conditions showed a substantial contribution to the disability burden in men and women across all smoking categories. Other important contributors were depression and cardiovascular diseases in never smokers; depression, chronic respiratory diseases, and diabetes in former smokers (cessation ≥2 years); chronic respiratory diseases, cancer, and cardiovascular diseases in daily light smokers; cardiovascular diseases and chronic respiratory diseases in men and depression and diabetes in women daily heavy smokers.ConclusionsBeyond the well-known effect of smoking on mortality, our findings showed an increasing trend of the disability prevalence and different contributors to the disability burden across smoking categories. This information can be useful from a public health perspective to define strategies to reduce disability in Belgium.     

Warmer winters increase the rhizosphere carbon flow to mycorrhizal fungi more than to other microorganisms in a temperate grassland

A decisive set of steps in the terrestrial carbon (C) cycle is the fixation of atmospheric C by plants and the subsequent C‐transfer to rhizosphere microorganisms. With climate change winters are expected to become milder in temperate ecosystems. Although the rate and pathways of rhizosphere C input to soil could be impacted by milder winters, the responses remain unknown. To address this knowledge‐gap, a winter‐warming experiment was established in a seminatural temperate grassland to follow the C flow from atmosphere, via the plants, to different groups of soil microorganisms. In situ ¹³CO₂ pulse labelling was used to track C into signature fatty acids of microorganisms. The winter warming did not result in any changes in biomass of any of the groups of microorganisms. However, the C flow from plants to arbuscular mycorrhizal (AM) fungi, increased substantially by winter warming. Saprotrophic fungi also received large amounts of plant‐derived C—indicating a higher importance for the turnover of rhizosphere C than biomass estimates would suggest—still, this C flow was unaffected by winter warming. AM fungi was the only microbial group positively affected by winter warming—the group with the closest connection to plants. Winter warming resulted in higher plant productivity earlier in the season, and this aboveground change likely induced plant nutrient limitation in warmed plots, thus stimulating the plant dependence on, and C allocation to, belowground nutrient acquisition. The preferential C allocation to AM fungi was at the expense of C flow to other microbial groups, which were unaffected by warming. Our findings imply that warmer winters may shift rhizosphere C‐fluxes to become more AM fungal‐dominated. Surprisingly, the stimulated rhizosphere C flow was matched by increased microbial turnover, leading to no accumulation of soil microbial biomass.