Dissimilatory Fe(III) reduction controls on arsenic mobilisation: a combined biogeochemical and NanoSIMS imaging approach

This is the final version. Available on open access from Frontiers Media via the DOI in this recordMicrobial metabolism plays a key role in controlling the fate of toxic groundwater contaminants such as arsenic. Dissimilatory metal reduction catalysed by subsurface bacteria can facilitate the mobilisation of arsenic via the reductive dissolution of As(V)-bearing Fe(III) mineral assemblages. The mobility of liberated As(V) can then be amplified via reduction to the more soluble As(III) by As(V)-respiring bacteria. This investigation focused on the reductive dissolution of As(V) sorbed onto Fe(III)-(oxyhydr)oxide by model Fe(III)- and As(V)-reducing bacteria, to elucidate the mechanisms underpinning these processes at the single cell scale. Axenic cultures of Shewanella sp. ANA-3 wild-type cells (able to respire both Fe(III) and As(V)) were grown using C-labelled lactate on an arsenical Fe(III)- (oxyhydr)oxide thin film, and after colonisation, the distribution of Fe and As in the solid phase was assessed using nanoscale secondary ion mass spectrometry (NanoSIMS), complemented with aqueous geochemistry analyses. Parallel experiments were conducted using an arrA mutant, able to respire Fe(III) but not As(V). NanoSIMS imaging showed that most metabolically active cells were not in direct contact with the Fe(III) mineral. Flavins were released by both strains, suggesting that these cell-secreted electron shuttles mediated extracellular Fe(III)-(oxyhydr)oxide reduction, but did not facilitate extracellular As(V) reduction, demonstrated by the presence of flavins yet lack of As(III) in the supernatants of the arrA deletion mutant strain. 3D reconstructions of NanoSIMS depth-profiled single cells revealed that As and Fe were associated with the cell surface in the wild-type cells, whereas for the arrA mutant only Fe was associated with the biomass. These data were consistent with Shewanella sp. ANA-3 respiring As(V) in a multistep process; first the reductive dissolution of the Fe(III) mineral released As(V), and once in solution, As(V) was respired by the cells to As(III). As well as highlighting Fe(III) reduction as the primary release mechanism for arsenic, our data also identified unexpected cellular As(III) retention mechanisms that require further investigation.Mexican National Council for Science and Technology (CONACyT)Natural Environment Research Council (NERC

Assessment of oxidative metabolism

Oxidative metabolism is one of the central physiological processes that regulate multiple functions in a cell including cell death and survival, proliferation, gene transcription, and protein modification. There are multitudes of techniques that are used to evaluate oxidative activity. Here, we summarize how to measure oxidative activity by flow cytometry. This versatile technique allows the evaluation of the level of oxidative activity within heterogeneous populations of cells and in cell culture. Flow cytometry is a quick method that yields highly reproducible results with small sample volumes. Therefore, it is an ideal technique for evaluating changes in oxidative activity in samples from mice

Open versus closed view autorefraction in young adults

Purpose: While there are numerous studies comparing open-view autorefractors to subjective refraction or other open-view autorefractors, most studies between closed and open-view autorefraction tend to focus on children rather than young adults. The aim of this study was to determine the concordance in non-cycloplegic refractive error between two modern objective autorefractors: the closed-view monocular Topcon TRK-2P and the binocular open-view Grand Seiko WR-5500. Methods: Fifty young adults aged 20–29 years (mean age 22 ± 1.6 years) underwent non-cycloplegic autorefraction using the Grand Seiko WAM-5500 (open view) and Topcon TRK-2P (closed-view) autorefractors on both eyes. Findings were expressed as the isolated spherical component and were also converted from clinical to vector notation: Mean Spherical Error (MSE) and the astigmatic components J0 and J45. Results: Mean MSE ± SD was −1.00 ± 2.40D for the Grand Seiko WAM-5500 compared to −1.23 ± 2.29D for the Topcon TRK-2P. Up to seventy-six percent of the cohort had mean spherical errors from the Topcon TRK-2P which fell within ±0.50D of the Grand Seiko reading and 58% fell within ±0.25D. Mean differences between the two instruments were statistically significant for all components (J0, spherical, and MSE) (p  0.05). Conclusions: The differences in non-cycloplegic MSE between these two instruments are small, but statistically significant. From a clinical perspective the Topcon TRK-2P may serve as a useful starting point for subjective refraction, but additional work is needed to help further minimise differences between the instruments

Ezrin interacts with the SARS coronavirus spike protein and restrains infection at the entry stage

© 2012 Millet et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.Background: Entry of Severe Acute Respiratory Syndrome coronavirus (SARS-CoV) and its envelope fusion with host cell membrane are controlled by a series of complex molecular mechanisms, largely dependent on the viral envelope glycoprotein Spike (S). There are still many unknowns on the implication of cellular factors that regulate the entry process. Methodology/Principal Findings: We performed a yeast two-hybrid screen using as bait the carboxy-terminal endodomain of S, which faces the cytosol during and after opening of the fusion pore at early stages of the virus life cycle. Here we show that the ezrin membrane-actin linker interacts with S endodomain through the F1 lobe of its FERM domain and that both the eight carboxy-terminal amino-acids and a membrane-proximal cysteine cluster of S endodomain are important for this interaction in vitro. Interestingly, we found that ezrin is present at the site of entry of S-pseudotyped lentiviral particles in Vero E6 cells. Targeting ezrin function by small interfering RNA increased S-mediated entry of pseudotyped particles in epithelial cells. Furthermore, deletion of the eight carboxy-terminal amino acids of S enhanced S-pseudotyped particles infection. Expression of the ezrin dominant negative FERM domain enhanced cell susceptibility to infection by SARS-CoV and S pseudotyped particles and potentiated S-dependent membrane fusion. Conclusions/Significance: Ezrin interacts with SARS-CoV S endodomain and limits virus entry and fusion. Our data present a novel mechanism involving a cellular factor in the regulation of S-dependent early events of infection.This work was supported by the Research Grant Council of Hong Kong (RGC#760208)and the RESPARI project of the International Network of Pasteur Institutes

A structural comparison of human serum transferrin and human lactoferrin

The transferrins are a family of proteins that bind free iron in the blood and bodily fluids. Serum transferrins function to deliver iron to cells via a receptor-mediated endocytotic process as well as to remove toxic free iron from the blood and to provide an anti-bacterial, low-iron environment. Lactoferrins (found in bodily secretions such as milk) are only known to have an anti-bacterial function, via their ability to tightly bind free iron even at low pH, and have no known transport function. Though these proteins keep the level of free iron low, pathogenic bacteria are able to thrive by obtaining iron from their host via expression of outer membrane proteins that can bind to and remove iron from host proteins, including both serum transferrin and lactoferrin. Furthermore, even though human serum transferrin and lactoferrin are quite similar in sequence and structure, and coordinate iron in the same manner, they differ in their affinities for iron as well as their receptor binding properties: the human transferrin receptor only binds serum transferrin, and two distinct bacterial transport systems are used to capture iron from serum transferrin and lactoferrin. Comparison of the recently solved crystal structure of iron-free human serum transferrin to that of human lactoferrin provides insight into these differences

Cryptosporidium Priming Is More Effective than Vaccine for Protection against Cryptosporidiosis in a Murine Protein Malnutrition Model

Cryptosporidium is a major cause of severe diarrhea, especially in malnourished children. Using a murine model of C. parvum oocyst challenge that recapitulates clinical features of severe cryptosporidiosis during malnutrition, we interrogated the effect of protein malnutrition (PM) on primary and secondary responses to C. parvum challenge, and tested the differential ability of mucosal priming strategies to overcome the PM-induced susceptibility. We determined that while PM fundamentally alters systemic and mucosal primary immune responses to Cryptosporidium, priming with C. parvum (106 oocysts) provides robust protective immunity against re-challenge despite ongoing PM. C. parvum priming restores mucosal Th1-type effectors (CD3+CD8+CD103+ T-cells) and cytokines (IFNγ, and IL12p40) that otherwise decrease with ongoing PM. Vaccination strategies with Cryptosporidium antigens expressed in the S. Typhi vector 908htr, however, do not enhance Th1-type responses to C. parvum challenge during PM, even though vaccination strongly boosts immunity in challenged fully nourished hosts. Remote non-specific exposures to the attenuated S. Typhi vector alone or the TLR9 agonist CpG ODN-1668 can partially attenuate C. parvum severity during PM, but neither as effectively as viable C. parvum priming. We conclude that although PM interferes with basal and vaccine-boosted immune responses to C. parvum, sustained reductions in disease severity are possible through mucosal activators of host defenses, and specifically C. parvum priming can elicit impressively robust Th1-type protective immunity despite ongoing protein malnutrition. These findings add insight into potential correlates of Cryptosporidium immunity and future vaccine strategies in malnourished children

International Veterinary Epilepsy Task Force consensus proposal: Medical treatment of canine epilepsy in Europe

In Europe, the number of antiepileptic drugs (AEDs) licensed for dogs has grown considerably over the last years. Nevertheless, the same questions remain, which include, 1) when to start treatment, 2) which drug is best used initially, 3) which adjunctive AED can be advised if treatment with the initial drug is unsatisfactory, and 4) when treatment changes should be considered. In this consensus proposal, an overview is given on the aim of AED treatment, when to start long-term treatment in canine epilepsy and which veterinary AEDs are currently in use for dogs. The consensus proposal for drug treatment protocols, 1) is based on current published evidence-based literature, 2) considers the current legal framework of the cascade regulation for the prescription of veterinary drugs in Europe, and 3) reflects the authors’ experience. With this paper it is aimed to provide a consensus for the management of canine idiopathic epilepsy. Furthermore, for the management of structural epilepsy AEDs are inevitable in addition to treating the underlying cause, if possible

Identification of mutations in the PYRIN-containing NLR genes (NLRP) in head and neck squamous cell carcinoma

Head and Neck Squamous Cell Carcinoma (HNSCC) encompasses malignancies that arise in the mucosa of the upper aerodigestive tract. Recent high throughput DNA sequencing revealed HNSCC genes mutations that contribute to several cancer cell characteristics, including dysregulation of cell proliferation and death, intracellular proinflammatory signaling, and autophagy. The PYRIN-domain containing NLR (Nucleotide-binding domain, Leucine rich Repeats - containing) proteins have recently emerged as pivotal modulators of cell death, autophagy, inflammation, and metabolism. Their close physiologic association with cancer development prompted us to determine whether mutations within the NLRP (PYRIN-containing NLR ) gene family were associated with HNSCC genome instability and their clinicopathologic correlations. Catastrophic mutational events underlie cancer cell genome instability and mark a point-of-no-return in cancer cell development and generation of heterogeneity. The mutation profiles of 62 patients with primary conventional type HNSCC excluding other histologic variants were analyzed. Associations were tested using Fisher's Exact test or Mann-Whitney U test. Mutations in NLRP were associated with elevated genome instability as characterized by higher mutation rates. Clinically, NLRP mutations were more frequently found in HNSCC arising in the floor of mouth (50.0%) in comparison with HNSCC at other head and neck locations (14.8%). These mutations were clustered at the leucine rich repeats region of NLRP proteins, and affected NLRP genes were mostly localized at chromosomes 11p15.4 and 19q13.42-19q13.43. Twenty novel NLRP mutations were identified in HNSCC, and mutations in this group of genes were correlated with increased cancer cell genome mutation rates, and such features could be a potential molecular biomarker of HNSCC genome instability. © 2014 Lei et al

S-100 protein positive cells in nasopharyngeal carcinoma (NPC): absence of prognostic significance. A clinicopathological and immunohistochemical study of 40 cases

An immunohistochemical study of S-100 protein in 43 nasopharyngeal carcinomas (NPC) of known clinical evolution (33 primary and 10 metastatic) is presented. Sixty per cent of primary site cases as well as all metastatic forms showed S-100 protein positive cells intermingled with tumour cells. These S-100 positive elements were identified as Langerhans cells. No significant differences were found when correlating S-100 protein positivity and histological NPC variants, neither in age nor in sex of patients. Statistical analysis failed to demonstrate any positive correlation between S-100 protein reactivity and clinical survival