Surface Percolation and Growth. An alternative scheme for breaking the diffraction limit in optical patterning

A nanopatterning scheme is presented by which the structure height can be controlled in the tens of nanometers range and the lateral resolution is a factor at least three times better than the point spread function of the writing beam. The method relies on the initiation of the polymerization mediated by a very inefficient energy transfer from a fluorescent dye molecule after single photon absorption. The mechanism has the following distinctive steps: the dye adsorbs on the substrate surface with a higher concentration than in the bulk, upon illumination it triggers the polymerization, then isolated islands develop and merge into a uniform structure (percolation), which subsequently grows until the illumination is interrupted. This percolation mechanism has a threshold that introduces the needed nonlinearity for the fabrication of structures beyond the diffraction limit.Comment: 10 pages, 8 figure

Detection of Parechovirus (P) and Enterovirus (E) Among Infants Evaluated for Late-Onset Sepsis in the Neonatal Intensive Care Unit (NICU): The VIRIoN-P-E Study

Background: Limited data exist on the role of human parechoviruses (HPeV) and enteroviruses (EV) as causes of late-onset sepsis (LOS) in the NICU. Objective: To determine the frequency of detection of parechoviruses and enteroviruses among infants >72 hr of age who were evaluated for LOS in 2 academic NICUs (Parkland Memorial Hospital [PMH], Dallas -shared bays; Women & Infants Hospital [W&I], RI -single patient rooms) Design/Methods: Prospective cohort study of inborn infants hospitalized in the NICU at PMH and WIH from 1/2012 to 1/2013 and were enrolled in the Viral Respiratory Infections in the Neonatal Intensive Care Unit (VIRIoN-I; J Pediatr 2014:165:690). Eligible subjects were infants of all gestational ages (GA) and birth weights (BW) who were >72 hrs of age, remained in the NICU since birth, and underwent evaluation with initiation of antibiotic therapy for suspected LOS. Nasopharyngeal specimens were obtained within 72 hrs of the sepsis evaluation using flexible flocked nylon swabs that were placed in universal transport medium and frozen at -80\ub0C until tested for parechovirus and enterovirus RNA by polymerase chain reaction (PCR) assay (Virology Laboratory, Nationwide Children\u2019s Hospital, Columbus, OH). Demographic, clinical, laboratory, and radiographic data were obtained. Results: Of the 100 infants enrolled in the VIRIoN-I study, nasopharyngeal specimens were available from 65 (59, PMH; 6, WIH) for parechovirus and enterovirus PCR testing. These 65 infants (38, male; 27, female; 49, Hispanic; 6, white; 9, Black; 1, unknown) had a mean \ub1SD gestational age of 30 \ub1 5 wks and birth weight of 1619 \ub1 929 g, and received empirical antibiotics for possible LOS. Infants had a total of 94 sepsis evaluations (65, 1 evaluation; 16, 2; 8, 3; 4, 4) at a mean age of 20 days. Reasons for the sepsis evaluations included fever (9), hypothermia (65), apnea (50),feeding intolerance (51), seizure (1), irritabilitiy (5), emesis (20), diarrhea (1), bloody stool (5), rhinorrhea/congestion/cough (6), and lethargy (9). Four infants died. None of the infants had parechovirus or enterovirus detected in nasopharygeal specimens either at the first or subsequent sepsis evaluations. Conclusion(s): The burden of disease due to parechovirus and enteroviruses among inborn infants who remain in the NICU since birth appears to be low in those evaluated for LOS. Larger, prospective studies are needed to fully determine their contribution to \u201cculture-negative\u201d sepsis in the NICU. Publication Number: 3860.53

Imaging the Electrocyte of Torpedo Marmorata by Scanning Force Microscopy

Scanning force microscopy (SFM) and scanning electron microscopy (SEM) were used to examine the tissue structure of the electric organ of Torpedo marmorata in air and in liquid after applying fracturing and cryosectioning techniques and chemical fixation. The electric organ is organized in columns of stacked electrocytes, arranged in a honeycomb pattern. The columns were cut along a plane normal to the cell stack and thin sections were transferred to polylysine coated glass coverslips. The polarity of the electrocytes was made apparent by immunofluorescence microscopy directed to different domains of the acetylcholine receptor (AChR), thus revealing the innervated face of the cell. SFM and SEM both showed cell surfaces to be overlaid by a network of collagen fibers by their characteristic banding pattern with about 64 nm periodicity and about 2.5 nm corrugation amplitude. In liquid, significantly lower structural resolution was achieved by SFM, probably due to sample elasticity

Interstitial lung disease in a newborn affected by mevalonic aciduria

Introduction: Mevalonic aciduria (MA) is the most severe phenotype of mevalonate-kinase deficiency (MKD), with a onset in early infancy and poor prognosis. MA diagnosis may be challenging in the neonatal period given its rarity and its unspecific symptoms that frequently recall those of other neonatal diseases. To our knowledge, interstial lung involvement has never been described as onset feature in a newborn with MKD. Objectives: We report the case of a newborn affected by MKD characterized by interstitial lung disease. Methods: The patient underwent laboratory and radiology evaluation as clinically indicated. Direct Sanger sequencing was used to screen the 10 exons of the MVK gene. Results: A female neonate born at term from consanguineous parents was referred to our hospital at 16 days of life (DOL) for mild hypotonia and persistent raised inflammatory markers despite antibiotic therapy. Infectious work-up was negative for both viral and bacterial infections. Chest x-ray revealed bilateral perihilar peribronchial thickening. Electroencephalography (EEG) reported moderate diffuse anomalies of background activity without major abnormalities. On DOL 20 the first episode of fever was recorded. Due to worsening tachypnea and persistent abnormal chest x-ray, a pulmonary CT scan was performed and showed diffuse ground-glass bilateral infiltrates consistent with alveolar-interstitial lung disease. On DOL 22 a palpable maculo-papular skin rash appeared on feet and hands, vanishing spontaneously 24 hours later. Bone marrow examination and levels of perforins, neuron-specific enolase and urinary catabolites of catecholamines were normal. A total body MRI was normal except for a mild cerebellar hypoplasia and the known interstitial lung disease. The patient kept presenting hypotonia, relapsing episodes of fever and skin rashes, developed anemia requiring blood transfusions and failure to thrive became evident. Type-I IFN signature was negative. A genetic test was requested, as well as quantification of urinary levels of mevalonic acid, which were markedly above the normal range. Direct Sanger sequencing allowed to detect a homozygous c.709A>T missense mutation in the exon 8 of the MVK gene, coding for a protein substitution p.T237S already classified as pathogenic in the INFEVERS database (http://fmf.igh.cnrs.fr/ISSAID/infevers/) and therefore consistent with the diagnosis of MKD. Both parents and her sister were found to be heterozygous carriers of the same mutation. On DOL 38 treatment with anakinra was started, with prompt regression of fever and skin rash, decrease in inflammatory markers, increase in reticulocytes count and weight gain. Hypotonia improved but persisted. The patient was discharged from hospital on DOL 56 in good clinical conditions, with acute phase reactants within the normal range and mild hypotonia. She is now 4 months old, still on anakinra treatment without adverse events. Conclusion: Autoinflammatory diseases in the neonatal period are a diagnostic challenge. Clinical suspicion is crucial in order to perform specific laboratory and genetic testing and start appropriate treatment. Interstitial lung involvement may be present in MKD and, together with increased inflammatory markers, could be the first manifestation of the disease

Determining the reference range of blood presepsin in term and preterm neonates

Introduction Sepsis is still a major cause of morbidity and mortality in neonates, especially in preterm infants. Mortality can reach 60-70% in very low birth weight infants (birthweight 0.20). This reduced model explains 3.8% of the total sum of squares. After adjustment for all the factors in the model, presepsin levels appear to be significantly lower in twins (496 pg/ml \uf0b1 65.5 vs 655 pg/ml \uf0b1 11.8) and in neonates with Apgar at 1 min 658 (644 pg/ml \uf0b1 11.8 vs 774 pg/ml \uf0b1 56.2). So none of the above factors seems worth to be taken into account in determining the reference limits for presepsin blood levels in healthy term neonates. Preterm neonates. The largest differences in presepsin level are observed between small for gestational age (SGA) (903 pg/ml \uf0b1 57.1) and adequate for gestational age (AGA) neonates (703 pg/ml \uf0b1 26.7), between neonates with and without mechanical ventilation at blood sampling (1090 pg/ml \uf0b1 86.9 vs 711 pg/ml \uf0b1 24.7) and at delivery (855 pg/ml \uf0b1 87.3 vs 729 pg/ml \uf0b1 25.8), between neonates with and without venous catether (801 pg/ml \uf0b1 47.5 vs 716 pg/ml \uf0b1 28.9), between neonates who underwent blood sampling after the 4th day or before (797 pg/ml \uf0b1 46.2 vs 716 pg/ml \uf0b1 29.2), between males and females (778 pg/ml \uf0b1 35.1 vs 701 pg/ml \uf0b1 34.7). All these factors, when simultaneously introduced into a multivariable linear model, explain only 18.8% of the total sum of squares. A second multivariable linear model was fitted after removing the factors that showed the lowest effect on presepsin level (those associated with a p-value >0.50). This reduced model explains 13.4% of the total sum of squares. A third and more parsimonious multivariable linear model was fitted after removing the factors that showed the lowest effect on presepsin level (those associated with a p-value >0.20). This reduced model explains 12.3% of the total sum of squares. After adjustment for all the factors in the model, presepsin levels result to be significantly lower in AGA neonates (706 pg/ml \uf0b1 25.7 vs 890 pg/ml \uf0b1 55.0) and between neonates with and without mechanical ventilation at blood sampling (1074 pg/ml \uf0b1 85.3 vs 712 pg/ml \uf0b1 24.2). Even in this case, none of the above factors is expected to substantially affect the reference limits for presepsin blood levels in preterm neonates. Conclusion Presepsin blood levels seem to be quite independent of most of maternal and neonatal conditions examined in this study both in preterm and term neonates. The factors exerting significant effects (multiple birth and Apgar at 1 min, in term neonates, weight by gestational age and mechanical ventilation in preterm neonates) are expected to affect presepsin reference limits only to minor extent. References [1] Evaluation of a newly identified soluble CD14 subtype as a marker for sepsis. Yaegashi Y., Shirakawa K., Sato N., Suzuki Y., Kojika M., Imai S., Takahashi G., Miyata M., Furusako S., Endo S. J Infect Chemother. 2005;11:234-8. [2] CDC/NHSN surveillance definition of health care-associated infection and criteria for specific types of infection in the acute care setting. Horan T.C., Andrus M., Dudeck M.A. Am J Infect Control. 2008;36:309-32

Rab27a controls HIV-1 assembly by regulating plasma membrane levels of phosphatidylinositol 4,5-bisphosphate

During the late stages of the HIV-1 replication cycle, the viral polyprotein Pr55(Gag) is recruited to the plasma membrane (PM), where it binds phosphatidylinositol 4,5-bisphosphate (PI(4,5)P2) and directs HIV-1 assembly. We show that Rab27a controls the trafficking of late endosomes carrying phosphatidylinositol 4-kinase type 2 α (PI4KIIα) toward the PM of CD4(+) T cells. Hence, Rab27a promotes high levels of PM phosphatidylinositol 4-phosphate and the localized production of PI(4,5)P2, therefore controlling Pr55(Gag) membrane association. Rab27a also controls PI(4,5)P2 levels at the virus-containing compartments of macrophages. By screening Rab27a effectors, we identified that Slp2a, Slp3, and Slac2b are required for the association of Pr55(Gag) with the PM and that Slp2a cooperates with Rab27a in the recruitment of PI4KIIα to the PM. We conclude that by directing the trafficking of PI4KIIα-positive endosomes toward the PM, Rab27a controls PI(4,5)P2 production and, consequently, HIV-1 replication.Universidad de Buenos Aires and CONICET doctoral fellowships, Agencia Nacional de Pro- moción Científica y Tecnológica (Argentina) grants: (2010-1681, 2012-00353), Creative and Novel Ideas in HIV Research Program, University of Alabama at Birmingham Center for AIDS Research funding grant P30 AI027767-24

Vascular Endothelium in Neonatal Sepsis: Basic Mechanisms and Translational Opportunities

Neonatal sepsis remains a major health issue worldwide, especially for low-birth weight and premature infants, with a high risk of death and devastating sequelae. Apart from antibiotics and supportive care, there is an unmet need for adjunctive treatments to improve the outcomes of neonatal sepsis. Strong and long-standing research on adult patients has shown that vascular endothelium is a key player in the pathophysiology of sepsis and sepsis-associated organ failure, through a direct interaction with pathogens, leukocytes, platelets, and the effect of soluble circulating mediators, in part produced by endothelial cells themselves. Despite abundant evidence that the neonatal immune response to sepsis is distinct from that of adults, comparable knowledge on neonatal vascular endothelium is much more limited. Neonatal endothelial cells express lower amounts of adhesion molecules compared to adult ones, and present a reduced capacity to neutralize reactive oxygen species. Conversely, available evidence on biomarkers of endothelial damage in neonates is not as robust as in adult patients, and endothelium-targeted therapeutic opportunities for neonatal sepsis are almost unexplored. Here, we summarize current knowledge on the structure of neonatal vascular endothelium, its interactions with neonatal immune system and possible endothelium-targeted diagnostic and therapeutic tools for neonatal sepsis. Furthermore, we outline areas of basic and translational research worthy of further study, to shed light on the role of vascular endothelium in the context of neonatal sepsis

Impact of different stages of intrauterine inflammation on outcome of preterm neonates: Gestational age-dependent and-independent effect

Objective: To investigate the impact of different stages of intrauterine inflammation (IUI) on neonatal outcomes, before and after adjusting for gestational age (GA) and other perinatal confounders. Methods: This was an observational, prospective, single-center cohort study including all eligible neonates with GA < 35 weeks and/or birth weight 64 1500 g born at a 3 rd level Neonatal Intensive Care Unit between 2011 and 2014. Pathological patterns of placenta, membranes and cord were classified according to Redline's criteria. Multivariable linear and logistic regression models were applied, either including or not GA among the covariates. Results: Of the 807 enrolled neonates, 134 (16.6%) had signs of IUI: among these, 54.5% showed just histological chorioamnionitis (HCA), 25.4% had HCA + funisitis (FUN) stage 1, and 20.1% had HCA + FUN stage 2-3. At univariate analysis, HCA increased the risk for retinopathy of prematurity (ROP) and bronchopulmonary dysplasia, while FUN (any stage) had a deleterious impact on all outcomes investigated. After adjustment for covariates not including GA, HCA was a risk factor only for ROP (OR = 2.8, CI: 1-7.8), while FUN (any stage) was still associated with increased ORs for all outcomes (p <0.01). Upon inclusion of GA in the regression model, the results differed remarkably. HCA was associated with lower risk for mechanical ventilation (OR = 0.3, CI: 0.1-0.7) and need for surfactant (OR = 0.5, CI: 0.2-0.9), while FUN (any stage) worsened clinical conditions at birth (p <0.05), increased the risk for early-onset sepsis (p <0.01), and increased the length of mechanical ventilation (FUN stage 2-3 only, RC = 6.5 days, CI: 2-11). No other outcome was affected. Conclusions: IUI, especially FUN, negatively impact most neonatal morbidities, but its effect is partially reverted adjusting for GA. Considered that GA is an intermediate variable interposed between prenatal causes of prematurity and outcomes, the appropriateness of adjusting for GA may be questionable

Prevalence, Outcome, and Prevention of Congenital Cytomegalovirus Infection in Neonates Born to Women With Preconception Immunity (CHILd Study)

Background. Human cytomegalovirus (HCMV) is the leading infectious cause of congenital disabilities. We designed a prospective study to investigate the rate, outcome, and risk factors of congenital CMV (cCMV) infection in neonates born to immune women, and the potential need and effectiveness of hygiene recommendations in this population.Methods. The study was composed of 2 sequential parts: an epidemiology (part 1) and a prevention (part 2) study. Performance of part 2 depended upon a cCMV rate >0.4%. Women enrolled in part 1 did not receive hygiene recommendations. Newborns were screened by HCMV DNA testing in saliva and cCMV was confirmed by urine testing.Results. Saliva swabs were positive for HCMV DNA in 45/9661 newborns and cCMV was confirmed in 18 cases. The rate of cCMV was .19% (95% confidence interval [CI]: .11-.29%), and 3 out of 18 infants with cCMV had symptoms of CMV at birth. Age, nationality, occupation, and contact with children were similar between mothers of infected and noninfected newborns. Twin pregnancy (odds ratio [OR]: 7.2; 95% CI: 1.7-32.2; P=.037) and maternal medical conditions (OR: 3.9; 95% CI: 1.5-10.1; P= .003) appeared associated with cCMV. Given the rate of cCMV was lower than expected, the prevention part of the study was cancelled.Conclusions. Newborns from women with preconception immunity have a low rate of cCMV, which appears to be mostly due to reactivation of the latent virus. Therefore, serological screening in childbearing age would be pivotal to identify HCMV-seropositive women, whose newborns have a low risk of cCMV