L-ergothioneine reduces nitration of lactoferrin and loss of antibacterial activity associated with nitrosative stress.

Lactoferrin (LF) is a multifunctional antimicrobial, anti-inflammatory, and antioxidant protein that occurs naturally in mammals, most notably in exocrine gland tissues and fluids, such as in the eye. Nitrosative stress can promote changes to tyrosine and other amino acid residues of the protein, which also reduces the activity of LF. l-ergothioneine (ET) is a potent anti-inflammatory antioxidant present in the eye and other tissues through nutrition or supplementation and that may play a role in the prevention or treatment of a variety of diseases. Here we investigated the ability of ET to reduce 3-nitrotyrosine (NTyr) formation using two separate substrates, with the goal of determining whether ET can protect the antibacterial function of LF and other proteins when exposed separately to peroxynitrite and tetranitromethane as nitrating reagents. Native human LF was used as a simple protein substrate, and lamb corneal lysate was chosen as one example of mammalian tissue with a more complex mixture of proteins and other biomolecules. Nitration was monitored by absorbance and fluorescence spectroscopy as well as sandwich (nitrated LF) and direct NTyr (corneal lysate) enzyme-linked immunosorbent assays (ELISAs). We found that pretreatment with ET reduced chemical modification of both native LF and corneal lysate samples and loss of antibacterial LF function due to exposure to the nitrating reagents. These initial results suggest that ET, raised to sufficiently elevated levels, could be tailored as a therapeutic agent to reduce effects of nitrosative stress on LF and in turn sustain the protein activity.

Room-level ventilation in schools and universities.

Ventilation is of primary concern for maintaining healthy indoor air quality and reducing the spread of airborne infectious disease, including COVID-19. In addition to building-level guidelines, increased attention is being placed on room-level ventilation. However, for many universities and schools, ventilation data on a room-by-room basis are not available for classrooms and other key spaces. We present an overview of approaches for measuring ventilation along with their advantages and disadvantages. We also present data from recent case studies for a variety of institutions across the United States, with various building ages, types, locations, and climates, highlighting their commonalities and differences, and examples of the use of this data to support decision making.

Heterogeneous nitration reaction of BSA protein with urban air: improvements in experimental methodology.

Gas-phase ozone (O3) and nitrogen dioxide (NO2) can react with environmentally exposed proteins to induce chemical modifications such as the formation of nitrotyrosine (NTyr). Certain proteins with these modifications have also been shown to promote adverse health effects and can trigger an immune response. It is hypothesized that proteinaceous material suspended in the atmosphere as particulate matter, e.g., embedded in pollen, can undergo heterogenous reactions to produce chemically modified proteins that impact human health, especially in urban areas. To investigate the protein modification process under ambient outdoor reaction conditions, bovine serum albumin (BSA) protein samples were loaded onto filters and exposed to urban air in Denver, Colorado (USA). Losses and measurement artifacts were measured independently to calculate nitration effects on the protein via high-performance liquid chromatography and to support the experimental methodology. O3 loss from inlet lines using three commonly used particulate filters was quantified, showing a range of ambient O3 concentration losses from 3.2% for Kynar® (polyvinylidene fluoride) filters to > 60% for commonly used HEPA filters. Protein mass extraction efficiency was calculated as a function of filter material and protein mass using both native and nitrated BSA. Finally, we show examples of BSA samples nitrated by exposure to urban air as a proof-of-concept for future studies, highlighting the potential for atmospherically relevant NTyr formation. The methodology vetted here provides support for a wide variety of experimental efforts related to exposure of analytes to O3 and more broadly to an expanding field of protein modification in ambient air.

Modification of lactoferrin by peroxynitrite reduces its antibacterial activity and changes protein structure.

Lactoferrin (LF) is a multifunctional protein that plays important physiological roles as one of the most concentrated proteins in many human and other mammalian fluids and tissues. In particular, LF provides antibacterial properties to human milk, saliva, and tear fluid. LF also protects against stress-induced lipid peroxidation at inflammation sites through its iron-binding ability. Previous studies have shown that LF can be efficiently nitrated via biologically relevant mediators such as peroxynitrite (ONOO- ), which are also present at high intracellular concentrations during inflammation and nitrosative stress. Here, we examine changes in antibacterial properties and structure of LF following ONOO- treatment. The reaction induces nitration of tyrosine and tryptophan residues, which are commonly used as biomarker molecules for several diseases. Treatment with ONOO- at a 10/1 M ratio of ONOO- to tyrosine inhibited all antibacterial activity exhibited by native LF. Secondary structural changes in LF were assessed using circular dichroism spectroscopy. Nitration products with and without the addition of Fe3+ show significant reduction in alpha-helical properties, suggesting partial protein unfolding. Iron-binding capacity of LF was also reduced after treatment with ONOO- , suggesting a decreased ability of LF to protect against cellular damage. LC-MS/MS spectrometry was used to identify LF peptide fragments nitrated by ONOO- , including tyrosine residue Y92 located in the iron-binding region. These results suggest that posttranslational modification of LF by ONOO- could be an important pathway to exacerbate infection, for example, in inflamed tissues and to reduce the ability of LF to act as an immune responder and decrease oxidative damage.

Examining the current standards for genetic discovery and replication in the era of mega-biobanks.

With the recent deluge of mega-biobank data, it is time to revisit what constitutes "replication" for genome-wide association studies. Many replication samples are unavailable or underpowered, therefore alternatives beyond strict statistical replication are needed until the required resources become available.

Development and characterization of an inexpensive single-particle fluorescence spectrometer for bioaerosol monitoring.

Laser-induced fluorescence (LIF) techniques to analyze atmospheric aerosols are commonly applied for research and human exposure monitoring, but are very expensive or offer poor spectral resolution. Here, we discuss how a recently proposed instrument can acquire resolved fluorescence spectra from many individual particles in a single camera image using four excitation wavelengths matched with common biological fluorophores for particle discrimination at lower cost. We discuss emission intensity calibration and demonstrate spectral differentiation among four species of pollen. These data provide context for how the instrument could be developed for pollen and mold-spore detection or for use by citizen scientists.

Outbreak of Salmonella Chailey infections linked to precut coconut pieces - United States and Canada, 2017†.

Foodborne salmonellosis causes an estimated one million illnesses and 400 deaths annually in the United States (US). During March-May 2017, an outbreak of 19 cases of Salmonella Chailey associated with precut coconut pieces from a single grocery store chain occurred in the United States and Canada. The chain voluntarily recalled precut coconut pieces. This was the first time that coconut has been associated with a Salmonella outbreak in the United States or Canada. In recent years, salmonellosis outbreaks have been caused by foods not typically associated with Salmonella. Raw coconut should now be considered in investigations of Salmonella outbreaks among fresh food consumers.

Development of a sandwich ELISA with potential for selective quantification of human lactoferrin protein nitrated through disease or environmental exposure.

Lactoferrin (LF) is an important multifunctional protein that comprises a large fraction of the protein mass in certain human fluids and tissues, and its concentration is often used to assess health and disease. LF can be nitrated by multiple routes, leading to changes in protein structure, and nitrated proteins can negatively impact physiological health via nitrosative stress. Despite an awareness of the detrimental effects of nitrated proteins and the importance of LF within the body, cost-effective methods for detecting and quantifying nitrated lactoferrin (NLF) are lacking. We developed a procedure to selectively quantify NLF using sandwich enzyme-linked immunosorbent assay (ELISA), utilizing a polyclonal anti-LF capture antibody paired with a monoclonal anti-nitrotyrosine detector antibody. The assay was applied to quantify NLF in samples of pure LF nitrated via two separate reactions at molar ratios of excess nitrating agent to the total number of tyrosine residues between 10/1 and 100/1. Tetranitromethane (TNM) was used as a laboratory surrogate for an environmental pathway selective for production of 3-nitrotyrosine, and sodium peroxynitrite (ONOO-) was used as a surrogate for an endogenous nitration pathway. UV-vis spectroscopy (increased absorbance at 350 nm) and fluorescence spectroscopy (emission decreased by > 96%) for each reaction indicate the production of NLF. A lower limit of NLF detection using the ELISA method introduced here was calculated to be 0.065 µg mL-1, which will enable the detection of human-physiologically relevant concentrations of NLF. Our approach provides a relatively inexpensive and practical way to assess NLF in a variety of systems. Graphical abstract We developed a procedure to selectively quantify nitrated lactoferrin (NLF) protein using a sandwich enzyme-linked immunosorbent assay (ELISA) and verified results against several spectroscopic techniques. Our approach provides an inexpensive and practical way to assess NLF in a variety of systems.

Polygenic loading for major depression is associated with specific medical comorbidity.

Major depressive disorder frequently co-occurs with medical disorders, raising the possibility of shared genetic liability. Recent identification of 15 novel genetic loci associated with depression allows direct investigation of this question. In cohorts of individuals participating in biobanks at two academic medical centers, we calculated polygenic loading for risk loci reported to be associated with depression. We then examined the association between such loading and 50 groups of clinical diagnoses, or topics, drawn from these patients' electronic health records, determined using a novel application of latent Dirichilet allocation. Three topics showed experiment-wide association with the depression liability score; these included diagnostic groups representing greater prevalence of mood and anxiety disorders, greater prevalence of cardiac ischemia, and a decreased prevalence of heart failure. The latter two associations persisted even among individuals with no mood disorder diagnosis. This application of a novel method for grouping related diagnoses in biobanks indicate shared genetic risk for depression and cardiac disease, with a pattern suggesting greater ischemic risk and diminished heart failure risk.

Sources and dynamics of fluorescent particles in hospitals.

Fluorescent particles can be markers of bioaerosols and are therefore relevant to nosocomial infections. To date, little research has focused on fluorescent particles in occupied indoor environments, particularly hospitals. In this study, we aimed to determine the spatial and temporal variation of fluorescent particles in two large hospitals in Brisbane, Australia (one for adults and one for children). We used an Ultraviolet Aerodynamic Particle Sizer (UVAPS) to identify fluorescent particle sources, as well as their contribution to total particle concentrations. We found that the average concentrations of both fluorescent and non-fluorescent particles were higher in the adults' hospital (0.06×106 and 1.20×106  particles/m3 , respectively) than in the children's hospital (0.03×106 and 0.33×106  particles/m3 , respectively) (P<.01). However, the proportion of fluorescent particles was higher in the children's hospital. Based on the concentration results and using activity diaries, we were able to identify sources of particle production within the two hospitals. We demonstrated that particles can be easily generated by a variety of everyday activities, which are potential sources of exposure to pathogens. Future studies to further investigate their role in nosocomial infection are warranted.

Psychological interventions to reduce suicidality in high-risk patients with major depression: a randomized controlled trial.

BACKGROUND: Positive psychological constructs have been associated with reduced suicidal ideation, and interventions to cultivate positive feelings have the potential to reduce suicide risk. This study compares the efficacy of a 6-week, telephone-based positive psychology (PP) intervention against a cognition-focused (CF) control intervention among patients recently hospitalized for depression and suicidal ideation or behavior. METHOD: A total of 65 adults with a current major depressive episode reporting suicidal ideation or a recent suicide attempt were enrolled from participating in-patient psychiatric units. Prior to discharge, participants were randomized to the PP (n = 32) or CF (n = 33) intervention. In both interventions, participants received a treatment manual, performed weekly PP (e.g. gratitude letter) or CF (e.g. recalling daily events) exercises, and completed weekly one-on-one telephone sessions over 6 weeks. Between-group differences in hopelessness (primary outcome), depression, suicidality and positive psychological constructs at 6 and 12 weeks were tested using mixed-effects models accounting for intensity of post-hospitalization psychiatric treatment. RESULTS: Compared with PP, the CF intervention was associated with significantly greater improvements in hopelessness at 6 weeks (ß = -3.15, 95% confidence interval -6.18 to -0.12, effect size = -0.84, p = 0.04), but not 12 weeks. Similarly, the CF intervention led to greater improvements in depression, suicidal ideation, optimism and gratitude at 6 and 12 weeks. CONCLUSIONS: Contrary to our hypothesis, the CF intervention was superior to PP in improving hopelessness, other suicide risk factors and positive psychological constructs during a key post-discharge period among suicidal patients with depression. Further study of this CF intervention is warranted in populations at high suicide risk.

Chamber bioaerosol study: human emissions of size-resolved fluorescent biological aerosol particles.

Humans are a prominent source of airborne biological particles in occupied indoor spaces, but few studies have quantified human bioaerosol emissions. The chamber investigation reported here employs a fluorescence-based technique to evaluate bioaerosols with high temporal and particle size resolution. In a 75-m(3) chamber, occupant emission rates of coarse (2.5-10 µm) fluorescent biological aerosol particles (FBAPs) under seated, simulated office-work conditions averaged 0.9 ± 0.3 million particles per person-h. Walking was associated with a 5-6× increase in the emission rate. During both walking and sitting, 60-70% or more of emissions originated from the floor. The increase in emissions during walking (vs. while sitting) was mainly attributable to release of particles from the floor; the associated increased vigor of upper body movements also contributed. Clothing, or its frictional interaction with human skin, was demonstrated to be a source of coarse particles, and especially of the highly fluorescent fraction. Emission rates of FBAPs previously reported for lecture classes were well bounded by the experimental results obtained in this chamber study. In both settings, the size distribution of occupant FBAP emissions had a dominant mode in the 3-5 µm diameter range.

A marine biogenic source of atmospheric ice-nucleating particles.

The amount of ice present in clouds can affect cloud lifetime, precipitation and radiative properties. The formation of ice in clouds is facilitated by the presence of airborne ice-nucleating particles. Sea spray is one of the major global sources of atmospheric particles, but it is unclear to what extent these particles are capable of nucleating ice. Sea-spray aerosol contains large amounts of organic material that is ejected into the atmosphere during bubble bursting at the organically enriched sea-air interface or sea surface microlayer. Here we show that organic material in the sea surface microlayer nucleates ice under conditions relevant for mixed-phase cloud and high-altitude ice cloud formation. The ice-nucleating material is probably biogenic and less than approximately 0.2 micrometres in size. We find that exudates separated from cells of the marine diatom Thalassiosira pseudonana nucleate ice, and propose that organic material associated with phytoplankton cell exudates is a likely candidate for the observed ice-nucleating ability of the microlayer samples. Global model simulations of marine organic aerosol, in combination with our measurements, suggest that marine organic material may be an important source of ice-nucleating particles in remote marine environments such as the Southern Ocean, North Pacific Ocean and North Atlantic Ocean.

Exploring the Balance between DNA Pressure and Capsid Stability in Herpesviruses and Phages.

UNLABELLED: We have recently shown in both herpesviruses and phages that packaged viral DNA creates a pressure of tens of atmospheres pushing against the interior capsid wall. For the first time, using differential scanning microcalorimetry, we directly measured the energy powering the release of pressurized DNA from the capsid. Furthermore, using a new calorimetric assay to accurately determine the temperature inducing DNA release, we found a direct influence of internal DNA pressure on the stability of the viral particle. We show that the balance of forces between the DNA pressure and capsid strength, required for DNA retention between rounds of infection, is conserved between evolutionarily diverse bacterial viruses (phages λ and P22), as well as a eukaryotic virus, human herpes simplex 1 (HSV-1). Our data also suggest that the portal vertex in these viruses is the weakest point in the overall capsid structure and presents the Achilles heel of the virus's stability. Comparison between these viral systems shows that viruses with higher DNA packing density (resulting in higher capsid pressure) have inherently stronger capsid structures, preventing spontaneous genome release prior to infection. This force balance is of key importance for viral survival and replication. Investigating the ways to disrupt this balance can lead to development of new mutation-resistant antivirals. IMPORTANCE: A virus can generally be described as a nucleic acid genome contained within a protective protein shell, called the capsid. For many double-stranded DNA viruses, confinement of the large DNA molecule within the small protein capsid results in an energetically stressed DNA state exerting tens of atmospheres of pressures on the inner capsid wall. We show that stability of viral particles (which directly relates to infectivity) is strongly influenced by the state of the packaged genome. Using scanning calorimetry on a bacterial virus (phage λ) as an experimental model system, we investigated the thermodynamics of genome release associated with destabilizing the viral particle. Furthermore, we compare the influence of tight genome confinement on the relative stability for diverse bacterial and eukaryotic viruses. These comparisons reveal an evolutionarily conserved force balance between the capsid stability and the density of the packaged genome.

Size-resolved fluorescent biological aerosol particle concentrations and occupant emissions in a university classroom.

UNLABELLED: This study is among the first to apply laser-induced fluorescence to characterize bioaerosols at high time and size resolution in an occupied, common-use indoor environment. Using an ultraviolet aerodynamic particle sizer, we characterized total and fluorescent biological aerosol particle (FBAP) levels (1-15 µm diameter) in a classroom, sampling with 5-min resolution continuously during eighteen occupied and eight unoccupied days distributed throughout a one-year period. A material-balance model was applied to quantify per-person FBAP emission rates as a function of particle size. Day-to-day and seasonal changes in FBAP number concentration (NF ) values in the classroom were small compared to the variability within a day that was attributable to variable levels of occupancy, occupant activities, and the operational state of the ventilation system. Occupancy conditions characteristic of lecture classes were associated with mean NF source strengths of 2 × 10(6) particles/h/person, and 9 × 10(4) particles per metabolic g CO2 . During transitions between lectures, occupant activity was more vigorous, and estimated mean, per-person NF emissions were 0.8 × 10(6) particles per transition. The observed classroom peak in FBAP size at 3-4 µm is similar to the peak in fluorescent and biological aerosols reported from several studies outdoors. PRACTICAL IMPLICATIONS: Coarse particles that exhibit fluorescence at characteristic wavelengths are considered to be proxies for biological particles. Recently developed instruments permit their detection and sizing in real time. In a mechanically ventilated classroom, emissions from human occupants were a strong determinant of coarse-mode fluorescent biological aerosol particle (FBAP) levels. Human FBAP emission rates were significant under quiet occupancy conditions and increased with activity level. Fluorescent particle emissions peaked at a diameter of 3­4 µm, which is the expected modal size of airborne particles with associated microbes. Human activity patterns, and associated coarse FBAP and total particle levels varied strongly on short timescales. Thus, the dynamic temporal behavior of aerosol concentrations must be considered when determining collection protocols for samples meant to be representative of average concentrations using time-integrated or 'snapshot' bioaerosol measurement techniques.

Bioprecipitation: a feedback cycle linking earth history, ecosystem dynamics and land use through biological ice nucleators in the atmosphere.

Landscapes influence precipitation via the water vapor and energy fluxes they generate. Biologically active landscapes also generate aerosols containing microorganisms, some being capable of catalyzing ice formation and crystal growth in clouds at temperatures near 0 °C. The resulting precipitation is beneficial for the growth of plants and microorganisms. Mounting evidence from observations and numerical simulations support the plausibility of a bioprecipitation feedback cycle involving vegetated landscapes and the microorganisms they host. Furthermore, the evolutionary history of ice nucleation-active bacteria such as Pseudomonas syringae supports that they have been part of this process on geological time scales since the emergence of land plants. Elucidation of bioprecipitation feedbacks involving landscapes and their microflora could contribute to appraising the impact that modified landscapes have on regional weather and biodiversity, and to avoiding inadvertent, negative consequences of landscape management.

Structure of the pseudorabies virus capsid: comparison with herpes simplex virus type 1 and differential binding of essential minor proteins.

The structure of pseudorabies virus (PRV) capsids isolated from the nucleus of infected cells and from PRV virions was determined by cryo-electron microscopy (cryo-EM) and compared to herpes simplex virus type 1 (HSV-1) capsids. PRV capsid structures closely resemble those of HSV-1, including distribution of the capsid vertex specific component (CVSC) of HSV-1, which is a heterodimer of the pUL17 and pUL25 proteins. Occupancy of CVSC on all PRV capsids is near 100%, compared to ~50% reported for HSV-1 C-capsids and 25% or less that we measure for HSV-1 A- and B-capsids. A PRV mutant lacking pUL25 does not produce C-capsids and lacks visible CVSC density in the cryo-EM-based reconstruction. A reconstruction of PRV capsids in which green fluorescent protein was fused within the N-terminus of pUL25 confirmed previous studies with a similar HSV-1 capsid mutant localizing pUL25 to the CVSC density region that is distal to the penton. However, comparison of the CVSC density in a 9-Å-resolution PRV C-capsid map with the available crystal structure of HSV-1 pUL25 failed to find a satisfactory fit, suggesting either a different fold for PRV pUL25 or a capsid-bound conformation for pUL25 that does not match the X-ray model determined from protein crystallized in solution. The PRV capsid imaged within virions closely resembles C-capsids with the addition of weak but significant density shrouding the pentons that we attribute to tegument proteins. Our results demonstrate significant structure conservation between the PRV and HSV capsids.

Leprosy epidemics during history increased protective allele frequency of PARK2/PACRG genes in the population of the Mljet Island, Croatia.

INTRODUCTION: Two regulatory polymorphisms (rs1040079 and rs9356058) shared by PARK2 and PACRG genes were identified as major risk variants for leprosy susceptibility. The aim of this study was to investigate if allele frequencies of these polymorphisms in the isolated population of the island of Mljet, which served as a quarantine for leprosy patients during past centuries, were different to allele frequencies in two control populations with no history of leprosy. SUBJECTS AND METHODS: This study included 88 unrelated Caucasian individuals from the island of Mljet while two control groups included 93 individuals from the island of Rab and 160 individuals from the region of Split. Genotyping for rs1040079 and rs9356058 was performed by "real-time" PCR analysis. We also compared the allele frequency of the rs9356058 polymorphism from the population of Mljet with allele frequencies derived from the existing genome wide association scans in two additional island populations, Vis (924 subjects) and Korcula (909 subjects). RESULTS: We found a significant increase in the frequency of rs9356058 allele C in the population of Mljet when compared to both control groups. We also observed a significant increase in the frequency of rs1040079 allele A in the population of Mljet when compared with the population of Rab, however this increase was not significant when compared with the population of Split. Allele frequencies of both examined polymorphisms did not differ between the two control populations. Protective haplotype rs9356058-rs1040079 CA was also more frequent in the population of Mljet compared with the Rab and Split populations. In addition, an increase of frequency of rs9356058 allele C was also observed in the population of Mljet when compared with the frequency in the Korcula population. CONCLUSION: The results of our study show the association of polymorphisms rs9356058 and rs1040079 in gene PARK2/PACRG with leprosy. The results of our study indicate that exposure to leprosy and mortality in the population caused by leprosy on Mljet resulted in the selection of rs9356058 "protective" C allele in the PARK2 gene, while this was not observed in the two control groups. This is the first study to assess the genetic susceptibility to leprosy in a European population.

Rainforest aerosols as biogenic nuclei of clouds and precipitation in the Amazon.

The Amazon is one of the few continental regions where atmospheric aerosol particles and their effects on climate are not dominated by anthropogenic sources. During the wet season, the ambient conditions approach those of the pristine pre-industrial era. We show that the fine submicrometer particles accounting for most cloud condensation nuclei are predominantly composed of secondary organic material formed by oxidation of gaseous biogenic precursors. Supermicrometer particles, which are relevant as ice nuclei, consist mostly of primary biological material directly released from rainforest biota. The Amazon Basin appears to be a biogeochemical reactor, in which the biosphere and atmospheric photochemistry produce nuclei for clouds and precipitation sustaining the hydrological cycle. The prevailing regime of aerosol-cloud interactions in this natural environment is distinctly different from polluted regions.