Digital Commons Network^™

Toxoplasma Gondii, Suicidal Behavior, And Intermediate Phenotypes For Suicidal Behavior, Teodor T. Postolache, Abhishek Wadhawan, Dan Rujescu, Andrew J. Hoisington, Aline Dagdag, Enrique Baca-Garcia, Christopher A. Lowry, Olaoluwa O. Okusaga, Lisa A. Brenner

Faculty Publications

Within the general literature on infections and suicidal behavior, studies on Toxoplasma gondii ( T. gondii ) occupy a central position. This is related to the parasite's neurotropism, high prevalence of chronic infection, as well as specific and non-specific behavioral alterations in rodents that lead to increased risk taking, which are recapitulated in humans by T. gondii's associations with suicidal behavior, as well as trait impulsivity and aggression, mental illness and traffic accidents. This paper is a detailed review of the associations between T. gondii serology and suicidal behavior, a field of study that started 15 years ago with our …

The Status Of Spotted Seatrout (Cynoscion Nebulosus) As A Technologically Feasible Species For U.S. Marine Aquaculture, Reginald Blaylock, Eric Saillant, Angelos Apeitos, David Abrego, Paul Cason, Robert Vega

Faculty Publications

Culture models and facilities for large-scale, commercial production of popular Gulf of Mexico species are unavailable. The spotted seatrout (Cynoscion nebulosus) is one of the most popular recreational fishes in the Gulf of Mexico. Seatrout culture techniques were adapted from red drum (Sciaenops ocellatus) protocols developed in the 1970s. Broodstock husbandry, spawning, and extensive pond rearing techniques using fertilized and bloomed brackish ponds were well-established by the 1980s. By 2018, approximately 80 million 25–30-day old seatrout had been produced, mainly for stock enhancement. Cannibalism and poor nutrition hindered intensive tank culture. Between 2005 and 2015, an …

Inferring Single-Cell 3d Chromosomal Structures Based On The Lennard-Jones Potential, Mengsheng Zha, Nan Wang, Chaoyang Zhang, Zheng Wang

Faculty Publications

Reconstructing three‐dimensional (3D) chromosomal structures based on single‐cell Hi‐C data is a challenging scientific problem due to the extreme sparseness of the single‐cell Hi‐C data. In this research, we used the Lennard‐Jones potential to reconstruct both 500 kb and high‐resolution 50 kb chromosomal structures based on single‐cell Hi‐C data. A chromosome was represented by a string of 500 kb or 50 kb DNA beads and put into a 3D cubic lattice for simulations. A 2D Gaussian function was used to impute the sparse single‐cell Hi‐C contact matrices. We designed a novel loss function based on the Lennard‐Jones potential, in which …

The Covid-19 Pandemic And International Law, Preston J. Lim, Oona A. Hathaway, Alasdair Phillips-Robins, Mark Stevens

Faculty Publications

How does the COVID-19 pandemic affect States’ obligations under international law? This is a question of not just academic interest but real importance for people’s lives. After all, whether States abide by international law—and whether international law is fit for purpose—is vitally important for everyone from refugees exposed to the virus in unsanitary detention centers to national leaders fighting disinformation campaigns and safeguarding vaccine supply chains. International law has been central to the world’s response to the pandemic from the start—even if the participants did not always realize it. International law, after all, required States to take certain actions to …

Effects Of Timber Harvest On Epigeous Fungal Fruiting Patterns And Community Structure In A Northern Hardwood Ecosystem, Benjamin W. Borgmann-Winter, Ryan B. Stephens, Anthony W. D'Amato, Serita D. Frey, Rebecca J. Rowe

Faculty Publications

Epigeous fungal fruiting has important impacts on fungal reproduction and ecosystem function. Forest disturbances, such as timber harvest, impact moisture, host availability, and substrate availability, which in turn may drive changes in fungal fruiting patterns and community structure. We surveyed mushrooms in 0.4 ha patch cuts (18 months post-harvest) and adjacent intact hardwood forest in northern New Hampshire, USA, to document the effects of timber harvest on summer fruiting richness, biomass, diversity, and community structure of ectomycorrhizal, parasitic, and saprobic mushroom taxa. Fungal fruiting richness, diversity, and community heterogeneity were greater in intact forests than patch cuts. Among functional groups, …

Disentangling The Effects Of Vapor Pressure Deficit And Soil Water Availability On Canopy Conductance In A Seasonal Tropical Forest During The 2015 El Niño Drought, Yilin Fang, L. Ruby Leung, Brett T. Wolfe, Matteo Detto, Ryan G. Knox, Nate G. Mcdowell, Charlotte Grossiord, Chonggang Xu, Bradley O. Christoffersen, Pierre Gentine, Charles D. Koven, Jeffrey Q. Chambers

Faculty Publications

Water deficit in the atmosphere and soil are two key interactive factors that constrain transpiration and vegetation productivity. It is not clear which of these two factors is more important for the water and carbon flux response to drought stress in ecosystems. In this study, field data and numerical modeling were used to isolate their impact on evapotranspiration (ET) and gross primary productivity (GPP) at a tropical forest site in Barro Colorado Island (BCI), Panama, focusing on their response to the drought induced by the El Niño event of 2015–2016. Numerical simulations were performed using a plant hydrodynamic scheme (HYDRO) …

Genomic Gc Content Drifts Downward In Most Bacterial Genomes, Ely Bert

Faculty Publications

In every kingdom of life, GC->AT transitions occur more frequently than any other type of mutation due to the spontaneous deamination of cytidine. In eukaryotic genomes, this slow loss of GC base pairs is counteracted by biased gene conversion which increases genomic GC content as part of the recombination process. However, this type of biased gene conversion has not been observed in bacterial genomes, so we hypothesized that GC->AT transitions cause a reduction of genomic GC content in prokaryotic genomes on an evolutionary time scale. To test this hypothesis, we used a phylogenetic approach to analyze triplets of …

Genomic Gc Content Drifts Downward In Most Bacterial Genomes, Bert Ely

Faculty Publications

In every kingdom of life, GC->AT transitions occur more frequently than any other type of mutation due to the spontaneous deamination of cytidine. In eukaryotic genomes, this slow loss of GC base pairs is counteracted by biased gene conversion which increases genomic GC content as part of the recombination process. However, this type of biased gene conversion has not been observed in bacterial genomes, so we hypothesized that GC->AT transitions cause a reduction of genomic GC content in prokaryotic genomes on an evolutionary time scale. To test this hypothesis, we used a phylogenetic approach to analyze triplets of …

Latewood Ring Width Reveals Ce 1734 Felling Dates For Walker House Timbers In Tupelo, Mississippi, Usa, Thomas W. Patterson, Grant L. Harley, David H. Holt, Raymond T. Doherty, Daniel J. King, Karen J. Heeter, Ashley L. Chavez, Alyssa C. Crowell, Ian M. Stewart

Faculty Publications

Dendroarchaeology is under-represented in the Gulf Coastal Plain region of the United States (US), and at present, only three published studies have precision dated a collection of 18th–19th-century structures. In this study, we examined the tree-ring data from pine, poplar, and oak timbers used in the Walker House in Tupelo, Mississippi. The Walker House was constructed ca. the mid-1800s with timbers that appeared to be recycled from previous structures. In total, we examined 30 samples (16 pines, 8 oaks, and 6 poplars) from the attic and crawlspace. We cross-dated latewood ring growth from the attic pine samples to the period …

Increased Antimicrobial And Multidrug Resistance Downstream Of Wastewater Treatment Plants In An Urban Watershed, Maitreyee Mukherjee, Edward Laird, Terry J. Gentry, John P. Brooks, Raghupathy Karthikeyan

Faculty Publications

Development and spread of antimicrobial resistance (AMR) and multidrug resistance (MDR) through propagation of antibiotic resistance genes (ARG) in various environments is a global emerging public health concern. The role of wastewater treatment plants (WWTPs) as hot spots for the dissemination of AMR and MDR has been widely pointed out by the scientific community. In this study, we collected surface water samples from sites upstream and downstream of two WWTP discharge points in an urban watershed in the Bryan-College Station (BCS), Texas area, over a period of nine months. E. coli isolates were tested for resistance to ampicillin, tetracycline, sulfamethoxazole, …

Ecosytem Services: Delivering Decision-Making For Salt Marshes, Philine S. E. Zu Ermgassen, Ronald Baker, Michael W. Beck, Kate Dodds, Sophus O. S. E. Zu Ermgassen, Debbrota Mallick, Matthew D. Taylor, R. Eugene Turner

Faculty Publications

No abstract provided.

Australian Smokers' Sensory Experiences And Beliefs Associated With Menthol And Non-Menthol Cigarettes, Serafino Mancuso, Emily Brennan, Kimberley Dunstone, Amanda Vittiglia, Sarah Durkin, Jim F. Thrasher Ph.D., Ma, Ms, Janet Hoek, Melanie Wakefield

Faculty Publications

Many current smokers incorrectly believe that menthol cigarettes are less harmful, likely due to the biological and sensory effects of menthol, which can lead smokers to have favourable sensory experiences. In this study, we measured the extent to which Australian smokers associate certain sensory experiences with smoking menthol and non-menthol cigarettes, and their beliefs about how damaging and enjoyable they find cigarettes with each of these sensory experiences. A sample of 999 Australian 18–69-year-old weekly smokers was recruited from a non-probability online panel; this study focuses on the 245 respondents who currently smoked menthol cigarettes at least once per week. …

Host–Guest Interactions In A Metal–Organic Framework Isoreticular Series For Molecular Photocatalytic Co2 Reduction, Philip M. Stanley, Johanna Haimerl, Christopher Thomas, Alexander Urstoeger, Michael Schuster Prof. Dr., Natalia B. Shustova Prof. Dr., Angela Casini Prof. Dr., Bernhard Rieger Prof. Dr., Julien Warnan Dr., Roland A. Fischer Prof. Dr.

Faculty Publications

A strategy to improve homogeneous molecular catalyst stability, efficiency, and selectivity is the immobilization on supporting surfaces or within host matrices. Herein, we examine the co-immobilization of a CO2 reduction catalyst [ReBr(CO)3(4,4′-dcbpy)] and a photosensitizer [Ru(bpy)2(5,5′-dcbpy)]Cl2 using the isoreticular series of metal–organic frameworks (MOFs) UiO-66, -67, and -68. Specific host pore size choice enables distinct catalyst and photosensitizer spatial location—either at the outer MOF particle surface or inside the MOF cavities—affecting catalyst stability, electronic communication between reaction center and photosensitizer, and consequently the apparent catalytic rates. These results allow for a rational understanding of an optimized supramolecular layout of catalyst, …

Variability Of Usa East Coast Surface Total Alkalinity Distributions Revealed By Automated Instrument Measurements, Christopher W. Hunt, Joseph Salisbury, Douglas Vandemark, Steffen Aßmann, Christopher Melrose, Rik Wanninkhof, Kumiko Azetsu-Scott

Faculty Publications

Seawater total alkalinity (TA) is one important determinant used to monitor the ocean carbon cycle, whose spatial distributions have previously been characterized along the United States East Coast via discrete bottle samples. Using these data, several regional models for TA retrievals based on practical salinity (S) have been developed. Broad-scale seasonal or interannual variations, however, are not well resolved in these models and existing data are highly seasonally biased. This study reports findings from the first long duration deployment of a new, commercially available TA titrator aboard a research vessel and the continuous underway surface TA measurements produced. The …

Brief Overview Of Approaches And Challenges In New Antibiotic Development: A Focus On Drug Repurposing, Natalie K. Boyd, Chengwen Teng Pharm.D., Ph.D., Christopher R. Frei

Faculty Publications

Drug repurposing, or identifying new uses for existing drugs, has emerged as an alternative to traditional drug discovery processes involving de novo synthesis. Drugs that are currently approved or under development for non-antibiotic indications may possess antibiotic properties, and therefore may have repurposing potential, either alone or in combination with an antibiotic. They might also serve as “antibiotic adjuvants” to enhance the activity of certain antibiotics.

Solvent Effect To The Uniformity Of Surfactant-Free Salmon-Dna Thin Films, Jake Richter, Moses Nnaji, Heungman Park

Faculty Publications

Fabrication of surfactant-modified DNA thin films with high uniformity, specifically DNA–CTMA, has been well considered via drop-casting and spin-coating techniques. However, the fabrication of thin films with pure DNA has not been sufficiently studied. We characterize the uniformity of thin films from aqueous salmon DNA solutions mixed with ethanol, methanol, isopropanol, and acetone. Measurements of thickness and macroscopic uniformity are made via a focused-beam ellipsometer. We discuss important parameters for optimum uniformity and note what the effects of solvent modifications are. We find that methanol- and ethanol-added solutions provide optimal fabrication methods, which more consistently produce high degrees of uniformity …

Ocean-Atmosphere Interactions During Hurricanes Marco And Laura (2020), Emily N. Eley, Bulusu Subrahmanyam, Corinne B. Trott

Faculty Publications

During August of the 2020 Atlantic Hurricane Season, the Gulf of Mexico (GoM) was affected by two subsequent storms, Hurricanes Marco and Laura. Hurricane Marco entered the GoM first (22 August) and was briefly promoted to a Category 1 storm. Hurricane Laura followed Marco closely (25 August) and attained Category 4 status after a period of rapid intensification. Typically, hurricanes do not form this close together; this study aims to explain the existence of both hurricanes through the analysis of air-sea fluxes, local thermodynamics, and upper-level circulation. The GoM and its quality of warm, high ocean heat content waters proved …

De Novo Transcriptome Assembly Of The Green Alga Ankistrodesmus Falcatus, Rachel A. Schomaker, Jeffry L. Dudycha

Faculty Publications

Ankistrodesmus falcatus is a globally distributed freshwater chlorophyte that is a candidate for biofuel production, is used to study the effects of toxins on aquatic communities, and is used as food in zooplankton research. Each of these research fields is transitioning to genomic tools. We created a reference transcriptome for of A. falcatus using NextGen sequencing and de novo assembly methods including Trinity, Velvet-Oases, and EvidentialGene. The assembled transcriptome has a total of 17,997 contigs, an N50 value of 2,462, and a GC content of 64.8%. BUSCO analysis recovered 83.3% of total chlorophyte BUSCOs and 82.5% of the eukaryotic …

Cognitive Empathy And Longitudinal Changes In Temporo-Parietal Junction Thickness In Schizophrenia, Derin J. Cobia, Tatiana Karpouzian-Rogers, Julie Petersen, Lei Wang, Vijay A. Mittal, John G. Csernansky, Matthew J. Smith

Faculty Publications

Objective: Deficits in cognitive empathy are well-documented in individuals with schizophrenia and are related to reduced community functioning. The temporoparietal junction (TPJ) is closely linked to cognitive empathy. We compared the relationship between baseline cognitive empathy and changes in TPJ thickness over 24 months between individuals with schizophrenia and healthy controls. Methods: Individuals with schizophrenia (n = 29) and healthy controls (n = 26) completed a cognitive empathy task and underwent structural neuroimaging at baseline and approximately 24 months later. Symmetrized percent change scores were calculated for right and left TPJ, as well as whole-brain volume, and compared between groups. …

Prion-Like C-Terminal Domain Of Tdp-43 And Α-Synuclein Interact Synergistically To Generate Neurotoxic Hybrid Fibrils, Shailendra Dhakal, Courtney E. Wyant, Hannah E. George, Sarah E. Morgan, Vijayaraghavan Rangachari

Faculty Publications

Aberrant aggregation and amyloid formation of tar DNA binding protein (TDP-43) and α-synuclein (αS) underlie frontotemporal dementia (FTD) and Parkinson’s disease (PD), respectively. Amyloid inclusions of TDP-43 and αS are also commonly co-observed in amyotrophic lateral sclerosis (ALS), dementia with Lewy bodies (DLB) and Alzheimer disease (AD). Emerging evidence from cellular and animal models show colocalization of the TDP-43 and αS aggregates, raising the possibility of direct interactions and co-aggregation between the two proteins. In this report, we set out to answer this question by investigating the interactions between αS and prion-like pathogenic C-terminal domain of TDP-43 (TDP-43 PrLD). PrLD …

Beyond Structural Motifs: The Frontier Of Actinide-Containing Metal–Organic Frameworks, Corey R. Martin, Gabrielle A. Leith, Natalia B. Shustova Prof. Dr.

Faculty Publications

In this perspective, we feature recent advances in the field of actinide-containing metal–organic frameworks (An-MOFs) with a main focus on their electronic, catalytic, photophysical, and sorption properties. This discussion deviates from a strictly crystallographic analysis of An-MOFs, reported in several reviews, or synthesis of novel structural motifs, and instead delves into the remarkable potential of An-MOFs for evolving the nuclear waste administration sector. Currently, the An-MOF field is dominated by thorium- and uranium-containing structures, with only a few reports on transuranic frameworks. However, some of the reported properties in the field of An-MOFs foreshadow potential implementation of these materials and …

Beyond Structural Motifs: The Frontier Of Actinide-Containing Metal–Organic Frameworks, Corey R. Martin, Gabrielle A. Leith, Natalia B. Shustova

Faculty Publications

In this perspective, we feature recent advances in the field of actinide-containing metal-organic frameworks (An-MOFs) with a main focus on their electronic, catalytic, photophysical, and sorption properties. This discussion deviates from a strictly crystallographic analysis of An-MOFs, reported in several reviews, or synthesis of novel structural motifs, and instead delves into the remarkable potential of An-MOFs for evolving the nuclear waste administration sector. Currently, the An-MOF field is dominated by thorium- and uranium-containing structures, with only a few reports on transuranic frameworks. However, some of the reported properties in the field of An-MOFs foreshadow potential implementation of these materials and …

Nanoantibiotics: Functions And Properties At The Nanoscale To Combat Antibiotic Resistance, M. Mustafa Mamun, Adeola Julian Sorinolu, Mariya Munir, Eric P. Vejerano

Faculty Publications

One primary mechanism for bacteria developing resistance is frequent exposure to antibiotics. Nanoantibiotics (nAbts) is one of the strategies being explored to counteract the surge of antibiotic resistant bacteria. nAbts are antibiotic molecules encapsulated with engineered nanoparticles (NPs) or artificially synthesized pure antibiotics with a size range of ≤100 nm in at least one dimension. NPs may restore drug efficacy because of their nanoscale functionalities. As carriers and delivery agents, nAbts can reach target sites inside a bacterium by crossing the cell membrane, interfering with cellular components, and damaging metabolic machinery. Nanoscale systems deliver antibiotics at enormous particle number concentrations. …

Environmentally Persistent Free Radicals As Sources Of Pops, Eric P. Vejerano, Jeonghyeon Ahn, Junaid Latif, Mustafa Mamun

Faculty Publications

Environmentally persistent free radicals (EPFRs) are a new class organic pollutant sharing some of the attributes of persistent organic pollutants (POPs). This opinion/short review aims to describe the properties of EPFRs that merit their recognition as an additional and potentially significant source of POPs. EPFRs are ubiquitous in diverse environments because of multiple factors: (1) organic precursors from anthropogenic, biogenic, and other natural emission sources are abundant; multiple mechanisms in PM and soils form (2) EPFRs; and (3) EPFRs are stable and persist for a long time, thereby, accumulate in the environment and potentially transported long range. The hazards of …

Cycling Phosphorus And Nitrogen Through Cropping Systems In An Intensive Dairy Production Region, April B. Leytem, Paula Williams, Shan Zuidema, Audrey Martinez, Yen Leng Chong, Alyssa Vincent, Aaron Vincent, Daniel Cronan, Andrew Kliskey, J. D. Wulfhorst, Lilian Alessa, David Bjorneberg

Faculty Publications

As pressure on the dairy industry to reduce its environmental impact increases, efficient recycling of manure nutrients through local cropping systems becomes crucial. The aim of this study was to calculate annual nitrogen (N) and phosphorus (P) budgets in six counties located in the Magic Valley, Idaho and estimate what distance manure would need to be transported to be in balance with crop nutrient demand given current dairy cattle populations and cropping systems. Our analysis suggests that crop N needs will not be met solely by manure, and synthetic fertilizer will need to be applied. However, to balance P with …

Covid-19: A Syndemic Requiring An Integrated Approach For Marginalized Populations, Rosemary M. Caron, Amanda Rodrigues Amorim Adegboye

Faculty Publications

The novel coronavirus, SARS-CoV-2, responsible for the COVID-19 pandemic, has challenged healthcare systems globally. The health inequities experienced by immigrants, refugees, and racial/ethnic minorities have been aggravated during the COVID-19 pandemic. The socioeconomic, political, and demographic profile of these vulnerable populations places them at increased risk of contracting COVID-19 and experiencing significant morbidity and mortality. Thus, the burden of the COVID-19 pandemic is disproportionally higher among these at-risk groups. The purpose of this perspective is to: (1) highlight the interactions among the social determinants of health (SDoH) and their bi-directional relationship with the COVID-19 pandemic which results in the current …

Fungal Community Response To Long-Term Soil Warming With Potential Implications For Soil Carbon Dynamics, Gregory J. Pec, Linda T. A. Van Diepen, Melissa A. Knorr, A. Stuart Grandy, Jerry M. Melillo, Kristen M. Deangelis, Jeffrey L. Blanchard, Serita D. Frey

Faculty Publications

The direction and magnitude of climate warming effects on ecosystem processes such as carbon cycling remain uncertain. Soil fungi are central to these processes due to their roles as decomposers of soil organic matter, as mycorrhizal symbionts, and as determinants of plant diversity. Yet despite their importance to ecosystem functioning, we lack a clear understanding of the long-term response of soil fungal communities to warming. Toward this goal, we characterized soil fungal communities in two replicated soil warming experiments at the Harvard Forest (Petersham, Massachusetts, USA) which had experienced 5°C above ambient soil temperatures for 5 and 20 yr at …

A Holistic Framework Integrating Plant-Microbe-Mineral Regulation Of Soil Bioavailable Nitrogen, Amanda B. Daly, Andrea Jilling, Timothy M. Bowles, Robert W. Buchowski, Serita D. Frey, Cynthia M. Kallenbach, Marco Keiluweit, Maria Mooshammer, Joshua P. Schimel, A. Stuart Grandy

Faculty Publications

Soil organic nitrogen (N) is a critical resource for plants and microbes, but the processes that govern its cycle are not well-described. To promote a holistic understanding of soil N dynamics, we need an integrated model that links soil organic matter (SOM) cycling to bioavailable N in both unmanaged and managed landscapes, including agroecosystems. We present a framework that unifies recent conceptual advances in our understanding of three critical steps in bioavailable N cycling: organic N (ON) depolymerization and solubilization; bioavailable N sorption and desorption on mineral surfaces; and microbial ON turnover including assimilation, mineralization, and the recycling of microbial …

Sodah: The Soils Data Harmonization Database, An Open-Source Synthesis Of Soil Data From Research Networks, Version 1.0, William R. Wieder, Derek Pierson, Stevan Earl, Kate Lajtha, Sara G. Baer, Ford Ballantyne, Asmeret Asefaw Berhe, Sharon A. Billings, Laurel M. Brigham, Stephany S. Chacon, Jennifer Fraterrigo, Serita D. Frey, Katerina Georgiou, Marie-Anne De Graaff, A. Stuart Grandy, Melannie D. Hartman, Sarah E. Hobbie, Chris Johnson, Jason Kaye, Emily Kyker-Snowman, Marcy E. Litvak, Michelle C. Mack, Avni Malhotra, Jessica A. M. Moore, Knute Nadelhoffer, Craig Rasmussen, Whendee L. Silver, Benjamin N. Sulman, Xanthe Walker, Samantha Weintraub

Faculty Publications

Data collected from research networks present opportunities to test theories and develop models about factors responsible for the long-term persistence and vulnerability of soil organic matter (SOM). Synthesizing datasets collected by different research networks presents opportunities to expand the ecological gradients and scientific breadth of information available for inquiry. Synthesizing these data is challenging, especially considering the legacy of soil data that have already been collected and an expansion of new network science initiatives. To facilitate this effort, here we present the SOils DAta Harmonization database (SoDaH; https://lter.github.io/som-website, last access: 22 December 2020), a flexible database designed to harmonize diverse …

The Impact Of Aquaculture On The Genetics And Distribution Of The Onuphid Annelid Diopatra Biscayensis, Matthew P. Galaska, David S. Wethey, Andrés Arias, Stanislas F. Dubois, Kenneth M. Halanych, Sarah A. Woodin

Faculty Publications

Aim

Evolutionary history of natural populations can be confounded by human intervention such as the case of decorator worm species Diopatra (Onuphidae), which have a history of being transported through anthropogenic activities. Because they build tubes and act as ecosystem engineers, they can have a large impact on the overall ecosystem in which they occur. One conspicuous member, Diopatra biscayensis, which was only described in 2012, has a fragmented distribution that includes the Bay of Biscay and the Normanno-Breton Gulf in the English Channel. This study explores the origin of these worms in the Normanno-Breton region, which has been …