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Background: The origin of divergent SARS-CoV-2 spike sequences found in wastewater, but not in clinical surveillance, remains unclear. These cryptic wastewater sequences have harbored many of the same mutations that later emerged in Omicron lineages. We first detected a cryptic lineage in municipal wastewater in Wisconsin in January 2022. Named the Wisconsin Lineage, we sought to determine the geographic origin of this virus and characterize its persistence and evolution over time. Methods: We systematically sampled maintenance holes to trace the origin of the Wisconsin Lineage. We sequenced spike RBD domains, and where possible, whole viral genomes, to characterize the evolution of this lineage over the 13 consecutive months that it was detectable. Findings: The persistence of the Wisconsin Lineage signal allowed us to trace it from a central wastewater plant to a single facility, with a high concentration of viral RNA. The viral sequences contained a combination of fixed nucleotide substitutions characteristic of Pango lineage B.1.234, which circulated in Wisconsin at low levels from October 2020 to February 2021, while mutations in the spike gene resembled those subsequently found in Omicron variants. Interpretation: We propose that prolonged detection of the Wisconsin Lineage in wastewater represents persistent shedding of SARS-CoV-2 from an infected individual, with ongoing within-host viral evolution leading to an ancestral B.1.234 virus accumulating Omicron-like mutations. Funding: The Rockefeller Foundation, Wisconsin Department of Health Services, Centers for Disease Control and Prevention (CDC), National Institute on Drug Abuse (NIDA), and the Center for Research on Influenza Pathogenesis and Transmission.

Urbanization exerts many pressures on species, yet little is known about how these pressures impact species interactions. Studies of urban plant-pollinator systems provide mounting evidence that urbanization impairs pollinator movement in fragmented urban landscapes, yet the consequences for pollinator-mediated plant reproduction remains unclear. In non-urban areas, habitat corridors can facilitate the movement of organisms including pollinators, but whether these corridors facilitate plant-pollinator interactions in urban areas remains understudied. To examine how urban environments and green corridors influence plant-pollinator interactions, we measured reproductive success in the native plant common milkweed (Asclepias syriaca), and the community structure of its pollinators, for two years along two urban-rural transects in the Greater Toronto Area, Canada, one of which followed a green corridor. We found that urbanization decreased male fitness (i.e., pollen removal), increased fruit set (i.e., mean no. of follicles per inflorescence), and inconsistently affected female fitness (i.e., no. of follicles) in A. syriaca. Urbanization simultaneously decreased pollinator abundance but increased pollinator richness. Proximity to a green corridor inconsistently affected male fitness but increased reproductive effort (i.e., no. of inflorescences) in A. syriaca, while pollinator diversity and richness was lower in corridors. Notably, there were no consistent relationships between pollinator community structure and reproductive success in A. syriaca in both the presence, and absence, of a green corridor. These results demonstrate the complexity with which urbanization, green corridors, and pollinator communities can shape the reproductive investment and fitness of native plant populations.

Legislative sovereignty is often discussed with one eye on the past and one eye on the procedural functions of law-making in the present. This limits the scope for a conceptual understanding of leg...

The first 2 years of the COVID-19 pandemic were mainly characterized by recurrent mutations of SARS-CoV-2 Spike protein at residues K417, L452, E484, N501 and P681 emerging independently across different variants of concern (Alpha, Beta, Gamma, and Delta). Such homoplasy is a marker of convergent evolution. Since Spring 2022 and the third year of the pandemic, with the advent of Omicron and its sublineages, convergent evolution has led to the observation of different lineages acquiring an additional group of mutations at different amino acid residues, namely R346, K444, N450, N460, F486, F490, Q493, and S494. Mutations at these residues have become increasingly prevalent during Summer and Autumn 2022, with combinations showing increased fitness. The most likely reason for this convergence is the selective pressure exerted by previous infection- or vaccine-elicited immunity. Such accelerated evolution has caused failure of all anti-Spike monoclonal antibodies, including bebtelovimab and cilgavimab. While we are learning how fast coronaviruses can mutate and recombine, we should reconsider opportunities for economically sustainable escape-proof combination therapies, and refocus antibody-mediated therapeutic efforts on polyclonal preparations that are less likely to allow for viral immune escape.

Objectives Little is known about the external validity of the Data-collection on Adverse Effects of Anti-HIV Drugs (D:A:D) model for predicting cardiovascular disease (CVD) risk among people living with HIV (PLWH). We aimed to evaluate the performance of the updated D:A:D model for 5-year CVD risk in a diverse group of PLWH engaged in HIV care. Methods We used data from an institutional HIV registry, which includes PLWH engaged in care at a safety-net HIV clinic. Eligible individuals had a baseline clinical encounter between 1 January 2013 and 31 December 2014, with follow-up through to 31 December 2019. We estimated 5-year predicted risks of CVD as a function of the prognostic index and baseline survival of the D:A:D model, which were used to assess model discrimination (C-index), calibration and net benefit. Results Our evaluable population comprised 1029 PLWH, of whom 30% were female, 50% were non-Hispanic black, and median age was 45 years. The C-index was 0.70 [95% confidence limits (CL): 0.64-0.75]. The predicted 5-year CVD risk was 3.0% and the observed 5-year risk was 8.9% (expected/observed ratio = 0.33, 95% CL: 0.26-0.54). The model had a greater net benefit than treating all or treating none at a risk threshold of 10%. Conclusions The D:A:D model was miscalibrated for CVD risk among PLWH engaged in HIV care at an urban safety-net HIV clinic, which may be related to differences in case-mix and baseline CVD risk. Nevertheless, the HIV D:A:D model may be useful for decisions about CVD intervention for high-risk patients.

This chapter takes a “Theory of Change” approach in examining the opportunities and challenges of blockchain in sustainable development and social impact. It addresses the manner in which both the discourse and the use cases of blockchain and initiatives related to sustainable development and social impact have evolved, from the discourse of the Pillar of the 4th Industrial Revolution and the effects of the silicon valley “style” ventures, the overestimation of “market opportunity”, the conflation and confounding of different of types of blockchain initiatives and offerings combined with key assumptions about readiness and capacity not only of the technological elements—as blockchain as a “ready service”—but also within the social impact and sustainable development landscape. It then examines key use cases and loops back to address the key needs and barriers for scaling up (As opposed to the issues surrounding technical transactional scaling issues related to the need for blockchain platforms to process hundreds of thousands of transactions per second as well as the increased need for miners, developers, businesses and other stakeholders) of blockchain solutions and impact in sustainable development.

David Brown sat at his desk in the stock room of his toy store, Huckle Buckle Beanstalk, looking at the three résumés in front of him. He needed a new store manager, and these three résumés were his top choices. Each offered a different mix of skills and experience, and he hoped he would have the unusual luxury of having one rise to the top in the afternoon interviews. He had fired his previous store manager just over a month earlier for theft, and he knew that the business would suffer if he had to continue managing the store himself.

CD39 (ENTPD1) is a key enzyme responsible for the degradation of extracellular adenosine triphosphate (ATP) and is upregulated in the tumor microenvironment (TME). High levels of extracellular ATP, often generated in the TME as a result of tissue damage and immunogenic cell death, can initiate proinflammatory responses that are blocked by the enzymatic activity of CD39. In addition, extracellular adenosine accumulates in cancerous tissues through the degradation of ATP by CD39 and other ectonucleotidases (eg, CD73), and constitutes an important mechanism of tumor immune escape, induction of angiogenesis, and metastasis. Thus, inhibition of CD39 can convert a suppressive TME to a proinflammatory environment. SRF617 is an investigational fully human anti-CD39 antibody that binds to human CD39 with nanomolar affinity and potently inhibits its enzymatic function. Results of the current studies show that CD39 is predominantly expressed in tumor stroma and on tumor-infiltrating immune cells in samples from patients with cancer. Similar expression patterns are observed in various murine tumor models. In vivo, SRF617 has significant single-agent anti-tumor activity in a variety of cell line-derived xenograft models that express CD39. Cancer therapies, such as immunogenic cell death agents, can increase inflammation and ATP levels in the TME; combination studies of SRF617 with these agents showed improved preclinical efficacy and led to significant improvement in survival. In addition, a mouse-specific anti-CD39 surrogate antibody demonstrated potent binding and enzymatic inhibition of murine CD39 in vitro and significantly decreased tumor growth in a syngeneic murine tumor model. Pharmacodynamic studies demonstrated mechanistic changes in the tumor-infiltrating leukocytes and plasma chemokine levels. Combination treatment of a murine anti-CD39 surrogate and an anti-mouse PD-1 displayed improved activity and an increase in overall survival in the CT-26 mouse model. In addition, administration of SRF617 in combination with other immunotherapies also demonstrated a similar improvement in activity and survival. In summary, these studies demonstrate that SRF617 is a potent inhibitor of CD39 enzymatic activity both in vitro and in vivo. Inhibition of CD39 potentiates the activity of chemotherapy and immunotherapy agents to improve tumor growth inhibition and survival in mice. These findings support future clinical studies of SRF617 as monotherapy and in combination with other therapeutic agents in treating patients with cancer. Citation Format: Sonia G. Das, Austin Dulak, Gege Tan, Marc Johnson, Tauqeer H. Zaidi, Michael C. Warren, Secil Koseoglu, Erik Devereaux, Marisa O. Peluso, Alison M. Paterson, Benjamin H. Lee, Vito J. Palombella, Pamela M. Holland, Andrew C. Lake. SRF617, a potent enzymatic inhibitor of CD39, demonstrates single-agent activity and cooperates with various cancer therapies in both solid tumor and hematologic malignancies [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 6639.

Canada and the European Union (EU) began negotiating a comprehensive economic and trade agreement (CETA) in May 2009. The authors survey both the process and the content of CETA from the perspective of the province of Quebec. For the first time, Canadian provinces and territories have fully participated in international trade negotiations that will impact them directly and engender important economic benefits. CETA will ensure that Canadian provinces and territories obtain real market access for their goods and services to the EU’s 500 million consumers. It will encourage and increase foreign investment as well as create further labour mobility between Canada and the EU. Quebec will benefit from a growing and deeper relationship with the EU through trade and cooperation, thereby fostering economic globalization and integration.

Purpose: Language sampling is an important technique that speech-language pathologists use to gather and analyze a child's language abilities. The current study examines the effects of student training on the quality of child language samples obtained by undergraduate speech- language pathology students. Method: A pretest-posttest, within-subjects design was used so that each student obtained two 50-utterance language samples from children with no diagnosed language dis- orders. In the first condition, students were given a brief explanation of the method of collecting a language sample. In the second condition, students were given a handout of more detailed sampling techniques and they role-played interacting with a child. Results: The language samples obtained by the students S

Pavement surface temperature is an essential requirement for the winter maintenance decision-making processes when determining suitable deicers and optimal application rates for transportation facilities such as parking lots. Currently, there are very few services/methods available to directly measure the pavement surface temperature. One method, the Road Weather Information System (RWIS), has less utility for parking lot and sidewalk maintenance purposes due to the operational constraints and high cost of using the RWIS service. This paper presents the results of a two-year field study focused on developing parametric and non-parametric models to forecast pavement surface temperature in parking lots based on a rigorous investigation of its correlation with common meteorological variables. The models were developed using available metrological data alongside measured pavement temperatures obtained through field observations. This study also evaluates the effects of different types of pavement (asphalt, interlocking bricks, and portland cement concrete) on pavement surface temperature. Evaluation results show that the developed models are able to predict pavement surface temperatures in different winter weather conditions with acceptable accuracy.

This multi-genre writing, multimedia, co-inquiry project produced successful end products. The middle school students gained experience in writing in a variety of genres, communicating through multiple media, and using education technology to do so. Likewise, the pre-service teachers developed their skills in these same areas. More importantly for our preservice teachers they gained teaching experience and had a chance to interact with students in the role of the teacher. This success would not have been achieved without the collaboration and cooperation of many individuals as well as access to the required technologies. The asynchronous discussion board was an important part of that success. This mode of telecommunications facilitated increased social and substantive interactions between the middle school students and the preservice teachers from the beginning to the end of the project. With careful consideration of the points and questions we present here, we hope that you will be able to adopt or adapt this mode of telecommunications to add value to your pre-service teachers’ teaching experiences and interactions with K-12 students.

Cross-linked amylose starch (CLA), hydroxypropylmethylcellulose (HPMC), and HPMC/CLA matrices were prepared by direct compression. HPMC was used to slow down the enzymatic degradation of CLA matrices. CLA was either granulated alone and mixed with HPMC or cogranulated with the latter. Compaction characteristics of the powder, hydration and mechanical properties of the resulting matrices, as well as the release profiles of three model drugs were investigated. The results showed that wet granulation of CLA in the presence of 10% HPMC improved significantly the flow properties of the powder without compromising its compactibility. Both CLA and HPMC deformed mainly by plastic flow (yield pressures are 75 and 124 MPa, respectively), but CLA exhibited a stronger elastic component (elastic recoveries are 18.4 and 11.5%, respectively). The values of yield pressure increased linearly with the concentration of HPMC. The addition of HPMC to CLA slightly decreased the resistance to consolidation but the crushing force of the final compacts was found to be proportional to the HPMC concentration. Mechanical studies on swollen matrices revealed that CLA formed a stronger gel than HPMC or CLA/HPMC mixture, and swelling and erosion of the tablets increased with HPMC content and incubation time. The in vitro release kinetics of three model drugs (pseudeoephedrine sulfate, sodium diclofenac, and prednisone) showed a clear effect of drug solubility and presence of alpha-amylase in the dissolution medium on the release rate. The addition of HPMC to CLA protected the tablets against alpha-amylase hydrolysis and reduced the release rate of prednisone and sodium diclofenac. The release of pseudoephedrine sulfate was fast and independent of HPMC and occurred mainly by diffusion.

Regulators of G protein signaling (RGS) constitute a family of proteins with a conserved RGS domain of approximately 120 amino acids that accelerate the intrinsic GTP hydrolysis of activated Galpha(i) and Galpha(q) subunits. The phosphorylation-dependent interaction of 14-3-3 proteins with a subset of RGS proteins inhibits their GTPase-accelerating activity in vitro. The inhibitory interaction between 14-3-3 and RGS7 requires phosphorylation of serine 434 of RGS7. We now show that phosphorylation of serine 434 is dynamically regulated by TNF-alpha. Cellular stimulation by TNF-alpha transiently decreased the phosphorylation of serine 434 of RGS7, abrogating the inhibitory interaction with 14-3-3. We examined the effect of 14-3-3 on RGS-mediated deactivation kinetics of G protein-coupled inwardly rectifying K(+) channels (GIRKs) in Xenopus oocytes. 14-3-3 inhibited the function of wild-type RGS7, but not that of either RSG7(P436R) or RGS4, two proteins that do not bind 14-3-3. Our findings are the first evidence that extracellular signals can modulate the activity of RGS proteins by regulating their interaction with 14-3-3.

The budding of enveloped retrovirus, such as HIV, involves oligomerization of the structural protein Gag at the plasma membrane and the incorporation of a patch of the host cell plasma membrane in the viral particle. Previous studies indicate that certain host proteins and viral envelope proteins are enriched in the membrane of virus particles. Since the viral membrane is derived from the host membrane, such specialized composition of viral membrane suggests viral assembly and budding involve differentiation of the host cell plasma-membrane at the assembly site. However, the mechanism of the membrane remodeling process during viral assembly is not understood. using a combination of quantitative superresolution microscopy (PALM and STORM) and diffraction-limited TIRF microscopy, we have investigated the spatial and temporal differentiation of the host cell plasma-membrane during budding of HIV. We find that in absence of Gag, the viral envelope protein is randomly distributed in the plasma membrane. However, oligomerization of Gag at the membrane leads to dramatic redistribution of the envelope proteins at the site of Gag assembly. We also see enrichment of GPI-anchored protein such as CD59 at the assembly site, while a model transmembrane protein EGFP-GT46 is actively excluded from the membrane patch. Gag is anchored to the plasma membrane via myristoylated lipid anchor and electrostatic interactions. Interestingly, we find that myristoylated inner leaflet anchored proteins are enriched while proteins with farnesyl and geranyl lipid anchors are depleted at the assembly site. Taken together, our results indicate that the oligomerization of HIV Gag at the plasma-membrane creates a specialized microenvironment, which leads to the differentiation of the local plasma membrane. This provides new insights about the temporal and spatial remodeling of plasma membrane during assembly and budding of retroviruses.

Background Human immunoficiency virus (HIV)–associated pericardial effusion is common. We present its clinical features, cause, and prognosis on the basis of a review of 40 cases at a single public hospital. Methods A retrospective study was conducted of 122 patients with pericardial effusion (of which 40 were HIV associated) admitted to Queens Hospital Center from January 1988 to April 1997. A review of the literature is also presented. Results Forty patients with HIV-associated pericardial effusion represent 33% of the 122 patients with pericardial effusion admitted during that period. The most common symptom of the 40 patients was dyspnea (75%). Echocardiogram detected small effusions in 18 (45%), moderate effusions in 10 (25%), and large effusions in 12 (30%). Sixteen (40%) patients had cardiac tamponade, in 15 of whom pericardiocentesis or pericardiostomy was performed. Causes of cardiac tamponade were Mycobacterium species in 3 (19%), Streptococcus pneumoniae in 1 (6%), Staphylococcus aureus in 1 (6%), Kaposi's sarcoma in 1 (6%), and unknown in 10 (63%). In comparison, causes of cardiac tamponade in 74 cases of acquired immunodeficiency syndrome in the literature were 45% idiopathic, 20% mycobacteria, 19% bacteria, 7% lymphoma, 5% Kaposi's sarcoma, 3% viruses, and 1% fungus. Thirteen of the 40 patients were lost to follow-up. Among the other 27, 11 (41%) were alive at 3 months and 5 (19%) at 1 year. Ten of the 27 patients had cardiac tamponade, of whom 5 (50%) were alive at 3 months and 3 (30%) at 1 year. Conclusions HIV-associated pericardial effusion is the most common type of pericardial effusion in our inner city hospital. Causes are diverse. The development of pericardial effusion predicts a poor prognosis in HIV infection. (Am Heart J 1999;137:516-21.)

It is often thought that affording rights is a progressive movement; rights are given to natural legal persons; the rights are normalised in societal expectations and they form part of a body of enforceable rights against the state. On 7 February 2018, Bermuda became the first state in modern history to withdraw the right of same-sex couples to marry, bucking the trend of progressively affording rights. In a recent judgment, the Bermudian Supreme Court has ruled that taking away the right of same-sex couples to marry is unconstitutional. This article will briefly consider the development of the right of same-sex couples to marry in Bermuda, the connection between Bermudian human rights law and the European Convention on Human Rights and ask whether rights afforded under a constitutional arrangement can be taken away.