Expanding chemistry through in vitro and in vivo biocatalysis.

Living systems contain a vast network of metabolic reactions, providing a wealth of enzymes and cells as potential biocatalysts for chemical processes. The properties of protein and cell biocatalysts-high selectivity, the ability to control reaction sequence and operation in environmentally benign conditions-offer approaches to produce molecules at high efficiency while lowering the cost and environmental impact of industrial chemistry. Furthermore, biocatalysis offers the opportunity to generate chemical structures and functions that may be inaccessible to chemical synthesis. Here we consider developments in enzymes, biosynthetic pathways and cellular engineering that enable their use in catalysis for new chemistry and beyond.

Biocatalytic control of site-selectivity and chain length-selectivity in radical amino acid halogenases.

Biocatalytic C-H activation has the potential to merge enzymatic and synthetic strategies for bond formation. FeII/αKG-dependent halogenases are particularly distinguished for their ability both to control selective C-H activation as well as to direct group transfer of a bound anion along a reaction axis separate from oxygen rebound, enabling the development of new transformations. In this context, we elucidate the basis for the selectivity of enzymes that perform selective halogenation to yield 4-Cl-lysine (BesD), 5-Cl-lysine (HalB), and 4-Cl-ornithine (HalD), allowing us to probe how site-selectivity and chain length selectivity are achieved. We now report the crystal structure of the HalB and HalD, revealing the key role of the substrate-binding lid in positioning the substrate for C4 vs C5 chlorination and recognition of lysine vs ornithine. Targeted engineering of the substrate-binding lid further demonstrates that these selectivities can be altered or switched, showcasing the potential to develop halogenases for biocatalytic applications.

Engineering nonphotosynthetic carbon fixation for production of bioplastics by methanogenic archaea.

The conversion of CO2 to value-added products allows both capture and recycling of greenhouse gas emissions. While plants and other photosynthetic organisms play a key role in closing the global carbon cycle, their dependence on light to drive carbon fixation can be limiting for industrial chemical synthesis. Methanogenic archaea provide an alternative platform as an autotrophic microbial species capable of non-photosynthetic CO2 fixation, providing a potential route to engineered microbial fermentation to synthesize chemicals from CO2 without the need for light irradiation. One major challenge in this goal is to connect upstream carbon-fixation pathways with downstream biosynthetic pathways, given the distinct differences in metabolism between archaea and typical heterotrophs. We engineered the model methanogen, Methanococcus maripaludis, to divert acetyl-coenzyme A toward biosynthesis of value-added chemicals, including the bioplastic polyhydroxybutyrate (PHB). A number of studies implicated limitations in the redox pool, with NAD(P)(H) pools in M. maripaludis measured to be <15% of that of Escherichia coli, likely since methanogenic archaea utilize F420 and ferredoxins instead. Multiple engineering strategies were used to precisely target and increase the cofactor pool, including heterologous expression of a synthetic nicotinamide salvage pathway as well as an NAD+-dependent formate dehydrogenase from Candida boidinii. Engineered strains of M. maripaludis with improved NADH pools produced up to 171 ± 4 mg/L PHB and 24.0 ± 1.9% of dry cell weight. The metabolic engineering strategies presented in this study broaden the utility of M. maripaludis for sustainable chemical synthesis using CO2 and may be transferable to related archaeal species.

Survival After Invasive or Conservative Management of Stable Coronary Disease.

BACKGROUND: The ISCHEMIA trial (International Study of Comparative Health Effectiveness With Medical and Invasive Approaches) compared an initial invasive versus an initial conservative management strategy for patients with chronic coronary disease and moderate or severe ischemia, with no major difference in most outcomes during a median of 3.2 years. Extended follow-up for mortality is ongoing. METHODS: ISCHEMIA participants were randomized to an initial invasive strategy added to guideline-directed medical therapy or a conservative strategy. Patients with moderate or severe ischemia, ejection fraction ≥35%, and no recent acute coronary syndromes were included. Those with an unacceptable level of angina were excluded. Extended follow-up for vital status is being conducted by sites or through central death index search. Data obtained through December 2021 are included in this interim report. We analyzed all-cause, cardiovascular, and noncardiovascular mortality by randomized strategy, using nonparametric cumulative incidence estimators, Cox regression models, and Bayesian methods. Undetermined deaths were classified as cardiovascular as prespecified in the trial protocol. RESULTS: Baseline characteristics for 5179 original ISCHEMIA trial participants included median age 65 years, 23% women, 16% Hispanic, 4% Black, 42% with diabetes, and median ejection fraction 0.60. A total of 557 deaths accrued during a median follow-up of 5.7 years, with 268 of these added in the extended follow-up phase. This included a total of 343 cardiovascular deaths, 192 noncardiovascular deaths, and 22 unclassified deaths. All-cause mortality was not different between randomized treatment groups (7-year rate, 12.7% in invasive strategy, 13.4% in conservative strategy; adjusted hazard ratio, 1.00 [95% CI, 0.85-1.18]). There was a lower 7-year rate cardiovascular mortality (6.4% versus 8.6%; adjusted hazard ratio, 0.78 [95% CI, 0.63-0.96]) with an initial invasive strategy but a higher 7-year rate of noncardiovascular mortality (5.6% versus 4.4%; adjusted hazard ratio, 1.44 [95% CI, 1.08-1.91]) compared with the conservative strategy. No heterogeneity of treatment effect was evident in prespecified subgroups, including multivessel coronary disease. CONCLUSIONS: There was no difference in all-cause mortality with an initial invasive strategy compared with an initial conservative strategy, but there was lower risk of cardiovascular mortality and higher risk of noncardiovascular mortality with an initial invasive strategy during a median follow-up of 5.7 years. REGISTRATION: URL: https://www. CLINICALTRIALS: gov; Unique identifier: NCT04894877.

Geospatial analysis of Plasmodium falciparum serological indicators: school versus community sampling in a low-transmission malaria setting.

BACKGROUND: Due to low numbers of active infections and persons presenting to health facilities for malaria treatment, case-based surveillance is inefficient for understanding the remaining disease burden in low malaria transmission settings. Serological data through the detection of IgG antibodies from previous malaria parasite exposure can fill this gap by providing a nuanced picture of where sustained transmission remains. Study enrollment at sites of gathering provides a potential approach to spatially estimate malaria exposure and could preclude the need for more intensive community-based sampling. METHODS: This study compared spatial estimates of malaria exposure from cross-sectional school- and community-based sampling in Haiti. A total of 52,405 blood samples were collected from 2012 to 2017. Multiplex bead assays (MBAs) tested IgG against P. falciparum liver stage antigen-1 (LSA-1), apical membrane antigen 1 (AMA1), and merozoite surface protein 1 (MSP1). Predictive geospatial models of seropositivity adjusted for environmental covariates, and results were compared using correlations by coordinate points and communes across Haiti. RESULTS: Consistent directional associations were observed between seroprevalence and environmental covariates for elevation (negative), air temperature (negative), and travel time to urban centers (positive). Spearman's rank correlation for predicted seroprevalence at coordinate points was lowest for LSA-1 (ρ = 0.10, 95% CI: 0.09-0.11), but improved for AMA1 (ρ = 0.36, 95% CI: 0.35-0.37) and MSP1 (ρ = 0.48, 95% CI: 0.47-0.49). CONCLUSIONS: In settings approaching P. falciparum elimination, case-based prevalence data does not provide a resolution of ongoing malaria transmission in the population. Immunogenic antigen targets (e.g., AMA1, MSP1) that give higher population rates of seropositivity provide moderate correlation to gold standard community sampling designs and are a feasible approach to discern foci of residual P. falciparum transmission in an area.

Reaction pathway engineering converts a radical hydroxylase into a halogenase.

FeII/α-ketoglutarate (FeII/αKG)-dependent enzymes offer a promising biocatalytic platform for halogenation chemistry owing to their ability to functionalize unactivated C-H bonds. However, relatively few radical halogenases have been identified to date, limiting their synthetic utility. Here, we report a strategy to expand the palette of enzymatic halogenation by engineering a reaction pathway rather than substrate selectivity. This approach could allow us to tap the broader class of FeII/αKG-dependent hydroxylases as catalysts by their conversion to halogenases. Toward this goal, we discovered active halogenases from a DNA shuffle library generated from a halogenase-hydroxylase pair using a high-throughput in vivo fluorescent screen coupled to an alkyne-producing biosynthetic pathway. Insights from sequencing halogenation-active variants along with the crystal structure of the hydroxylase enabled engineering of a hydroxylase to perform halogenation with comparable activity and higher selectivity than the wild-type halogenase, showcasing the potential of harnessing hydroxylases for biocatalytic halogenation.

Engineering site-selective incorporation of fluorine into polyketides.

Although natural products and synthetic small molecules both serve important medicinal functions, their structures and chemical properties are relatively distinct. To expand the molecular diversity available for drug discovery, one strategy is to blend the effective attributes of synthetic and natural molecules. A key feature found in synthetic compounds that is rare in nature is the use of fluorine to tune drug behavior. We now report a method to site-selectively incorporate fluorine into complex structures to produce regioselectively fluorinated full-length polyketides. We engineered a fluorine-selective trans-acyltransferase to produce site-selectively fluorinated erythromycin precursors in vitro. We further demonstrated that these analogs could be produced in vivo in Escherichia coli on engineering of the fluorinated extender unit pool. By using engineered microbes, elaborate fluorinated compounds can be produced by fermentation, offering the potential for expanding the identification and development of bioactive fluorinated small molecules.

Readability of online patient education material for foregut surgery.

INTRODUCTION: Health literacy is the ability of individuals to use basic health information and services to make well-informed decisions. Low health literacy among surgical patients has been associated with nonadherence to preoperative and/or discharge instructions as well as poor comprehension of surgery. It likely poses as a barrier to patients considering foregut surgery which requires an understanding of different treatment options and specific diet instructions. The objective of this study was to assess and compare the readability of online patient education materials (PEM) for foregut surgery. METHODS: Using Google, the terms "anti-reflux surgery, "GERD surgery," and "foregut surgery" were searched and a total of 30 webpages from universities and national organizations were selected. The readability of the text was assessed with seven instruments: Flesch Reading Ease formula (FRE), Gunning Fog (GF), Flesch-Kincaid Grade Level (FKGL), Coleman Liau Index (CL), Simple Measure of Gobbledygook (SMOG), Automated Readability Index (ARI), and Linsear Write Formula (LWF). Mean readability scores were calculated with standard deviations. We performed a qualitative analysis gathering characteristics such as, type of information (preoperative or postoperative), organization, use of multimedia, inclusion of a version in another language. RESULTS: The overall average readability of the top PEM for foregut surgery was 12th grade. There was only one resource at the recommended sixth grade reading level. Nearly half of PEM included some form of multimedia. CONCLUSIONS: The American Medical Association and National Institute of Health have recommended that PEMs to be written at the 5th-6th grade level. The majority of online PEM for foregut surgery is above the recommended reading level. This may be a barrier for patients seeking foregut surgery. Surgeons should be aware of the potential gaps in understanding of their patients to help them make informed decisions and improve overall health outcomes.

Death-is-life-enhancing: Adaptation and validation of the Norwegian Death Mindsets Measure (NDMM).

While existing psychological frameworks and their accompanying measures focus on death as anxiety-inducing and debilitating, we highlight an overlooked perspective of death-that death can be a basis for living with more meaning and presence. The present research adapts and validates the Death Mindsets Measure (DMM), which assesses the mindset that "death-is-life-enhancing," for a Norwegian context. Firstly, we translated the DMM and consulted with Norwegian bereavement experts and bereaved Norwegians on items' clarity and relevance to cultural perspectives of death. Secondly, we validated the Norwegian DMM (NDMM) on a predominantly bereaved community sample of Norwegians (N = 241). Using structural equation modeling, we confirmed the hierarchical two-factor structure of our measure. The NDMM also demonstrated high internal consistency and discriminant validity with existing death anxiety and death attitudinal measures. Finally, our measure explained additional variance in psychological well-being beyond existing death anxiety and attitudinal measures.

Biosynthesis of Strained Amino Acids by a PLP-Dependent Enzyme through Cryptic Halogenation.

Amino acids (AAs) are modular building blocks which nature uses to synthesize both macromolecules, such as proteins, and small molecule natural products, such as alkaloids and non-ribosomal peptides. While the 20 main proteinogenic AAs display relatively limited side chain diversity, a wide range of non-canonical amino acids (ncAAs) exist that are not used by the ribosome for protein synthesis, but contain a broad array of structural features and functional groups. In this communication, we report the discovery of the biosynthetic pathway for a new ncAA, pazamine, which contains a cyclopropane ring formed in two steps. In the first step, a chlorine is added onto the C4 position of lysine by a radical halogenase, PazA. The cyclopropane ring is then formed in the next step by a pyridoxal-5'-phosphate-dependent enzyme, PazB, via an SN2-like attack at C4 to eliminate chloride. Genetic studies of this pathway in the native host, Pseudomonas azotoformans, show that pazamine potentially inhibits ethylene biosynthesis in growing plants based on alterations in the root phenotype of Arabidopsis thaliana seedlings. We further show that PazB can be utilized to make an alternative cyclobutane-containing AA. These discoveries may lead to advances in biocatalytic production of specialty chemicals and agricultural biotechnology.

Synergistic Binding of the Halide and Cationic Prime Substrate of l-Lysine 4-Chlorinase, BesD, in Both Ferrous and Ferryl States.

An aliphatic halogenase requires four substrates: 2-oxoglutarate (2OG), halide (Cl- or Br-), the halogenation target ("prime substrate"), and dioxygen. In well-studied cases, the three nongaseous substrates must bind to activate the enzyme's Fe(II) cofactor for efficient capture of O2. Halide, 2OG, and (lastly) O2 all coordinate directly to the cofactor to initiate its conversion to a cis-halo-oxo-iron(IV) (haloferryl) complex, which abstracts hydrogen (H•) from the non-coordinating prime substrate to enable radicaloid carbon-halogen coupling. We dissected the kinetic pathway and thermodynamic linkage in binding of the first three substrates of the l-lysine 4-chlorinase, BesD. After addition of 2OG, subsequent coordination of the halide to the cofactor and binding of cationic l-Lys near the cofactor are associated with strong heterotropic cooperativity. Progression to the haloferryl intermediate upon the addition of O2 does not trap the substrates in the active site and, in fact, markedly diminishes cooperativity between halide and l-Lys. The surprising lability of the BesD•[Fe(IV)=O]•Cl•succinate•l-Lys complex engenders pathways for decay of the haloferryl intermediate that do not result in l-Lys chlorination, especially at low chloride concentrations; one identified pathway involves oxidation of glycerol. The mechanistic data imply (i) that BesD may have evolved from a hydroxylase ancestor either relatively recently or under weak selective pressure for efficient chlorination and (ii) that acquisition of its activity may have involved the emergence of linkage between l-Lys binding and chloride coordination following the loss of the anionic protein-carboxylate iron ligand present in extant hydroxylases.

A systematic review of robotic surgery curricula using a contemporary educational framework.

BACKGROUND: There has been a rising trend in robotic surgery. Thus, there is demand for a robotic surgery curriculum (RSC) for training surgical trainees and practicing surgeons. There are limited data available about current curricular designs and the extent to which they have incorporated educational frameworks. Our aim was to study the existing robotic surgery curricula using Kern's 6-step approach in curriculum development. METHODS: A systematic review was conducted using PubMed, PubMed Central, Cochrane, Embase, and Scopus (we searched studies from 2001 to 2021). PRISMA Guidelines was used to guide the search. Curriculum designed for general surgery and its subspecialties were included. Urology and gynecology were excluded. The articles were reviewed by five reviewers. RESULTS: Our review yielded 71 articles, including 39 curricula at 9 different settings. Using Kern's framework, we demonstrated that the majority of robotic surgery curricula contained all the elements of Kern's curricular design. However, there were significant deficiencies in important aspects of these curricula i.e., implementation, the quality of assessment tools for measurement of performance and evaluation of the educational value of these interventions. Most institutions used commercial virtual reality simulators (VRS) as the main component of their RSC and 23% of curricula only used VRS. CONCLUSIONS: Although majority of these studies contained all the elements of Kern's framework, there are critical deficiencies in the components of existing curricula. Future curricula should be designed using established educational frameworks to improve the quality of robotic surgery training.

Early postoperative recovery comparisons of superior capsule reconstruction to tendon transfers.

BACKGROUND: The management of massive posterosuperior rotator cuff tears is controversial, with no gold standard. Two recently developed techniques that have shown promising initial results include arthroscopic superior capsular reconstruction (SCR) and tendon transfers (latissimus or lower trapezius). However, there remains a scarcity of studies examining each procedure's early postoperative clinical outcomes individually or in comparison to each other. The purpose of this study is to compare the early postoperative recovery outcomes of tendon transfers (TTs) to SCR. METHODS: Using the surgical outcomes system global database (Arthrex Inc.), we assessed the postoperative recovery outcomes for all patients who had outcomes recorded at least 6 months after SCR or TT. The time points analyzed included preoperative and postoperative (2 weeks, 6 weeks, 3 months, 6 months, 1 year, 2 years). The outcomes analyzed included pain visual analog scale (VAS) score, American Shoulder and Elbow Surgeons Standardized Shoulder Assessment Form (ASES) score, VR-12 physical, and Single Assessment Numeric Evaluation (SANE). RESULTS: Overall, 163 patients underwent SCR and 24 arthroscopically assisted TT. The mean age for SCR and TT was 60 and 56 years, respectively. Postoperative recovery curves demonstrate that both procedures produced improved outcomes at each postoperative time point compared to preoperative. The pain and functional outcomes measures, including VAS, ASES, SANE, and VR-12 physical, were comparable for TT and SCRs, with similar recovery curves between the 2 techniques. Ultimately at 2 years postoperatively, there were no significant differences between the 2 techniques. CONCLUSIONS: Analysis of the early outcomes associated with arthroscopic treatment of massive posterosuperior rotator cuff tears demonstrated that the arthroscopically assisted tendon transfers and arthroscopic superior capsular reconstruction had similar pain and functional outcomes throughout the 2-year postoperative recovery period. Overall, the process of recovery appears equivalent between the 2 techniques. Future studies are needed to assess the outcomes of each technique and specific indications in an attempt to delineate an algorithm for the treatment of irreparable rotator cuff tears.

The Immediate Effects of a Combined Mass Drug Administration and Indoor Residual Spraying Campaign to Accelerate Progress Toward Malaria Elimination in Grande-Anse, Haiti.

BACKGROUND: Haiti is planning targeted interventions to accelerate progress toward malaria elimination. In the most affected department (Grande-Anse), a combined mass drug administration (MDA) and indoor residual spraying (IRS) campaign was launched in October 2018. This study assessed the intervention's effectiveness in reducing Plasmodium falciparum prevalence. METHODS: An ecological quasi-experimental study was designed, using a pretest and posttest with a nonrandomized control group. Surveys were conducted in November 2017 in a panel of easy access groups (25 schools and 16 clinics) and were repeated 2-6 weeks after the campaign, in November 2018. Single-dose sulfadoxine-pyrimethamine and primaquine was used for MDA, and pirimiphos-methyl as insecticide for IRS. RESULTS: A total of 10 006 participants were recruited. Fifty-two percent of the population in the intervention area reported having received MDA. Prevalence diminished between 2017 and 2018 in both areas, but the reduction was significantly larger in the intervention area (ratio of adjusted risk ratios, 0.32 [95% confidence interval, .104-.998]). CONCLUSIONS: Despite a moderate coverage, the campaign was effective in reducing P. falciparum prevalence immediately after 1 round. Targeted MDA plus IRS is useful in preelimination settings to rapidly decrease the parasite reservoir, an encouraging step to accelerate progress toward malaria elimination.

Factors Associated With Human IgG Antibody Response to Anopheles albimanus Salivary Gland Extract, Artibonite Department, Haiti, 2017.

Serological data can provide estimates of human exposure to both malaria vector and parasite based on antibody responses. A multiplex bead-based assay was developed to simultaneously detect IgG to Anopheles albimanus salivary gland extract (SGE) and 23 Plasmodium falciparum antigens among 4185 participants enrolled in Artibonite department, Haiti in 2017. Logistic regression adjusted for participant- and site-level covariates and found children under 5 years and 6-15 years old had 3.7- and 5.4-fold increase in odds, respectively, of high anti-SGE IgG compared to participants >15 years. Seropositivity to P. falciparum CSP, Rh2_2030, and SEA-1 antigens was significantly associated with high IgG response against SGE, and participant enrolment at elevations under 200 m was associated with higher anti-SGE IgG levels. The ability to approximate population exposure to malaria vectors through SGE serology data is very dependent by age categories, and SGE antigens can be easily integrated into a multiplex serological assay.

Postoperative recovery comparisons of arthroscopic Bankart to open Latarjet for the treatment of anterior glenohumeral instability.

BACKGROUND: Recurrent anterior glenohumeral instability is a disabling pathology that can be successfully treated by arthroscopic Bankart repair or open Latarjet. However, there is a paucity of studies comparing the postoperative recovery. The purpose of this study is to evaluate the postoperative pain and functional recovery following arthroscopic Bankart versus open Latarjet. METHODS: This is a retrospective analysis of a multicenter prospective outcomes registry database. Postoperative recovery outcomes of either a primary or revision arthroscopic Bankart and open Latarjet procedures were compared. A minimum of 1-year follow-up was required. Outcomes measures included pain visual analog scale (VAS), American Shoulder and Elbow Surgeons (ASES) function score, ASES index score, and single assessment numeric evaluation (SANE) score. Overall, 787 patients underwent primary arthroscopic Bankart, 36 underwent revision arthroscopic Bankart and 75 underwent an open Latarjet procedure. RESULTS: When compared to primary arthroscopic Bankart, open Latarjet demonstrated significantly lower VAS scores at 6 weeks (p = 0.03), 3 months (p = 0.01), and 2 years (p < 0.05). Medium-term outcomes for ASES scores and SANE score, at 1 and 2 years showed no difference. Latarjet demonstrated significantly lower (p < 0.05) preoperative early postoperative VAS pain scores with no difference at 1 year or 2 years when compared to primary Bankart. There was no difference in ASES function or index between Bankart and Latarjet. Revision Bankart provided inferior outcomes for VAS, ASES function, and ASES index when compared to primary Bankart and Latarjet at 1 year and 2 years. CONCLUSIONS: Primary arthroscopic Bankart repair and open Latarjet provided nearly equivalent improvements in pain (VAS) and functional outcomes (ASES, SANE, VR-12) during the early recovery phase (2 years). This study supports the use of either procedure in the primary treatment of anterior glenohumeral instability. Revision arthroscopic Bankart repair demonstrated deteriorating outcomes at 1 and 2 years postoperatively.

Substrate-Triggered µ-Peroxodiiron(III) Intermediate in the 4-Chloro-l-Lysine-Fragmenting Heme-Oxygenase-like Diiron Oxidase (HDO) BesC: Substrate Dissociation from, and C4 Targeting by, the Intermediate.

The enzyme BesC from the ß-ethynyl-l-serine biosynthetic pathway in Streptomyces cattleya fragments 4-chloro-l-lysine (produced from l-Lysine by BesD) to ammonia, formaldehyde, and 4-chloro-l-allylglycine and can analogously fragment l-Lys itself. BesC belongs to the emerging family of O2-activating non-heme-diiron enzymes with the "heme-oxygenase-like" protein fold (HDOs). Here, we show that the binding of l-Lys or an analogue triggers capture of O2 by the protein's diiron(II) cofactor to form a blue µ-peroxodiiron(III) intermediate analogous to those previously characterized in two other HDOs, the olefin-installing fatty acid decarboxylase, UndA, and the guanidino-N-oxygenase domain of SznF. The ∼5- and ∼30-fold faster decay of the intermediate in reactions with 4-thia-l-Lys and (4RS)-chloro-dl-lysine than in the reaction with l-Lys itself and the primary deuterium kinetic isotope effects (D-KIEs) on decay of the intermediate and production of l-allylglycine in the reaction with 4,4,5,5-[2H4]-l-Lys suggest that the peroxide intermediate or a reversibly connected successor complex abstracts a hydrogen atom from C4 to enable olefin formation. Surprisingly, the sluggish substrate l-Lys can dissociate after triggering intermediate formation, thereby allowing one of the better substrates to bind and react. The structure of apo BesC and the demonstrated linkage between Fe(II) and substrate binding suggest that the triggering event involves an induced ordering of ligand-providing helix 3 (α3) of the conditionally stable HDO core. As previously suggested for SznF, the dynamic α3 also likely initiates the spontaneous degradation of the diiron(III) product cluster after decay of the peroxide intermediate, a trait emerging as characteristic of the nascent HDO family.

Multistate Outbreak of SARS-CoV-2 Infections, Including Vaccine Breakthrough Infections, Associated with Large Public Gatherings, United States.

During July 2021, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) B.1.617.2 variant infections, including vaccine breakthrough infections, occurred after large public gatherings in Provincetown, Massachusetts, USA, prompting a multistate investigation. Public health departments identified primary and secondary cases by using coronavirus disease surveillance data, case investigations, and contact tracing. A primary case was defined as SARS-CoV-2 detected <14 days after travel to or residence in Provincetown during July 3-17. A secondary case was defined as SARS-CoV-2 detected <14 days after close contact with a person who had a primary case but without travel to or residence in Provincetown during July 3-August 10. We identified 1,098 primary cases and 30 secondary cases associated with 26 primary cases among fully and non-fully vaccinated persons. Large gatherings can have widespread effects on SARS-CoV-2 transmission, and fully vaccinated persons should take precautions, such as masking, to prevent SARS-CoV-2 transmission, particularly during substantial or high transmission.

Prolonged severe acute respiratory syndrome coronavirus 2 persistence, attenuated immunologic response, and viral evolution in a solid organ transplant patient.

Unlike immunocompetent hosts, the duration of viral persistence after infection with severe acute respiratory syndrome coronavirus 2 can be prolonged in immunosuppressed patients. Here, we present a case of viral persistence for over 19 weeks in a patient with a history of solid organ transplant and explore the clinical, virologic, and immunologic course. Our patient still demonstrated viral persistence at 138 days with low polymerase chain reaction cycle threshold values and evidence of continuing viral sequence evolution indicative of ongoing virus replication. These findings have important implications for infection prevention and control recommendations in immunosuppressed patients. Immune response, including neutralizing antibody titers, T cell activity, and cytokine levels, peaked around days 44-72 after diagnosis. Anti-S trimer antibodies were low at all time points, and T cell response was attenuated by day 119. As immune response waned and viral load increased, increased genetic diversity emerged, suggesting a mechanism for the development of viral variants.

ISCHEMIA-EXTEND studies: Rationale and design.

BACKGROUND: The ISCHEMIA and the ISCHEMIA-CKD trials found no statistical difference in the primary clinical endpoint between initial invasive management and initial conservative management of patients with chronic coronary disease and moderate to severe ischemia on stress testing without or with advanced chronic kidney disease (CKD). In ISCHEMIA, there was numerically lower cardiovascular mortality but higher non-cardiovascular mortality with no significant difference in all-cause death with an initial invasive strategy when compared with a conservative strategy. However, an invasive strategy increased peri-procedural myocardial infarction (MI) but decreased spontaneous MI with continued separation of curves over time, which potentially may lead to reduced risk of cardiovascular and all-cause mortality. Thus, the long-term effect of invasive management strategy on mortality remains unclear. In ISCHEMIA-CKD, the treatment and cause-specific mortality rates were similar during follow-up. METHODS: Funded by the National Heart, Lung, and Blood Institute, the ISCHEMIA-EXTEND observational study is the long-term follow-up of surviving participants (projected median of 10 years) with chronic coronary disease from the ISCHEMIA trial. In the ISCHEMIA trial, 5,179 participants with moderate or severe stress-induced ischemia were randomized to initial invasive management with angiography, revascularization when feasible, and guideline-directed medical therapy (GDMT), or initial conservative management with GDMT alone and angiography reserved for failure of medical therapy. ISCHEMIA-CKD EXTEND is the long-term follow-up of surviving participants (projected median of 9 years) from the ISCHEMIA-CKD trial, a companion trial that included 777 patients with advanced CKD. Ascertainment of death will be conducted via direct participant contact, medical record review, and/or vital status registry search. The overarching objective of long-term follow-up is to assess whether there are between-group differences in long-term all-cause, cardiovascular, and non-cardiovascular mortality, and increase precision around the treatment effect estimates for risk of all-cause, cardiovascular, and non-cardiovascular mortality. We will conduct Bayesian survival modeling to take advantage of rich inferences using the posterior distribution of the treatment effect. CONCLUSIONS: The long-term effect of an initial invasive versus conservative strategy on all-cause, cardiovascular, and non-cardiovascular mortality will be assessed. The findings of ISCHEMIA-EXTEND and ISCHEMIA-CKD EXTEND will inform patients, practitioners, practice guidelines, and health policy.