SARS-CoV-2 RBD antibodies that maximize breadth and resistance to escape.

An ideal therapeutic anti-SARS-CoV-2 antibody would resist viral escape1-3, have activity against diverse sarbecoviruses4-7, and be highly protective through viral neutralization8-11 and effector functions12,13. Understanding how these properties relate to each other and vary across epitopes would aid the development of therapeutic antibodies and guide vaccine design. Here we comprehensively characterize escape, breadth and potency across a panel of SARS-CoV-2 antibodies targeting the receptor-binding domain (RBD). Despite a trade-off between in vitro neutralization potency and breadth of sarbecovirus binding, we identify neutralizing antibodies with exceptional sarbecovirus breadth and a corresponding resistance to SARS-CoV-2 escape. One of these antibodies, S2H97, binds with high affinity across all sarbecovirus clades to a cryptic epitope and prophylactically protects hamsters from viral challenge. Antibodies that target the angiotensin-converting enzyme 2 (ACE2) receptor-binding motif (RBM) typically have poor breadth and are readily escaped by mutations despite high neutralization potency. Nevertheless, we also characterize a potent RBM antibody (S2E128) with breadth across sarbecoviruses related to SARS-CoV-2 and a high barrier to viral escape. These data highlight principles underlying variation in escape, breadth and potency among antibodies that target the RBD, and identify epitopes and features to prioritize for therapeutic development against the current and potential future pandemics.

Effects of prepartum metabolizable protein supply and management strategy on lactational performance and blood biomarkers in dairy cows during early lactation.

Our objective was to investigate the effects of prepartum metabolizable protein (MP) supply and management strategy on milk production and blood biomarkers in early lactation dairy cows. Ninety-six multigravida Holstein cows were used in a randomized complete block design study, blocked by calving date, and then assigned randomly to 1 of 3 treatments within block. Cows on the first treatment were fed a far-off lower MP diet [MP = 83 g/kg of dry matter (DM)] between -55 and -22 d before expected calving and then a close-up lower MP diet (MP = 83 g/kg of DM) until parturition (LPLP). Cows on the second treatment were fed the far-off lower MP diet between -55 to -22 d before expected parturition and then a prepartum higher MP diet (MP = 107 g/kg of DM) until calving (LPHP). Cows on the third treatment had a shortened 43-d dry period and were fed the prepartum higher MP diet from dry-off to parturition (SDHP). After calving, cows received the same fresh diet from d 0 to 14 and the same high diet from d 15 to 84. Data were analyzed separately for wk -6 to -1 and wk 1 to 12, relative to parturition. Dry matter intake from wk -6 to -1 was not different between LPHP and LPLP and increased for SDHP compared with LPLP. In contrast, dry matter intake for wk 1 to 12 postpartum did not change for LPHP versus LPLP or for SDHP versus LPLP. Compared with LPLP cows, LPHP cows had lower energy-corrected milk yield and tended to have decreased milk fat yield during wk 1 to 12 of lactation. Conversely, yields of energy-corrected milk and milk fat and protein were similar for SDHP compared with LPLP. Plasma urea N during wk -3 to -1 increased for LPHP versus LPLP and for SDHP versus LPLP; however, no differences in plasma urea N were observed postpartum. Elevated prepartum MP supply did not modify circulating total fatty acids, ß-hydroxybutyrate, total protein, albumin, or aspartate aminotransferase during the prepartum and postpartum periods. Increased MP supply prepartum combined with a shorter dry period (SDHP vs. LPLP) tended to increase whole-blood ß-hydroxybutyrate postpartum; however, other blood metabolites were not affected. Taken together, under the conditions of this study, elevated MP supply in close-up diets reduced milk production without affecting blood metabolites in multiparous dairy cows during early lactation. A combination of a shorter dry period and increased prepartum MP supply (i.e., SDHP vs. LPLP) improved prepartum dry matter intake without modifying energy-corrected milk yield and blood biomarkers in early lactation cows.

"Empowerment for Us by Us (E4UBU)": Developing a Model of Empowerment Using Feminist Participatory Methods with LBQT+ Persons Assigned Female at Birth in Western Kenya.

Lesbian, bisexual, queer, trans and other gender diverse persons assigned female at birth (heretofore referred to as "LBQT+ persons") in Western Kenya experience intersectional oppression and stigma. This stigma can manifest in acts of sexual and gender-based violence (SGBV) and sexual and gender minority (SGM)-based violence, as well as various forms of discrimination-all of which have been linked to disproportionately higher levels of negative health outcomes for this group. Despite these challenges, many LBQT+ persons have been able to gain personal and collective power and thrive in this oppressive environment. The Empowerment for Us by Us (E4UBU) project is a mixed methods feminist participatory research study focused on exploring how LBQT+ persons conceptualize and define empowerment for themselves, and to understand their perspectives on how feelings of power and powerlessness influence their physical and mental health. This paper focuses on data from the first phase of the study, in which qualitative in-depth interviews were conducted with 40 LBQT+ persons (ages 19 to 50) from Kisumu and Homa Bay in Western Kenya. A participatory interpretive phenomenological analysis was conducted to understand the lived experiences of LBQT+ persons as they navigate intersectional oppression and its influence on their experiences of empowerment and subsequent health outcomes. Findings from this analysis were presented to two different focus groups composed of participants who had participated in the in-depth interviews to gather their insights on the interpretations of the interviews as a form of member checking. Findings revealed that "empowerment" was not experienced and viewed by LBQT+ persons as a monolithic construct, but rather a process through which LBQT+ persons are able to transform negative forces of intersectional oppression and powerlessness into experiences of power and subsequent individual and collective action and impact-all leading to improved mental health and well-being. This process is facilitated at several junctures by participatory seeking and attainment of community-appropriate resources at multiple socio-ecological levels that, when accessed with sufficient intensity, frequency, and duration, enhance one's journey through the process of empowerment. These facilitation junctures are viewed as likely points of focus for public health intervention. Analysis also revealed that the process of empowerment is dependent on the context within which the process is occurring, the specific issues being faced, and the population of focus. Recommendations for how this model can be used for future research and practice to improve the lives of LBQT+ persons in Kenya are discussed.

A Retrospective Application of the Arbon and Hartman Models to the Union Cycliste International Mountain Bike World Cup.

INTRODUCTION: Outdoor activities have accelerated in the past several years. The authors were tasked with providing medical care for the Union Cycliste International (UCI) mountain biking World Cup in Snowshoe, West Virginia (USA) in September 2021. The Hartman and Arbon models were designed to predict patient presentation and hospital transport rates as well as needed medical resources at urban mass-gathering events. However, there is a lack of standardized methods to predict injury, illness, and insult severity at rural mass gatherings. STUDY OBJECTIVE: This study aimed to determine whether the Arbon model would predict, within 10%, the number of patient presentations to be expected and to determine if the event classification provided by the Hartman model would adequately predict resources needed during the event. METHODS: Race data were collected from UCI event officials and injury data were collected from participants at time of presentation for medical care. Predicted presentation and transport rates were calculated using the Arbon model, which was then compared to the actual observed presentation rates. Furthermore, the event classification provided by the Hartman model was compared to the resources utilized during the event. RESULTS: During the event, 34 patients presented for medical care and eight patients required some level of transport to a medical facility. The Arbon predictive model for the 2021 event yielded 30.3 expected patient presentations. There were 34 total patient presentations during the 2021 race, approximately 11% more than predicted. The Hartman model yielded a score of four. Based on this score, this race would be classified as an "intermediate" event, requiring multiple Advanced Life Support (ALS) and Basic Life Support (BLS) personnel and transport units. CONCLUSION: The Arbon model provided a predicted patient presentation rate within reasonable error to allow for effective pre-event planning and resource allocation with only a four patient presentation difference from the actual data. While the Arbon model under-predicted patient presentations, the Hartman model under-estimated resources needed due to the high-risk nature of downhill cycling. The events staffed required physician skills and air medical services to safely care for patients. Further evaluation of rural events will be needed to determine if there is a generalized need for physician presence at smaller events with inherently risky activities, or if this recurring cycling event is an outlier.

Safety and Efficacy of Low-Dose Versus High-Dose Parenteral Ketorolac for Acute Pain Relief in Patients 65 Years and Older in the Emergency Department.

Background There is limited data surrounding acute pain management in elderly ED patients. Ketorolac is a potent non-steroidal anti-inflammatory drug (NSAID) with dose/duration-dependent side effects. There is evidence that an analgesic ceiling effect exists for parenteral ketorolac doses greater than 10 milligrams (mg); however, this has not been studied in patients 65 years and older. Methods This was a retrospective chart review of ED patients 65 years and older who received at least one dose of parenteral ketorolac. Patients were separated into two cohorts based on the ketorolac dose received: 15 mg IV or 30 mg intramuscular (IM) and 30 mg IV or 60 mg IM. The primary objective was to evaluate the analgesic efficacy of parenteral ketorolac doses measured as needing rescue analgesia from 30 minutes to 2 hours after ketorolac administration. Secondary objectives included changes in pain scores and the occurrence of adverse drug events commonly associated with ketorolac. Results Two-hundred and sixty patients received ketorolac doses of 15 mg IV or 30 mg IM, and 52 received 30 mg IV or 60 mg IM. The primary outcome occurred in seven of 52 patients who received ketorolac 30 mg IV or 60 mg IM and 17 of 260 patients who received ketorolac 15 mg IV or 30 mg IM (13.5% vs. 6.5%, p=0094; OR: 2.22, 95% CI: 0.87-5.67). The average change in pain scores were 2.9 (±3.1) and 2.8 (±2.9) for patients who received doses 30 mg IV or 60 mg IM compared to doses 15 mg IV or 30 mg IM, respectively (p=0.154). The occurrence of adverse events was low in both groups. Conclusion Parenteral ketorolac doses of 15 mg IV or 30 mg IM did not demonstrate a greater need for rescue analgesia compared to doses of 30 mg IV or 60 mg IM.

Evaluation of peripartum supplementation of methionine hydroxy analogue on beef cow-calf performance.

The objective was to evaluate the effects of peripartum supplementation of a methionine hydroxy analogue (MHA) to primiparous, spring-calving beef females on dam and progeny performance. Angus heifers (n = 60) were blocked by expected parturition date, stratified by body weight (BW) and body condition score (BCS), and randomized to 1 of 15 pens. Pens were randomly assigned to 1 of 3 dietary treatments: a basal diet supplemented with 0 (M0), 15 (M15), or 30 (M30) g/animal/d of MHA (provided as MFP feed supplement, Novus International Inc., St. Charles, MO). Diets were fed from 45 ±â€…13 (SD) d pre-calving through 81 ±â€…13 d postpartum (DPP), after which all cow-calf pairs were managed as a single group on pasture until weaning (199 ±â€…13 DPP). Dam BW, BCS, and blood samples were taken at 6 predetermined timepoints. Progeny data were collected at birth, 2 intermediate timepoints, and at weaning. Milk samples were collected for composition analysis at 7 ±â€…2 DPP and at 55 ±â€…5 DPP. Serial progesterone samples were analyzed to establish resumption of cyclicity, and ultrasonography was performed at 55 ±â€…5 DPP to evaluate ovarian function. Cows were bred via artificial insemination at 82 ±â€…13 DPP and subsequently exposed to bulls for a 55-d breeding season. Pen was the experimental unit, and preplanned orthogonal contrasts were tested (linear effect and M0 vs. M15 + M30). Dam BW and BCS were not affected by treatment (P ≥ 0.29) throughout the study. Week 1 milk fat concentration increased linearly (P = 0.05) and total solids tended to increase linearly (P = 0.07) as MHA increased; however, no other milk components were affected (P ≥ 0.16). Treatment did not affect (P ≥ 0.16) dam reproductive parameters or progeny growth from birth until weaning. Post-calving, circulating methionine equivalents tended to linearly increase (P = 0.10) with increasing MHA supplementation. At breeding, plasma urea N linearly decreased (P = 0.03) with increased supplementation of MHA, and plasma non-esterified fatty acids were less (P = 0.04) in MHA-supplemented dams compared with dams receiving no MHA. Maternal circulating glucose, glutathione peroxidase, and thiobarbituric acid-reactive substances were not affected (P ≥ 0.15) by treatment at any point. These data indicate that peripartum supplementation of MHA may increase milk fat composition shortly after calving, but MHA supplementation did not improve progeny growth or dam reproductive performance in the current study.

Effects of copper, zinc, and manganese source and inclusion during late gestation on beef cow-calf performance, mineral transfer, and metabolism.

To determine effects of Cu, Zn, and Mn source and inclusion during late gestation, multiparous beef cows [n = 48; 649 ±â€…80 kg body weight (BW); 5.3 ±â€…0.5 body condition score (BCS)] were individually-fed hay and supplement to meet or exceed all nutrient recommendations except Cu, Zn, and Mn. From 91.2 ±â€…6.2 d pre-calving to 11.0 ±â€…3.2 d post-calving, cows received: no additional Cu, Zn, or Mn (control, CON), sulfate-based Cu, Zn, and Mn (inorganic, ITM) or metal methionine hydroxy analogue chelates (MMHAC) of Cu, Zn, and Mn at 133% recommendations, or a combination of inorganic and chelated Cu, Zn, and Mn (reduce and replace, RR) to meet 100% of recommendations. Data were analyzed with treatment and breeding group (and calf sex if P < 0.25 for offspring measures) as fixed effects, animal as experimental unit, and sampling time as a repeated effect for serum, plasma, and milk measures over time. Post-calving cow liver Cu was greater (P ≤ 0.07) in MMHAC compared with all other treatments. Calves born to RR had greater (P ≤ 0.05) liver Cu than ITM and CON, and MMHAC had greater (P = 0.06) liver Cu than CON. Liver Mn was less (P ≤ 0.08) for RR calves than all other treatments. Calf plasma Zn was maintained (P ≥ 0.15) from 0 to 48 h of age in ITM and MMHAC but decreased (P ≤ 0.03) in CON and RR. Gestational cow BW, BCS, and metabolites were not affected (P ≥ 0.13) by treatment, but gestational serum thiobarbituric acid reactive substances (TBARS) were greater (P = 0.01) for CON than MMHAC. Treatment did not affect (P ≥ 0.13) calf birth size, vigor, placental size and minerals, or transfer of passive immunity. Neonatal calf serum Ca was greater (P ≤ 0.05) for MMHAC than all other treatments; other calf serum chemistry and plasma cortisol were not affected (P ≥ 0.12). Pre-suckling colostrum yield, and lactose concentration and content, were greater (P ≤ 0.06) for MMHAC compared with ITM and RR. Colostral triglyceride and protein concentrations were greater (P ≤ 0.08) for RR than MMHAC and CON. Cow lactational BW and BCS, milk yield and composition, and pre-weaning calf BW and metabolism were not affected (P ≥ 0.13) by treatment. Lactational serum TBARS were greater (P = 0.04) for RR than CON at day 35 and greater (P ≤ 0.09) for MMHAC at day 60 than all other treatments. Source and inclusion of Cu, Zn, and Mn altered maternal and neonatal calf mineral status, but calf size and vigor at birth, passive transfer, and pre-weaning growth were not affected in this study.

Antibodies to the SARS-CoV-2 receptor-binding domain that maximize breadth and resistance to viral escape.

An ideal anti-SARS-CoV-2 antibody would resist viral escape 1-3 , have activity against diverse SARS-related coronaviruses 4-7 , and be highly protective through viral neutralization 8-11 and effector functions 12,13 . Understanding how these properties relate to each other and vary across epitopes would aid development of antibody therapeutics and guide vaccine design. Here, we comprehensively characterize escape, breadth, and potency across a panel of SARS-CoV-2 antibodies targeting the receptor-binding domain (RBD), including S309 4 , the parental antibody of the late-stage clinical antibody VIR-7831. We observe a tradeoff between SARS-CoV-2 in vitro neutralization potency and breadth of binding across SARS-related coronaviruses. Nevertheless, we identify several neutralizing antibodies with exceptional breadth and resistance to escape, including a new antibody (S2H97) that binds with high affinity to all SARS-related coronavirus clades via a unique RBD epitope centered on residue E516. S2H97 and other escape-resistant antibodies have high binding affinity and target functionally constrained RBD residues. We find that antibodies targeting the ACE2 receptor binding motif (RBM) typically have poor breadth and are readily escaped by mutations despite high neutralization potency, but we identify one potent RBM antibody (S2E12) with breadth across sarbecoviruses closely related to SARS-CoV-2 and with a high barrier to viral escape. These data highlight functional diversity among antibodies targeting the RBD and identify epitopes and features to prioritize for antibody and vaccine development against the current and potential future pandemics.

Ultrapotent human antibodies protect against SARS-CoV-2 challenge via multiple mechanisms.

Efficient therapeutic options are needed to control the spread of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) that has caused more than 922,000 fatalities as of 13 September 2020. We report the isolation and characterization of two ultrapotent SARS-CoV-2 human neutralizing antibodies (S2E12 and S2M11) that protect hamsters against SARS-CoV-2 challenge. Cryo-electron microscopy structures show that S2E12 and S2M11 competitively block angiotensin-converting enzyme 2 (ACE2) attachment and that S2M11 also locks the spike in a closed conformation by recognition of a quaternary epitope spanning two adjacent receptor-binding domains. Antibody cocktails that include S2M11, S2E12, or the previously identified S309 antibody broadly neutralize a panel of circulating SARS-CoV-2 isolates and activate effector functions. Our results pave the way to implement antibody cocktails for prophylaxis or therapy, circumventing or limiting the emergence of viral escape mutants.