Immune response to SARS-CoV-2 variants after immunization with different vaccines in Mexico.

There is limited information on the antibody responses against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in subjects from developing countries with populations having a high incidence of co-morbidities. Here, we analysed the immunogenicity of homologous schemes using the ChAdOx1-S, Sputnik V, or BNT162b2 vaccines and the effect of a booster dose with ChAdOx1-S in middle-aged adults who were seropositive or seronegative to the SARS-CoV-2 spike protein before vaccination. The study was conducted post-vaccination with a follow-up of 4 months for antibody titre using enzyme-linked immunosorbent assay (ELISA) and pseudovirus (PV) neutralization assays (PNAs). All three vaccines elicited a superior IgG anti-receptor-binding domain (RBD) and neutralization response against the Alpha and Delta variants when administered to individuals with a previous infection by SARS-CoV-2. The booster dose spiked the neutralization activity among individuals with and without a prior SARS-CoV-2 infection. The ChAdOx1-S vaccine induced weaker antibody responses in infection-naive subjects. A follow-up of 4 months post-vaccination showed a drop in antibody titre, with about 20% of the infection-naive and 100% of SARS-CoV-2 pre-exposed participants with detectable neutralization capacity against Alpha pseudovirus (Alpha-PV) and Delta PV (Delta-PV). Our observations support the use of different vaccines in a country with high seroprevalence at the vaccination time.

The Pre-Existing Human Antibody Repertoire to Computationally Optimized Influenza H1 Hemagglutinin Vaccines.

Computationally optimized broadly reactive Ag (COBRA) hemagglutinin (HA) immunogens have previously been generated for several influenza subtypes to improve vaccine-elicited Ab breadth. As nearly all individuals have pre-existing immunity to influenza viruses, influenza-specific memory B cells will likely be recalled upon COBRA HA vaccination. We determined the epitope specificity and repertoire characteristics of pre-existing human B cells to H1 COBRA HA Ags. Cross-reactivity between wild-type HA and H1 COBRA HA proteins P1, X6, and Y2 were observed for isolated mAbs. The mAbs bound five distinct epitopes on the pandemic A/California/04/2009 HA head and stem domains, and most mAbs had hemagglutination inhibition and neutralizing activity against 2009 pandemic H1 strains. Two head-directed mAbs, CA09-26 and CA09-45, had hemagglutination inhibition and neutralizing activity against a prepandemic H1 strain. One mAb, P1-05, targeted the stem region of H1 HA, but did not compete with a known stem-targeting H1 mAb. We determined that mAb P1-05 recognizes a recently discovered HA epitope, the anchor epitope, and we identified similar mAbs using B cell repertoire sequencing. In addition, the trimerization domain distance from HA was critical to recognition of this epitope by mAb P1-05, suggesting the importance of protein design for vaccine formulations. Overall, these data indicate that seasonally vaccinated individuals possess a population of functional H1 COBRA HA-reactive B cells that target head, central stalk, and anchor epitopes, and they demonstrate the importance of structure-based assessment of subunit protein vaccine candidates to ensure accessibility of optimal protein epitopes.

Spreading Standardized Documentation of Pediatric Occupational and Physical Therapy Dose: A Quality Improvement Initiative.

IMPORTANCE: Clinicians and researchers can leverage clinical documentation of therapy services for quality improvement and research purposes. However, documentation is often institution specific and may not adequately capture the dose of therapy delivered, thus limiting collaboration. OBJECTIVE: To implement documentation of pediatric occupational and physical therapy dose from one institution to another institution. Dose documentation includes the frequency, intensity, time, and type of interventions delivered (FITT Epic® Flowsheet) at each session. DESIGN: Prospective time-series quality improvement study. SETTING: Two large urban pediatric hospitals. PARTICIPANTS: Occupational and physical therapy staff members. INTERVENTION: Staff training and feedback loops utilizing existing groups. OUTCOMES AND MEASURES: The process measure (number of available staff trained and using the FITT Epic Flowsheet over time) and the outcome measure (percentage of FITT Epic Flowsheets used for treatment visits each month) were analyzed using a statistical process control chart. The balancing measure (percentage of notes closed before 7 p.m. on the same day as the encounter) was analyzed using mean per month across three time periods. RESULTS: Fifty-seven staff members (68%) attended formal training. On average, clinicians documented 90% of sessions using the FITT Epic Flowsheet after implementation. There was no change observed in the balancing measure. CONCLUSIONS AND RELEVANCE: Documentation of dose was spread from one institution to another. Shared documentation will facilitate future collaboration for quality improvement and research purposes. Occupational therapy practitioners and leaders should consider implementing documentation with common dose elements. Plain-Language Summary: Occupational therapy documentation is often institution specific and may not adequately capture the dose of therapy (frequency, intensity, time) or types of interventions that were delivered, thus limiting opportunities for collaboration between institutions. This article adds to the literature on administrative supports for clinical and quality improvement research by illustrating a specific example of how documentation of dose can be shared from one institution to another. The data show that clinicians at one institution started using a new style of documentation using the FITT Epic® Flowsheet and shared discrete dose elements with another institution, creating new opportunities for collaboration. Shared documentation can facilitate future collaboration for quality improvement and research purposes.

Structure-Based Design and Antigenic Validation of Respiratory Syncytial Virus G Immunogens.

Respiratory syncytial virus (RSV) is a leading cause of severe lower respiratory tract disease of children, the elderly, and immunocompromised individuals. Currently, there are no FDA-approved RSV vaccines. The RSV G glycoprotein is used for viral attachment to host cells and impairment of host immunity by interacting with the human chemokine receptor CX3CR1. Antibodies that disrupt this interaction are protective against infection and disease. Nevertheless, development of an RSV G vaccine antigen has been hindered by its low immunogenicity and safety concerns. A previous study described three engineered RSV G proteins containing single-point mutations that induce higher levels of IgG antibodies and have improved safety profiles compared to wild-type RSV G (H. C. Bergeron, J. Murray, A. M. Nuñez Castrejon, et al., Viruses 13:352, 2021, https://doi.org/10.3390/v13020352). However, it is unclear if the mutations affect RSV G protein folding and display of its conformational epitopes. In this study, we show that the RSV G S177Q protein retains high-affinity binding to protective human and mouse monoclonal antibodies and has equal reactivity as wild-type RSV G protein to human reference immunoglobulin to RSV. Additionally, we determined the high-resolution crystal structure of RSV G S177Q protein in complex with the anti-RSV G antibody 3G12, further validating its antigenic structure. These studies show for the first time that an engineered RSV G protein with increased immunogenicity and safety retains conformational epitopes to high-affinity protective antibodies, supporting its further development as an RSV vaccine immunogen. IMPORTANCE Respiratory syncytial virus (RSV) causes severe lower respiratory diseases of children, the elderly, and immunocompromised populations. There currently are no FDA-approved RSV vaccines. Most vaccine development efforts have focused on the RSV F protein, and the field has generally overlooked the receptor-binding antigen RSV G due to its poor immunogenicity and safety concerns. However, single-point mutant RSV G proteins have been previously identified that have increased immunogenicity and safety. In this study, we investigate the antibody reactivities of three known RSV G mutant proteins. We show that one mutant RSV G protein retains high-affinity binding to protective monoclonal antibodies, is equally recognized by anti-RSV antibodies in human sera, and forms the same three-dimensional structure as the wild-type RSV G protein. Our study validates the structure-guided design of the RSV G protein as an RSV vaccine antigen.

Trauma radiology teaching for foundation doctors working within the Scottish Trauma Network improves radiology requests and patient safety: a multidepartmental quality improvement project.

The importance of radiology in trauma management is particularly relevant today as Scotland develops its Scottish Trauma Network. Trauma and radiology are scarcely covered in the 2016 and 2021 Foundation Programme Curriculum. Trauma is a significant and endemic public health challenge, while radiology is ever growing in use as a diagnostic and interventional tool. Currently, the majority of radiological investigation requests in trauma are made by foundation doctors. Accordingly, there is an urgent need to ensure foundation doctors are adequately trained in trauma radiology. This prospective, multidepartmental quality improvement project at a single major trauma centre primarily analysed the impact of trauma radiology teaching on the quality of foundation doctors' radiology requests according to Ionising Radiation Medical Exposure Regulations (IRMER) criteria. As a secondary outcome, the impact of teaching on patient safety was also evaluated. 50 foundation doctors across three departments dealing with trauma patients had their trauma radiology requests analysed before and after the intervention of trauma-focused radiology teaching. Results showed cancelled and altered radiology requests decreased from 20% to 5% and from 25% to 10%, respectively, with a p value of 0.01. This translated to fewer delays in trauma patients receiving radiological investigations. The foundation curriculum would benefit from the introduction of trauma radiology teaching for its foundation doctors, in parallel to the demands of a growing national trauma network. Education globally improves radiology request quality by raising awareness and respect of IRMER criteria and ultimately leads to positive changes for patient safety.

Implementing Parent Coaching in Hospital-Based Pediatric Occupational Therapy: A Multisite Quality Improvement Project.

IMPORTANCE: Parent coaching (PC) is a best practice for young children with, or at high risk for, cerebral palsy (CP). Occupational therapy practitioners in outpatient settings encounter barriers to implementing PC. OBJECTIVE: To increase the documented use of PC in outpatient occupational therapy visits for children younger than age 2 yr with, or at high risk for, CP from 0% to 80%. DESIGN: Multicenter quality improvement (QI) initiative with a time-series design. SETTING: Three pediatric tertiary-care institutions, each with multiple outpatient occupational therapy clinics. PARTICIPANTS: Practitioners in the outpatient clinics and patients <2 yr old with, or at high risk for, cerebral palsy. INTERVENTION: Plan-do-study-act cycles included interventions packaged as a toolkit: education sessions, quick references, electronic medical record (EMR) supports, and site-specific strategies. OUTCOMES AND MEASURES: The primary outcome measure was the use of PC in outpatient sessions. Process measures included pre- and posteducation practitioner knowledge scores and an EMR checklist. Balancing measures (ensuring that changes do not cause problems in other areas) of parent satisfaction/experience and practitioner productivity were measured pre- and postintervention. RESULTS: The primary outcome measure goal (80% documented use of PC in sessions) was attained in the seventh month of the study, sustained for 4 mo, and settled at 79.1% for the remaining 6 mo. Practitioner knowledge scores increased from 83.1% to 87.9% after initial education sessions, t[56] = 3.289, p = .001. Parent satisfaction/experience and practitioner productivity scores did not change. CONCLUSIONS AND RELEVANCE: QI methodology can support PC implementation in pediatric outpatient practice. What This Article Adds: This multisite QI initiative shows that outpatient occupational therapy practitioners can implement PC as a best practice with the use of a toolkit. Results suggest that education alone does not result in changes to practitioner behavior and that QI methods can help when implementing best practices in a clinical setting.

Targeting the Conserved Stem Loop 2 Motif in the SARS-CoV-2 Genome.

RNA structural elements occur in numerous single-stranded positive-sense RNA viruses. The stem-loop 2 motif (s2m) is one such element with an unusually high degree of sequence conservation, being found in the 3' untranslated region (UTR) in the genomes of many astroviruses, some picornaviruses and noroviruses, and a variety of coronaviruses, including severe acute respiratory syndrome coronavirus (SARS-CoV) and SARS-CoV-2. The evolutionary conservation and its occurrence in all viral subgenomic transcripts imply a key role for s2m in the viral infection cycle. Our findings indicate that the element, while stably folded, can nonetheless be invaded and remodeled spontaneously by antisense oligonucleotides (ASOs) that initiate pairing in exposed loops and trigger efficient sequence-specific RNA cleavage in reporter assays. ASOs also act to inhibit replication in an astrovirus replicon model system in a sequence-specific, dose-dependent manner and inhibit SARS-CoV-2 replication in cell culture. Our results thus permit us to suggest that the s2m element is readily targeted by ASOs, which show promise as antiviral agents. IMPORTANCE The highly conserved stem-loop 2 motif (s2m) is found in the genomes of many RNA viruses, including SARS-CoV-2. Our findings indicate that the s2m element can be targeted by antisense oligonucleotides. The antiviral potential of this element represents a promising start for further research into targeting conserved elements in RNA viruses.

CRISPR/Cas9 gene editing for the creation of an MGAT1-deficient CHO cell line to control HIV-1 vaccine glycosylation.

Over the last decade, multiple broadly neutralizing monoclonal antibodies (bN-mAbs) to the HIV-1 envelope protein (Env) gp120 have been described. Many of these recognize epitopes consisting of both amino acid and glycan residues. Moreover, the glycans required for binding of these bN-mAbs are early intermediates in the N-linked glycosylation pathway. This type of glycosylation substantially alters the mass and net charge of Envs compared to molecules with the same amino acid sequence but possessing mature, complex (sialic acid-containing) carbohydrates. Since cell lines suitable for biopharmaceutical production that limit N-linked glycosylation to mannose-5 (Man5) or earlier intermediates are not readily available, the production of vaccine immunogens displaying these glycan-dependent epitopes has been challenging. Here, we report the development of a stable suspension-adapted Chinese hamster ovary (CHO) cell line that limits glycosylation to Man5 and earlier intermediates. This cell line was created using the clustered regularly interspaced short palindromic repeat (CRISPR)/CRISPR-associated protein 9 (Cas9) gene editing system and contains a mutation that inactivates the gene encoding Mannosyl (Alpha-1,3-)-Glycoprotein Beta-1,2-N-Acetylglucosaminyltransferase (MGAT1). Monomeric gp120s produced in the MGAT1- CHO cell line exhibit improved binding to prototypic glycan-dependent bN-mAbs directed to the V1/V2 domain (e.g., PG9) and the V3 stem (e.g., PGT128 and 10-1074) while preserving the structure of the important glycan-independent epitopes (e.g., VRC01). The ability of the MGAT1- CHO cell line to limit glycosylation to early intermediates in the N-linked glycosylation pathway without impairing the doubling time or ability to grow at high cell densities suggests that it will be a useful substrate for the biopharmaceutical production of HIV-1 vaccine immunogens.

Identification and CRISPR/Cas9 Inactivation of the C1s Protease Responsible for Proteolysis of Recombinant Proteins Produced in CHO Cells.

Proteolysis associated with recombinant protein expression in Chinese Hamster Ovary (CHO) cells has hindered the development of biologics including HIV vaccines. When expressed in CHO cells, the recombinant HIV envelope protein, gp120, undergoes proteolytic clipping by a serine protease at a key epitope recognized by neutralizing antibodies. The problem is particularly acute for envelope proteins from clade B viruses that represent the major genetic subtype circulating in much of the developed world, including the US and Europe. In this paper, we have identified complement Component 1's (C1s), a serine protease from the complement cascade, as the protease responsible for the proteolysis of gp120 in CHO cells. CRISPR/Cas9 knockout of the C1s protease in a CHO cell line was shown to eliminate the proteolytic activity against the recombinantly expressed gp120. In addition, the C1s-/- MGAT1- CHO cell line, with the C1s protease and the MGAT1 glycosyltransferase knocked out, enabled the production of unclipped gp120 from a clade B isolate (BaL-rgp120) and enriched for mannose-5 glycans on gp120 that are required for the binding of multiple broadly neutralizing monoclonal antibodies (bN-mAbs). The availability of this technology will allow for the scale-up and testing of multiple vaccine concepts in regions of the world where clade B viruses are in circulation. Furthermore, the proteolysis issues caused by the C1s protease suggests a broader need for a C1s-deficient CHO cell line to express other recombinant proteins that are susceptible to serine protease activity in CHO cells. Similarly, the workflow described here to identify and knockout C1s in a CHO cell line can be applied to remedy the proteolysis of biologics by other CHO proteases.

Pediatric Nasal Reconstruction by Washio Procedure.

The use of the Washio retroauricular flap for nasal reconstruction has been infrequently covered in recent literature, particularly concerning the pediatric population. A retrospective study was conducted between 2014 and 2018 and included all pediatric patients who underwent a Washio retroauricular flap procedure for nasal reconstruction operated on by the same surgeon at a referral center for pediatric plastic and maxillofacial surgery. The mean age at the time of the first stage of the Washio procedure was just under 8 years of age (range: 6 years 3 months-8 years 10 months). The Washio retroauricular flap procedure was successfully employed in three patients with three different anatomical defects, including the nasal alae, nasal tip, and columella, without postoperative healing complications. Arguably, the Washio method is sufficiently versatile to be used in various defect types, allows space and planning for subsequent surgical corrections, avoids additional visible scarring of the face, and spares flaps that may be required at the end of the growth, such as the pedicled forehead flap. It is a safe procedure, provided that at least a two-stage procedure is performed, and a progressive postoperative verticalization is prescribed to limit venous drainage complications.

HIV-1 envelope proteins and V1/V2 domain scaffolds with mannose-5 to improve the magnitude and quality of protective antibody responses to HIV-1.

Two lines of investigation have highlighted the importance of antibodies to the V1/V2 domain of gp120 in providing protection from HIV-1 infection. First, the recent RV144 HIV-1 vaccine trial documented a correlation between non-neutralizing antibodies to the V2 domain and protection. Second, multiple broadly neutralizing monoclonal antibodies to the V1/V2 domain (e.g. PG9) have been isolated from rare infected individuals, termed elite neutralizers. Interestingly, the binding of both types of antibodies appears to depend on the same cluster of amino acids (positions 167­171) adjacent to the junction of the B and C strands of the four-stranded V1/V2 domain ß-sheet structure. However, the broadly neutralizing mAb, PG9, additionally depends on mannose-5 glycans at positions 156 and 160 for binding. Because the gp120 vaccine immunogens used in previous HIV-1 vaccine trials were enriched for complex sialic acid-containing glycans, and lacked the high mannose structures required for the binding of PG9-like mAbs, we wondered if these immunogens could be improved by limiting glycosylation to mannose-5 glycans. Here, we describe the PG9 binding activity of monomeric gp120s from multiple strains of HIV-1 produced with mannose-5 glycans. We also describe the properties of glycopeptide scaffolds from the V1/V2 domain also expressed with mannose-5 glycans. The V1/V2 scaffold from the A244 isolate was able to bind the PG9, CH01, and CH03 mAbs with high affinity provided that the proper glycans were present. We further show that immunization with A244 V1/V2 fragments alone, or in a prime/boost regimen with gp120, enhanced the antibody response to sequences in the V1/V2 domain associated with protection in the RV144 trial.

Discovery of three novel neutralizing antibody epitopes on the human astrovirus capsid spike and mechanistic insights into virus neutralization.

Human astroviruses (HAstVs) are a leading cause of viral childhood diarrhea that infect nearly every individual during their lifetime. Although human astroviruses are highly prevalent, no approved vaccine currently exists. Antibody responses appear to play an important role in protection from HAstV infection, however knowledge about the neutralizing epitope landscape is lacking, as only 3 neutralizing antibody epitopes have previously been determined. Here, we structurally define the epitopes of 3 uncharacterized HAstV-neutralizing monoclonal antibodies: antibody 4B6 with X-ray crystallography to 2.67 Å, and antibodies 3H4 and 3B4 simultaneously with single-particle cryogenic-electron microscopy to 3.33 Å. We assess the epitope locations relative to conserved regions on the capsid spike and find that while antibodies 4B6 and 3B4 target the upper variable loop regions of the HAstV spike protein, antibody 3H4 targets a novel region near the base of the spike that is more conserved. Additionally, we found that all 3 antibodies bind with high affinity, and they compete with receptor FcRn binding to the capsid spike. These studies inform which regions of the HAstV capsid can be targeted by monoclonal antibody therapies and could aid in rational vaccine design. Importance: Human astroviruses infect nearly every child in the world, causing diarrhea, vomiting, and fever. Despite the prevalence of human astroviruses, little is known about how antibodies block virus infection. Here, we determined high-resolution structures of the astrovirus capsid protein in complex with three virus-neutralizing antibodies. The antibodies bind distinct sites on the capsid spike domain. We find that the antibodies block virus attachment to human cells and prevent capsid spike interaction with the human neonatal Fc receptor. These findings support the use of the human astrovirus capsid spike as an antigen in a vaccine to prevent astrovirus disease.

Advancing long-read nanopore genome assembly and accurate variant calling for rare disease detection.

More than 50% of families with suspected rare monogenic diseases remain unsolved after whole genome analysis by short read sequencing (SRS). Long-read sequencing (LRS) could help bridge this diagnostic gap by capturing variants inaccessible to SRS, facilitating long-range mapping and phasing, and providing haplotype-resolved methylation profiling. To evaluate LRS's additional diagnostic yield, we sequenced a rare disease cohort of 98 samples, including 41 probands and some family members, using nanopore sequencing, achieving per sample ∼36x average coverage and 32 kilobase (kb) read N50 from a single flow cell. Our Napu pipeline generated assemblies, phased variants, and methylation calls. LRS covered, on average, coding exons in ∼280 genes and ∼5 known Mendelian disease genes that were not covered by SRS. In comparison to SRS, LRS detected additional rare, functionally annotated variants, including SVs and tandem repeats, and completely phased 87% of protein-coding genes. LRS detected additional de novo variants, and could be used to distinguish postzygotic mosaic variants from prezygotic de novos . Eleven probands were solved, with diverse underlying genetic causes including de novo and compound heterozygous variants, large-scale SVs, and epigenetic modifications. Our study demonstrates LRS's potential to enhance diagnostic yield for rare monogenic diseases, implying utility in future clinical genomics workflows.

Sequences in glycoprotein gp41, the CD4 binding site, and the V2 domain regulate sensitivity and resistance of HIV-1 to broadly neutralizing antibodies.

The swarm of quasispecies that evolves in each HIV-1-infected individual represents a source of closely related Env protein variants that can be used to explore various aspects of HIV-1 biology. In this study, we made use of these variants to identify mutations that confer sensitivity and resistance to the broadly neutralizing antibodies found in the sera of selected HIV-1-infected individuals. For these studies, libraries of Env proteins were cloned from infected subjects and screened for infectivity and neutralization sensitivity. The nucleotide sequences of the Env proteins were then compared for pairs of neutralization-sensitive and -resistant viruses. In vitro mutagenesis was used to identify the specific amino acids responsible for the neutralization phenotype. All of the mutations altering neutralization sensitivity/resistance appeared to induce conformational changes that simultaneously enhanced the exposure of two or more epitopes located in different regions of gp160. These mutations appeared to occur at unique positions required to maintain the quaternary structure of the gp160 trimer, as well as conformational masking of epitopes targeted by neutralizing antibodies. Our results show that sequences in gp41, the CD4 binding site, and the V2 domain all have the ability to act as global regulators of neutralization sensitivity. Our results also suggest that neutralization assays designed to support the development of vaccines and therapeutics targeting the HIV-1 Env protein should consider virus variation within individuals as well as virus variation between individuals.

Pediatric Outpatient Occupational Therapy: Transitioning From In-Person Services to Telehealth.

The COVID-19 pandemic necessitated rapid adoption of telehealth for outpatient pediatric occupational therapy practice. The dose of therapy may have varied across diagnostic and geographical groups despite efforts to ensure access for all patients. The objective of the study was to describe the visit length of outpatient pediatric occupational therapy practice for three diagnostic groups at one institution both during and prior to the COVID-19 pandemic. Retrospective review of electronic health records for two time periods using both practitioner-entered and telecommunications data. Data were analyzed using descriptive statistics and generalized linear mixed model. Prior to the pandemic, average treatment length did not vary by primary diagnosis. During the pandemic, average visit length varied by primary diagnosis, with feeding disorder (FD) visits significantly shorter than cerebral palsy (CP) and autism spectrum disorder (ASD) visits. During the pandemic, visit length was associated with rurality for the whole sample and for patients with ASD and CP, but not FD. Patients with FD may have been seen for shorter durations during telehealth visits. The technology gap may affect services for patients living in rural communities.

Structural insights into the broad protection against H1 influenza viruses by a computationally optimized hemagglutinin vaccine.

Influenza virus poses an ongoing human health threat with pandemic potential. Due to mutations in circulating strains, formulating effective vaccines remains a challenge. The use of computationally optimized broadly reactive antigen (COBRA) hemagglutinin (HA) proteins is a promising vaccine strategy to protect against a wide range of current and future influenza viruses. Though effective in preclinical studies, the mechanistic basis driving the broad reactivity of COBRA proteins remains to be elucidated. Here, we report the crystal structure of the COBRA HA termed P1 and identify antigenic and glycosylation properties that contribute to its immunogenicity. We further report the cryo-EM structure of the P1-elicited broadly neutralizing antibody 1F8 bound to COBRA P1, revealing 1F8 to recognize an atypical receptor binding site epitope via an unexpected mode of binding.

Clinical Focus: Findings and Clinical Implications for Thickening Formula With Infant Cereal Using the International Dysphagia Diet Standardisation Initiative Flow Test.

PURPOSE: The International Dysphagia Diet Standardisation Initiative (IDDSI) framework was established to provide standardized terminology and objective measures to assess foods and liquids for persons with swallowing difficulties. This clinical focus article reports the findings and clinical implications of the flow testing of infant formulas thickened with infant cereal completed as part of the transition process to IDDSI for one large pediatric quaternary care hospital. METHOD: To determine a common recipe that could be used to thicken formulas with infant cereal to the appropriate IDDSI levels, three clinicians completed flow testing on 94 infant formulas. To examine intra- and interclinician variability in the process, they repeated flow testing with three commonly used formulas and infant cereal. RESULTS: Clinicians were unable to identify a standard recipe (infant formula + infant cereal combination) that consistently thickened different formula brands to a desired IDDSI thickness level, as there was pronounced variability across and within infant formulas. Reliability testing revealed that, overall, clinician mixers were consistent in replicating similar results to themselves and to each other and that, instead, greater variability lies within the formula (and infant formula + infant cereal combination). CONCLUSIONS: Based on findings of pronounced variability within and across infant formulas, our institution determined that the creation of a standard recipe for achieving IDDSI thickness levels of formula mixed with infant cereal was not feasible or clinically appropriate. We offer recommendations for similar institutions for advancing clinical management of infant dysphagia using the IDDSI flow test and directions for future research.

Loss to patient-reported outcome measure follow-up after hip arthroplasty and knee arthroplasty : patient satisfaction, associations with non-response, and maximizing returns.

AIMS: The aim of this study was to determine satisfaction rates after hip and knee arthroplasty in patients who did not respond to postoperative patient-reported outcome measures (PROMs), characteristics of non-responders, and contact preferences to maximize response rates. METHODS: A prospective cohort study of patients planned to undergo hip arthroplasty (n = 713) and knee arthroplasty (n = 737) at a UK university teaching hospital who had completed preoperative PROMs questionnaires, including the EuroQol five-dimension health-related quality of life score, and Oxford Hip Score (OHS) and Oxford Knee Score (OKS). Follow-up questionnaires were sent by post at one year, including satisfaction scoring. Attempts were made to contact patients who did not initially respond. Univariate, logistic regression, and receiver operator curve analysis was performed. RESULTS: At one year, 667 hip patients (93.5%) and 685 knee patients (92.9%) had undergone surgery and were alive. No response was received from 151/667 hip patients (22.6%), 83 (55.0%) of whom were ultimately contacted); or from 108/685 knee patients (15.8%), 91 (84.3%) of whom were ultimately contacted. There was no difference in satisfaction after arthroplasty between initial non-responders and responders for hips (74/81 satisfied vs 476/516 satisfied; p = 0.847) or knees (81/93 satisfied vs 470/561 satisfied; p = 0.480). Initial non-response and persistent non-response was associated with younger age, higher BMIs, and worse preoperative PROMs for both hip and knee patients (p < 0.050). Being in employment was associated with persistent non-response for hip patients (p = 0.047). Multivariate analysis demonstrated that younger age (p < 0.038), higher BMI (p = 0.018), and poorer preoperative OHS (p = 0.031) were independently associated with persistent non-response to hip PROMs. No independent associations were identified for knees. Using a threshold of > 66.4 years predicted a preference for contact by post (area under the curve 0.723 (95% confidence interval (CI) 0.647 to 0.799; p < 0.001, though this CI crosses the 0.7 limit considered reliable). CONCLUSION: The majority of initial non-responders were ultimately contactable with effort. Satisfaction rates were not inferior in patients who did not initially respond to PROMs. Cite this article: Bone Jt Open 2022;3(4):275-283.

Protease cleavage sites in HIV-1 gp120 recognized by antigen processing enzymes are conserved and located at receptor binding sites.

The identification of vaccine immunogens able to elicit broadly neutralizing antibodies (bNAbs) is a major goal in HIV vaccine research. Although it has been possible to produce recombinant envelope glycoproteins able to adsorb bNAbs from HIV-positive sera, immunization with these proteins has failed to elicit antibody responses effective against clinical isolates of HIV-1. Thus, the epitopes recognized by bNAbs are present on recombinant proteins, but they are not immunogenic. These results led us to consider the possibility that changes in the pattern of antigen processing might alter the immune response to the envelope glycoprotein to better elicit protective immunity. In these studies, we have defined protease cleavage sites on HIV gp120 recognized by three major human proteases (cathepsins L, S, and D) important for antigen processing and presentation. Remarkably, six of the eight sites identified in gp120 were highly conserved and clustered in regions of the molecule associated with receptor binding and/or the binding of neutralizing antibodies. These results suggested that HIV may have evolved to take advantage of major histocompatibility complex (MHC) class II antigen processing enzymes in order to evade or direct the antiviral immune response.

Mutation at a single position in the V2 domain of the HIV-1 envelope protein confers neutralization sensitivity to a highly neutralization-resistant virus.

Understanding the determinants of neutralization sensitivity and resistance is important for the development of an effective human immunodeficiency virus type 1 (HIV-1) vaccine. In these studies, we have made use of the swarm of closely related envelope protein variants (quasispecies) from an extremely neutralization-resistant clinical isolate in order to identify mutations that conferred neutralization sensitivity to antibodies in sera from HIV-1-infected individuals. Here, we describe a virus with a rare mutation at position 179 in the V2 domain of gp120, where replacement of aspartic acid (D) by asparagine (N) converts a virus that is highly resistant to neutralization by multiple polyclonal and monoclonal antibodies, as well as antiviral entry inhibitors, to one that is sensitive to neutralization. Although the V2 domain sequence is highly variable, D at position 179 is highly conserved in HIV-1 and simian immunodeficiency virus (SIV) and is located within the LDI/V recognition motif of the recently described α4ß7 receptor binding site. Our results suggest that the D179N mutation induces a conformational change that exposes epitopes in both the gp120 and the gp41 portions of the envelope protein, such as the CD4 binding site and the MPER, that are normally concealed by conformational masking. Our results suggest that D179 plays a central role in maintaining the conformation and infectivity of HIV-1 as well as mediating binding to α4ß7.