Germinal center responses to SARS-CoV-2 mRNA vaccines in healthy and immunocompromised individuals.

Vaccine-mediated immunity often relies on the generation of protective antibodies and memory B cells, which commonly stem from germinal center (GC) reactions. An in-depth comparison of the GC responses elicited by SARS-CoV-2 mRNA vaccines in healthy and immunocompromised individuals has not yet been performed due to the challenge of directly probing human lymph nodes. Herein, through a fine-needle aspiration-based approach, we profiled the immune responses to SARS-CoV-2 mRNA vaccines in lymph nodes of healthy individuals and kidney transplant recipients (KTXs). We found that, unlike healthy subjects, KTXs presented deeply blunted SARS-CoV-2-specific GC B cell responses coupled with severely hindered T follicular helper cell, SARS-CoV-2 receptor binding domain-specific memory B cell, and neutralizing antibody responses. KTXs also displayed reduced SARS-CoV-2-specific CD4 and CD8 T cell frequencies. Broadly, these data indicate impaired GC-derived immunity in immunocompromised individuals and suggest a GC origin for certain humoral and memory B cell responses following mRNA vaccination.

Efficient recall of Omicron-reactive B cell memory after a third dose of SARS-CoV-2 mRNA vaccine.

We examined antibody and memory B cell responses longitudinally for ∼9-10 months after primary 2-dose SARS-CoV-2 mRNA vaccination and 3 months after a 3rd dose. Antibody decay stabilized between 6 and 9 months, and antibody quality continued to improve for at least 9 months after 2-dose vaccination. Spike- and RBD-specific memory B cells remained durable over time, and 40%-50% of RBD-specific memory B cells simultaneously bound the Alpha, Beta, Delta, and Omicron variants. Omicron-binding memory B cells were efficiently reactivated by a 3rd dose of wild-type vaccine and correlated with the corresponding increase in neutralizing antibody titers. In contrast, pre-3rd dose antibody titers inversely correlated with the fold-change of antibody boosting, suggesting that high levels of circulating antibodies may limit the added protection afforded by repeat short interval boosting. These data provide insight into the quantity and quality of mRNA-vaccine-induced immunity over time through 3 or more antigen exposures.

Prior vaccination promotes early activation of memory T cells and enhances immune responses during SARS-CoV-2 breakthrough infection.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection of vaccinated individuals is increasingly common but rarely results in severe disease, likely due to the enhanced potency and accelerated kinetics of memory immune responses. However, there have been few opportunities to rigorously study early recall responses during human viral infection. To better understand human immune memory and identify potential mediators of lasting vaccine efficacy, we used high-dimensional flow cytometry and SARS-CoV-2 antigen probes to examine immune responses in longitudinal samples from vaccinated individuals infected during the Omicron wave. These studies revealed heightened spike-specific responses during infection of vaccinated compared to unvaccinated individuals. Spike-specific cluster of differentiation (CD)4 T cells and plasmablasts expanded and CD8 T cells were robustly activated during the first week. In contrast, memory B cell activation, neutralizing antibody production and primary responses to nonspike antigens occurred during the second week. Collectively, these data demonstrate the functionality of vaccine-primed immune memory and highlight memory T cells as rapid responders during SARS-CoV-2 infection.

Rapid induction of antigen-specific CD4+ T cells is associated with coordinated humoral and cellular immunity to SARS-CoV-2 mRNA vaccination.

SARS-CoV-2 mRNA vaccines have shown remarkable clinical efficacy, but questions remain about the nature and kinetics of T cell priming. We performed longitudinal antigen-specific T cell analyses on healthy SARS-CoV-2-naive and recovered individuals prior to and following mRNA prime and boost vaccination. Vaccination induced rapid antigen-specific CD4+ T cell responses in naive subjects after the first dose, whereas CD8+ T cell responses developed gradually and were variable in magnitude. Vaccine-induced Th1 and Tfh cell responses following the first dose correlated with post-boost CD8+ T cells and neutralizing antibodies, respectively. Integrated analysis revealed coordinated immune responses with distinct trajectories in SARS-CoV-2-naive and recovered individuals. Last, whereas booster vaccination improved T cell responses in SARS-CoV-2-naive subjects, the second dose had little effect in SARS-CoV-2-recovered individuals. These findings highlight the role of rapidly primed CD4+ T cells in coordinating responses to the second vaccine dose in SARS-CoV-2-naive individuals.

SARS-CoV-2 mRNA Vaccines Foster Potent Antigen-Specific Germinal Center Responses Associated with Neutralizing Antibody Generation.

The deployment of effective vaccines against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is critical to eradicate the coronavirus disease 2019 (COVID-19) pandemic. Many licensed vaccines confer protection by inducing long-lived plasma cells (LLPCs) and memory B cells (MBCs), cell types canonically generated during germinal center (GC) reactions. Here, we directly compared two vaccine platforms-mRNA vaccines and a recombinant protein formulated with an MF59-like adjuvant-looking for their abilities to quantitatively and qualitatively shape SARS-CoV-2-specific primary GC responses over time. We demonstrated that a single immunization with SARS-CoV-2 mRNA, but not with the recombinant protein vaccine, elicited potent SARS-CoV-2-specific GC B and T follicular helper (Tfh) cell responses as well as LLPCs and MBCs. Importantly, GC responses strongly correlated with neutralizing antibody production. mRNA vaccines more efficiently induced key regulators of the Tfh cell program and influenced the functional properties of Tfh cells. Overall, this study identifies SARS-CoV-2 mRNA vaccines as strong candidates for promoting robust GC-derived immune responses.

PD-1 combination therapy with IL-2 modifies CD8+ T cell exhaustion program.

Combination therapy with PD-1 blockade and IL-2 is highly effective during chronic lymphocytic choriomeningitis virus infection1. Here we examine the underlying basis for this synergy. We show that PD-1 + IL-2 combination therapy, in contrast to PD-1 monotherapy, substantially changes the differentiation program of the PD-1+TCF1+ stem-like CD8+ T cells and results in the generation of transcriptionally and epigenetically distinct effector CD8+ T cells that resemble highly functional effector CD8+ T cells seen after an acute viral infection. The generation of these qualitatively superior CD8+ T cells that mediate viral control underlies the synergy between PD-1 and IL-2. Our results show that the PD-1+TCF1+ stem-like CD8+ T cells, also referred to as precursors of exhausted CD8+ T cells, are not fate-locked into the exhaustion program and their differentiation trajectory can be changed by IL-2 signals. These virus-specific effector CD8+ T cells emerging from the stem-like CD8+ T cells after combination therapy expressed increased levels of the high-affinity IL-2 trimeric (CD25-CD122-CD132) receptor. This was not seen after PD-1 blockade alone. Finally, we show that CD25 engagement with IL-2 has an important role in the observed synergy between IL-2 cytokine and PD-1 blockade. Either blocking CD25 with an antibody or using a mutated version of IL-2 that does not bind to CD25 but still binds to CD122 and CD132 almost completely abrogated the synergistic effects observed after PD-1 + IL-2 combination therapy. There is considerable interest in PD-1 + IL-2 combination therapy for patients with cancer2,3, and our fundamental studies defining the underlying mechanisms of how IL-2 synergizes with PD-1 blockade should inform these human translational studies.

Evaluating large language models on medical, lay-language, and self-reported descriptions of genetic conditions.

Large language models (LLMs) are generating interest in medical settings. For example, LLMs can respond coherently to medical queries by providing plausible differential diagnoses based on clinical notes. However, there are many questions to explore, such as evaluating differences between open- and closed-source LLMs as well as LLM performance on queries from both medical and non-medical users. In this study, we assessed multiple LLMs, including Llama-2-chat, Vicuna, Medllama2, Bard/Gemini, Claude, ChatGPT3.5, and ChatGPT-4, as well as non-LLM approaches (Google search and Phenomizer) regarding their ability to identify genetic conditions from textbook-like clinician questions and their corresponding layperson translations related to 63 genetic conditions. For open-source LLMs, larger models were more accurate than smaller LLMs: 7b, 13b, and larger than 33b parameter models obtained accuracy ranges from 21%-49%, 41%-51%, and 54%-68%, respectively. Closed-source LLMs outperformed open-source LLMs, with ChatGPT-4 performing best (89%-90%). Three of 11 LLMs and Google search had significant performance gaps between clinician and layperson prompts. We also evaluated how in-context prompting and keyword removal affected open-source LLM performance. Models were provided with 2 types of in-context prompts: list-type prompts, which improved LLM performance, and definition-type prompts, which did not. We further analyzed removal of rare terms from descriptions, which decreased accuracy for 5 of 7 evaluated LLMs. Finally, we observed much lower performance with real individuals' descriptions; LLMs answered these questions with a maximum 21% accuracy.

Tet2 deletion in CD4+ T cells disrupts Th1 lineage commitment in memory cells and enhances T follicular helper cell recall responses to viral rechallenge.

Following viral clearance, antigen-specific CD4+ T cells contract and form a pool of distinct Th1 and Tfh memory cells that possess unique epigenetic programs, allowing them to rapidly recall their specific effector functions upon rechallenge. DNA methylation programing mediated by the methylcytosine dioxygenase Tet2 contributes to balancing Th1 and Tfh cell differentiation during acute viral infection; however, the role of Tet2 in CD4+ T cell memory formation and recall is unclear. Using adoptive transfer models of antigen-specific wild type and Tet2 knockout CD4+ T cells, we find that Tet2 is required for full commitment of CD4+ T cells to the Th1 lineage and that in the absence of Tet2, memory cells preferentially recall a Tfh like phenotype with enhanced expansion upon secondary challenge. These findings demonstrate an important role for Tet2 in enforcing lineage commitment and programing proliferation potential, and highlight the potential of targeting epigenetic programing to enhance adaptive immune responses.

Approximating facial expression effects on diagnostic accuracy via generative AI in medical genetics.

Artificial intelligence (AI) is increasingly used in genomics research and practice, and generative AI has garnered significant recent attention. In clinical applications of generative AI, aspects of the underlying datasets can impact results, and confounders should be studied and mitigated. One example involves the facial expressions of people with genetic conditions. Stereotypically, Williams (WS) and Angelman (AS) syndromes are associated with a "happy" demeanor, including a smiling expression. Clinical geneticists may be more likely to identify these conditions in images of smiling individuals. To study the impact of facial expression, we analyzed publicly available facial images of approximately 3500 individuals with genetic conditions. Using a deep learning (DL) image classifier, we found that WS and AS images with non-smiling expressions had significantly lower prediction probabilities for the correct syndrome labels than those with smiling expressions. This was not seen for 22q11.2 deletion and Noonan syndromes, which are not associated with a smiling expression. To further explore the effect of facial expressions, we computationally altered the facial expressions for these images. We trained HyperStyle, a GAN-inversion technique compatible with StyleGAN2, to determine the vector representations of our images. Then, following the concept of InterfaceGAN, we edited these vectors to recreate the original images in a phenotypically accurate way but with a different facial expression. Through online surveys and an eye-tracking experiment, we examined how altered facial expressions affect the performance of human experts. We overall found that facial expression is associated with diagnostic accuracy variably in different genetic conditions.

Innate lymphoid cells support regulatory T cells in the intestine through interleukin-2.

Interleukin (IL)-2 is a pleiotropic cytokine that is necessary to prevent chronic inflammation in the gastrointestinal tract1-4. The protective effects of IL-2 involve the generation, maintenance and function of regulatory T (Treg) cells4-8, and the use of low doses of IL-2 has emerged as a potential therapeutic strategy for patients with inflammatory bowel disease9. However, the cellular and molecular pathways that control the production of IL-2 in the context of intestinal health are undefined. Here we show, in a mouse model, that IL-2 is acutely required to maintain Treg cells and immunological homeostasis throughout the gastrointestinal tract. Notably, lineage-specific deletion of IL-2 in T cells did not reduce Treg cells in the small intestine. Unbiased analyses revealed that, in the small intestine, group-3 innate lymphoid cells (ILC3s) are the dominant cellular source of IL-2, which is induced selectively by IL-1ß. Macrophages in the small intestine produce IL-1ß, and activation of this pathway involves MYD88- and NOD2-dependent sensing of the microbiota. Our loss-of-function studies show that ILC3-derived IL-2 is essential for maintaining Treg cells, immunological homeostasis and oral tolerance to dietary antigens in the small intestine. Furthermore, production of IL-2 by ILC3s was significantly reduced in the small intestine of patients with Crohn's disease, and this correlated with lower frequencies of Treg cells. Our results reveal a previously unappreciated pathway in which a microbiota- and IL-1ß-dependent axis promotes the production of IL-2 by ILC3s to orchestrate immune regulation in the intestine.

The unstructured linker of Mlh1 contains a motif required for endonuclease function which is mutated in cancers.

Eukaryotic DNA mismatch repair (MMR) depends on recruitment of the Mlh1-Pms1 endonuclease (human MLH1-PMS2) to mispaired DNA. Both Mlh1 and Pms1 contain a long unstructured linker that connects the N- and carboxyl-terminal domains. Here, we demonstrated the Mlh1 linker contains a conserved motif (Saccharomyces cerevisiae residues 391-415) required for MMR. The Mlh1-R401A,D403A-Pms1 linker motif mutant protein was defective for MMR and endonuclease activity in vitro, even though the conserved motif could be >750 Å from the carboxyl-terminal endonuclease active site or the N-terminal adenosine triphosphate (ATP)-binding site. Peptides encoding this motif inhibited wild-type Mlh1-Pms1 endonuclease activity. The motif functioned in vivo at different sites within the Mlh1 linker and within the Pms1 linker. Motif mutations in human cancers caused a loss-of-function phenotype when modeled in S. cerevisiae. These results suggest that the Mlh1 motif promotes the PCNA-activated endonuclease activity of Mlh1-Pms1 via interactions with DNA, PCNA, RFC, or other domains of the Mlh1-Pms1 complex.

Immunologic Predictors of Vaccine Responsiveness in Patients With Lymphoma and Chronic Lymphocytic Leukemia.

Patients with B-cell lymphomas have altered cellular components of vaccine responses due to malignancy and therapy, and the optimal timing of vaccination relative to therapy remains unknown. Severe acute respiratory syndrome coronavirus 2 vaccines created an opportunity for new insights in vaccine timing because patients were challenged with a novel antigen across multiple phases of treatment. We studied serologic messenger RNA vaccine response in retrospective and prospective cohorts with lymphoma and chronic lymphocytic leukemia, paired with clinical and research immune parameters. Reduced serologic response was observed more frequently during active treatment, but nonresponse was also common within observation and posttreatment groups. Total immunoglobulin A and immunoglobulin M correlated with successful vaccine response. In individuals treated with anti-CD19-directed chimeric antigen receptor-modified T cells, nonresponse was associated with reduced B and T follicular helper cells. Predictors of vaccine response varied by disease and therapeutic group, and therefore further studies of immune health during and after cancer therapies are needed to individualize vaccine timing.

Cross-Platform Synaptic Network Analysis of Human Entorhinal Cortex Identifies TWF2 as a Modulator of Dendritic Spine Length.

Proteomic studies using postmortem human brain tissue samples have yielded robust assessments of the aging and neurodegenerative disease(s) proteomes. While these analyses provide lists of molecular alterations in human conditions, like Alzheimer's disease (AD), identifying individual proteins that affect biological processes remains a challenge. To complicate matters, protein targets may be highly understudied and have limited information on their function. To address these hurdles, we sought to establish a blueprint to aid selection and functional validation of targets from proteomic datasets. A cross-platform pipeline was engineered to focus on synaptic processes in the entorhinal cortex (EC) of human patients, including controls, preclinical AD, and AD cases. Label-free quantification mass spectrometry (MS) data (n = 2260 proteins) was generated on synaptosome fractionated tissue from Brodmann area 28 (BA28; n = 58 samples). In parallel, dendritic spine density and morphology was measured in the same individuals. Weighted gene co-expression network analysis was used to construct a network of protein co-expression modules that were correlated with dendritic spine metrics. Module-trait correlations were used to guide unbiased selection of Twinfilin-2 (TWF2), which was the top hub protein of a module that positively correlated with thin spine length. Using CRISPR-dCas9 activation strategies, we demonstrated that boosting endogenous TWF2 protein levels in primary hippocampal neurons increased thin spine length, thus providing experimental validation for the human network analysis. Collectively, this study describes alterations in dendritic spine density and morphology as well as synaptic proteins and phosphorylated tau from the entorhinal cortex of preclinical and advanced stage AD patients.SIGNIFICANCE STATEMENT Proteomic studies can yield vast lists of molecules that are altered under various experimental or disease conditions. Here, we provide a blueprint to facilitate mechanistic validation of protein targets from human brain proteomic datasets. We conducted a proteomic analysis of human entorhinal cortex (EC) samples spanning cognitively normal and Alzheimer's disease (AD) cases with a comparison of dendritic spine morphology in the same samples. Network integration of proteomics with dendritic spine measurements allowed for unbiased discovery of Twinfilin-2 (TWF2) as a regulator of dendritic spine length. A proof-of-concept experiment in cultured neurons demonstrated that altering Twinfilin-2 protein level induced corresponding changes in dendritic spine length, thus providing experimental validation for the computational framework.

Effect of capacity building interventions on classroom teacher and early childhood educator perceived capabilities, knowledge, and attitudes relating to physical activity and fundamental movement skills: a systematic review and meta-analysis.

BACKGROUND: Capacity building may play an important role in improving classroom teachers' and early childhood educators' (ECE) capacity to implement physical activity and FMS interventions. Capacity building is the development of knowledge, skills, and structures to improve the capability of individuals and organisations to achieve effective health promotion. This review aimed to determine the efficacy of capacity building interventions on teachers' and ECEs' perceived capabilities, knowledge, and attitudes relating to physical activity and fundamental movement skills. METHODS: An exhaustive literature search of six electronic databases was conducted. Controlled, single-group pre-post studies were included if they measured the effect of a capacity building intervention on in-service or pre-service classroom teachers' (primary or secondary) or ECEs' physical activity or fundamental skills related perceived capabilities, knowledge, or attitudes. The effects of interventions were synthesised using random effects meta-analysis. Subgroup analysis and meta-regression was conducted to determine if the effects differed based on study design, type of teacher (ECE vs. primary school), or teacher level (pre-service vs. in-service). RESULTS: A total of 22 studies reporting on 25 unique samples were included in the meta-analyses. Only studies reporting on ECEs and primary school teachers were identified. Interventions most commonly included training/professional development, resources and toolkits, communities of practice, mentorships, and ongoing support. Results showed that capacity building interventions significantly improved teachers' and ECEs' perceived capabilities (g = 0.614, 95% CI = 0.442, 0.786), knowledge (g = 0.792 95% CI = 0.459, 1.125), and attitudes (g = 0.376 95% CI = 0.181, 0.571). The effects did not differ significantly as a function of any of the moderators examined. CONCLUSION: Findings from this review provide strong support that capacity building interventions are efficacious at improving teachers' and ECEs' perceived capabilities, knowledge, and attitudes related to promoting physical activity and teaching fundamental movement skills. Pre-service teachers and ECEs should be provided training in physical activity and fundamental movement skills as part of their degrees, and continual professional development and capacity building should be offered to in-service teachers and ECEs to promote physical activity and fundamental movement skills in children.

Visual Assessment of Phony Peach Disease: Evaluating Rater Accuracy and Reliability.

Phony peach disease (PPD), found predominantly in central and southern Georgia, is a re-emerging disease caused by Xylella fastidiosa (Xf) subsp. multiplex. Accurate detection and rapid removal of symptomatic trees are crucial to effective disease management. Currently, peach producers rely solely on visual identification of symptoms to confirm PPD, which can be ambiguous if early in development. We compared visual assessment to quantitative PCR (qPCR) for detecting Xf in 'Julyprince' in 2019 and 2020 (JP2019 and JP2020) and in 'Scarletprince' in 2020 (SP2020). With no prior knowledge of qPCR results, all trees in each orchard were assessed by a cohort of five experienced and five inexperienced raters in the morning and afternoon. Visual identification accuracy of PPD was variable, but experienced raters were more accurate when identifying PPD trees. In JP2019, the mean rater accuracy for experienced and inexperienced raters was 0.882 and 0.805, respectively. For JP2020, the mean rater accuracy for experienced and inexperienced raters was 0.914 and 0.816, respectively. For SP2020, the mean rater accuracy for experienced and inexperienced raters was 0.898 and 0.807, respectively. All raters had false positive (FP) and false negative (FN) observations, but experienced raters had significantly lower FN rates compared with the inexperienced group. Almost all raters overestimated the incidence of PPD in the orchards. Reliability of visual assessments was demonstrated as moderate to good, regardless of experience. Further research is needed to develop accurate and reliable methods of detection to aid management of PPD as both FPs and FNs are costly to peach production.

Scaling a common assessment of associative ability: Development and validation of a multiple-choice compound remote associates task.

The assessment of creativity as an individual difference has historically focused on divergent thinking, which is increasingly viewed as involving the associative processes that are also understood to be a key component of creative potential. Research on associative processes has proliferated in many sub-fields, often using Compound Remote Associates (CRA) tasks with an open response format and relatively small participant samples. In the present work, we introduce a new format that is more amenable to large-scale data collection in survey designs, and present evidence for the reliability and validity of CRA measures in general using multiple large samples. Study 1 uses a large, representative dataset (N = 1,323,480) to demonstrate strong unidimensionality and internal consistency (α = .97; ωt = .87), as well as links to individual differences in temperament, cognitive ability, occupation, and job characteristics. Study 2 uses an undergraduate sample (N = 685) to validate the use of a multiple-choice format relative to the traditional approach. Study 3 uses a crowdsourced sample (N = 357) to demonstrate high test-retest reliability of the items (r =.74). Finally, Study 4 uses a sample that overlaps with Study 1 (N = 1,502,922) to provide item response theory (IRT) parameters for a large set of high-quality CRA items that use a multiple-choice response mode, thus facilitating their use in future research on creativity, insight, and related topics.

Demand creation for HIV testing services: A systematic review and meta-analysis.

BACKGROUND: HIV testing services (HTS) are the first steps in reaching the UNAIDS 95-95-95 goals to achieve and maintain low HIV incidence. Evaluating the effectiveness of different demand creation interventions to increase uptake of efficient and effective HTS is useful to prioritize limited programmatic resources. This review was undertaken to inform World Health Organization (WHO) 2019 HIV testing guidelines and assessed the research question, "Which demand creation strategies are effective for enhancing uptake of HTS?" focused on populations globally. METHODS AND FINDINGS: The following electronic databases were searched through September 28, 2021: PubMed, PsycInfo, Cochrane CENTRAL, CINAHL Complete, Web of Science Core Collection, EMBASE, and Global Health Database; we searched IAS and AIDS conferences. We systematically searched for randomized controlled trials (RCTs) that compared any demand creation intervention (incentives, mobilization, counseling, tailoring, and digital interventions) to either a control or other demand creation intervention and reported HTS uptake. We pooled trials to evaluate categories of demand creation interventions using random-effects models for meta-analysis and assessed study quality with Cochrane's risk of bias 1 tool. This study was funded by the WHO and registered in Prospero with ID CRD42022296947. We screened 10,583 records and 507 conference abstracts, reviewed 952 full texts, and included 124 RCTs for data extraction. The majority of studies were from the African (N = 53) and Americas (N = 54) regions. We found that mobilization (relative risk [RR]: 2.01, 95% confidence interval [CI]: [1.30, 3.09], p < 0.05; risk difference [RD]: 0.29, 95% CI [0.16, 0.43], p < 0.05, N = 4 RCTs), couple-oriented counseling (RR: 1.98, 95% CI [1.02, 3.86], p < 0.05; RD: 0.12, 95% CI [0.03, 0.21], p < 0.05, N = 4 RCTs), peer-led interventions (RR: 1.57, 95% CI [1.15, 2.15], p < 0.05; RD: 0.18, 95% CI [0.06, 0.31], p < 0.05, N = 10 RCTs), motivation-oriented counseling (RR: 1.53, 95% CI [1.07, 2.20], p < 0.05; RD: 0.17, 95% CI [0.00, 0.34], p < 0.05, N = 4 RCTs), short message service (SMS) (RR: 1.53, 95% CI [1.09, 2.16], p < 0.05; RD: 0.11, 95% CI [0.03, 0.19], p < 0.05, N = 5 RCTs), and conditional fixed value incentives (RR: 1.52, 95% CI [1.21, 1.91], p < 0.05; RD: 0.15, 95% CI [0.07, 0.22], p < 0.05, N = 11 RCTs) all significantly and importantly (≥50% relative increase) increased HTS uptake and had medium risk of bias. Lottery-based incentives and audio-based interventions less importantly (25% to 49% increase) but not significantly increased HTS uptake (medium risk of bias). Personal invitation letters and personalized message content significantly but not importantly (<25% increase) increased HTS uptake (medium risk of bias). Reduced duration counseling had comparable performance to standard duration counseling (low risk of bias) and video-based interventions were comparable or better than in-person counseling (medium risk of bias). Heterogeneity of effect among pooled studies was high. This study was limited in that we restricted to randomized trials, which may be systematically less readily available for key populations; additionally, we compare only pooled estimates for interventions with multiple studies rather than single study estimates, and there was evidence of publication bias for several interventions. CONCLUSIONS: Mobilization, couple- and motivation-oriented counseling, peer-led interventions, conditional fixed value incentives, and SMS are high-impact demand creation interventions and should be prioritized for programmatic consideration. Reduced duration counseling and video-based interventions are an efficient and effective alternative to address staffing shortages. Investment in demand creation activities should prioritize those with undiagnosed HIV or ongoing HIV exposure. Selection of demand creation interventions must consider risks and benefits, context-specific factors, feasibility and sustainability, country ownership, and universal health coverage across disease areas.

Online protein digestion in membranes between capillary electrophoresis and mass spectrometry.

This research employs pepsin-containing membranes to digest proteins online after a capillary electrophoresis (CE) separation and prior to tandem mass spectrometry. Proteolysis after the separation allows the peptides from a given protein to enter the mass spectrometer in a single plug. Thus, migration time can serve as an additional criterion for confirming the identification of a peptide. The membrane resides in a sheath-flow electrospray ionization (ESI) source to enable digestion immediately before spray into the mass spectrometer, thus limiting separation of the digested peptides. Using the same membrane, digestion occurred reproducibly during 20 consecutive CE analyses performed over a 10 h period. Additionally, after separating a mixture of six unreduced proteins with CE, online digestion facilitated protein identification with at least 2 identifiable peptides for all the proteins. Sequence coverages were >75% for myoglobin and carbonic anhydrase II but much lower for proteins containing disulfide bonds. Development of methods for efficient separation of reduced proteins or identification of cross-linked peptides should enhance sequence coverages for proteins with disulfide bonds. Migration times for the peptides identified from a specific protein differed by <∼30 s, which allows for rejection of some spurious peptide identifications.

Expert peer facilitation of the EVERYbody Project: A randomized-controlled evaluation of a diversity-focused, dissonance-based, universal body image program for college students.

OBJECTIVE: A novel peer facilitation model was used to deliver a two session, dissonance-based, inclusive body image intervention that critically examines how internalized size-based oppression intersects with race, class, gender, sexuality, and ability. METHOD: The EVERYbody Project was open to all college students and delivered by "expert" peer facilitators with body image and diversity experience and advanced facilitation skills. Recruitment was halted due to COVID-19; 90 students in the Northwest US (M age = 19.83 years, SD = 2.38; 80% female-identified, 13% male-identified, 7% gender expansive) were randomized to receive the EVERYbody Project or a video-based, low-dissonance comparison intervention. RESULTS: Around half of students (56%) held one or more specific socially marginalized identity (26% with a racial or ethnic identity other than white and non-Hispanic, 39% with a sexual identity other than straight, 7% with a gender identity other than cisgender). The EVERYbody Project produced greater reductions in three outcomes associated with poor body image (internalized appearance norms, body dissatisfaction, and negative affect) compared to the video intervention through 3-month follow-up (ps < .003) with medium between group effects. Both conditions experienced small reductions in eating disorder psychopathology over time. DISCUSSION: Expert peer facilitation may be a viable delivery model for inclusive, diversity-focused dissonance-based body image programs. PUBLIC SIGNIFICANCE: This study explored a novel facilitation approach for a diversity-focused body image program for college students. A brief (4 h) dissonance-based program was open to all college students and delivered by "expert" diverse peer leaders who were screened for facilitation readiness. Body image and related outcomes were improved through 3-month follow-up relative to a comparison condition, suggesting that expert peer facilitation may be a viable option for delivering universal, inclusive body image programming.

Prevalence and Distribution of Phony Peach Disease (Caused by Xylella fastidiosa) in the United States.

Peach is an important specialty fruit crop in the United States, and phony peach disease (PPD), caused by Xylella fastidiosa subsp. multiplex, has been a major cause of yield loss since it was first observed in 1885. Under a federal eradication program, surveys of PPD were conducted from 1929 to 1972, when the program was terminated. No surveys have been conducted in approximately 50 years; therefore, the current prevalence of PPD in the United States is unknown, especially in the Southeast, where damage was previously most severe. To ascertain the status of PPD, we surveyed orchards in Alabama, Florida, Georgia, and South Carolina from June to August 2020 and, except for South Carolina and northern Georgia, PPD was prevalent. Trees in 17 orchards were subjected to confirmation of X. fastidiosa using the AmplifyRP XRT+ for X. fastidiosa to corroborate our visual assessments; based on these tests, PPD incidence in the orchards ranged from 0 to 30.5%. Ancillary written surveys of relative PPD presence and prevalence were sent to fruit pathologists from universities in 20 states where PPD was historically reported. Only 35.0% of respondents reported that PPD either currently or recently occurred in their state and, of these, three reported PPD to be of significant concern. The results of the physical and written surveys indicate that PPD remains prevalent mainly in the southeastern region of the United States but, in other states where previously reported, it is either not present or has very low prevalence when compared with historical accounts of the disease.