Enhancing Vascular Health and Lowering Blood Pressure in Spontaneously Hypertensive Rats through Syrah Grape (Vitis vinifera) Pomace: The Role of Phenolic Compounds.

BACKGROUND: The beneficial properties of wine by-products include actions that help prevent and treat cardiovascular conditions such as hypertension, primarily due to their antioxidant effects. Novel pharmacotherapies are being developed to treat arterial hypertension, including investigations into natural products exhibiting biological activity, necessitating rigorous evaluation of their efficacy and safety. This study aimed to identify and quantify phenolic compounds in Syrah (Vitis vinifera) grapes grown in the Brazilian Cerrado and their presence in winemaking by-products. It also examined the effects of grape pomace on blood pressure. METHODS: Fresh grapes, pomace, and lees, were subjected to spectrophotometric determination of total phenolic compounds, followed by identification and quantification using HPLC-DAD-ESI-MSn. Normotensive male rats (Wistar) and spontaneously hypertensive rats (SHR) received grape pomace-enriched (150 or 300 mg/kg/day, 14 days) or standard chow. Indirect arterial pressure was assessed, while vascular reactivity was evaluated in mesenteric resistance arteries. RESULTS: Pomace samples exhibited higher total phenolic compound concentrations than grapes or lees. Seven derivatives of hydroxycinnamic acids and twenty-one flavonols were identified. Quercetin-3-glucoside and ethyl caffeate were the most abundant phenolic compounds. Grape pomace-enriched chow demonstrated a dose-dependent hypotensive effect in rats. CONCLUSION: the abundance of flavonols and hydroxycinnamic acids, combined with their hypotensive effects, underscores the therapeutic potential of fine wine-making by-products produced in the Brazilian Cerrado.

HIV/HBV coinfection remodels the immune landscape and natural killer cell ADCC functional responses.

BACKGROUND AND AIMS: HBV and HIV coinfection is a common occurrence globally, with significant morbidity and mortality. Both viruses lead to immune dysregulation including changes in natural killer (NK) cells, a key component of antiviral defense and a promising target for HBV cure strategies. Here we used high-throughput single-cell analysis to explore the immune cell landscape in people with HBV mono-infection and HIV/HBV coinfection, on antiviral therapy, with emphasis on identifying the distinctive characteristics of NK cell subsets that can be therapeutically harnessed. APPROACH AND RESULTS: Our data show striking differences in the transcriptional programs of NK cells. HIV/HBV coinfection was characterized by an over-representation of adaptive, KLRC2 -expressing NK cells, including a higher abundance of a chemokine-enriched ( CCL3/CCL4 ) adaptive cluster. The NK cell remodeling in HIV/HBV coinfection was reflected in enriched activation pathways, including CD3ζ phosphorylation and ZAP-70 translocation that can mediate stronger antibody-dependent cellular cytotoxicity responses and a bias toward chemokine/cytokine signaling. By contrast, HBV mono-infection imposed a stronger cytotoxic profile on NK cells and a more prominent signature of "exhaustion" with higher circulating levels of HBsAg. Phenotypic alterations in the NK cell pool in coinfection were consistent with increased "adaptiveness" and better capacity for antibody-dependent cellular cytotoxicity compared to HBV mono-infection. Overall, an adaptive NK cell signature correlated inversely with circulating levels of HBsAg and HBV-RNA in our cohort. CONCLUSIONS: This study provides new insights into the differential signature and functional profile of NK cells in HBV and HIV/HBV coinfection, highlighting pathways that can be manipulated to tailor NK cell-focused approaches to advance HBV cure strategies in different patient groups.

A pan-influenza monoclonal antibody neutralizes H5 strains and prophylactically protects through intranasal administration.

Avian A(H5N1) influenza virus poses an elevated zoonotic threat to humans, and no pharmacological products are currently registered for fast-acting pre-exposure protection in case of spillover leading to a pandemic. Here, we show that an epitope on the stem domain of H5 hemagglutinin is highly conserved and that the human monoclonal antibody CR9114, targeting that epitope, potently neutralizes all pseudotyped H5 viruses tested, even in the rare case of substitutions in its epitope. Further, intranasal administration of CR9114 fully protects mice against A(H5N1) infection at low dosages, irrespective of pre-existing immunity conferred by the quadrivalent seasonal influenza vaccine. These data provide a proof-of-concept for broad, pre-exposure protection against a potential future pandemic using the intranasal administration route. Studies in humans should assess if autonomous administration of a broadly-neutralizing monoclonal antibody is safe and effective and can thus contribute to pandemic preparedness.

Serological evidence of virus infection in Eidolon helvum fruit bats: implications for bushmeat consumption in Nigeria.

Introduction: The Eidolon helvum fruit bat is one of the most widely distributed fruit bats in Africa and known to be a reservoir for several pathogenic viruses that can cause disease in animals and humans. To assess the risk of zoonotic spillover, we conducted a serological survey of 304 serum samples from E. helvum bats that were captured for human consumption in Makurdi, Nigeria. Methods: Using pseudotyped viruses, we screened 304 serum samples for neutralizing antibodies against viruses from the Coronaviridae, Filoviridae, Orthomyxoviridae and Paramyxoviridae families. Results: We report the presence of neutralizing antibodies against henipavirus lineage GH-M74a virus (odds ratio 6.23; p < 0.001), Nipah virus (odds ratio 4.04; p = 0.00031), bat influenza H17N10 virus (odds ratio 7.25; p < 0.001) and no significant association with Ebola virus (odds ratio 0.56; p = 0.375) in this bat cohort. Conclusion: The data suggest a potential risk of zoonotic spillover including the possible circulation of highly pathogenic viruses in E. helvum populations. These findings highlight the importance of maintaining sero-surveillance of E. helvum, and the necessity for further, more comprehensive investigations to monitor changes in virus prevalence, distribution over time, and across different geographic locations.

Natural killer cell responses during SARS-CoV-2 infection and vaccination in people living with HIV-1.

Natural killer (NK) cell subsets with adaptive properties are emerging as regulators of vaccine-induced T and B cell responses and are specialized towards antibody-dependent functions contributing to SARS-CoV-2 control. Although HIV-1 infection is known to affect the NK cell pool, the additional impact of SARS-CoV-2 infection and/or vaccination on NK cell responses in people living with HIV (PLWH) has remained unexplored. Our data show that SARS-CoV-2 infection skews NK cells towards a more differentiated/adaptive CD57+FcεRIγ- phenotype in PLWH. A similar subset was induced following vaccination in SARS-CoV-2 naïve PLWH in addition to a CD56bright population with cytotoxic potential. Antibody-dependent NK cell function showed robust and durable responses to Spike up to 148 days post-infection, with responses enriched in adaptive NK cells. NK cell responses were further boosted by the first vaccine dose in SARS-CoV-2 exposed individuals and peaked after the second dose in SARS-CoV-2 naïve PLWH. The presence of adaptive NK cells associated with the magnitude of cellular and humoral responses. These data suggest that features of adaptive NK cells can be effectively engaged to complement and boost vaccine-induced adaptive immunity in potentially more vulnerable groups such as PLWH.

Working conditions of nursing professionals in the context of COVID-19 / Condições de trabalho dos profissionais de enfermagem no contexto da COVID-19

Introduction: the COVID-19 pandemic has increased the number of hospitalizations and is responsible for increasing the workload of nursing professionals with a deficiency in human resources and personal protective equipment.Objective: to analyze aspects related to the working conditions of Nursing professionals in the context of COVID-19.Methods: documentary research recorded under narratives in the databases of the Federal Nursing Council, the Regional Nursing Councils of Espírito Santo and Pernambuco, the Federal Public University and Social Media.Results: are presented and analyzed according to the categories: a) nursing work conditions, threatening the worker's health and life; b) the autonomy of nurses in the full exercise of their profession and the cultural aspect of the dominant ideology; c) the Federal Nursing Council and the Regional Nursing Councils of Espírito Santo and Pernambuco as disciplining, normalizing, managing and controlling bodies for the professional practice of nursing.Conclusion: the narratives found in this study demonstrate the precarious working conditions, exacerbated by the pandemic, and the role of nursing in coping with COVID-19

Introdução: a pandemia pela COVID-19 tem elevado o número de internações hospitalares sendo responsável pelo aumento da carga de trabalho dos profissionais de enfermagem com deficiência de recursos humanos e de equipamentos de proteção individual. Objetivo: analisar aspectos relativos às condições de trabalho dos profissionais da Enfermagem no contexto da COVID-19. Método: pesquisa documental registrada sob narrativas nas bases de dados do Conselho Federal de Enfermagem, dos Conselhos Regionais de Enfermagem do Espírito Santo e de Pernambuco, de Universidade Pública Federal e das Mídias Sociais. Resultados: são apresentados e analisados mediante as categorias: a) as condições de trabalho da enfermagem, ameaçando a saúde e vida do trabalhador; b) a autonomia do enfermeiro/a no exercício pleno de sua profissão e aspecto cultural da ideologia dominante; c) o Conselho Federal de Enfermagem e os Conselhos Regionais de Enfermagem do Espírito Santo e de Pernambuco como órgãos disciplinadores, normalizadores, gestores e de controle do exercício profissional da enfermagem. Conclusão: as narrativas encontradas nesse estudo demonstraram as precárias condições de trabalho, agudizadas pela pandemia, e o protagonismo da enfermagem no enfrentamento da COVID-19.

Comparison of the Permeability of Potential Ballast Rocks from Northern Rio de Janeiro State under Different Fouling Rates after Sodium Sulfate Attack.

This paper aimed to analyze the reduction in the ballast layer permeability simulated in a laboratory in saturated conditions by the presence of rock dust as a contaminant of three types of rocks explored in different deposits in the northern region of the state of Rio de Janeiro, Brazil, through laboratory testing relating the physical properties of rock particles before and after sodium sulfate attack. Sodium sulfate attack is justified by the proximity of some sections of the planned EF-118 Vitória-Rio railway line to the coast and of the sulfated water table to the ballast bed, which could degrade the material used and compromise the railway track. Granulometry and permeability tests were performed to compare ballast samples with fouling rates of 0, 10, 20, and 40% rock dust by volume. A constant head permeameter was used to analyze hydraulic conductivity and establish correlations between the petrography and mercury intrusion porosimetry of the rocks, namely two types of metagranite (Mg1 and Mg3) and a gneisse (Gn2). Rocks, such as Mg1 and Mg3, with a larger composition of minerals susceptible to weathering according to petrography analyses, tend to be more sensitive to weathering tests. This, in conjunction with the climate in the region studied, with average annual temperature and rainfall of 27 °C and 1200 mm, could compromise track safety and user comfort. Additionally, the Mg1 and Mg3 samples showed greater percentage variation in wear after the Micro-Deval test, which could damage the ballast due to the considerable changeability of the material. The mass loss caused by abrasion due to the passage of rail vehicles was assessed by the Micro-Deval test, with Mg3 (intact rock) declining from 8.50 ± 1.5 to 11.04 ± 0.5% after chemical attack. However, Gn2, which exhibited the greatest mass loss among the samples, showed no significant variation in average wear, and its mineralogical characteristics remained almost unchanged after 60 sodium sulfate cycles. These aspects, combined with its satisfactory hydraulic conductivity rate, indicate that Gn2 is suitable for use as railway ballast in the EF-118 railway line.

Novel intranasal vaccine targeting SARS-CoV-2 receptor binding domain to mucosal microfold cells and adjuvanted with TLR3 agonist Riboxxim™ elicits strong antibody and T-cell responses in mice.

SARS-CoV-2 continues to circulate in the human population necessitating regular booster immunization for its long-term control. Ideally, vaccines should ideally not only protect against symptomatic disease, but also prevent transmission via asymptomatic shedding and cover existing and future variants of the virus. This may ultimately only be possible through induction of potent and long-lasting immune responses in the nasopharyngeal tract, the initial entry site of SARS-CoV-2. To this end, we have designed a vaccine based on recombinantly expressed receptor binding domain (RBD) of SARS-CoV-2, fused to the C-terminus of C. perfringens enterotoxin, which is known to target Claudin-4, a matrix molecule highly expressed on mucosal microfold (M) cells of the nasal and bronchial-associated lymphoid tissues. To further enhance immune responses, the vaccine was adjuvanted with a novel toll-like receptor 3/RIG-I agonist (Riboxxim™), consisting of synthetic short double stranded RNA. Intranasal prime-boost immunization of mice induced robust mucosal and systemic anti-SARS-CoV-2 neutralizing antibody responses against SARS-CoV-2 strains Wuhan-Hu-1, and several variants (B.1.351/beta, B.1.1.7/alpha, B.1.617.2/delta), as well as systemic T-cell responses. A combination vaccine with M-cell targeted recombinant HA1 from an H1N1 G4 influenza strain also induced mucosal and systemic antibodies against influenza. Taken together, the data show that development of an intranasal SARS-CoV-2 vaccine based on recombinant RBD adjuvanted with a TLR3 agonist is feasible, also as a combination vaccine against influenza.

Pseudotyped Viruses for Influenza.

We have developed an influenza hemagglutinin (HA) pseudotype (PV) library encompassing all influenza A (IAV) subtypes from HA1-HA18, influenza B (IBV) subtypes (both lineages), representative influenza C (ICV), and influenza D (IDV) viruses. These influenza HA (or hemagglutinin-esterase fusion (HEF) for ICV and IDV) pseudotypes have been used in a pseudotype microneutralization assay (pMN), an optimized luciferase reporter assay, that is highly sensitive and specific for detecting neutralizing antibodies against influenza viruses. This has been an invaluable tool in detecting the humoral immune response against specific hemagglutinin or hemagglutinin-esterase fusion proteins for IAV to IDV in serum samples and for screening antibodies for their neutralizing abilities. Additionally, we have also produced influenza neuraminidase (NA) pseudotypes for IAV N1-N9 subtypes and IBV lineages. We have utilized these NA-PV as surrogate antigens in in vitro assays to assess vaccine immunogenicity. These NA PV have been employed as the source of neuraminidase enzyme activity in a pseudotype enzyme-linked lectin assay (pELLA) that is able to measure neuraminidase inhibition (NI) titers of reference antisera, monoclonal antibodies, and postvaccination sera. Here we show the production of influenza HA, HEF, and NA PV and their employment as substitutes for wild-type viruses in influenza serological and neutralization assays. We also introduce AutoPlate, an easily accessible web app that can analyze data from pMN and pELLA quickly and efficiently, plotting inhibition curves and calculating half-maximal concentration (IC50) neutralizing antibody titers. These serological techniques coupled with user-friendly analysis tools are faster, safer, inexpensive alternatives to classical influenza assays while also offering the reliability and reproducibility to advance influenza research and make it more accessible to laboratories around the world.

Uso e adesão de equipamentos de proteção individual por residentes: validação de instrumento de medida / Use and adherence to personal protective equipment by residents: validation of a measuring instrument / Uso y adhesión de equipos de protección individual por parte de los residentes: validación de un instrumento de medida

RESUMO Objetivo: Validar o instrumento de medida que avalia o uso e a adesão de equipamentos de proteção individual entre os residentes de saúde. Método: Estudo metodológico desenvolvido em ambiente virtual entre agosto de 2020 e março de 2021 com residentes da atenção primária e hospitalar das cinco regiões do Brasil. Os participantes responderam a versão adaptada para residentes do "E.P.I.-APS". Utilizou-se a análise fatorial confirmatória para validação do instrumento. Resultados: Participaram 227 residentes, sendo maioria mulher cis (82,8%), atuavam na região Sudeste (58,1%) e em atenção hospitalar (47,6%). A estrutura em oito domínios (gorro/touca; luvas; comportamento de segurança; máscara N95; higienização das mãos; avental/capote; máscara cirúrgica; óculos de proteção/protetor facial) foi confirmada. Apenas dois itens com carga fatorial inferior a 0,5 foram mantidos. Conclusão: O instrumento é válido para mensurar o uso e adesão aos equipamentos de proteção individual entre residentes, o que contribui para direcionar a formação e a segurança laboral.

ABSTRACT Objective: To validate the measuring instrument that assesses healthcare residents' use and adherence to personal protective equipment. Method: Methodological study developed in a virtual environment between August 2020 and March 2021 with primary care and hospital residents from the five regions of Brazil. The participants answered the adapted version of the "P.P.E.-PHC" for residents. Confirmatory factor analysis was used to validate the instrument. Results: 227 residents took part, the majority of whom were cis women (82.8%), worked in the Southeast region (58.1%), and in-hospital care (47.6%). The structure in eight domains (cap, gloves, safety behavior, N95 mask, hand hygiene, apron/coat, surgical mask, goggles/face shield) was confirmed. Only two items with a factor load of less than 0.5 were kept. Conclusion: The tool is valid for measuring residents' use of and adherence to personal protective equipment, which helps to guide training and occupational safety.

RESUMEN Objetivo: Validar el instrumento de medición que evalúa el uso y la adhesión de los equipos de protección individual entre los residentes de salud. Método: Estudio metodológico desarrollado en un entorno virtual entre agosto de 2020 y marzo de 2021 con residentes de atención primaria y hospitalaria de las cinco regiones de Brasil. Los participantes respondieron a la versión adaptada del "E.P.I.-APS" para residentes. Se utilizó un análisis factorial confirmatorio para validar el instrumento. Resultados: Participaron 227 residentes, en su mayoría mujeres cis (82,8%), que trabajaban en el sureste (58,1%) y en cuidados hospitalarios (47,6%). Se confirmó la estructura en ocho dominios (gorro; guantes; comportamiento de seguridad; mascarilla N95; higiene de las manos; delantal/abrigo; mascarilla quirúrgica; gafas/protector facial). Sólo se mantuvieron dos ítems con una carga factorial inferior a 0,5. Conclusión: La herramienta es válida para medir el uso y la adhesión de los equipos de protección individual entre los residentes, lo que ayuda a orientar la formación y la seguridad laboral.

Sequential events during the quiescence to proliferation transition establish patterns of follicle cell differentiation in the Drosophila ovary.

Stem cells cycle between periods of quiescence and proliferation to promote tissue health. In Drosophila ovaries, quiescence to proliferation transitions of follicle stem cells (FSCs) are exquisitely feeding-dependent. Here, we demonstrate feeding-dependent induction of follicle cell differentiation markers, eyes absent (Eya) and castor (Cas) in FSCs, a patterning process that does not depend on proliferation induction. Instead, FSCs extend micron-scale cytoplasmic projections that dictate Eya-Cas patterning. We identify still life and sickie as necessary and sufficient for FSC projection growth and Eya-Cas induction. Our results suggest that sequential, interdependent events establish long-term differentiation patterns in follicle cell precursors, independently of FSC proliferation induction.

Anticipatory postural adjustments in older versus young adults: a systematic review and meta-analysis.

BACKGROUND: Anticipatory postural adjustments (APAs) are a feedforward mechanism triggered in advance to a predictable perturbation, to help the individual counteract mechanical effects that the disturbance may cause. Whether or not this strategy is compromised in the elderly is not a consensus in the literature. METHODS: In this systematic review with meta-analysis, we investigated aging effects on postural control, based on anticipatory postural adjustments (APAs). We selected 11 eligible articles of the following databases: Lilacs, SciELO, PubMed, Cochrane Central, Embase, and CINAHL, involving 324 research participants, assessing their methodological quality and extracting electromyographic, posturographic, and kinematic measurements. We included studies that investigated the occurrence of APAs in healthy younger and older adults, published before 10th August 2022, in English. Studies involving participant with conditions that may affect balance or that did not report measures of onset or amplitude of electromyography (EMG), COP, or kinematics were excluded. To analyze the aggregated results from these studies, we performed the analysis based on the outcome measures (EMG, COP, or kinematic measures) used in individual studies. We calculated differences between younger and older adult groups as the mean differences between the groups and the estimated effect. Egger's test was conducted to evaluate whether this meta-analysis had publication bias. RESULTS: Through this review, older adults showed no significant difference in the velocity to perform a movement compared to the younger adults (MD 0.95, 95% CI -0.86, 2.76, I2 = 82%), but both muscle onset and center of pressure (COP) onset were significantly more delayed in older than in younger adults: erector spinae (MD -31.44, 95% CI -61.79, -1.09, I2 = 95%); rectus abdominis (RA) (MD -31.51, 95% CI -70.58, -3.57, I2 = 85%); tibialis anterior (TA) (MD -44.70, 95% CI -94.30, 4.91, I2 = 63%); soleus (SOL) (MD -37.74, 95% CI -65.43, -10.05, I2 = 91%); gastrocnemius (GAS) (MD -120.59, 95% CI -206.70, -34.49, I2 = 94%); quadriceps (Q) (MD -17.42, 95% CI -34.73, -0.12, I2 = 0%); biceps femoris (BF) (MD -117.47, 95% CI -192.55, -42.70, I2 = 97%); COP onset (MD -45.28, 95% CI -89.57, -0.98, I2 = 93%), and COP apa (COPapa) (MD 2.35, 95% CI -0.09, 4.79, I2 = 64%). These changes did not seem to be linked to the speed of movement but possibly to age-related physiological changes that indicated decreased motor control during APAs in older adults. CONCLUSIONS: Older adults use different postural strategies that aim to increase the safety margin and stabilize the body to perform the movement, according to the requirements imposed, and this should be considered in rehabilitation protocols. SYSTEMATIC REVIEW REGISTRATION: PROSPERO CRD420119143198.

Comparison of Static Balance Control in Infected Htlv-1 Subjects with Different Tsp/Ham Diagnosis.

(1) Background: Tropical spastic paraparesis (TSP/HAM) associated with the T cell lymphotropic virus in type I humans (HTLV-1) is a slow, chronic, and progressive disease that causes balance changes. TSP/HAM diagnosis can be classified as probable, possible, and definite. We compared the static balance control of HTLV-1-infected patients with different TSP/HAM diagnosis. (2) Methods: Our sample consisted of 13 participants infected with HTLV-1 and 16 healthy participants. The center of pressure was recorded using a force platform with open and closed eyes. We divided the recordings into three intervals, period T1 (corresponds to the first 10 s); period T2 (from 10 to 45 s); period T3 (from 45 to 55 s). (3) Results: Eight participants infected with HTLV-1 were classified as probable TSP/HAM and five participants infected with HTLV-1 were classified as definite TSP/HAM. There was a significant increase in postural instability in patients with definite PET/MAH considering the structural and global variables of body sway compared to the control and the probable TSP/HAM. (4) Conclusions: We concluded that the severity of balance is directly related to the degree of signs and symptoms of TSP/HAM.

Postural Adjustments in HTLV-1 Infected Patients during a Self-Initiated Perturbation.

BACKGROUND: Human T-cell lymphotropic virus type 1 (HTLV-1) infection can be associated with tropical spastic paraparesis (TSP/HAM), which causes neurological myelopathy and sensory and muscle tone alterations, leading to gait and balance impairments. Once trunk perturbation is predicted, the motor control system uses anticipatory and compensatory mechanisms to maintain balance by recruiting postural muscles and displacement of the body's center of mass. METHODS: Twenty-six participants (control or infected) had lower limb muscle onset and center of pressure (COP) displacements assessed prior to perturbation and throughout the entire movement. RESULTS: Semitendinosus (ST) showed delayed onset in the infected group compared to the control group. The percentage of trials with detectable anticipatory postural adjustment was also lower in infected groups in the tibialis anterior and ST. In addition, COP displacement in the infected group was delayed, had a smaller amplitude, and took longer to reach the maximum displacement. CONCLUSIONS: HTLV-1 infected patients have less efficient anticipatory adjustments and greater difficulty recovering their postural control during the compensatory phase. Clinical assessment of this population should consider postural stability during rehabilitation programs.

Antibody correlates of protection from SARS-CoV-2 reinfection prior to vaccination: A nested case-control within the SIREN study.

OBJECTIVES: To investigate serological differences between SARS-CoV-2 reinfection cases and contemporary controls, to identify antibody correlates of protection against reinfection. METHODS: We performed a case-control study, comparing reinfection cases with singly infected individuals pre-vaccination, matched by gender, age, region and timing of first infection. Serum samples were tested for anti-SARS-CoV-2 spike (anti-S), anti-SARS-CoV-2 nucleocapsid (anti-N), live virus microneutralisation (LV-N) and pseudovirus microneutralisation (PV-N). Results were analysed using fixed effect linear regression and fitted into conditional logistic regression models. RESULTS: We identified 23 cases and 92 controls. First infections occurred before November 2020; reinfections occurred before February 2021, pre-vaccination. Anti-S levels, LV-N and PV-N titres were significantly lower among cases; no difference was found for anti-N levels. Increasing anti-S levels were associated with reduced risk of reinfection (OR 0·63, CI 0·47-0·85), but no association for anti-N levels (OR 0·88, CI 0·73-1·05). Titres >40 were correlated with protection against reinfection for LV-N Wuhan (OR 0·02, CI 0·001-0·31) and LV-N Alpha (OR 0·07, CI 0·009-0·62). For PV-N, titres >100 were associated with protection against Wuhan (OR 0·14, CI 0·03-0·64) and Alpha (0·06, CI 0·008-0·40). CONCLUSIONS: Before vaccination, protection against SARS-CoV-2 reinfection was directly correlated with anti-S levels, PV-N and LV-N titres, but not with anti-N levels. Detectable LV-N titres were sufficient for protection, whilst PV-N titres >100 were required for a protective effect. TRIAL REGISTRATION NUMBER: ISRCTN11041050.

Influenza A (N1-N9) and Influenza B (B/Victoria and B/Yamagata) Neuraminidase Pseudotypes as Tools for Pandemic Preparedness and Improved Influenza Vaccine Design.

To better understand how inhibition of the influenza neuraminidase (NA) protein contributes to protection against influenza, we produced lentiviral vectors pseudotyped with an avian H11 hemagglutinin (HA) and the NA of all influenza A (N1-N9) subtypes and influenza B (B/Victoria and B/Yamagata). These NA viral pseudotypes (PV) possess stable NA activity and can be utilized as target antigens in in vitro assays to assess vaccine immunogenicity. Employing these NA PV, we developed an enzyme-linked lectin assay (pELLA) for routine serology to measure neuraminidase inhibition (NI) titers of reference antisera, monoclonal antibodies and post-vaccination sera with various influenza antigens. We also show that the pELLA is more sensitive than the commercially available NA-Fluor™ in detecting NA inhibition in these samples. Our studies may lead to establishing the protective NA titer that contributes to NA-based immunity. This will aid in the design of superior, longer lasting and more broadly protective vaccines that can be employed together with HA-targeted vaccines in a pre-pandemic approach.

SARS-CoV-2 immunity and vaccine strategies in people with HIV.

Current severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) vaccines, based on the ancestral Wuhan strain, were developed rapidly to meet the needs of a devastating global pandemic. People living with Human Immunodeficiency Virus (PLWH) have been designated as a priority group for SARS-CoV-2 vaccination in most regions and varying primary courses (two- or three-dose schedule) and additional boosters are recommended depending on current CD4+ T cell count and/or detectable HIV viraemia. From the current published data, licensed vaccines are safe for PLWH, and stimulate robust responses to vaccination in those well controlled on antiretroviral therapy and with high CD4+ T cell counts. Data on vaccine efficacy and immunogenicity remain, however, scarce in PLWH, especially in people with advanced disease. A greater concern is a potentially diminished immune response to the primary course and subsequent boosters, as well as an attenuated magnitude and durability of protective immune responses. A detailed understanding of the breadth and durability of humoral and T cell responses to vaccination, and the boosting effects of natural immunity to SARS-CoV-2, in more diverse populations of PLWH with a spectrum of HIV-related immunosuppression is therefore critical. This article summarizes focused studies of humoral and cellular responses to SARS-CoV-2 infection in PLWH and provides a comprehensive review of the emerging literature on SARS-CoV-2 vaccine responses. Emphasis is placed on the potential effect of HIV-related factors and presence of co-morbidities modulating responses to SARS-CoV-2 vaccination, and the remaining challenges informing the optimal vaccination strategy to elicit enduring responses against existing and emerging variants in PLWH.

Exploiting Pan Influenza A and Pan Influenza B Pseudotype Libraries for Efficient Vaccine Antigen Selection.

We developed an influenza hemagglutinin (HA) pseudotype library encompassing Influenza A subtypes HA1-18 and Influenza B subtypes (both lineages) to be employed in influenza pseudotype microneutralization (pMN) assays. The pMN is highly sensitive and specific for detecting virus-specific neutralizing antibodies against influenza viruses and can be used to assess antibody functionality in vitro. Here we show the production of these viral HA pseudotypes and their employment as substitutes for wildtype viruses in influenza neutralization assays. We demonstrate their utility in detecting serum responses to vaccination with the ability to evaluate cross-subtype neutralizing responses elicited by specific vaccinating antigens. Our findings may inform further preclinical studies involving immunization dosing regimens in mice and may help in the creation and selection of better antigens for vaccine design. These HA pseudotypes can be harnessed to meet strategic objectives that contribute to the strengthening of global influenza surveillance, expansion of seasonal influenza prevention and control policies, and strengthening pandemic preparedness and response.

Development of Lentiviral Vectors Pseudotyped With Influenza B Hemagglutinins: Application in Vaccine Immunogenicity, mAb Potency, and Sero-Surveillance Studies.

Influenza B viruses (IBV) cause respiratory disease epidemics in humans and are therefore components of seasonal influenza vaccines. Serological methods are employed to evaluate vaccine immunogenicity prior to licensure. However, classical methods to assess influenza vaccine immunogenicity such as the hemagglutination inhibition assay (HI) and the serial radial hemolysis assay (SRH), have been proven to have many limitations. As such, there is a need to develop innovative methods that can improve on these traditional assays and provide advantages such as ease of production and access, safety, reproducibility, and specificity. It has been previously demonstrated that the use of replication-defective viruses, such as lentiviral vectors pseudotyped with influenza A hemagglutinins in microneutralization assays (pMN) is a safe and sensitive alternative to study antibody responses elicited by natural influenza infection or vaccination. Consequently, we have produced Influenza B hemagglutinin-pseudotypes (IBV PV) using plasmid-directed transfection. To activate influenza B hemagglutinin, we have explored the use of proteases in increasing PV titers via their co-transfection during pseudotype virus production. When tested for their ability to transduce target cells, the influenza B pseudotypes produced exhibit tropism for different cell lines. The pseudotypes were evaluated as alternatives to live virus in microneutralization assays using reference sera standards, mouse and human sera collected during vaccine immunogenicity studies, surveillance sera from seals, and monoclonal antibodies (mAbs) against IBV. The influenza B pseudotype pMN was found to effectively detect neutralizing and cross-reactive responses in all assays and shows promise as an effective and versatile tool in influenza research.

AutoPlate: Rapid Dose-Response Curve Analysis for Biological Assays.

The emergence of COVID-19 has emphasised that biological assay data must be analysed quickly to develop safe, effective and timely vaccines/therapeutics. For viruses such as SARS-CoV-2, the primary way of measuring immune correlates of protection is through assays such as the pseudotype microneutralisation (pMN) assay, thanks to its safety and versatility. However, despite the presence of existing tools for data analysis such as PRISM and R the analysis of these assays remains cumbersome and time-consuming. We introduce an open-source R Shiny web application and R library (AutoPlate) to accelerate data analysis of dose-response curve immunoassays. Using example data from influenza studies, we show that AutoPlate improves on available analysis software in terms of ease of use, flexibility and speed. AutoPlate (https://philpalmer.shinyapps.io/AutoPlate/) is a tool for the use of laboratories and wider scientific community to accelerate the analysis of biological assays in the development of viral vaccines and therapeutics.