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Chicken TCRγδ+CD8α+T cells are antigen-specific and protective in H9N2 AIV infection.
TCRγδ+ T cells are a major lymphocyte population of chickens, but their response or contribution to immunity against avian influenza virus (AIV) remains unknown. Here, we report an increase in the proportion and activation state of TCRγδ+CD8α+ T cells in the PBMCs of 3 chicken lines (MHC homozygous H-B2 and H-B21 lines and outbred G-WL line) with the strongest responses observed in the more resistant H-B2 chickens. H9N2 AIV infection induced mRNA upregulation of interferon (IFN)-γ and cytotoxicity-associated molecules, including, Granzyme A, Granzyme K, and perforin in sorted TCRγδ+CD8α+ T cells. Moreover, in ex vivo cultured TCRγδ+CD8α+ T cells in response to H9N2 AIV infected splenocytes, strongly indicates the activation of these cells' cytolytic potential via detection of transcription levels of cytotoxic genes with quantitative reverse transcription polymerase chain reaction (qRT-PCR), and IFN-γ protein level with ELISPOT and an intracellular cytokine staining assays. Most importantly, in vivo depletion of γδ T cells led to reduced H9N2 AIV control, which was particularly evident in the early phase of infection. Taken together, these results indicate that strong TCRγδ+CD8α+T cell response plays a critical role in protecting chicken against H9N2 AIV infection.
Hypothetical performance of syndrome-based management of acute paediatric admissions of children aged more than 60 days in a Kenyan district hospital
OBJECTIVE: To investigate whether the outpatient, syndrome-based approach of the Integrated Management of Childhood Illness (IMCI) protocol could be extended to the inpatient arena to give clear and simple minimum standards of care for poorly resourced facilities. METHODS: A prospective, one-year admission cohort retrospectively compared hypothetical performance of syndrome-based management with paediatrician-defined final diagnosis. Admission syndrome definitions were based on local adaptations to the IMCI protocol that encompassed 20 clinical features, measurement of oxygen saturation, and malaria microscopy. FINDINGS: After 315 children with clinically obvious diagnoses (e.g. sickle cell disease and burns) were excluded, 3705 admission episodes were studied. Of these, 2334 (63%) met criteria for at least one severe syndrome (mortality 8% vs <1% for "non-severe" cases), and half of these had features of two or more severe syndromes. No cases of measles were seen. Syndrome-based treatment would have been appropriate (sensitivity >95%) for severe pneumonia, severe malaria, and diarrhoea with severe dehydration, and probably for severe malnutrition (sensitivity 71%). Syndrome-directed treatment suggested the use of broad-spectrum antibiotics in 75/133 (56% sensitivity) children with bacteraemic and 63/71 (89% sensitivity) children with meningitis. CONCLUSIONS: Twenty clinical features, oxygen saturation measurements, and results of malaria blood slides could be used for inpatient, syndrome-based management of acute paediatric admissions. The addition of microscopy of the cerebrospinal fluid and haemoglobin measurements would improve syndrome-directed treatment considerably. This approach might rationalize admission policy and standardize inpatient paediatric care in resource-poor countries, although the clinical detection of bacteraemia remains a problem.
Clarifying the taxonomy of the Finch Louse Fly Ornithomya fringillina (Curtis) (Diptera: Hippoboscidae) – an analysis of morphotypes
The louse flies in the genus Ornithomya are avian ectoparasites. The patterns of alar microtrichia on the wings of the Ornithomyae are commonly used to help distinguish the various species, with the patterns in most species found to be constant between individuals. The Finch Louse Fly Ornithomya fringillina (Curtis) in the United Kingdom, Ireland and the Isle of Man, is unusual in that the several patterns have been described. Consequently it has a complicated taxonomic history and there is some confusion about species identification. Louse flies were collected by licensed bird ringers and an analysis of the simple morphological features, phenology and geographical distribution of these traits was performed. No significant differences were found between the three main types, and it was concluded that the differences were not due to sexual dimorphism and did not provide evidence that the different forms were separate species. Analysis of COX1 DNA sequences confirmed this result and proved that these are indeed morphotypes and not distinct species. There was no geographical separation between COX1 sequences from the United Kingdom and those from flies sampled in other parts of the world. The molecular analysis also suggested that Ornithomya bequaerti (Maa) and Ornithomya candida (Maa) may not be valid species, but represent morphotypes of O. fringillina.
An artificial intelligence-based approach to identify volume status in patients with severe dengue using wearable PPG data.
Dengue shock syndrome (DSS) is a serious complication of dengue infection which occurs when critical plasma leakage results in haemodynamic shock. Treatment is challenging as fluid therapy must balance the risk of hypoperfusion with volume overload. In this study, we investigate the potential utility of wearable photoplethysmography (PPG) to determine volume status in DSS. In this prospective observational study, we enrolled 250 adults and children with a clinical diagnosis of dengue admitted to the Hospital for Tropical Diseases, Ho Chi Minh City. PPG monitoring using a wearable device was applied for a 24-hour period. Clinical events were then matched to the PPG data by date and time. We predefined two clinical states for comparison: (1) the 2-hour period before a shock event was an "empty" volume state and (2) the 2-hour period between 1 and 3 hours after a fluid initiation event was a "full" volume state. PPG data were sampled from these states for analysis. Variability and waveform morphology features were extracted and analyzed using principal components analysis and random forest. Waveform images were used to develop a computer vision model. Of the 250 patients enrolled, 90 patients experienced the predefined outcomes, and had sufficient data for the analysis. Principal components analysis identified four principal components (PCs), from the 23 pulse wave features. Logistic regression using these PCs showed that the empty state is associated with PCs 1 (p = 0.016) and 4 (p = 0.036) with both PCs denoting increased sympathetic activity. Random forest showed that heart rate and the LF-HF ratio are the most important features. A computer vision model had a sensitivity of 0.81 and a specificity of 0.70 for the empty state. These results provide proof of concept that an artificial intelligence-based approach using continuous PPG monitoring can provide information on volume states in DSS.
MetE: a promising protective antigen for tuberculosis vaccine development
IntroductionTuberculosis (TB), caused by Mycobacterium tuberculosis (MTB), remains a significant global health concern. The existing vaccine, Bacillus Calmette-Guérin (BCG), provides inconsistent protection, highlighting the pressing need for a more effective vaccine. We aimed to identify novel MTB antigens and assess their protective efficacy as TB vaccine candidates.MethodsUsing immunopeptidomics, we identified 64 and 80 unique mycobacterial antigens derived from BCG and MTB, respectively. We prioritised antigens based on HLA allele coverage through an immunoinformatics approach.ResultsThe candidates, hisD, metE, and mmpL12, delivered as DNA vaccines, were evaluated for efficacy in mice using the ex vivo Mycobacterial Growth Inhibition Assay (MGIA) and metE was identified as a promising candidate. In vivo murine MTB challenge experiments confirmed the protective efficacy conferred by metE when formulated as recombinant protein with AS01™ or AddaS03™ adjuvants, compared to the naïve group. The immunogenic profiles of metE formulated in the two different adjuvants differed, with metE-AS01™ inducing antigen-specific IFN-γ, TNF-α, IL-2, IL-17, IgG1 and IgG2a-c, while metE-AddaS03™ induced TNF-α, IL-2, IL-17, IL-4, IgM, IgG1, IgG2b.ConclusionOur findings highlight metE as a promising protective antigen for future TB vaccine development.
Risk factors of metabolic dysfunction-associated steatotic liver disease in a cohort of patients with chronic hepatitis B.
BACKGROUND AND AIMS: Chronic hepatitis B (CHB) and metabolic dysfunction-associated steatotic liver disease (MASLD) commonly co-exist, with conflicting data in prevalence and disease severity. We aimed to investigate these discrepancies. METHODS: This multicentre study included consecutive CHB patients from 19 European centres. A survey on standard of care for MASLD screening in CHB was circulated. RESULTS: 1709 CHB patients were included; median age: 53 (42-64), males 60.7%, BMI 25.6 (14-63), 57.3% White. MASLD prevalence (1510 consecutive patients) was 42.3%. BMI (OR=1.27, 95% CI:1.19-1.36), ferritin (OR=1.00, 95% CI:1.00-1.00) and type-2-diabetes (T2DM) (OR=2.60, 95% CI:1.12-6.02) were independently associated with MASLD. The prevalence of advanced fibrosis was 18% (255/1420) in the whole cohort, 25.4% (162/639) among CHB with MASLD, and 13.7% in those without MASLD. Independent predictors of advanced fibrosis were MASLD (OR:2.76, 95%CI:1.50-5.05), BMI (OR:1.08, 95%CI:1.02-1.15), ALT (OR:1.01, 95%CI:1.00-1.03), lower PLTs (OR:0.99, 95%CI:0.98-0.99), insulin-treatment (OR:13.88, 95%CI:2.95-65.28) and long-term antivirals (OR:4.86, 95%CI:2.40-9.85). During follow-up (48 months), only patients without MASLD showed significant LSM improvement over time (p<0.001). Among patients with MASLD, FIB-4 and LSM performed moderately at predicting advanced fibrosis (AUROC 0.71 vs 0.70, p=0.38), against histology. As standard of care, 68.4% centres screened all CHB patients for MASLD. 52.6% followed the same treatment indication in those with CHB and MASLD vs CHB only. CONCLUSION: In this large European cohort, MASLD and fibrosis were highly prevalent among CHB, while MASLD aggravated liver fibrosis. Though screening strategies remain inconsistent, ferritin levels, increased BMI and T2DM may inform on the presence of MASLD. Biomarkers showed modest performance in predicting fibrosis.
Origin and evolution of bacterial periplasmic force transducers.
In double-membraned bacteria, non-equilibrium processes that occur at the outer membrane (OM) are typically coupled to the chemiosmotically-energised inner membrane (IM). TolA and TonB are homologous proteins which energetically couple IM motor proteins to the essential processes of OM-stabilisation and substrate import, respectively. The evolutionary trajectories of these proteins have been difficult to elucidate due to low sequence conservation, yet they are thought to transduce force similarly. Here, this problem was addressed using structural prediction approaches to identify and annotate force transduction operons to trace their distribution and evolutionary origins. In the process, we identify a novel OM-tethering system and a previously unknown family of monomeric force transducers. This approach revealed putative tolA genes, and thus the core organisational principles of the tol-pal operon throughout diverse bacterial taxa. We discovered that the α-helical structure of the periplasm-spanning domain II of TolA previously thought its hallmark, is anomalous amongst most Tol-Pal systems. This structure is mainly prevalent in γ-proteobacteria, likely in adaptation to their lifestyle. Comparison of Tol-Pal and Ton system distribution suggests that TolA emerged from a TonB paralogue and co-emerged with Pal, the OM-tethering lipoprotein that functionalises the Tol-Pal system. We also determined that TolB, the Pal-mobilising protein, likely emerged from a family of outer membrane proteins (OMPs); and CpoB, a periplasmic factor that coordinates peptidoglycan remodelling with cell division, was originally a lipoprotein present in the ancestral Tol-Pal system. The extensive conservation of the Tol-Pal system throughout Gracilicutes highlights its significance in bacterial cell biology.
Distinct neutralization sensitivity between adult and infant transmitted/founder HIV-1 subtype C viruses to broadly neutralizing monoclonal antibodies.
Broadly neutralizing antibodies (bnAbs), passively administered or elicited through vaccination, are a promising strategy for novel HIV prevention, treatment or inducing ART-free remission. However, HIV diversity and evolution are a barrier to the efficacy of bnAbs and there is therefore an urgent need for continuous virus surveillance to identify bnAbs with optimal neutralization breadth and potency against transmitted/founder (TF) viruses, especially in high-burden regions. We determined the neutralization sensitivity of TF viruses isolated within seven days after first detection of heterosexually acquired infection from young women 18-23 years old (n = 39) and within 1 month after birth from in-utero infected infants (n = 21) from FRESH and Baby Cure cohorts respectively, in KwaZulu-Natal, South Africa, where HIV-1 subtype C predominates. Neutralization sensitivities of 47 viruses from FRESH and 21 viruses from Baby Cure were assessed against nine bnAbs targeting different regions on the HIV-1 Env trimer. HIV-1 env sequences within and between bnAb epitopes were compared with database. The bnAbs VRC07-523LS, CAP256-VRC26.25, PGDM1400, 10E8 and PGT151 displayed higher neutralization breadth and potency than other bnAbs against FRESH TF viruses (>70% coverage, starting concentration of 10 μg/ml). Furthermore, VRC07-523LS showed higher neutralization breadth and potency than other bnAbs against Baby Cure TF viruses (p = 0.02). Interestingly, CAP256-VRC26.25 and PGT151 had lower neutralization coverage against infant TF viruses (<60% coverage). Moreover, 40% of infants TF had escape mutations within the V2 loop compared to 28% observed in FRESH and these mutations may explain the observed differences in neutralization sensitivities. However, few mutations were observed in gp120-gp41 interface in both adults and infants. Our findings suggest that intervention studies may have to consider different antibody combinations in adult versus paediatric settings. Moreover, high transmission of escape variants in both vertical and heterosexual transmissions is of concern. This information may be important in the selection of bnAbs that will undergo clinical testing in subtype C settings.
Immunogenicity of standard and extended dosing intervals of BNT162b2 mRNA vaccine.
Extension of the interval between vaccine doses for the BNT162b2 mRNA vaccine was introduced in the United Kingdom to accelerate population coverage with a single dose. At this time, trial data were lacking, and we addressed this in a study of United Kingdom healthcare workers. The first vaccine dose induced protection from infection from the circulating alpha (B.1.1.7) variant over several weeks. In a substudy of 589 individuals, we show that this single dose induces severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) neutralizing antibody (NAb) responses and a sustained B and T cell response to the spike protein. NAb levels were higher after the extended dosing interval (6-14 weeks) compared with the conventional 3- to 4-week regimen, accompanied by enrichment of CD4+ T cells expressing interleukin-2 (IL-2). Prior SARS-CoV-2 infection amplified and accelerated the response. These data on dynamic cellular and humoral responses indicate that extension of the dosing interval is an effective immunogenic protocol.
Acute Plasmodium yoelii 17XNL Infection During BCG Vaccination Limits T Cell Responses and Mycobacterial Growth Inhibition.
Tuberculosis and malaria overlap in many sub-Saharan African countries where Bacillus Calmette Guérin (BCG) vaccination is routinely administered. The aim of this study was to determine whether the timing of BCG vaccination in relation to a malaria infection has implications for BCG vaccine efficacy. Mice were intradermally vaccinated with BCG either 4 weeks before infection with blood-stage Plasmodium yoelii 17XNL, at 13 days post-infection (during an acute blood-stage malaria infection) or 21 days post-infection (after clearance of P. yoelii 17XNL infection). Ex vivo control of mycobacterial growth by splenocytes was used as a surrogate of protective efficacy, and PPD-specific T-cell responses were quantified by flow cytometry. No differences in mycobacterial growth control were detected between BCG vaccinated mice and groups receiving vaccination prior to or after clearance of P. yoelii 17XNL infection. Poorer control of mycobacterial growth was observed following BCG vaccination administered during an acute malarial infection compared to BCG vaccination only or BCG vaccination after blood-stage malaria infection, and mycobacterial growth negatively correlated with the magnitude of total cytokine production from PPD-specific CD4+ T cells (p
Origin and Evolution of Bacterial Periplasmic Force Transducers.
In double-membraned bacteria, non-equilibrium processes that occur at the outer membrane are typically coupled to the chemiosmotically energized inner membrane. TolA and TonB are homologous proteins which energetically couple inner membrane motor proteins to the essential processes of outer membrane stabilization and substrate import, respectively. The evolutionary trajectories of these proteins have been difficult to elucidate due to low-sequence conservation, yet they are thought to transduce force similarly. Here, this problem was addressed using structural prediction approaches to identify and annotate force transduction operons to trace their distribution and evolutionary origins. In the process, we identify a novel outer membrane-tethering system and a previously unknown family of monomeric force transducers. This approach revealed putative tolA genes, and thus the core organizational principles of the tol-pal operon throughout diverse bacterial taxa. We discovered that the α-helical structure of the periplasm-spanning domain II of TolA previously thought its hallmark, is anomalous amongst most Tol-Pal systems. This structure is mainly prevalent in γ-proteobacteria, likely in adaptation to their lifestyle. Comparison of Tol-Pal and Ton system distribution suggests that TolA emerged from a TonB paralogue and co-emerged with Pal, the outer membrane-tethering lipoprotein that functionalizes the Tol-Pal system. We also determined that TolB, the Pal-mobilizing protein, likely emerged from a family of outer membrane proteins; and CpoB, a periplasmic factor that coordinates peptidoglycan remodeling with cell division, was originally a lipoprotein present in the ancestral Tol-Pal system. The extensive conservation of the Tol-Pal system throughout Gracilicutes highlights its significance in bacterial cell biology.
Iron deficiency causes aspartate-sensitive dysfunction in CD8+ T cells.
Iron is an irreplaceable co-factor for metabolism. Iron deficiency affects >1 billion people and decreased iron availability impairs immunity. Nevertheless, how iron deprivation impacts immune cell function remains poorly characterised. We interrogate how physiologically low iron availability affects CD8+ T cell metabolism and function, using multi-omic and metabolic labelling approaches. Iron limitation does not substantially alter initial post-activation increases in cell size and CD25 upregulation. However, low iron profoundly stalls proliferation (without influencing cell viability), alters histone methylation status, gene expression, and disrupts mitochondrial membrane potential. Glucose and glutamine metabolism in the TCA cycle is limited and partially reverses to a reductive trajectory. Previous studies identified mitochondria-derived aspartate as crucial for proliferation of transformed cells. Despite aberrant TCA cycling, aspartate is increased in stalled iron deficient CD8+ T cells but is not utilised for nucleotide synthesis, likely due to trapping within depolarised mitochondria. Exogenous aspartate markedly rescues expansion and some functions of severely iron-deficient CD8+ T cells. Overall, iron scarcity creates a mitochondrial-located metabolic bottleneck, which is bypassed by supplying inhibited biochemical processes with aspartate. These findings reveal molecular consequences of iron deficiency for CD8+ T cell function, providing mechanistic insight into the basis for immune impairment during iron deficiency.
Features influencing the health and economic impact of preventing COVID-19 in immunocompromised individuals.
Many immunocompromised individuals mount inadequate immune responses following COVID-19 vaccination, thus relying on other social distancing behaviours, particularly shielding, for protection, impacting their quality of life. However, little is known about historical/current levels and effectiveness of shielding or factors influencing individuals' decision to continue shielding. Long-acting antibody pre-exposure prophylaxis (LAAB-PrEP) provides direct protection against COVID-19 in immunocompromised individuals who have been and may continue to shield. However, the proportion and incidence of circulating variants for which LAAB-PrEP would be effective is unpredictable. Given this uncertain behavioural and immuno-epidemiological context, we developed a modelling framework to explore features that most impact health outcomes and cost effectiveness of long-term administration of LAAB-PrEP against COVID-19 infection in immunocompromised individuals in the English context. The model predicted that the incremental cost-effectiveness ratio (ICER) of LAAB-PrEP against COVID-19 in immunocompromised individuals will be largely driven by features of utility of shielding, current/future shielding behaviour, cost of shielding, risk of COVID-19 hospitalisation among immunocompromised individuals and the time horizon used for the cost-effectiveness analysis. The model estimated that for realistic ranges of influential factors, it is possible for LAAB-PrEP to be cost effective under the conditions that most immunocompromised individuals would shield indefinitely if it were not available but would switch to LAAB-PrEP if it were. Thus, if individuals stop shielding when taking LAAB-PrEP, then LAAB-PrEP is cost effective.