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Abstract Commensal relationships between wild plants and their dispersers play a key ecological and evolutionary role in community structure and function. While non-human dispersers are often considered critical to plant recruitment, human dispersers have received much less attention, especially when it comes to non-domesticated plants. Australia, as a continent historically characterized by economies reliant on non-domesticated plants, is thus a key system for exploring the ecological role of people as seed dispersers in the absence of agriculture. Here, we utilize a controlled observation research design, employing ecological surveys and ethnographic observations to examine how seed dispersal and landscape burning by Martu Aboriginal people affects the distribution of three preferred plants and one (edible, but non-preferred) control species. Using an information theoretic approach, we find that the three preferred plants show evidence of human dispersal, with the strongest evidence supporting anthropogenic dispersal for the wild bush tomato, Solanum diversiflorum.

Summary: Vγ9Vδ2 T cells constitute a homogeneous effector T cell population that lyses tumors of different origin, including the prostate. We generated a bispecific T cell engager (bsTCE) to direct Vγ9Vδ2 T cells to PSMA+ prostate cancer (PCa) cells. The PSMA-Vδ2 bsTCE triggered healthy donor and PCa patient-derived Vγ9Vδ2 T cells to lyse PSMA+ PCa cell lines and patient-derived tumor cells while sparing normal prostate cells and enhanced Vγ9Vδ2 T cell antigen cross-presentation to CD8+ T cells. Vγ9Vδ2 T cell expressed NKG2D and DNAM-1 contributed to Vγ9Vδ2 T cell activation and tumor lysis at low PSMA-Vδ2 bsTCE concentrations. In vivo models confirmed the antitumor efficacy of the bsTCE and demonstrated a half-life of 6–7 days. Tissue-cross reactivity analysis was in line with known tissue distribution of PSMA and Vγ9Vδ2 T cells. Together these data show the PSMA-Vδ2 bsTCE to represent a promising anti-tumor strategy and supports its ongoing evaluation in a phase 1/2a clinical trial in therapy refractory metastatic castration-resistant PCa.

Background: Despite considerable advancements in cancer immunotherapy, advanced melanoma still presents a substantial clinical challenge. In an effort to explore treatment options, we examined the immunotherapeutic potential of effector Vγ9Vδ2-T cells in vitro in a three-dimensional (3D) human organotypic melanoma-in-skin (Mel-RhS) model. Materials and methods: Vγ9Vδ2-T cells were introduced into Mel-RhS via intradermal injection and cultured within the tissue microenvironment for up to 3 days. Results: Vγ9Vδ2-T cells remained viable for up to 3 days and were in close proximity to or within tumor nests. Upon Mel-RhS dissociation, a fraction was shown to be decorated by melanoma-associated chondroitin sulfate proteoglycan (MCSP), demonstrating their ability to actively navigate the tumor microenvironment and trogocytose cancer cells. Investigation into the apparent trogocytosis revealed an enhanced activated state of MCSP-decorated Vγ9Vδ2-T cells, evidenced by increased expression levels of 4-1BB, NKp44, programmed cell death protein-1 (PD-1), and programmed death-ligand 1 (PD-L1), compared with their MCSP− counterpart. These findings suggest that Vγ9Vδ2-T cells, upon successfully contacting melanoma cells, actively recognize and acquire MCSP from these malignant cells. Evidence of actual tumor cell elimination, although not significant, was only obtained after preincubation of Mel-RhS with pamidronate, a phosphoantigen-inducing agent, indicating the need for additional T cell receptor-mediated signaling for Vγ9Vδ2-T cells to reach their full oncolytic potential. Conclusions: This study highlights the viability and persistence of Vγ9Vδ2-T cells within the 3D microenvironment, their migratory and antitumor functionality, and the suitability of the model for testing T cell-based therapies, contributing both to the understanding of Vγ9Vδ2-T cell biology and their application in cancer immunotherapy.

BackgroundVγ9Vδ2 T-cells are antitumor immune effector cells that can detect metabolic dysregulation in cancer cells through phosphoantigen-induced conformational changes in the butyrophilin (BTN) 2A1/3A1 complex. In order to clinically exploit the anticancer properties of Vγ9Vδ2 T-cells, various approaches have been studied including phosphoantigen stimulation, agonistic BTN3A-specific antibodies, adoptive transfer of expanded Vγ9Vδ2 T-cells, and more recently bispecific antibodies. While Vγ9Vδ2 T-cells constitute a sizeable population, typically making up ~1-10% of the total T cell population, lower numbers have been observed with increasing age and in the context of disease. MethodsWe evaluated whether bivalent single domain antibodies (VHHs) that link Vδ2-TCR specific VHHs with different affinities could support Vγ9Vδ2 T-cell expansion and could be incorporated in a bispecific engager format when additionally linked to a tumor antigen specific VHH. ResultsBivalent VHHs that link a high and low affinity Vδ2-TCR specific VHH can support Vγ9Vδ2 T-cell expansion. The majority of Vγ9Vδ2 T-cells that expanded following exposure to these bivalent VHHs had an effector or central memory phenotype and expressed relatively low levels of PD-1. Bispecific engagers that incorporated the bivalent Vδ2-TCR specific VHH as well as a tumor antigen specific VHH triggered antitumor effector functions and supported expansion of Vγ9Vδ2 T-cells in vitro and in an in vivo model in NOG-hIL-15 mice.ConclusionBy enhancing the number of Vγ9Vδ2 T-cells available to exert antitumor effector functions, these novel Vδ2-bivalent bispecific T cell engagers may promote the overall efficacy of bispecific Vγ9Vδ2 T-cell engagement, particularly in patients with relatively low levels of Vγ9Vδ2 T-cells.

Background CD1d is a monomorphic major histocompatibility complex class I-like molecule that presents lipid antigens to distinct T-cell subsets and can be expressed by various malignancies. Antibody-mediated targeting of CD1d on multiple myeloma cells was reported to induce apoptosis and could therefore constitute a novel therapeutic approach.Methods To determine how a CD1d-specific single-domain antibody (VHH) enhances binding of the early apoptosis marker annexin V to CD1d+ tumor cells we use in vitro cell-based assays and CRISPR-Cas9-mediated gene editing, and to determine the structure of the VHH1D17-CD1d(endogenous lipid) complex we use X-ray crystallography.Results Anti-CD1d VHH1D17 strongly enhances annexin V binding to CD1d+ tumor cells but this does not reflect induction of apoptosis. Instead, we show that VHH1D17 enhances presentation of phosphatidylserine (PS) in CD1d and that this is saposin dependent. The crystal structure of the VHH1D17-CD1d(endogenous lipid) complex demonstrates that VHH1D17 binds the A′-pocket of CD1d, leaving the lipid headgroup solvent exposed, and has an electro-negatively charged patch which could be involved in the enhanced PS presentation by CD1d. Presentation of PS in CD1d does not trigger phagocytosis but leads to greatly enhanced binding of T-cell immunoglobulin and mucin domain containing molecules (TIM)-1 to TIM-3, TIM-4 and induces TIM-3 signaling.Conclusion Our findings reveal the existence of an immune modulatory CD1d(PS)-TIM axis with potentially unexpected implications for immune regulation in both physiological and pathological conditions.

Abstract Specimen identification is the backbone of archeozoological research. The challenge of differentiating postcranial skeletal elements of closely related wild animals in biodiverse regions can prove a barrier to understanding past human foraging behaviours. Morphometrics are increasingly being employed to classify paleozoological animal remains, however, the potential of these methods to discriminate between wild animal groups has yet to be fully realised. Here we demonstrate the applicability of a traditional morphometric approach to taxonomically classify foot and ankle bones of kangaroos, a large and highly diverse marsupial family. Using multiple discriminant analysis, we classify archaeological specimens from Boodie Cave, in northwest Australia and identify the presence of two locally extinct macropod species during the terminal Pleistocene. The appearance of the banded hare-wallaby and northern nail-tail wallaby in the Pilbara region at this time provides independent evidence of the ecological and human responses to a changing climate at the end of the last Ice Age. Traditional morphometrics provides an accessible, inexpensive, and non-destructive tool for paleozoological specimen classification and has substantial potential for applications to other diverse wild faunas.

While immunoglobulin A (IgA) is well known for its neutralizing and anti-inflammatory function, it is becoming increasingly clear that IgA can also induce human inflammatory responses by various different immune cells. Yet, little is known about the relative role of induction of inflammation by the two IgA subclasses i.e. IgA1, most prominent subclass in circulation, and IgA2, most prominent subclass in the lower intestine. Here, we set out to study the inflammatory function of IgA subclasses on different human myeloid immune cell subsets, including monocytes, and in vitro differentiated macrophages and intestinal CD103+ dendritic cells (DCs). While individual stimulation with IgA immune complexes only induced limited inflammatory responses by human immune cells, both IgA subclasses strongly amplified pro-inflammatory cytokine production upon co-stimulation with Toll-like receptor (TLR) ligands such as Pam3CSK4, PGN, and LPS. Strikingly, while IgA1 induced slightly higher or similar levels of pro-inflammatory cytokines by monocytes and macrophages, respectively, IgA2 induced substantially more inflammation than IgA1 by CD103+ DCs. In addition to pro-inflammatory cytokine proteins, IgA2 also induced higher mRNA expression levels, indicating that amplification of pro-inflammatory cytokine production is at least partially regulated at the level of gene transcription. Interestingly, cytokine amplification by IgA1 was almost completely dependent on Fc alpha receptor I (FcαRI), whilst blocking this receptor only partially reduced cytokine induction by IgA2. In addition, IgA2-induced amplification of pro-inflammatory cytokines was less dependent on signaling through the kinases Syk, PI3K, and TBK1/IKKϵ. Combined, these findings indicate that IgA2 immune complexes, which are most abundantly expressed in the lower intestine, particularly promote inflammation by human CD103+ intestinal DCs. This may serve an important physiological function upon infection, by enabling inflammatory responses by this otherwise tolerogenic DC subset. Since various inflammatory disorders are characterized by disturbances in IgA subclass balance, this may also play a role in the induction or exacerbation of chronic intestinal inflammation.

Summary: The emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants poses continuous challenges in combating the virus. Here, we describe vaccination strategies to broaden SARS-CoV-2 and sarbecovirus immunity by combining spike proteins based on different viruses or viral strains displayed on two-component protein nanoparticles. First, we combined spike proteins based on ancestral and Beta SARS-CoV-2 strains to broaden SARS-CoV-2 immune responses. Inclusion of Beta spike improved neutralizing antibody responses against SARS-CoV-2 Beta, Gamma, and Omicron BA.1 and BA.4/5. A third vaccination with ancestral SARS-CoV-2 spike also improved cross-neutralizing antibody responses against SARS-CoV-2 variants, in particular against the Omicron sublineages. Second, we combined SARS-CoV and SARS-CoV-2 spike proteins to broaden sarbecovirus immune responses. Adding SARS-CoV spike to a SARS-CoV-2 spike vaccine improved neutralizing responses against SARS-CoV and SARS-like bat sarbecoviruses SHC014 and WIV1. These results should inform the development of broadly active SARS-CoV-2 and pan-sarbecovirus vaccines and highlight the versatility of two-component nanoparticles for displaying diverse antigens.

DNA profiling is just one of many tools available to police in a criminal investigation. However, unlike any other criminal investigative tool, DNA profiling has captured the public imagination. Its portrayal in popular fiction has given rise to the “CSI effect”, whereby the weight and credibility of science, combined with the predictive potential of DNA, produce an aura of infallibility and certainty only amplified by creative license. Genetic material holds particular significance for Indigenous peoples, establishing identity and group membership, as well as associated rights. This taonga (treasure) status entails particular sensitivities regarding its handling and use; these have been well explored in health, but less so in the forensic context. This article presents professional and lay Māori perspectives on forensic DNA technologies in New Zealand, highlighting the inseparability of these from Indigenous experiences of criminalization but also their value in informing operational, ethical and justice-oriented considerations.

Advanced ecological modelling reveals how Sahul (Australia and New Guinea) was first peopled, suggesting the most probable routes and surprisingly rapid early settlement of this continent by anatomically modern humans starting 50,000 to 75,000 years ago.

We show how the replica method can be used to compute the asymptotic eigenvalue spectrum of a real Wishart product matrix. For unstructured factors, this provides a compact, elementary derivation of a polynomial condition on the Stieltjes transform first proved by Müller [IEEE Trans. Inf. Theory. 48, 2086-2091 (2002)]. We then show how this computation can be extended to ensembles where the factors are drawn from matrix Gaussian distributions with general correlation structure. For both unstructured and structured ensembles, we derive polynomial conditions on the average values of the minimum and maximum eigenvalues, which in the unstructured case match the results obtained by Akemann, Ipsen, and Kieburg [Phys. Rev. E 88, 052118 (2013)] for the complex Wishart product ensemble.

Abstract DMD is a rare disorder characterized by progressive muscle degeneration and premature death. Therapy development is delayed by difficulties to monitor efficacy non‐invasively in clinical trials. In this study, we used RNA‐sequencing to describe the pathophysiological changes in skeletal muscle of 3 dystrophic mouse models. We show how dystrophic changes in muscle are reflected in blood by analyzing paired muscle and blood samples. Analysis of repeated blood measurements followed the dystrophic signature at five equally spaced time points over a period of seven months. Treatment with two antisense drugs harboring different levels of dystrophin recovery identified genes associated with safety and efficacy. Evaluation of the blood gene expression in a cohort of DMD patients enabled the comparison between preclinical models and patients, and the identification of genes associated with physical performance, treatment with corticosteroids and body measures. The presented results provide evidence that blood RNA‐sequencing can serve as a tool to evaluate disease progression in dystrophic mice and patients, as well as to monitor response to (dystrophin‐restoring) therapies in preclinical drug development and in clinical trials.

BackgroundPeriodontitis is a chronic multifactorial inflammatory disease of the supportive tissues of the teeth. In more recent years, remarkable epidemiological and pathophysiological associations between periodontitis and cardiovascular disease (CVD) have been presented. Whether or not treatment of periodontitis is valuable for primary or secondary prevention of cardiovascular disease, has not yet been fully established. In this practice-based pilot study we focused on primary prevention of cardiovascular disease, by investigating the effect of periodontal treatment on the earliest detectable stage of CVD; endothelial dysfunction.MethodsOtherwise healthy periodontitis and non-periodontitis participants 45–70 years of age were included in the study. One year after completing periodontal (non-surgical and surgical) treatment of the periodontitis patients and 1 year after inclusion of the controls, all baseline measurements were repeated. Full-mouth examinations were performed by a periodontist to determine their Periodontal Inflamed Surface Area (PISA) score and other dental parameters. To assess the cardiovascular conditions, endothelial function through the reactive hyperemia index (RHI) assessed by the EndoPAT™, and several physical and biochemical parameters were measured.Results21 patients with diagnosed, untreated periodontitis and 21 participants without periodontitis were included in this follow-up study. After periodontal therapy in the periodontitis patients, the PISA reduced significantly. The RHI did not show a significant improvement after treatment of the periodontitis patients (−0.1 ± 0.8, p = 0.524). Similarly, other secondary cardiovascular outcome measurements, hsCRP, total cholesterol, HDL cholesterol, triglycerides, HbA1c, and systolic blood pressure did not improve significantly after periodontal treatment. Controls did not show any significant changes in the RHI, in other CVD parameters and in the PISA after 1-year follow-up.ConclusionIn this practice-based pilot study, periodontal treatment did not improve the endothelial function in otherwise healthy adults with periodontitis. Future studies are needed to be of larger size and could focus on periodontitis patients with co-morbidities to investigate whether periodontal treatment has secondary preventive effect on endothelial function and other CVD parameters.Clinical Trial Registration[www.ClinicalTrials.gov], identifier [ISRCTN55656827].

Background: Periodontitis is a chronic multifactorial inflammatory disease of the supportive tissues of the teeth. Pathophysiological evidence suggests a possible common inflammatory background between periodontitis and cardiovascular diseases (CVD). Pathological and epidemiological associations between these two diseases have been presented, but are still debated. This study aimed to investigate the association between the inflammatory burden of periodontitis and the presence and extent of coronary calcification. Secondary aims were to study other cardiovascular parameters and cardiovascular risk predictors in relation to periodontitis and dental health. Methods: Healthy periodontitis or non-periodontitis patients 45–70 years of age were included in a prospective cross-sectional study. Full-mouth examinations were performed by a periodontist to determine their Periodontal Inflamed Surface Area (PISA) score and other dental parameters. To assess the cardiovascular conditions, Coronary Artery Calcium (CAC) scores, endothelial function assessments by the EndoPAT ™, and several physical and biochemical examinations were performed. Results: Seventy-one patients were included. Elevated CAC scores and endothelial dysfunction were not significantly related to PISA or dental health. PISA was significantly related to the Framingham and Reynolds CVD risk predictors, but were no longer significant after correction for confounders. The same applied to the significant relations between tooth loss, dental plaque and bleeding scores and the CVD risk predictors. Conclusions: Periodontitis is associated with increased CVD risk, but is not an independent risk factor. This link is still important to make to bridge the gap between dentistry and general medicine and to identify patients at risk for CVD in an earlier stage.

Interleukin (IL)-4 is a cytokine that affects both adaptive and innate immune responses. In the central nervous system, microglia express IL-4 receptors and it has been described that IL-4-exposed microglia acquire anti-inflammatory properties. We here demonstrate that IL-4 exposure induces changes in the cell surface protein expression profile of primary rhesus macaque microglia and enhances their potential to induce proliferation of T cells with a regulatory signature. Moreover, we show that Toll like receptor (TLR)-induced cytokine production is broadly impaired in IL-4-exposed microglia at the transcriptional level. IL-4 type 2 receptor-mediated signaling is shown to be crucial for the inhibition of microglial innate immune responses. TLR-induced nuclear translocalization of NF-κB appeared intact, and we found no evidence for epigenetic modulation of target genes. By contrast, nuclear extracts from IL-4-exposed microglia contained significantly less NF-κB capable of binding to its DNA consensus site. Further identification of the molecular mechanisms that underlie the inhibition of TLR-induced responses in IL-4-exposed microglia may aid the design of strategies that aim to modulate innate immune responses in the brain, for example in gliomas.

Online texts rapidly are becoming central to children's out-of-school literate lives. However, children's engagement with such unofficial texts rarely is addressed in schools, or is addressed in limited ways. To address this gap, we argue for conceptualizing popular media, including websites, as the new literatures of childhood, tracking how popular websites maintain many of the values of children's literature but extend beyond the genres, purposes, and textual practices commonly discussed in children's literature. Through analyses of cultural models of gender in four popular websites--including cultural models of ideal girls and boys, appropriate activities for each gender, and characterizations of the opposite sex--we illustrate how a literary approach to new media texts can be used to refocus literacy education.

The temporal resolution of optical measurements of neural activity has traditionally been limited by the image or volume acquisition rate. Here, the authors describe an analysis that exploits the short duration of neural measurements within each image to extract neural responses at higher temporal resolution than the acquisition rate.