The Eurasian lynx is of special conservation concern based on the European Union's Habitat Directive and its populations need to be maintained or restored at favourable conservation status. To ...evaluate lynx population status, appropriate monitoring needs to be in place. We modelled the distribution dynamics of lynx in the Alps (200 000 km2) during 1994–2014 at a resolution of 100 km2. Lynx distribution and detection probability varied by year, country, forest cover, elevation and distance to the nearest release site. Occupancy of neighbouring quadrats had a strong positive effect on colonization and persistence rates. Our analyses demonstrate the importance of accounting for imperfect detection: the raw data underestimated the lynx range by 55% on average, depending on country and winter. Over the past 20 years the Alpine lynx range has expanded at an average rate of 4% per year, which was partly due to the lynx translocations to new areas. Our approach to large‐scale distribution modelling and analysing trends using site occupancy models can be applied retrospectively and is useful in many cases where a network of trained people is established to report the presence of target species, for example, in Europe where member states of the European Union have to report conservation status of species of community interest. Hence, dynamic occupancy models are an appealing framework for inference about the large‐scale range dynamics based on opportunistic data and a useful tool for large‐scale management and conservation programmes.
We analysed range dynamics of a reintroduced large carnivore, the Eurasian lynx, in the Alps (200 000 km2) over 20 years, combining a cutting edge occupancy model with citizen science. Lynx distribution and detection probability varied by distance to the nearest release site, year, forest cover, elevation and country. Occupancy of neighbouring quadrats had a strong positive effect on colonization and persistence rates. Our analyses demonstrate the importance of accounting for imperfect detection: the raw data underestimated the lynx range by 55% on average. Over the past 20 years the Alpine lynx range expanded at an average rate of 4% per year, which was partly due to the lynx translocations to new areas. Our approach to large‐scale distribution modelling and analysing trends using site occupancy models can be applied retrospectively and is useful in many cases where a network of trained people is established to report the presence of target species.
Inferring the distribution and abundance of a species from field records must deal with false‐negative and false‐positive errors. False‐negative errors occur if a species present goes undetected, ...while false‐positive errors are typically a consequence of species misidentification. False‐positive observations in studies of rare species may cause an overestimation of the distribution or abundance of the species and distort trend indices. We illustrate this issue with the monitoring of the Eurasian lynx in the Alps. We developed a three‐level classification of field records according to their reliability as inferred from whether they were validated or not. The first category (C1) represents ‘hard fact’ data (e.g. dead lynx); the second category (C2) includes confirmed data (e.g. tracks verified by an expert); and the third category (C3) are unconfirmed data (e.g. any kind of direct visual observation). For lynx, which is a comparatively well‐known species in the Alps, we use site‐occupancy modelling to estimate its distribution and show that the inferred lynx distribution is highly sensitive to presence sign category: it is larger if based on C3 records compared with the more reliable C1 and C2 records. We believe that the reason for this is a fairly high frequency of false‐positive errors among C3 records. This suggests that distribution records for many lesser‐known species may be similarly unreliable, because they are mostly or exclusively based on unconfirmed and thus soft data. Nevertheless, such soft data form a considerable part of species assessments as presented, for example in the International Union for Conservation of Nature Red List. However, C3 records can often not be discarded because they may be the only information available. When inferring the distribution of rare carnivores, especially for species with an expanding or shrinking range, we recommend a rigorous discrimination between fully reliable and un‐ or only partly reliable data, in order to identify possible methodological problems in the distribution maps related to false‐positive records.
Abstract
Objectives
To investigate ceftolozane/tazobactam pharmacokinetics (PK) in plasma and interstitial space fluid (ISF) of muscle and subcutaneous tissue and establish a population PK model.
...Methods
Eight healthy volunteers received four IV doses of 1000/500 mg ceftolozane/tazobactam q8h in a prospective, open-labelled PK study. ISF concentration–time profiles were determined via in vivo microdialysis up to 8 h post-dose and efficacy of unbound ceftolozane and tazobactam was estimated using the time above MIC (%ƒT>MIC) and time above threshold concentration (%T>CT), respectively. A population PK model was established by merging derived plasma and soft tissue PK data.
Results
Ceftolozane reached %ƒT>MIC values of 100% in plasma, muscle and subcutaneous ISF for Enterobacteriaceae and 87%, 89% and 87%, respectively, for Pseudomonas aeruginosa. Tazobactam %T>CT was 21%, 22% and 21% in plasma, muscle and subcutaneous ISF, respectively. Plasma protein binding was 6.3% for ceftolozane and 8.0% for tazobactam. Multiple-dose ceftolozane AUC0–8 ISF/plasma ratios were 0.92 ± 0.17 in muscle and 0.88 ± 0.18 in subcutis, and tazobactam ratios were 0.89 ± 0.25 in muscle and 0.87 ± 0.21 in subcutis, suggesting substantial soft tissue penetration.
Conclusions
Tazobactam %T>CT values were distinctly below proposed target values, indicating that tazobactam might be underdosed in the investigated drug combination. However, ISF/unbound plasma ratios of ceftolozane and tazobactam support their use in soft tissue infections. A plasma and soft tissue PK model adds important information on the PK profile of ceftolozane/tazobactam. Further investigations in patients suffering from wound infections are needed to confirm these findings.
Dendritic cells (DC) form a specialized system for presenting Ag to naive or quiescent T cells and consequently play a central role in the induction of T and B cell immunity. In this study we used DC ...generated from peripheral progenitors to analyze the effect of IL-10 on the accessory function of human DC. We demonstrate that immature DC, harvested on days 9 to 11 and exposed to IL-10 for the last 2 days of culture, show a strongly reduced capacity to stimulate a CD4+ T cell response in an allogeneic MLR in a dose-dependent manner. In contrast, fully mature DC are completely resistant to the effects of IL-10. These results were obtained in both an alloantigen-induced MLR and an anti-CD3 mAb-induced response of primed and naive (CD45RA+) CD4+ T cells. FACS analysis revealed inhibition of the up-regulation of the costimulatory molecules CD58 and CD86 and the specific DC marker CD83 in DC pretreated with IL-10. These data suggest that IL-10 inhibited the development of fully mature DC. Furthermore, DC precultured with IL-10, but not controls, induced a state of alloantigen-specific anergy in CD4+ T cells and of peptide-specific anergy in the influenza hemagglutinin-specific T cell clone HA1.7. Analysis of the supernatants of these anergic T cells revealed a reduced production of IL-2 and IFN-gamma compared with that in control cells. Collectively, these data suggest that IL-10 converts immature DC into tolerogenic APC, which might be a useful tool in the therapy of patients with autoimmune or allergic diseases.
Relapse remains a leading cause of death after allogeneic hematopoietic cell transplantation (HCT) for patients with high-risk leukemias. The potentially beneficial donor T cell-mediated ...graft-versus-leukemia (GVL) effect is often mitigated by concurrent graft-versus-host disease (GVHD). Providing T cells that can selectively target Wilms tumor antigen 1 (WT1), a transcription factor overexpressed in leukemias that contributes to the malignant phenotype, represents an opportunity to promote antileukemic activity without inducing GVHD. HLA-A*0201-restricted WT1-specific donor-derived CD8 cytotoxic T cell (CTL) clones were administered after HCT to 11 relapsed or high-risk leukemia patients without evidence of on-target toxicity. The last four treated patients received CTL clones generated with exposure to interleukin-21 (IL-21) to prolong in vivo CTL survival, because IL-21 can limit terminal differentiation of antigen-specific T cells generated in vitro. Transferred cells exhibited direct evidence of antileukemic activity in two patients: a transient response in one patient with advanced progressive disease and the induction of a prolonged remission in a patient with minimal residual disease (MRD). Additionally, three treated patients at high risk for relapse after HCT survive without leukemia relapse, GVHD, or additional antileukemic treatment. CTLs generated in the presence of IL-21, which were transferred in these latter three patients and the patient with MRD, all remained detectable long-term and maintained or acquired in vivo phenotypic and functional characteristics associated with long-lived memory CD8 T cells. This study supports expanding efforts to immunologically target WT1 and provides insights into the requirements necessary to establish potent persistent T cell responses.
Chimeric antigen receptor (CAR)-T cell therapy is a revolutionary approach which produced remarkable clinical responses in patients with B-cell malignancies, however challenges limit their efficacy ...in other tumours. It has been shown that this limited efficacy is due to the nature of these tumours and the presence of a hostile tumour microenvironment reducing/abrogating their functionality and persistence.
Aim of the study was to identify metabolic mechanisms responsible for loosing CAR-T and other cellular products’ response and develop strategies to improve their efficacy and persistence.
Adoptive cellular products were characterized molecularly (real-time PCR/western blot/RNAseq/proteomics), structurally (electron microscopy (EM)), immunophenotypically (facs/immunofluorescence (IF)), metabolically (seahorse/IF) and functionally (in vitro short- and long-term co-cultures in 2D and 3D models/in a murine model).
Analysing GD2.CAR-T cells from our clinical trial (NCT03373097), we observed that the ex-vivo T-cell manipulation induces a deep and lasting blockage of autophagy pathways independently of the kind of activation, stimulation or cytokines used. We extend our analysis to other cellular products (antigen-specific cytotoxic T cells and NK cells) observing also a strong autophagy downregulation. Then, we explored whether autophagy can be restored by the over-expression of autophagy key-proteins. This results in an autophagy restoration, increased mitochondrial potential and frequency of naïve cells, better oxygen consumption and extracellular acidification rate. Further, autophagy restored T cells showed a significant reduction in apoptosis, improved response to external stimulus, cell mobility, cell-to-cell communication and intracellular organelle organization. EM analysis also revealed mitochondria colocalization, autophagosomes and micro-vesicles. At the functional level, autophagy-restored CAR-T cells eradicate more efficiently tumour at low effector:target ratios in both short and long-term co-cultures using 2D and 3D models as well as in a murine model without signs of exhaustion or toxicity.
In conclusion, autophagy restoration in CAR T cells and adoptive cellular products could revolutionize the effectiveness of these approaches securing better persistence, anti-tumour activity and ability to resist to negative stimuli induced by the tumour and its TME.
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