Selection of an antibody library identifies a pathway to induce immunity by targeting CD36 on steady state CD8\u3b1+ dendritic cells

Due to their prominent role in the orchestration of a broad range of immune responses, dendritic cells (DCs) have emerged in the past decade as central target for cancer immunotherapy. Recent advance in the knowledge of DCs functions and subset specialization led to the design of novel immunotherapeutic approaches based on the possibility to target DCs directly in vivo, thus avoiding their ex vivo manipulation. A promising strategy is to use antibodies to target antigens to cell-surface molecules expressed by DCs, in order to increase T-cell mediated immune responses. The studies performed so far, have revealed that the efficacy of in vivo DC vaccination depend on several factors including the specific DC subset targeted, their maturation status and the nature and biological properties of the receptor targeted. Therefore, the identification of the most appropriate ligand/receptor pairing it is a requisite to improve the modality of delivery tumor Ags to DCs. To this aim, we screened a library of Ab fragments on mouse DCs to isolate new potential antibodies capable of targeting DCs in vivo and able to induce T-cell mediated immune responses against specific antigens. In this study, we provide the proof of principle that the phage display technology can be successfully used to isolate internalizing antibodies on mouse DCs. We further develop such technology by engineering the selected molecules to create antigen fusion proteins to use in vaccination protocols. In particular, we focus on a high affinity Ab against CD36, a multiligand scavenger receptor primarily expressed by the CD8\u3b1+ subset of conventional DCs. We characterize the antigen presenting properties of this receptor which help to delineate a novel function of CD36 in adaptive immunity. We show that targeting CD36 on DCs results in the delivery of exogenous Ags to both the MHC class-I and MHC class-II processing pathways. In addition, immunization with the recombinant anti CD36-Ag fusion Ab induces the robust activation of na\uefve CD4+ and CD8+ Ag specific T lymphocytes and the differentiation of primed CD8+ T cells into long term effector CTLs. Finally, we demonstrate that in vivo targeting of CD8\u3b1+ DCs with anti-CD36 Ab elicits humoral and cell mediated protection from the growth of an Ag specific tumor. Collectively, these results identify CD36 as an appropriate receptor to better elucidate the properties of the lymphoid organ resident CD8+ DCs and indicate it as a novel potential target for cancer immunotherapy

Update on the diagnosis of tuberculosis

Background Tuberculosis remains a global public health threat, and the development of rapid and precise diagnostic tools is the key to enabling the early start of treatment, monitoring response to treatment, and preventing the spread of the disease. Objective An overview of recent progress in host- and pathogen-based tuberculosis diagnostics. Sources We conducted a PubMed search of recent relevant articles and guidelines on tuberculosis screening and diagnosis. Content An overview of currently used methods and perspectives in the following areas of tuberculosis diagnostics is provided: immune-based diagnostics, X-ray, clinical symptoms and scores, cough detection, culture of Mycobacterium tuberculosis and identifying its resistance profile using phenotypic and genotypic methods, including next generation sequencing, sputum- and non-sputum-based molecular diagnosis of tuberculosis and monitoring of response to treatment. Implications A brief overview of the most relevant advances and changes in international guidelines regarding screening and diagnosing tuberculosis is provided in this review. It aims at reviewing all relevant areas of diagnostics, including both pathogen- and host-based methods.PostprintPeer reviewe

Content, mineral allocation and leaching behavior of heavy metals in urban PM2.5

This work provides a novel perspective in the field of urban airborne particle investigation that is not currently found in the literature. Four sampling campaigns were performed in the urban area of Rome (Central Italy) during the winter and summer seasons (February and July 2013 and 2014, respectively). The measured concentrations of the regulated 34 elements of As, Cd, Ni and Pb were consistent with those reported by the local Environmental Agency (ARPA Lazio), but non-regulated heavy metals, including Fe, Cu, Cr and Zn, were also found in PM2.5 and analyzed in detail. As an novelty, heavy metals were associated with the host-identified mineral phases, primarily oxides and alloys, and to a lesser extent, other minerals, such as sulfates, carbonates and silicates. Leaching tests of the collected samples were conducted in a buffered solution mimicking the bodily physiological environment. Despite the highest concentration of heavy metals found during the winter sampling period, all of the elements showed a leaching trend leading to major mobility during the summer period. To explain this result, an interesting comparative analysis between the leaching test behavior and innovative mineral allocation was conducted. Both the heavy metal content and mineral allocation in PM2.5 might contribute to the bioavailability of toxic elements in the pulmonary environment. Hence, for regulatory purposes, the non-linear dependency of heavy metal bioavailability on the total metal content should be taken into account

Towards standardisation:comparison of five whole genome sequencing (WGS) analysis pipelines for detection of epidemiologically linked tuberculosis cases

BackgroundWhole genome sequencing (WGS) is a reliable tool for studying tuberculosis (TB) transmission. WGS data are usually processed by custom-built analysis pipelines with little standardisation between them.AimTo compare the impact of variability of several WGS analysis pipelines used internationally to detect epidemiologically linked TB cases.MethodsFrom the Netherlands, 535 Mycobacterium tuberculosis complex (MTBC) strains from 2016 were included. Epidemiological information obtained from municipal health services was available for all mycobacterial interspersed repeat unit-variable number of tandem repeat (MIRU-VNTR) clustered cases. WGS data was analysed using five different pipelines: one core genome multilocus sequence typing (cgMLST) approach and four single nucleotide polymorphism (SNP)-based pipelines developed in Oxford, United Kingdom; Borstel, Germany; Bilthoven, the Netherlands and Copenhagen, Denmark. WGS clusters were defined using a maximum pairwise distance of 12 SNPs/alleles.ResultsThe cgMLST approach and Oxford pipeline clustered all epidemiologically linked cases, however, in the other three SNP-based pipelines one epidemiological link was missed due to insufficient coverage. In general, the genetic distances varied between pipelines, reflecting different clustering rates: the cgMLST approach clustered 92 cases, followed by 84, 83, 83 and 82 cases in the SNP-based pipelines from Copenhagen, Oxford, Borstel and Bilthoven respectively.ConclusionConcordance in ruling out epidemiological links was high between pipelines, which is an important step in the international validation of WGS data analysis. To increase accuracy in identifying TB transmission clusters, standardisation of crucial WGS criteria and creation of a reference database of representative MTBC sequences would be advisable

Coordinate regulation of tissue macrophage and dendritic cell population dynamics by CSF-1

CSF-1 drives the homeostatic expansion of macrophages within the growing myometrium of pregnant mice by stimulating in situ proliferation and inducing monocyte precursor recruitment from the blood

A cluster of multidrug-resistant Mycobacterium tuberculosis among patients arriving in Europe from the Horn of Africa: a molecular epidemiological study

SummaryBackground The risk of tuberculosis outbreaks among people fleeing hardship for refuge in Europe is heightened. We describe the cross-border European response to an outbreak of multidrug-resistant tuberculosis among patients from the Horn of Africa and Sudan. Methods On April 29 and May 30, 2016, the Swiss and German National Mycobacterial Reference Laboratories independently triggered an outbreak investigation after four patients were diagnosed with multidrug-resistant tuberculosis. In this molecular epidemiological study, we prospectively defined outbreak cases with 24-locus mycobacterial interspersed repetitive unit-variable number tandem repeat (MIRU-VNTR) profiles; phenotypic resistance to isoniazid, rifampicin, ethambutol, pyrazinamide, and capreomycin; and corresponding drug resistance mutations. We whole-genome sequenced all Mycobacterium tuberculosis isolates and clustered them using a threshold of five single nucleotide polymorphisms (SNPs). We collated epidemiological data from host countries from the European Centre for Disease Prevention and Control. Findings Between Feb 12, 2016, and April 19, 2017, 29 patients were diagnosed with multidrug-resistant tuberculosis in seven European countries. All originated from the Horn of Africa or Sudan, with all isolates two SNPs or fewer apart. 22 (76%) patients reported their travel routes, with clear spatiotemporal overlap between routes. We identified a further 29 MIRU-VNTR-linked cases from the Horn of Africa that predated the outbreak, but all were more than five SNPs from the outbreak. However all 58 isolates shared a capreomycin resistance-associated tlyA mutation. Interpretation Our data suggest that source cases are linked to an M tuberculosis clone circulating in northern Somalia or Djibouti and that transmission probably occurred en route before arrival in Europe. We hypothesise that the shared mutation of tlyA is a drug resistance mutation and phylogenetic marker, the first of its kind in M tuberculosis sensu stricto. Funding The Swiss Federal Office of Public Health, the University of Zurich, the Wellcome Trust, National Institute for Health Research (NIHR) Oxford Biomedical Research Centre (BRC), the Medical Research Council, BELTA-TBnet, the European Union, the German Center for Infection Research, and Leibniz Science Campus Evolutionary Medicine of the Lung (EvoLUNG)

Updating the approaches to define susceptibility and resistance to anti-tuberculosis agents: implications for diagnosis and treatment

11 páginas, 2 figuras, 1 tablaInappropriately high breakpoints have resulted in systematic false-susceptible AST results to anti-TB drugs. MIC, PK/PD and clinical outcome data should be combined when setting breakpoints to minimise the emergence and spread of antimicrobial resistance.I. Comas was supported by PID2019-104477RB-I00 from the Spanish Science Ministry and by ERC (CoG 101001038)Peer reviewe

Reaching consensus on drug resistance conferring mutations (Part 1)

Objective/background: Molecular-based, rapid drug-susceptibility tests are needed to guide the appropriate use of new drugs and new therapeutic regimens at the programmatic level, and to prevent a further increase in the incidence of drug-resistant tuberculosis (TB). Experts have recognized the need for a global, curated, and standardized analysis and data-sharing platform that provides a one-stop data source for clinically relevant genotypic and phenotypic information on Mycobacterium tuberculosis. Methods: To this purpose, the Relational Sequencing TB Data Platform (ReSeqTB) consortium has critically reviewed the most inclusive set of published data on mutations associated with drug resistance in M. tuberculosis to date, and has graded a comprehensive list of globally prevalent mutations based on the strength of their association with drug resistance. Results and Conclusions: ReSeqTB serves as a single repository for the compilation, curation, and validation of existing and newly created sequences and metadata on M. tuberculosis strains and will use the currently reviewed data set, validated by international experts, as a starting point until sufficient new sequence data are accumulated. This initiative is supported by a global partnership of academic institutions, public health agencies, and nongovernmental organizations including the Critical Path Institute, FIND, the World Health Organization, the New Diagnostics Working Group, the U.S. Centers for Disease Control and Prevention, and the National Institute of Allergy and Infectious Diseases and it is financially supported by the Bill & Melinda Gates Foundation. Key strengths of the ReSeqTB Database include the following: A user-friendly interface designed for nonexpert or expert operability.A standardized and validated analysis pipeline for variant analyses of M. tuberculosis next-generation sequencing (NGS) data.Access to data beyond the published literature with dynamic and iterative updates of new data generated by global surveillance and clinical trials.A well-developed legal structure to ensure intellectual property rights and data ownership remain with contributors.A structured data-sharing architecture to restrict access to sensitive or unpublished data sets.Metadata standardization using CDISC: supports global, platform-independent data standards that enable information system interoperability.An emphasis on data quality and rigorous, expert curation with multiple quality control checks for whole-genome sequencing and other metadata.Validation of NGS analysis output by an expert committee with grading of resistance conferring mutations based on rigorous statistical standards.Regulatory-compliant analysis pipeline and database architecture. Successful execution of such an extensive database platform requires substantial collaboration from scientists investigating the genetic basis for drug resistance worldwide, and from developers with expertise in database design and implementation