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Abstract In the tumour microenvironment, IL-1α promotes neoangiogenesis, matrix remodelling, tumour proliferation, chemoresistance, and metastases. Highly expressed in human colorectal cancers, IL-1α is associated with poor prognosis. XB2001, a fully human monoclonal antibody neutralizing IL-1α, was evaluated for safety and preliminary efficacy with trifluridine/tipiracil (FTD/TPI) and bevacizumab in metastatic colorectal cancer patients previously treated with oxaliplatin- and irinotecan-based chemotherapies. This single institution, phase 1 study used a 3 + 3 design to assess XB2001 at doses of 250 mg, 500 mg and 1000 mg every 14 days, associated with FTD/TPI 35 mg/m² (days 1–5 and 8-12, every 28 days) (NCT05201352). The Maximum Tolerated Dose of XB2001 + FTD/TPI was then associated in combination with bevacizumab (5 mg/kg, days 1 and 15). Safety, efficacy, pharmacokinetics and pharmacodynamics were assessed. Seventeen patients (median age: 67.4 years) were enroled. No patient exhibited dose-limiting toxicity at any dose. The most common treatment-related adverse events (TRAE) of any grade (G) were diarrhoea (35.3%), nausea (47.1%) and anaemia (35.3%). G3-4 TRAE were neutropenia (17.6%) hypertension and infection (5.9% each). The RP2D (recommended phase 2 dose) of XB2001 was 1000 mg. The disease control rate was 76%, with 23% of patients achieving an objective response, including one complete response. Response and longer progression-free survival were associated with a decrease in serum IL-6 levels during therapy. High intratumoral IL-1α expression at baseline and CD8/PD-L1 infiltration are associated with a better progression-free survival. The combination of XB2001 with FTD/TPI and bevacizumab is feasible and safe, and showed encouraging clinical activity in chemotherapy-resistant mCRC.

Abstract Glioblastoma multiforme (GBM) is the most common adult primary brain tumor. The standard of care involves maximal surgery followed by radiotherapy and concomitant chemotherapy with temozolomide (TMZ), in addition to adjuvant TMZ. However, the recurrence rate of GBM within 1–2 years post-diagnosis is still elevated and has been attributed to the accumulation of multiple factors including the heterogeneity of GBM, genomic instability, angiogenesis, and chronic tumor hypoxia. Tumor hypoxia activates downstream signaling pathways involved in the adaptation of GBM to the newly oxygen-deprived environment, thereby contributing to the resistance and recurrence phenomena, despite the multimodal therapeutic approach used to eradicate the tumor. Therefore, in this review, we will focus on the development and implication of chronic or limited-diffusion hypoxia in tumor persistence through genetic and epigenetic modifications. Then, we will detail the hypoxia-induced activation of vital biological pathways and mechanisms that contribute to GBM resistance. Finally, we will discuss a proteomics-based approach to encourage the implication of personalized GBM treatments based on a hypoxia signature.

Respiratory infections with respiratory syncytial virus (RSV) account for an important part of hospital admissions for acute respiratory infections. Nirsevimab has been developed to reduce the hospital burden of RSV infections. Compared with the product previously used, it has a stronger binding capacity to RSV F protein and a high affinity for FcRn (neonatal receptor for the Fc fragment of IgG), which extends its lifespan. Nirsevimab has been shown to be highly effective in reducing hospitalization rates of RSV infections but a large or unknown number of treated subjects have been excluded in clinical and post-marketing studies. However, analysis of these studies cannot exclude that, in rare cases, nirsevimab facilitates and worsens RSV infection (or other respiratory infections). This could be attributable to antibody-dependent enhancement (ADE) which has been observed with RSV F protein antibodies in inactivated vaccine trials. This risk has been incompletely assessed in pre-clinical and clinical trials (incomplete exploration of nirsevimab effector functions and pharmacokinetics). ADE by disruption of the immune system (not studied and due to FcRn binding) could explain why there is no reduction in all-cause hospital admissions in treated age groups. Given the high price of nirsevimab, the cost-effectiveness of mass immunization campaigns may therefore be debated from an economic as well as a scientific point of view.

Abstract Background Microsatellite instability (MSI) due to mismatch repair deficiency (dMMR) is common in colorectal cancer (CRC). These cancers are associated with somatic coding events, but the noncoding pathophysiological impact of this genomic instability is yet poorly understood. Here, we perform an analysis of coding and noncoding MSI events at the different steps of colorectal tumorigenesis using whole exome sequencing and search for associated splicing events via RNA sequencing at the bulk-tumor and single-cell levels. Results Our results demonstrate that MSI leads to hundreds of noncoding DNA mutations, notably at polypyrimidine U2AF RNA-binding sites which are endowed with cis-activity in splicing, while higher frequency of exon skipping events are observed in the mRNAs of MSI compared to non-MSI CRC. At the DNA level, these noncoding MSI mutations occur very early prior to cell transformation in the dMMR colonic crypt, accounting for only a fraction of the exon skipping in MSI CRC. At the RNA level, the aberrant exon skipping signature is likely to impair colonic cell differentiation in MSI CRC affecting the expression of alternative exons encoding protein isoforms governing cell fate, while also targeting constitutive exons, making dMMR cells immunogenic in early stage before the onset of coding mutations. This signature is characterized by its similarity to the oncogenic U2AF1-S34F splicing mutation observed in several other non-MSI cancer. Conclusions Overall, these findings provide evidence that a very early RNA splicing signature partly driven by MSI impairs cell differentiation and promotes MSI CRC initiation, far before coding mutations which accumulate later during MSI tumorigenesis.

How can inter-individual variability be quantified? Measuring many features per experiment raises the question of choosing them to recapitulate high-dimensional data. Tackling this challenge on spindle elongation phenotypes, we showed that only three typical elongation patterns describe spindle elongation in C. elegans one-cell embryo. These archetypes, automatically extracted from the experimental data using principal component analysis (PCA), accounted for more than 95% of inter-individual variability of more than 1600 experiments across more than 100 different conditions. The two first archetypes were related to spindle average length and anaphasic elongation rate. The third archetype, accounting for 6% of the variability, was novel and corresponded to a transient spindle shortening in late metaphase, reminiscent of kinetochore function-defect phenotypes. Importantly, these three archetypes were robust to the choice of the dataset and were found even considering only non-treated conditions. Thus, the inter-individual differences between genetically perturbed embryos have the same underlying nature as natural inter-individual differences between wild-type embryos, independently of the temperatures. We thus propose that beyond the apparent complexity of the spindle, only three independent mechanisms account for spindle elongation, weighted differently in the various conditions. Interestingly, the spindle-length archetypes covered both metaphase and anaphase, suggesting that spindle elongation in late metaphase is sufficient to predict the late anaphase length. We validated this idea using a machine-learning approach. Finally, given amounts of these three archetypes could represent a quantitative phenotype. To take advantage of this, we set out to predict interacting genes from a seed based on the PCA coefficients. We exemplified this firstly on the role of tpxl-1 whose homolog tpx2 is involved in spindle microtubule branching, secondly the mechanism regulating metaphase length, and thirdly the central spindle players which set the length at anaphase. We found novel interactors not in public databases but supported by recent experimental publications.

French Guiana, located in the Guiana Shield, is a natural reservoir for many zoonotic pathogens that are of considerable medical or veterinary importance. Until now, there has been limited data available on the description of parasites circulating in this area, especially on protozoan belonging to the phylum Apicomplexa; conversely, the neighbouring countries describe a high parasitic prevalence in animals and humans. Epidemiological surveillance is necessary, as new potentially virulent strains may emerge from these forest ecosystems, such as Amazonian toxoplasmosis. However, there is no standard tool for detecting protozoa in wildlife. In this study, we developed Meat-Borne-Parasite, a high-throughput meta-barcoding workflow for detecting Apicomplexa based on the Oxford Nanopore Technologies sequencing platform using the 18S gene of 14 Apicomplexa positive samples collected in French Guiana. Sequencing reads were then analysed with MetONTIIME pipeline. Thanks to a scoring rule, we were able to classify 10 samples out of 14 as Apicomplexa positive and reveal the presence of co-carriages. The same samples were also sequenced with the Illumina platform for validation purposes. For samples identified as Apicomplexa positive by both platforms, a strong positive correlation at up to the genus level was reported. Overall, the presented workflow represents a reliable method for Apicomplexa detection, which may pave the way for more comprehensive biomonitoring of zoonotic pathogens.

Amyotrophic Lateral Sclerosis (ALS) is the most common neurodegenerative disease affecting motor neurons. The pathophysiology of ALS is not well understood but TDP-43 proteinopathy (aggregation and mislocalization) is one of the major phenomena described. Several factors can influence TDP-43 behavior such as mild pH alterations that can induce conformational changes in recombinant TDP-43, increasing its propensity to aggregate. However to our knowledge, no studies have been conducted yet in a cellular setting, in the context of ALS. We therefore tested the effect of cellular pH alterations on the localization, aggregation, and phosphorylation of TDP-43. HEK293T cells overexpressing wildtype TDP-43 were incubated for 1 h with solutions of different pH (6.4, 7.2, and 8). Incubation of cells for 1 h in solutions of pH 6.4 and 8 led to an increase in TDP-43-positive puncta. This was accompanied by the mislocalization of TDP-43 from the nucleus to the cytoplasm. Our results suggest that small alterations in cellular pH affect TDP-43 and increase its mislocalization into cytoplasmic TDP-43-positive puncta, which might suggest a role of TDP-43 in the response of cells to pH alterations.

Circadian rhythms are intrinsic 24-hour oscillations found in nearly all life forms. They orchestrate key physiological and behavioral processes, allowing anticipation and response to daily environmental changes. These rhythms manifest across entire organisms, in various organs, and through intricate molecular feedback loops that govern cellular oscillations. Recent studies describe circadian regulation of pathogens, including parasites, bacteria, viruses, and fungi, some of which have their own circadian rhythms while others are influenced by the rhythmic environment of hosts. Pathogens target specific tissues and organs within the host to optimize their replication. Diverse cellular compositions and the interplay among various cell types create unique microenvironments in different tissues, and distinctive organs have unique circadian biology. Hence, residing pathogens are exposed to cyclic conditions, which can profoundly impact host-pathogen interactions. This review explores the influence of circadian rhythms and mammalian tissue-specific interactions on the dynamics of pathogen-host relationships. Overall, this demonstrates the intricate interplay between the body's internal timekeeping system and its susceptibility to pathogens, which has implications for the future of infectious disease research and treatment.

Abstract Fluctuations of consciousness and their rhythmicities have been rarely studied in patients with a disorder of consciousness after acute brain injuries. 24-h assessment of brain (EEG), behaviour (eye-opening), and circadian (clock-controlled hormones secretion from urine) functions was performed in acute brain-injured patients. The distribution, long-term predictability, and rhythmicity (circadian/ultradian) of various EEG features were compared with the initial clinical status, the functional outcome, and the circadian rhythmicities of behaviour and clock-controlled hormones. Here we show that more physiological and favourable patterns of fluctuations are associated with a higher 24 h predictability and sharp up-and-down shape of EEG switches, reminiscent of the Flip-Flop model of sleep. Multimodal rhythmic analysis shows that patients with simultaneous circadian rhythmicity for brain, behaviour, and hormones had a favourable outcome. Finally, both re-emerging EEG fluctuations and homogeneous 24-h cycles for EEG, eye-opening, and hormones appeared as surrogates for preserved functionality in brainstem and basal forebrain, which are key prognostic factors for later improvement. While the recovery of consciousness has previously been related to a high short-term complexity, we suggest in this exploratory study the importance of the high predictability of the 24 h long-term generation of brain rhythms and highlight the importance of circadian body-brain rhythms in awakening.

The mucosal pellicle (MP) is a biological film protecting the oral mucosa. It is composed of bounded salivary proteins and transmembrane mucin MUC1 expressed by oral epithelial cells. Previous research indicates that MUC1 expression enhances the binding of the main salivary protein forming the MP, MUC5B. This study investigated the influence of MUC1 structure on MP formation. A TR146 cell line, which does not express MUC1 natively, was stably transfected with genes coding for three MUC1 isoforms differing in the structure of the two main extracellular domains: the VNTR domain, exhibiting a variable number of tandem repeats, and the SEA domain, maintaining the two bound subunits of MUC1. Semi-quantification of MUC1 using dot blot chemiluminescence showed comparable expression levels in all transfected cell lines. Semi-quantification of MUC5B by immunostaining after incubation with saliva revealed that MUC1 expression significantly increased MUC5B adsorption. Neither the VNTR domain nor the SEA domain was influenced MUC5B anchoring, suggesting the key role of the MUC1 N-terminal domain. AFM-IR nanospectroscopy revealed discernible shifts indicative of changes in the chemical properties at the cell surface due to the expression of the MUC1 isoform. Furthermore, the observed chemical shifts suggest the involvement of hydrophobic effects in the interaction between MUC1 and salivary proteins.

Cancer stem cells are a subpopulation of tumor cells characterized by their ability to self-renew, induce tumors upon engraftment in animals and exhibit strong resistance to chemotherapy and radiotherapy. These cells exhibit numerous characteristics in common with embryonic stem cells, expressing some of their markers, typically absent in non-pathological adult differentiated cells. The aim of this study was to investigate the potential of conditioned media from cancer stem cells to modulate the fate of Leukemia Inhibitory Factor (LIF)-dependent murine embryonic stem cells (mESCs) as a way to obtain a direct readout of the secretome of cancer cells. A functional assay, “the StemDif sensor test”, was developed with two types of cancer stem cells derived from grade IV glioblastoma (adult and pediatric) or from gastric adenocarcinoma. We show that conditioned media from the selection of adult but not pediatric Glioma-Inducing Cells (GICs) maintain mESCs’ pluripotency in correlation with LIF secretion and activation of STAT3 protein. In contrast, conditioned media from gastric adenocarcinoma cells display LIF-independent stemness and differentiation activities on mESC. Our test stands out for its user-friendly procedures, affordability and straightforward output, positioning it as a pioneering tool for in-depth exploration of cancer stem cell secretome characteristics.

Platelet-rich plasma (PRP) is a rising therapy treating locomotor system lesions such as knee osteoarthritis. The objective of this study was to evaluate patients’ satisfaction 6 to 12 months after a PRP injection for cartilage lesions of their knee under real-life conditions. Patients’ satisfaction was assessed by a specific questionnaire named SATMED-Q©, which explores six different dimensions of a given treatment. In addition, pain and function were assessed thanks to VAS pain, WOMAC, and IKDC scores. Responders were identified through the OMERACT-OARSI criteria. We observed excellent satisfaction after a PRP injection with a SATMED-Q© score of 80.81% 6 to 12 months after the procedure. Even when there was no significant improvement in pain and function scores, 52% of the evaluated population fulfilled the OMERACT-OARSI criteria and were considered responders. According to the sub-group analysis, patients with less osteoarthritis damage (i.e., Kellgren–Lawrence grades 1–2) and older study subjects (i.e., >40 years old) with focal chondropathy had benefited most from their PRP injection. Thus, platelet-rich plasma seems to be a well-tolerated and efficient therapy for cartilage lesions of the knee.

Cellular senescence is a reprogrammed cell state triggered as an adaptative response to a variety of stresses, most often those affecting the genome integrity. Senescent cells accumulate in most tissues with age and contribute to the development of several pathologies. Studying molecular pathways involved in senescence induction and maintenance, or in senescence escape, can be hindered by the heterogeneity of senescent cell populations. Here, we describe a flow cytometry strategy for sorting senescent cells according to three senescence canonical markers whose thresholds can be independently adapted to be more or less stringent: (i) the senescence-associated-β-galactosidase (SA-β-Gal) activity, detected using 5-dodecanoylaminofluorescein Di-β-D-galactopyranoside (C12FDG), a fluorigenic substrate of β-galactosidase; (ii) cell size, proportional to the forward scatter value, since increased size is one of the major changes observed in senescent cells; and (iii) cell granularity, proportional to the side scatter value, which reflects the accumulation of aggregates, lysosomes, and altered mitochondria in senescent cells. We applied this protocol to the sorting of normal human fibroblasts at the replicative senescence plateau. We highlighted the challenge of sorting these senescent cells because of their large sizes, and established that it requires using sorters equipped with a nozzle of an unusually large diameter: at least 200 µm. We present evidence of the sorting efficiency and sorted cell viability, as well as of the senescent nature of the sorted cells, confirmed by the detection of other senescence markers, including the expression of the CKI p21 and the presence of 53BP1 DNA damage foci. Our protocol makes it possible, for the first time, to sort senescent cells from contaminating proliferating cells and, at the same time, to sort subpopulations of senescent cells featuring senescent markers to different extents.Graphical abstract

Chromosome segregation requires both the separation of sister chromatids and the sustained condensation of chromatids during anaphase. In yeast cells, cohesin is not only required for sister chromatid cohesion but also plays a major role determining the structure of individual chromatids in metaphase. Separase cleavage is thought to remove all cohesin complexes from chromosomes to initiate anaphase. It is thus not clear how the length and organisation of segregating chromatids is maintained during anaphase in the absence of cohesin. Here, we show that degradation of cohesin at the anaphase onset causes aberrant chromatid segregation. Hi-C analysis on segregating chromatids demonstrates that cohesin depletion causes loss of intrachromatid organisation. Surprisingly, tobacco etch virus (TEV)-mediated cleavage of cohesin does not dramatically disrupt chromatid organisation in anaphase, explaining why bulk segregation is achieved. In addition, we identified a small pool of cohesin complexes bound to telophase chromosomes in wild-type cells and show that they play a role in the organisation of centromeric regions. Our data demonstrates that in yeast cells cohesin function is not over in metaphase, but extends to the anaphase period when chromatids are segregating.

Radiation therapy and platinum-based chemotherapy are common treatments for lung cancer patients. Several factors are considered for the low overall survival rate of lung cancer, such as the patient’s physical state and the complex heterogeneity of the tumor, which leads to resistance to the treatment. Consequently, precision medicines are needed for the patients to improve their survival and their quality of life. Until now, no patient-derived tumoroid model has been reported to predict the efficiency of radiation therapy in non-small-cell lung cancer. Using our patient-derived tumoroid model, we report that this model could be used to evaluate the efficiency of radiation therapy and cisplatin-based chemotherapy in non-small-cell lung cancer. In addition, these results can be correlated to clinical outcomes of patients, indicating that this patient-derived tumoroid model can predict the response to radiotherapy and chemotherapy in non-small-cell lung cancer.

Jeremy Gauthier et al. present the chromosome scale assembly of the genome of the parasitic wasp C. congregata and show that bracovirus genes have colonized all ten chromosomes. Comparison with genome scaffolds of another wasp reveals a striking stability of these regions over ~53 million years, suggesting strong evolutionary constraints.

Abstract Background A key step in domestication of the grapevine was the transition from separate sexes (dioecy) in wild Vitis vinifera ssp. sylvestris (V. sylvestris) to hermaphroditism in cultivated Vitis vinifera ssp. sativa (V. vinifera). It is known that V. sylvestris has an XY system and V. vinifera a modified Y haplotype (Yh) and that the sex locus is small, but it has not previously been precisely characterized. Results We generate a high-quality de novo reference genome for V. sylvestris, onto which we map whole-genome re-sequencing data of a cross to locate the sex locus. Assembly of the full X, Y, and Yh haplotypes of V. sylvestris and V. vinifera sex locus and examining their gene content and expression profiles during flower development in wild and cultivated accessions show that truncation and deletion of tapetum and pollen development genes on the X haplotype likely causes male sterility, while the upregulation of a Y allele of a cytokinin regulator (APRT3) may cause female sterility. The downregulation of this cytokinin regulator in the Yh haplotype may be sufficient to trigger reversal to hermaphroditism. Molecular dating of X and Y haplotypes is consistent with the sex locus being as old as the Vitis genus, but the mechanism by which recombination was suppressed remains undetermined. Conclusions We describe the genomic and evolutionary characterization of the sex locus of cultivated and wild grapevine, providing a coherent model of sex determination in the latter and for transition from dioecy to hermaphroditism during domestication.

Summary: During normal mammary gland development, s-SHIP promoter expression marks a distinct type of mammary stem cells, at two different stages, puberty and early mid-pregnancy. To determine whether s-SHIP is a marker of mammary cancer stem cells (CSCs), we generated bitransgenic mice by crossing the C3(1)-SV40 T-antigen transgenic mouse model of breast cancer, and a transgenic mouse (11.5kb-GFP) expressing green fluorescent protein from the s-SHIP promoter. Here we show that in mammary tumors originating in these bitransgenic mice, s-SHIP promoter expression enriches a rare cell population with CSC activity as demonstrated by sphere-forming assays in vitro and limiting dilution transplantation in vivo. These s-SHIP-positive CSCs are characterized by lower expression of Delta-like non-canonical Notch ligand 1 (DLK1), a negative regulator of the Notch pathway. Inactivation of Dlk1 in s-SHIP-negative tumor cells increases their tumorigenic potential, suggesting a role for DLK1 in mammary cancer stemness. : Bourette and colleagues assess renewal and differentiation potential of s-SHIP-expressing cells in mouse mammary tumors, by in vitro sphere-forming assay and in vivo transplantation. Their data indicate that s-SHIP expression marks a distinct population of mammary cancer stem cells and unveils a potential role of Dlk1, a negative regulator of the Notch pathway, in the regulation of mammary tumor cell stemness. Keywords: breast cancer, CSC, s-SHIP, transgenic mouse, mammary tumor, DLK1, notch, SHIP1

Abstract Objective We recently investigated the pathways of immunoreactive bradykinin (iBK) formation in fresh blood of normal volunteers and of patients with hereditary angioedema due to C1-esterase inhibitor deficiency (HAE-1/-2). Herein, we adapted the techniques to small volumes (200 μl) of previously frozen citrated plasma and further analyzed the mechanisms of iBK formation with additional biotechnological inhibitors. Results Measurable iBK formation was observed under stimulation with tissue kallikrein (KLK-1, 10 nM), the particulate material Kontact-APTT (concentration reduced to 2% v/v) or recombinant tissue plasminogen activator (tPA, 169 nM), with little background in unstimulated plasma incubated for up to 2 h. Plasma samples from HAE-1/-2 patients responded earlier to tPA than those from controls, as previously reported with whole blood. Lanadelumab inhibited iBK formation induced by Kontact-APTT and tPA. A highly specific plasmin inhibitor, DX-1000, abolished tPA-induced iBK formation in plasma but had no effect against Kontact-APTT, confirming the role of fibrinolysis in tPA-induced kinin formation. The anti-lanadelumab neutralizing antibody M293-D02 reversed the inhibitory effects of lanadelumab. Frozen plasma is a suitable material for measuring iBK formation kinetics, with possible applications such as investigating the effect of rare disease states on the kallikrein–kinin system and monitoring the effect of HAE prophylactic treatments.

Summary: Neurodegenerative disorders are an increasingly common and irreversible burden on society, often affecting the aging population, but their etiology and disease mechanisms are poorly understood. Studying monogenic neurodegenerative diseases with known genetic cause provides an opportunity to understand cellular mechanisms also affected in more complex disorders. We recently reported that loss-of-function mutations in the autophagy adaptor protein SQSTM1/p62 lead to a slowly progressive neurodegenerative disease presenting in childhood. To further elucidate the neuronal involvement, we studied the cellular consequences of loss of p62 in a neuroepithelial stem cell (NESC) model and differentiated neurons derived from reprogrammed p62 patient cells or by CRISPR/Cas9-directed gene editing in NESCs. Transcriptomic and proteomic analyses suggest that p62 is essential for neuronal differentiation by controlling the metabolic shift from aerobic glycolysis to oxidative phosphorylation required for neuronal maturation. This shift is blocked by the failure to sufficiently downregulate lactate dehydrogenase expression due to the loss of p62, possibly through impaired Hif-1α downregulation and increased sensitivity to oxidative stress. The findings imply an important role for p62 in neuronal energy metabolism and particularly in the regulation of the shift between glycolysis and oxidative phosphorylation required for normal neurodifferentiation. : SQSTM1/p62 is a known autophagy adaptor that, if lost, causes childhood-onset neurodegeneration. Data from Wredenberg et al. show that loss of p62 in a neuronal stem cell model does not affect mitophagy but instead leads to impaired differentiation. The authors suggest p62 finely tunes LDHA expression and thus controls the metabolic shift to OXPHOS required for proper differentiation. Keywords: SQSTM1, p62, hypoxia, mitochondria, neurodifferentiation, neuroepithelial-like stem cells, neuronal development, oxidative stress, mitophagy, neurodegeneration

After death, diagenesis takes place. Numerous processes occur concomitantly, which makes it difficult to identify the diagenetic processes. The diagenetic processes refer to all processes (chemical or physical) that modify the skeletal remains. These processes are highly variable depending on the environmental factors (weather, temperature, age, sex, etc.), especially in the early stages. Numerous studies have evaluated bone diagenetic processes over long timescales (~millions of years), but fewer have been done over short timescales (between days and thousands of years). The objective of the study is to assess the early stages of diagenetic processes by Raman microspectroscopy over 12 months. The mineral and organic matrix modifications are monitored through physicochemical parameters. Ribs from six humans were buried in soil. The modifications of bone composition were followed by Raman spectroscopy each month. The decrease in the mineral/organic ratio and carbonate type-B content and the increase in crystallinity reveal that minerals undergo dissolution–recrystallization. The decrease in collagen cross-linking indicates that collagen hydrolysis induces the fragmentation of collagen fibres over 12 months.

Background: Patients living with HIV (PLWHIV) can develop autoimmune diseases (AD) needing immunosuppressive treatments (IST). This study aims to describe the impact of IST in PLWHIV. Methods: This was a multicentric retrospective observational study in six HIV referral centers on PLWHIV under IST for AD. Demographic factors, viral co-infections, immunovirological status before and under IST, infectious events, and their descriptions were collected and described focusing on infectious events, immunovirological variations, and IST effectiveness. Results: 9480 PLWHIV were screened for inclusion. Among them, 138 (1.5%) had a history of auto-immune disease, among which 32 (23%) received IST. There was mainly spondyloarthropathy (28%) and the most commonly used IST was methotrexate. The median follow-up under IST was 3.8 years (2.7; 5.9). There were 15 infectious events (0.5 events/individuals) concerning nine patients. At the last medical follow-up, 81% of these were in remission of their AD. Under IST, there was an increase in CD4 during follow-up (629 vs. 827 CD4/mm3, p = 0.04). No HIV virological failure was noted. Conclusions: This study supports a growing evidence base that IST can be used safely and effectively in PLWHIV with careful monitoring.

Diverse biomarkers and pathological alterations have been found in muscle of patients with Amyotrophic lateral sclerosis (ALS), but the relation between such alterations and dysfunction in energetic metabolism remains to be investigated. We established the metabolome of muscle and serum of ALS patients and correlated these findings with the clinical status and pathological alterations observed in the muscle. We obtained data from 20 controls and 17 ALS patients (disease duration: 9.4 ± 6.8 months). Multivariate metabolomics analysis identified a distinct serum metabolome for ALS compared to controls (p-CV-ANOVA < 0.035) and revealed an excellent discriminant profile for muscle metabolome (p-CV-ANOVA < 0.0012). Citramalate was discriminant for both muscle and serum. High lauroylcarnitine levels in muscle were associated with low Forced Vital Capacity. Transcriptomics analysis of key antioxidant enzymes showed an upregulation of SOD3 (p = 0.0017) and GLRX2(1) (p = 0.0022) in ALS muscle. Analysis of mitochondrial enzymatic activity in muscle revealed higher complex II/CS (p = 0.04) and lower LDH (p = 0.03) activity in ALS than in controls. Our study showed, for the first time, a global dysfunction in the muscle of early-stage ALS patients. Furthermore, we identified novel metabolites to be employed as biomarkers for diagnosis and prognosis of ALS patients.

White-handed gibbons produce loud and acoustically complex songs when interacting with their neighbours or when encountering predators. In both contexts, songs are assembled from a small number of units although their composition differs in context-specific ways. Here, we investigated whether wild gibbons could infer the ‘meaning’ when hearing exemplars recorded in both contexts (i.e. ‘duet songs’ vs. ‘predator songs’). We carried out a playback experiment by which we simulated the presence of a neighbouring group producing either its duet or a predator song in order to compare subjects’ vocal and locomotor responses. When hearing a recording of a duet song, subjects reliably responded with their own duet song, which sometimes elicited further duet songs in adjacent groups. When hearing a recording of a predator song, however, subjects typically remained silent, apart from one of six groups which replied with its own predator song. Moreover, in two of six trials, playbacks of predator songs elicited predator song replies in non-adjacent groups. Finally, all groups showed strong anti-predator behaviour to predator songs but never to duet songs. We concluded that white-handed gibbons discriminated between the two song types and were able to infer meaning from them. We discuss the implications of these findings in light of the current debate on the evolutionary origins of syntax.

Background Quantitative real-time PCR (qPCR) is a well-established method for detecting and quantifying bacteria, and it is progressively replacing culture-based diagnostic methods in food microbiology. High-throughput qPCR using microfluidics brings further advantages by providing faster results, decreasing the costs per sample and reducing errors due to automatic distribution of samples and reagents. In order to develop a high-throughput qPCR approach for the rapid and cost-efficient quantification of microbial species in complex systems such as fermented foods (for instance, cheese), the preliminary setup of qPCR assays working efficiently under identical PCR conditions is required. Identification of target-specific nucleotide sequences and design of specific primers are the most challenging steps in this process. To date, most available tools for primer design require either laborious manual manipulation or high-performance computing systems. Results We developed the SpeciesPrimer pipeline for automated high-throughput screening of species-specific target regions and the design of dedicated primers. Using SpeciesPrimer, specific primers were designed for four bacterial species of importance in cheese quality control, namely Enterococcus faecium, Enterococcus faecalis, Pediococcus acidilactici and Pediococcus pentosaceus. Selected primers were first evaluated in silico and subsequently in vitro using DNA from pure cultures of a variety of strains found in dairy products. Specific qPCR assays were developed and validated, satisfying the criteria of inclusivity, exclusivity and amplification efficiencies. Conclusion In this work, we present the SpeciesPrimer pipeline, a tool to design species-specific primers for the detection and quantification of bacterial species. We use SpeciesPrimer to design qPCR assays for four bacterial species and describe a workflow to evaluate the designed primers. SpeciesPrimer facilitates efficient primer design for species-specific quantification, paving the way for a fast and accurate quantitative investigation of microbial communities.

Autism Spectrum Disorders (ASDs) are neurodevelopmental disorders defined by impaired social interactions and communication with repetitive behaviors, activities, or interests. Gastrointestinal (GI) disturbances and gut microbiota dysbiosis are frequently associated with ASD in childhood. However, it is not known whether microbiota dysbiosis in ASD patients also occurs in adulthood. Further, the consequences of altered gut microbiota on digestive functions and the enteric nervous system (ENS) remain unexplored. Therefore, we studied, in mice, the ability offecal supernatant (FS) from adult ASD patients to induce GI dysfunctions and ENS remodeling. First, the analyses of the fecal microbiota composition in adult ASD patients indicated a reduced α-diversity and increased abundance of three bacterial 16S rRNA gene amplicon sequence variants compared to healthy controls (HC). The transfer of FS from ASD patients (FS–ASD) to mice decreased colonic barrier permeability by 29% and 58% compared to FS–HC for paracellular and transcellular permeability, respectively. These effects are associated with the reduced expression of the tight junction proteins JAM-A, ZO-2, cingulin, and proinflammatory cytokines TNFα and IL1β. In addition, the expression of glial and neuronal molecules was reduced by FS–ASD as compared to FS-HC in particular for those involved in neuronal connectivity (βIII-tubulin and synapsin decreased by 31% and 67%, respectively). Our data suggest that changes in microbiota composition in ASD may contribute to GI alterations, and in part, via ENS remodeling.

Activated fibroblasts are considered major drivers of fibrotic disease progression through the production of excessive extracellular matrix (ECM) in response to signals from damaged epithelial and inflammatory cells. Nevertheless, epithelial cells are capable of expressing components of the ECM, cross-linking enzymes that increase its stability and are sensitive to factors involved in the early stages of fibrosis. We therefore wanted to test the hypothesis that epithelial cells can deposit ECM in response to stimulation in a comparable manner to fibroblasts. We performed immunofluorescence analysis of components of stable, mature extracellular matrix produced by primary human renal proximal tubular epithelial cells and renal fibroblasts in response to cytokine stimulation. Whilst fibroblasts produced a higher basal level of extracellular matrix components, epithelial cells were able to deposit significant levels of fibronectin, collagen I, III and IV in response to cytokine stimulation. In response to hypoxia, epithelial cells showed an increase in collagen IV deposition but not in response to the acute stress stimuli aristolochic acid or hydrogen peroxide. When epithelial cells were in co-culture with fibroblasts we observed significant increases in the level of matrix deposition which could be reduced by transforming growth factor beta (TGF-β) blockade. Our results highlight the role of epithelial cells acting as efficient producers of stable extracellular matrix which could contribute to renal tubule thickening in fibrosis.

Sirtuin-1 deacetylates proteins and has emerged as a critical regulator of different cellular processes, particularly inflammation. Basal SIRT1 activity was previously found to limit Th9 and enhance Th17 differentiation in mice, but the effect of pharmacological SIRT1 activation on T cell differentiation and antitumor responses remains unclear. Here, we find that SIRT1 pharmacological agonists selectively impede mouse and human Th17 cell differentiation. SIRT1 activation induces STAT3 deacetylation, thus reducing its ability to translocate into the nucleus, bind to Rorc promoter, and induce its transcription. SIRT1 agonists reduce tumor growth in mice by blocking Th17 cell differentiation. In cancer patients, the SIRT1 agonist metformin reduced the frequency of Th17 cells and STAT3 acetylation levels. Altogether, these data underscore that SIRT1 activation impedes Th17 cell differentiation and thereby limits tumor growth and suggest that SIRT1 activators may directly target IL-17A functions.

The ability of Mycobacterium tuberculosis (Mtb) to persist in its host is central to the pathogenesis of tuberculosis, yet the underlying mechanisms remain incompletely defined. PerM, an integral membrane protein, is required for persistence of Mtb in mice. Here, we show that perM deletion caused a cell division defect specifically during the chronic phase of mouse infection, but did not affect Mtb’s cell replication during acute infection. We further demonstrate that PerM is required for cell division in chronically infected mice and in vitro under host-relevant stresses because it is part of the mycobacterial divisome and stabilizes the essential divisome protein FtsB. These data highlight the importance of sustained cell division for Mtb persistence, define condition-specific requirements for cell division and reveal that survival of Mtb during chronic infection depends on a persistence divisome.

The research was carried out in order to assess the effects of nine cultivars in factorial combination with an open field or greenhouse growing on yield, quality indicators, antioxidants and elemental composition of leek in Moscow region. Greenhouse management resulted in higher yield compared to open field cultivation, due to higher mean pseudo-stem weight, and cultivar Giraffe gave the highest production. Pseudo-stem dry matter was better affected by greenhouse cultivation, whereas the content of monosaccharides, total sugars, nitrates, ascorbic acid, and polyphenols was enhanced by open field growing. The cultivars Vesta and Summer breeze showed the highest dry matter and total sugar content, whereas Goliath had the highest antioxidant, selenium, and potassium concentration. Among the mineral elements, K and Mg in pseudo-stems were better affected by greenhouse conditions, whereas Ca attained a higher concentration under open field growing. The antioxidant system of A porrum was characterized by significant positive correlations between Se, polyphenols, ascorbic acid, and potassium

Deforestation is a major threat to biodiversity, particularly within tropical forest habitats. Some of the fastest diminishing tropical forest habitats in the world occur in Indonesia, where fragmentation is severely impacting biodiversity, including on the island of Java, which holds many endemic species. Extreme fragmentation on the western part of the island, especially due to small-scale agriculture, impacts animal movement and increases mortality risk for mainly arboreal taxa. To mitigate this risk in an agroforest environment in Garut District, West Java, we installed 10 canopy bridges and monitored them through camera trapping between 2017 and 2019. Five of the monitored bridges were made of waterlines and five of rubber hose. We recorded Javan palm civets using the waterline bridges 938 times, while Javan slow lorises used the waterlines 1079 times and the rubber bridges 358 times. At least 19 other species used the bridges for crossing or perching. Our results demonstrate that relatively simple and cost-effective materials can be used to mitigate the effects of habitat fragmentation. We also recommend the use of camera traps to monitor the effectiveness of these interventions.

Summary: The lack of disease-modifying treatments for neurodegenerative disease stems in part from our rudimentary understanding of disease mechanisms and the paucity of targets for therapeutic intervention. Here we used an integrated discovery paradigm to identify a new therapeutic target for diseases caused by α-synuclein (α-syn), a small lipid-binding protein that misfolds and aggregates in Parkinson’s disease and other disorders. Using unbiased phenotypic screening, we identified a series of compounds that were cytoprotective against α-syn-mediated toxicity by inhibiting the highly conserved enzyme stearoyl-CoA desaturase (SCD). Critically, reducing the levels of unsaturated membrane lipids by inhibiting SCD reduced α-syn toxicity in human induced pluripotent stem cell (iPSC) neuronal models. Taken together, these findings suggest that inhibition of fatty acid desaturation has potential as a therapeutic approach for the treatment of Parkinson’s disease and other synucleinopathies. : There are no treatments that target the underlying cause of synucleinopathies such as Parkinson’s disease. Using unbiased small-molecule phenotypic screening in yeast, Vincent et al. identify a potential therapeutic target, stearoyl-CoA desaturase (Ole1/SCD). Inhibiting SCD in human-derived neurons enhances their survival in the prescence of toxic α-synuclein. Keywords: α-synuclein, Parkinson’s disease, stearoyl-CoA desaturase, fatty acid desaturation, phenotypic drug screen, target identification, chemical genetics, vesicle trafficking

Phospholipid membranes form cellular barriers but need to be flexible enough to divide by fission. Phospholipids generally contain a saturated fatty acid (FA) at position sn1 whereas the sn2-FA is saturated, monounsaturated or polyunsaturated. Our understanding of the impact of phospholipid unsaturation on membrane flexibility and fission is fragmentary. Here, we provide a comprehensive view of the effects of the FA profile of phospholipids on membrane vesiculation by dynamin and endophilin. Coupled to simulations, this analysis indicates that: (i) phospholipids with two polyunsaturated FAs make membranes prone to vesiculation but highly permeable; (ii) asymmetric sn1-saturated-sn2-polyunsaturated phospholipids provide a tradeoff between efficient membrane vesiculation and low membrane permeability; (iii) When incorporated into phospholipids, docosahexaenoic acid (DHA; omega-3) makes membranes more deformable than arachidonic acid (omega-6). These results suggest an explanation for the abundance of sn1-saturated-sn2-DHA phospholipids in synaptic membranes and for the importance of the omega-6/omega-3 ratio on neuronal functions.

Listeria monocytogenes causes listeriosis, a foodborne disease that poses serious risks to fetuses, newborns and immunocompromised adults. This intracellular bacterial pathogen proliferates in the host cytosol and exploits the host actin polymerization machinery to spread from cell-to-cell and disseminate in the host. Here, we report that during several days of infection in human hepatocytes or trophoblast cells, L. monocytogenes switches from this active motile lifestyle to a stage of persistence in vacuoles. Upon intercellular spread, bacteria gradually stopped producing the actin-nucleating protein ActA and became trapped in lysosome-like vacuoles termed Listeria-Containing Vacuoles (LisCVs). Subpopulations of bacteria resisted degradation in LisCVs and entered a slow/non-replicative state. During the subculture of host cells harboring LisCVs, bacteria showed a capacity to cycle between the vacuolar and the actin-based motility stages. When ActA was absent, such as in ΔactA mutants, vacuolar bacteria parasitized host cells in the so-called "viable but non-culturable" state (VBNC), preventing their detection by conventional colony counting methods. The exposure of infected cells to high doses of gentamicin did not trigger the formation of LisCVs, but selected for vacuolar and VBNC bacteria. Together, these results reveal the ability of L. monocytogenes to enter a persistent state in a subset of epithelial cells, which may favor the asymptomatic carriage of this pathogen, lengthen the incubation period of listeriosis, and promote bacterial survival during antibiotic therapy.

Sunflower is an important oilseed crop, as well as a model system for evolutionary studies, but its 3.6 gigabase genome has proven difficult to assemble, in part because of the high repeat content of its genome. Here we report on the sequencing, assembly, and analyses of 96 randomly chosen BACs from sunflower to provide additional information on the repeat content of the sunflower genome, assess how repetitive elements in the sunflower genome are organized relative to genes, and compare the genomic distribution of these repeats to that found in other food crops and model species. We also examine the expression of transposable element-related transcripts in EST databases for sunflower to determine the representation of repeats in the transcriptome and to measure their transcriptional activity. Our data confirm previous reports in suggesting that the sunflower genome is >78% repetitive. Sunflower repeats share very little similarity to other plant repeats such as those of Arabidopsis, rice, maize and wheat; overall 28% of repeats are “novel” to sunflower. The repetitive sequences appear to be randomly distributed within the sequenced BACs. Assuming the 96 BACs are representative of the genome as a whole, then approximately 5.2% of the sunflower genome comprises non TE-related genic sequence, with an average gene density of 18kbp/gene. Expression levels of these transposable elements indicate tissue specificity and differential expression in vegetative and reproductive tissues, suggesting that expressed TEs might contribute to sunflower development. The assembled BACs will also be useful for assessing the quality of several different draft assemblies of the sunflower genome and for annotating the reference sequence.

Abstract Living systems may have evolved probabilistic bet hedging strategies that generate cell‐to‐cell phenotypic diversity in anticipation of environmental catastrophes, as opposed to adaptation via a deterministic response to environmental changes. Evolution of bet hedging assumes that genotypes segregating in natural populations modulate the level of intraclonal diversity, which so far has largely remained hypothetical. Using a fluorescent Pmet17‐GFP reporter, we mapped four genetic loci conferring to a wild yeast strain an elevated cell‐to‐cell variability in the expression of MET17, a gene regulated by the methionine pathway. A frameshift mutation in the Erc1p transmembrane transporter, probably resulting from a release of laboratory strains from negative selection, reduced Pmet17‐GFP expression variability. At a second locus, cis‐regulatory polymorphisms increased mean expression of the Mup1p methionine permease, causing increased expression variability in trans. These results demonstrate that an expression quantitative trait locus (eQTL) can simultaneously have a deterministic effect in cis and a probabilistic effect in trans. Our observations indicate that the evolution of transmembrane transporter genes can tune intraclonal variation and may therefore be implicated in both reactive and anticipatory strategies of adaptation.

Maximakinin (MK), an amphibian peptide possessing the C-terminal sequence of bradykinin (BK), is a BK B2 receptor (B2R) agonist eliciting prolonged signaling. We reinvestigated this 19-mer for species-specific pharmacologic profile, in vivo confirmation of resistance to inactivation by angiotensin converting enzyme (ACE), value as a module for the design of fusion proteins that bind to the B2R in mammalian species and potential activity as a histamine releaser. Competition of the binding of [3H]BK to recombinant human myc-B2Rs in cells that express these receptors revealed that MK possessed a tenuous fraction (

Summary Changes in the physical properties of the lipid matrix of cell membranes have repeatedly been proposed to underlie stresses associated with e.g. drought, cold and xenobiotics. Therefore, the ability to experimentally monitor such properties is central to the fundamental physiological understanding of adaptive changes. Here, we test the analysis of shape fluctuations in membranes composed of lipid extracts from two soil invertebrates, and show that theories and experimental approaches previously developed for simpler liposomes may be applied directly to reconstituted membrane lipids. Specifically, we show how the bending rigidity of giant unilamellar liposomes of lipid extracts can be determined precisely. We suggest that future measurements of this parameter could elucidate mechanisms of adaptive processes such as changes in lipid composition and accumulation of protective osmolytes.

When small phosphatidylcholine liposomes are added to perforated cells, they bind preferentially to the Golgi suggesting an exceptional avidity of this organelle for curved membranes without stereospecific interactions. We show that the cis golgin GMAP-210 accounts for this property. First, the liposome tethering properties of the Golgi resembles that of the amphipathic lipid-packing sensor (ALPS) motif of GMAP-210: both preferred small (radius < 40 nm) liposomes made of monounsaturated but not saturated lipids. Second, reducing GMAP-210 levels or redirecting its ALPS motif to mitochondria decreased liposome capture by the Golgi. Extensive mutagenesis analysis suggests that GMAP-210 tethers authentic transport vesicles via the same mechanism whereby the ALPS motif senses lipid-packing defects at the vesicle surface through its regularly spaced hydrophobic residues. We conclude that the Golgi uses GMAP-210 as a filter to select transport vesicles according to their size and bulk lipid composition.

ISG15 is an interferon-stimulated, linear di-ubiquitin-like protein, with anti-viral activity. The role of ISG15 during bacterial infection remains elusive. We show that ISG15 expression in nonphagocytic cells is dramatically induced upon Listeria infection. Surprisingly this induction can be type I interferon independent and depends on the cytosolic surveillance pathway, which senses bacterial DNA and signals through STING, TBK1, IRF3 and IRF7. Most importantly, we observed that ISG15 expression restricts Listeria infection in vitro and in vivo. We made use of stable isotope labeling in tissue culture (SILAC) to identify ISGylated proteins that could be responsible for the protective effect. Strikingly, infection or overexpression of ISG15 leads to ISGylation of ER and Golgi proteins, which correlates with increased secretion of cytokines known to counteract infection. Together, our data reveal a previously uncharacterized ISG15-dependent restriction of Listeria infection, reinforcing the view that ISG15 is a key component of the innate immune response.

The identification of Mycobacterium tuberculosis genes necessary for persistence in vivo provides insight into bacterial biology as well as host defense strategies. We show that disruption of M. tuberculosis membrane protein PerM (Rv0955) resulted in an IFN-γ-dependent persistence defect in chronic mouse infection despite the mutant's near normal growth during acute infection. The perM mutant required increased magnesium for replication and survival; incubation in low magnesium media resulted in cell elongation and lysis. Transcriptome analysis of the perM mutant grown in reduced magnesium revealed upregulation of cell division and cell wall biosynthesis genes, and live cell imaging showed PerM accumulation at the division septa in M. smegmatis. The mutant was acutely sensitive to β-lactam antibiotics, including specific inhibitors of cell division-associated peptidoglycan transpeptidase FtsI. Together, these data implicate PerM as a novel player in mycobacterial cell division and pathogenesis, and are consistent with the hypothesis that immune activation deprives M. tuberculosis of magnesium.

Sleep spindles and K-complexes (KCs) define stage 2 NREM sleep (N2) in humans. We recently showed that KCs are isolated downstates characterized by widespread cortical silence. We demonstrate here that KCs can be quasi-synchronous across scalp EEG and across much of the cortex using electrocorticography (ECOG) and localized transcortical recordings (bipolar SEEG). We examine the mechanism of synchronous KC production by creating the first conductance based thalamocortical network model of N2 sleep to generate both spontaneous spindles and KCs. Spontaneous KCs are only observed when the model includes diffuse projections from restricted prefrontal areas to the thalamic reticular nucleus (RE), consistent with recent anatomical findings in rhesus monkeys. Modeled KCs begin with a spontaneous focal depolarization of the prefrontal neurons, followed by depolarization of the RE. Surprisingly, the RE depolarization leads to decreased firing due to disrupted spindling, which in turn is due to depolarization-induced inactivation of the low-threshold Ca2+ current (IT). Further, although the RE inhibits thalamocortical (TC) neurons, decreased RE firing causes decreased TC cell firing, again because of disrupted spindling. The resulting abrupt removal of excitatory input to cortical pyramidal neurons then leads to the downstate. Empirically, KCs may also be evoked by sensory stimuli while maintaining sleep. We reproduce this phenomenon in the model by depolarization of either the RE or the widely-projecting prefrontal neurons. Again, disruption of thalamic spindling plays a key role. Higher levels of RE stimulation also cause downstates, but by directly inhibiting the TC neurons. SEEG recordings from the thalamus and cortex in a single patient demonstrated the model prediction that thalamic spindling significantly decreases before KC onset. In conclusion, we show empirically that KCs can be widespread quasi-synchronous cortical downstates, and demonstrate with the first model of stage 2 NREM sleep a possible mechanism whereby this widespread synchrony may arise.

Listeria monocytogenes (Lm) is an invasive foodborne pathogen that leads to severe central nervous system and maternal-fetal infections. Lm ability to actively cross the intestinal barrier is one of its key pathogenic properties. Lm crosses the intestinal epithelium upon the interaction of its surface protein internalin (InlA) with its host receptor E-cadherin (Ecad). InlA-Ecad interaction is species-specific, does not occur in wild-type mice, but does in transgenic mice expressing human Ecad and knock-in mice expressing humanized mouse Ecad. To study listeriosis in wild-type mice, InlA has been "murinized" to interact with mouse Ecad. Here, we demonstrate that, unexpectedly, murinized InlA (InlA(m)) mediates not only Ecad-dependent internalization, but also N-cadherin-dependent internalization. Consequently, InlA(m)-expressing Lm targets not only goblet cells expressing luminally-accessible Ecad, as does Lm in humanized mice, but also targets villous M cells, which express luminally-accessible N-cadherin. This aberrant Lm portal of entry results in enhanced innate immune responses and intestinal barrier damage, both of which are not observed in wild-type Lm-infected humanized mice. Murinization of InlA therefore not only extends the host range of Lm, but also broadens its receptor repertoire, providing Lm with artifactual pathogenic properties. These results challenge the relevance of using InlA(m)-expressing Lm to study human listeriosis and in vivo host responses to this human pathogen.

Fibers are widely used in different industrial processes, for example in paper manufacturing or lost circulation problems in the oil industry. Recently, interest towards the use of fibers at the microscale has grown, driven by research in bio-medical applications or drug delivery systems. Microfluidic systems are not only directly relevant for lab-on-chip applications, but have also proven to be good model systems to tackle fundamental questions about the flow of fiber suspensions. It has therefore become necessary to provide fiber-like particles with an excellent control of their properties. We present here two complementary in situ methods to fabricate controlled micro-fibers allowing for an embedded fabrication and flow-on-a-chip platform. The first one, based on a photo-lithography principle, can be used to make isolated fibers and dilute fiber suspensions at specific locations of interest inside a microchannel. The self-assembly property of super-paramagnetic colloids is the principle of the second fabrication method, which enables the fabrication of concentrated suspensions of more flexible fibers. We propose a flow gallery with several examples of fiber flow illustrating the two methods’ capabilities and a range of recent laminar flow results.

The human opportunistic pathogen Bacillus cereus belongs to the B. cereus group that includes bacteria with a broad host spectrum. The ability of these bacteria to colonize diverse hosts is reliant on the presence of adaptation factors. Previously, an IVET strategy led to the identification of a novel B. cereus protein (IlsA, Iron-regulated leucine rich surface protein), which is specifically expressed in the insect host or under iron restrictive conditions in vitro. Here, we show that IlsA is localized on the surface of B. cereus and hence has the potential to interact with host proteins. We report that B. cereus uses hemoglobin, heme and ferritin, but not transferrin and lactoferrin. In addition, affinity tests revealed that IlsA interacts with both hemoglobin and ferritin. Furthermore, IlsA directly binds heme probably through the NEAT domain. Inactivation of ilsA drastically decreases the ability of B. cereus to grow in the presence of hemoglobin, heme and ferritin, indicating that IlsA is essential for iron acquisition from these iron sources. In addition, the ilsA mutant displays a reduction in growth and virulence in an insect model. Hence, our results indicate that IlsA is a key factor within a new iron acquisition system, playing an important role in the general virulence strategy adapted by B. cereus to colonize susceptible hosts.

Herpesviruses constitute a family of large DNA viruses widely spread in vertebrates and causing a variety of different diseases. They possess dsDNA genomes ranging from 120 to 240 kbp encoding between 70 to 170 open reading frames. We previously reported the protein interaction networks of two herpesviruses, varicella-zoster virus (VZV) and Kaposi's sarcoma-associated herpesvirus (KSHV). In this study, we systematically tested three additional herpesvirus species, herpes simplex virus 1 (HSV-1), murine cytomegalovirus and Epstein-Barr virus, for protein interactions in order to be able to perform a comparative analysis of all three herpesvirus subfamilies. We identified 735 interactions by genome-wide yeast-two-hybrid screens (Y2H), and, together with the interactomes of VZV and KSHV, included a total of 1,007 intraviral protein interactions in the analysis. Whereas a large number of interactions have not been reported previously, we were able to identify a core set of highly conserved protein interactions, like the interaction between HSV-1 UL33 with the nuclear egress proteins UL31/UL34. Interactions were conserved between orthologous proteins despite generally low sequence similarity, suggesting that function may be more conserved than sequence. By combining interactomes of different species we were able to systematically address the low coverage of the Y2H system and to extract biologically relevant interactions which were not evident from single species.