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Cytotoxic-T-lymphocyte (CTL) mediated control of HIV-1 is enhanced by targeting highly networked epitopes in complex with human-leukocyte-antigen-class-I (HLA-I). However, the extent to which the presenting HLA allele contributes to this process is unknown. Here we examine the CTL response to QW9, a highly networked epitope presented by the disease-protective HLA-B57 and disease-neutral HLA-B53. Despite robust targeting of QW9 in persons expressing either allele, T cell receptor (TCR) cross-recognition of the naturally occurring variant QW9_S3T is consistently reduced when presented by HLA-B53 but not by HLA-B57. Crystal structures show substantial conformational changes from QW9-HLA to QW9_S3T-HLA by both alleles. The TCR-QW9-B53 ternary complex structure manifests how the QW9-B53 can elicit effective CTLs and suggests sterically hindered cross-recognition by QW9_S3T-B53. We observe populations of cross-reactive TCRs for B57, but not B53 and also find greater peptide-HLA stability for B57 in comparison to B53. These data demonstrate differential impacts of HLAs on TCR cross-recognition and antigen presentation of a naturally arising variant, with important implications for vaccine design.

Follicular CD8 + T cells (fCD8) mediate surveillance in lymph node (LN) germinal centers against lymphotropic infections and cancers, but the precise mechanisms by which these cells mediate immune control remain incompletely resolved. To address this, we investigated functionality, clonotypic compartmentalization, spatial localization, phenotypic characteristics, and transcriptional profiles of LN-resident virus-specific CD8 + T cells in persons who control HIV without medications. Antigen-induced proliferative and cytolytic potential consistently distinguished spontaneous controllers from noncontrollers. T cell receptor analysis revealed complete clonotypic overlap between peripheral and LN-resident HIV-specific CD8 + T cells. Transcriptional analysis of LN CD8 + T cells revealed gene signatures of inflammatory chemotaxis and antigen-induced effector function. In HIV controllers, the cytotoxic effectors perforin and granzyme B were elevated among virus-specific CXCR5 + fCD8s proximate to foci of HIV RNA within germinal centers. These results provide evidence consistent with cytolytic control of lymphotropic infection supported by inflammatory recruitment, antigen-specific proliferation, and cytotoxicity of fCD8s.

International audience; Caffeine is the most consumed psychoactive substance worldwide. Previous studies suggested higher caffeine consumption in subjects with schizophrenia spectrum disorders (SSD) as well as associations with symptoms, medication and medication side-effects. In a large and well-characterized sample of SSD subjects we explored the association between caffeine consumption and clinical (psychosis related, severity, general health) as well as pharmacological (antipsychotic treatment, sedation potential) variables. Eight hundred four subjects with data on their caffeine (coffee and tea) consumption successively recruited were included in this study. After controlling for potential confounders (demographic variables, smoking) only the negative dimension of psychosis was associated with the amount of caffeine ingested. Less severe negative symptoms were associated with higher caffeine consumption. The effect size of this association was small (partial correlation coefficient = −0.12) but significant.

International audience; Background: Aggressiveness is a stigma frequently associated with schizophrenia. The role of insight as a risk factor of aggressiveness remains contradictory; mainly because single measures of these states mask their complexity and heterogeneity.Methods: This study was conducted on 666 patients aged 15 and above with a DSM-IV-TR diagnosis of schizophrenia spectrum disorder, drawn from the French national network of schizophrenia expert center database. Collected data comprised socio-demographics and standardized psychiatric assessments. Aggressiveness was evaluated using the Buss-Perry Aggression Questionnaire and insight using the Scale to assess Unawareness of Mental Disorder (SUMD) and Birchwood Insight Scale (BIS).Results: Hostility was the aggressiveness dimension the most strongly associated with SUMD insight dimensions. Patients aware of their illness were nearly twice as likely to show hostility than those seriously unaware (OR = 1.95, 95% CI.: 1.08-3.5), but not when further adjusting for depression. Similarly, those aware of the consequences of their illness and of their symptoms were more hostile. Patients moderately aware of illness consequences had a higher risk of both anger and physical aggressiveness than those unaware (OR = 2.63, 95% CI.: 1.42-4.86, OR = 2.47, 95% CI.: 1.33-4.60, respectively), even when adjusting for depression for anger.Conclusion: Our study confirms that a multi-dimensional approach to insight and aggressiveness is essential to understand the types of links between these clinical states. Insight may trigger the expression of an underlying hostile tendency, maybe via depression and self-stigmatisation. This should be taken into account in therapeutic approaches to improve insight.

Structure-based immunogen design Vaccine design for highly mutable pathogens is hindered by a paucity of conserved immunogenic epitopes. Gaiha et al. employed a structure-based technique using network theory to assign scores to protein structure in order to infer mutational constraints (see the Perspective by McMichael and Carrington). The authors validated the method on proteins with published functional outcomes and then assessed mutational constraints within the HIV proteome. Highly networked residues strongly associated with immune control of HIV infection and may lead to protective immunogens for pathogens for which there is currently no efficient vaccine. Science , this issue p. 480 ; see also p. 438

Peste des petits ruminants (PPR) is an acute transboundary infectious viral disease of small ruminants, mainly sheep and goats. Host susceptibility varies considerably depending on the PPR virus (PPRV) strain, the host species and breed. The effect of strains with different levels of virulence on the modulation of the immune system has not been thoroughly compared in an experimental setting so far. In this study, we used a multi-omics approach to investigate the host cellular factors involved in different infection phenotypes. Peripheral blood mononuclear cells (PBMCs) from Saanen goats were activated with a T-cell mitogen and infected with PPRV strains of different virulence: Morocco 2008 (high virulence), Ivory Coast 1989 (low virulence) and Nigeria 75/1 (live attenuated vaccine strain). Our results showed that the highly virulent strain replicated better than the other two in PBMCs and rapidly induced cell death and a stronger inhibition of lymphocyte proliferation. However, all the strains affected lymphocyte proliferation and induced upregulation of key antiviral genes and proteins, meaning a classical antiviral response is orchestrated regardless of the virulence of the PPRV strain. On the other hand, the highly virulent strain induced stronger inflammatory responses and activated more genes related to lymphocyte migration and recruitment, and inflammatory processes. Both transcriptomic and proteomic approaches were successful in detecting viral and antiviral effectors under all conditions. The present work identified key immunological factors related to PPRV virulencein vitro. &nbsp

Treatment-resistant schizophrenia (TRS) affects around 30% of patients with schizophrenia (SZ) resulting in poor functioning, relapses, and reduced quality of life. Convergent findings show that inflammation could contribute to resistance. We thus search for immune signatures of patients with TRS/ultra TRS (UTRS) in a sample of community-dwelling outpatients with SZ. In total, 195 stabilized SZ patients (mean age = 31.2 years, 73% male gender) were consecutively included in the network of the FondaMental Expert Centers for Schizophrenia in France and received a thorough clinical assessment. At inclusion, psychotic symptomatology was evaluated by the Positive and Negative Syndrome Scale (PANSS) for schizophrenia. Circulating serum/plasma levels of a large panel of markers reflecting the main inflammatory pathways were evaluated. TRS was defined by current treatment by clozapine (CLZ) and UTRS by current CLZ treatment + PANSS total score ≥ 70. The frequency of TRS and UTRS patients was, respectively, 20% and 7.7% and was defined using multivariable analysis elevated by high levels of interleukin (IL)-12/IL-23p40, IL-17A, IL-10, and beta 2 microglobulin (B2M) and IL-12/IL-23p40, IL-17A, IL-6, IL-10, IFNγ, and B2M, respectively. These observations suggest that resistance and ultra resistance to CLZ treatment are underpinned by pro-inflammatory molecules mainly belonging to the T helper 17 pathway, a finding making sense given the interplay between inflammation and antipsychotic treatment responses. If confirmed, our findings may allow us to consider IL-23/IL-17 pathway as a therapeutic target for patients with resistance to antipsychotics.

Summary Defining factors that govern CD8+ T cell immunodominance is critical for the rational design of vaccines for viral pathogens. Here, we assess the contribution of human leukocyte antigen (HLA) class-I-peptide stability for 186 optimal HIV epitopes across 18 HLA alleles using transporter associated with antigen processing (TAP)-deficient mono-allelic HLA-expressing cell lines. We find that immunodominant HIV epitopes increase surface stabilization of HLA class-I molecules in comparison to subdominant epitopes. HLA class-I-peptide stability is also strongly correlated with overall immunodominance hierarchies, particularly for epitopes from high-abundance proteins (e.g., Gag). Moreover, HLA alleles associated with HIV protection are preferentially stabilized by epitopes derived from topologically important viral regions at a greater frequency than neutral and risk alleles. These findings indicate that relative stabilization of HLA class-I is a key factor for CD8+ T cell epitope immunodominance hierarchies, with implications for HIV control and the design of T-cell-based vaccines., Graphical abstract, Highlights • TAP-deficient HLA-expressing cells provide rapid assessment of HLA-peptide stability • Immunodominant HIV CD8+ T cell epitopes are superior stabilizers of HLA molecules • HLA class-I-peptide stability correlates with overall immunodominance hierarchies • Protective HLA alleles are preferentially stabilized by highly networked epitopes, Kaseke et al. describe a cell-based HLA-peptide stability assay that leverages TAP-deficient mono-allelic HLA-expressing cell lines. Robust HLA stabilization is a common feature of immunodominant epitopes, and HLA-peptide stability correlates with immunodominance hierarchies. Epitopes derived from topologically constrained regions of the HIV proteome preferentially stabilize protective HLA class-I alleles.

Summary Spontaneous control of HIV infection has been repeatedly linked to antiviral CD8+ T cells but is not always permanent. To address mechanisms of durable and aborted control of viremia, we evaluated immunologic and virologic parameters longitudinally among 34 HIV-infected subjects with differential outcomes. Despite sustained recognition of autologous virus, HIV-specific proliferative and cytolytic T cell effector functions became selectively and intrinsically impaired prior to aborted control. Longitudinal transcriptomic profiling of functionally impaired HIV-specific CD8+ T cells revealed altered expression of genes related to activation, cytokine-mediated signaling, and cell cycle regulation, including increased expression of the antiproliferative transcription factor KLF2 but not of genes associated with canonical exhaustion. Lymphoid HIV-specific CD8+ T cells also exhibited poor functionality during aborted control relative to durable control. Our results identify selective functional impairment of HIV-specific CD8+ T cells as prognostic of impending aborted HIV control, with implications for clinical monitoring and immunotherapeutic strategies.

International audience; The aim of our study was to compare the performance of three different instruments measuring clinical and cognitive dimensions of insight. Data on 182 outpatients with schizophrenia and one-year follow-up assessments was drawn from the FACE-SZ cohort. Awareness of clinical state (« clinical insight ») was measured using both a clinician-rated measure (the Scale to assess Unawareness of Mental Disorder (SUMD)), and a self-report measure (the Birchwood Insight Scale (BIS). Cognitive insight was measured using a self-report measure (the Beck Cognitive Insight Scale (BCIS)). For each scale, change in insight was examined at the follow-up. Correlations between SUMD and BIS subscales measuring same dimensions were significant. BIS-BCIS correlations were weak for all combinations except between BIS illness dimension and BCIS composite score. At the follow-up, BIS and SUMD awareness of treatment need improved whereas illness and symptom awareness increased only on the SUMD. Conversely, cognitive insight composite scores decreased. Despite relatively good overall agreement between the two clinical insight instruments, considerable variability for similar insight dimensions measured by different instruments was found. Agreement between cognitive and clinical insight is moderate. Our study strengthens the argument that insight is harder to operationalize than other symptoms and may explain why it is so seldom explicitly targeted in schizophrenia treatment.

Summary: Defining factors that govern CD8+ T cell immunodominance is critical for the rational design of vaccines for viral pathogens. Here, we assess the contribution of human leukocyte antigen (HLA) class-I-peptide stability for 186 optimal HIV epitopes across 18 HLA alleles using transporter associated with antigen processing (TAP)-deficient mono-allelic HLA-expressing cell lines. We find that immunodominant HIV epitopes increase surface stabilization of HLA class-I molecules in comparison to subdominant epitopes. HLA class-I-peptide stability is also strongly correlated with overall immunodominance hierarchies, particularly for epitopes from high-abundance proteins (e.g., Gag). Moreover, HLA alleles associated with HIV protection are preferentially stabilized by epitopes derived from topologically important viral regions at a greater frequency than neutral and risk alleles. These findings indicate that relative stabilization of HLA class-I is a key factor for CD8+ T cell epitope immunodominance hierarchies, with implications for HIV control and the design of T-cell-based vaccines.

There are intriguing parallels between plants and animals, with respect to the structures of their innate immune receptors, that suggest universal principles of innate immunity. The cytosolic nucleotide binding site-leucine rich repeat (NBS-LRR) resistance proteins of plants (R-proteins) and the so-called NOD-like receptors of animals (NLRs) share a domain architecture that includes a STAND (signal transduction ATPases with numerous domains) family NTPase followed by a series of LRRs, suggesting inheritance from a common ancestor with that architecture. Focusing on the STAND NTPases of plant R-proteins, animal NLRs, and their homologs that represent the NB-ARC (nucleotide-binding adaptor shared by APAF-1, certain R gene products and CED-4) and NACHT (named for NAIP, CIIA, HET-E, and TEP1) subfamilies of the STAND NTPases, we analyzed the phylogenetic distribution of the NBS-LRR domain architecture, used maximum-likelihood methods to infer a phylogeny of the NTPase domains of R-proteins, and reconstructed the domain structure of the protein containing the common ancestor of the STAND NTPase domain of R-proteins and NLRs. Our analyses reject monophyly of plant R-proteins and NLRs and suggest that the protein containing the last common ancestor of the STAND NTPases of plant R-proteins and animal NLRs (and, by extension, all NB-ARC and NACHT domains) possessed a domain structure that included a STAND NTPase paired with a series of tetratricopeptide repeats. These analyses reject the hypothesis that the domain architecture of R-proteins and NLRs was inherited from a common ancestor and instead suggest the domain architecture evolved at least twice. It remains unclear whether the NBS-LRR architectures were innovations of plants and animals themselves or were acquired by one or both lineages through horizontal gene transfer.

International audience; Objectives The present article is a synthesis of the first 10 years of follow-up of the FondaMental Academic Center of Expertise for Schizophrenia (FACE-SZ) cohort. Methods More than 700 community-dwelling stabilized subjects have been recruited and evaluated to date. The mean age was 32 years with 75 % males, the mean illness duration was 11 years, the mean age at illness onset was 21 years, the mean duration of untreated psychosis was 1.5 years and 55 % were current daily tobacco smokers. Results The major findings of the FACE-SZ cohort may be summarized as follows: the metabolic syndrome is twice more frequent in schizophrenia as compared to the general population and is not correctly assessed and treated; cognitive disturbances have been found in benzodiazepine consumers and in patients with chronic low-grade peripheral inflammation; major depressive disorder (MDD) is a common current comorbid condition in about 20% of the subjects at the evaluation. MDD is associated with impaired quality of life and with increased nicotine dependency in SZ daily tobacco smokers. Improving depression and negative symptoms may be the most effective strategies to improve quality of life in schizophrenia; the duration of untreated psychosis is much longer in cannabis smokers and in subjects with an age at illness onset < 19 years. Adherence to treatment is diminished in subjects who report a subjective negative feeling after treatment intake independent of objective side effects (extrapyramidal syndrome and weight gain). Akathisia has been found in 18% of the subjects and has been associated with antipsychotic polytherapy. Conclusions In the light of these results, some recommendations for clinical care may be suggested. The early detection of schizophrenia should be specifically increased in adolescents and/or cannabis smokers. All patients should be administered a comprehensive neuropsychological evaluation at the beginning of the illness and after stabilization under treatment. Improving metabolic parameters and lifestyle (diet and physical activity) should be reinforced. The benefit/risk ratio of benzodiazepine and antipsychotic polytherapy should be regularly reevaluated and withdrawn as soon as possible. If MDD remains underdiagnosed and undertreated, improving depression may strongly improve the quality of life of SZ subjects. In the end, Cognitive Remediation Therapy and anti-inflammatory strategies should be more frequently included in therapeutic strategies.; Objectifs Le présent article propose de faire un retour sur les 10 ans d’expérience du réseau national des 10 centres experts schizophrénie Français, à travers une revue des principales publications scientifiques qui en sont issues. Méthodes À ce jour, les données de plus de 700 patients ont pu être analysées. Globalement, cette population est composée de trois quarts d’hommes, est âgée en moyenne de 32 ans, la durée moyenne de maladie est d’environ 11 ans, l’âge moyen de début de maladie est de 21 ans, la durée de psychose non traitée est d’environ un an et demi. 55 % des sujets sont consommateurs de tabac au moment de l’évaluation. Enfin, un focus sera fait sur les projets de recherche multicentriques collaboratifs déployés au sein de ce réseau. Résultats Les principales données obtenues peuvent être résumées comme suit : le syndrome métabolique reste deux fois plus fréquent dans la schizophrénie qu’en population générale et n’est pas correctement pris en charge ; des troubles cognitifs spécifiques ont été identifiés chez les patients consommateurs de benzodiazépines et chez les patients présentant une inflammation périphérique. Des spécificités liées au sexe ont également été observées ; la dépression comorbide reste très fréquente, elle est associée à une diminution de la qualité de vie et à une augmentation de la dépendance nicotinique chez les fumeurs. Prendre en charge la dépression et les symptômes négatifs pourraient fortement améliorer la qualité de vie des patients ; le délai à l’instauration du traitement est plus long pour les schizophrénies se déclenchant avant 19 ans et chez les consommateurs de cannabis. L’adhésion au traitement est diminuée chez les patients rapportant un ressenti subjectif négatif du traitement, indépendamment de la prise de poids ou d’un syndrome extrapyramidal. Enfin, l’akathisie est très fréquente et fortement associée à la polythérapie antipsychotique. Conclusions Ces résultats conduisent à formuler plusieurs recommandations. Le dépistage du trouble et le traitement devraient être renforcés chez les adolescents et les consommateurs de cannabis. Tous les patients devraient bénéficier d’une évaluation neuropsychologique au début de leur trouble, après stabilisation sous traitement, puis de façon plus espacée au cours de leur suivi. La prise en charge des paramètres métaboliques, de l’alimentation et de l’activité physique devrait être renforcée. La balance bénéfice/risque de la prescription de benzodiazépines doit être réévaluée régulièrement au regard de son impact notamment sur la cognition. La monothérapie antipsychotique est recommandée autant que possible pour limiter les effets indésirables. La dépression reste sous-diagnostiquée et doit faire l’objet d’une évaluation systématique et d’un traitement spécifique. La remédiation cognitive doit également être plus largement proposée chez les patients présentant des troubles cognitifs.

Introduction This study aimed was to investigate the relationship between different types of childhood trauma and the level of insight (i.e., awareness of having a psychiatric disorder) in subjects suffering from schizophrenia, as well as the putative role of clinical mediators. Methods 294 community-dwelling subjects with stable schizophrenia were included into FACE-SZ, a multicentre cross-sectional study. All patients were assessed by specialized multidisciplinary teams. The level of insight was assessed by the Scale to assess Unawareness of Mental Disorder (SUMD), and childhood trauma by the Childhood Trauma Questionnaire (CTQ). Path analyses from the five CTQ subscales (physical abuse and neglect, emotional abuse and neglect, and sexual abuse) and the SUMD, with current symptomatology (i.e., positive, negative, global psychopathology and depression) as mediator, was performed. Results Physical neglect (β = 0.14) and abuse (β = 0.13) were significantly associated with poor insight. Negative symptoms were a clinical mediator of the relationship between physical neglect and poor insight. Moreover, positive (β = 0.21) and negative (β = 0.30) symptoms were associated with poor insight, whereas depression (β = −0.14) was associated with higher levels of insight. Discussion For the first time, this study shows a significant relationship between childhood trauma, specifically physical neglect and abuse, and poor insight. The level of insight was linked to different clinical dimensions. Among subjects with schizophrenia, these results provide support for a role of childhood trauma in poorer management outcomes, and the need to provide treatment, including psycho-education that better targets the consequences of childhood trauma.

Objectives Reducing the duration of untreated psychosis (DUP) may improve the prognosis of schizophrenia. This study investigated the prevalence, and associated risk factors, of long DUP in a large, non-selected sample of community-dwelling schizophrenia patients (SZ). Method 478 community-dwelling stable SZ participants (122 women and 356 men; mean age 32.37 ± 9.86 years) were recruited between 2010 and 2016. The mean retrospective DUP was evaluated from both patient and family reports, as well as hospital/psychiatrists records. Long DUP was defined as > 2 years. Results The mean DUP was 1.5 years. 80 participants (16.7%) had a DUP > 2 years. In multivariate analyses, after adjustment for sex, education level, history of childhood trauma and history of maternal schizophrenia or bipolar disorder, long DUP was associated with a younger age of illness onset (19.3 ± 6.67 years vs. 22.0 ± 6.51 years, adjusted odd ratio aOR = 0.91, 95%CI [0.86; 0.97], p = 0.003) and cannabis use disorder (20.0% vs. 10.3%, aOR = 2.41, 95%CI [1.14–5.09], p = 0.02). Conclusion A high proportion of SZ patients still have a long DUP. The present results suggest that illness onset before age 19 years and cannabis use are associated with long DUP in schizophrenia patients. Early psychosis detection programs should prioritize the targeting of these populations.

The present article is a synthesis of the first 10 years of follow-up of the FondaMental Academic Center of Expertise for Schizophrenia (FACE-SZ) cohort.More than 700 community-dwelling stabilized subjects have been recruited and evaluated to date. The mean age was 32 years with 75 % males, the mean illness duration was 11 years, the mean age at illness onset was 21 years, the mean duration of untreated psychosis was 1.5 years and 55 % were current daily tobacco smokers.The major findings of the FACE-SZ cohort may be summarized as follows: the metabolic syndrome is twice more frequent in schizophrenia as compared to the general population and is not correctly assessed and treated; cognitive disturbances have been found in benzodiazepine consumers and in patients with chronic low-grade peripheral inflammation; major depressive disorder (MDD) is a common current comorbid condition in about 20% of the subjects at the evaluation. MDD is associated with impaired quality of life and with increased nicotine dependency in SZ daily tobacco smokers. Improving depression and negative symptoms may be the most effective strategies to improve quality of life in schizophrenia; the duration of untreated psychosis is much longer in cannabis smokers and in subjects with an age at illness onset19 years. Adherence to treatment is diminished in subjects who report a subjective negative feeling after treatment intake independent of objective side effects (extrapyramidal syndrome and weight gain). Akathisia has been found in 18% of the subjects and has been associated with antipsychotic polytherapy.In the light of these results, some recommendations for clinical care may be suggested. The early detection of schizophrenia should be specifically increased in adolescents and/or cannabis smokers. All patients should be administered a comprehensive neuropsychological evaluation at the beginning of the illness and after stabilization under treatment. Improving metabolic parameters and lifestyle (diet and physical activity) should be reinforced. The benefit/risk ratio of benzodiazepine and antipsychotic polytherapy should be regularly reevaluated and withdrawn as soon as possible. If MDD remains underdiagnosed and undertreated, improving depression may strongly improve the quality of life of SZ subjects. In the end, Cognitive Remediation Therapy and anti-inflammatory strategies should be more frequently included in therapeutic strategies.

International audience; The functional outcome in schizophrenia spectrum disorders is affected by multiple factors such as cognitive performance and clinical symptoms. Psychiatric disability may be another important determinant of functional outcome. The purpose of this study was to test whether schizophrenia symptoms and psychiatric disability mediated the association between cognition and functioning. Between April 2013 and July 2017, we included 108 community-dwelling adults with stable schizophrenia spectrum disorder in a multicenter study. Psychiatric disability was assessed with the Evaluation of Cognitive Processes involved in Disability in Schizophrenia (ECPDS) scale by relatives of patients. ECPDS focused on the broad array of motivational, neurocognitive, sociocognitive, and metacognitive impairments that result in activity restrictions. We used a battery of tests to assess seven cognition domains (processing speed, attention/vigilance, working, verbal and visual memory, reasoning and problem solving, and executive functioning) and cross-sectional structural equation modeling (SEM) for the mediation analyses. We estimated the one-year temporal stability of ECPDS scores in 45 participants. The model provided showed good fit and explained 43.9% of the variance in functioning. The effect of neurocognition on functioning was fully mediated by symptoms (proportion mediated: 36.5%) and psychiatric disability (proportion mediated: 31.3%). The ECPDS score had acceptable one-year temporal stability. The ECPDS scale has satisfactory psychometric properties, and shows significant convergence with neurocognition and functioning, suggesting a role for this tool in the routine evaluation of cognitive remediation needs. Our model validates psychiatric disability as a crucial step from cognitive impairment to restricted participation in life situations.

Low-grade inflammation has repeatedly been associated with schizophrenia (SZ) and in particular with cognitive impairment. Female gender, overweight and tobacco smoking have been suggested as risk factors to increase inflammation while preclinical inconsistent findings have been found regarding the association with psychotropic drugs. The aim of this study was to explore if psychotropic drugs were associated with inflammation in SZ and to determine which psychotropic drug was associated with inflammation in stable SZ subjects while considering clinical confounding factors. Participants were consecutively included in the network of the FondaMental Expert Centers for Schizophrenia and received a thorough clinical assessment, including recording of current treatment. High-sensitivity CRP (hs-CRP) was measured for each participant as a proxy to define peripheral low-grade inflammation. The zero-inflated Poisson regression model estimated the relationship between low-grade inflammation and psychotropic drug. Four hundred and five stabilized, community-dwelling SZ subjects (mean age = 32.6 years, 74% male gender) have been included. In total, 148 participants (36.5%) were found with undetectable blood hs-CRP level. The probability of having an undetectable CRP was associated with a lower body mass index (p 0.0001) and no cyamemazine add-on antipsychotic therapy (p = 0.001). The other 257 participants (63.5%) were found to have low-grade inflammation (hs-CRP 0 mg/L). Low-grade inflammation was significantly associated with female gender (p = 0.004), higher body mass index (p 0.0001), current tobacco smoking (p 0.0001), clomipramine (p = 0.04), quetiapine (p 0.0001) and hypnotic (p = 0.0006) consumption while decreased hs-CRP blood levels was associated with aripiprazole (p = 0.004) and valproate/valpromide (p = 0.03) consumption. The present study suggests that some psychotropic drugs (quetiapine, cyamemazine, clomipramine) may be associated with increased peripheral low-grade inflammation in SZ patients while others (aripiprazole, valproate) may be associated with decreased peripheral low-grade inflammation. These results should be replicated in SZ and non-SZ populations and the biological underpinnings should be further explored.

Supplemental material, Supplemental_Material for Screening for cognitive deficits with the Evaluation of Cognitive Processes involved in Disability in Schizophrenia scale by Paul Roux, Mathieu Urbach, Sandrine Fonteneau, Fabrice Berna, Lore Brunel, Delphine Capdevielle, Isabelle Chereau, Julien Dubreucq, Catherine Faget-Agius, Guillaume Fond, Sylvain Leignier, Claire-Cécile Perier, Raphaëlle Richieri, Priscille Schneider, Franck Schürhoff, Anne Marie Tronche, Hanan Yazbek, Anna Zinetti-Bertschy, the FondaMental Advanced Centers of Expertise in Schizophrenia (FACE-SCZ) Collaborators, Christine Passerieux, Eric Brunet-Gouet, M Andrianarisoa, B Aouizerate, N Bazin, F Berna, O Blanc, L Brunel, E Bulzacka, D Capdevielle, I Chereau-Boudet, G Chesnoy-Servanin, JM Danion, T D’Amato, A Deloge, C Delorme, H Denizot, JM Dorey, C Dubertret, J Dubreucq, C Faget, C Fluttaz, G Fond, S Fonteneau, F Gabayet, E Giraud-Baro, D Lacelle, C Lançon, H Laouamri, M Leboyer, T Le Gloahec, Y Le Strat, PM Llorca, J Mallet, E Metairie, D Misdrahi, I Offerlin-Meyer, C Passerieux, P Peri, S Pires, C Portalier, L Ramet, R Rey, C Roman, A Schandrin, F Schürhoff, A Tessier, AM Tronche, M Urbach, F Vaillant, A Vehier, P Vidailhet, E Vilà, H Yazbek and A Zinetti-Bertschy in Clinical Rehabilitation

Objective Latent Toxoplasma infection has been associated with widespread brain immune activation, increased blood brain barrier permeability, neural disruption, increased dopamine release in dopaminergic neurons, with NMDA activation and with schizophrenia (SZ) onset risk. Toxoplasma has been suggested to be a source of chronic low-grade inflammation and this inflammation has been associated with cognitive impairment in SZ. The objective of the present study were (i) to determine if latent Toxoplasma infection was associated with specific clinical features in stabilized SZ subjects, with cognitive impairment and with increased low-grade peripheral inflammation and (ii) to determine if Treatments with Anti-Toxoplasmic Activity (TATA) were associated with improved outcomes in subjects with latent Toxoplasma infection. Methods A comprehensive 2 daylong clinical and neuropsychological battery was administered in 250 SZ subjects included between 2015 and 2017 in the national FondaMental Expert Center (FACE-SZ) Cohort. Solid phase-enzyme microplate immunoassay methods were used to measure IgG class of antibodies to T. gondii in blood sample. Latent Toxoplasma infection was defined by T . gondii IgG ratio ≥0.8, equivalent to ≥10 international units. Chronic peripheral inflammation was defined by highly sensitive C reactive protein blood level ≥ 3 mg/L. Results Latent Toxoplasma infection has been found in 184 (73.6%) of this national multicentric sample. In the multivariate analyses, latent Toxoplasma infection has been significantly associated with higher PANSS negative (aOR = 1.1 [1.1–1.1], p = 0.04) and excitement subscores (aOR = 1.3 [1.1–1.6], p = 0.01), with two specific symptoms (i.e., reference delusion (aOR = 3.6 [1.2–10.6] p = 0.01) and alogia (aOR = 16.7 [2.0–134.7], p = 0.008)) and with chronic low-grade peripheral inflammation (27.2% vs. 7.6%, aOR = 3.8 [1.4–10.3], p = 0.004). Extrapyramidal symptoms remained significantly associated with latent Toxoplasma infection. On the opposite, no significant association of latent Toxoplasma infection with age, gender, age at SZ onset, suicide behavior or cognitive deficits has been found in these models (all p > 0.05). TATA were associated with lower depressive symptoms (aOR = 0.8[0.7–0.9], p = 0.01), and with lower rates of chronic peripheral inflammation (20.9% vs. 48.6%, aOR = 3.5 [1.5–7.9], p = 0.003) but not with higher cognitive scores (p > 0.05). Conclusion The present findings suggest that Toxoplasma is almost 3 times more frequent in SZ population compared to general population in France. The potential cerebral underpinnings of the association of latent Toxoplasma infection and the above-mentioned outcomes have been discussed. Future studies should confirm that TATA may be effective to reduce Toxoplasma-associated depressive symptoms and low-grade peripheral inflammation.

La schizophrenie et le trouble obsessionnel-compulsif appartiennent, depuis leurs toutes premieres descriptions, a deux entites nosographiques distinctes. Selon la theorie analytique, ces deux troubles sont censes emerger sur des « structures » fondamentalement differentes : nevrotique ou psychotique. Neanmoins, un lien entre la « nevrose obsessionnelle » et la « psychose » a rapidement ete decrit. Les nomenclatures actuelles permettent de poser les deux diagnostics chez un meme sujet, mais elles restent en partie heritieres des categories psychanalytiques. Pourtant, il arrive qu’un doute persiste sur la nature du symptome obsessionnel observe. Tout d’abord parce que les symptomes obsessionnels sont frequents dans la schizophrenie, au point que certains auteurs ont fait l’hypothese d’une sous-categorie specifique, le « schizo-obsessive disorder » (Poyurovsky et al., 2012 ; Scotti-Muzzi et Saide, 2017). Mais aussi parce que les etudes cliniques ont conduit a un tournant majeur de la description du TOC a partir du DSM-IV : la possibilite d’un TOC avec un mauvais insight (Foa et al., 1995). Ce qui fait desormais la difference entre une obsession et une idee delirante n’est plus la reconnaissance de son caractere absurde, mais la lutte anxieuse qu’elle provoque (Oulis et al., 2013). Cette approche brouille les distinctions entre troubles anxieux et troubles psychotiques, nous forcant a reinterroger nos criteres diagnostiques. De plus elle vient ebranler de facon profonde nos representations de la pathologie mentale et souligne les tensions entre les approches dimensionnelles et categorielles.

Background Tobacco use is common in patients with schizophrenia (SZ) but little is known on the role of tobacco in the physiopathology or on the course of the disease. Only few studies embrace an extensive examination of clinical and therapeutic characteristics in stabilized patients. The objective of the present study was to determine the prevalence of tobacco smoking in stabilized SZ outpatients and the clinical and treatment characteristics associated with daily tobacco use in a large community-dwelling sample of patients. Methods Three-hundred-and-sixty-one patients were included in the network of the FondaMental Expert Centers for Schizophrenia. Current tobacco status was self-declared. Results 53.7% were smokers. Mean age at tobacco onset was 17.2 years old. In multivariate analyses, after adjustment for confounding factors, positive symptoms and mean daily antipsychotic dose were associated with a higher frequency of tobacco use (OR = 1.06 95%IC[1.02–1.12], for positive symptoms, OR = 1.1, 95%IC[1.02–1.18] for daily antipsychotic dose). Education level, negative symptoms, anticholinergic agents, clozapine or aripiprazole administration were independently associated with a lower frequency of tobacco use (respectively OR = 0.87, 95%IC [0.79, 0.95], OR = 0.95, 95%IC[0.91–0.98], OR = 0.41, 95%IC[0.22–0.76], OR = 0.56, 95%IC = [0.32, 0.99] and OR = 0.49, 95%IC [0.26–0.91]). Conclusion The prevalence of current tobacco smoking in a French community-dwelling SZ patients is higher that observed in the general population. Patients with tobacco use present clinical and therapeutic specificities that may involve interaction between cholinergic-nicotinic and dopaminergic systems. The present study suggests that some therapeutics may improve daily smoking behavior in smokers. These results should be confirmed in longitudinal studies.

International audience; OBJECTIVES:Depression and negative symptoms have been associated with impaired Quality of life (QoL) in schizophrenia (SZ). However, childhood trauma may influence both QoL and depression in SZ patients, with consequences for the management of impaired QoL in SZ patients. The aim of the present study was to determine if childhood trauma was associated with impaired QoL in schizophrenia.METHOD:A sample of 544 community-dwelling stabilized SZ patients enrolled in FACE-SZ cohort were utilized in this study (74.1% males, mean aged 32.3years, mean illness duration 10.6years). QoL was self-reported with the S-QoL18 questionnaire. Childhood trauma was self-reported with the Childhood Trauma Questionnaire. Depression was measured by the Calgary Depression Rating Scale for Schizophrenia. Psychotic severity was measured by the Positive and Negative Syndrome Scale for Schizophrenia (PANSS). Other clinical factors, treatments, comorbidities, functioning and sociodemographical variables were also recorded, with validated scales.RESULTS:Overall, 151 participants (27.8%) had a current major depressive episode and 406 (82.5%) reported at least one episode of historical childhood trauma. In multivariate analyses, lower QoL total score was associated with a history of childhood trauma (β=-0.21, p

Objectives Sex differences can yield important clues regarding illness pathophysiology and its treatment. Schizophrenia (SZ) has a lower incidence rate, and a better prognosis, in women versus men. The present study investigated the cognitive profiles of both sexes in a large multi-centre sample of community-dwelling SZ patients. Method 544 community-dwelling stable SZ subjects (141 women and 403 men; mean age 34.5 ± 12.1 and 31.6 ± 8.7 years, respectively) were tested with a comprehensive battery of neuropsychological tests. Results Although community-dwelling SZ men had more risk factors for impaired cognition (including first-generation antipsychotics administration and comorbid addictive disorders), women had lower scores on a wide range of cognitive functions, including current and premorbid intellectual functioning, working memory, semantic memory, non-verbal abstract thinking and aspects of visual exploration. However, women scored higher in tests of processing speed and verbal learning, as well as having a lower verbal learning bias. No sex difference were evident for visuospatial learning abilities, cued verbal recall, sustained attention and tests of executive functions, including cognitive flexibility, verbal abstract thinking, verbal fluency and planning abilities. Conclusion Sex differences are evident in the cognitive profiles of SZ patients. The impact on daily functioning and prognosis, as well as longitudinal trajectory, should be further investigated in the FACE-SZ follow-up study. Sex differences in cognition have implications for precision-medicine determined therapeutic strategies. Limits Given the restricted age range of the sample, future research will have to determine cognitive profiles across gender in late onset SZ.

Plants and animals must avoid becoming a free meal to microbes, which vastly outnumber eukaryotic life in both quantity and diversity. Adaptive immunity in the strict sense, whereby the host creates an immunological memory after exposure to a pathogen, is limited to vertebrates. Both plants and animals (including insects and mammals) have an innate immune system, which helps protect hosts from the majority of microbes they encounter during their lifetime. Plant and animal innate immune systems recognize an overlapping set of conserved microbe-associated molecular patterns (MAMPs). This observation suggests that the innate immune system in plants and animals may have been derived from a common ancestor. However, the majority of data indicate that innate immunity has arisen independently in plants and animals and that functional overlap is the result of convergent evolution: confronted with the same problem, and given the same molecular tools, plants and animals have independently derived similar solutions. This review discusses the functional and mechanistic details of the innate immune system in plants and animals including receptor-mediated immunity, endolysosomal immunity, and the interplay of the innate immune system and host-associated microbial communities.

Une des missions des centres expert de la fondation fondamental est d’apporter de l’aide aux psychiatres correspondants dans l’evaluation therapeutique, mais egalement en cas de doute diagnostique. Ce symposium reunit trois specialistes des troubles de l’humeur et des troubles schizophreniques, pour aborder certaines difficultes des presentations cliniques et l’aide que peuvent apporter les donnees recueillies par la Fondation. Le Dr Urbach evoquera les doutes diagnostiques entre trouble obsessionnel-compulsif et schizophrenie, deux entites nosographiques pourtant distinctes. Les TOC sont frequents dans la schizophrenie. Certains auteurs ont suppose l’existence d’une sous-categorie specifique, et les etudes cliniques ont conduit a un tournant majeur a partir du DSM-IV : la possibilite d’un TOC avec un mauvais insight. L’auteur insistera sur cette categorie, qui brouille les distinctions entre troubles anxieux et troubles psychotiques. Cette entite interroge de facon profonde nos representations de la pathologie mentale et souligne les tensions entre les approches dimensionnelles et categorielles. Le Dr Ducasse s’interessera au trouble bipolaire, et au trouble de personnalite borderline : les interfaces entre ces deux troubles, et les manieres de les differencier. Deux troubles frequents, engendrant un impact individuel et societal negatifs considerables, co-occurents dans 20 % des cas, et souvent confondus. Dans une perspective de depasser les limites de nos representations categorielles, une comprehension dimensionnelle du trouble de la personnalite borderline sera presentee. Mieux comprendre le vecu interieur, pour mieux aider. L’auteur s’interessera egalement a l’evolution des deux troubles, et les influences potentielles de l’un sur l’autre. Le Dr Fond abordera les enjeux cliniques du diagnostic differentiel entre symptomes depressifs et negatifs souvent confondus chez les patients, ainsi que les facteurs associes a la depression et a la non-remission. Il presentera egalement les dernieres donnees issues de la cohorte FACE-SZ, une cohorte de 1000 patients avec schizophrenie suivis sur 3 ans. Ces analyses ont revele le role majeur de la depression, identifiee chez un tiers des patients, et non diagnostiquee et non traitee dans la moitie des cas, alors qu’elle est associee a une alteration importante de la qualite de vie des patients. Les antidepresseurs ont pourtant montre leur efficacite, meme si 44 % des patients traites n’entrent pas en remission.

The genetically tractable model organism Caenorhabditis elegans was first used to model bacterial virulence in vivo a decade ago. Since then, great strides have been made in the identification of host response pathways that are involved in the defence against infection. Strikingly, C. elegans seems to detect and respond to infection without the involvement of its Toll-like receptor homologue, in contrast to the well-established role for these proteins in innate immunity in mammals. What, therefore, do we know about host defence mechanisms in C. elegans, and what can they tell us about innate immunity in higher organisms?

In case of a severe accident in one of the nuclear power plants in northern Germany consequences may reach across the borders of counties and Länder and several local and regional authorities will have responsibilities for disaster response measures. Consistent assessment of the situation and coordinated action are essential for protection of the population and task personnel as effective as possible. Disaster response managements have to come to a comprehensive assessment of the developing radiological situation at an early stage to be able to initiate appropriate protective measures. The results of a working group from several local and regional authorities from Schleswig-Holstein, Hamburg, Lower Saxony and Mecklenburg-West Pomerania on this topic will be presented here. The working group has compiled sets of data and information which are relevant in decision making for protective measures and has developed a concept for the efficient distribution of data and information to the disaster response managements and other recipients with responsibilities in emergency control.

International audience; The main objective of this study was to determine the prevalence of akathisia in a community-dwelling sample of patients with schizophrenia, and to determine the effects of treatments and the clinical variables associated with akathisia. 372 patients with schizophrenia or schizoaffective disorder were systematically included in the network of FondaMental Expert Center for Schizophrenia and assessed with validated scales. Akathisia was measured with the Barnes Akathisia Scale (BAS). Ongoing psychotropic treatment was recorded. The global prevalence of akathisia (as defined by a score of 2 or more on the global akathisia subscale of the BAS) in our sample was 18.5%. Patients who received antipsychotic polytherapy were at higher risk of akathisia and this result remained significant (adjusted odd ratio=2.04, p=.025) after controlling the influence of age, gender, level of education, level of psychotic symptoms, substance use comorbidities, current administration of antidepressant, anticholinergic drugs, benzodiazepines, and daily-administered antipsychotic dose. The combination of second-generation antipsychotics was associated with a 3-fold risk of akathisia compared to second-generation antipsychotics used in monotherapy. Our results indicate that antipsychotic polytherapy should be at best avoided and suggest that monotherapy should be recommended in cases of akathisia. Long-term administration of benzodiazepines or anticholinergic drugs does not seem to be advisable in cases of akathisia, given the potential side effects of these medications.

Objective Abdominal obesity was suggested to be a better predictor than Metabolic Syndrome (MetS) for cardiovascular mortality, however this is has not been extensively studied in schizophrenia. Hyperuricemia (HU) was also suggested to be both an independent risk factor for greater somatic comorbidity and a global metabolic stress marker in patients with schizophrenia. The aim of this study was to estimate the prevalence of MetS, abdominal obesity and HU, to examine the association between metabolic parameters with HU in a cohort of French patients with schizophrenia or schizo-affective disorder (SZ), and to estimate the prevalence rates of treatment of cardio-vascular risk factors. Method 240 SZ patients (age = 31.4 years, male gender 74.3%) were systematically included. Metabolic syndrome was defined according to the International Diabetes Federation and HU if serum uric acid level was above 360 μmol/L. Results MetS, abdominal obesity and HU were found respectively in 24.2%, 21.3% and 19.6% of patients. In terms of risk factors, multiple logistic regression showed that after taking into account the potential confounders, the risk for HU was higher in males (OR = 5.9, IC95 [1.7–21.4]) and in subjects with high waist circumference (OR = 3.1, IC95 [1.1–8.3]) or hypertriglyceridemia (OR = 4.9, IC95 [1.9–13]). No association with hypertension, low HDL cholesterol or high fasting glucose was observed. Only 10% of patients with hypertension received a specific treatment, 18% for high fasting glucose and 8% for dyslipidemia. Conclusions The prevalence of MetS, abdominal obesity and hyperuricemia is elevated in French patients with schizophrenia, all of which are considerably under-diagnosed and undertreated. HU is strongly associated with abdominal obesity but not with psychiatric symptomatology.

Pseudomonas syringae strains deliver variable numbers of type III effector proteins into plant cells during infection. These proteins are required for virulence, because strains incapable of delivering them are nonpathogenic. We implemented a whole-genome, high-throughput screen for identifying P. syringae type III effector genes. The screen relied on FACS and an arabinose-inducible hrpL σ factor to automate the identification and cloning of HrpL-regulated genes. We determined whether candidate genes encode type III effector proteins by creating and testing full-length protein fusions to a reporter called Δ79AvrRpt2 that, when fused to known type III effector proteins, is translocated and elicits a hypersensitive response in leaves of Arabidopsis thaliana expressing the RPS2 plant disease resistance protein. Δ79AvrRpt2 is thus a marker for type III secretion system-dependent translocation, the most critical criterion for defining type III effector proteins. We describe our screen and the collection of type III effector proteins from two pathovars of P. syringae . This stringent functional criteria defined 29 type III proteins from P. syringae pv. tomato , and 19 from P. syringae pv. phaseolicola race 6. Our data provide full functional annotation of the hrpL -dependent type III effector suites from two sequenced P. syringae pathovars and show that type III effector protein suites are highly variable in this pathogen, presumably reflecting the evolutionary selection imposed by the various host plants.

Sequencing the insertion sites of 8,865 Tn917insertions inEnterococcus faecalisstrain OG1RF identified a hot spot in the replication terminus region corresponding to 6% of the genome where 65% of the transposons had inserted. InE. faecalis, Tn917preferentially inserted at a 29-bp consensus sequence centered on TATAA, a 5-bp sequence that is duplicated during insertion. The regional insertion site preference at the chromosome terminus was not observed in another low-G+C gram-positive bacterium,Listeria monocytogenes, although the consensus insertion sequence was the same. The 8,865 Tn917insertion sites sequenced inE. faecaliscorresponded to only ∼610 different open reading frames, far fewer than the predicted number of 2,400, assuming random insertion. There was no significant preference in orientation of the Tn917insertions with either transcription or replication. Even though OG1RF has a smaller genome than strain V583 (2.8 Mb versus 3.2 Mb), the onlyE. faecalisstrain whose sequence is in the public domain, over 10% of the Tn917insertions appear to be in a OG1RF-specific sequence, suggesting that there are significant genomic differences amongE. faecalisstrains.

This chapter discusses the relevance of model host pathogenesis as a third general approach to studying microbial virulence, in which infections are studied in the context of nonvertebrate whole animal hosts. The chapter explains the combination of four key steps: (i) the development of the model host-pathogen system, (ii) the development of genomic tools in both the pathogen and host, (iii) the distribution and use of these tools by the greater research community beyond the laboratories involved in their initial development, and (iv) the collection and ultimate integration of experimental data from a wide variety of research groups made possible by the widespread use of a common resource and its accompanying Web-accessible public database. Pseudomonas aeruginosa PA14 was chosen for the construction of a non-redundant mutant library because it is remarkably virulent in the greatest number of model hosts tested, as well as in a number of murine systems of infection. There are four major advantages to construction of a non-redundant mutant library approach as opposed to the traditional approach of screening a random collection of strains for avirulent or attenuated mutants. A microarray experiment in a Caenhorhabidits elegans mutant defective in a newly defined host defense response gene identified candidate downstream genes important in the response to pathogens.

The synthesis of 1,5-dideoxy-1,5-imino-D-xylitol 2 (and the enantiomeric iminoalditols 3 and 4) is described. Alditol 2 compares favorably with 1-deoxynojirimycin 1 as a potent inhibitor of β-glucosidase.

PATIMDB is a software package for facilitating the generation of transposon mutant insertion libraries. The software has two main functions: process tracking and automated sequence analysis. The process tracking function specifically includes recording the status and fates of multiwell plates and samples in various stages of library construction. Automated sequence analysis refers specifically to the pipeline of sequence analysis starting with ABI files from a sequencing facility and ending with insertion location identifications. The protocols in this unit describe installation and use of PATIMDB software.

Putative essential genes can be identified by comparing orthologs not disrupted in multiple near-saturated transposon insertion mutation libraries in related strains of the same bacterial species. Methods for identifying all orthologs between two bacterial strains and putative essential orthologs are described. In addition, protocols detailing near-saturation transposon insertion mutagenesis of bacteria are presented, including (1) conjugation-mediated mutagenesis, (2) automated colony picking and liquid handling of mutant cultures, and (3) arbitrary polymerase chain reaction amplification and sequencing of genomic DNA adjacent to transposon insertion sites.

The growing number of sequenced bacterial genomes, including those of important pathogenic isolates, has made a significant impact on the field of bacterial pathogenesis (Raskin et al., 2006). Combined with other technical advances in the laboratory, this wealth of information has made possible the development and widespread adoption of genomic tools for the study of infectious disease. This chapter focuses on the sequencing and functional analysis of PA14, a clinical isolate of the ubiquitous environmental bacterium and important opportunistic human pathogen, Pseudomonas aeruginosa. By comparing PA14 to the sequence of the less virulent P. aeruginosa isolate, PAO1, we have identified genomic sequences absent in one or the other strain in order to examine the relationship between genomic content and pathogenicity. To assess the importance of strain-specific genes and virulence, we tested 20 diverse P. aeruginosa strains, including PA14 and PAO1, in a Caenorhabdidits elegans pathogenesis model and observed a wide range of pathogenic potential; however, genotyping these strains using a custom microarray showed that the presence of genes present in PA14 and absent in PAO1 did not correlate with the virulence of these strains. To further examine the roles in virulence of PA14 genes absent in PAO1, we utilized a full-genome non-redundant mutant library of PA14 to identify five ORFs (absent in PAO1) required for C. elegans killing. Surprisingly, although these five genes are present in many other P. aeruginosa strains, they do not correlate with virulence in C. elegans. Genes required for pathogenicity in one strain are neither required for nor predictive of virulence in other strains. We therefore propose that virulence in this organism is a complex process that is both multifactorial and combinatorial. Not only are multiple pathogenicity determinants acting in parallel within a given strain, but also, when comparing different strains, different combinations of pathogenicity factors may be selected to determine the ultimate virulence

Putative essential genes can be identified by comparing orthologs not disrupted in multiple near-saturated transposon insertion mutation libraries in related strains of the same bacterial species. Methods for identifying all orthologs between two bacterial strains and putative essential orthologs are described. In addition, protocols detailing near-saturation transposon insertion mutagenesis of bacteria are presented, including (1) conjugation-mediated mutagenesis, (2) automated colony picking and liquid handling of mutant cultures, and (3) arbitrary polymerase chain reaction amplification and sequencing of genomic DNA adjacent to transposon insertion sites.

The synthesis of 1,5-dideoxy-1,5-imino-D-xylitol 2 (and the enantiomeric iminoalditols 3 and 4) is described. Alditol 2 compares favorably with 1-deoxynojirimycin 1 as a potent inhibitor of β-glucosidase.

microbiology . For the article “Analysis of Pseudomonas aeruginosa diguanylate cyclases and phosphodiesterases reveals a role for bis-(3′-5′)-cyclic-GMP in virulence,” by Hemantha Kulesekara, Vincent …

The opportunistic pathogen Pseudomonas aeruginosa is responsible for systemic infections in immunocompromised individuals and chronic respiratory disease in patients with cystic fibrosis. Cyclic nucleotides are known to play a variety of roles in the regulation of virulence-related factors in pathogenic bacteria. A set of P. aeruginosa genes, encoding proteins that contain putative domains characteristic of diguanylate cyclases (DGCs) and phosphodiesterases (PDEs) that are responsible for the maintenance of cellular levels of the second messenger bis-(3′-5′)-cyclic dimeric GMP (c-di-GMP) was identified in the annotated genomes of P. aeruginosa strains PAO1 and PA14. Although the majority of these genes are components of the P. aeruginosa core genome, several are located on presumptive horizontally acquired genomic islands. A comprehensive analysis of P. aeruginosa genes encoding the enzymes of c-di-GMP metabolism (DGC- and PDE-encoding genes) was carried out to analyze the function of c-di-GMP in two disease-related phenomena, cytotoxicity and biofilm formation. Analysis of the phenotypes of DGC and PDE mutants and overexpressing clones revealed that certain virulence-associated traits are controlled by multiple DGCs and PDEs through alterations in c-di-GMP levels. A set of mutants in selected DGC- and PDE-encoding genes exhibited attenuated virulence in a mouse infection model. Given that insertions in different DGC and PDE genes result in distinct phenotypes, it seems likely that the formation or degradation of c-di-GMP by these enzymes is in highly localized and intimately linked to particular targets of c-di-GMP action.

The ubiquitous bacterium Pseudomonas aeruginosa is the quintessential opportunistic pathogen. Certain isolates infect a broad range of host organisms, from plants to humans. The pathogenic promiscuity of particular variants may reflect an increased virulence gene repertoire beyond the core P. aeruginosa genome. We have identified and characterized two P. aeruginosa pathogenicity islands (PAPI-1 and PAPI-2) in the genome of PA14, a highly virulent clinical isolate. The 108-kb PAPI-1 and 11-kb PAPI-2, which are absent from the less virulent reference strain PAO1, exhibit highly modular structures, revealing their complex derivations from a wide array of bacterial species and mobile elements. Most of the genes within these islands that are homologous to known genes occur in other human and plant bacterial pathogens. For example, PAPI-1 carries a complete gene cluster predicted to encode a type IV group B pilus, a well known adhesin absent from strain PAO1. However, >80% of the PAPI-1 DNA sequence is unique, and 75 of its 115 predicted ORF products are unrelated to any known proteins or functional domains. Significantly, many PAPI-1 ORFs also occur in several P. aeruginosa cystic fibrosis isolates. Twenty-three PAPI ORFs were mutated, and 19 were found to be necessary for full plant or animal virulence, with 11 required for both. The large set of “extra” virulence functions encoded by both PAPIs may contribute to the increased promiscuity of highly virulent P. aeruginosa strains, by directing additional pathogenic functions.

Pseudomonas aeruginosa strain PA14 is an opportunistic human pathogen capable of infecting a wide range of organisms including the nematode Caenorhabditis elegans. We used a non-redundant transposon mutant library consisting of 5,850 clones corresponding to 75% of the total and approximately 80% of the non-essential PA14 ORFs to carry out a genome-wide screen for attenuation of PA14 virulence in C. elegans. We defined a functionally diverse 180 mutant set (representing 170 unique genes) necessary for normal levels of virulence that included both known and novel virulence factors. Seven previously uncharacterized virulence genes (ABC transporters PchH and PchI, aminopeptidase PepP, ATPase/molecular chaperone ClpA, cold shock domain protein PA0456, putative enoyl-CoA hydratase/isomerase PA0745, and putative transcriptional regulator PA14_27700) were characterized with respect to pigment production and motility and all but one of these mutants exhibited pleiotropic defects in addition to their avirulent phenotype. We examined the collection of genes required for normal levels of PA14 virulence with respect to occurrence in P. aeruginosa strain-specific genomic regions, location on putative and known genomic islands, and phylogenetic distribution across prokaryotes. Genes predominantly contributing to virulence in C. elegans showed neither a bias for strain-specific regions of the P. aeruginosa genome nor for putatively horizontally transferred genomic islands. Instead, within the collection of virulence-related PA14 genes, there was an overrepresentation of genes with a broad phylogenetic distribution that also occur with high frequency in many prokaryotic clades, suggesting that in aggregate the genes required for PA14 virulence in C. elegans are biased towards evolutionarily conserved genes., Author Summary Pseudomonas aeruginosa is an opportunistic human pathogen that can also infect a wide range of model organisms, including the nematode Caenorhabditis elegans. To identify P. aeruginosa genes that play key roles in the pathogenic process, we performed a screen for mutants that exhibited reduced ability to kill C. elegans using a previously constructed non-redundant library representing approximately 80% of the non-essential P. aeruginosa PA14 genes. We defined a functionally diverse set of 180 P. aeruginosa mutants (representing 170 unique genes) necessary for normal levels of virulence that included both known and novel virulence factors. The major contributors to P. aeruginosa virulence in the C. elegans infection model were not secretion systems or their corresponding effectors, but rather regulators (particularly ones that are involved in quorum sensing) and genes likely to play key roles in survival of P. aeruginosa within the host intestine. Moreover, these putative P. aeruginosa virulence genes are neither overrepresented in strain-specific regions nor in horizontally acquired genomic islands and furthermore tend to have orthologs that are widely distributed across sequenced prokaryotic species. These data underscore the diversity of pathways involved in virulence, and especially the importance of highly conserved genes for P. aeruginosa virulence in the C. elegans host model.