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Background and objectives: Serum neurofilament light (sNfL) is a biomarker for neuro-axonal damage in multiple sclerosis (MS). Clinical implementation remains limited. We investigated the impact of implementation on clinical decisions using questionnaires at the MS Center Amsterdam, a tertiary outpatient clinic. Methods: sNfL assessments were added to routine clinical practice (August 2021–December 2022). Before and after the results, clinicians filled in questionnaires on context of testing, clinical decisions, certainty herein, expectation of magnetic resonance imaging (MRI) activity, urgency, and motivation to receive the sNfL result and perceived value of sNfL. Results: sNfL was assessed in 166 cases (age 41 ± 12 years, 68% female, 64% disease-modifying therapy (DMT) use) for the following contexts: "DMT monitoring" (55%), "new symptoms" (18%), "differential diagnosis" (17%), and "DMT baseline" (11%). Clinical decisions changed in 19.3% of cases post-disclosure, particularly in context "new symptoms" (38%) and with higher sNfL levels (β = 0.03, p = 0.04). Certainty increased (p = 0.004), while expectation of MRI activity decreased with disclosure of low sNfL levels (p = 0.01). Motivation was highest in context "differential diagnosis" (p < 0.001); perceived value and urgency were highest in context "new symptoms" (p = 0.02). Conclusion: In this study, sNfL implementation had considerable impact on clinical decision-making and certainty herein. Standard implementation may complement patient care but warrants caution and more exploration in diverse clinical settings [ABSTRACT FROM AUTHOR]

Background: Substantial physical-disability worsening in relapsing-remitting multiple sclerosis (RRMS) occurs outside of clinically recorded relapse. This phenomenon, termed progression independent of relapse activity (PIRA), is yet to be established for cognitive decline. Methods: Retrospective analysis of RRMS patients. Cognitive decline was defined using reliable-change-index cut-offs for each test (Symbol Digit Modalities Test, Brief Visuospatial Memory Test–Revised, California Verbal Learning Test–II). Decline was classified as PIRA if the following conditions were met: no relapse observed between assessments nor within 9 months of cognitive decline. Results: The study sample (n = 336) was 80.7% female with a mean (standard deviation (SD)) age, disease duration, and observation period of 43.1 (9.5), 10.8 (8.4), and 8.1 (3.1) years, respectively. A total of 169 (50.3%) subjects were cognitively impaired at baseline relative to age-, sex-, and education-matched HCs. Within subjects who experienced cognitive decline (n = 167), 89% experienced cognitive PIRA. A total of 141 (68.1%) cognitive decline events were observed independent of EDSS worsening. Cognitive PIRA was more likely to be observed with increased assessments (p < 0.001) and lower assessment density (p < 0.001), accounting for baseline clinical factors. Conclusion: These results establish the concept of cognitive PIRA and further our understanding of progressive cognitive decline in RRMS. [ABSTRACT FROM AUTHOR]

Background: Higher age is associated with less inflammatory disease activity in relapsing-remitting multiple sclerosis (RRMS). It is unknown whether age itself or disease duration underlies this association. Objectives: This study investigated the effects of age, disease duration, and inflammatory disease activity in people with RRMS. Methods: Individual patient-level data from five large phase III randomized controlled trials (RCTs) was utilized to investigate the association of both age and disease duration with annualized relapse rate (ARR), contrast-enhancing lesions (CELs), and new T2 lesions on magnetic resonance imaging (MRI) at baseline and follow-up. Results: The data set included 5626 participants. Higher age was associated with lower ARRs, lower CEL number on MRI at baseline and follow-up, and lower new T2 lesion numbers at follow-up. This effect was present in all disease duration groups. For example, we found a lower number of new T2 lesions on MRI during follow-up in higher age groups compared to lower age groups, independent of disease duration. Conclusion: Aging in RRMS is associated with a lower risk of inflammatory disease activity, across different disease durations. Age should be taken into account when designing clinical trials and future research should investigate how age should be integrated into personalized predictions of treatment response and risk profiling. [ABSTRACT FROM AUTHOR]

Background: Humoral responses after SARS-CoV-2 vaccination are greatly impaired in multiple sclerosis (MS) patients on fingolimod. Effects of repeated vaccination and infections on long-term responses are unclear. Methods: Prospective study in 60 MS patients on fingolimod measuring humoral responses after up to four vaccinations and 8 months after fourth vaccination. Results: Anti-WH1 antibody titers increased with each additional vaccination. At long-term follow-up titers increased further and most patients developed new humoral responses against the BA.1 omicron variant. Conclusion: Repeated SARS-CoV-2 vaccinations boost humoral immunity and, probably together with SARS-CoV-2 infections, induce humoral responses on the long-term in almost all patients. [ABSTRACT FROM AUTHOR]

Background: Extended interval dosing (EID) of natalizumab treatment is increasingly used in multiple sclerosis. Besides the clear anti-inflammatory effect, natalizumab is considered to have neuroprotective properties as well. Objectives: This study aimed to study the longitudinal effects of EID compared to standard interval dosing (SID) and natalizumab drug concentrations on brain atrophy. Methods: Patients receiving EID or SID of natalizumab with a minimum radiological follow-up of 2 years were included. Changes in brain atrophy measures over time were derived from clinical routine 3D-Fluid Attenuated Inversion Recovery (FLAIR)-weighted magnetic resonance imaging (MRI) scans using SynthSeg. Results: We found no differences between EID (n = 32) and SID (n = 50) for whole brain (−0.21% vs −0.16%, p = 0.42), ventricular (1.84% vs 1.13%, p = 0.24), and thalamic (−0.32% vs −0.32%, p = 0.97) annualized volume change over a median follow-up of 3.2 years. No associations between natalizumab drug concentration and brain atrophy rate were found. Conclusion: We found no clear evidence that EID compared to SID or lower natalizumab drug concentrations have a negative impact on the development of brain atrophy over time. [ABSTRACT FROM AUTHOR]

Background: Whether the degree of inflammation (and its resolution) and neurodegeneration after treatment initiation predicts disease progression in multiple sclerosis (MS) remains unclear. Objectives: To assess the predictive value of magnetic resonance imaging (MRI)-derived brain and lesion volume (LV) changes in years 1 and 2 of treatment for disease progression. Methods: Patients receiving early interferon beta-1a treatment in REFLEX/REFLEXION (N = 262) were included. Predictive regression models included new/enlarging LV (positive activity), disappearing/shrinking LV (negative activity), and global/central atrophy during years 1 and 2. Results: Faster global atrophy and/or pseudo-atrophy and positive lesion activity in years 1 and 2 related to an increased probability and faster conversion to clinically definite multiple sclerosis (CDMS). Negative lesion activity in year 1 and slower central atrophy in year 2 were predictive of confirmed disability progression (9-Hole Peg Test). Positive lesion activity in year 2 was predictive of faster global atrophy, while positive lesion activity in years 1 and 2 was predictive of faster central atrophy. Conclusions: A higher degree of global atrophy and/or pseudo-atrophy in year 1 was predictive of CDMS. Positive lesion activity in any year was related to CDMS and neurodegeneration. Disability was related to negative lesion activity in year 1 and slower central atrophy in year 2. [ABSTRACT FROM AUTHOR]

Background: Clinical relapses are the defining feature of relapsing forms of multiple sclerosis (MS), but relatively little is known about the time course of relapse recovery. Objective: The aim of this study was to investigate the time course of and patient factors associated with the speed and success of relapse recovery in people with relapsing–remitting MS (RRMS). Methods: Using data from CombiRx, a large RRMS trial (clinicaltrials.gov identifier NCT00211887), we measured the time to recovery from the first on-trial relapse. We used Kaplan–Meier survival analyses and Cox regression models to investigate the association of patient factors with the time to unconfirmed and confirmed relapse recovery. Results: CombiRx included 1008 participants. We investigated 240 relapses. Median time to relapse recovery was 111 days. Most recovery events took place within 1 year of relapse onset: 202 of 240 (84%) individuals recovered during follow-up, 161 of 202 (80%) by 180 days, and 189 of 202 (94%) by 365 days. Relapse severity was the only factor associated with relapse recovery. Conclusion: Recovery from relapses takes place up to approximately 1 year after the event. Relapse severity, but no other patient factors, was associated with the speed of relapse recovery. Our findings inform clinical practice and trial design in RRMS. [ABSTRACT FROM AUTHOR]

Background: There is a need in Relapsing-Remitting Multiple Sclerosis (RRMS) treatment for biomarkers that monitor neuroinflammation, neurodegeneration, treatment response, and disease progression despite treatment. Objective: To assess the value of serum glial fibrillary acidic protein (sGFAP) as a biomarker for clinical disease progression and brain volume measurements in natalizumab-treated RRMS patients. Methods: sGFAP and neurofilament light (sNfL) were measured in an observational cohort of natalizumab-treated RRMS patients at baseline, +3, +12, and +24 months and at the last sample follow-up (median 5.17 years). sGFAP was compared between significant clinical progressors and non-progressors and related to magnetic resonance imaging (MRI)-derived volumes of the whole brain, ventricle, thalamus, and lesion. The relationship between sGFAP and sNfL was assessed. Results: A total of 88 patients were included, and 47.7% progressed. sGFAP levels at baseline were higher in patients with gadolinium enhancement (1.3-fold difference, p = 0.04) and decreased in 3 months of treatment (adj. p < 0.001). No association was found between longitudinal sGFAP levels and progressor status. sGFAP at baseline and 12 months was significantly associated with normalized ventricular (positively), thalamic (negatively), and lesion volumes (positively). Baseline and 12-month sGFAP predicted annualized ventricle volume change rate after 1 year of treatment. sGFAP correlated with sNfL at baseline (p < 0.001) and last sample follow-up (p < 0.001) but stabilized earlier. Discussion: sGFAP levels related to MRI markers of neuroinflammation and neurodegeneration. [ABSTRACT FROM AUTHOR]

Background: Suboptimal performance during neuropsychological testing frequently occurs in multiple sclerosis (MS), leading to unreliable cognitive outcomes. Neurophysiological alterations correlate with MS-related cognitive impairment, but studies have not yet considered performance validity. Objectives: To investigate neurophysiological markers of cognitive impairment in MS, while explicitly addressing performance validity. Methods: Magnetoencephalography recordings, neuropsychological assessments, and performance validity testing were obtained from 90 MS outpatients with cognitive complaints. Spectral and resting-state functional connectivity (rsFC) properties were compared between cognitively impaired (CI), cognitively preserved (CP), and suboptimally performing (SUB) patients using regression models and permutation testing. Results: CI had higher power in low-frequency bands and lower power in high bands compared to CP, indicating neuronal slowing. CI also showed lower beta power compared to SUB. Overall power spectra visually differed between CI and CP, and SUB showed overlap with both groups. CI had lower rsFC than CP and SUB patients. CP and SUB patients showed no differences. Conclusion: Neuronal slowing and altered rsFC can be considered cognitive markers in MS. Patients who performed suboptimally showed resemblance with patients with and without cognitive impairments, and although their overall neurophysiological profile was more similar to patients without impairments, it suggests heterogeneity regarding their pathophysiology. [ABSTRACT FROM AUTHOR]

Background: Patient-reported outcome measures (PROMs) are often used in clinical research, but little is known about their performance as longitudinal outcomes. Methods: We used data from ASCEND, a large SPMS trial (n = 889), to investigate changes on the Short Form Health Survey 36 (SF-36 v2) and the Multiple Sclerosis Impact Scale (MSIS-29) over 2 years of follow-up. Results: PROM scores changed little over the 2 years of follow-up. In contrast to physical disability measures, there was no consistent trend in PROM change: significant worsening occurred about as often as improvement. Using a 6-month confirmation reduced the number of both worsening and improvement events without altering their relative balance. There was no clear difference in worsening events in groups based on population characteristics, nor was there a noticeable effect using different thresholds for clinically significant change. Conclusion: We found little consistent change in MSIS-29 and SF-36 over 2 years of follow-up in people with SPMS. Our findings show a disconnect between disability worsening and PROM change in this population. Our findings raise caution about the use of these PROMs as primary outcome measures in SPMS trials and call for a critical reappraisal of the longitudinal use of these measures in SPMS trials. [ABSTRACT FROM AUTHOR]

In this observational study, 159 patients with multiple sclerosis received personalized dosing of ocrelizumab incentivized by the COVID-19 pandemic. Re-dosing was scheduled when CD19 B-cell count was ⩾10 cells/µL (starting 24 weeks after the previous dose, repeated 4-weekly). Median interval until re-dosing or last B-cell count was 34 [30–38] weeks. No clinical relapses were reported and a minority of patients showed Expanded Disability Status Scale (EDSS) progression. Monthly serum neurofilament light levels remained stable during extended intervals. Two (1.9%) of 107 patients with a follow-up magnetic resonance imaging (MRI) scan showed radiological disease activity. Personalized dosing of ocrelizumab could significantly extend intervals with low short-term disease activity incidence, encouraging future research on long-term safety and efficacy. [ABSTRACT FROM AUTHOR]

In women with very active multiple sclerosis (MS), natalizumab can be continued during pregnancy to prevent rebound disease activity. Our aim was to evaluate changes in serum natalizumab trough concentrations during pregnancy. Blood samples of 3 patients were collected before, during, and after pregnancy. Natalizumab trough concentrations gradually decreased during pregnancy. The patient with the lowest trough concentrations during the third trimester was treated with extended interval dosing (EID). After delivery, natalizumab concentrations increased to similar levels as before pregnancy. All patients remained clinically and radiologically stable. MS neurologists should be aware of decreasing natalizumab concentrations during pregnancy, especially in patients with low initial trough concentrations and patients with EID. [ABSTRACT FROM AUTHOR]

Background: Cognitive decline remains difficult to predict as structural brain damage cannot fully explain the extensive heterogeneity found between MS patients. Objective: To investigate whether functional brain network organization measured with magnetoencephalography (MEG) predicts cognitive decline in MS patients after 5 years and to explore its value beyond structural pathology. Methods: Resting-state MEG recordings, structural MRI, and neuropsychological assessments were analyzed of 146 MS patients, and 100 patients had a 5-year follow-up neuropsychological assessment. Network properties of the minimum spanning tree (i.e. backbone of the functional brain network) indicating network integration and overload were related to baseline and longitudinal cognition, correcting for structural damage. Results: A more integrated beta band network (i.e. smaller diameter) and a less integrated delta band network (i.e. lower leaf fraction) predicted cognitive decline after 5 years ( R adj 2 = 15 %), independent of structural damage. Cross-sectional analyses showed that a less integrated network (e.g. lower tree hierarchy) related to worse cognition, independent of frequency band. Conclusions: The level of functional brain network integration was an independent predictive marker of cognitive decline, in addition to the severity of structural damage. This work thereby indicates the promise of MEG-derived network measures in predicting disease progression in MS. [ABSTRACT FROM AUTHOR]

Background: Neurophysiological measures of brain function, such as magnetoencephalography (MEG), are widely used in clinical neurology and have strong relations with cognitive impairment and dementia but are still underdeveloped in multiple sclerosis (MS). Objectives: To demonstrate the value of clinically applicable MEG-measures in evaluating cognitive impairment in MS. Methods: In eyes-closed resting-state, MEG data of 83 MS patients and 34 healthy controls (HCs) peak frequencies and relative power of six canonical frequency bands for 78 cortical and 10 deep gray matter (DGM) areas were calculated. Linear regression models, correcting for age, gender, and education, assessed the relation between cognitive performance and MEG biomarkers. Results: Increased alpha1 and theta power was strongly associated with impaired cognition in patients, which differed between cognitively impaired (CI) patients and HCs in bilateral parietotemporal cortices. CI patients had a lower peak frequency than HCs. Oscillatory slowing was also widespread in the DGM, most pronounced in the thalamus. Conclusion: There is a clinically relevant slowing of neuronal activity in MS patients in parietotemporal cortical areas and the thalamus, strongly related to cognitive impairment. These measures hold promise for the application of resting-state MEG as a biomarker for cognitive disturbances in MS in a clinical setting. [ABSTRACT FROM AUTHOR]

Only a small proportion of all paediatric fractures is caused by child abuse or neglect, especially in highly prevalent long bone fractures. It can be d ifficu lt to differentiate abusive fractures from non-abusive fractures. This article focuses on femoral fractures in young children. Based on three cases, this article presents a forensic evidence-based approach to differentiate between accidental and non-accidental causes of femoral fractures. We describe three cases of young children who were presented to the emergency department because of a suspected femur fracture. Although in all cases, the fracture had a similar location and appearance, the clinical history and developmental stage of the child led to three different conclusions. In the first two cases, an accidental mechanism was a plausible conclusion, although in the second case, neglect of parental supervision was the cause for concern. In the third case, a non-accidental injury was diagnosed and appropriate legal prosecution followed. Any do ctor treating children should always be aware of the possibility of child abuse and neglect in children with injuries, especially in young and non-mobile children presenting with an unknown trauma mechanism. If a suspicion of child abuse or neglect arises, a thorough diagnostic work-up should be performed, including a full skeletal survey according to the guidelines of the Royal College of Radiologists and the Royal College of Paediatrics and Child Health. In order to make a good assessment, the radiologist reviewing the skeletal survey needs access to all relevant clinical and social information. [ABSTRACT FROM AUTHOR]

The article presents a study on the implication of serum natalizumab concentration and anti-natalizumab antibodies in multiple sclerosis. It mentions that 73 patients were used in the prospective observational cohort study, who underwent treatment of natalizumab. It states that radioimmunoassay was used to measure antibodies opposing natalizumab. The study found that high antibody titers and low natalizumab serum concentration are related with natalizumab's limited efficacy.

The article discusses research which examines the possible associations of glutamate related single nucleotide polymorphisms (SNPs) and multiple magnetic resonance imaging (MRI)-derived phenotypes in multiple sclerosis (MS). The study analysis seven phenotypes related to brain and lesion volumes using the sparse reduced-rank regression (sRRR) statistical model. It reveals the associations between single nucleotide polymorphisms (SNPs) and phenotypic variation in normalized brain volume (NBV).

Background: Biomarkers of neuronal and axonal damage (serum neurofilament light (sNfL) and serum glial fibrillary acidic protein (sGFAP)) may provide insight into the aetiology of natalizumab wearing-off symptoms (WoSs)., Objectives: We investigated the longitudinal association between and predictive value of sNfL and sGFAP and the occurrence of WoS in MS patients treated with natalizumab., Methods: We performed longitudinal measurements of sNfL and sGFAP in NEXT-MS trial participants who completed a questionnaire about WoS., Results: A total of 364 participants were included. In total, 55.5% presented with WoS and 44.5% without WoS during natalizumab treatment. Longitudinal analyses showed no association between sNfL and sGFAP levels and WoS at any timepoint. Biomarker levels at baseline did not predict first-time WoS occurrence., Conclusion: Acute and chronic neuronal and axonal damage are most likely not the underlying cause of WoS., Competing Interests: Declaration of Conflicting InterestsThe author(s) declared the following potential conflicts of interest with respect to the research, authorship and/or publication of this article: A.A.T. has nothing to disclose. M.H.J.W. has nothing to disclose. L.B. has nothing to disclose. L.M.Y.G. has nothing to disclose. E.H. has accepted (speaker and congress) fees from Merck Serono, Biogen Idec, Roche and Sanofi Genzyme. E.M.P.E.Z. reports advisory boards/consultancy fees for Merck, Novartis, Genzyme and Roche. L.C.v.R. has nothing to disclose. C.E.P.v.M. has nothing to disclose. A.V. has nothing to disclose. J.P.M. has nothing to disclose. B.H.A.W. has nothing to disclose. N.F.K. has nothing to disclose. E.L.J.H. has nothing to disclose. J.J.J.v.E. reports honoraria for advisory boards and/or speakers fee from Merck Serono, Biogen Idec, Sanofi Genzyme, Roche and Novartis. C.M.R. has nothing to disclose. J.J.K. has nothing to disclose. M.E. has nothing to disclose. J.N. has nothing to disclose. J.v.G. has nothing to disclose. L.G.F.S. has nothing to disclose. M.E.K. has nothing to disclose. E.P.J.A. has nothing to disclose. G.W.v.D. has nothing to disclose. W.H.B. has nothing to disclose. E.M.M.S. has nothing to disclose. B.W.v.O. has nothing to disclose. B.A.d.J. has nothing to disclose. B.M.J.U. reports research support and/or consultancy fees from Genzyme, Biogen Idec, Novartis, Teva Pharmaceutical Industries, Merck Serono, Roche and Immunic Therapeutics. T.R. received funding for research from Genmab and consultancy fees from Novartis. J.K. received research grants for multicentre investigator-initiated trials DOT-MS trial, ClinicalTrials.gov Identifier: NCT04260711 (ZonMW) and BLOOMS trial (ZonMW and Treatmeds) (ClinicalTrials.gov Identifier: NCT05296161); received consulting fees for F. Hoffmann-La Roche Ltd, Biogen, Teva, Merck, Novartis and Sanofi/Genzyme (all payments to institution); reports speaker relationships with F. Hoffmann-La Roche Ltd, Biogen, Immunic, Teva, Merck, Novartis and Sanofi/Genzyme (all payments to institution) and adjudication committee of MS clinical trial of Immunic (payments to institution only). Z.L.E.v.K. has nothing to disclose. C.E.T. reports funding from National MS Society (Progressive MS alliance) and Innovative Medicines Initiatives 3TR (Horizon 2020, grant no. 831434); has a research contract with Celgene; serves on editorial boards of Medidact Neurologie/Springer, Neurology: Neuroimmunology & Neuroinflammation and is editor of a Neuromethods book Springer.

Background: Extended interval dosing (EID) of natalizumab is a promising strategy to optimise treatment in multiple sclerosis (MS). Personalised EID by therapeutic drug monitoring can enable further extension of treatment intervals., Methods: The NEXT-MS trial is an investigator-initiated prospective phase IV non-randomised study. Adults with a diagnosis of relapsing-remitting MS who received ≥6 natalizumab infusions were included in three groups: personalised EID with a target drug trough concentration of 10 µg/mL (EID10), an exploratory group of personalised EID with a target of 5 µg/mL (EID5) and standard interval dosing (SID) of 4 weeks. The primary outcome is radiological disease activity (new/newly enlarged T2 lesions) comparing the EID10 group to a historical cohort of SID (HSID)., Results: Results of the first phase of the NEXT-MS trial are reported here (n=376) as the study will continue with an amended protocol. In the EID10 group (n=251), incidence rate of radiological activity was 10.0 per 1000 person-years, which was non-inferior to the HSID cohort (24.7 per 1000 person-years (n=87), incidence rate difference 14.7, 90% CI -4.5 to 34.0). Incidence rate of radiological activity was 10.0 per 1000 person-years in the EID5 group (n=65), and 47.0 per 1000 person-years in the SID group (n=60). Serum neurofilament light levels did not increase over time within the EID groups. There were no cases of progressive multifocal leukoencephalopathy., Conclusions: MS disease activity is adequately controlled with personalised natalizumab EID. Interval extension to a drug trough concentration of 5 µg/mL is likely a safe target to extend natalizumab treatment intervals >6 weeks., Trial Registration Number: NCT04225312., Competing Interests: Competing interests: EH: has accepted (speaker and congress) fees from Merck Serono, Biogen Idec, Roche and Sanofi Genzyme. EMPEZ: reports advisory boards/consultancy fees for Merck, Novartis, Genzyme and Roche. JvE: reports honoraria for advisory boards and/or speakers fee from Merck Serono, Biogen Idec, Sanofi Genzyme, Roche and Novartis. FB: Steering committee and iDMC for Biogen, Merck, Roche, EISAI. Consultant to Roche, Biogen, Merck, IXICO, Jansen, Combinostics. Research agreements with Novartis, Merck, Biogen, GE, Roche. Co-founder of Queen Square Analytics. Funding by NIHR-UCLH-BRC, Novartis, GE, UKMSS, MAGNIMS-ECTRIMS, EC-H2020, EC-JU (IMI), EPSRC. Editorial board member for Brain, MSJ, Neurology, Radiology and Neuroradiology (section editor). CET: reports funding from National MS Society (Progressive MS alliance) and Innovative Medicines Initiatives 3TR (Horizon 2020, grant no 831434); has a research contract with Celgene; serves on editorial boards of Medidact Neurologie/Springer, Neurology: Neuroimmunology and Neuroinflammation, and is editor of a Neuromethods book Springer. TR received funding for research from Genmab and consultancy fees from Novartis. BMJU: reports research support and/or consultancy fees from Genzyme, Biogen Idec, Novartis, Teva Pharmaceutical Industries, Merck Serono, Roche, and Immunic Therapeutics. JK: received research grants for multicentre investigator initiated trials DOT-MS trial, ClinicalTrials.gov Identifier: NCT04260711 (ZonMW) and BLOOMS trial (ZonMW and Treatmeds), ClinicalTrials.gov Identifier: NCT05296161); received consulting fees for F. Hoffmann-La Roche Ltd, Biogen, Teva, Merck, Novartis and Sanofi/Genzyme (all payments to institution); reports speaker relationships with F. Hoffmann-La Roche, Biogen, Immunic, Teva, Merck, Novartis and Sanofi/Genzyme (all payments to institution); adjudication committee of MS clinical trial of Immunic (payments to institution only)., (© Author(s) (or their employer(s)) 2024. No commercial re-use. See rights and permissions. Published by BMJ.)

Salmonella is an intracellular bacterium found in the gastrointestinal tract of mammalian, avian, and reptilian hosts. Mouse models have been extensively used to model in vivo distinct aspects of human Salmonella infections and have led to the identification of several host susceptibility genes. We have investigated the susceptibility of Collaborative Cross strains to intravenous infection with Salmonella enterica serovar Typhimurium as a model of human systemic invasive infection. In this model, strain CC042/GeniUnc (CC042) mice displayed extreme susceptibility with very high bacterial loads and mortality. CC042 mice showed lower spleen weights and decreased splenocyte numbers before and after infection, affecting mostly CD8 + T cells, B cells, and all myeloid cell populations, compared with control C57BL/6J mice. CC042 mice also had lower thymus weights with a reduced total number of thymocytes and double-negative and double-positive (CD4 + , CD8 + ) thymocytes compared to C57BL/6J mice. Analysis of bone marrow-resident hematopoietic progenitors showed a strong bias against lymphoid-primed multipotent progenitors. An F2 cross between CC042 and C57BL/6N mice identified two loci on chromosome 7 ( Stsl6 and Stsl7 ) associated with differences in bacterial loads. In the Stsl7 region, CC042 carried a loss-of-function variant, unique to this strain, in the integrin alpha L ( Itgal ) gene, the causative role of which was confirmed by a quantitative complementation test. Notably, Itgal loss of function increased the susceptibility to S. Typhimurium in a (C57BL/6J × CC042)F1 mouse background but not in a C57BL/6J mouse inbred background. These results further emphasize the utility of the Collaborative Cross to identify new host genetic variants controlling susceptibility to infections and improve our understanding of the function of the Itgal gene., (Copyright © 2019 American Society for Microbiology.)

Salmonella presents a global public health concern. Central to Salmonella pathogenicity is an ability to subvert host defences through strategically targeting host proteins implicated in restricting infection. Therefore, to gain insight into the host-pathogen interactions governing Salmonella infection, we performed an in vivo genome-wide mutagenesis screen to uncover key host defence proteins. This revealed an uncharacterized role of CYRI (FAM49B) in conferring host resistance to Salmonella infection. We show that CYRI binds to the small GTPase RAC1 through a conserved domain present in CYFIP proteins, which are known RAC1 effectors that stimulate actin polymerization. However, unlike CYFIP proteins, CYRI negatively regulates RAC1 signalling, thereby attenuating processes such as macropinocytosis, phagocytosis and cell migration. This enables CYRI to counteract Salmonella at various stages of infection, including bacterial entry into non-phagocytic and phagocytic cells as well as phagocyte-mediated bacterial dissemination. Intriguingly, to dampen its effects, the bacterial effector SopE, a RAC1 activator, selectively targets CYRI following infection. Together, this outlines an intricate host-pathogen signalling interplay that is crucial for determining bacterial fate. Notably, our study also outlines a role for CYRI in restricting infection mediated by Mycobacterium tuberculosis and Listeria monocytogenes. This provides evidence implicating CYRI cellular functions in host defence beyond Salmonella infection.

Evidence from prior studies has shown an advantage in recognition memory for emotional compared to neutral words. Whether this advantage is short-lived or rather extends over longer periods, as well as whether the effect depends on words' valence (i.e., positive or negative), remains unknown. In the present ERP/EEG study, we investigated this issue by manipulating the lag distance (LAG-2, LAG-8 and LAG-16) between the presentation of old and new words in an online recognition memory task. LAG differences were observed at behavior, ERPs and in the theta frequency band. In line with previous studies, negative words were associated with faster reaction times, higher hit rates and increased amplitude in a positive ERP component between 386 and 564 ms compared to positive and neutral words. Remarkably, the interaction of LAG by EMOTION revealed that negative words were associated with better performance and larger ERPs amplitudes only at LAG-2. Also in the LAG-2 condition, emotional words (i.e., positive and negative words) induced a stronger desynchronization in the beta band between 386 and 542 ms compared to neutral words. These early enhanced memory effects for emotional words are discussed in terms of the Negative Emotional Valence Enhances Recapitulation (NEVER) model and the mobilization-minimization hypothesis., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Viral respiratory tract infections are the main causative agents of the onset of infection-induced asthma and asthma exacerbations that remain mechanistically unexplained. Here we found that deficiency in signaling via type I interferon receptor led to deregulated activation of group 2 innate lymphoid cells (ILC2 cells) and infection-associated type 2 immunopathology. Type I interferons directly and negatively regulated mouse and human ILC2 cells in a manner dependent on the transcriptional activator ISGF3 that led to altered cytokine production, cell proliferation and increased cell death. In addition, interferon-γ (IFN-γ) and interleukin 27 (IL-27) altered ILC2 function dependent on the transcription factor STAT1. These results demonstrate that type I and type II interferons, together with IL-27, regulate ILC2 cells to restrict type 2 immunopathology.

The outcome of infection with Salmonella Typhimurium in mouse models of human typhoid fever is dependent upon a coordinated complex immune response. A panel of recombinant congenic strains (RCS) derived from reciprocal backcross of A/J and C57BL/6J mice was screened for their susceptibility to Salmonella infection and two susceptibility loci, Ity4 (Immunity to Typhimurium locus 4) and Ity5, were identified. We validated Ity5 in a genetic environment free of the impact of Ity4 using a cross between A/J and 129S6. Using a time-series analysis of genome-wide transcription during infection, comparing A/J with AcB60 mice having a C57BL/6J-derived Ity5 interval, we have identified the differential expression of the positional candidate gene Cd40, Cd40-associated signaling pathways, and the differential expression of numerous genes expressed in neutrophils. CD40 is known to coordinate T cell-dependent B-cell responses and myeloid cell activation. In fact, CD40 signaling is altered in A/J mice as seen by impaired IgM upregulation during infection, decreased Ig class switching, neutropenia, reduced granulocyte recruitment in response to infection and inflammation, and decreased ERK1/2 activity. These results suggest that altered CD40 signaling and granulocyte recruitment in response to infection are responsible for the Ity5-associated Salmonella susceptibility of A/J mice.

Infectious diseases are responsible for over 25% of deaths globally, but many more individuals are exposed to deadly pathogens. The outcome of infection results from a set of diverse factors including pathogen virulence factors, the environment, and the genetic make-up of the host. The completion of the human reference genome sequence in 2004 along with technological advances have tremendously accelerated and renovated the tools to study the genetic etiology of infectious diseases in humans and its best characterized mammalian model, the mouse. Advancements in mouse genomic resources have accelerated genome-wide functional approaches, such as gene-driven and phenotype-driven mutagenesis, bringing to the fore the use of mouse models that reproduce accurately many aspects of the pathogenesis of human infectious diseases. Treatment with the mutagen N-ethyl-N-nitrosourea (ENU) has become the most popular phenotype-driven approach. Our team and others have employed mouse ENU mutagenesis to identify host genes that directly impact susceptibility to pathogens of global significance. In this review, we first describe the strategies and tools used in mouse genetics to understand immunity to infection with special emphasis on chemical mutagenesis of the mouse germ-line together with current strategies to efficiently identify functional mutations using next generation sequencing. Then, we highlight illustrative examples of genes, proteins, and cellular signatures that have been revealed by ENU screens and have been shown to be involved in susceptibility or resistance to infectious diseases caused by parasites, bacteria, and viruses.

Host genetics has a key role in susceptibility to Salmonella Typhimurium infection. We previously used N-ethyl-N-nitrosourea (ENU) mutagenesis to identify a loss-of-function mutation within the gene ubiquitin-specific peptidase 18 (Usp18(Ity9)), which confers increased susceptibility to Salmonella Typhimurium. USP18 functions to regulate type I interferon (IFN) signaling and as a protease to remove ISG15 from substrate proteins. Usp18(Ity9) mice are susceptible to infection with Salmonella Typhimurium and have increased expression and function of ISG15, but Usp18(Ity9) mice lacking Isg15 do not show improved survival with Salmonella challenge. Type I IFN signaling is increased in Usp18(Ity9) mice and inhibition of type I IFN signaling is associated with improved survival in mutant mice. Hyperactivation of type I IFN signaling leads to increased IL-10, deregulated expression of autophagy markers and elevated interleukin (IL)-1β and IL-17. Furthermore, Usp18(Ity9) mice are more susceptible to infection with Mycobacterium tuberculosis, have increased bacterial load in the lung and spleen, elevated inflammatory cytokines and more severe lung pathology. These findings demonstrate that regulation of type I IFN signaling is the predominant mechanism affecting the susceptibility of Usp18(Ity9) mice to Salmonella infection and that hyperactivation of signaling leads to increased IL-10, deregulation of autophagic markers and increased proinflammatory cytokine production.

Salmonella enterica is a ubiquitous Gram-negative intracellular bacterium that continues to pose a global challenge to human health. The etiology of Salmonella pathogenesis is complex and controlled by pathogen, environmental, and host genetic factors. In fact, patients immunodeficient in genes in the IL-12, IL-23/IFN-γ pathway are predisposed to invasive nontyphoidal Salmonella infection. Using a forward genomics approach by N-ethyl-N-nitrosourea (ENU) germline mutagenesis in mice, we identified the Ity14 (Immunity to Typhimurium locus 14) pedigree exhibiting increased susceptibility following in vivo Salmonella challenge. A DNA-binding domain mutation (p.G418&#95;E445) in Stat4 (Signal Transducer and Activator of Transcription Factor 4) was the causative mutation. STAT4 signals downstream of IL-12 to mediate transcriptional regulation of inflammatory immune responses. In mutant Ity14 mice, the increased splenic and hepatic bacterial load resulted from an intrinsic defect in innate cell function, IFN-γ-mediated immunity, and disorganized granuloma formation. We further show that NK and NKT cells play an important role in mediating control of Salmonella in Stat4(Ity14/Ity14) mice. Stat4(Ity14/Ity14) mice had increased expression of genes involved in cell-cell interactions and communication, as well as increased CD11b expression on a subset of splenic myeloid dendritic cells, resulting in compromised recruitment of inflammatory cells to the spleen during Salmonella infection. Stat4(Ity14/Ity14) presented upregulated compensatory mechanisms, although inefficient and ultimately Stat4(Ity14/Ity14) mice develop fatal bacteremia. The following study further elucidates the pathophysiological impact of STAT4 during Salmonella infection.

Background: Although it is often performed in clinical practice, the diagnostic value of a screening physical examination to detect maltreatment in children without prior suspicion has not been reviewed. This article aims to evaluate the diagnostic value of a complete physical examination as a screening instrument to detect maltreatment in children without prior suspicion., Methods: We systematically searched the databases of MEDLINE, EMBASE, PsychINFO, CINAHL, and ERIC, using a sensitive search strategy. Studies that i) presented medical findings of a complete physical examination for screening purposes in children 0-18 years, ii) specifically recorded the presence or absence of signs of child maltreatment, and iii) recorded child maltreatment confirmed by a reference standard, were included. Two reviewers independently performed study selection, data extraction, and quality appraisal using the QUADAS-2 tool., Results: The search yielded 4,499 titles, of which three studies met the eligibility criteria. The prevalence of confirmed signs of maltreatment during screening physical examination varied between 0.8% and 13.5%. The designs of the studies were inadequate to assess the diagnostic accuracy of a screening physical examination for child maltreatment., Conclusions: Because of the lack of informative studies, we could not draw conclusions about the diagnostic value of a screening physical examination in children without prior suspicion of child maltreatment.

Using the N-ethyl-N-nitrosourea (ENU) mutagenesis screen, we have identified two skeletal morphology mutants, Skm1 and Skm2. Positional cloning and candidate gene sequencing localized the causative point mutations within the genes coding for natriuretic peptide receptor C (NPR-C) and filamin b (FLNB), respectively. Mice that carry a mutation in Npr3 exhibit a skeletal overgrowth phenotype, resulting in an elongated body and kyphosis. Skm2 mice, carrying a mutation in Flnb, present with scoliosis and lordosis. These mutant mice will serve as useful models for the study of vertebral malformations.

The Gram-negative bacteria, Salmonella, cause a broad spectrum of clinical diseases in humans, ranging from asymptomatic carriage to life-threatening sepsis. We have designed an experimental model to study the contribution of genetic factors to the persistence of Salmonella Enteritidis during the late phase of infection in 129S6/SvEvTac and C57BL/6J mice. C57BL/6J mice cleared the bacteria from their reticuloendothelial system within a period of 42 days, whereas the 129S6 mice still presented a high bacterial load. Using this model, we have identified ten Salmonella Enteritidis susceptibility loci (Ses1, Ses1.1, and Ses3-Ses10) associated with bacterial persistence in target organs of 129S6/SvEvTac mice using a two-locus epistasis QTL linkage mapping approach. Significant statistical interactions were detected between Ses1 on chromosome 1 and Ses5 on chromosome 7 and between Ses1 and Ses4 on chromosome X. In this study, we functionally validated the genetic architecture of Salmonella persistence in 129S6 mice using single- (129S6.B6-Ses1.2 that combines Ses1 and Ses1.1 loci, 129S6.B6-Ses4, and 129S6.B6-Ses5) and double-congenic mice (129S6.B6-Ses1.2/Ses4 and 129S6.B6-Ses1.2/Ses5). These experiments demonstrate functional interactions between Ses1.2 and Ses4 or Ses5 that improve Salmonella Enteritidis clearance, validating the critical role played by gene-gene interactions in the contribution to bacterial clearance heritability. Improved bacterial clearance in double-congenic mice could be explained by the impact of Ses4 and Ses5 in combination with Ses1.2 on TH polarization since a TH2 bias (decreased Ifng and increased Il4 mRNA levels and reduced IgG2a immunoglobulins in the serum) was observed in 129S6.B6-Ses1.2/Ses5 mice and a TH17 (high Il17 expression) bias in 129S6.B6-Ses1.2/Ses4.

Salmonella, a ubiquitous Gram-negative intracellular bacterium, is a food borne pathogen that infects a broad range of hosts. Infection with Salmonella Typhimurium in mice is a broadly recognized experimental model resembling typhoid fever in humans. Using a N-ethyl-N-nitrosurea (ENU) mutagenesis recessive screen, we report the identification of Ity16 (Immunity to Typhimurium locus 16), a locus responsible for increased susceptibility to infection. The position of Ity16 was refined on chromosome 8 and a nonsense mutation was identified in the ankyrin 1 (Ank1) gene. ANK1 plays an important role in the formation and stabilization of the red cell cytoskeleton. The Ank1(Ity16/Ity16) mutation causes severe hemolytic anemia in uninfected mice resulting in splenomegaly, hyperbilirubinemia, jaundice, extramedullary erythropoiesis and iron overload in liver and kidneys. Ank1(Ity16/Ity16) mutant mice demonstrated low levels of hepcidin (Hamp) expression and significant increases in the expression of the growth differentiation factor 15 (Gdf15), erythropoietin (Epo) and heme oxygenase 1 (Hmox1) exacerbating extramedullary erythropoiesis, tissue iron deposition and splenomegaly. As the infection progresses in Ank1(Ity16/Ity16), the anemia worsens and bacterial load were high in liver and kidneys compared to wild type mice. Heterozygous Ank1(+/Ity16) mice were also more susceptible to Salmonella infection although to a lesser extent than Ank1(Ity16/Ity16) and they did not inherently present anemia and splenomegaly. During infection, iron accumulated in the kidneys of Ank1(+/Ity16) mice where bacterial loads were high compared to littermate controls. The critical role of HAMP in the host response to Salmonella infection was validated by showing increased susceptibility to infection in Hamp-deficient mice and significant survival benefits in Ank1(+/Ity16) heterozygous mice treated with HAMP peptide. This study illustrates that the regulation of Hamp and iron balance are crucial in the host response to Salmonella infection in Ank1 mutants.