Prevalence and antimicrobial resistance of Salmonella enterica subsp. diarizonae serovar 61:k:1,5,(7) in Swiss sheep flocks.

Salmonella (S.) enterica subspecies diarizonae (IIIb) serovar 61:k:1,5,(7) (S. IIIb 61:k:1,5,(7)) is considered to be sheep-associated, as it can be found in the intestine, tonsils and nose of clinically healthy sheep, but it has also been described in separate clinical disorders in sheep. In particular, S. IIIb 61:k:1,5,(7) is described as the causative agent of chronic proliferative rhinitis (CPR) in sheep. In Switzerland, CPR in sheep due to S. IIIb 61:k:1,5,(7) was first described in 2017 in a flock of Texel sheep. Therefore, we assessed the prevalence of S. IIIb 61:k:1,5,(7) within the Swiss sheep population using a representative sampling strategy. From May 2017 to June 2018 a total of 681 nasal swabs from individual clinically healthy sheep of 141 different flocks throughout Switzerland were taken. Swabs were analysed by selective enrichment for the presence of S. IIIb 61:k:1,5,(7). Additionally, antimicrobial resistance of the isolates was determined by broth microdilution. A total of 146 out of 681 nasal swabs tested positive for S. IIIb 61:k:1,5,(7), which corresponds to a prevalence on animal level of 21% (95%CI 18%-25%). In 73 out of 141 flocks tested, at least one sheep tested positive for S. IIIb 61:k:1.5,(7), resulting in a minimal prevalence on flock level of 52% (95%CI 43%-60%). Positive flocks were found in all cantons except the canton of Jura. Adults were significantly more affected than sheep under one year/lambs and positive sheep were found in several breeds. No microbiologically resistant isolates were detected, except for one isolate showing resistance against ampicillin. Because of its widespread occurrence in the Swiss sheep population, further research should focus on the pathogenic impact of S. IIIb 61:k:1,5,(7) on the health status of sheep.

Perceived importance of pandemic interventions for attending cultural events - findings from Germany.

BACKGROUND: During the first waves of the COVID-19 pandemic, many cultural and sporting events were held without spectators or had to be cancelled. Therefore, several containment strategies to provide requirements for safe events were developed and tested. Nonetheless, every second (50.7%) is afraid of becoming infected on an event. We therefore investigated which hygiene and containment measures are perceived to be important from the visitor's point of view and thus might increase subjective sense of safety. METHODS: This online study was carried out in November 2020. A total of 1,004 persons, who regularly attended events before the pandemic, took part in the study. The importance of different hygiene and containment measures was evaluated using a 5-point Likert-scale (1 "unimportant" to 5 "extremely important"). Potential statistical differences in socio-demographical aspects (age, gender, net disposable income for leisure activities) and attendance on events were tested with analyses of variance. RESULTS: Participants perceived the use of disinfectant (M = 4.10) as the most important element of containment strategies, followed by transparent information on the hygiene strategy (M = 4.00), reduced occupancy (M = 3.98), and optimized ventilation (M = 3.97). Body temperature measurement at the entrance (M = 3.27), a negative SARS-CoV-2 test (M = 3.11), completion of a health questionnaire (M = 3.05), and abandoning breaks and catering (M = 2.98) were considered as less important. Analyses of group differences in socio-demographical aspects found abandoning breaks and catering to be more important to men than to women. This strategy is also more important to people aged 66 and above than to younger age groups (e.g., age 20-40). For women, the use of disinfectant is considerably more important. No other significant differences exist. CONCLUSION: Combining relevant measures appears to be important to provide a safe containment strategy. Measures aimed at positively influencing people's sense of safety do not fully correspond to researched knowledge of effectiveness. There are also target group-specific differences in the rating of measures, which should be considered while preparing containment strategies. To describe the dynamic development of changes in subjective rating of containment strategies, continuing research is needed.

Influenza A virus NS1 optimises virus infectivity by enhancing genome packaging in a dsRNA-binding dependent manner.

BACKGROUND: The non-structural protein 1 (NS1) of influenza A virus (IAV) is a key player in inhibiting antiviral response in host cells, thereby facilitating its replication. However, other roles of NS1, which are independent of antagonising host cells' antiviral response, are less characterised. METHODS: To investigate these unidentified roles, we used a recombinant virus, which lacks NS1 expression, and observed its phenotypes during the infection of antiviral defective cells (RIG-I KO cells) in the presence or absence of exogeneous NS1. Moreover, we used virus-like particle (VLP) production system to further support our findings. RESULTS: Our experiments demonstrated that IAV deficient in NS1 replicates less efficiently than wild-type IAV in RIG-I KO cells and this replication defect was complemented by ectopic expression of NS1. As suggested previously, NS1 is incorporated in the virion and participates in the regulation of viral transcription and translation. Using the VLP production system, in which minigenome transcription or viral protein production was unaffected by NS1, we demonstrated that NS1 facilitates viral genome packaging into VLP, leading to efficient minigenome transfer by VLP. Furthermore, the incorporation of NS1 and the minigenome into VLP were impaired by introducing a point mutation (R38A) in the double stranded RNA-binding domain of NS1. CONCLUSION: These results suggest a novel function of NS1 in improving genome packaging in a dsRNA binding-dependent manner. Taken together, NS1 acts as an essential pro-viral regulator, not only by antagonizing host immunity but also by facilitating viral replication and genome packaging.

Attitudes Towards Adapted Lifestyle-Integrated Functional Exercise Developed for 60-70-Year-Olds: Perceptions of Participants and Trainers.

BACKGROUND: Time commitments, limited access, or unwillingness to join a group are some of the many reasons for low adherence to structured exercise in older adults. A promising alternative approach is integrating exercise into daily routines. OBJECTIVE: This study tested whether an adapted Lifestyle-integrated Functional Exercise (aLiFE) programme is suitable for adults aged 60-70 years. METHODS: The aLiFE approach was evaluated by interviews and focus-groups with participants and trainers following 4-week pre-post intervention pilot study. For data analyses, Framework Approach was used. Coding was managed using NVivo, and subsequently organised into overarching themes. RESULTS: Twenty women and 11 men (mean age 66.4 ± 2.7 years) and 6 trainers (30.0 ± 6.2 years; 5 women) participated. Both participants and trainers were positive about the programme. Participants understood the concept of integrating balance, strength and physical activities into daily lives and valued the individual tailoring in the programme, the preventive approach, and the support of trainers. Trainers valued the flexible approach and peer support between trainers. However, both participants and trainers disliked the extensive study paperwork and reported some challenges to integrate activities into daily routines during the compressed intervention: busy and varied lifestyles, embarrassment performing activities in public, pain, difficulty of specific activities. Participants noted habitualisation of some activities within the short intervention period, even without continuous self-monitoring. CONCLUSIONS: aLiFE is a highly acceptable intervention amongst adults aged 60-70 years. Trainers are especially relevant as motivators and support providers. The effectiveness of the aLiFE approach should be tested in a randomised controlled trial.

The Adapted Lifestyle-Integrated Functional Exercise Program for Preventing Functional Decline in Young Seniors: Development and Initial Evaluation.

BACKGROUND: The Lifestyle-integrated Functional Exercise (LiFE) program is an intervention integrating balance and strength activities into daily life, effective at reducing falls in at-risk people ≥70 years. There is potential for LiFE to be adapted to young seniors in order to prevent age-related functional decline. OBJECTIVE: We aimed to (1) develop an intervention by adapting Lifestyle-integrated Functional Exercise (aLiFE) to be more challenging and suitable for preventing functional decline in young seniors in their 60s and (2) perform an initial feasibility evaluation of the program. Pre-post changes in balance, mobility, and physical activity (PA) were also explored. METHODS: Based on a conceptual framework, a multidisciplinary expert group developed an initial aLiFE version, including activities for improving strength, neuromotor performances, and PA. Proof-of-concept was evaluated in a 4-week pre-post intervention study measuring (1) feasibility including adherence, frequency of practice, adverse events, acceptability (i.e., perceived helpfulness, adaptability, level of difficulty of single activities), and safety, and (2) changes in balance/mobility (Community Balance and Mobility Scale) and PA (1 week activity monitoring). The program was refined based on the study results. RESULTS: To test the initial aLiFE version, 31 young seniors were enrolled and 30 completed the study (mean age 66.4 ± 2.7 years, 60% women). Of a maximum possible 16 activities, participants implemented on average 12.1 ± 1.8 activities during the intervention, corresponding to mean adherence of 76%. Implemented activities were practiced 3.6-6.1 days/week and 1.8-7.8 times/day, depending on the activity type. One noninjurious fall occurred during practice, although the participant continued the intervention. The majority found the activities helpful, adaptable to individual lifestyle, appropriately difficult, and safe. CMBS score increased with medium effect size (d = 0.72, p = 0.001). Increase in daily walking time (d = 0.36) and decrease in sedentary time (d = -0.10) were nonsignificant. Refinements included further increasing the task challenge of some strength activities and defining the most preferred activities in the trainer's manual to facilitate uptake of the program. CONCLUSION: aLiFE has the potential to engage young seniors in regular lifestyle-integrated activities. Effectiveness needs to be evaluated in a randomized controlled trial.

Performance-based clinical tests of balance and muscle strength used in young seniors: a systematic literature review.

BACKGROUND: Many balance and strength tests exist that have been designed for older seniors, often aged ≥70 years. To guide strategies for preventing functional decline, valid and reliable tests are needed to detect early signs of functional decline in young seniors. Currently, little is known about which tests are being used in young seniors and their methodological quality. This two-step review aims to 1) identify commonly used tests of balance and strength, and 2) evaluate their measurement properties in young seniors. METHODS: First, a systematic literature search was conducted in MEDLINE to identify primary studies that employed performance-based tests of balance and muscle strength, and which aspects of balance and strength these tests assess in young seniors aged 60-70. Subsequently, for tests used in ≥3 studies, a second search was performed to identify method studies evaluating their measurement properties. The quality of included method studies was evaluated using the Consensus-based Standards for selection of health Measurement Instruments (COSMIN) checklist. RESULTS: Of 3454 articles identified, 295 met the inclusion criteria. For the first objective, 69 balance and 51 muscle strength tests were identified, with variations in administration mode and outcome reporting. Twenty-six balance tests and 15 muscle strength tests were used in ≥3 studies, with proactive balance tests and functional muscle power tests used most often. For the second objective, the search revealed 1880 method studies, of which nine studies (using 5 balance tests and 1 strength test) were included for quality assessment. The Timed Up and Go test was evaluated the most (4 studies), while the Community Balance and Mobility (CBM) scale was the second most assessed test (3 studies). For strength, one study assessed the reliability of the Five times sit-to-stand. CONCLUSION: Commonly used balance and muscle strength tests in young seniors vary greatly with regards to administration mode and outcome reporting. Few studies have evaluated measurement properties of these tests when used in young seniors. There is a need for standardisation of existing tests to improve their informative value and comparability. For measuring balance, the CBM is a new and promising tool to detect even small balance deficits in balance in young seniors.

German version of the Community Balance and Mobility Scale : Translation and evaluation of measurement properties.

BACKGROUND: Tools to detect subtle balance deficits in high-functioning community-dwelling older adults are lacking. The Community Balance and Mobility Scale (CBM) is a valuable tool to measure balance deficits in this group; however, it is not yet available in the German language. OBJECTIVE: The aim was 1) to translate and cross-culturally adapt the CBM into the German language and 2) to investigate the measurement properties of the German CBM (G-CBM). MATERIAL AND METHODS: The original CBM was translated into the German language according to established guidelines. A total of 51 older adults (mean age 69.9 ± 7.1 years) were recruited to measure construct validity by comparing the G­CBM against standardized balance and/or mobility assessments including the Fullerton Advanced Balance Scale (FAB), Berg Balance Scale (BBS), 3 m Tandem Walk (3MTW), 8 Level Balance Scale (8LBS), 30 s Chair Stand Test (30CST), Timed Up and Go (TUG) test, gait speed, and the Falls Efficacy Scale International (FES-I). Intrarater and interrater reliability and internal consistency reliability were estimated using intraclass correlations (ICC) and Cronbach's alpha, respectively. Ceiling effects were calculated as the percentage of the sample scoring the maximum score. RESULTS: The G­CBM correlated excellently with FAB and BBS (ρ = 0.78-0.85; P < 0.001), good with 3MTW, TUG, and FES-I (ρ = -0.55 to -0.61; P < 0.001), and moderately with 8LBS, 30CST, and habitual gait speed (ρ = 0.32-0.46; P < 0.001). Intrarater (ICC3,k = 0.998; P < 0.001) and interrater (ICC2,k = 0.996; P < 0.001) reliability, and internal consistency reliability (α = 0.998) were also high. The G­CBM did not show ceiling effects. CONCLUSION: The G­CBM is a valid and reliable tool for measuring subtle balance deficits in older high-functioning adults. The absence of ceiling effects emphasizes the use of this scale in this cohort. The G­CBM can now be utilized in clinical practice.

Concurrent validity and reliability of the Community Balance and Mobility scale in young-older adults.

BACKGROUND: With the growing number of young-older adults (baby-boomers), there is an increasing demand for assessment tools specific for this population, which are able to detect subtle balance and mobility deficits. Various balance and mobility tests already exist, but suffer from ceiling effects in higher functioning older adults. A reliable and valid challenging balance and mobility test is critical to determine a young-older adult's balance and mobility performance and to timely initiate preventive interventions. The aim was to evaluate the concurrent validity, inter- and intrarater reliability, internal consistency, and ceiling effects of a challenging balance and mobility scale, the Community Balance and Mobility Scale (CBM), in young-older adults aged 60 to 70 years. METHODS: Fifty-one participants aged 66.4 ± 2.7 years (range, 60-70 years) were assessed with the CBM. The Fullerton Advanced Balance scale (FAB), 3-Meter Tandem Walk (3MTW), 8-level balance scale, Timed-Up-and-Go (TUG), and 7-m habitual gait speed were used to estimate concurrent validity, examined by Spearman correlation coefficient (ρ). Inter- and intrarater reliability were calculated as Intra-class-correlations (ICC), and internal consistency by Cronbach alpha and item-total correlations (ρ). Ceiling effects were determined by obtaining the percentage of participants reaching the highest possible score. RESULTS: The CBM significantly correlated with the FAB (ρ = 0.75; p < .001), 3MTW errors (ρ = - 0.61; p < .001), 3MTW time (ρ = - 0.35; p = .05), the 8-level balance scale (ρ = 0.35; p < .05), the TUG (ρ = - 0.42; p < .01), and 7-m habitual gait speed (ρ = 0.46, p < .001). Inter- (ICC2,k = 0.97), intrarater reliability (ICC3,k = 1.00) were excellent, and internal consistency (α = 0.88; ρ = 0.28-0.81) was good to satisfactory. The CBM did not show ceiling effects in contrast to other scales. CONCLUSIONS: Concurrent validity of the CBM was good when compared to the FAB and moderate to good when compared to other measures of balance and mobility. Based on this study, the CBM can be recommended to measure balance and mobility performance in the specific population of young-older adults. TRIAL REGISTRATION: Trial number: ISRCTN37750605 . (Registered on 21/04/2016).

Feasibility and Effectiveness of Intervention Programmes Integrating Functional Exercise into Daily Life of Older Adults: A Systematic Review.

BACKGROUND: Traditionally, exercise programmes for improving functional performance and reducing falls are organised as structured sessions. An alternative approach of integrating functional exercises into everyday tasks has emerged in recent years. OBJECTIVES: Summarising the current evidence for the feasibility and effectiveness of interventions integrating functional exercise into daily life. METHODS: A systematic literature search was conducted including articles based on the following criteria: (1) individuals ≥60 years; (2) intervention studies of randomised controlled trials (RCTs) and non-randomised studies (NRS); (3) using a lifestyle-integrated approach; (4) using functional exercises to improve strength, balance, or physical functioning; and (5) reporting outcomes on feasibility and/or effectiveness. Methodological quality of RCTs was evaluated using the PEDro scale. RESULTS: Of 4,415 articles identified from 6 databases, 14 (6 RCTs) met the inclusion criteria. RCT quality was moderate to good. Intervention concepts included (1) the Lifestyle-integrated Functional Exercise (LiFE) programme integrating exercises into everyday activities and (2) combined programmes using integrated and structured training. Three RCTs evaluated LiFE in community dwellers and reported significantly improved balance, strength, and functional performance compared with controls receiving either no intervention, or low-intensity exercise, or structured exercise. Two of these RCTs reported a significant reduction in fall rate compared with controls receiving either no intervention or low-intensity exercise. Three RCTs compared combined programmes with usual care in institutionalised settings and reported improvements for some (balance, functional performance), but not all (strength, falls) outcomes. NRS showed behavioural change related to LiFE and feasibility in more impaired populations. One NRS comparing a combined home-based programme to a gym-based programme reported greater sustainability of effects in the combined programme. CONCLUSIONS: This review provides evidence for the effectiveness of integrated training for improving motor performances in older adults. Single studies suggest advantages of integrated compared with structured training. Combined programmes are positively evaluated in institutionalised settings, while little evidence exists in other populations. In summary, the approach of integrating functional exercise into daily life represents a promising alternative or complement to structured exercise programmes. However, more RCTs are needed to evaluate this concept in different target populations and the potential for inducing behavioural change.

Influenza virus adaptation PB2-627K modulates nucleocapsid inhibition by the pathogen sensor RIG-I.

The cytoplasmic RNA helicase RIG-I mediates innate sensing of RNA viruses. The genomes of influenza A virus (FLUAV) are encapsidated by the nucleoprotein and associated with RNA polymerase, posing potential barriers to RIG-I sensing. We show that RIG-I recognizes the 5'-triphosphorylated dsRNA on FLUAV nucleocapsids but that polymorphisms at position 627 of the viral polymerase subunit PB2 modulate RIG-I sensing. Compared to mammalian-adapted PB2-627K, avian FLUAV nucleocapsids possessing PB2-627E are prone to increased RIG-I recognition, and RIG-I-deficiency partially restores PB2-627E virus infection of mammalian cells. Heightened RIG-I sensing of PB2-627E nucleocapsids correlates with previously established lower affinity of 627E-containing PB2 for nucleoprotein and is increased by further nucleocapsid instability. The effect of RIG-I on PB2-627E nucleocapsids is independent of antiviral signaling, suggesting that RIG-I-nucleocapsid binding alone can inhibit infection. These results indicate that RIG-I is a direct avian FLUAV restriction factor and highlight nucleocapsid disruption as an antiviral strategy.

Specific behavioral comorbidity in a large sample of children with functional incontinence: Report of 1,001 cases.

AIMS: Psychological comorbidity among children with functional incontinence is high: 20-30% of children with nocturnal enuresis (NE), 20-40% of those with daytime urinary incontinence (DUI) and 30-50% of those with fecal incontinence (FI) have clinically relevant comorbid disorders. The aim of this study was to analyze specific comorbid behavioral symptoms for different subtypes of incontinence in a large group of children. METHODS: All 1,001 consecutive children and adolescents (67.5% boys) with a mean age of 8.5 years presented at a tertiary outpatient department between 2004 and 2011 were examined with a full pediatric and child psychiatric assessment. Prevalence of different subforms of incontinence and associated behavioral symptoms were analyzed. The internalizing, externalizing, and total problem scores of the Child Behavior Checklist (CBCL) were evaluated. RESULTS: Of all children, 70.1% (702 children) had NE, 36.1% (361 children) had DUI, and 36.8% (368 children) were affected by FI. More than 43% of all children had clinically relevant psychological symptoms (CBCL total score, cut-off at 90th percentile). Children with non-retentive FI had highest rates of clinically relevant psychological symptoms (58.8%). Children with combined subtypes of incontinence (any combination of NE, DUI, and FI) were more affected by psychological comorbidities than children with isolated subtypes (NE or DUI or FI). CONCLUSIONS: Children with incontinence have high rates of comorbid behavioral symptoms-three to six times higher than norms. Especially children with FI and combined subtypes of incontinence were affected. As behavioral symptoms and disorders will interfere with incontinence treatment, a general screening is recommended.

Monitoring activation of the antiviral pattern recognition receptors RIG-I and PKR by limited protease digestion and native PAGE.

Host defenses to virus infection are dependent on a rapid detection by pattern recognition receptors (PRRs) of the innate immune system. In the cytoplasm, the PRRs RIG-I and PKR bind to specific viral RNA ligands. This first mediates conformational switching and oligomerization, and then enables activation of an antiviral interferon response. While methods to measure antiviral host gene expression are well established, methods to directly monitor the activation states of RIG-I and PKR are only partially and less well established. Here, we describe two methods to monitor RIG-I and PKR stimulation upon infection with an established interferon inducer, the Rift Valley fever virus mutant clone 13 (Cl 13). Limited trypsin digestion allows to analyze alterations in protease sensitivity, indicating conformational changes of the PRRs. Trypsin digestion of lysates from mock infected cells results in a rapid degradation of RIG-I and PKR, whereas Cl 13 infection leads to the emergence of a protease-resistant RIG-I fragment. Also PKR shows a virus-induced partial resistance to trypsin digestion, which coincides with its hallmark phosphorylation at Thr 446. The formation of RIG-I and PKR oligomers was validated by native polyacrylamide gel electrophoresis (PAGE). Upon infection, there is a strong accumulation of RIG-I and PKR oligomeric complexes, whereas these proteins remained as monomers in mock infected samples. Limited protease digestion and native PAGE, both coupled to western blot analysis, allow a sensitive and direct measurement of two diverse steps of RIG-I and PKR activation. These techniques are relatively easy and quick to perform and do not require expensive equipment.

Segmented negative-strand RNA viruses and RIG-I: divide (your genome) and rule.

The group of negative-stranded RNA viruses (NSVs) with a segmented genome comprises pathogens like influenza virus (eight segments), Rift Valley fever virus and Hantavirus (three segments), or Lassa virus (two segments). Partitioning the genome allows rapid evolution of new strains by reassortment. Each segment carries a short double-stranded (ds) 'panhandle' structure which serves as promoter. Similar dsRNA structures, however, represent the optimal ligand for RIG-I, a cytoplasmic pathogen sensor of the antiviral interferon response. Thus, segmenting a virus genome can entail an increased RIG-I sensitivity. Here, we outline the astonishingly diverse and efficient strategies by which segmented NSVs are compensating for the elevated number of RIG-I ligands in their genome.

RIG-I-like receptors and negative-strand RNA viruses: RLRly bird catches some worms.

Negative strand RNA viruses with a nonsegmented genome (ns-NSVs) or a segmented genome (s-NSVs) are an important source of human and animal diseases. Survival of the host from those infections is critically dependent on rapidly reacting innate immune responses. Two cytoplasmic RNA helicases, RIG-I and MDA5 (collectively termed RIG-I-like receptors, RLRs), are essential for recognizing virus-specific RNA structures to initiate a signalling cascade, resulting in the production of the antiviral type I interferons. Here, we will review the current knowledge and views on RLR agonists, RLR signalling, and the wide variety of countermeasures ns-NSVs and s-NSVs have evolved. Specific aspects include the consequences of genome segmentation for RLR activation and a discussion on the physiological ligands of RLRs.

IFIT2 is an effector protein of type I IFN-mediated amplification of lipopolysaccharide (LPS)-induced TNF-α secretion and LPS-induced endotoxin shock.

Type I IFN signaling amplifies the secretion of LPS-induced proinflammatory cytokines such as TNF-α or IL-6 and might thus contribute to the high mortality associated with Gram-negative septic shock in humans. The underlying molecular mechanism, however, is ill defined. In this study, we report the generation of mice deficient in IFN-induced protein with tetratricopeptide repeats 2 (Ifit2) and demonstrate that Ifit2 is a critical signaling intermediate for LPS-induced septic shock. Ifit2 expression was significantly upregulated in response to LPS challenge in an IFN-α receptor- and IFN regulatory factor (Irf)9-dependent manner. Also, LPS induced secretion of IL-6 and TNF-α by bone marrow-derived macrophages (BMDMs) was significantly enhanced in the presence of Ifit2. In accordance, Ifit2-deficient mice exhibited significantly reduced serum levels of IL-6 and TNF-α and reduced mortality in an endotoxin shock model. Investigation of the underlying signal transduction events revealed that Ifit2 upregulates Irf3 phosphorylation. In the absence of Irf3, reduced Ifn-ß mRNA expression and Ifit2 protein expression after LPS stimulation was found. Also, Tnf-α and Il-6 secretion but not Tnf-α and Il-6 mRNA expression levels were reduced. Thus, IFN-stimulated Ifit2 via enhanced Irf3 phosphorylation upregulates the secretion of proinflammatory cytokines. It thereby amplifies LPS-induced cytokine production and critically influences the outcome of endotoxin shock.

Incoming RNA virus nucleocapsids containing a 5'-triphosphorylated genome activate RIG-I and antiviral signaling.

Host defense to RNA viruses depends on rapid intracellular recognition of viral RNA by two cytoplasmic RNA helicases: RIG-I and MDA5. RNA transfection experiments indicate that RIG-I responds to naked double-stranded RNAs (dsRNAs) with a triphosphorylated 5' (5'ppp) terminus. However, the identity of the RIG-I stimulating viral structures in an authentic infection context remains unresolved. We show that incoming viral nucleocapsids containing a 5'ppp dsRNA "panhandle" structure trigger antiviral signaling that commences with RIG-I, is mediated through the adaptor protein MAVS, and terminates with transcription factor IRF-3. Independent of mammalian cofactors or viral polymerase activity, RIG-I bound to viral nucleocapsids, underwent a conformational switch, and homo-oligomerized. Enzymatic probing and superresolution microscopy suggest that RIG-I interacts with the panhandle structure of the viral nucleocapsids. These results define cytoplasmic entry of nucleocapsids as the proximal RIG-I-sensitive step during infection and establish viral nucleocapsids with a 5'ppp dsRNA panhandle as a RIG-I activator.

Human cell tropism and innate immune system interactions of human respiratory coronavirus EMC compared to those of severe acute respiratory syndrome coronavirus.

Infections with human coronavirus EMC (HCoV-EMC) are associated with severe pneumonia. We demonstrate that HCoV-EMC resembles severe acute respiratory syndrome coronavirus (SARS-CoV) in productively infecting primary and continuous cells of the human airways and in preventing the induction of interferon regulatory factor 3 (IRF-3)-mediated antiviral alpha/beta interferon (IFN-α/ß) responses. However, HCoV-EMC was markedly more sensitive to the antiviral state established by ectopic IFN. Thus, HCoV-EMC can utilize a broad range of human cell substrates and suppress IFN induction, but it does not reach the IFN resistance of SARS-CoV.

Inclusion bodies are a site of ebolavirus replication.

Inclusion bodies are a characteristic feature of ebolavirus infections in cells. They contain large numbers of preformed nucleocapsids, but their biological significance has been debated, and they have been suggested to be aggregates of viral proteins without any further biological function. However, recent data for other viruses that produce similar structures have suggested that inclusion bodies might be involved in genome replication and transcription. In order to study filovirus inclusion bodies, we fused mCherry to the ebolavirus polymerase L, which is found in inclusion bodies. The resulting L-mCherry fusion protein was functional in minigenome assays and incorporated into virus-like particles. Importantly, L-mCherry fluorescence in transfected cells was readily detectable and distributed in a punctate pattern characteristic for inclusion bodies. A recombinant ebolavirus encoding L-mCherry instead of L was rescued and showed virtually identical growth kinetics and endpoint titers to those for wild-type virus. Using this virus, we showed that the onset of inclusion body formation corresponds to the onset of viral genome replication, but that viral transcription occurs prior to inclusion body formation. Live-cell imaging further showed that inclusion bodies are highly dynamic structures and that they can undergo dramatic reorganization during cell division. Finally, by labeling nascent RNAs using click technology we showed that inclusion bodies are indeed the site of viral RNA synthesis. Based on these data we conclude that, rather than being inert aggregates of nucleocapsids, ebolavirus inclusion bodies are in fact complex and dynamic structures and an important site at which viral RNA replication takes place.

Tacaribe virus but not junin virus infection induces cytokine release from primary human monocytes and macrophages.

The mechanisms underlying the development of disease during arenavirus infection are poorly understood. However, common to all hemorrhagic fever diseases is the involvement of macrophages as primary target cells, suggesting that the immune response in these cells may be of paramount importance during infection. Thus, in order to identify features of the immune response that contribute to arenavirus pathogenesis, we have examined the growth kinetics and cytokine profiles of two closely related New World arenaviruses, the apathogenic Tacaribe virus (TCRV) and the hemorrhagic fever-causing Junin virus (JUNV), in primary human monocytes and macrophages. Both viruses grew robustly in VeroE6 cells; however, TCRV titres were decreased by approximately 10 fold compared to JUNV in both monocytes and macrophages. Infection of both monocytes and macrophages with TCRV also resulted in the release of high levels of IL-6, IL-10 and TNF-α, while levels of IFN-α, IFN-ß and IL-12 were not affected. However, we could show that the presence of these cytokines had no direct effect on growth of either TCRV of JUNV in macrophages. Further analysis also showed that while the production of IL-6 and IL-10 are dependent on viral replication, production of TNF-α also occurs after exposure to UV-inactivated TCRV particles and is thus independent of productive virus infection. Surprisingly, JUNV infection did not have an effect on any of the cytokines examined indicating that, in contrast to other viral hemorrhagic fever viruses, macrophage-derived cytokine production is unlikely to play an active role in contributing to the cytokine dysregulation observed in JUNV infected patients. Rather, these results suggest that an early, controlled immune response by infected macrophages may be critical for the successful control of infection of apathogenic viruses and prevention of subsequent disease, including systemic cytokine dysregulation.

The effects of therapeutic climbing in patients with chronic low back pain: a randomized controlled study.

STUDY DESIGN: A randomized controlled study investigated the effects of therapeutic climbing in patients with chronic low back pain. Before and after 4 weeks of training, physical and mental well-being were measured by two questionnaires (36-Item Short Form Health Survey [SF-36]; Hannover Functional Ability Questionnaire for measuring back pain-related disability [FFbH-R]). OBJECTIVE: Therapeutic climbing has been suggested to increase muscular strength and perceived physical and mental well-being. This study focused on the psychological effects of therapeutic climbing and compared it with standard exercise therapy. SUMMARY OF BACKGROUND DATA: Therapeutic climbing has become increasingly popular in rehabilitation and its effects on muscular strengthening have been shown. Therapeutic climbing has also been suggested to yield psychological effects such as changes in attentional focus from pain to physical capabilities. To date, no controlled clinical trial has investigated these psychological effects and it is unclear whether therapeutic climbing is comparable or superior to other forms of exercise. METHODS: Twenty-eight patients with chronic low back pain conducted either a therapeutic climbing or a standard exercise regime. Each program took 4 weeks, including four guided training sessions per week. Before and after the program, patients answered two questionnaires assessing their physical and mental well-being. RESULTS: For the Hannover Functional Ability Questionnaire for measuring back pain-related disability, there was no difference before versus after or between the treatments. For the SF-36, both treatments showed significant improvements in 3/8 subscales of the SF-36. In 2/8 subscales, only the participants of the therapeutic climbing improved and in 1/8 subscales the converse was true. Comparing both groups, significantly larger improvements were found after therapeutic climbing in two subscales of the SF-36: physical functioning and general health perception. CONCLUSION: The benefits of therapeutic climbing were comparable with those of a standard exercise regime. In two subscales of the SF-36, the benefits of therapeutic climbing exceeded those of standard exercise therapy, primarily in perceived health and physical functioning of the patients. This finding demonstrates that therapeutic climbing is equivalent and partly superior to standard exercise therapy for patients with chronic low back pain.