Detection of Autoantibodies Against the Acetylcholine Receptor, Evaluation of Commercially Available Methodologies: Fixed Cell-Based Assay, Radioimmunoprecipitation Assay and Enzyme-Linked Immunosorbent Assay1.

Background/Objective: Myasthenia Gravis (MG) is an autoimmune disorder characterized by pathogenic autoantibodies (AAbs) targeting nicotinic acetylcholine receptors (AChR), disrupting neuromuscular communication. RadioImmunoPrecipitation Assay (RIPA) is recommended to detect AChR AAbs, but its complexity and radioactive requirements limit widespread use. We compare non-RIPA anti-AChR immunoassays, including Cell-Based Assay (CBA) and two ELISA kits, against the gold standard RIPA. Methods/Results: 145 samples were included with medical indication for anti-AChR testing. By the RIPA method, 63 were negative (RIPA-Neg < 0.02 nmol/L), 18 were classified as Borderline (≥0.02 -1 nmol/L), and 64 were positive (RIPA-Pos > 1 nmol/L). The competitive ELISA showed poor agreement with RIPA (Kappa = 0.216). The indirect ELISA demonstrated substantial agreement with RIPA (Kappa = 0.652), with ∼76% sensitivity and ∼94% specificity for MG diagnostic. The CBA, where fixed cells expressing clustered AChR were used as substrate, exhibited almost perfect agreement with RIPA (Kappa = 0.984), yielding ∼98% sensitivity and 96% specificity for MG. In addition, a semiquantitative analysis showed a strong correlation between CBA titration, indirect ELISA, and RIPA levels (r = 0.793 and r = 0.789, respectively). Conclusions: The CBA displayed excellent analytical performance for MG diagnostic when compared to RIPA, making it a potential replacement for RIPA in clinical laboratories. Some solid-phase assays (such as the indirect ELISA applied here), as well as CBA titration, offer reliable options to estimate anti-AChR AAb levels after confirming positivity by the CBA.∥.

Birth weight and its relationship with endothelial function and pattern of endothelium-derived microparticles during childhood: New insight about early vascular damage.

AIMS: To investigate whether a specific endothelium-derived microparticles (EMPs) phenotype could be associated with birth weight and microvascular endothelial function in children. MATERIALS AND METHODS: A total of 95 children aged 6-14 years were recruited. Anthropometric measurements, blood pressure measurement, microvascular endothelial function testing, and biochemical profile analyses were performed. Standardized flow cytometry methods were used to identify and quantify the circulating CD144+, CD31+/annexin V+, and CD62E+ EMPs. KEY FINDINGS: The circulating number of CD31+/annexin V+ EMPs and CD144+ EMP levels were correlated with birth weight, systolic blood pressure, microvascular endothelial function, total cholesterol, and low-density lipoprotein cholesterol (LDL-C) level. In the multivariable logistic regression models, we identified strong evidence of a higher risk of microvascular endothelial dysfunction among children with low birth weight (LBW) and increased levels of both CD31+/annexin V+ EMPs and LDL-C; LBW and elevated LDL-C levels were independent predictors of high circulating numbers of CD31+/annexin V+ and CD144+ > 75th percentile EMPs. SIGNIFICANCE: Our data provide evidence that children with LBW values showed greater numbers of circulating CD31+/annexin V+ and CD144+ EMPs. In addition, LBW and high levels of CD31+/annexin V+ and LDL-C were significant risk factors for the presence of microvascular endothelial dysfunction.

Anopheles albimanus (Diptera: Culicidae) Ensemble Distribution Modeling: Applications for Malaria Elimination.

In the absence of entomological information, tools for predicting Anopheles spp. presence can help evaluate the entomological risk of malaria transmission. Here, we illustrate how species distribution models (SDM) could quantify potential dominant vector species presence in malaria elimination settings. We fitted a 250 m resolution ensemble SDM for Anopheles albimanus Wiedemann. The ensemble SDM included predictions based on seven different algorithms, 110 occurrence records and 70 model projections. SDM covariates included nine environmental variables that were selected based on their importance from an original set of 28 layers that included remotely and spatially interpolated locally measured variables for the land surface of Costa Rica. Goodness of fit for the ensemble SDM was very high, with a minimum AUC of 0.79. We used the resulting ensemble SDM to evaluate differences in habitat suitability (HS) between commercial plantations and surrounding landscapes, finding a higher HS in pineapple and oil palm plantations, suggestive of An. albimanus presence, than in surrounding landscapes. The ensemble SDM suggested a low HS for An. albimanus at the presumed epicenter of malaria transmission during 2018-2019 in Costa Rica, yet this vector was likely present at the two main towns also affected by the epidemic. Our results illustrate how ensemble SDMs in malaria elimination settings can provide information that could help to improve vector surveillance and control.

High circulating levels of CD62E+ and CD31+/Annexin V+ endothelium-derived microparticles in children with overweight/obesity: Evidence of early vascular damage.

BACKGROUND: Obesity perturbs endothelium integrity, leading to endothelial activation, which predisposes the release of endothelium-derived microparticles (EMP). We measured the CD31+/annexin V+ and CD62E+ EMP levels to improve our understanding of their contribution to endothelial damage in children with overweight/obesity. SUBJECT AND METHODS: In this cross-sectional study, 107 children with normal weight and 35 children with overweight/obesity were evaluated. Anthropometric measurement, blood pressure, biochemical profile was performed. Standardized flow cytometry methods were used to identify and quantify circulating CD31+/annexin V+ and CD62E+ EMP. RESULTS: Children with overweight/obesity had significantly higher circulating levels of CD31+/annexin V+ (750 [600]) and CD62E+ (1400 [700]) EMP than those with normal weight (P < 0.001 for both). We found that EMP levels were positively correlated with body mass index (BMI), waist circumference, blood pressure, total cholesterol, low-density lipoprotein cholesterol (LDLc), and triglycerides. The multivariable logistic regression model revealed that the risks of having high EMP levels (> 75th percentile) were high in children with both large waist circumference and elevated LDLc level. Receiver operating characteristic (ROC) curves demonstrated that the LDLc levels showed significantly greater discrimination than waist circumference for both CD31+/annexin V+ (P = 0.031) as CD62E+ EMPs (P = 0.041). CONCLUSIONS: Children with overweight/obesity have high circulating CD31+/annexin V+ and CD62E+ EMP levels, which may be an early sign of endothelial apoptosis and inflammatory activation in response to injury. These EMP levels were positively associated with several cardiometabolic risk factors. Our data underscore the negative influence of high-risk metabolic profiles on endothelial integrity in the early stages of childhood obesity.

Calidad de la atención de servicios de internación psiquiátrica en hospitales generales de Argentina, 2018 / Quality of psychiatric hospitalization services in general hospitals in Argentina, 2018

RESUMEN Objetivo: evaluar la calidad de atención en servicios de internación psiquiátrica de hospitales generales desde los enfoques de derechos y comunitario. Métodos: estudio multicéntrico, observacional descriptivo. Se seleccionaron cuatro servicios de internación psiquiátrica de hospitales generales ubicados en centros urbanos de distinta población en Argentina. En cada uno de ellos se analizaron los registros estadísticos, se realizaron observaciones no participantes y se entrevistó a gestores, trabajadores y usuarios. La guía de observación y entrevista se elaboró a partir de la revisión de instrumentos de la OMS y del proyecto Idea. Los resultados se analizaron a partir de las categorías de enfoque de derechos y orientación comunitaria de los servicios. Resultados: respecto del enfoque de derechos se observa que el que requiere mayor desarrollo es el de capacidad jurídica. En relación a la orientación comunitaria, se observa un mayor desarrollo de esta perspectiva a través de la implementación de diversas estrategias y el trabajo con las familias como la más común. Conclusiones: en términos de procesos y resultados de la atención se observan algunas diferencias entre los servicios que cuentan con sala especializada y los que no, y que estos últimos requieren de otros estudios para poder ser analizados.

ABSTRACT Objective: To evaluate, based on the human rights and community care frameworks, the quality of hospitalization psychiatric services in general hospitals. Methods: Multisite, descriptive observational study. There were analyzed four psychiatric hospitalization services, located in general hospitals of urban districts with different sizes in Argentina. Data collection included analysis of statistical records, non-participant observations of services, and interviews with service managers, service staff and users. The observation and interview guides were based on two international tools: Idea project interview guide, and QRTK of the WHO. Results were analyzed using two main categories: human rights-based care approach and community orientation of care delivery. Results: Regarding human rights-based care, legal capacity is the right that requires more improvement in services. About community orientation of care, different strategies were observed, with the work with the families as the most common. Conclusions: One aspect that requires further research is the specific type of psychiatric hospitalization service in the general hospital, given the fact that some differences in process and results were observed between general and specialized wards.

Interkit Reproducibility of the Indirect Immunofluorescence Assay on HEp-2 Cells Depends on the Immunofluorescence Reactivity Intensity and Pattern.

Introduction: The indirect immunofluorescence assay on HEp-2 cells (HEp-2/IFA) is used worldwide for screening for autoantibodies to cellular antigens. Cell culture and fixation methods influence the cell distribution of autoantigens and the preservation of epitopes. Therefore, discrepancy of results obtained using different HEp-2/IFA kits (interkit nonreproducibility) is a common phenomenon in the clinical laboratory routine. Objective: This study evaluated the interkit nonreproducibility of HEp-2/IFA results using samples from patients with systemic autoimmune disease (SAD), nonautoimmune diseases (NAD), and healthy blood donors (HBD). Methods: Serum from 275 SAD patients, 293 NAD patients, and 300 HBD were processed at 1:80 dilution using four HEp-2 kits according to the manufacturers' instructions. Interkit reproducibility was determined for positive/negative results and patterns. The agreement of positive/negative results among kits for each sample was determined as the reactivity agreement score (RAS). The pattern reproducibility score (PRS) in each sample was calculated as a function of the number of kits showing equivalent patterns. Qualitative variables and ordinal variables were analyzed by the Chi-square and Mann-Whitney U tests, respectively. Results: A total of 402 samples were nonreactive in all kits and were considered devoid of autoantibodies. Further analysis included the 466 reactive samples (238 SAD, 119 NAD, 109 HBD). Reactivity to the nucleus had the highest interkit reproducibility (RAS = 83.6), followed by the metaphase plate (RAS = 78.9), cytoplasm (RAS = 77.4), and nucleolus (RAS = 72.4). Interkit reproducibility was higher in SAD (RAS = 78.0) than in NAD (RAS = 70.6) and HBD (RAS = 71.3) groups. Samples with strong reactivity (++++/4 and +++/4) had higher interkit reproducibility than those with weak reactivity (+/4). In the SAD group, RAS for nuclear reactivity was 87.5% for strongly reactive samples as opposed to 4.4% for weakly reactive samples, and the same was observed for NAD and HBD samples. The most robust patterns were the centromere AC-3 (PRS = 78.4), multiple nuclear dots AC-6 (PRS = 73.6), nuclear coarse speckled AC-5 (PRS = 71.3), nuclear homogeneous AC-1 (PRS = 67.9), and the reticular cytoplasmic AC-21 (PRS = 68.6). Conclusion: Interkit nonreproducibility in HEp-2/IFA is prevalent and occurs with the highest frequency with weakly reactive samples. International initiatives with the engagement of in vitro diagnostic industry are encouraged to promote the harmonization of the properties and performance of HEp-2/IFA commercial kits.

Malaria Elimination in Costa Rica: Changes in Treatment and Mass Drug Administration.

Costa Rica is a candidate to eliminate malaria by 2020. The remaining malaria transmission hotspots are located within the Huétar Norte Region (HNR), where 90% of the country's 147 malaria cases have occurred since 2016, following a 33-month period without transmission. Here, we examine changes in transmission with the implementation of a supervised seven-day chloroquine and primaquine treatment (7DCPT). We also evaluate the impact of a focal mass drug administration (MDA) in January 2019 at Boca Arenal, the town in HNR reporting the greatest local transmission. We found that the change to a seven-day treatment protocol, from the prior five-day program, was associated with a 98% reduction in malaria transmission. The MDA helped to reduce transmission, keeping the basic reproduction number, RT, significantly below 1, for at least four months. However, following new imported cases from Nicaragua, autochthonous transmission resumed. Our results highlight the importance of appropriate treatment delivery to reduce malaria transmission, and the challenge that highly mobile populations, if their malaria is not treated, pose to regional elimination efforts in Mesoamerica and México.

Changes in the result of antinuclear antibody immunofluorescence assay on HEp-2 cells reflect disease activity status in systemic lupus erythematosus.

Background The objective of the study was to determine whether the staining pattern and titer of indirect immunofluorescence assay on HEp-2 cells (HEp-2 IFA) are associated with systemic lupus erythematosus (SLE) disease activity. Methods A total of 269 consecutive patients meeting the ACR and SLICC criteria for SLE were classified into three groups according to the SLE Disease Activity Index 2000 (SLEDAI2K): Remission (SLEDAI2K = 0; n = 47); Intermediate (SLEDAI2K = 1-5; n = 111); Active (SLEDAI2K ≥ 6; n = 111). All subjects were assessed for HEp-2 IFA titer and staining pattern and nine traditional parameters of SLE disease activity. After a 6 to 12-month interval, 101 of the 269 patients were reassessed. Results HEp-2 IFA homogeneous nuclear pattern (AC-1) occurred more frequently in the Active Group compared to the Remission Group (p < 0.001). Fine speckled nuclear pattern (AC-4) tended to occur more frequently in the Remission Group compared to the Active Group (p = 0.054). Subjects with AC-1 pattern had higher SLEDAI (8.8 ± 7.6) than those with AC-4 (4.8 ± 5.2) (p < 0.001). HEp-2 IFA titer and anti-nuclear antibody by enzyme-linked immunosorbent assay (ANA-ELISA) values were lower in the Remission Group compared to the other two groups (p < 0.001). Multivariate analyses identified only ELISA anti-dsDNA as an independent variable associated with disease activity. In follow-up analysis, HEp-2 IFA titer decreased significantly in the 33 subjects with decreased disease activity (p = 0.002). Receiver operator characteristic (ROC) curve analysis for determination of disease activity showed equivalent areas under the curve (AUC) for HEp-2 IFA titer and traditional disease activity parameters. Conclusions HEp-2 IFA pattern and titer can reflect SLE disease activity and may be considered in conjunction with other laboratory and clinical parameters in the assessment of SLE disease activity.

Polysaccharide κ-Carrageenan as Doping Agent in Conductive Coatings for Electrochemical Controlled Release of Dexamethasone at Therapeutic Doses.

Smart conductive materials are developed in regenerative medicine to promote a controlled release profile of charged bioactive agents in the vicinity of implants. The incorporation and the active electrochemical release of the charged compounds into the organic conductive coating is achieved due to its intrinsic electrical properties. The anti-inflammatory drug dexamethasone was added during the polymerization, and its subsequent release at therapeutic doses was reached by electrical stimulation. In this work, a Poly (3,4-ethylenedioxythiophene): κ-carrageenan: dexamethasone film was prepared, and κ-carrageenan was incorporated to keep the electrochemical and physical stability of the electroactive matrix. The presence of κ-carrageenan and dexamethasone in the conductive film was confirmed by µ-Raman spectroscopy and their effect in the topographic was studied using profilometry. The dexamethasone release process was evaluated by cyclic voltammetry and High-Resolution mass spectrometry. In conclusion, κ-carrageenan as a doping agent improves the electrical properties of the conductive layer allowing the release of dexamethasone at therapeutic levels by electrochemical stimulation, providing a stable system to be used in organic bioelectronics systems.

Health policy impacts on malaria transmission in Costa Rica.

Costa Rica is near malaria elimination. This achievement has followed shifts in malaria health policy. Here, we evaluate the impacts that different health policies have had on malaria transmission in Costa Rica from 1913 to 2018. We identified regime shifts and used regression models to measure the impact of different health policies on malaria transmission in Costa Rica using annual case records. We found that vector control and prophylactic treatments were associated with a 50% malaria case reduction in 1929-1931 compared with 1913-1928. DDT introduction in 1946 was associated with an increase in annual malaria case reduction from 7.6% (1942-1946) to 26.4% (1947-1952). The 2006 introduction of 7-day supervised chloroquine and primaquine treatments was the most effective health policy between 1957 and 2018, reducing annual malaria cases by 98% (2009-2018) when compared with 1957-1968. We also found that effective malaria reduction policies have been sensitive to natural catastrophes and extreme climatic events, both of which have increased malaria transmission in Costa Rica. Currently, outbreaks follow malaria importation into vulnerable areas of Costa Rica. This highlights the need to timely diagnose and treat malaria, while improving living standards, in the affected areas.

Effects of salts and sucrose on the rheological behavior, thermal stability, and molecular structure of the Pereskia aculeata Miller mucilage.

The rheological behavior, thermogravimetric analysis (TGA), and atomic force microscopy (AFM) of the Pereskia aculeata Miller (OPN) mucilage treated with sodium chloride (NaCl), calcium chloride (CaCl2), and sucrose were evaluated. The experimental design was divided in a fractional factorial 25-1 for the screening of factors (OPN, sucrose, NaCl, CaCl2, and pH) and then in a 5 × 3 × 3 full factorial (OPN, sucrose, and NaCl). The model solutions used for the screening of factors presented shear-thinning behavior and the OPN mucilage concentrations were the factors that had significant effect on the apparent viscosity. Sucrose addition increased the thermal stability of the OPN mucilage solutions. OPN mucilage, sucrose, and NaCl were the variables with significant effect on thermogravimetric responses. The samples presented Newtonian behavior in 0-1.25% OPN mucilage concentrations and non-Newtonian behavior adjusted by power-law in 2.50-5.00% OPN mucilage concentrations with predominance of elastic behavior, contributing to the formation of stronger gels. The presence of sucrose in the systems containing OPN mucilage changed their rheological properties and salt additions caused reduction in viscosity. The AFM results provided a better understanding of the mechanism of OPN mucilage interactions in different solutions that justify the changes in viscosities.

Biological anti-TNF drugs: immunogenicity underlying treatment failure and adverse events.

INTRODUCTION: Genetically engineered monoclonal antibodies and fusion proteins directed against cytokines or their receptors represent a breakthrough in the treatment of various chronic immune-inflammatory diseases. Areas covered: Studies show high remission rates in several diseases, but clinical practice shows a significant percentage of individuals who do not exhibit the desired response. Loss of therapeutic benefit after initial successful response is designated secondary failure. Immune-biological agents are not self-antigens and are therefore potentially immunogenic. Secondary failure is frequently caused by antibodies against immune-biologicals, known as anti-drug antibodies (ADA). ADA that neutralize circulating immune-biologicals and/or promote their clearance can reduce treatment efficacy. Furthermore, ADA can induce adverse events by diverse immunological mechanisms. This review provides a comprehensive overview of ADA in rheumatoid arthritis patients treated with anti-TNF immune-biologicals, and explores the concept of therapeutic drug monitoring (TDM) as an effective strategy to improve therapeutic management. Expert opinion: Monitoring circulating ADA and therapeutic immune-biological drugs is helpful when evaluating patients with secondary failure. However, immunological tests for ADA vary considerably regarding their ability to detect different types of ADA. Several assays are not designed to determine ADA-induced drug neutralizing capacity, and they may report clinically non-relevant data, especially if drug is present in test samples.

Shedding of host autophagic proteins from the parasitophorous vacuolar membrane of Plasmodium berghei.

The hepatic stage of the malaria parasite Plasmodium is accompanied by an autophagy-mediated host response directly targeting the parasitophorous vacuolar membrane (PVM) harbouring the parasite. Removal of the PVM-associated autophagic proteins such as ubiquitin, p62, and LC3 correlates with parasite survival. Yet, it is unclear how Plasmodium avoids the deleterious effects of selective autophagy. Here we show that parasites trap host autophagic factors in the tubovesicular network (TVN), an expansion of the PVM into the host cytoplasm. In proliferating parasites, PVM-associated LC3 becomes immediately redirected into the TVN, where it accumulates distally from the parasite's replicative centre. Finally, the host factors are shed as vesicles into the host cytoplasm. This strategy may enable the parasite to balance the benefits of the enhanced host catabolic activity with the risk of being eliminated by the cell's cytosolic immune defence.

Calidad de la atención en internación psiquiátrica en hospitales generales de cuatro provincias argentinas / Quality of care in psychiatric hospitalization in general hospitals of four argentine provinces

Argentina está en un proceso de transformación de la atención en salud mental, lo que implica el desarrollo de nuevos dispositivos de atención y el fortalecimiento de aquellos consonantes con la reforma, entre los que se encuentran los servicios de internación psiquiátrica en hospitales generales. La situación de dichos servicios en el país es heterogénea cuantitativa y cualitativamente, y son pocos los estudios al respecto. A partir de lo anterior, el objetivo general de este estudio fue evaluar la calidad de la atención de servicios de salud mental con internación psiquiátrica en hospitales generales, desde los ejes de la orientación comunitaria y el respeto y salvaguarda de derechos. Se realizó un estudio observacional, enmarcado en la investigación evaluativa y en la investigación-acción, en cuatro servicios ubicados en diferentes provincias del país. Se realizó análisis documental de datos estadísticos de los servicios, y entrevistas con el responsable, trabajadores y usuarios de los servicios. El análisis de los datos fue mixto. Los resultados indican que los derechos de los usuarios tienden a ser respetados durante la internación, en especial la conservación de los vínculos. En cuanto a la información sobre el diagnóstico y tratamiento, aún se requiere afianzar lo referido al consentimiento informado formal (escrito). Sobre la toma de decisiones, éstas tienden a incluir a los usuarios, aunque se desconocen las decisiones anticipadas. Los usuarios por su parte desconocen sus derechos, y las asociaciones que los nuclean. La orientación comunitaria se desarrolla mediante diversas estrategias como el trabajo con redes (formales e informales), la preparación para el alta, y las acciones para motivar el mantenimiento de los vínculos sociales y afectivos. Se observa un reconocimiento sobre la importancia y utilidad de la evaluación, pero la insuficiencia de herramientas para implementarla. Los datos estadísticos tienden a ser de baja calidad

25-Hydroxivitamin D Serum Concentration, Not Free and Bioavailable Vitamin D, Is Associated with Disease Activity in Systemic Lupus Erythematosus Patients.

We aim to evaluate the prevalence of vitamin D deficiency in patients with systemic lupus erythematosus (SLE) and investigate the association between total, free and bioavailable vitamin D serum concentrations and disease activity. Patients with SLE (ACR 1997) consecutively seen at UNIFESP's outpatient's clinics had disease activity measured after clinical and laboratory evaluation using SLEDAI (Systemic Lupus Erythematosus Disease Activity Index). 25-hydroxyvitamin D (25(OH)D) serum concentrations measured by chemiluminescence and vitamin D binding protein (DBP) measured by ELISA were used to calculate free and bioavailable vitamin D. Healthy blood donors were used as controls. A total of 142 patients (71.4%) had 25(OH)D serum concentrations below 30 ng/mL. Total 25(OH)D serum concentration was associated with disease activity categorized in 5 continuous groups of SLEDAI. 25(OH)D serum concentrations were higher among patients with SLEDAI 1-5 and lower in those with severe activity (SLEDAI≥20) (p <0.05). On the other hand, no statistically significant difference was observed for DBP, free and bioavailable vitamin D measurements in the disease activity subgroups evaluated. Vitamin D deficiency is highly prevalent among patients with SLE and was associated with higher disease activity. DBP serum level and calculation of free and bioavailable vitamin D were not associated with SLE disease activity.

Differential capacity of therapeutic drugs to induce Rods/Rings structures in vitro and in vivo and generation of anti-Rods/Rings autoantibodies.

Some HCV patients using ribavirin and interferon alpha (IFN-α) develop anti-rods and rings (RR) autoantibodies, the main target of which is inosine monophosphate dehydrogenase (IMPDH), the rate-determining enzyme in de novo GTP biosynthesis. In vitro inhibition of IMPDH by ribavirin induces RR formation. Here we investigate whether other commonly used drugs that interfere with GTP biosynthesis can induce RR structures in vitro and vivo and elicit generation of autoantibodies. HEp-2 cells treated for 24h with ribavirin, mycophenolic acid (MPA), azathioprine, methotrexate or acyclovir were positive for RR structures. However, adefovir, entecavir, tenofovir and lamivudine did not induce RR structures in these cells. Structures induced by ribavirin in HEp-2 cells are stable after 24h drug-washout, while structures induced by other drugs are relatively labile, disappearing within 2h. Looking at patients treated with these drugs, HCV patients treated with ribavirin (n=17) showed higher average percentage of RR-positive peripheral mononuclear cells than autoimmune patients treated with RR-inducing immunosuppressant drugs (n=21). Serum from 173 autoimmune patients who had been treated with MPA, azathioprine or methotrexate was tested for presence of anti-RR autoantibodies, and only one sample was found to be positive. Conversely, of 48 anti-RR autoantibody positive samples identified at Fleury Laboratories over 30months, 94% were from HCV patients treated with ribavirin plus IFN-α. These data indicate that RR structures can be induced by a variety of drugs in vitro and in vivo, but anti-RR autoantibody production is mostly restricted to HCV patients under ribavirin+IFN-α treatment.

The machinery underlying malaria parasite virulence is conserved between rodent and human malaria parasites.

Sequestration of red blood cells infected with the human malaria parasite Plasmodium falciparum in organs such as the brain is considered important for pathogenicity. A similar phenomenon has been observed in mouse models of malaria, using the rodent parasite Plasmodium berghei, but it is unclear whether the P. falciparum proteins known to be involved in this process are conserved in the rodent parasite. Here we identify the P. berghei orthologues of two such key factors of P. falciparum, SBP1 and MAHRP1. Red blood cells infected with P. berghei parasites lacking SBP1 or MAHRP1a fail to bind the endothelial receptor CD36 and show reduced sequestration and virulence in mice. Complementation of the mutant P. berghei parasites with the respective P. falciparum SBP1 and MAHRP1 orthologues restores sequestration and virulence. These findings reveal evolutionary conservation of the machinery underlying sequestration of divergent malaria parasites and support the notion that the P. berghei rodent model is an adequate tool for research on malaria virulence.

Long-term live imaging reveals cytosolic immune responses of host hepatocytes against Plasmodium infection and parasite escape mechanisms.

Plasmodium parasites are transmitted by Anopheles mosquitoes to the mammalian host and actively infect hepatocytes after passive transport in the bloodstream to the liver. In their target host hepatocyte, parasites reside within a parasitophorous vacuole (PV). In the present study it was shown that the parasitophorous vacuole membrane (PVM) can be targeted by autophagy marker proteins LC3, ubiquitin, and SQSTM1/p62 as well as by lysosomes in a process resembling selective autophagy. The dynamics of autophagy marker proteins in individual Plasmodium berghei-infected hepatocytes were followed by live imaging throughout the entire development of the parasite in the liver. Although the host cell very efficiently recognized the invading parasite in its vacuole, the majority of parasites survived this initial attack. Successful parasite development correlated with the gradual loss of all analyzed autophagy marker proteins and associated lysosomes from the PVM. However, other autophagic events like nonselective canonical autophagy in the host cell continued. This was indicated as LC3, although not labeling the PVM anymore, still localized to autophagosomes in the infected host cell. It appears that growing parasites even benefit from this form of nonselective host cell autophagy as an additional source of nutrients, as in host cells deficient for autophagy, parasite growth was retarded and could partly be rescued by the supply of additional amino acid in the medium. Importantly, mouse infections with P. berghei sporozoites confirmed LC3 dynamics, the positive effect of autophagy activation on parasite growth, and negative effects upon autophagy inhibition.