Schistosoma mansoni cercarial elastase (SmCE): differences in immunogenic properties of native and recombinant forms

The Schistosoma mansoni cercarial elastase (SmCE) has previously been shown to be poorly immunogenic in mice. However, a minority of mice were able to produce antibodies against SmCE after multiple immunizations with crude pre- parations containing the enzyme. These mice were partially protected against challenge infections of S. mansoni. In the present study, we show that in contrast to the poor immunogenicity of the enzymatically active native form of SmCE derived from a crude preparation (cercarial transformation fluid), immunization of CBA/Ca mice with two enzymatically inactive forms, namely purified native SmCE or a recombinant SmCE fused to recombinant Schistosoma japonicum gluta- thione S-transferase (rSmCE-SjGST), after adsorption onto aluminum hydroxide adjuvant, induced specific anti-SmCE immunoglobulin G (IgG) in all mice within 2 weeks of the second immunization. The IgG antibody response to rSmCE- SjGST was mainly of the IgG1 subclass. These results suggest that inactive forms of the antigen could be used to obtain the optimum immunogenic effects as a vaccine candidate against schistosomiasis. Mice immunized with the rSmCE- SjGST on alum had smaller mean worm burdens and lower tissue egg counts when compared with adjuvant alone- and recombinant SjGST-injected controls. The native SmCE was antigenically cross-reactive with homologous enzymes of Schistosoma haematobium and Schistosoma margrebowiei

Global, regional, and national burden of disorders affecting the nervous system, 1990–2021: a systematic analysis for the Global Burden of Disease Study 2021

BackgroundDisorders affecting the nervous system are diverse and include neurodevelopmental disorders, late-life neurodegeneration, and newly emergent conditions, such as cognitive impairment following COVID-19. Previous publications from the Global Burden of Disease, Injuries, and Risk Factor Study estimated the burden of 15 neurological conditions in 2015 and 2016, but these analyses did not include neurodevelopmental disorders, as defined by the International Classification of Diseases (ICD)-11, or a subset of cases of congenital, neonatal, and infectious conditions that cause neurological damage. Here, we estimate nervous system health loss caused by 37 unique conditions and their associated risk factors globally, regionally, and nationally from 1990 to 2021.MethodsWe estimated mortality, prevalence, years lived with disability (YLDs), years of life lost (YLLs), and disability-adjusted life-years (DALYs), with corresponding 95% uncertainty intervals (UIs), by age and sex in 204 countries and territories, from 1990 to 2021. We included morbidity and deaths due to neurological conditions, for which health loss is directly due to damage to the CNS or peripheral nervous system. We also isolated neurological health loss from conditions for which nervous system morbidity is a consequence, but not the primary feature, including a subset of congenital conditions (ie, chromosomal anomalies and congenital birth defects), neonatal conditions (ie, jaundice, preterm birth, and sepsis), infectious diseases (ie, COVID-19, cystic echinococcosis, malaria, syphilis, and Zika virus disease), and diabetic neuropathy. By conducting a sequela-level analysis of the health outcomes for these conditions, only cases where nervous system damage occurred were included, and YLDs were recalculated to isolate the non-fatal burden directly attributable to nervous system health loss. A comorbidity correction was used to calculate total prevalence of all conditions that affect the nervous system combined.FindingsGlobally, the 37 conditions affecting the nervous system were collectively ranked as the leading group cause of DALYs in 2021 (443 million, 95% UI 378–521), affecting 3·40 billion (3·20–3·62) individuals (43·1%, 40·5–45·9 of the global population); global DALY counts attributed to these conditions increased by 18·2% (8·7–26·7) between 1990 and 2021. Age-standardised rates of deaths per 100 000 people attributed to these conditions decreased from 1990 to 2021 by 33·6% (27·6–38·8), and age-standardised rates of DALYs attributed to these conditions decreased by 27·0% (21·5–32·4). Age-standardised prevalence was almost stable, with a change of 1·5% (0·7–2·4). The ten conditions with the highest age-standardised DALYs in 2021 were stroke, neonatal encephalopathy, migraine, Alzheimer's disease and other dementias, diabetic neuropathy, meningitis, epilepsy, neurological complications due to preterm birth, autism spectrum disorder, and nervous system cancer.InterpretationAs the leading cause of overall disease burden in the world, with increasing global DALY counts, effective prevention, treatment, and rehabilitation strategies for disorders affecting the nervous system are needed

Global burden of 288 causes of death and life expectancy decomposition in 204 countries and territories and 811 subnational locations, 1990–2021: a systematic analysis for the Global Burden of Disease Study 2021

BACKGROUND Regular, detailed reporting on population health by underlying cause of death is fundamental for public health decision making. Cause-specific estimates of mortality and the subsequent effects on life expectancy worldwide are valuable metrics to gauge progress in reducing mortality rates. These estimates are particularly important following large-scale mortality spikes, such as the COVID-19 pandemic. When systematically analysed, mortality rates and life expectancy allow comparisons of the consequences of causes of death globally and over time, providing a nuanced understanding of the effect of these causes on global populations. METHODS The Global Burden of Diseases, Injuries, and Risk Factors Study (GBD) 2021 cause-of-death analysis estimated mortality and years of life lost (YLLs) from 288 causes of death by age-sex-location-year in 204 countries and territories and 811 subnational locations for each year from 1990 until 2021. The analysis used 56 604 data sources, including data from vital registration and verbal autopsy as well as surveys, censuses, surveillance systems, and cancer registries, among others. As with previous GBD rounds, cause-specific death rates for most causes were estimated using the Cause of Death Ensemble model-a modelling tool developed for GBD to assess the out-of-sample predictive validity of different statistical models and covariate permutations and combine those results to produce cause-specific mortality estimates-with alternative strategies adapted to model causes with insufficient data, substantial changes in reporting over the study period, or unusual epidemiology. YLLs were computed as the product of the number of deaths for each cause-age-sex-location-year and the standard life expectancy at each age. As part of the modelling process, uncertainty intervals (UIs) were generated using the 2·5th and 97·5th percentiles from a 1000-draw distribution for each metric. We decomposed life expectancy by cause of death, location, and year to show cause-specific effects on life expectancy from 1990 to 2021. We also used the coefficient of variation and the fraction of population affected by 90% of deaths to highlight concentrations of mortality. Findings are reported in counts and age-standardised rates. Methodological improvements for cause-of-death estimates in GBD 2021 include the expansion of under-5-years age group to include four new age groups, enhanced methods to account for stochastic variation of sparse data, and the inclusion of COVID-19 and other pandemic-related mortality-which includes excess mortality associated with the pandemic, excluding COVID-19, lower respiratory infections, measles, malaria, and pertussis. For this analysis, 199 new country-years of vital registration cause-of-death data, 5 country-years of surveillance data, 21 country-years of verbal autopsy data, and 94 country-years of other data types were added to those used in previous GBD rounds. FINDINGS The leading causes of age-standardised deaths globally were the same in 2019 as they were in 1990; in descending order, these were, ischaemic heart disease, stroke, chronic obstructive pulmonary disease, and lower respiratory infections. In 2021, however, COVID-19 replaced stroke as the second-leading age-standardised cause of death, with 94·0 deaths (95% UI 89·2-100·0) per 100 000 population. The COVID-19 pandemic shifted the rankings of the leading five causes, lowering stroke to the third-leading and chronic obstructive pulmonary disease to the fourth-leading position. In 2021, the highest age-standardised death rates from COVID-19 occurred in sub-Saharan Africa (271·0 deaths [250·1-290·7] per 100 000 population) and Latin America and the Caribbean (195·4 deaths [182·1-211·4] per 100 000 population). The lowest age-standardised death rates from COVID-19 were in the high-income super-region (48·1 deaths [47·4-48·8] per 100 000 population) and southeast Asia, east Asia, and Oceania (23·2 deaths [16·3-37·2] per 100 000 population). Globally, life expectancy steadily improved between 1990 and 2019 for 18 of the 22 investigated causes. Decomposition of global and regional life expectancy showed the positive effect that reductions in deaths from enteric infections, lower respiratory infections, stroke, and neonatal deaths, among others have contributed to improved survival over the study period. However, a net reduction of 1·6 years occurred in global life expectancy between 2019 and 2021, primarily due to increased death rates from COVID-19 and other pandemic-related mortality. Life expectancy was highly variable between super-regions over the study period, with southeast Asia, east Asia, and Oceania gaining 8·3 years (6·7-9·9) overall, while having the smallest reduction in life expectancy due to COVID-19 (0·4 years). The largest reduction in life expectancy due to COVID-19 occurred in Latin America and the Caribbean (3·6 years). Additionally, 53 of the 288 causes of death were highly concentrated in locations with less than 50% of the global population as of 2021, and these causes of death became progressively more concentrated since 1990, when only 44 causes showed this pattern. The concentration phenomenon is discussed heuristically with respect to enteric and lower respiratory infections, malaria, HIV/AIDS, neonatal disorders, tuberculosis, and measles. INTERPRETATION Long-standing gains in life expectancy and reductions in many of the leading causes of death have been disrupted by the COVID-19 pandemic, the adverse effects of which were spread unevenly among populations. Despite the pandemic, there has been continued progress in combatting several notable causes of death, leading to improved global life expectancy over the study period. Each of the seven GBD super-regions showed an overall improvement from 1990 and 2021, obscuring the negative effect in the years of the pandemic. Additionally, our findings regarding regional variation in causes of death driving increases in life expectancy hold clear policy utility. Analyses of shifting mortality trends reveal that several causes, once widespread globally, are now increasingly concentrated geographically. These changes in mortality concentration, alongside further investigation of changing risks, interventions, and relevant policy, present an important opportunity to deepen our understanding of mortality-reduction strategies. Examining patterns in mortality concentration might reveal areas where successful public health interventions have been implemented. Translating these successes to locations where certain causes of death remain entrenched can inform policies that work to improve life expectancy for people everywhere. FUNDING Bill & Melinda Gates Foundation

Can antibody conjugated nanomicelles alter the prospect of antibody targeted therapy against schistosomiasis mansoni?

BackgroundCLA (conjugated linoleic acid)-mediated activation of the schistosome tegument-associated sphingomyelinase and consequent disruption of the outer membrane might allow host antibodies to access the apical membrane antigens. Here, we investigated a novel approach to enhance specific antibody delivery to concealed surface membrane antigens of Schistosoma mansoni utilising antibody-conjugated-CLA nanomicelle technology.Methodology/principal findingsWe invented and characterised an amphiphilic CLA-loaded whey protein co-polymer (CLA-W) as an IV injectable protein nanocarrier. Rabbit anti-Schistosoma mansoni infection (anti-SmI) and anti-Schistosoma mansoni alkaline phosphatase specific IgG antibodies were purified from rabbit sera and conjugated to the surface of CLA-W co-polymer to form antibody-conjugated-CLA-W nanomicelles (Ab-CLA-W). We investigated the schistosomicidal effects of CLA-W and Ab-CLA-W in a mouse model of Schistosoma mansoni against early and late stages of infection. Results showed that conjugation of nanomicelles with antibodies, namely anti-SmI, significantly enhanced the micelles' schistosomicidal and anti-pathology activities at both the schistosomula and adult worm stages of the infection resulting in 64.6%-89.9% reductions in worm number; 72.5-94% and 66.4-85.2% reductions in hepatic eggs and granulomas, respectively. Treatment induced overall improvement in liver histopathology, reducing granuloma size and fibrosis and significantly affecting egg viability. Indirect immunofluorescence confirmed CLA-W-mediated antigen exposure on the worm surface. Electron microscopy revealed extensive ultrastructural damage in worm tegument induced by anti-SmI-CLA-W.Conclusion/significanceThe novel antibody-targeted nano-sized CLA delivery system offers great promise for treatment of Schistosoma mansoni infection and control of its transmission. Our in vivo observations confirm an immune-mediated enhanced effect of the schistosomicidal action of CLA and hints at the prospect of nanotechnology-based immunotherapy, not only for schistosomiasis, but also for other parasitic infections in which chemotherapy has been shown to be immune-dependent. The results propose that the immunodominant reactivity of the anti-SmI serum, Schistosoma mansoni fructose biphosphate aldolase, SmFBPA, merits serious attention as a therapeutic and vaccine candidate

Treatment of <i>Schistosoma mansoni</i> with miltefosine <i>in vitro</i> enhances serological recognition of defined worm surface antigens

<div><p>Background</p><p>Miltefosine, an anti-cancer drug that has been successfully repositioned for treatment of <i>Leishmania</i> infections, has recently also shown promising effects against <i>Schistosoma</i> spp targeting all life cycle stages of the parasite. The current study examined the effect of treating <i>Schistosoma mansoni</i> adult worms with miltefosine on exposure of worm surface antigens <i>in vitro</i>.</p><p>Methodology/Principal findings</p><p>In an indirect immunofluorescence assay, rabbit anti-<i>S</i>.<i>mansoni</i> adult worm homogenate and anti-<i>S</i>. <i>mansoni</i> infection antisera gave strong immunofluorescence of the <i>S</i>. <i>mansoni</i> adult worm surface after treatment with miltefosine, the latter antiserum having previously been shown to synergistically enhance the schistosomicidal activity of praziquantel. Rabbit antibodies that recognised surface antigens exposed on miltefosine-treated worms were recovered by elution off the worm surface in low pH buffer and were used in a western immunoblotting assay to identify antigenic targets in a homogenate extract of adult worms (SmWH). Four proteins reacting with the antibodies in immunoblots were purified and proteomic analysis (MS/MS) combined with specific immunoblotting indicated they were the <i>S</i>. <i>mansoni</i> proteins: fructose-1,6 bisphosphate aldolase (SmFBPA), Sm22.6, alkaline phosphatase and malate dehydrogenase. These antibodies were also found to bind to the surface of 3-hour schistosomula and induce immune agglutination of the parasites, suggesting they may have a role in immune protection.</p><p>Conclusion/Significance</p><p>This study reveals a novel mode of action of miltefosine as an anti-schistosome agent. The immune-dependent hypothesis we investigated has previously been lent credence with praziquantel (PZQ), whereby treatment unmasks parasite surface antigens not normally exposed to the host during infection. Antigens involved in this molecular mechanism could have potential as intervention targets and antibodies against these antigens may act to increase the drug’s anti-parasite efficacy and be involved in the development of resistance to re-infection.</p></div

Characterization of worm surface antigens recognized by rabbit anti-SmI antibodies after miltefosine- and PZQ-treatment.

<p>(i) Immunofluorescent staining of PZQ- (6 μg/ml for 24 hours) (A and C) and miltefosine- (40 μg/ml for 48 hours) (B and D) treated <i>S</i>. <i>mansoni</i> adult worms detected with rabbit anti-SmI IgG antibodies (A and B) and IgG antibodies from a normal rabbit serum (C and D). Scale bar = 100 μm. (ii) Western immunoblots of a <i>S</i>. <i>mansoni</i> crude worm homogenate preparation (SmWH) were probed with rabbit anti-SmI antiserum (lane 1), rabbit anti-SmI antibodies eluted from PZQ- (6 μg/ml for 24 hours) (lane 2) and miltefosine- (40 μg/ml for 48 hours) (lane 3) treated worms. A blot of SmWH probed with a normal rabbit serum (lane 4) was used as a control. Lane M, protein molecular weight markers. Antigenic extract was loaded onto the gel with 10 μg protein/lane. Blots were detected using HRP-conjugated anti-rabbit IgG secondary antibodies. Detected antigens with the most intense reactivities are arrowed.</p

Detection by rabbit anti-SmWH antibodies of worm surface antigens exposed by miltefosine or PZQ.

<p>(i) Immunofluorescent staining of PZQ- (6 μg/ml for 24 hours) (A and C) and miltefosine- (40 μg/ml for 48 hours) (B and D) treated <i>S</i>. <i>mansoni</i> adult worms detected with rabbit anti-SmWH IgG antibodies (A and B) and IgG antibodies from a rabbit that received adjuvant alone (C and D). Scale bar = 100 μm. (ii) Western immunoblots of <i>S</i>. <i>mansoni</i> crude worm homogenate preparation (SmWH) were probed with rabbit anti-SmWH antiserum (lane 1), rabbit anti-SmWH antibodies eluted from PZQ- (6 μg/ml for 24 hours) (lane 2) and miltefosine- (40 μg/ml for 48 hours) (lane 3) treated worms. A blot of SmWH probed with a serum from a rabbit injected with Freund’s adjuvant alone (lane 4) was used as a control. Lane M, protein molecular weight markers. Antigenic extract was loaded onto the gel with 10 μg protein/lane. Blots were detected using HRP-conjugated anti-rabbit IgG secondary antibodies. Detected antigens with the most intense reactivities are arrowed.</p

<i>In vitro</i> lethal effect of miltefosine on <i>S</i>. <i>mansoni</i> adult worms.

<p>LC50s of the drug (concentrations that kill 50% of worms) 24, 48, 72 and 96 hours after <i>in vitro</i> exposure to miltefosine were calculated from concentration-response curves at the respective time points and LC50 values were plotted against length of incubation.</p

<i>In vitro</i> activity of miltefosine against <i>S</i>. <i>mansoni</i> adult worms.

<p><i>In vitro</i> activity of miltefosine against <i>S</i>. <i>mansoni</i> adult worms.</p